bims-aporos Biomed news
on Apoptosis and reactive oxygen species
Issue of 2019‒01‒27
sixty-one papers selected by
Gavin McStay
Staffordshire University

  1. Cell Biosci. 2019 ;9 12
    Hu X, Wu X, Zhao B, Wang Y.
      Background: Proliferative vitreoretinopathy (PVR) is a severe blinding complication of retinal detachment surgery. Increasing evidence demonstrate that PVR is associated with oxidative stress. Scutellarin is a natural flavone compound that has been reported to have anti-oxidative activity. However, the effect of scutellarin on PVR remains unknown. In the current study, we assessed the effect of scutellarin on hydrogen peroxide (H2O2)-induced oxidative injury in human retinal pigment epithelium cells (ARPE-19).Methods: ARPE-19 cells were pretreated with different concentrations of scutellarin for 2 h, and then challenged with H2O2 (1 mM) for 24 h. The levels of reactive oxygen species (ROS), malondialdehyde (MDA) and superoxide dismutase (SOD) and glutathione (GSH) activity were measured to assess the level of oxidative stress. Flow cytometry was performed to detect the apoptosis rate of ARPE-19 cells. Expression levels of bcl-2, bax, cleaved-caspase-3, p-JAK2, JAK2, p-STAT3, and STAT3 were measured using western blot.
    Results: Our results revealed that scutellarin improved the cell viability of H2O2-induced ARPE-19 cells. Scutellarin alleviated the H2O2-induced oxidative stress in ARPE-19 cells, which was illustrated by reduced levels of ROS and MDA, accompanied by increased SOD activity and GSH level. The increased apoptosis rate of ARPE-19 cells caused by H2O2 induction was significantly decreased after scutellarin treatment. H2O2 treatment resulted in significant increase in bax expression and decrease in bcl-2 expression, while the changes in the expressions of bax and bcl-2 were reversed by scutellarin treatment. In addition, scutellarin promoted the activation of JAK2/STAT3 signaling pathway in H2O2-induced ARPE-19 cells. Suppression of JAK2/STAT3 signaling pathway abolished the protective effects of scutellarin on H2O2-induced ARPE-19 cells.
    Conclusion: These findings suggested that scutellarin was capable for alleviating H2O2-induced oxidative damage in ARPE-19 cells, which might be ascribed to the activation of JAK2/STAT3 signaling pathway.
    Keywords:  JAK2/STAT3 signaling pathway; Oxidative stress; Proliferative vitreoretinopathy (PVR); Retinal pigment epithelium (RPE) cells; Scutellarin
  2. Phytomedicine. 2018 Sep 19. pii: S0944-7113(18)30481-1. [Epub ahead of print]54 224-230
    Firdaus F, Zafeer MF, Anis E, Ahmad F, Hossain MM, Ali A, Afzal M.
      BACKGROUND: It is evaluated that a few million individuals worldwide are experiencing Arsenic (As) harmfulness coming about because of anthropogenic discharges. There is likewise proof to propose that As can affect the peripheral, as well as, the central nervous system (CNS). On the contrary, thymoquinone (TQ), a biologically active ingredient of Nigella sativa has exhibited numerous neuro-pharmacological traits since ancient times.HYPOTHESIS/PURPOSE: In the present study, the neuroprotective efficacy of TQ was explored by primarily studying its antioxidant and anti-apoptotic potential against Arsenic trioxide (As2O3) induced toxicity in SH-SY5Y human neuroblastoma cell lines.
    STUDY DESIGN: For experimentation, cells were seeded in 96 well tissue culture plates and kept undisturbed for 24 h to attain proper adhesion. After 75-80% confluence, cells were pretreated with 10 µM and 20 µM thymoquinone (TQ) for 1 h After adding 2 µM As, cells were set aside for incubation for 24 h without changing the medium.
    METHODS: The mitigatory effects of TQ with particular reference to cell viability and cytotoxicity, the generation of reactive oxygen species, DNA damage, and mitochondrial dynamics were studied.
    RESULTS: Pretreatment of SH-SY5Y cells with TQ (10 and 20 μM) for an hour and subsequent exposure to 2 μM As2O3 protected the SH-SY5Y cells against the neuro-damaging effects of the latter. Also, the SH-SY5Y cells were better preserved with increased viability, repaired DNA, less free radical generation and balanced transmembrane potential than those exposed to As2O3 alone. TQ pretreatment also inhibited As2O3-induced exacerbation in protein levels of BAX and PARP-1 and restored the loss of Bcl2 levels.
    CONCLUSION: The findings of this study suggest that TQ may prevent neurotoxicity and As2O3-induced apoptosis and cytotoxicity. It is, therefore, worth studying further for its potential to reduce the risks of arsenic-related neurological implications.
    Keywords:  Abbreviations: As, Arsenic; Apoptosis; As(2)O(3), Arsenic trioxide; DCFDA, 2′, 7′–dichlorofluorescein diacetate; Flow cytometry; MMP, Mitochondrial Membrane Potential; Mitochondria; ROS, Reactive Oxygen Species; Reactive Oxygen Species; Rh-123, Rhodamine-123; TQ, Thymoquinone
  3. Int J Med Sci. 2019 ;16(1): 145-155
    Jeong JY, Cha HJ, Choi EO, Kim CH, Kim GY, Yoo YH, Hwang HJ, Park HT, Yoon HM, Choi YH.
      Baicalein, a flavonoid extracted from the roots of Scutellaria baicalensis Georgi., has various pharmacological effects due to its high antioxidant activity. However, no study has yet been conducted on the protective efficacy of baicalein against oxidative stress in Schwann cells. In this study, we evaluated the protective effect of baicalein on DNA damage and apoptosis induced by hydrogen peroxide (H2O2) in HEI193 Schwann cells. For this purpose, HEI193 cells exposed to H2O2 in the presence or absence of baicalein were applied to cell viability assay, immunoblotting, Nrf2-specific small interfering RNA (siRNA) transfection, comet assay, and flow cytometry analyses. Our results showed that baicalein effectively inhibited H2O2-induced cytotoxicity and DNA damage associated with the inhibition of reactive oxygen species (ROS) accumulation. Baicalein also weakened H2O2-induced mitochondrial dysfunction, increased the Bax/Bcl-2 ratio, activated caspase-9 and -3, and degraded poly(ADP-ribose) polymerase. In addition, baicalein increased not only the expression but also the phosphorylation of nuclear factor-erythroid 2 related factor 2 (Nrf2) and promoted the expression of heme oxygenase-1 (HO-1), a critical target enzyme of Nrf2, although the expression of kelch-like ECH-associated protein-1 was decreased. However, the inhibition of Nrf2 expression by transfection with Nrf2-siRNA transfection abolished the expression of HO-1 and antioxidant potential of baicalein. These results demonstrate that baicalein attenuated H2O2-induced apoptosis through the conservation of mitochondrial function while eliminating ROS in HEI193 Schwann cells, and the antioxidant efficacy of baicalein implies at least a Nrf2/HO-1 signaling pathway-dependent mechanism. Therefore, it is suggested that baicalein may have a beneficial effect on the prevention and treatment of peripheral neuropathy induced by oxidative stress.
    Keywords:  Baicalein; DNA damage; Nrf2/HO-1; Schwann cells; apoptosis; oxidative stress
  4. Neuroreport. 2019 Jan 22.
    Li H, Li X, Luo W, Zhu X.
      This study investigated the neuroprotective effects of (Z)-7,4'-dimethoxy-6-hydroxy-aurone-4-O-β-glucopyranoside (DHG) against hydrogen peroxide-induced cell damage in PC12 cells and further evaluated its effects on doxorubicin-induced anxiety-like behavior in rats. PC12 cells were treated with different concentrations of DHG (1-10 µM) for 24 h and then exposed to 0.2 mM hydrogen peroxide for 12 h. Cell viability was measured using the 3-(4,5-dimethylthiazol)-2,5-diphenyltetrazolium-bromide and lactate dehydrogenase assay. The apoptosis was detected by annexin V-PI staining. Oxidative stress was confirmed in PC12 cells by enzyme-linked immunosorbent assay, real-time PCR, and western blot. The anxiolytic effects of DHG were evaluated in the elevated plus maze test. Our results showed that DHG treatment significantly increased cell viability and decreased PC12 cell apoptosis induced by hydrogen peroxide by increasing the mitochondrial membrane potential, decreasing the cytochrome c release, inhibiting the activities of caspase-3 and caspase-9, and regulating the expression of apoptosis-related proteins. Moreover, DHG treatment effectively attenuated the redox imbalance in PC12 cells by enhancing the activity of superoxide dismutase and increasing the level of glutathione, as well as decreasing levels of malondialdehyde and intracellular reactive oxygen species. Furthermore, DHG treatment significantly mitigated the doxorubicin-induced adverse behavioral changes in rats. These results indicate the neuroprotective and antiapoptotic properties of DHG exerted by counteracting the oxidative stress and highlight DHG as a potential therapeutic regimen for behavior impairment of anxiety.
  5. Nanomaterials (Basel). 2019 Jan 22. pii: E139. [Epub ahead of print]9(2):
    Gurunathan S, Kang MH, Jeyaraj M, Kim JH.
      Graphene oxide (GO) is an common nanomaterial and has attracted unlimited interest in academia and industry due to its physical, chemical, and biological properties, as well as for its tremendous potential in applications in various fields, including nanomedicine. Whereas studies have evaluated the size-dependent cytotoxicity of GO in cancer cells, there have been no studies on the biological behavior of ultra-small graphene nanosheets in germ cells. To investigate, for the first time, the cyto- and geno- toxic effects of different sizes of GO in two different cell types, Leydig (TM3) and Sertoli (TM4) cells, we synthesized different sized GO nanosheets with an average size of 100 and 20 nm by a modification of Hummers' method, and characterized them by various analytical techniques. Cell viability and proliferation assays showed significant size- and dose-dependent toxicity with GO-20 and GO-100. Interestingly, GO-20 induced significant loss of cell viability and cell proliferation, higher levels of leakage of lactate dehydrogenase (LDH) and reactive oxygen species (ROS) generation compared to GO-100. Both GO-100 and GO-20 induced significant loss of mitochondrial membrane potential (MMP) in TM3 and TM4 cells, which is a critical factor for ROS generation. Furthermore, GO-100 and GO-20 caused oxidative damage to DNA by increasing the levels of 8-oxo-dG, which is formed by direct attack of ROS on DNA; GO-100 and GO-20 upregulate various genes responsible for DNA damage and apoptosis. We found that phosphorylation levels of EGFR/AKT signaling molecules, which are related to cell survival and apoptosis, were significantly altered after GO-100 and GO-20 exposure. Our results showed that GO-20 has more potent toxic effects than GO-100, and that the loss of MMP and apoptosis are the main toxicity responses to GO-100 and GO-20 treatments, which likely occur due to EGFR/AKT pathway regulation. Collectively, our results suggest that both GO-100 and GO-20 exhibit size-dependent germ cell toxicity in male somatic cells, particularly TM3 cells, which seem to be more sensitive compared to TM4, which strongly suggests that applications of GO in commercial products must be carefully evaluated.
    Keywords:  DNA damage; Leydig cells; Sertoli cells; apoptosis; graphene oxide; mitochondrial membrane potential; oxidative stress
  6. Int J Mol Med. 2019 Jan 10.
    Yu H, Yuan B, Chu Q, Wang C, Bi H.
      By analyzing the L‑glutamic acid (L‑Glu)‑induced apoptosis of PC12 cells and an AlCl3 combined with D‑galactose (D‑gal)‑developed Alzheimer's disease (AD) mouse model, the protective effects of isoastilbin (IAB) against AD were systematically investigated in the present study. Pre‑incubation with IAB for 3 h prior to treatment with 25 mM L‑Glu decreased cell viability and inhibited apoptosis, suppressed the accumulation of intracellular reactive oxygen species, and restored mitochondrial membrane potential in PC12 cells induced by L‑Glu. In mice with AD, the reduced escape latency time in the water maze test, suppressed chronic movement in the center area of an open field test and enhanced ability to seek hidden food in a Y maze test indicated that abnormal behaviors had improved after 28 days of treatment with IAB. Furthermore, IAB reduced the deposition of amyloid β (Aβ) and the expression of phosphorylated‑Tau in the mouse brain and enhanced the serum levels of Aβ. IAB ameliorated the oxidative stress via modulating the levels of associated enzymes and improved the functioning of the central cholinergic system, as indicated by an increase in acetylcholine and choline acetyltransferase concentrations. The expression levels of acetylcholine esterase were reduced in the mouse brain in response to IAB pre‑treatment. In cells and brain tissue, IAB regulated the expression levels of pro‑ and anti‑apoptotic proteins and enhanced the nuclear levels of NF‑E2p45‑related factor 2 (Nrf2); subsequently, IAB further enhanced the expression of superoxide dismutase 1, catalase, and heme oxygenase‑1 and ‑2. The findings of the present study indicated that the protection of IAB against AD is at least partially associated with its antioxidation and anti‑apoptotic properties.
  7. J Cell Mol Med. 2019 Jan 22.
    Zhao X, Cai A, Peng Z, Liang W, Xi H, Li P, Chen G, Yu J, Chen L.
      As a nitric oxide (NO) donor prodrug, JS-K inhibits cancer cell proliferation, induces the differentiation of human leukaemia cells, and triggers apoptotic cell death in various cancer models. However, the anti-cancer effect of JS-K in gastric cancer has not been reported. In this study, we found that JS-K inhibited the proliferation of gastric cancer cells in vitro and in vivo and triggered mitochondrial apoptosis. Moreover, JS-K induced a significant accumulation of reactive oxygen species (ROS), and the clearance of ROS by antioxidant reagents reversed JS-K-induced toxicity in gastric cancer cells and subcutaneous xenografts. Although JS-K triggered significant NO release, NO scavenging had no effect on JS-K-induced toxicity in vivo and in vitro. Therefore, ROS, but not NO, mediated the anti-cancer effects of JS-K in gastric cancer. We also explored the potential mechanism of JS-K-induced ROS accumulation and found that JS-K significantly down-regulated the core proteins of mitochondria respiratory chain (MRC) complex I and IV, resulting in the reduction of MRC complex I and IV activity and the subsequent ROS production. Moreover, JS-K inhibited the expression of antioxidant enzymes, including copper-zinc-containing superoxide dismutase (SOD1) and catalase, which contributed to the decrease of antioxidant enzymes activity and the subsequent inhibition of ROS clearance. Therefore, JS-K may target MRC complex I and IV and antioxidant enzymes to exert ROS-dependent anti-cancer function, leading to the potential usage of JS-K in the prevention and treatment of gastric cancer.
