bims-supasi 2022-05-08 papers

bims-supasi

Biomed News

on Sulfation pathways and signalling

Issue of 2022–05–08
nineteen papers selected by
Jonathan Wolf Mueller, University of Birmingham

N-acetylneuraminic acid and chondroitin sulfate modified nanomicelles with ROS-sensitive H2S donor via targeting E-selectin receptor and CD44 receptor for the efficient therapy of atherosclerosis.
Building Block Analysis of ATIII Affinity Fractions of Heparins: Application to the ATIII Binding Capacity of Non-conventional 3-O-Sulfated Sequences.
The interaction between oxytocin and heparin.
Sulfation Pathways During Neurodevelopment.
The therapeutic potential of chondroitin sulfate in Aspergillus fumigatus keratitis.
Nanozyme-natural enzymes cascade catalyze cholesterol consumption and reverse cancer multidrug resistance.
PCB Sulfates in Serum from Mothers and Children in Urban and Rural U.S. Communities.
Sulfated Naphthopyrones and Anthraquinones from the Vietnamese Crinoid Comanthus delicata.
Oxysterols and Oxysterol Sulfates in Alzheimer's Disease Brain and Cerebrospinal Fluid.
Heparan sulfate mimetic fucoidan restores the endothelial glycocalyx and protects against dysfunction induced by serum of COVID-19 patients in the intensive care unit.
Soluble α-klotho and heparin modulate the pathologic cardiac actions of fibroblast growth factor 23 in chronic kidney disease.
Development and Validation of an LC-MS/MS-Based Method for Quantifying Urinary Endogenous 6-Hydroxymelatonin.
Human milk glycosaminoglycans inhibit cytomegalovirus and respiratory syncytial virus infectivity by impairing cell binding.
The Effects of Sacran, a Sulfated Polysaccharide, on Gut Microbiota Using Chronic Kidney Disease Model Rats.
Investigation of the individual human sulfatome in plasma and urine samples reveals an age-dependency.
Structural insights into choline-O-sulfatase reveal the molecular determinants for ligand binding.
Short-term high-fat diet alters acetaminophen metabolism in healthy individuals.
The N-Terminus of Human Sulfotransferase 2B1b─a Sterol-Sensing Allosteric Site.
Differential genetic associations and expression of PAPST1/SLC35B2 in bipolar disorder and schizophrenia.

Int J Biol Macromol. 2022 May 02. pii: S0141-8130(22)00894-7. [Epub ahead of print]

N-acetylneuraminic acid and chondroitin sulfate modified nanomicelles with ROS-sensitive H2S donor via targeting E-selectin receptor and CD44 receptor for the efficient therapy of atherosclerosis.

Qiang Chen, Chunjing Guo, Xiudi Zhou, Yanguo Su, Huimin Guo, Min Cao, Jing Li, Yue Zhang, Weiyi Zhao, Xin Gao, Shuqi Mi, Daquan Chen.

  Currently, very limited therapeutic approaches are available for the drug treatment of atherosclerosis(AS). H2S-donor is becoming a common trend in much life-threatening research. Several studies have documented that H2S-lyase is predominantly present in endothelial cells. N-Acetylneuraminic acid (SA), natural carbohydrate, binds specifically to the E-selectin receptor of endothelial cells. Meanwhile, recent studies related to Chondroitin sulfate have excellent target binding ability with CD44 receptor. We conjecture that the N-Acetylneuraminic acid and Chondroitin sulfate modified nanomicelles not only enhances the accumulation of the drug but also cleaves the H2S donor in the lesion, thus one stone two birds. Given these findings, we synthesized two kinds of nanoparticles, Carrier I (SCCF) and Carrier II (SCTM), for atherosclerosis to validate our guesses. Initially, S-allyl-L-cysteine and 4-methoxyphenylthiourea were used as H2S donors for SCCF and SCTM, respectively. After the introduction of ROS-sensitive groups. Then, micelles with N-Acetylneuraminic acid and Chondroitin sulfate were prepared to load rapamycin(RAP). Further, in atherosclerosis Oil Red O staining (ORO) results confirmed remarkable treatment effect with SCCF@RAP and SCTM@RAP. Thus, we conclude that the effect of dual-targeting nanomicelles with ROS-sensitive H2S donor based on N-Acetylneuraminic acid and Chondroitin sulfate will have a better role in atherosclerosis.

