bims-supasi 2025-03-16 papers

bims-supasi

Biomed News

on Sulfation pathways and signalling

Issue of 2025–03–16
fourteen papers selected by
Jonathan Wolf Mueller, University of Birmingham

Highly selective peptide-based fluorescent probe with aggregation induced emission (AIE) for detection of chondroitin sulfate and its application in living cells and zebrafish imaging.
Online PGC-LC-MS analysis of colonic mucin O-glycans in ovalbumin-induced food allergy in Balb/c mice by treatment with sea cucumber chondroitin sulfate polysaccharide.
Cartilaginous fishes-derived chondroitin sulfates potentially suppress lipid droplet accumulation in the differentiated 3T3-L1 adipocytes.
Structure, functionality and bioactivity of sulfated polysaccharide extracted from rainbow trout byproducts: pH-shift method vs enzymatic hydrolysis.
Green Sulfation of Arabinogalactan in the Melt of a Sulfamic Acid-Urea Mixture.
Pregnenolone Sulfate Permeation at the Blood-Brain Barrier is Independent of OATP2B1-In Vivo and In Vitro Insights.
A dual role for pleiotrophin in modulating inflammation and myelination in the presence of chondroitin sulfate proteoglycans after nervous system injury.
Human CSPG4-targeting CAR-macrophages inhibit melanoma growth.
Innovative injectable, self-healing, exosome cross-linked biomimetic hydrogel for cartilage regeneration.
Identification of the metabolomic alterations associated with the formation of bisphenol-A sulfate metabolite in HepG2 cells.
Light-Induced Conjugation of Inorganic Phosphate and Sulfate with Aminocyclobutenediones under Aqueous Conditions.
Febuxostat facilitates neovasculogenesis in chronic kidney disease through xanthine oxidase/NADPH oxidase/c-Jun signaling pathways.
Trastuzumab Decreases the Expression of G1/S Regulators and Syndecan-4 Proteoglycan in Human Rhabdomyosarcoma.
Profiling neuroactive steroids in pregnancy. A non-derivatised liquid chromatography tandem mass spectrometry method for the quantitation of allopregnanolone and four related isomers in maternal serum.

Spectrochim Acta A Mol Biomol Spectrosc. 2025 Mar 11. pii: S1386-1425(25)00340-3. [Epub ahead of print]336 126034

Highly selective peptide-based fluorescent probe with aggregation induced emission (AIE) for detection of chondroitin sulfate and its application in living cells and zebrafish imaging.

Xinlin Cao, Shiyang Li, Chunmei Pu, Weiliang Deng, Peng Wang, Yong An.

  Chondroitin sulfate (CS), as a kind of acid mucosaccharide with natural activity, has shown various physiological activities in human body due to its advantages of high biocompatibility, good degradability and little side effects. Herein, a novel and simple peptide probe (TPE-ASRH) based on tetraphenylethene (TPE) and tetrapeptide (Ala-Ser-Arg-His-NH2) was designed and synthesized. TPE-ASRH displayed remarkable aggregation induced emission (AIE) characteristic in DMSO/water binary mixture. Based on electrostatic attraction, TPE-ASRH displayed highly selective and sensitive detection to chondroitin sulfate with large Stokes shift (156 nm), and the limit of detection (LOD) for chondroitin sulfate was calculated to be 0.11 nM based on 3σ/k. In addition, the colour change of TPE-ASRH was observed significantly from midnightblue to steelblue after adding chondroitin sulfate using naked eyes under 365 nm UV irradiation. Meanwhile, TPE-ASRH was able to achieve a rapid response to chondroitin sulfate (less than 30 s) with a pH response range of 3-12, which indicated that TPE-ASRH can detect chondroitin sulfate rapidly under physiological conditions. The response mechanism of TPE-ASRH to chondroitin sulfate was demonstrated using Zeta particle size and potential, UV-vis titration spectroscopy, FTIR spectra and CD spectroscopy. Most importantly, TPE-ASRH revealed the considerably low cytotoxic effects and good biological permeability, and was successfully applied to image chondroitin sulfate in living cells and zebrafish.

Keywords:  AIE characteristic; Bioimaging; Chondroitin sulfate; Fluorescent probe; Tetrapeptide

DOI:  https://doi.org/10.1016/j.saa.2025.126034
Int J Biol Macromol. 2025 Mar 05. pii: S0141-8130(25)02359-1. [Epub ahead of print] 141808

Online PGC-LC-MS analysis of colonic mucin O-glycans in ovalbumin-induced food allergy in Balb/c mice by treatment with sea cucumber chondroitin sulfate polysaccharide.