    Keywords:  JS-K; apoptosis; gastric cancer; mitochondria respire chain complex; nitric oxide; reactive oxygen species
  8. Aquat Toxicol. 2019 Jan 15. pii: S0166-445X(18)31068-3. [Epub ahead of print]208 168-178
    Liu Q, Tang X, Wang Y, Yang Y, Zhang W, Zhao Y, Zhang X.
      As a newly emerging environmental contaminant, tributyl phosphate (TBP) is an additive flame retardant of high production volume that is frequently detected in biota and the environment. Despite evidence that TBP is a potential threat to marine organisms, ecotoxicology data for TBP in marine organisms at low trophic levels are scarce. In this study, the acute toxicological effect of TBP on the marine phytoplankton Phaeodactylum tricornutum was thoroughly investigated, and the possible mechanism was explored. The results showed that TBP at concentrations ≥0.2 mg L-1 significantly inhibited P. tricornutum growth in a clear dose-response manner, with 72-h EC10, EC20, EC50 and EC90 values of 0.067, 0.101, 0.219 and 0.716 mg L-1, respectively. Algal cells treated with TBP exhibited distorted shapes, ruptured cell membranes and damaged organelles, especially mitochondria. Additionally, apoptosis was triggered, followed by a decrease in mitochondrial membrane potential, indicating that cellular damage occurred during exposure. Although the activities of two antioxidant enzymes, superoxide peroxidase and catalase, were upregulated by TBP at 1.2 mg L-1, excess reactive oxygen species (ROS) and malondialdehyde still accumulated in algal cells after exposure, suggesting that the cells experienced oxidative stress. Moreover, both growth inhibition and apoptosis were positively correlated with ROS levels and were ameliorated by pretreatment with the ROS scavenger N-acetyl-l-cysteine. Taken together, the results indicate that TBP exposure leads to growth inhibition and cellular damage in P. tricornutum, and a ROS-mediated pathway might contribute to these observed toxicological effects.
    Keywords:  Acute toxicity; Apoptosis; Phaeodactylum tricornutum; Reactive oxygen species (ROS); Tributyl phosphate
  9. Int J Med Sci. 2019 ;16(1): 8-16
    Park C, Choi EO, Kim GY, Hwang HJ, Kim BW, Yoo YH, Park HT, Choi YH.
      Background: Due to its high antioxidant activity, baicalein, a kind of flavonoid present in Radical Scutellariae, has various pharmacological effects. However, the protective effect against oxidative stress in Schwann cells, which plays an important role in peripheral neuropathy, has not yet been studied. In this study, the effects of baicalein on hydrogen peroxide (H2O2)-induced DNA damage and apoptosis in RT4-D6P2T Schwann cells were evaluated. Methods: Cell viability assay was performed using MTT assay and colony formation assay. Apoptosis was assessed by flow cytometry analysis and DNA fragmentation assay. The effects on DNA damage and ATP content were analyzed by comet method and luminometer. In addition, changes in protein expression were observed by Western blotting. Results: Our results show that baicalein significantly inhibits H2O2-induced cytotoxicity through blocking reactive oxygen species (ROS) generation. We also demonstrate that baicalein is to block H2O2-induced DNA damage as evidenced by inhibition of DNA tail formation and γH2AX phosphorylation. Moreover, baicalein significantly attenuated H2O2-induced apoptosis and mitochondrial dysfunction, and restored inhibition of ATP production. The suppression of apoptosis by baicalein in H2O2-stimulated cells was associated with reduction of increased Bax/Bcl-2 ratio, activation of caspase-9 and -3, and degradation of poly (ADP-ribose) polymerase. Conclusions: These results demonstrate that baicalein eliminates H2O2-induced apoptosis through conservation of mitochondrial function by the removal of ROS. Therefore, it is suggested that baicalein protects Schwann cells from oxidative stress, and may be beneficial for the prevention and treatment of peripheral neuropathy induced by oxidative stress.
    Keywords:  Baicalein; DNA damage; Schwann cells; apoptosis; oxidative stress
  10. Phytomedicine. 2018 Oct 31. pii: S0944-7113(18)30556-7. [Epub ahead of print]56 136-146
    He L, Zhang Y, Kang N, Wang Y, Zhang Z, Zha Z, Yang S, Xu Q, Liu Y.
      BACKGROUND: Cisplatin is a highly effective chemotherapeutic agent commonly used in the treatment of a wide variety of malignancies. However, its clinical usage is severely limited by its serious side effects, especially nephrotoxicity. Anemoside B4, is a major saponins, rich in root of Pulsatilla chinensis (Bunge), has anti-inflammation in vitro. However, the antioxidant or anti-inflammatory effects of anemoside B4 in cisplatin-induced nephrotoxicity have not been clearly demonstrated.PURPOSE: In this study, we investigated whether anemoside B4 exhibits protective effects against cisplatin-induced nephrotoxicity involving antioxidant or anti-apoptosis effects.
    METHOD: To clarify it, the effects of anemoside B4 on HEK 293 cell viability was measured by CCK8 kits, intracellular antioxidant capacity including glutathione reduced (GSH), catalase (CAT) were estimated using chemical kits, apoptosis rate and intracellular reactive oxygen species (ROS) was analyzed by flow cytometry, apoptosis protein was measured by western blotting. In vivo model of cisplatin-induced mice acute renal failure was performed to evaluate the properties of anemoside B4. Besides, to evaluate the effect of anemoside B4 on the anti-tumor activity of cisplatin, S180 xenograft models were used.
    RESULTS: Anemoside B4 potently increased cisplatin-treated HEK 293T cells viability on the concentration and time manners and inhibited cells apoptosis, as demonstrated by the decreased cleaved PARP protein expressions. Anemoside B4 decreased reactive oxygen species (ROS) content and improved superoxide dismutase (SOD) activity. In vivo experiment showed that pretreatment with anemoside B4 effectively adjusted body weight and kidney index, and reduced cisplatin-elevated blood urea nitrogen (BUN) and creatinine (CREA) levels, as well as ameliorated the histopathological damage. Further studies showed that anemoside B4 did not reduce antitumor activity of cisplatin in murine S180 cancer xenograft tumor models. In addition, anemoside B4 per set showed low toxicity in mice.
    CONCLUSION: The strong antioxidant and anti-apoptosis effects of anemoside B4 may provide therapeutic potential for cisplatin-induced nephrotoxicity without compromising its therapeutic efficiency.
    Keywords:  Anemoside B4; Antioxidant; Apoptosis; Cisplatin; Nephrotoxicity
  11. Mol Med Rep. 2019 Jan 18.
    Yu N, Song N, Liu CY, Yang GL.
      Previous studies have demonstrated that ovariectomy may lead to a reduction in antioxidative biomarkers in the myocardium, thus suggesting that estrogens may serve a protective role in the suppression of oxidative stress. Lycium barbarum polysaccharides (LBP) are a well‑known antioxidant Chinese traditional medicine, which appear to have a similar function to estrogens with regards to the regulation of cardiac function. In the present study, 30 Sprague‑Dawley rats were randomly divided into the following groups: Sham operation group, ovariectomized (OVX) group, estradiol valerate group, high‑dose LBP (LBP‑H) group and low‑dose LBP (LBP‑L) group. All of the rats were provided tap water, estradiol valerate or LBP for 12 weeks. In addition, all rats were ovariectomized, with the exception of rats in the sham operation group, which underwent fat removal only. Reactive oxygen species (ROS), malondialdehyde (MDA), glutathione peroxidase (GSH‑px), catalase (CAT) and superoxide dismutase activities were subsequently examined. The protein expression levels of cleaved caspase‑9, cleaved caspase‑3 and phosphorylated‑protein kinase B (p‑Akt) were also assessed. The results demonstrated that high‑dose LBP decreased the enhanced levels of ROS and MDA in OVX rats, whereas GSH‑px and CAT activities were increased in the LBP‑H group compared with in OVX rats. Furthermore, the expression levels of cleaved caspase‑9 and cleaved caspase‑3 were significantly upregulated in the OVX group, whereas high‑dose LBP exerted protective effects on OVX rats by decreasing the expression of apoptotic proteins. Conversely, p‑Akt expression was decreased in the OVX group and was increased in the LBP‑H group. These results indicated that LBP is essentially involved in cardiac protection by inhibiting apoptosis in response to oxidative stress. In addition, improvement of antioxidant status by LBP is associated with the Akt signaling pathway in the myocardium of OVX rats.
  12. Theranostics. 2019 ;9(1): 167-178
    Chen Y, Li N, Wang J, Zhang X, Pan W, Yu L, Tang B.
      Radiotherapy is an extensively used treatment modality in the clinic and can kill malignant cells by generating cytotoxic reactive oxygen species (ROS). Unfortunately, excessive dosages of radiation are typically required because only a small proportion of the radiative energy is adsorbed by the soft tissues of a tumor, which results in the nonselective killing of normal cells and severe systemic side effects. An efficient nanosensitizer that makes cancer cells more sensitive to radiotherapy under a relatively low radiation dose would be highly desirable. Methods: In this study, we developed a Gd-doped titania nanosensitizer that targets mitochondria to achieve efficient radiotherapy. Upon X-ray irradiation, the nanosensitizer triggers a "domino effect" of ROS accumulation in mitochondria. This overabundance of ROS leads to mitochondrial permeability transition and ultimately irreversible cell apoptosis. Confocal laser imaging, western blotting and flow cytometry analysis were used to explore the biological process of intrinsic apoptosis induced by the nanosensitizer. Clonogenic survival assay, cell migration and invasion experiments were employed to evaluate the radiosensitizing effect of the nanosensitizer in vitro. Finally, to evaluate the therapeutic outcome of the nanosensitizer in vivo, MCF-7 tumor model was used. Results: Confocal laser images and western blotting data demonstrated that the nanosensitizer in conjunction with X-ray irradiation could induce cell apoptosis in ROS-mediated apoptotic signal pathways. A clonogenic survival assay revealed that cells treated with the prepared nanosensitizer exhibited a lower number of viable cell colonies than that of the nontargeted group under X-ray irradiation. Notably, with only a single dose of radiotherapy, the mitochondria-targeted nanosensitizer elicited the complete ablation of tumors in a mouse model. Conclusion: The designed nanosensitizer in combination with X-ray radiation exposure could be used for radiotherapy against cancer in living cells and in vivo. Moreover, the nanosensitizer with mitochondria targeting played a pivotal role in triggering a "domino effect" of ROS and cell apoptosis. The current strategy could provide new opportunities in designing efficient radiosensitizers for future cancer therapy.
    Keywords:  cancer; mitochondria-targeting; nanosensitizer; radiotherapy; reactive oxygen species
  13. Phytomedicine. 2018 Sep 05. pii: S0944-7113(18)30312-X. [Epub ahead of print]53 171-181
    Wu H, Gao H, Gao S, Lei Z, Dai L, Wang X, Han Y, Wang Z, Han L.
      BACKGROUND: Although the protective effects of Yiqi-Huoxue granule (YQHX), a Chinese 4-herb formula, on patients with ischemic heart diseases are related to the attenuation of oxidative stress injury, the mechanism(s) underlying these actions remains poorly understood.PURPOSE: Our aim was to investigate the potential protective effects of YQHX treatment against oxidative stress induced by hydrogen peroxide (H2O2) in rat H9c2 cells.
    METHODS: H9c2 cells were treated with YQHX for 16 h before exposed to 200 μM H2O2 for 6 h. The apoptosis induced by H2O2 was measured using hoechst 33,342 staining and Annexin-V FITC/PI assay. The expression of uncoupling protein 2 (UCP2), Bcl-2, Bax, and caspase-3 were observed using western blot. The effects of UCP2 knockdown on cell apoptosis and intracellular ROS production were also investigated.
    RESULTS: H2O2 exposure led to significant activation of oxidative stress followed by increased apoptosis and ROS production, as well as decreased UCP2 expression in H9c2 cells. YQHX treatment at the concentration of 0.75 and 1.5 mg/ml remarkably reduced the expression of Bax and caspase-3, whereas increased the protein expression of Bcl-2 and UCP2. These changes were attenuated by transgenic knockdown of UCP2 with Lenti-shUCP2 vector.
    CONCLUSIONS: Taken together, our study demonstrated that YQHX attenuates H2O2-induced apoptosis by upregulating UCP2 expression in H9c2 Cells, suggesting that YQHX is a promising therapeutic approach for the treatment of I/R injury-mediated apoptosis.
    Keywords:  Cardiomyocyte apoptosis; Oxidative stress; Uncoupling protein 2 (UCP2); Yiqi-Huoxue granule (YQHX)
  14. Phytomedicine. 2018 Dec 11. pii: S0944-7113(18)30606-8. [Epub ahead of print]57 117-128
    Wang L, Liu Y, Zhao TL, Li ZZ, He JY, Zhang BJ, Du HZ, Jiang JW, Yuan ST, Sun L.
      BACKGROUND: Topotecan (TPT) is a Topo I inhibitor and shows obvious anti-cancer effects on gastric cancer. Cancer cells reprogram their metabolic pathways to increase nutrients uptake, which has already been a hallmark of cancer. But the effect of TPT on metabolism in gastric cancer remains unknown.PURPOSE: To investigate the effect of TPT on metabolism in gastric cancer.