Keywords:  Anti-atherosclerosis; CD44 receptor; Chondroitin sulfate; E-selectin receptor; N-Acetylneuraminic acid; ROS-sensitive H(2)S donor

DOI:  https://doi.org/10.1016/j.ijbiomac.2022.04.180
Front Med (Lausanne). 2022 ;9 841738

Building Block Analysis of ATIII Affinity Fractions of Heparins: Application to the ATIII Binding Capacity of Non-conventional 3-O-Sulfated Sequences.

Pierre Mourier.

  In heparin, some 3-O-sulfated sequences do not meet the structural requirements of the ATIII binding pentasaccharide. These "non-conventional" sequences are the object of this study. In a previous paper (Mourier P. Heparinase digestion of 3-O-sulfated sequences: selective heparinase II digestion for separation and identification of binding sequences present in ATIII affinity fractions of bovine intestine heparins), we demonstrated that unsaturated 3-O-sulfated disaccharides detected in exhaustive heparin digests were specifically cleaved by heparinase I. Consequently, building blocks analyses of heparins using heparinases I+II+III digestion could be compared with experiments where only heparinase II is used. In these latter conditions of depolymerization, the 3-O-sulfated sequences digested into unsaturated 3-O-sulfated disaccharides with heparinases I+II+III, were heparinase II-resistant on their non-reducing side, resulting in longer new building blocks. These properties were used to study the structural neighborhood of these 3-O-sulfated moieties, which have still-undefined biological functions. In this part, heparinases I+II+III and heparinase II digestions of porcine mucosa, bovine mucosa and bovine lung heparins were compared in six fractions of increasing affinity for ATIII. Tagging of building blocks by reductive amination with sulfanilic acid was used. The distribution of 3-O-sulfated building blocks in the ATIII affinity fractions was used to examine the ATIII binding of these sequences.

Keywords:  3-O-sulfated disaccharides; ATIII affinity; heparin; heparinase digestion; sulfanilic tagging

DOI:  https://doi.org/10.3389/fmed.2022.841738
RSC Adv. 2020 Jul 27. 10(47): 28300-28313

The interaction between oxytocin and heparin.

Einat Schnur, Timothy R Rudd.

Oxytocin (OXT) is a small cyclic peptide that is administered to pregnant women to induce birth in cases where labour is prolonged. It has previously been observed that patients taking a low molecular weight heparin, dalteparin (DAL), and then prescribed, OXT experienced a swifter labour compared to women given OXT alone. Herein are described the interactions between OXT and a number of heparin-based oligosaccharides; DAL; fondaparinux (FP), which is a synthetic heparin oligosaccharide that represents the predominant antithrombin binding-site, and a family of chemically-derived heparin hexasaccharides. The latter oligosaccharides were chosen as they represent sequences found within the polysaccharide dalteparin. Furthermore, the carbohydrate chemical space was investigated by comparing the interaction between OXT and four chemically derivatived heparin hexasaccharides; I2S-A6S NS (DP6), I2OH-A6S NS (DP6-2OH, de-2-O-sulfated hexasaccharide), I2S-A6OH NS (DP6-6OH, de-6-O-sulfated hexasaccharide) and I2S-A6S NAc (DP6-NAc, de-N-sulfated hexasaccharide). The interactions between the peptide and oligosaccharides were studied using a series of 13C-1H and 15N-1H HSQC NMR experiments, at a range of temperatures. This approach allowed the binding epitopes of the peptide and oligosaccharides to be identified, highlighting that 6-O- and N-sulfation substituent groups of heparin are important for the interaction between the peptide and carbohydrate. This is an important observation as de-N-sulfation is a traditional method for decreasing the anticoagulation properties of heparin. Furthermore, low temperature experiments of the OXT : FP complex indicate that hydrogen-bonding is very important for the interaction between the peptide and oligosaccharide.

DOI: https://doi.org/10.1039/d0ra04204h
Front Mol Biosci. 2022 ;9 866196

Sulfation Pathways During Neurodevelopment.

Taylor Clarke, Francesca E Fernandez, Paul A Dawson.