Cheng Li, Yu Lu, Zhijun Zhang, Linjuan Huang, Zhongfu Wang.

  The highly sulfated polysaccharide sea cucumber chondroitin sulfate (SCCS) can alleviate intestinal damage and display strong anti-food-allergic activity. The O-glycopattern levels in colonic mucin are closely related to the its protective effect on function of the intestinal barrier. However, the effect of the SCCS on colonic mucin O-glycan has not been investigated. In this study, ovalbumin (OVA)-sensitized allergic mice and SCCS treatment were used. Mouse colonic mucin O-glycome was released and analyzed through reductive β-elimination combined with PGC-LC-MS. A total of presumptive 20 neutral and 28 acidic O-glycan structures were identified, in which the core 2 type acidic O-glycan structure is predominant in Balb/c female mice. Treatment with OVA and SCCS did not change the numbers of colon mucin O-glycan type, but the expression level of total O-glycosylation was more abundant in the SCCS group mice than in the OVA group (1.8-fold), especially for acidic O-glycans (co-modified by fucose and sulfate groups). Furthermore, supplementation with SCCS reversed most of the O-glycan decreasing trend, which may be associated with a return to healthy levels of gut microbiota. In conclusion, our results demonstrate that SCCS could restore colonic mucin O-glycosylation levels and intestinal homeostasis and contribute to enhancing intestinal barrier function.

Keywords:  Colonic mucin O-glycosylation; Food allergy; Sea cucumber chondroitin sulfate

DOI:  https://doi.org/10.1016/j.ijbiomac.2025.141808
Glycoconj J. 2025 Mar 10.

Cartilaginous fishes-derived chondroitin sulfates potentially suppress lipid droplet accumulation in the differentiated 3T3-L1 adipocytes.

Danang Dwi Cahyadi, Katsuhiko Warita, Naoko Takeda-Okuda, Jun-Ichi Tamura, Yoshinao Z Hosaka.

  In this study, we investigated for cell proliferative and adipogenic differentiation inhibitory activities of chondroitin sulfate (CS) from cartilaginous fish: mako shark (Isurus oxyrinchus, spine part, Ms-CS), blue shark (Prionace glauca, spine part, Bs-CS), sharpspine skate (Okamejei acutispina, head and tail parts, Sp-CS) and stingray (Dasyatis akajei, head part, St-CS) on 3T3-L1 cells. Most of the CSs from cartilaginous fish showed concentration-dependent cell proliferative activity of 3T3-L1 cells within the retrieved concentration range (0-1,000 μg/mL), while under induction of adipocyte differentiation, they inhibited lipid accumulation. In particular, Ms-CS and Sp-CS were highly active in inhibiting lipid accumulation in the cells. The present study revealed that cartilaginous fish-derived CS has inhibitory activity on 3T3-L1 adipocyte differentiation by suppressing lipid droplet accumulation, although the degree of suppression varied depending on the composition of the CS and its origin. In addition, a significant increase in chondroitin sulfate N-acetylgalactosaminyltransferase 2 (Csgalnact2) expression of the Sp-CS group at the concentration of 500 µg/mL was observed. Csgalnact2 expression is associated with chondroitin N-acetylgalactosaminyltransferase-2 (ChGn-2), one of the glycosyltransferases that catalyzes the chain initiation and elongation of the CS backbone in its biosynthesis. Exogenous CS from cartilaginous fishes increased Csgalnact2 expression, although further studies are needed to confirm changes in CS biosynthesis. We observed reduced lipid accumulation in differentiated 3T3-L1 cells. Our findings highlight the role of CS polysaccharides, in inhibiting adipogenesis, even though further investigation is required to understand the underlying mechanism.

Keywords:  3T3-L1; Adipogenesis; Chondrichthyes; Chondroitin sulfate; Fat; Glycosaminoglycans

DOI:  https://doi.org/10.1007/s10719-025-10183-0
Food Chem. 2025 Mar 05. pii: S0308-8146(25)00916-1. [Epub ahead of print]479 143665

Structure, functionality and bioactivity of sulfated polysaccharide extracted from rainbow trout byproducts: pH-shift method vs enzymatic hydrolysis.