    METHODS: ATP production was measured by ATP Assay kit. Glucose and glutamine uptake were measured by Glucose (HK) Assay Kit and Glutamine/Glutamate Determination Kit respectively. To detect glutathione (GSH) concentration and reactive oxygen species (ROS) generation, GSH and GSSG Assay Kit and ROS Assay Kit were adopted. Apoptosis rates, mitochondrial membrane potential (MMP) were determined by flow cytometry and protein levels were analyzed by immumohistochemical staining and western blotting.
    RESULTS: TPT increased ATP production. TPT promoted glucose uptake possibly via up-regulation of hexokinase 2 (HK2) or glucose transporter 1 (GLUT1) expression, while decreased glutamine uptake by down-regulation of ASCT2 expression. ASCT2 inhibitor GPNA and ASCT2 knockdown significantly suppressed the growth of gastric cancer cells. Inhibition of ASCT2 reduced glutamine uptake which led to decreased production of GSH and increased ROS level. ASCT2 knockdown induced apoptosis via the mitochondrial pathway and weakened anti-cancer effect of TPT.
    CONCLUSION: TPT inhibits glutamine uptake via down-regulation of ASCT2 which causes oxidative stress and induces apoptosis through the mitochondrial pathway. Moreover, TPT inhibits proliferation partially via ASCT2. These observations reveal a previously undescribed mechanism of ASCT2 regulated gastric cancer proliferation and demonstrate ASCT2 is a potential anti-cancer target of TPT.
    Keywords:  ASCT2; Apoptosis; Gastric cancer; Oxidative stress; Topotecan
  15. Biochim Biophys Acta Mol Basis Dis. 2019 Jan 16. pii: S0925-4439(19)30016-X. [Epub ahead of print]
    Mao M, Zhang T, Wang Z, Wang H, Xu J, Yin F, Wang G, Sun M, Wang Z, Hua Y, Cai Z.
      Osteosarcoma (OS) is ranked as the most common primary bone malignancy in children and adolescents worldwide, and the 5-year overall survival rate of OS is not optimistic. Constitutive activation of signal transducer and activator of transcription 3 (STAT3) has been implicated in tumor cell growth, proliferation, and anti-apoptosis in OS. Therefore, the discovery of novel molecular compounds that can effectively block STAT3 activation, is essential for the treatment of OS and improving prognosis. Here, we investigate whether Glaucocalyxin A (GLA), derived from Rabdosia japonica, exhibit the potential anticancer effects in OS. First of all, we identify that GLA potently suppressed cell proliferation, induced G2/M phase arrest and promoted substantial apoptosis in OS. Next, we conclude that GLA could induce Reactive oxygen species (ROS)-mediated oxidative stress via an imbalance of GSH and GSSG. Then, we elucidate for the first time that GLA could significantly inhibit both constitutive and IL-6-inducible activation of STAT3 (Tyr705) and JAK2, the upstream regulator of STAT3. Furthermore, we elucidate that the inhibition of STAT3 is mainly induced by ROS-mediated oxidative stress. Overall, our findings demonstrate that GLA could exhibit potent anticancer effects through effectively blocking the STAT3 signaling pathway, which was induced by ROS-mediated oxidative stress in OS in vitro and in vivo.
    Keywords:  Glaucocalyxin A; Osteosarcoma; Oxidative stress; ROS; STAT3
  16. Food Funct. 2019 Jan 21.
    Wang L, Li C, Huang Q, Fu X.
      Defective glucose-stimulated insulin secretion (GSIS) induced by chronic exposure to reactive oxygen species (ROS) is a hallmark of type 2 diabetes mellitus (T2DM). Therefore, it is of great interest to search for biofunctional agents with antioxidant activity to protect pancreatic islet cells from oxidative damage. In the present study, selenium nanoparticles (SeNPs) functionalized with a novel polysaccharide (RTFP-3) extracted from Rosa roxburghii fruit were first prepared via a facile, single-step and green in situ synthesis method. The in vitro protective effects of RP3-SeNPs on INS-1 cells against H2O2-induced cell apoptosis were investigated. Structural characterization indicated that RTFP-3-functionalized SeNPs (RP3-SeNPs) with an average diameter of 104.5 nm were highly uniform and extremely stable in comparison with bare SeNPs. The results of bioassays revealed that RP3-SeNPs possessed much higher protective and suppressive activities against H2O2-induced apoptosis of INS-1 cells in comparison with their individual components. After treatment with an RP3-SeNPs solution (2 μg mL-1), the cell viability of INS-1 cells reached about 89.34%. Mechanistic studies demonstrated that RP3-SeNPs effectively blocked the overproduction of intracellular ROS, mitochondrial damage, and the activation of caspase-3, caspase-8, and caspase-9 in INS-1 cells, which indicated that RP3-SeNPs functioned via attenuating oxidative stress and downregulating the expression of uncoupling protein-2 (UCP-2). Our findings suggest that RP3-SeNPs can function as a promising candidate to prevent or limit the dysfunction of β-cells.
  17. Exp Mol Pathol. 2019 Jan 17. pii: S0014-4800(18)30277-6. [Epub ahead of print]
    Greenshields AL, Fernando W, Hoskin DW.
      Breast cancer is the most prevalent cancer diagnosis in women, with triple-negative and human epidermal growth factor 2 (HER2)-enriched advanced breast cancers having the poorest prognoses. The morbidity and mortality associated with advanced disease, as well as the emergence of multi-drug resistant variants, highlights the urgency to develop novel therapeutic agents. Artesunate (ART) is a semi-synthetic derivative of artemisinin from the Chinese herb sweet wormwood. ART is widely used in the treatment of malaria and is well tolerated by patients. Importantly, ART also has anti-cancer activities and may therefore represent a less toxic alternative to conventional chemotherapy. In this study, we demonstrate a dose- and time-dependent inhibitory effect of ART on the growth of triple-negative MDA-MB-468 and HER2-enriched SK-BR-3 breast cancer cells, which was the result of both anti-proliferative and cytotoxic activities. ART inhibited breast cancer cell proliferation via a reactive oxygen species (ROS)-dependent G2/M arrest and ROS-independent G1 arrest. ART-treated MDA-MB-468 and SK-BR-3 cells also experienced apoptotic cell death, which was both ROS- and iron-dependent. ART-induced oxidative stress caused the loss of mitochondrial outer membrane integrity and damage to the cellular DNA of MDA-MB-468 and SK-BR-3 cells. In addition, exposure to low-dose ART sensitized MDA-MB-468 and SK-BR-3 cells to chemotherapeutic drugs. On the basis of our findings, we suggest that ART may have clinical utility in the treatment of triple-negative and HER2-enriched breast cancers.
    Keywords:  Artesunate; Cell cycle arrest; Cytotoxicity, breast cancer, reactive oxygen species
  18. Int J Biochem Cell Biol. 2019 Jan 17. pii: S1357-2725(19)30016-0. [Epub ahead of print]
    Li E, Sun Y, Lv G, Li Y, Zhang Z, Hu Z, Cao W.
      Sonodynamic therapy (SDT) is a noninvasive therapeutic method via the activation of certain chemical sensitizers using low intensity ultrasound. In this work, we evaluated the antitumor effect of sinoporphyrin sodium (DVDMS) mediated SDT (DVDMS-SDT) on Hepatocellular carcinoma (HCC) cell lines both in vitro and in vivo. The results indicated that DVDMS-SDT was significantly more efficacious than PpIX-SDT in treating hepatocellular cell line Hep-G2. DVDMS-SDT also increased the ratio of cells in the G2/M phase and decreased the CDK1 and Cyclin B1 protein level. DVDMS-SDT markedly increased intracellular reactive oxygen species (ROS) in vitro. The increased ROS production up-regulated the expression of p53 and Bax, and down-regulated Bcl-2 expression, which led to the activation of caspase-3, ultimately initiated cell apoptosis. These effects could be partially reversed by the ROS scavenger N-acetylcysteine (NAC). In vivo experiments revealed that the DVDMS-SDT resulted in an effective inhibition of tumor growth and prolonged the survival time of tumor-bearing mice. More importantly, no obvious signs of side effects were observed. These results suggested that DVDMS-SDT is very effective in treating Hepatocellular carcinoma without side effects. The primary mechanism of SDT is due to the increased ROS activated the p53/Caspase 3 axis of apoptosis.
    Keywords:  Apptosis; Hepatocellular carcinoma; Reactive oxygen species; Sinoporphyrin sodium; Sonodynamic therapy
  19. Nanomedicine (Lond). 2019 Jan 24.
    Roy S, Sarkar A, Jaiswal A.
      AIM: To develop near-infrared (NIR) light-responsive reduced graphene oxide (RGO)-based nanocomposites with improved stability, biocompatibility and enhanced in vitro chemo-photothermal therapeutic efficiency.MATERIALS & METHODS: Poly(allylamine hydrochloride)-functionalized RGO-based nanocomposites (RGO-PAH) were synthesized and thoroughly characterized. In vitro biocompatibility, cellular uptake and in vitro synergistic chemo-photothermal therapeutic efficiency of drug-loaded RGO-PAH nanocomposites were evaluated along with elucidation of cell death mechanism.
    RESULTS: RGO-PAH nanocomposites showed excellent photothermal transduction, pH-dependent drug release, rapid internalization, high biocompatibility and highly efficient synergistic in vitro chemo-photothermal therapy via apoptosis induction through increase in intracellular reactive oxygen species (ROS) production followed by oxidative DNA damage.
    CONCLUSION: Excellent biocompatibility and highly efficient chemo-photothermal killing of cancer cells at a very low concentration reflects the potential of RGO-PAH as a NIR-responsive therapeutic agent for cancer therapy.
    Keywords:  DNA fragmentation; apoptosis; doxorubicin; near-infrared light; poly(allylamine hydrochloride); reactive oxygen species; reduced graphene oxide; synergistic chemo-photothermal therapy
  20. Mater Sci Eng C Mater Biol Appl. 2019 Apr;pii: S0928-4931(18)31302-X. [Epub ahead of print]97 467-478
    Dhanwal V, Katoch A, Singh A, Chakraborty S, Faheem MM, Kaur G, Nayak D, Singh N, Goswami A, Kaur N.
      Organic nanoparticles (ONPs) possess great research interests for their promising effects in the enhancement of bioactivity including anticancer activity with less toxicity. The present study describes the preparation, characterization and biological evaluation of aqueous phase ONPs of potent 1,2-disubstituted benzimidazole derivative (BZ6) for anticancer activity. BZ6-ONPs were characterized through UV-absorption and fluorescence spectroscopic analysis for their photo-physical properties. DLS, TEM and SEM studies were carried out for morphological and structural analysis. Cytotoxicity determination on a panel of four different cancer cell lines (MCF-7, MiaPaca-2, HT-29 and HCT-116) revealed that the BZ6-ONPs show highest activity in human breast cancer MCF-7 cells. Surprisingly, the BZ6-ONPs were found to be non-toxic towards normal breast epithelial fR2 cells. Additionally, the FITC-ONPs showed enhanced uptake in 3D tumor spheroids of MCF-7 cells compared to the free FITC. BZ6-ONPs strongly halted cell proliferation and induced apoptosis, possibly through oxidative stress-mediated reactive oxygen species (ROS) generation and loss of mitochondrial membrane potential (MMP) in MCF-7 cells. Moreover, molecular mechanism-based studies revealed that BZ6-ONPs downregulated AKT/NF-κB/vimentin/survivin-mediated oncogenic signaling pathway promoting cell proliferation and malignancy. In a nutshell, BZ6-ONPs are therapeutically efficacious, which needs further development as a treatment option in human mammary gland carcinomas.
    Keywords:  3D spheroid uptake; Apoptosis; Benzimidazole; Breast cancer; Organic nanoparticles; Reactive oxygen species (ROS)
  21. J Pharm Investig. 2018 ;48(2): 143-151
    Hwang HS, Shin H, Han J, Na K.
      Photodynamic therapy (PDT) is performed using a photosensitizer and light of specific wavelength in the presence of oxygen to generate singlet oxygen and reactive oxygen species(ROS) in the cancer cells. The accumulated photosensitizers in target sites induce ROS generation upon light activation, then the generated cytotoxic reactive oxygen species lead to tumor cell death via apoptosis or necrosis, and damages the target sites which results tumor destruction. As a consequence, the PDT-mediated cell death is associated with anti-tumor immune response. In this paper, the effects of PDT and immune response on tumors are reviewed. Activation of an immune response regarding the innate and adaptive immune response, interaction with immune cells and tumor cells that associated with antitumor efficacy of PDT are also discussed.
    Keywords:  Antitumor immune response; Cancer; Photodynamic therapy; Reactive oxygen species
  22. Mol Med Rep. 2019 Jan 15.
    Ma Y, Xiong L.
      Endoplasmic reticulum stress (ERS) serves a vital role in the pathological development of Alzheimer's disease (AD). ERS can promote programmed cell death (apoptosis) during AD; however, the specific molecular mechanisms that lead to ERS remain unclear. It is very important that a drug for the treatment of AD is identified. Our previous studies indicated that astragaloside IV (AST IV) has anti‑inflammatory effects and helps cells resist oxidative stress. In the present study, western blotting and reverse transcription semi‑quantitative polymerase chain reaction were used to detect protein and mRNA expression levels, flow cytometry was used to measure intracellular reactive oxygen species (ROS) levels, and superoxide dismutase (SOD) and malondialdehyde (MDA) activity was detected using commercially available kits. The results demonstrated that SOD activity was decreased, and MDA content, ROS levels, and the expression levels of p38 mitogen‑activated protein kinase (MAPK) and ERS‑associated proteins, including binding immunoglobulin protein/glucose‑regulated protein and growth arrest‑ and DNA damage ‑inducible gene 153/C/EBP homologous protein, were increased in amyloid β (Aβ)25‑35‑treated PC12 cells. Furthermore, to investigate the role of p38 MAPK and the effects of AST IV in an in vitro model of AD, SB203580, a p38 MAPK signaling pathway inhibitor, and AST IV were administered to Aβ25‑35‑treated PC12 cells. The results revealed that AST IV protected the cells against AD. This effect may be caused by decreases in ROS levels, which may inhibit the p38 MAPK signaling pathway and thereby suppress ERS in Aβ25‑35‑treated PC12 cells.