  Sulfate is an important nutrient that modulates a diverse range of molecular and cellular functions in mammalian physiology. Over the past 2 decades, animal studies have linked numerous sulfate maintenance genes with neurological phenotypes, including seizures, impaired neurodevelopment, and behavioral abnormalities. Despite sulfation pathways being highly conserved between humans and animals, less than one third of all known sulfate maintenance genes are clinically reportable. In this review, we curated the temporal and spatial expression of 91 sulfate maintenance genes in human fetal brain from 4 to 17 weeks post conception using the online Human Developmental Biology Resource Expression. In addition, we performed a systematic search of PubMed and Embase, identifying those sulfate maintenance genes linked to atypical neurological phenotypes in humans and animals. Those findings, together with a search of the Online Mendelian Inheritance in Man database, identified a total of 18 candidate neurological dysfunction genes that are not yet considered in clinical settings. Collectively, this article provides an overview of sulfate biology genes to inform future investigations of perturbed sulfate homeostasis associated with neurological conditions.

Keywords:  brain; embryological; fetal; gene expression; neurological dysfunction; sulfate

DOI:  https://doi.org/10.3389/fmolb.2022.866196
Mol Immunol. 2022 Apr 30. pii: S0161-5890(22)00193-6. [Epub ahead of print]147 50-61

The therapeutic potential of chondroitin sulfate in Aspergillus fumigatus keratitis.

Junjie Luan, Xudong Peng, Jing Lin, Yingxue Zhang, Xue Tian, Lu Zhan, Guiqiu Zhao.

   PURPOSE: To explore the therapeutic effect of chondroitin sulfate (CS) on Aspergillus fumigatus (A. fumigatus) keratitis.
METHODS: The nontoxic concentration of CS was determined using the Draize eye test and cell counting kit-8. Cell scratch test and cell proliferation test were evaluated the impact of CS on the proliferation and migration of human corneal epithelial cells (HCECs). Adherence assay and plate counting were used to detect fungal load in vivo and in vitro, respectively. Clinical score and hematoxylin-eosin (HE) staining were applied to assess the therapeutic effects of CS in an A. fumigatus keratitis mice model. The neutrophil infiltration and activity were detected by flow cytometry (FCM), immunofluorescence staining, and myeloperoxidase (MPO) assay. Toll-like receptor 4 (TLR-4) expression in RAW 264.7 cells and mouse cornea was detected by immunofluorescence staining. Real-time PCR (RT-PCR), western blot, and enzyme-linked immunosorbent assay (ELISA) were applied to examine the expression of inflammatory mediators.
RESULTS: CS at 400 μg/mL (non-cytotoxic) significantly promoted proliferation and migration of HCECs. In an A. fumigatus keratitis mice model, CS treatment alleviated fungal keratitis (FK) severity by decreasing corneal fungal load and inhibiting neutrophil infiltration. In RAW 264.7 cells, the mRNA and protein levels of TLR-4, phosphorylated nuclear factor (NF)-κB (p-NF-κB), interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-a (TNF-α), cyclooxygenase 2 (COX-2), and macrophage inflammatory protein-2 (MIP-2) were remarkably lower in the siTLR-4 treated group, while higher in rTLR-4 treated group than in the corresponding control group. CS treatment suppressed rTLR-4 induced the mRNA and protein expression of TLR-4, p-NF-κB, IL-1β, IL-6, COX-2, TNF-α, and MIP-2 in RAW cells.
CONCLUSION: CS may ameliorate the prognosis of A. fumigatus keratitis by promoting corneal epithelial proliferation, inhibiting the recruitment and activity of neutrophils, and inhibiting the inflammatory response by down-regulation of the TLR-4/NF-κB signaling.

Keywords:  Anti-inflammatory; Chondroitin sulfate (CS); Fungal keratitis; TLR-4/NF-κB

DOI:  https://doi.org/10.1016/j.molimm.2022.04.007
J Nanobiotechnology. 2022 May 02. 20(1): 209

Nanozyme-natural enzymes cascade catalyze cholesterol consumption and reverse cancer multidrug resistance.

Bin Du, Mei Zheng, Huizhen Ma, Jingshu Huang, Qingqing Jiao, Yimeng Bai, Mengmeng Zhao, Jie Zhou.

  Multidrug resistance is still a major obstacle to cancer treatment. The most studies are to inhibit the activity of the drug transporter P-glycoprotein (P-gp), but the effect is not ideal. Herein, a nanosystem was built based on cascade catalytic consumption of cholesterol. Cholesterol oxidase (natural enzyme, COD) was immobilized on the carrier (NH2-MIL-88B, MOF) through amide reaction, COD catalyzed the consumption of cholesterol, the reaction product H2O2 was further produced by the MOF with its peroxidase-like activity to produce hydroxyl radicals (•OH) with killing effect. Due to the high expression of CD44 receptor on the surface of tumor cells, we encapsulated chondroitin sulfate gel shell (CS-shell) with CD44 targeting and apoptosis promoting effect on the surface of DOX@MOF-COD nanoparticles, which can accurately and efficiently deliver the drugs to the tumor site and improve the effect of reversing drug resistance. Taking drug-resistant cell membrane as "breakthrough", this paper will provide a new idea for reversing multidrug resistance of tumor.