Shahab Naghdi, Masoud Rezaei, Mehdi Tabarsa, Mehdi Abdollahi.

  Here, a novel method for sequentially extracting sulfated polysaccharides (SPs) from Oncorhynchus mykiss byproducts using alkaline/acid solubilization followed by isoelectric precipitation is compared with conventional enzymatic hydrolysis. Alkaline solubilization (SP-Alk) yielded SPs (2.46 %) comparable to the enzymatic method (SP-Enz, 2.77 %), while acidic solubilization (SP-Aci) yielded 1.96 %. SP-Alk showed comparable carbohydrate and sulfate content but lower protein than SP-Enz. Additionally, SP-Alk showed the highest monosaccharide content of rhamnose, mannose, glucose, and galactose. The extraction method affected the Molecular weight of SPs with SP-Enz having the lowest (44.95 kDa). Structural and thermal properties of the SPs were similar as revealed by FTIR/XRD and DSC, respectively. While SP-Enz exhibited slightly better antioxidant and functional properties (foaming, stability, emulsifying activity), SP-Alk showed a considerable performance with similar antimicrobial activity. Altogether, the pH-shift method can be a promising alternative for sequential extraction of SPs compared with enzymatic hydrolysis, avoiding enzymatic degradation of proteins.

Keywords:  Bioactivity; Biorefinery; Functional properties; Marine side streams; Sulfated polysaccharides

DOI:  https://doi.org/10.1016/j.foodchem.2025.143665
Polymers (Basel). 2025 Feb 27. pii: 642. [Epub ahead of print]17(5):

Green Sulfation of Arabinogalactan in the Melt of a Sulfamic Acid-Urea Mixture.

Vladimir A Levdansky, Alexander V Levdansky, Yuriy N Malyar, Timur Yu Ivanenko, Olga Yu Fetisova, Aleksandr S Kazachenko, Boris N Kuznetsov.

  Sulfation of arabinogalactan (AG) from larch wood (Larix sibirica Ledeb.) in the melt of a sulfamic acid-urea mixture has been first examined. The impact of the AG sulfation temperature on the AG sulfate yield and the sulfur content has been established. The high sulfur content (11.3-11.6%) in sulfated AG has been obtained in the temperature range of 115-120 °C for a sulfation time of 0.5 h. The process effectively prevents molecular degradation under these conditions. The incorporation of sulfate groups into the arabinogalactan structure has been confirmed by the appearance of absorption bands in the FTIR spectrum that are typical of sulfate group vibrations. The 13C NMR spectroscopy study has proven that the AG sulfation in the melt of a sulfamic acid-urea mixture leads to the substitution of some free hydroxyl groups for C6, C4, and C2 carbon atoms of the AG β-D-galactopyranose units. The advantage of the proposed AG sulfation method is that the reaction occurs without solvent, and the reaction time is only 0.5 h. The kinetics of the thermal decomposition of the initial AG and sulfated AG samples have been studied. It has been found that the sulfated AG samples have a lower thermal resistance than the initial AG. The kinetic analysis has revealed a decrease in the activation energy of the thermal degradation of the sulfated samples as compared to the initial AG.

Keywords:  arabinogalactan; green sulfation; melt; sulfamic acid–urea mixture; sulfated arabinogalactan

DOI:  https://doi.org/10.3390/polym17050642
Biopharm Drug Dispos. 2025 Mar 12.

Pregnenolone Sulfate Permeation at the Blood-Brain Barrier is Independent of OATP2B1-In Vivo and In Vitro Insights.

Valerio Taggi, Anima M Schäfer, Stefan Oswald, Jonny Hanna Kinzi, Isabell Seibert, Alaa Al-Khatib, Henriette E Meyer Zu Schwabedissen.