  23. Cancer Lett. 2019 Jan 17. pii: S0304-3835(19)30023-0. [Epub ahead of print]
    Jeong S, Yun HK, Jeong YA, Jo MJ, Kang SH, Kim JL, Kim DY, Park SH, Kim BR, Na YJ, Lee SI, Kim HD, Kim DH, Oh SC, Lee DH.
      Cannabidiol (CBD), one of the compounds present in the marijuana plant, has anti-tumor properties, but its mechanism is not well known. This study aimed to evaluate the apoptotic action of CBD in colorectal cancer (CRC) cells, and focused on its effects on the novel pro-apoptotic Noxa-reactive oxygen species (ROS) signaling pathway. CBD experiments were performed using the CRC cell lines HCT116 and DLD-1. CBD induced apoptosis by regulating many pro- and anti-apoptotic proteins, of which Noxa showed significantly higher expression. To understand the relationship between Noxa and CBD-induced apoptosis, Noxa levels were downregulated using siRNA, and the expression of apoptosis markers decreased. After ROS production was blocked, the level of Noxa also decreased, suggesting that ROS is involved in the regulation of Noxa, which along with ROS is a well-known pro-apoptotic signaling agents. As a result, CBD induced apoptosis in a Noxa-and-ROS-dependent manner. Taken together, the results obtained in this study re-demonstrated the effects of CBD treatment in vivo, thus confirming its role as a novel, reliable anticancer drug.
    Keywords:  Bcl-2 protein family; Marijuana extract; ROS; apoptotic cell death; colon cancer
  24. Cell Transplant. 2019 Jan 22. 963689718824559
    Li F, Yang Z, Stone C, Ding JY, Previch L, Shen J, Ji Y, Geng X, Ding Y.
      In vitro liver conservation is an issue of ongoing critical importance in graft transplantation. In this study, we investigated the possibility of augmenting the standard pre-transplant liver conservation protocol (University of Wisconsin (UW) cold solution) with the phenothiazines chlorpromazine and promethazine. Livers from male Sprague-Dawley rats were preserved either in UW solution alone, or in UW solution plus either 2.4, 3.6, or 4.8 mg chlorpromazine and promethazine (C+P, 1:1). The extent of liver injury following preservation was determined by alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, the ratio of AST/ALT, morphological changes as assessed by hematoxylin-eosin staining, apoptotic cell death as determined by ELISA, and by expression of the apoptotic regulatory proteins BAX and Bcl-2. Levels of glucose (GLU) and lactate dehydrogenase (LDH) in the preservation liquid were determined at 3, 12, and 24 h after incubation to assess glucose metabolism. Oxidative stress was assessed by levels of superoxide dismutase (SOD), reactive oxygen species (ROS), and malondialdehyde (MDA), and inflammatory cytokine expression was evaluated with Western blotting. C+P augmentation induced significant reductions in ALT and AST activities; the AST/ALT ratio; as well as in cellular swelling, vacuolar degeneration, apoptosis, and BAX expression. These changes were associated with lowered levels of GLU and LDH; decreased expression of SOD, MDA, ROS, TNF-α, and IL-1β; and increased expression of Bcl-2. We conclude that C+P augments hypothermic preservation of liver tissue by protecting hepatocytes from ischemia-induced oxidative stress and metabolic dysfunction. This result provides a basis for improvement of the current preservation strategy, and thus for the development of a more effective graft conservation method.
    Keywords:  graft; hypothermic preservation; liver conservation; organ transplantation; oxidative injury
  25. Chem Biol Interact. 2019 Jan 17. pii: S0009-2797(18)31633-8. [Epub ahead of print]
    Yu J, Sun W, Song Y, Liu J, Xue F, Gong K, Yang X, Kang Q.
      Oxidative stress-induced damage of retinal ganglion cells (RGCs) is a major contributor to retinal degenerative diseases, such as glaucoma. Sirtuin 6 (SIRT6) has emerged as a cytoprotective protein against various insults. However, whether SIRT6 exerts a protective effect against oxidative stress-damaged RGCs remains unknown. In this study, we aimed to investigate the potential role and regulatory mechanism of SIRT6 in hydrogen peroxide (H2O2)-induced oxidative damage of RGCs in vitro. We found that SIRT6 expression was significantly downregulated in RGCs with H2O2 treatment. Functional experiments showed that overexpression of SIRT6 improved survival and reduced apoptosis and the production of reactive oxygen species (ROS) in H2O2-treated RGCs. In contrast, SIRT6 knockdown had the opposite effect. Moreover, we found that SIRT6 overexpression promoted the nuclear accumulation of nuclear factor erythroid 2-related factor 2 (Nrf2) and increased the activity of antioxidant response element (ARE). In addition, we found that the promotional effect of SIRT6 on Nrf2/ARE signaling was associated with inhibition of BTB and CNC homology 1 (Bach1), an inhibitor of Nrf2. However, overexpression of Bach1 or inhibition of Nrf2/ARE signaling partially reversed the SIRT6-mediated protective effect. Taken together, these results demonstrate that SIRT6 protects RGCs from oxidative stress-induced damage by promoting the activation of Nrf2/ARE signaling via inhibition of Bach1, suggesting a potential role of SIRT6 in retinal degenerative diseases.
    Keywords:  Bach1; Nrf2; Retinal ganglion cells; SIRT6
  26. Chemosphere. 2019 Jan 16. pii: S0045-6535(19)30074-8. [Epub ahead of print]221 693-702
    Yu X, Yin H, Peng H, Lu G, Liu Z, Dang Z.
      Organophosphate flame retardants (OPFRs) and brominated flame retardants (BFRs) are frequently detected in indoor environment at high levels, posing health risks to humans. However, the potential cytotoxicity mediated by OPFRs and BFRs in relevant human cell models is limited. In current study, non-small cell lung cancer A549 cell was employed to investigate toxicity mechanisms of typical OPFRs (i.e., tris (2-chloroethyl) phosphate (TCEP), tris-(2-chloropropyl) phosphate (TCPP), tricresy phosphate (TCP), triphenyl phosphate (TPHP) and BFRs (i.e., 2,2',4,4'-tetrabromodiphenyl ether (BDE-47), 3,3', 5,5'-tetrabromobisphenol A (TBBPA)). It was found that BDE-47 exhibited the strongest cytotoxicity, followed by TBBPA, TPHP, TCP, TCPP and TCEP. OPFRs and BFRs could cause the reduction of cell viability of A549 cell in both dose- and time-dependent manner after exposure for 24 and 48 h. Simultaneously, excessive generation of reactive oxygen species (ROS), mitochondrial membrane potential (MMP) dysfunction, cell apoptosis and overload of intracellular free Ca2+ demonstrated that cytotoxicity induced by OPFRs and BFRs were mediated by oxidative stress. Of note, the survival rate of cell significantly increased when pretreated with Ac-DEVD-CHO, suggesting that caspase-3 dependent mitochondrial pathway may have played a primary role in the process of A549 cell apoptosis.
    Keywords:  BFRs; Caspase-dependent pathway; Cell apoptosis; MMP dysfunction; OPFRs
  27. Mol Med Rep. 2019 Jan 14.
    Zuo Y, Hu J, Xu X, Gao X, Wang Y, Zhu S.
      Sodium azide (NaN3), an inhibitor of cytochrome oxidase, induces the release of excitotoxins via an energy impairment and this, in turn, results in neurodegeneration. The present study aimed to investigate the toxic effects NaN3 on apoptosis of PC12 cells and its mechanism of action in peroxisome proliferator‑activated receptor γ co‑activator 1‑α (Pgc‑1α)‑associated signaling pathways. To induce apoptosis, PC12 cells were exposed to NaN3 (0, 5, 10, 20, 40 and 80 mM) for 12, 24, 48 and 72 h. Cell viability was determined by CCK‑8 assay. DAPI staining was employed to additionally examine apoptotic cells and their nuclear changes. Production of reactive oxygen species (ROS), mitochondrial membrane potential (ΔΨm) and apoptotic rate were also assessed by flow cytometry. Cellular ATP content was estimated by firefly luciferase assay. In addition, the expression levels of B‑cell lymphoma 2 (Bcl‑2), Bcl‑2‑associated X protein (Bax), phosphorylated (p)‑Ca2+/calmodulin‑dependent protein kinase (CaMK), p‑p38 mitogen‑activated protein kinase (p38 MAPK), Pgc‑1α, nuclear respiratory factor (Nrf)‑1, mitochondrial transcription factor A (Tfam), p‑extracellular signal‑regulated kinase (Erk)1/2, Nrf‑2 and complex IV (Cox IV) were determined by western blot analysis. The data suggested that NaN3 may induce PC12 cell injury and dose‑dependently decrease the cell viability. The expression levels of pro‑apoptotic proteins Bax and cytochrome c were upregulated, while the expression levels of anti‑apoptotic proteins procaspase‑3 and Bcl‑2 were downregulated. In addition, the phosphorylation of MAPK and Ca2+/calmodulin‑dependent protein kinase II (CaMKⅡ) family members including pan‑calcineurin A was increased, in particular the ratios of p‑p38/p38 and p‑CaMKⅡ/CaMKⅡ. However, the expression levels of Pgc‑1α and its associated proteins, including Nrf‑1/2, Tfam and p‑Erk1/2 were decreased. In addition, mitochondria were the target organelles of NaN3‑induced toxicity in PC12 cells, which moderated the dissipation of ΔΨm, preserved the cellular ATP content, promoted the production of ROS and increased the apoptotic rate. These results suggested that NaN3 induced cell death in PC12 cells via Pgc‑1α‑associated signaling pathways and provided a theoretical basis for additional investigation of the neurotoxic mechanism of NaN3, with applications in neurodegenerative disorders.
  28. Oxid Med Cell Longev. 2018 ;2018 8938207
    Ma Y, Ma B, Shang Y, Yin Q, Wang D, Xu S, Hong Y, Hou X, Liu X.
      Aging is a major factor that contributes to neurological impairment and neuropathological changes, such as inflammation, oxidative stress, neuronal apoptosis, and synaptic dysfunction. Flavonoids act as protective antioxidant and anti-inflammatory agents against various age-related neurodegenerative diseases. Here, we investigated the protective effect and mechanisms of the flavonoid-rich ethanol extract from the leaves of Diospyros kaki (FELDK) in the cortex and hippocampus of D-galactose- (gal-) aged mice. Our results showed that FELDK treatment restored memory impairment in mice as determined by the Y-maze and Morris water maze tests. FELDK decreased oxidative stress levels via inhibiting reactive oxygen species (ROS) and malondialdehyde (MDA) production and elevating antioxidative enzymes. FELDK also alleviated D-gal-induced neuroinflammation via suppressing the expression of advanced glycation end products (AGEs) and receptor for AGEs (RAGE) and activating microgliosis and astrocytosis, nuclear factor kappa B (NF-κB) nuclear translocation, and downstream inflammatory mediators. Moreover, FELDK inhibited the phosphatidylinositol 3-kinase (PI3K)/Akt and C-jun N-terminal kinase (JNK) apoptotic signaling pathways and ameliorated the impairment of synapse-related proteins. Hence, these results indicate that FELDK exerts neuroprotective effects on D-gal-induced brain aging. Thus, FELDK may be a potential therapeutic strategy for preventing and treating age-related neurodegenerative diseases such as Alzheimer's disease.
  29. J Cell Physiol. 2019 Jan 19.
    Huang YT, Liu CH, Yang YC, Aneja R, Wen SY, Huang CY, Kuo WW.
      The prevalence of chronic hyperglycemia and its complications, imposing a critical burden on the worldwide economy and the global healthcare system, is a pressing issue. Mounting evidence indicates that oxidative stress and hypoxia, two noticeable features of hyperglycemia, play a joint crucial role in mediating cellular apoptosis. However, the underlying detailed molecular mechanism remains elusive. Triggered by the observation that insulin-like growth factor (IGF1)-binding protein 3 (IGFBP3) can mediate, in renal cells, high-glucose-induced apoptosis by elevating oxidative stress, we wish to, in this study, know whether or not the similar scenario holds in cardiac cells and, if so, to find its relevant molecular key players, thereby dissecting the underlying molecular pathway. Specifically, we used a combination of three different cellular sources (H9c2 cells, diabetic rats, and neonatal rat ventricular cardiomyocytes) as our model systems of study. We made use of Co-IP assay and western blot analysis in conjunction with loss-of-function reasoning, gain-of-function logic, and inhibitor treatment as our main analytical tools. As a result, briefly, our main findings are that hyperglycemia can induce cardiac IGFBP3 overexpression and secretion, that high levels of IGFBP3 can sequester IGF1 from IGF1 survival pathway, leading to apoptosis, and that IGFBP3 gene upregulation is hypoxia-inducible factor (HIF)1α-dependent and reactive oxygen species dependent. Piecing these findings together allows us to propose the improved molecular regulatory mechanism. In conclusion, we have established the molecular roles of IGFBP3, HIF1, and prolyl hydroxylase domain in connecting oxidative stress with hypoxia and in cellular apoptosis under hyperglycemia.
    Keywords:  HIF1α; PHD; PI3K/Akt; diabetic cardiomyopathy; hyperglycemia; hypoxia
  30. J Mol Neurosci. 2019 Jan 25.
    Li Q, Wang P, Huang C, Chen B, Liu J, Zhao M, Zhao J.