Keywords:  Apoptosis; Cholesterol; Cholesterol oxidase; Chondroitin sulfate; Fluidity; Lipid raft; Reverse drug resistance

DOI:  https://doi.org/10.1186/s12951-022-01406-9
Environ Sci Technol. 2022 May 02.

PCB Sulfates in Serum from Mothers and Children in Urban and Rural U.S. Communities.

Duo Zhang, Panithi Saktrakulkla, Rachel F Marek, Hans-Joachim Lehmler, Kai Wang, Peter S Thorne, Keri C Hornbuckle, Michael W Duffel.

  Serum samples from 24 subjects (6 mother-daughter and 6 mother-son dyads) in a rural community (Columbus Junction, Iowa) and 24 subjects (6 mother-daughter and 6 mother-son dyads) in an urban community (East Chicago, Indiana) were analyzed for 74 sulfated metabolites of polychlorinated biphenyls (PCBs). We detected significantly higher mean concentrations of total assessed PCB sulfates in the urban group (110-8900 ng/g fresh weight of serum, mean = 3400 ng/g, standard error = 300) than in the rural cohort (530-6700 ng/g fresh weight of serum, mean = 1800 ng/g, standard error = 500). Eight PCB sulfate congeners (4-PCB 2 sulfate, 4'-PCB 2 sulfate, 2'-PCB 3 sulfate, 4'-PCB 3 sulfate, 4-PCB 11 sulfate, 4'-PCB 18 sulfate, 4'-PCB 25 sulfate, and 4-PCB 52 sulfate) contributed over 90% of the total assessed PCB sulfates in most individuals. The serum samples were enriched in PCB sulfates with fewer than 5 chlorine atoms, and this congener distribution differed from those of PCBs and hydroxylated PCBs in previous studies in the same communities. Regression analysis indicated several significant congener-specific correlations in mother-child dyads, and these relationships differed by location and by mother-daughter or mother-son dyads. This is the first study reporting a broad range of PCB sulfates in populations from urban and rural areas.

Keywords:  PCB metabolites; PCB sulfates; PCBs; human serum; organic pollutants

DOI:  https://doi.org/10.1021/acs.est.2c00223
Chem Pharm Bull (Tokyo). 2022 ;70(5): 408-412

Sulfated Naphthopyrones and Anthraquinones from the Vietnamese Crinoid Comanthus delicata.

Le Thi Vien, Tran Thi Hong Hanh, Tran Hong Quang, Do Cong Thung, Nguyen Xuan Cuong, Nguyen Hoai Nam, Pham Van Cuong, Phan Van Kiem, Chau Van Minh.

  Three new sulfated naphthopyrone derivatives namely delicapyrons F-H (1-3) and two new sulfated anthraquinone derivatives namely delicaquinons A (4) and B (5), together with 6-methoxycomaparvin-5-methylether-8-O-sodium sulfate (6), 6-methoxycomaparvin-8-O-sodium sulfate (7), comaparvin-8-O-sodium sulfate (8), and 3-propyl-1,6,8-trihydroxy-9,10-anthraquinone-6-O-sodium sulfate (9) were isolated from the Vietnamese crinoid Comanthus delicata. Their chemical structures were elucidated by extensive analysis of the one dimensional (1D) and 2D-NMR, high resolution electrospray ionization quadrupole time-of-flight (HR-ESI-QTOF) mass spectra as well as calculation of optical rotation. In addition, significant cytotoxicity was observed for 6 against LNCaP (prostate cancer) cell line with IC50 value of 20.29 ± 2.43 µM, whereas moderate or weak cytotoxic effects were observed for 1-3 and 5-8 on SK-Mel-2 (melanoma) cell line and 7 and 8 against LNCaP cell line, with IC50 values ranging from 49.96 ± 1.74 to 76.92 ± 5.85 µM.

Keywords:  Comanthus delicata; crinoid; cytotoxic activity; sulfated anthraquinone; sulfated naphthopyrone

DOI:  https://doi.org/10.1248/cpb.c21-00904
J Alzheimers Dis. 2022 Apr 27.

Oxysterols and Oxysterol Sulfates in Alzheimer's Disease Brain and Cerebrospinal Fluid.