  Sulfated steroids such as pregnenolone sulfate (PregS) are important for neuronal development and cognitive functions. Given the hydrophilic sulfate moiety, it is assumed that PregS requires an active transport mechanism to cross the blood-brain barrier (BBB). The human organic anion transporting polypeptide (OATP)2B1 has been previously shown to transport sulfated steroids and is therefore a proposed candidate for the transport of PregS. In this study, we aimed to investigate the role of OATP2B1 in the uptake of PregS in the brain. Tritium-labeled PregS was intravenously administered to humanized (SLCO2B1+/+), knockout (rSlco2b1-/-), and wildtype (WT) rats. Accumulation of the radiotracer was analyzed in rat brain, liver, small intestine, kidney, heart, and muscle. Moreover, transporter expression in brain microvessels was assessed through targeted proteomics and Western blot analysis. The involvement of hOATP2B1 in PregS transport across the BBB was further studied using a hBMEC-based in vitro BBB model. Our results indicated no significant difference in accumulation of the radiotracer among the different rat genotypes in the brain or in other tissues. In line with what we observed in the rat model, the subsequent in vitro study showed no involvement of hOATP2B1 in the transport of PregS. Taken together, our findings highlight the species-specific differences in transporter expression and suggest that OATP2B1 does not mediate PregS uptake across the BBB.

Keywords:  OATP2B1; blood–brain barrier; blood–brain barrier model; pregnenolone sulfate; rat model

DOI:  https://doi.org/10.1002/bdd.70002
Front Cell Neurosci. 2025 ;19 1549433

A dual role for pleiotrophin in modulating inflammation and myelination in the presence of chondroitin sulfate proteoglycans after nervous system injury.

Somnath J Gupta, Matthew A Churchward, Kathryn G Todd, Ian R Winship.

  Chondroitin sulfate proteoglycans (CSPGs), key components of the extracellular matrix and the glial scar that forms around central nervous system (CNS) injuries, are recognized for hindering neuronal regeneration. We previously demonstrated the potential of pleiotrophin (PTN) to induce neurite outgrowth even in the presence of inhibitory CSPGs. The effects of PTN on microglia and oligodendrocytes are not well described. Here, we examined how PTN administration alters the differentiation of oligodendrocyte precursor cells (OPCs) into mature oligodendrocytes in the presence of CSPGs using in vitro cell culture model. Moreover, we explored the effects of PTN on the inflammatory activity of microglia with and without inflammatory stimulation (IFN-γ) in a CSPG-rich environment. The data showed that the CSPG matrix inhibited the differentiation of OPCs into mature oligodendrocytes. PTN induced dose-dependent differentiation of OPCs into mature oligodendrocytes, with an optimal effect at 10 nM PTN. Moreover, PTN modified the immunological response of microglia in the presence of CSPGs, with reduced proinflammatory activity that was further reduced by PTN administration, in contrast to the increased release of matrix metalloproteinases (MMP 9). However, when IFN-γ-activated microglia were treated with PTN, proinflammatory signaling was stimulated at higher PTN concentrations (10 nM and 100 nM). Overall, our results indicate that PTN can overcome the inhibitory effect of CSPGs on the differentiation of OPCs into oligodendrocytes and can modulate inflammation mediated by CSPGs from microglia. Collectively, these findings demonstrate that PTN can effectively counteract the inhibitory effects of CSPGs on the differentiation of OPCs into mature oligodendrocytes while also modulating microglial responses to reduce proinflammatory activity and increase MMP-9 release. Thus, PTN has great potential to improve remyelination and neuroprotective strategies in the treatment of demyelinating diseases or any injury.

Keywords:  CSPGs; OPCs; PTN; microglia; oligodendrocytes; stroke

DOI:  https://doi.org/10.3389/fncel.2025.1549433
Oncogene. 2025 Mar 13.

Human CSPG4-targeting CAR-macrophages inhibit melanoma growth.

Daniel Greiner, Qian Xue, Trinity Qa Waddell, Elena Kurudza, Piyush Chaudhary, Rachel L Belote, Gianpietro Dotti, Robert L Judson-Torres, Melissa Q Reeves, Samuel H Cheshier, Minna Roh-Johnson.

Approximately half of melanoma patients relapse or fail to respond to current standards of care, highlighting the need for new treatment options. Engineering T-cells with chimeric antigen receptors (CARs) has revolutionized the treatment of hematological malignancies but has been clinically less effective in solid tumors. We therefore sought to engineer alternative immune cell types to inhibit melanoma progression. Engineering macrophages with CARs has emerged as a promising approach to overcome some of the challenges faced by CAR-T cells; however, whether these engineered macrophages can effectively inhibit melanoma growth is unknown. To determine whether CAR-macrophages (CAR-Ms) specifically target and kill melanoma cells, we engineered CAR-Ms targeting chondroitin sulfate proteoglycan 4 (CSPG4), an antigen expressed in melanoma. CSPG4-targeting CAR-Ms exhibited specific phagocytosis of CSPG4-expressing melanoma cells. We developed 3D approaches to show that CSPG4-targeting CAR-Ms efficiently infiltrated melanoma spheroids. Furthermore, combining CSPG4-targeting CAR-Ms with strategies inhibiting CD47/SIRPα "don't eat me" signaling synergistically enhanced CAR-M-mediated phagocytosis and robustly inhibited melanoma spheroid growth in 3D. Importantly, CSPG4-targeting CAR-Ms inhibited melanoma tumor growth in mouse models. These results suggest engineering macrophages against melanoma antigens is a promising solid tumor immunotherapy approach for treating melanoma.