      In this study, with primary mouse neural progenitor cells (NPCs), we investigated the neuroprotective effect of a tropomyosin-related kinase receptor B (TrkB) agonist, N-acetyl serotonin (NAS), against hydrogen peroxide (H2O2)-induced toxicity. We found that pre-incubation with NAS not only ameliorates H2O2-induced cell viability loss, lactate dehydrogenase (LDH) release, and proliferative and migratory capacity impairments, but counteracts H2O2-triggered production of nitric oxide (NO), reactive oxygen species (ROS), malondialdehyde (MDA), and 8-hydroxy-deoxyguanosine (8-OHdG) in a dose-dependent manner. Additionally, pre-treatment with NAS was able to attenuate H2O2-induced apoptosis in NPCs, evidenced by the decreased percentage of apoptotic cells and altered expression of apoptosis-related factors. Furthermore, in differentiated NPCs, NAS improves H2O2-induced reduction in neurite growth. Mechanistic studies revealed that the protective effects of NAS in NPCs may be mediated by the TrkB/PI3K/Akt/ cAMP response element binding protein (CREB) signaling cascades. In a mouse traumatic brain injury (TBI) model, we found that systemic administration of 30 mg/kg NAS could improve hippocampal neurogenesis, manifested by the increased number of SOX-2-positive cells and increased expression of phosphorylated CREB in the dentate gyrus (DG) area. Treatment with NAS also ameliorates cognitive impairments caused by TBI, as assessed by Y-maze and contextual and cued fear conditioning tests. Taken together, these results provide valuable insights into the neuroprotective and neuroregenerative effects of NAS, suggesting it may have therapeutic potential for the treatment of TBI.
    Keywords:  Apoptosis; Hippocampal neurogenesis; N-acetyl serotonin; Neural progenitor cell; Oxidative stress; Traumatic brain injury
  31. Molecules. 2019 Jan 23. pii: E407. [Epub ahead of print]24(3):
    Sibiya MA, Raphoko L, Mangokoana D, Makola R, Nxumalo W, Matsebatlela TM.
      Despite major advancements in the development of various chemotherapeutic agents, treatment for lung cancer remains costly, ineffective, toxic to normal non-cancerous cells, and still hampered by a high level of remissions. A novel cohort of quinoxaline derivatives designed to possess a wide spectrum of biological activities was synthesized with promising targeted and selective anticancer drug activity. Hence, this study was aimed at determining in vitro anticancer activity effects of a newly synthesized class of 3-(quinoxaline-3-yl) prop-2-ynyl quinoxaline derivatives on A549 lung cancer cells. An assessment of the quinoxaline derivatives ferric reducing power, free radical scavenging activity, cytotoxic activity, and ability to induce reactive oxygen species (ROS) production was performed using the Ferric Reducing Antioxidant Power (FRAP), 2,2-diphenyl-1-picryl-hydrazyl (DPPH), 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and 2',7'-dichlorodihydrofluorescein diacetate (H₂DCFDA) assays, respectively. The ability of the quinoxaline derivatives to induce apoptosis in A549 cells was assessed using the Acridine Orange/Ethidium Bromide (AO/EB) and Annexin V-FITC/Dead Cell Assay. Of the four quinoxaline derivatives tested, 3-(quinoxaline-3-yl) prop-2-ynyl methanosulphate (LA-39B) and 3-(quinoxaline-3-yl) prop-2-yn-1-ol (LA-55) displayed a dose-dependent reducing power, free-radical scavenging activity, inhibition of cell viability, and stimulation of ROS production which was accompanied by induction of apoptosis in A549 lung cancer cells. None of the quinoxaline derivatives induced cell death or ROS production in non-cancerous Raw 267.4 macrophage cells. Cytotoxicity was observed in A549 lung cancer, HeLa cervical cancer, and MCF-7 breast cancer cells albeit inhibition was more pronounced in A549 cells. The results of the study suggest that 3-(quinoxaline-3-yl) prop-2-ynyl methanosulphate and 3-(quinoxaline-3-yl) prop-2-yn-1-ol induce apoptotic cell death in A549 lung cancer cells.
    Keywords:  anticancer; antioxidant; apoptosis; free radicals; lung cancer; quinoxalines
  32. Int J Mol Sci. 2019 Jan 23. pii: E478. [Epub ahead of print]20(3):
    Liang Z, Yuan Z, Guo J, Wu J, Yi J, Deng J, Shan Y.
      Ganoderma lucidum polysaccharide (GLP) extracted from Ganoderma lucidum (Leyss. ex Fr.) Karst, a traditional Chinese medicine, is a biologically active substance reported to possess anti-oxidative, anti-apoptotic, and neurological protection. However, it is unknown whether GLP have any protective effect against high-fat constituents-induced epithelial cell injury. The aim of this study was to investigate the protection and molecular mechanism of GLP on injury induced by palmitic acid (PA) in the intestinal porcine epithelial cell line (IPEC-J2). First, we tested whether the treatment of GLP attenuate PA-induced IPEC-J2 cell death. GLP markedly blocked PA-caused cytotoxicity and apoptosis in IPEC-J2 cells. Moreover, GLP recovered the decreased mitochondrial function and inhibited activation of caspase-dependent apoptotic pathway. Interestingly, PA promoted cell apoptosis and autophagy through stimulation of phosphorylation of mitogen-activated protein kinases (MAPKs), AMP-activated protein kinase (AMPK), and inhibition of phosphorylation of Akt and mammalian target of rapamycin (mTOR), which was reversed by GLP. Taken together, this study revealed a protective effect of GLP against PA-evoked IPEC-J2 cell death through anti-apoptotic and anti-autophagic properties.
    Keywords:  Ganoderma lucidum; apoptosis; autophagy; mitochondria; polysaccharides
  33. Phytomedicine. 2018 Sep 04. pii: S0944-7113(18)30314-3. [Epub ahead of print]53 124-133
    Li X, Qu Z, Jing S, Li X, Zhao C, Man S, Wang Y, Gao W.
      BACKGROUND: Lung cancer is the leading cause of global cancer-related mortality. Dioscin-6'-O-acetate (DA), a novel natural steroidal saponin, was firstly isolated from the rhizomes of Dioscorea althaeoides R. Knuth. Until now, there were no studies on its pharmacological activities.PURPOSE: Here, we investigated the growth inhibitory effect and explored the underlying molecular mechanisms of DA against lung cancer cells.
    METHODS/STUDY DESIGNS: NSCLC H460, H1299, H520 cells and SCLC H446 cells were treated with DA. To display the cytotoxic effects and possible mechanism of DA on these cells, MTT assay, flow cytometry and western blot analysis were carried out.
    RESULTS: Our results showed that DA exerted strong anti-proliferative activity against lung cancer cells in a concentration- and time-dependent manner. Flow cytometry demonstrated DA induced the cell cycle arrest at S-phase (NCI-H460, NCI-H1299, NCI-H520) or G1-phase (NCI-H446), caused cellular apoptosis, generation of reactive oxygen species (ROS) and loss of mitochondrial membrane potential. Western blotting analysis showed DA treatment increased the levels of caspase 3, 8, 9, Bax, p21, p53, phosphorylated JNK and p38 MAPK and markedly decreased the expression of Bcl-2, p-ERK, p-PI3K, p-AKT and NF-κB. Blockade of caspases with Z-VAD-FMK converted apoptosis-related proteins. Suppression of p53 with pifithrin-α (PFT) attenuated cell cycle-related protein. Inhibition of ROS with N-acetyl-cysteine (NAC) adjusted apoptosis-related proteins and phosphorylated MAPK and PI3K, as well as NF-κB.
    CONCLUSION: Overall, our study indicated that DA suppressed lung cancer cells proliferation via inducing cell-cycle arrest and enhancing caspase-dependent apoptosis, at least partly, through ROS-mediated PI3K/AKT, MAPK and NF-κB signaling pathways.
    Keywords:  Apoptosis; Caspase; Dioscin-6′-O-acetate; Lung cancer; MAPK; PI3K/AKT
  34. Free Radic Res. 2019 Jan 22. 1-11
    Zhao QL, Ito H, Kondo T, Uehara T, Ikeda M, Abe H, Saitoh JI, Noguchi K, Suzuki M, Kurachi M.
      Antioxidant activity has been reported for some atypical antipsychotic drugs; however, the detailed mechanism is not well known. Here we investigated the effects of atypical antipsychotic drugs on ·OH radical formation, intracellular reactive oxygen species (ROS), and apoptosis induced by ionizing radiation. The reaction rate constants with ·OH radicals were determined for five antipsychotic drugs as follows, in descending order: olanzapine, aripiprazole, clozapine, haloperidol, and risperidone. Experiments with aminophenyl fluorescein, a fluorescent dye, showed that olanzapine and clozapine could scavenge intracellular ROS. However, experiments with hydroxyphenyl fluorescein showed that only olanzapine inhibited ROS generation. X-irradiation-induced apoptosis in human lymphoma U937 cells was inhibited by clozapine at relatively low concentrations and by olanzapine at higher concentrations. Clozapine inhibited caspase-8 and caspase-3 activation and prevented loss of mitochondrial membrane potential. In contrast, olanzapine inhibited X-irradiation-induced p-JNK activation. Although the atypical antipsychotic drugs used here have relatively high reaction rate constants with ·OH radicals in aqueous solutions, inhibition of intracellular ROS was not due to ·OH radical scavenging. In addition, suppression of X-irradiation-induced apoptosis was not directly linked with intracellular ROS scavenging. When apoptosis signaling pathways were studied, clozapine-mediated inhibition of apoptosis was dependent on caspase-3 and caspase-8. In contrast, olanzapine inhibited apoptosis via down regulation of X-irradiation-induced p-JNK. These results suggested that both olanzapine and clozapine have antioxidative and antiapoptotic activities via distinct pathways, and provide useful information for better understanding of drug characteristics.
    Keywords:  Apoptosis; ROS; clozapine; olanzapine; radiation
  35. Curr Med Chem. 2019 Jan 21.
    Chen D, Liu JR, Cheng Y, Cheng H, He P, Sun Y.
      Rhaponticin is a stilbenoid glucoside compound, can be found in medicinal plant of rhubarb rhizomes. Rhapontigenin (RHAG), the stilbene aglycone metabolite of rhaponticin, has shown various biological activities including anticancer activities to act a potential human cytochrome P450 inhibitor, antihyperlipidemic effect, anti-allergic action, antioxidant and antibacterial activities. Moreover, it was reported to scavenge intravellular reactive oxygen species (ROS), the 1,1-diphenyl-2-picrylliydrazyl (DPPH) radical, and hydrogen peroxide (H2O2). Meanwhile, RHAG exhibited the inhibitory activity for synthesis of DNA, RNA and protein, and also presented the capacity of inducing morphological changes and apoptosis of C. albicans. Here, the structure, pharmacokinetics, pharmacological effects as well as underlying mechanisms of rhaponticin and its metabolite, RHAG, have been extensively reviewed. This review will provide a certain reference value for developing the therapeutic drug of rhaponticin or RHAG.
    Keywords:  Cancer; Cardiovascular disease; Metabolism; Pharmacokinetics; Rhaponticin; Rhapontigenin
  36. Int Immunopharmacol. 2019 Jan 18. pii: S1567-5769(19)30114-6. [Epub ahead of print]69 19-26
    Khajevand-Khazaei MR, Azimi S, Sedighnejad L, Salari S, Ghorbanpour A, Baluchnejadmojarad T, Mohseni-Moghaddam P, Khamse S, Roghani M.
      Sepsis is a serious and life-threatening medical condition with a higher rate of patients' morbidity and mortality and with complications such as acute kidney injury (AKI). S-allyl cysteine (SAC) is the active constituent of the medicinal plant garlic (Allium sativum) with multiple beneficial effects including anti-inflammatory and antioxidant properties. In this research, we tried to determine the protective effect of SAC pretreatment in a mouse model of AKI. To induce AKI, lipopolysaccharide (LPS) was injected once (10 mg/kg, i.p.) and SAC was administered at doses of 25, 50, or 100 mg/kg (p.o.) 1 h before LPS. Treatment of LPS-challenged C56BL/6 animals with SAC lowered serum level of creatinine and blood urea nitrogen (BUN), partially restored renal oxidative stress-related biomarkers including malondialdehyde (MDA), glutathione (GSH), and activity of superoxide dismutase (SOD) and catalase in addition to improvement of mitochondrial membrane potential (MMP). Furthermore, SAC was capable to bring renal nuclear factor-kappaB (NF-κB), nuclear factor (erythroid-derived 2)-like 2 (Nrf2), toll-like receptor 4 (TLR4), cyclooxygenase-2 (COX2), tumor necrosis factor α (TNFα), interleukin-1β (IL-1β), interleukin-6 (IL-6), Annexin V, and DNA fragmentation partially back to their control levels. Additionally, SAC pretreatment was capable to exert a protective effect, as shown histologically by lower tubular injury and pathologic changes in the kidney. In summary, SAC is capable to alleviate LPS-induced AKI through mitigation of renal oxidative stress, inflammation, and apoptosis in addition to preservation of mitochondrial integrity and its favorable effect exhibits a dose-dependent pattern.
    Keywords:  Acute kidney injury; Apoptosis; Inflammation; Lipopolysaccharide; Oxidative stress; S-allyl cysteine
  37. Food Sci Nutr. 2019 Jan;7(1): 120-131
    Gao C, Wang F, Yuan L, Liu J, Sun D, Li X.
      Appropriate germination can improve the nutritional value and bioactivity of soybeans; however, few studies have assessed the effect of germination on soybean proteins. This study examined the physicochemical property, antioxidation, and cytoprotective effect of the germinated soybean proteins (Gsp). Gsp was extracted from soybeans which germinated for 0-3 days using the method of alkali-solution and acid-isolation extraction. The results showed that germination could digest soybean proteins into the smaller molecules; enhance the degree of hydrolysis, emulsifiability, and foaming capacity; increase the removal rate of ABTS, DPPH, O 2 - ˙, and ˙OH radical; and decrease the reducing power and lipid peroxidation of Gsp. Additionally, Gsp was able to protect HL-7702 human hepatocyte cells against benzo(a)pyrene (BaP)-induced cytotoxicity through mediating the cell cycle arrest, suppressing apoptosis, and increasing reactive oxygen species (ROS) levels. This work demonstrated that germination could enhance the physicochemical property and antioxidant activity of Gsp, which also displayed the remarkable cytoprotective effect. This study provided a fundamental basis for substantiating dietary of Gsp used for resistance to oxidation and hepatic injury.