Irundika H K Dias, Hala Shokr, Freya Shephard, Lisa Chakrabarti.

   BACKGROUND: Brain cholesterol levels are tightly regulated but increasing evidence indicates that cholesterol metabolism may drive Alzheimer's disease (AD)-associated pathological changes. Recent advances in understanding of mitochondrial dysfunction in AD brain have presented a vital role played by mitochondria in oxysterol biosynthesis and their involvement in pathophysiology. Oxysterol accumulation in brain is controlled by various enzymatic pathways including sulfation. While research into oxysterol is under the areas of active investigation, there is less evidence for oxysterol sulfates levels in human brain.
OBJECTIVE: This study investigates the hypothesis that AD brain oxysterols detoxification via sulfation is impaired in later stages of disease resulting in oxysterol accumulation.
METHODS: Lipids were extracted from postmortem frozen brain tissue and cerebrospinal (CSF) from late- (Braak stage III-IV) and early- (Braak stage I-II) stage AD patients. Samples were spiked with internal standards prior to lipid extraction. Oxysterols were enriched with a two-step solid phase extraction using a polymeric SPE column and further separation was achieved by LC-MS/MS.
RESULTS: Oxysterols, 26-hydroxycholesterol (26-OHC), 25-hydroxycholesterol (25-OHC), and 7-oxycholesterol levels were higher in brain tissue and mitochondria extracted from late-stage AD brain tissue except for 24S-hydroxycholesterol, which was decreased in late AD. However, oxysterol sulfates are significantly lower in the AD frontal cortex. Oxysterols, 25-OHC, and 7-oxocholesterol was higher is CSF but 26-OHC and oxysterol sulfate levels were not changed.
CONCLUSION: Our results show oxysterol metabolism is altered in AD brain mitochondria, favoring synthesis of 26-OHC, 25-OHC, and 7-oxocholesterol, and this may influence brain mitochondrial function and acceleration of the disease.

Keywords:  Alzheimer’s disease; brain; cholesterol; mitochondria; oxidative stress; oxysterols

DOI:  https://doi.org/10.3233/JAD-220083
ERJ Open Res. 2022 Apr;pii: 00652-2021. [Epub ahead of print]8(2):

Heparan sulfate mimetic fucoidan restores the endothelial glycocalyx and protects against dysfunction induced by serum of COVID-19 patients in the intensive care unit.

Lushun Yuan, Shuzhen Cheng, Wendy M P J Sol, Anouk I M van der Velden, Hans Vink, Ton J Rabelink, Bernard M van den Berg.

Accumulating evidence proves that endothelial dysfunction is involved in coronavirus disease 2019 (COVID-19) progression. We previously demonstrated that the endothelial surface glycocalyx has a critical role in maintenance of vascular integrity. Here, we hypothesised that serum factors of severe COVID-19 patients affect the glycocalyx and result in endothelial dysfunction. We included blood samples of 32 COVID-19 hospitalised patients at the Leiden University Medical Center, of which 26 were hospitalised in an intensive care unit (ICU) and six on a non-ICU hospital floor; 18 of the samples were obtained from convalescent patients 6 weeks after hospital discharge, and 12 from age-matched healthy donors (control) during the first period of the outbreak. First, we determined endothelial (angiopoietin 2 (ANG2)) and glycocalyx degradation (soluble thrombomodulin (sTM) and syndecan-1 (sSDC1)) markers in plasma. In the plasma of COVID-19 patients, circulating ANG2 and sTM were elevated in patients in the ICU. Primary lung microvascular endothelial cell (HPMEC) and human glomerular microvascular endothelial cell (GEnC) cultured in the presence of these sera led to endothelial cell glycocalyx degradation, barrier disruption, inflammation and increased coagulation on the endothelial surface, significantly different compared to healthy control and non-ICU patient sera. These changes could all be restored in the presence of fucoidan. In conclusion, our data highlight the link between endothelial glycocalyx degradation, barrier failure and induction of a procoagulant surface in COVID-19 patients in ICU which could be targeted earlier in disease by the presence of heparan sulfate mimetics.

DOI: https://doi.org/10.1183/23120541.00652-2021
Kidney Int. 2022 May 02. pii: S0085-2538(22)00347-7. [Epub ahead of print]

Soluble α-klotho and heparin modulate the pathologic cardiac actions of fibroblast growth factor 23 in chronic kidney disease.