DOI: https://doi.org/10.1038/s41388-025-03332-0
J Control Release. 2025 Mar 05. pii: S0168-3659(25)00217-2. [Epub ahead of print]381 113608

Innovative injectable, self-healing, exosome cross-linked biomimetic hydrogel for cartilage regeneration.

Chenlin Tu, Xiang Gao, Hong Zheng, Rui Huang, Fengkai Yang, Yeying Dong, Kaipeng Jing, Thomas Groth, Mingyan Zhao.

  The limited self-healing capacity of cartilage hinders its repair and regeneration at the defect sites. Recent research into small-molecular compounds has shown promise in achieving a better regeneration of cartilage. In this study, we encapsulate kartogenin (KGN) and transforming growth factor β1 (TGF-β1) within mesenchymal stem cells derived exosomes (EKT), and then coated them with succinylated chitosan (sCH) to create positively charged exosomes, termed CEKT. These CEKT exhibit exceptional chondrogenic promoting capabilities shown during in vitro studies with bone marrow derived mesenchymal stem cells (BMSCs). They also can penetrate deep into cartilage tissue derived from porcine knee joints guided by their positive charge. Subsequently, a dynamic exosomes-crosslinked hydrogel (Gel-CEKT) is fabricated by crosslinking CEKT with oxidized chondroitin sulfate (oCS) and Wharton's jelly (WJ) through imine bond formation. Physicochemical studies revealed the injectability, excellent adhesive, and self-healing abilities of this hydrogel, which enables minimally invasive and precise treatment of cartilage defects, assisted by the enriched and sustained administration of CEKT. In vitro cell experiments show that Gel-CEKT can efficiently recruit BMSCs and significantly promote the gene expression of Sox9 and protein expression of collagen II and aggrecan. Furthermore, we show in a rat model of cartilage defect that the Gel-CEKT demonstrates superior performance compared to Gel@EKT, which has freely encapsulated exosomes in the hydrogel. The freely encapsulated exosomes are rapidly released, whereas the exosome-crosslinked gel structure ensures sustained retention and functionality at the site of defect. This leads to impressive outcomings, including extensive new cartilage tissue formation, a smoother cartilage surface, significant chondrocyte production, and seamless integration with orderly and continuous structure formation between cartilage and subchondral bone. This study underscores the potential of exosomes-crosslinked hydrogels as a novel and promising therapeutic approach for clinical cartilage regeneration.

Keywords:  Cartilage regeneration; Charge guidance; Dynamic hydrogel; Exosomes; Mesenchymal stem cells; Sustained release

DOI:  https://doi.org/10.1016/j.jconrel.2025.113608
Food Chem Toxicol. 2025 Mar 07. pii: S0278-6915(25)00149-8. [Epub ahead of print]200 115382

Identification of the metabolomic alterations associated with the formation of bisphenol-A sulfate metabolite in HepG2 cells.

Hui-Jun Jin, Zi-Dong Qiu, Cen Qian, Chun-Yun Zhang, Yu Peng.

  The elucidation of the causal relationship between bisphenol-A (BPA) exposure and hepatoxic outcomes is challenging because of the complexity in both the BPA-derived metabolites formed in the liver and the associated endogenous molecular responses. We performed parallel metabolism experiments with BPA to characterize the BPA sulfate formation and the associated alterations in the metabolome level in HepG2 cells using mass spectrometry-based metabolome wide association study. Briefly, HepG2 cells were exposed for 8 or 24 h to 1 or 10 μM BPA in DMSO or DMSO alone. The levels of BPA sulfate in the cell culture media were quantified, and the sulfation efficiency was about 0.4 % observed for both 1 and 10 μM BPA in HepG2 cells. Targeted metabolomic analyses revealed alterations belonging to forty metabolic pathways following BPA exposure. Featured by the decreasing of estrone sulfate, estrogen metabolism was observed as the top 1 enriched pathway in response to BPA exposure. MWAS suggests that BPA sulfate formation in HepG2 cells resulted in vitamin B6 deficiency and dysregulated vitamin B6-dependent processes, for example, the kynurenine pathway in tryptophan metabolism. These findings collectively provide insights into the linkage between exogenous and endogenous metabolism and the potential initial events in BPA exposure-relevant hepatoxicity.