    Keywords:  antioxidant activity; apoptosis; benzo(a)pyrene; cytoprotective effect; germination soybean protein
  38. Sci Rep. 2019 Jan 21. 9(1): 245
    Siddiqui S, Ahmad R, Khan MA, Upadhyay S, Husain I, Srivastava AN.
      Ajwa dates (Phoenix dactylifera L.) are used by traditional therapeutic practitioners for several health benefits but most remain to be scientifically validated. In this study, we evaluated the apoptosis-inducing effect of ethanolic extract of Ajwa date pulp (ADP) on human hepatocellular carcinoma (HCC) HepG2 cells. High performance liquid chromatography analysis revealed the presence of polysaccharide β-D-glucan in ADP extract. Treated HCC cells revealed morphological characteristics of apoptosis under phase contrast microscopy. MTT assay demonstrated significant (p < 0.05) dose- and time-dependent inhibition of HCC cell growth. HCC cells were found to be in late apoptotic stage on treatment with higher doses of ADP extract as depicted by acridine orange/ethidium bromide and Annexin V-FITC/PI double stain. Importantly, ADP extract increased the reactive oxygen species level and decreased the mitochondrial membrane potential in treated HCC cells. Flow cytometry analysis demonstrated that ADP extract induced elevation of S and G2/M phases of cell cycle. Moreover, ADP extract induced apoptosis in HCC cells independent of tumor suppressor genes viz. CHEK2, ATM and TP53. Interestingly, ADP extract did not display any significant effect on normal cell line Vero. This study provides validation that ADP extract can be considered as a safe and natural potential drug candidate against human liver cancer.
  39. Radiol Oncol. 2019 Jan 19. pii: /j/raon.ahead-of-print/raon-2019-0002/raon-2019-0002.xml. [Epub ahead of print]
    Simenc J, Juric DM, Lipnik-Stangelj M.
      Background Astrocytes maintain central nerve system homeostasis and are relatively resistant to cell death. Dysfunction of cell death mechanisms may underlie glioblastoma genesis and resistance to cancer therapy; therefore more detailed understanding of astrocytic death modalities is needed in order to design effective therapy. The purpose of this study was to determine the effect of VAS2870, a pan-NADPH oxidase inhibitor, on staurosporine-induced cell death in astrocytes. Materials and methods Cultured rat astrocytes were treated with staurosporine as activator of cell death. Cell viability, production of reactive oxygen species (ROS), and mitochondrial potential were examined using flow cytometric analysis, while chemiluminescence analysis was performed to assess caspase 3/7 activity and cellular ATP. Results We show here for the first time, that VAS2870 is able to prevent staurosporine-induced cell death. Staurosporine exerts its toxic effect through increased generation of ROS, while VAS2870 reduces the level of ROS. Further, VAS2870 partially restores mitochondrial inner membrane potential and level of ATP in staurosporine treated cells. Conclusions Staurosporine induces cell death in cultured rat astrocytes through oxidative stress. Generation of ROS, mitochondrial membrane potential and energy level are sensitive to VAS2870, which suggests NADPH oxidases as an important effector of cell death. Consequently, NADPH oxidases activation pathway could be an important target to modulate astrocytic death.
    Keywords:  ATP; VAS2870; astrocytes; cell death; mitochondrial potential; reactive oxygen species
  40. Climacteric. 2019 Jan 24. 1-7
    Li Y, Ruan X, Liebenthron J, Montag M, Zhou Q, Kong W, Du J, Jin F, Li S, Cheng J, Wang H, Mueck AO.
      OBJECTIVE: Premature ovary insufficiency is frequent after chemotherapy/radiotherapy in cancer patients. Ovarian tissue (OT) cryopreservation and later retransplantation, the routine method in Europe, has recently been implemented at the first center in China. We investigated the protective effect of the antioxidant N-acetyl-l-cysteine (NAC) during the decisive freezing-thawing steps.METHODS: Fifteen OT samples were obtained from each of 13 cancer patients prospectively and randomly assigned to a control group and four groups with different NAC concentrations (Group 1, 0 mM NAC; Group 2, 0.5 mM NAC; Group 3, 1 mM NAC; Group 4, 5 mM NAC; Group 5, 25 mM NAC). After thawing, the follicle viability, DNA fragmentation, levels of reactive oxygen species (ROS), and total antioxidant capacity (TAC) were evaluated.
    RESULTS: OT cryopreserved and thawed with 25 mM NAC (Group 5) has the lowest proportion of apoptotic stroma cells, but the worst follicle viability. The other four groups show similar anti-apoptosis and good follicle viability. Group 4 presented the lowest ROS and highest TAC levels.
    CONCLUSIONS: OT cryopreserved and thawed in medium supplemented with 5 mM NAC shows the highest antioxidant and lowest ROS capability, good apoptotic parameters, and follicle viability. Our results need to be confirmed in larger patient cohorts prior to being accepted as a standard protocol.
    Keywords:  -acetyl--cysteine; DNA fragmentation; Ovarian tissue cryopreservation; follicle viability; reactive oxygen species; total antioxidant capability
  41. Exp Ther Med. 2019 Feb;17(2): 1312-1320
    Shen H, Yao Z, Zhao W, Zhang Y, Yao C, Tong C.
      Effective therapies to reduce ischemia/reperfusion and hypoxia/reoxygenation injury are currently lacking. Furthermore, the effects of loperamide and microRNA (miR)-21 on hypoxia/reoxygenation injury of cardiomyocytes have remained to be elucidated. Therefore, the present study aimed to investigate the effect of loperamide and miR-21 on cardiomyocytes during hypoxia/reoxygenation injury, and to explore the underlying molecular mechanisms. H9c2 rat cardiomyocytes were pre-treated with loperamide prior to hypoxia/reoxygenation. The viability of H9c2 cells was measured with a cell counting kit 8 and apoptosis was detected with an Annexin V-phycoerythrin/7-aminoactinomycin D apoptosis kit. Furthermore, reactive oxygen species were detected with a specific kit. Genes regulated by miR-21 were screened with an mRNA chip and confirmed using reverse-transcription quantitative polymerase chain reaction analysis. The direct targeting relationship of miR-21 with certain mRNAs was then confirmed using a Dual-Luciferase Reporter Assay system. The results indicated that the apoptotic rate and reactive oxygen species levels in rat cardiomyocytes were markedly increased after hypoxia/reoxygenation treatment. Pre-treatment with loperamide significantly protected H9c2 cells against apoptosis and reactive oxygen species production after hypoxia/reoxygenation. The protection was markedly decreased by miR-21 inhibitor and enhanced by miR-21 mimics. Screening for genes associated with cardiomyocyte apoptosis revealed that the relative expression of A-kinase anchoring protein 8 (Akap8) and BRCA1 associated RING domain 1 (Bard1) was consistent with the experimental results on apoptosis and reactive oxygen species. Compared with the group treated by hypoxia/reoxygenation alone, pre-treatment with loperamide markedly decreased the expression of BRCA1-interacting protein C-terminal helicase 1, Akap8 and Bard1 after hypoxia/reoxygenation. The decrease in the expression of Akap8 and Bard1 was markedly attenuated by miR-21 inhibitor and enhanced by miR-21 mimics. miR-21 mimics directly targeted the 3'-untranslated region (UTR) of Akap8 and Bard1 mRNA to thereby decrease their expression. In conclusion, the protection of rat cardiomyocytes against hypoxia/reoxygenation-induced apoptosis and reactive oxygen species production by loperamide was markedly enhanced by miR-21. miR-21 directly targets the 3'-UTR of Akap8 and Bard1 mRNA and enhances the inhibitory effects of loperamide on Akap8 and Bard1 expression in rat cardiomyocytes after hypoxia/reoxygenation.
    Keywords:  cardiomyocytes; hypoxia; loperamide; miR-21; reoxygenation
  42. Mol Neurobiol. 2019 Jan 24.
    Islam A, Kagawa Y, Miyazaki H, Shil SK, Umaru BA, Yasumoto Y, Yamamoto Y, Owada Y.
      Fatty acid-binding proteins (FABPs) bind and internalize long-chain fatty acids, controlling lipid dynamics. Recent studies have proposed the involvement of FABPs, particularly FABP7, in lipid droplet (LD) formation in glioma, but the physiological significance of LDs is poorly understood. In this study, we sought to examine the role of FABP7 in primary mouse astrocytes, focusing on its protective effect against reactive oxygen species (ROS) stress. In FABP7 knockout (KO) astrocytes, ROS induction significantly decreased LD accumulation, elevated ROS toxicity, and impaired thioredoxin (TRX) but not peroxiredoxin 1 (PRX1) signalling compared to ROS induction in wild-type astrocytes. Consequently, activation of apoptosis signalling molecules, including p38 mitogen-activated protein kinase (MAPK) and stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK), and increased expression of cleaved caspase 3 were observed in FABP7 KO astrocytes under ROS stress. N-acetyl L-cysteine (NAC) application successfully rescued the ROS toxicity in FABP7 KO astrocytes. Furthermore, FABP7 overexpression in U87 human glioma cell line revealed higher LD accumulation and higher antioxidant defence enzyme (TRX, TRX reductase 1 [TRXRD1]) expression than mock transfection and protected against apoptosis signalling (p38 MAPK, SAPK/JNK and cleaved caspase 3) activation. Taken together, these data suggest that FABP7 protects astrocytes from ROS toxicity through LD formation, providing new insights linking FABP7, lipid homeostasis, and neuropsychiatric/neurodegenerative disorders, including Alzheimer's disease and schizophrenia.
    Keywords:  Astrocytes; Fatty acid-binding protein 7; Lipid droplet; Thioredoxin; U87
  43. Free Radic Res. 2019 Jan 22. 1-191
    He W, Cao P, Xia Y, Hong L, Zhang T, Shen X, Zheng P, Shen H, Zhao Y, Zou P.
      Thioredoxin reductase 1 (TrxR1) has emerged as a potential target for cancer therapy, because it is overexpressed in several types of cancers and associated with increased tumor growth and poor patient prognosis. Alantolactone (ALT), a natural sesquiterpene lactone originated from traditional folk medicine Inula helenium L., has been reported to exert antitumor activity in various tumors. However, the effect of ALT on human gastric cancer cells and its underlying mechanism remains unknown. In this study, we showed that ALT inhibited cell proliferation and induced cell apoptosis in gastric cancer cells. Mechanistically, our data found that ALT induced reactive oxygen species (ROS) production by inhibiting TrxR1 activity, resulting in the activation of p38 mitogen-activated protein kinase (MAPK) pathway and eventually cell apoptosis in gastric cancer cells. And the effects of ALT were reversed by pretreatment with NAC (a scavenger of ROS). Further investigation revealed that ALT displayed synergistic lethality with erastin against gastric cancer cells, which demonstrating combined inhibition of TrxR1 and glutathione (GSH) leads to a synergistic effect in gastric cancer cells. More importantly, ALT treatment markedly reduced the activity of TrxR1 in vivo, and inhibited the growth of gastric cancer xenografts without exhibiting significant toxicity. Taken together, these findings suggest that ALT may be used as a novel therapeutic agent against human gastric cancer.
    Keywords:  Alantolactone; erastin; p38 MAPK; reactive oxygen species; thioredoxin reductase 1
  44. Neurotox Res. 2019 Jan 22.
    Ahmad Rather M, Justin-Thenmozhi A, Manivasagam T, Saravanababu C, Guillemin GJ, Essa MM.
      Asiatic acid (AA), a triterpenoid present in Centella asiatica, possesses the ability to cross blood brain barrier and received considerable attention for its neuroprotective role. We have reported the benefit of AA against aluminum chloride (AlCl3)-induced amyloid pathology, enhanced acetylcholine esterase (AChE) activity, and inflammation in Alzheimer's disease (AD) like model rats. Based on that, to find the exact mechanism of action of AA, the present study was designed to evaluate the oxidative stress, tau pathology, apoptosis, and Akt/GSK3β signaling pathway on AlCl3-induced neurotoxicity in Wistar rats. AD-like pathology was induced by oral administration of AlCl3 (100 mg/kg b.w.) for 6 weeks, which demonstrated a significant reduction in spatial memory performance, anxiety, and motor dysfunction and diminished the expression of cyclin-dependent kinase 5 (CDK 5-enzyme implicated in the phosphorylation of tau proteins), pTau, oxidative stress, and apoptosis, whereas oral ingestion of AA (75 mg/kg b.w.) for 7 weeks attenuated the above-said indices, which could be by activating Akt/GSK3β pathway. Current results suggested that AA could be able to modulate various pathological features of AD and could hold promise in AD treatment.
    Keywords:  Aluminum chloride; Apoptosis; Asiatic acid; Oxidative stress; Tau pathology
  45. Phytomedicine. 2018 Sep 19. pii: S0944-7113(18)30470-7. [Epub ahead of print]56 147-155
    Garg S, Malhotra RK, Khan SI, Sarkar S, Susrutha PN, Singh V, Goyal S, Nag TC, Ray R, Bhatia J, Arya DS.
      BACKGROUND: The therapeutic options for the reducing the damage caused by myocardial ischemia are limited and not devoid of adverse effects. The role of the flavanoid, fisetin, predominantly found in strawberry and apple, is yet to be explored in the heart.STUDY DESIGN: Male Wistar rats (n = 48) were administered fisetin (10, 20 & 40 mg/kg/day, orally) or vehicle for 28 days while ISO, 85 mg/kg, subcutaneously, was also administered at 24 h interval on the 27th and 28th day. On the 29th day, rats were anaesthetized and right carotid artery was cannulated to record hemodynamic parameters. Subsequently, blood sample was collected and heart was removed to evaluate various parameters.
    RESULTS: Fisetin at doses of 10 and 20 mg/kg reversed ISO induced detrimental alterations in blood pressure and left ventricular pressures and reduced the myocardial injury markers CK-MB and LDH in the serum. These findings were supported by amelioration of ISO induced histological and ultrastructural damage by fisetin. The disequilibrium in the levels of pro and anti oxidants in the myocardial tissue caused by ISO was also normalized Furthermore, apoptosis was evident from enhanced DNA fragmentation and raised pro-apoptotic proteins (bax, caspase-3, cytochrome-c) as well as suppressed anti-apoptotic protein (Bcl-2) in case of ISO treatment which again was reversed by fisetin. A molecular mechanism for this protection was elucidated as downregulation of RAGE and NF-κB However fisetin at 40 mg/kg revealed a deteriorating effect which was similar to ISO group of rats.
    CONCLUSION: Hence, through our study, the role of fisetin in cardioprotection has been uncovered via a molecular pathway.
    Keywords:  Apoptosis; Fisetin; Inflammation; Isoproterenol; Oxidative stress; RAGE
  46. J Ginseng Res. 2019 Jan;43(1): 10-19
    Hu JN, Xu XY, Li W, Wang YM, Liu Y, Wang Z, Wang YP.
      Background: Frequent overdose of paracetamol (APAP) has become the major cause of acute liver injury. The present study was designed to evaluate the potential protective effects of ginsenoside Rk1 on APAP-induced hepatotoxicity and investigate the underlying mechanisms for the first time.Methods: Mice were treated with Rk1 (10 mg/kg or 20 mg/kg) by oral gavage once per d for 7 d. On the 7th d, all mice treated with 250 mg/kg APAP exhibited severe liver injury after 24 h, and hepatotoxicity was assessed.
    Results: Our results showed that pretreatment with Rk1 significantly decreased the levels of serum alanine aminotransferase, aspartate aminotransferase, tumor necrosis factor, and interleukin-1β compared with the APAP group. Meanwhile, hepatic antioxidants, including superoxide dismutase and glutathione, were elevated compared with the APAP group. In contrast, a significant decrease in levels of the lipid peroxidation product malondialdehyde was observed in the ginsenoside Rk1-treated group compared with the APAP group. These effects were associated with a significant increase of cytochrome P450 E1 and 4-hydroxynonenal levels in liver tissues. Moreover, ginsenoside Rk1 supplementation suppressed activation of apoptotic pathways by increasing Bcl-2 and decreasing Bax protein expression levels, which was shown using western blotting analysis. Histopathological observation also revealed that ginsenoside Rk1 pretreatment significantly reversed APAP-induced necrosis and inflammatory infiltration in liver tissues. Biological indicators of nitrative stress, such as 3-nitrotyrosine, were also inhibited after pretreatment with Rk1 compared with the APAP group.
    Conclusion: The results clearly suggest that the underlying molecular mechanisms in the hepatoprotection of ginsenoside Rk1 in APAP-induced hepatotoxicity may be due to its antioxidation, antiapoptosis, anti-inflammation, and antinitrative effects.
    Keywords:  APAP-induced hepatotoxicity; anti-apoptosis; anti-inflammation; ginsenoside Rk1; oxidative stress
  47. Nanoscale. 2019 Jan 22.
    Gao F, He G, Yin H, Chen J, Liu Y, Lan C, Zhang S, Yang B.
      The development of efficient nanomedicines to improve anticancer therapeutic effects is highly attractive. In this work, we firstly report titania-coated Au nanoplate (Au NPL@TiO2) heterostructures, which play dual roles as nanoagents for synergistic photothermal/sonodynamic therapy in the second near-infrared (NIR) window. On the one hand, because the controlled TiO2 shells endow the Au NPL@TiO2 nanostructures with a red shift to the NIR II region, the as-prepared Au NPL@TiO2 nanostructures possess a high photothermal conversion efficiency of 42.05% when irradiated by a 1064 nm laser and are anticipated to be very promising candidates as photothermal agents. On the other hand, the Au nanoplates (Au NPLs), as electron traps, vastly improve the generation of reactive oxygen species (ROS) by the Au NPL@TiO2 nanostructures in contrast with pure TiO2 shell nanoparticles upon activation by ultrasound (US) via a sonodynamic process. Moreover, the toxicity and therapeutic effect of the Au NPL@TiO2 nanostructures were relatively systemically evaluated in vitro. The Au NPL@TiO2 nanostructures generate a large amount of intracellular ROS and exhibit laser power density-dependent toxicity, which eventually induces apoptosis of cancer cells. Furthermore, a synergistic therapeutic effect, with a cell viability of only 20.3% upon both photothermal and sonodynamic activation, was achieved at low concentrations of the Au NPL@TiO2 nanostructures. Experiments on mice also demonstrate the superiority of the combination of PTT and SDT, with the total elimination of tumors. This work provides a way of applying two-dimensional (2D) gold nanoplate core/TiO2 shell nanostructures as novel nanoagents for advanced multifunctional anticancer therapies in the second NIR window.
  48. Front Cell Infect Microbiol. 2018 ;8 445
    Jia C, Zhang J, Yu L, Wang C, Yang Y, Rong X, Xu K, Chu M.
      Coumarin (1,2-benzopyrone), an aromatic oxygen-containing heterocyclic compound, has various biological functions. Previous studies have demonstrated that coumarin and its derivatives exhibit antifungal activity against Candida albicans. In this study, we investigated the exact mechanism by which coumarin works against this fungus using Annexin V-FITC/PI double staining, TUNEL assay, and DAPI staining, and found that it induced a series of apoptotic features, including phosphatidylserine (PS) externalization, DNA fragmentation, and nuclear condensation. Moreover, it also induced cytochrome c release from the mitochondria to the cytoplasm and metacaspase activation. Further study revealed that intracellular reactive oxygen species (ROS) levels were increased and mitochondrial functions, such as mitochondrial membrane potential and mitochondrial morphology, were altered after treatment with coumarin. Cytosolic and mitochondrial Ca2+ levels were also found to be elevated. However, pretreatment with ruthenium red (RR), a known mitochondrial Ca2+ channel inhibitor, attenuated coumarin-mediated DNA fragmentation and metacaspase activity, indicating that the coumarin-induced C. albicans apoptosis is associated with mitochondrial Ca2+ influx. Finally, coumarin was found to be low-toxic and effective in prolonging the survival of C. albicans-infected mice. This study highlights the antifungal activity and mechanism of coumarin against C. albicans and provides a potential treatment strategy for C. albicans infection.
    Keywords:  Ca2+; Candida albicans; ROS; apoptosis; coumarin
  49. Apoptosis. 2019 Jan 24.
    Jiang X, Liao XH, Huang LL, Sun H, Liu Q, Zhang L.
      Ischemia/reperfusion is a major cause of acute kidney injury and can induce apoptosis in renal epithelial tubule (HK-2) cells. Accumulating evidence indicates that endoplasmic reticulum (ER) stress is a major contributor to apoptosis in acute kidney injury. We previously reported that augmenter of liver regeneration (ALR) functions as an anti-apoptotic factor in H2O2-treated HK-2 cells although the precise mechanism underlying this action remains unclear. In the present study, we investigate the role of ALR in H2O2-induced ER stress-mediated apoptosis. We overexpressed ALR and established a H2O2-induced ER stress model in HK-2 cells. Overexpression of ALR reduced the level of reactive oxygen species and the rate of apoptosis in H2O2-treated HK-2 cells. Using confocal microscopy and western blot, we observed that ALR colocalized with the ER and mitochondria compartment. Moreover, ALR suppressed ER stress by maintaining the morphology of the ER and reducing the levels of the ER-related proteins, glucose-regulated protein 78 (GRP78), phospho-protein kinase-like ER kinase (p-PERK), phospho-eukaryotic initiation factor 2α (p-eIF2α) and C/EBP-homologous protein (CHOP) significantly (p < 0.05). Mechanistically, ALR promoted Bcl-2 expression and suppressed Bax and cleaved-caspase-3 expression significantly during ER-stress induced apoptosis (p < 0.05). Furthermore, ALR attenuated calcium release from the ER, and transfer to mitochondria, under ER stress. To conclude, ALR alleviates H2O2-induced ER stress-mediated apoptosis in HK-2 cells by suppressing ER stress response and by maintaining calcium homeostasis. Consequently, ALR may protect renal tubule epithelial cells from ischemia/reperfusion induced acute kidney injury.
    Keywords:  Acute kidney injury; Apoptosis; Augmenter of liver regeneration; Endoplasmic reticulum stress
  50. Med Sci Monit. 2019 Jan 25. 25 739-745
    He J, Wei W, Yang Q, Wang Y.
      BACKGROUND Esophageal cancer causes considerable mortality and is ranked as the 6th most prevalent type of cancer across the world. At present, there is no effective esophageal cancer chemotherapy without adverse effects. Moreover, emergence of drug resistance among cancer is another obstacle in the treatment of esophageal cancer. Novel molecules of plant origin may prove beneficial in the development of chemotherapy for esophageal carcinoma. In this study we examined the anticancer effects of phillygenin against the vindesine-resistant esophageal cancer cell line SH-1-V1. MATERIAL AND METHODS The proliferation rate of SH-1-V1 cells was determined by WST-1 assay. Apoptosis was confirmed by propidium iodide (PI) staining. Cell cycle analysis, ROS, and MMP determination were performed by flow cytometery. Protein expression was assessed by Western blot analysis. RESULTS We found that phillygenin inhibited the growth of SH-1-V1 cells and exhibited an IC50 of 6 µM. Investigation of the underlying mechanism revealed that phillygenin triggered apoptotic cell death of the SH-1-V1 cells, which was also associated with enhancement of Bax expression and decreased expression of Bcl-2. Moreover, the expression of cleaved caspase 3 and 9 also increased upon phillygenin treatment. Phillygenin also caused a significant increase in ROS production, concomitant with decreased MMP levels. Phillygenin also caused arrest of cells in the G2/M phase of the cell cycle. In vivo evaluation of phillygenin revealed that it can inhibit tumor weight and volume, suggesting the anticancer potential of phillygenin. CONCLUSIONS In brief, phillygenin inhibited in vitro and in vivo cancer cell growth in drug-resistant human esophageal cancer cells, and these effects were mediated via apoptosis, ROS generation, mitochondrial membrane potential loss, and activation of the NF-kB signalling pathway.
  51. Cancer Lett. 2019 Jan 20. pii: S0304-3835(19)30014-X. [Epub ahead of print]
    Tian J, Guo F, Chen Y, Li Y, Yu B, Li Y.
      Globally, prostate cancer remains a challenging health burden for men as it is the second leading cause of cancer death in men and about one in nine will be diagnosed with prostate cancer in his lifetime. Enhanced expression of COX-2 and Glut-1 proteins are reported as major factors leading to the origin and progress of prostate cancer through modulating the associated signaling pathways. In this study, we have synthesized a multifunctional liposomal system containing celecoxib and genistein drugs. The combinatorial effect of these drugs leads to the selectively induce the apoptosis of prostate cancer cells than normal fibroblast cells. The mechanistic study suggests that enhanced reactive oxygen species (ROS) formation and a decrease in cellular GSH concentration, along with inhibition of COX-2 synthesis and Glut-1 receptors are the key processes behind the inhibition of prostate cancer cells. Overall, these results provide strong evidence for the role of COX-2 and Glut-1 proteins for the progression of prostate cancer and highlighting the potential of celecoxib and genistein as a useful and combinatorial pharmacological agent for chemotherapeutic purposes in prostate cancer.
    Keywords:  Celecoxib; Drug delivery; Genistein; Nanoliposomes; Prostate cancer
  52. J Nanobiotechnology. 2019 Jan 25. 17(1): 18
    Zhang CX, Cheng Y, Liu DZ, Liu M, Cui H, Zhang BL, Mei QB, Zhou SY.
      BACKGROUND: Cyclosporin A (CsA) is a promising therapeutic drug for myocardial ischemia reperfusion injury (MI/RI) because of its definite inhibition to the opening of mitochondrial permeability transition pore (mPTP). However, the application of cyclosporin A to treat MI/RI is limited due to its immunosuppressive effect to other normal organ and tissues. SS31 represents a novel mitochondria-targeted peptide which can guide drug to accumulate into mitochondria. In this paper, mitochondria-targeted nanoparticles (CsA@PLGA-PEG-SS31) were prepared to precisely deliver cyclosporin A into mitochondria of ischemic cardiomyocytes to treat MI/RI.RESULTS: CsA@PLGA-PEG-SS31 was prepared by nanoprecipitation. CsA@PLGA-PEG-SS31 showed small particle size (~ 50 nm) and positive charge due to the modification of SS31 on the surface of nanoparticles. CsA@PLGA-PEG-SS31 was stable for more than 30 days and displayed a biphasic drug release pattern. The in vitro results showed that the intracellular uptake of CsA@PLGA-PEG-SS31 was significantly enhanced in hypoxia reoxygenation (H/R) injured H9c2 cells. CsA@PLGA-PEG-SS31 delivered CsA into mitochondria of H/R injured H9c2 cells and subsequently increased the viability of H/R injured H9c2 cell through inhibiting the opening of mPTP and production of reactive oxygen species. In vivo results showed that CsA@PLGA-PEG-SS31 accumulated in ischemic myocardium of MI/RI rat heart. Apoptosis of cardiomyocyte was alleviated in MI/RI rats treated with CsA@PLGA-PEG-SS31, which resulted in the myocardial salvage and improvement of cardiac function. Besides, CsA@PLGA-PEG-SS31 protected myocardium from damage by reducing the recruitment of inflammatory cells and maintaining the integrity of mitochondrial function in MI/RI rats.
    CONCLUSION: CsA@PLGA-PEG-SS31 exhibited significant cardioprotective effects against MI/RI in rats hearts through protecting mitochondrial integrity, decreasing apoptosis of cardiomyocytes and myocardial infract area. Thus, CsA@PLGA-PEG-SS31 offered a promising therapeutic method for patients with acute myocardial infarction.
    Keywords:  Cyclosporin A; Mitochondria-targeted peptide; Mitochondrial permeability transition pore; Mitochondrial targeting; Myocardial ischemia/reperfusion injury
  53. Cell Death Dis. 2019 Jan 22. 10(2): 57
    Ross JA, Dungen KV, Bressler KR, Fredriksen M, Khandige Sharma D, Balasingam N, Thakor N.