Keywords:  BPA sulfate; HepG2 cells; Metabolome wide association study; Targeted metabolomics; Tryptophan metabolism; Vitamin B6 deficiency

DOI:  https://doi.org/10.1016/j.fct.2025.115382
Chemistry. 2025 Mar 11. e202500187

Light-Induced Conjugation of Inorganic Phosphate and Sulfate with Aminocyclobutenediones under Aqueous Conditions.

Koki Ogawa, Rena Kobayashi, Nanami Okada, Kenji Mishiro.

  Phosphate and sulfate groups play an important role in controlling the physical properties of biomolecules and artificial materials. However, despite their significance, the incorporation of phosphate or sulfate groups into aqueous organic compounds using non-enzymatic methods has been unprecedented. In this study, we have successfully conjugated inorganic phosphate and sulfate with aminocyclobutenedione derivatives via photochemical reactions under aqueous conditions. A plausible mechanism for the conjugation reactions was proposed on the basis of DFT calculations. These reactions will promote life science research based on organophosphates and organosulfates which have been difficult to access up to now.

Keywords:  Aqueous Conditions; Cyclobutenedione; Sulfate; phosphate; photochemical reaction

DOI:  https://doi.org/10.1002/chem.202500187
Biomed Pharmacother. 2025 Mar 10. pii: S0753-3322(25)00146-5. [Epub ahead of print]185 117952

Febuxostat facilitates neovasculogenesis in chronic kidney disease through xanthine oxidase/NADPH oxidase/c-Jun signaling pathways.

Hsin-Jou Lee, Chih-Hung Chiang, Jung-Hung Hsieh, Su-Chu Lin, Jaw-Wen Chen, Ting-Ting Chang.

  The accumulation of uremic toxins in circulation contributes to the cardiovascular diseases that result from chronic kidney disease (CKD). Indoxyl sulfate (IS), which is a protein-bound uremic toxin, promotes cardiovascular diseases with impaired neovascularization by increasing the reactive oxygen species (ROS). This study aimed to investigate febuxostat, a potent xanthine oxidase (XO) inhibitor, for its potential effects on the mechanisms of neovasculogenesis in CKD. CKD mice were generated by 5/6 subtotal nephrectomy and orally administered with febuxostat. Human aortic endothelial cells (HAECs) were used and treated with IS to simulate the CKD conditions in vitro. In the CKD mice, febuxostat reduced systemic ROS and preserved kidney function, as evidenced by the reduced levels of serum blood urea nitrogen, creatinine, urinary albumin-to-creatinine ratios, and renal inflammatory proteins. Furthermore, febuxostat improved neovasculogenesis in an aortic ring assay, a Matrigel plug assay, and a wound healing assay, as evidenced by increased microvascular sprouting in the aortic rings, hemoglobin contents, and capillary density in the CKD mice. In IS-stimulated HAECs, the antioxidative, pro-angiogenic, and anti-inflammatory effects of febuxostat enhanced the tube formation and migration abilities via the XO/p47/c-Jun signaling pathways. In summary, febuxostat might provide renal protection and facilitate neovasculogenesis in CKD. Further clinical study may need to be conducted to verify the effects of febuxostat in CKD patients with vascular complications.

Keywords:  Chronic kidney disease; Endothelial cells; Febuxostat; Neovasculogenesis; Xanthine oxidase

DOI:  https://doi.org/10.1016/j.biopha.2025.117952
Int J Mol Sci. 2025 Feb 27. pii: 2137. [Epub ahead of print]26(5):

Trastuzumab Decreases the Expression of G1/S Regulators and Syndecan-4 Proteoglycan in Human Rhabdomyosarcoma.

Dora Julianna Szabo, Eniko Toth, Kitti Szabo, Zsofia Kata Hegedus, Noemi Bozsity-Farago, Istvan Zupko, Laszlo Rovo, Xue Xiao, Lin Xu, Aniko Keller-Pinter.