      Physiological stress conditions attenuate global mRNA translation via modifications of key eukaryotic initiation factors. However, non-canonical translation initiation mechanisms allow cap-independent translation of certain mRNAs. We have previously demonstrated that eIF5B promotes cap-independent translation of the mRNA encoding the antiapoptotic factor, XIAP, during cellular stress. Here, we show that depletion of eIF5B sensitizes glioblastoma multiforme cells to TRAIL-induced apoptosis by a pathway involving caspases-8, -9, and -7, with no significant effect on cell cycle progression. eIF5B promotes evasion of apoptosis by promoting the translation of several IRES-containing mRNAs, encoding the antiapoptotic proteins XIAP, Bcl-xL, cIAP1, and c-FLIPS. We also show that eIF5B promotes translation of nuclear factor erythroid 2-related factor 2 and suggest that reactive oxygen species contribute to increased apoptosis under conditions of eIF5B depletion. Finally, eIF5B depletion leads to decreased activation of the canonical NF-κB pathway. Taken together, our data suggest that eIF5B represents a regulatory node, allowing cancer cells to evade apoptosis by promoting the translation of pro-survival proteins from IRES-containing mRNAs.
  54. J Biomed Sci. 2019 Jan 21. 26(1): 11
    Chaudhary GR, Yadav PK, Yadav AK, Tiwari M, Gupta A, Sharma A, Pandey AN, Pandey AK, Chaube SK.
      Stress is deeply rooted in the modern society due to limited resources and large competition to achieve the desired goal. Women are more frequently exposed to several stressors during their reproductive age that trigger generation of reactive oxygen species (ROS). Accumulation of ROS in the body causes oxidative stress (OS) and adversely affects ovarian functions. The increased OS triggers various cell death pathways in the ovary. Beside apoptosis and autophagy, OS trigger necroptosis in granulosa cell as well as in follicular oocyte. The OS could activate receptor interacting protein kinase-1(RIPK1), receptor interacting protein kinase-3 (RIPK3) and mixed lineage kinase domain-like protein (MLKL) to trigger necroptosis in mammalian ovary. The granulosa cell necroptosis may deprive follicular oocyte from nutrients, growth factors and survival factors. Under these conditions, oocyte becomes more susceptible towards OS-mediated necroptosis in the follicular oocytes. Induction of necroptosis in encircling granulosa cell and oocyte may lead to follicular atresia. Indeed, follicular atresia is one of the major events responsible for the elimination of majority of germ cells from cohort of ovary. Thus, the inhibition of necroptosis could prevent precautious germ cell depletion from ovary that may cause reproductive senescence and early menopause in several mammalian species including human.
    Keywords:  MLKL; Necroptosis; Ovary; Oxidative stress; RIPK; Stress; TNF
  55. Int J Biol Macromol. 2019 Jan 16. pii: S0141-8130(18)36102-6. [Epub ahead of print]
    Zhang W, Wu J, Zhang F, Dou X, Ma A, Zhang X, Shao H, Zhao S, Ling P, Liu F, Han G.
      LRWXG has previously been reported to have a protective effect on chondrocytes, preventing apoptosis induced by oxidative stress. In this study, we were aimed at determining whether LRWXG exerts its anti-apoptotic activity through the MAPK signaling pathways in chondrocytes. Our results show that, at the cellular level, apoptosis of chondrocytes in the groups treated by LRWXG decreases compared with groups treated by inhibitors alone and model group under conditions of oxidative stress in a dose-dependent manner. Mechanistically at the molecular level, LRWXG regulates the MAPK pathway induced by oxidative stress: The levels of phosphorylation of JNK and p38 proteins in the groups treated by LRWXG are lower than model group, while compared with corresponding groups of inhibitors, there are no significant difference; For other related proteins, LRWXG reduces the levels of the apoptosis-related proteins BAX and cleaved caspase-3, and increases the level of anti-apoptotic protein BCL2. In addition, LRWXG can significantly reduce the levels of inflammatory-related factors such as COX2, PEG2, TNFα and IL1β, and inhibits the expression of MMPs, increasing the content of type II collagen. The results of this research strongly suggest that LRWXG exerts its anti-apoptotic activity via regulating the MAPK signaling pathways in vitro.
    Keywords:  Apoptosis; Lower range of molecular weight of xanthan gum; Mitogen-activated protein kinase
  56. J Cell Mol Med. 2019 Jan 25.
    Hu S, Cheng M, Guo X, Wang S, Liu B, Jiang H, Huang C, Wu G.
      BACKGROUND: MicroRNAs (miRNAs) have been shown to commonly contribute to cardiac hypertrophy (CH). The aim of this study was to test the hypothesis that miR-200c plays an important role in the progression of CH by targeting myosin light chain kinase (MLCK/MYLK).METHODS AND RESULTS: Cardiac hypertrophy was induced by aortic banding (AB) in rats. Cellular hypertrophy in neonatal rat cardiomyocytes (NCMs) was induced by AngII treatment. Echocardiography, histology and molecular measurements were used to assess the results of the experiments. The levels of apoptosis and reactive oxygen species (ROS) were also measured. Quantitative real-time PCR (qRT-PCR) and Western blotting were used to measure mRNA and protein levels respectively. The present results showed that miR-200c expression was increased in response to CH both in vivo and in vitro. The down-regulation of miRNA-200c by a specific inhibitor markedly ameliorated CH resulting from AngII treatment, and the mRNA levels of atrial natriuretic peptide, brain natriuretic peptide and β-myosin heavy chain were simultaneously decreased. Notably, minimal apoptosis and ROS accumulation were identified in AngII-induced hypertrophic cardiomyocytes. Conversely, the up-regulation of miR-200c using specific mimics reversed these effects. Mechanistic investigations demonstrated that the MLCK gene is a direct target of miR-200c; an increase in miR-200c levels led to a decrease in the expression of MLCK and its downstream effector, p-MLC2, while miR-200c inhibition increased the expression of these proteins. Furthermore, inhibiting MLCK impaired the anti-hypertrophic effects contributions produced by the knockdown of miR-200c.
    CONCLUSION: Our studies suggest that miR-200c may serve as a potential therapeutic target that could delay hypertrophy. We have also uncovered a relationship between miR-200c and MLCK, identifying MLCK as a direct mediator of miR-200c.
    Keywords:  apoptosis; cardiac hypertrophy; miR-200c; myosin light chain kinase; reactive oxygen species
  57. Cell Tissue Res. 2019 Jan 21.
    Sohel MMH, Akyuz B, Konca Y, Arslan K, Sariozkan S, Cinar MU.
      Despite its essential role in ovulation, oxidative stress (OS) has been found to be cytotoxic to cells, while microRNAs (miRNAs) are known as a major regulator of genes involved in cellular defense against cytotoxicity. However, a functional link between OS and miRNA expression changes in granulosa cells (GCs) remains to be investigated. Here, we investigate the OS modulation of apoptosis-associated miRNAs and their biological relevance in bovine GCs. Following the evaluation of cell viability, accumulation of reactive oxygen species (ROS), cytotoxicity and mitochondrial activity, we used a ready-to-use miRNA PCR array to identify differentially regulated miRNAs. The results showed that exposure to 150 μM H2O2 for 4 h creates remarkable signs of OS in GCs characterized by more than 50% loss of cell viability, higher nuclear factor erythroid 2-related factor 2 (NRF2) nuclear translocation, significantly (p < 0.05) higher abundance of antioxidant genes, significantly (p < 0.001) higher accumulation of ROS, lower mitochondrial activity and a higher (p < 0.001) number of apoptotic nuclei compared to that of the control group. miRNA expression analysis revealed that a total of 69 miRNAs were differentially regulated in which 47 and 22 miRNAs were up- and downregulated, respectively, in stressed GCs. By applying the 2-fold and p < 0.05 criteria, we found 16 miRNAs were upregulated and 10 miRNAs were downregulated. Target prediction revealed that up- and downregulated miRNAs potentially targeted a total of 6210 and 3575 genes, respectively. Pathway analysis showed that upregulated miRNAs are targeting the genes involved mostly in cell survival, intracellular communication and homeostasis, cellular migration and growth control and disease pathways. Our results showed that OS modulates the expression of apoptosis-associated miRNAs that might have effects on cellular or molecular damages.
    Keywords:  Apoptosis; Granulosa cells; Oxidative stress; Signaling pathways; microRNA
  58. Mediators Inflamm. 2018 ;2018 6571676
    Kim JY, Park JH, Kim K, Jo J, Leem J, Park KK.
      Caspase-1 is a proinflammatory caspase responsible for the proteolytic conversion of the precursor forms of interleukin-1β to its active form and plays an important role in the pathogenesis of various inflammatory diseases. It was reported that genetic deficiency of caspase-1 prevented cisplatin-induced nephrotoxicity. However, whether pharmacological inhibition of caspase-1 also has a preventive effect against cisplatin-induced kidney injury has not been evaluated. In this study, we examined the effect of Ac-YVAD-cmk, a potent caspase-1-specific inhibitor, on renal function and histology in cisplatin-treated mice and explored its underlying mechanisms. We found that administration of Ac-YVAD-cmk effectively attenuated cisplatin-induced renal dysfunction, as evidenced by reduced plasma levels of blood urea nitrogen and creatinine, and histological abnormalities, such as tubular cell death, dilatation, and cast formation. Administration of Ac-YVAD-cmk inhibited caspase-3 activation as well as caspase-1 activation and attenuated apoptotic cell death, as assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling, in the kidneys of cisplatin-treated mice. Cisplatin-induced G2/M arrest of renal tubular cells was also reduced by caspase-1 inhibition. In addition, administration of Ac-YVAD-cmk reversed increased oxidative stress and depleted antioxidant capacity after cisplatin treatment. Moreover, increased macrophage accumulation and elevated expression of cytokines and chemokines were attenuated by caspase-1 inhibition. Taken together, these results suggest that caspase-1 inhibition by Ac-YVAD-cmk protects against cisplatin-induced nephrotoxicity through inhibition of renal tubular cell apoptosis, oxidative stress, and inflammatory responses. Our findings support the idea that caspase-1 may be a promising pharmacological target for the prevention of cisplatin-induced kidney injury.
  59. Nephron. 2019 Jan 23. 1-11
    Yuan L, Liao PP, Song HC, Zhou JH, Chu HC, Lyu L.
      AIMS: Hyperbilirubinemia is associated with postoperative acute kidney injury in patients undergoing cardiac surgeries. A high concentration of bilirubin could induce oxidative stress and cell apoptosis. The aim of this study was to investigate whether hyperbilirubinemia aggravated the renal tubule cells injury and the pro-apoptotic potential of bilirubin on renal ischemia reperfusion injury (RIRI).METHODS: The human proximal tubular epithelial cell line HK-2 cells were challenged with a gradient concentration of bilirubin for 24 h. Cell injury was assessed by flow cytometry and MTT assay. Bilirubin was injected intraperitoneally into male Sprague-Dawley rats once every 12 h (100 mg/kg), 3 times in total. The same solvent volume without bilirubin powder was used as vehicle in non-bilirubin injection groups. The RIRI surgical procedure was a bilateral renal pedicles clamping (45 min) followed by 30 h reperfusion. The rats were divided into 4 groups: negative control (NC), similar surgical procedures without clamping; Bil, bilirubin injection for 36 h, then rats were sacrificed; RIRI, RIRI surgical procedures; Bil + RIRI, RIRI applied 6 h later than the first bilirubin injection, rats were sacrificed after another 30 h.
    RESULTS: In vitro, bilirubin induced cell apoptosis and significantly decreased the cell viability of HK-2 cells. Bilirubin induced the active caspase 3 and phosphorylation of p38 in HK-2 cells. In vivo, serum creatinine was higher in Bil + RIRI compared with RIRI (p < 0.01). The tubular injury scores of hematoxylin and eosin and tubular necrosis scores of periodic acid-Schiff were higher in Bil + RIRI than these in RIRI (All p < 0.05). The number of Tunel-positive nuclei was higher in Bil + RIRI compared to RIRI (p < 0.001). The active caspase 3 and phosphorylation of p38 were higher and the Bcl2 was lower in Bil + RIRI compared to RIRI. Moreover, the apoptosis level was higher in Bil compared to NC.
    CONCLUSIONS: Our results reveal that the hyperbilirubinemia induces pro-apoptotic effects and aggravates RIRI.
    Keywords:  Apoptotic; Hyperbilirubinemia; Renal ischemia reperfusion injury
  60. Sci Rep. 2019 Jan 23. 9(1): 429
    Suekane A, Saito Y, Nakahata S, Ichikawa T, Ogoh H, Tsujikawa K, Morishita K.
      Ecotropic viral integration site-1 (EVI1) has a critical role in normal and malignant hematopoiesis. Since we previously identified high expression of calcitonin receptor like receptor (CRLR) in acute myeloid leukemia (AML) with high EVI1 expression, we here characterized the function of CRLR in hematopoiesis. Since higher expression of CRLR and receptor activity modifying protein 1 (RAMP1) was identified in immature hematopoietic bone marrow (BM) cells, we focused on calcitonin gene-related peptide (CGRP), a specific ligand for the CRLR/RAMP1 complex. To elucidate the role of CGRP in hematopoiesis, Ramp1-deficient (Ramp1-/-) mice were used. The steady-state hematopoiesis was almost maintained in Ramp1-/- mice; however, the BM repopulation capacity of Ramp1-/- mice was significantly decreased, and the transplanted Ramp1-/- BM mononuclear cells had low proliferation capacity with enhanced reactive oxygen species (ROS) production and cell apoptosis. Thus, CGRP is important for maintaining hematopoiesis during temporal exposures with proliferative stress. Moreover, continuous CGRP exposure to mice for two weeks induced a reduction in the number of BM immature hematopoietic cells along with differentiated myeloid cells. Since CGRP is known to be increased under inflammatory conditions to regulate immune responses, hematopoietic exhaustion by continuous CGRP secretion under chronic inflammatory conditions is probably one of the important mechanisms of anti-inflammatory responses.