  Rhabdomyosarcoma (RMS), the most common soft tissue sarcoma in children, arises from skeletal muscle cells that fail to differentiate terminally. Two subgroups of RMS, fusion-positive and fusion-negative RMS (FPRMS and FNRMS, respectively), are characterized by the presence or absence of the PAX3/7-FOXO1 fusion gene. RMSs frequently exhibit increased expression of human epidermal growth factor receptor-2 (HER2). Trastuzumab is a humanized monoclonal antibody targeting HER2, and its potential role in RMS treatment remains to be elucidated. Syndecan-4 (SDC4) is a heparan sulfate proteoglycan (HSPG) affecting myogenesis via Rac1-mediated actin remodeling. Previously, we demonstrated that the SDC4 gene is amplified in 28% of human FNRMS samples, associated with high mRNA expression, suggesting a tumor driver role. In this study, after analyzing the copy numbers and mRNA expressions of other HSPGs in human RMS samples, we found that in addition to SDC4, syndecan-1, syndecan-2, and glypican-1 were also amplified and highly expressed in FNRMS. In RD (human FNRMS) cells, elevated SDC4 expression was accompanied by low levels of phospho-Ser179 of SDC4, leading to high Rac1-GTP activity. Notably, this high SDC4 expression in RD cells decreased following trastuzumab treatment. Trastuzumab decreased the levels of G1/S checkpoint regulators cyclin E and cyclin D1 and reduced the cell number; however, it also downregulated the cyclin-dependent kinase inhibitor p21. The level of MyoD, a transcription factor essential for RMS cell survival, also decreased following trastuzumab administration. Our findings contribute to the understanding of the role of SDC4 in FNRMS. Since HER2 is expressed in about half of RMSs, the trastuzumab-mediated changes observed here may have therapeutic implications.

Keywords:  MyoD; cell cycle; proteoglycan; rhabdomyosarcoma; syndecan-4; trastuzumab

DOI:  https://doi.org/10.3390/ijms26052137
J Chromatogr B Analyt Technol Biomed Life Sci. 2025 Feb 27. pii: S1570-0232(25)00093-5. [Epub ahead of print]1256 124541

Profiling neuroactive steroids in pregnancy. A non-derivatised liquid chromatography tandem mass spectrometry method for the quantitation of allopregnanolone and four related isomers in maternal serum.

Edward Hinchliffe, Alexander Heazell.

  Neuroactive steroids are metabolites of progesterone, synthesised during pregnancy by the placenta. Here, we describe development of a novel liquid chromatography tandem mass spectrometry (LC-MS/MS) assay for quantitation of allopregnanolone, pregnanolone, isopregnanolone, epipregnanolone and allopregnan-20α-ol-3-one in maternal serum. Following addition of deuterated internal standards, 200 μL of serum was subjected to solid phase extraction. Chromatography was performed using a pentafluorophenyl column, and LC-MS/MS on a Sciex 6500+. Sample injection volume was 20 μL, and injection-to-injection time 10.0 min. The assay was validated according to published guidelines; assay linearity and lower limit of quantification were suitable for analysis of each steroid in maternal serum, for all analytes mean recoveries were 100 % ± 15 %, intra- and inter-assay imprecision <15 %, and matrix effects negligible, and specificity experiments confirmed nil interference from a wide range of endogenous metabolites of progesterone. The method was applied to human serum samples obtained from a large cohort of third trimester pregnancies which were subsequently characterised by normal fetal and maternal outcomes, and relationships between maternal neuroactive steroid concentrations and fetal gestational age assessed. Positive correlations between maternal serum concentration and fetal gestational age were observed for isopregnanolone, allopregnanolone and allopregnan-20α-ol-3-one. The LC-MS/MS method offers significant advantages over previously published approaches for quantitation of neuroactive steroids in human maternal serum, notably obviating the need for derivatisation, whilst achieving exceptional specificity. Characterisation of normal maternal neuroactive steroid concentrations will aid future research as dysregulated placental progesterone metabolism is observed in pregnancies with poor outcomes.

Keywords:  Allopregnanolone; Epipregnanolone; Isopregnanolone; Liquid chromatography tandem mass spectrometry; Neuroactive steroids; Pregnanolone

DOI:  https://doi.org/10.1016/j.jchromb.2025.124541