Christopher Yanucil, Dominik Kentrup, Isaac Campos, Brian Czaya, Kylie Heitman, David Westbrook, Gunars Osis, Alexander Grabner, Adam R Wende, Julian Vallejo, Michael J Wacker, Jose Alberto Navarro-Garcia, Gema Ruiz-Hurtado, Fuming Zhang, Yuefan Song, Robert J Linhardt, Kenneth White, Michael Kapiloff, Christian Faul.

  Fibroblast growth factor (FGF) 23 is a phosphate-regulating hormone that is elevated in patients with chronic kidney disease and associated with cardiovascular mortality. Experimental studies showed that elevated FGF23 levels induce cardiac hypertrophy by targeting cardiac myocytes via FGF receptor isoform 4 (FGFR4). A recent structural analysis revealed that the complex of FGF23 and FGFR1, the physiologic FGF23 receptor in the kidney, includes soluble α-klotho (klotho) and heparin, which both act as co-factors for FGF23/FGFR1 signaling. Here, we investigated whether soluble klotho, a circulating protein with cardio-protective properties, and heparin, a factor that is routinely infused into patients with kidney failure during the hemodialysis procedure, regulate FGF23/FGFR4 signaling and effects in cardiac myocytes. We developed a plate-based binding assay to quantify affinities of specific FGF23/FGFR interactions, and found that soluble klotho and heparin mediate FGF23 binding to distinct FGFR isoforms. Heparin specifically mediated FGF23 binding to FGFR4, and increased FGF23 stimulatory effects on hypertrophic growth and contractility in isolated cardiac myocytes. When repetitively injected into two different mouse models with elevated serum FGF23 levels, heparin aggravated cardiac hypertrophy. We also developed a novel procedure for the synthesis and purification of recombinant soluble klotho, which showed anti-hypertrophic effects in FGF23-treated cardiac myocytes. Thus, soluble klotho and heparin act as independent FGF23 co-receptors with opposite effects on the pathologic actions of FGF23, with soluble klotho reducing and heparin increasing FGF23-induced cardiac hypertrophy. Hence, whether heparin injections during hemodialysis in patients with extremely high serum FGF23 levels contribute to their high rates of cardiovascular events and mortality remains to be studied.

Keywords:  cardiac hypertrophy; chronic kidney disease; fibroblast growth factor 23; heparin; klotho; phosphate metabolism

DOI:  https://doi.org/10.1016/j.kint.2022.03.028
Chem Pharm Bull (Tokyo). 2022 ;70(5): 375-382

Development and Validation of an LC-MS/MS-Based Method for Quantifying Urinary Endogenous 6-Hydroxymelatonin.

Seiya Ohki, Miyu Kunimatsu, Shingo Ogawa, Hiroki Takano, Tomomi Furihata, Hiromi Shibasaki, Akitomo Yokokawa.

  Evaluation of endogenous melatonin (MEL) secretion using its urinary metabolites is useful for the treatment of circadian rhythm sleep disorders. The primary melatonin metabolites excreted in the urine are 6-hydroxymelatonin (6-O-MEL) sulfate (S-O-MEL) and 6-O-MEL glucuronate, which result from sequential MEL metabolism by phases I and II drug metabolizing enzymes. To determine the accurate MEL secretion level, these urinary metabolites should be enzymatically deconjugated and converted into MEL. Furthermore, the use of LC-tandem mass spectrometry (LC-MS/MS) is preferable for the precision of this determination. Therefore, as part of our ongoing efforts to ultimately determine the level of MEL secretion, we herein aimed to develop an LC-MS/MS-based quantification method for 6-O-MEL and optimize deconjugation conditions. We determined the LC-MS/MS conditions of 6-O-MEL measurement and optimized the conditions of enzymatic reactions. The most efficient S-O-MEL deconjugation (102.1%) was achieved with Roche Glucuronidase/Arylsulfatase (from Helix pomatia) at 37 °C, pH-4.0 reaction buffer, and 60 min of reaction time. For human urine samples, the minimum amount of the enzyme required was 5944 units. Under these conditions, the accuracy and precision values of the 6-O-MEL determination (relative errors and standard deviation) were -3.60--0.47% and <6.80%, respectively. Finally, we analyzed the total amount of MEL metabolites excreted in 24-h urine samples; it was 6.70-11.28 µg in three subjects, which is comparable with the values reported till date. Thus, we have established a new method of measuring the total 6-O-MEL in human urine samples using an LC-MS/MS coupled with the prerequisite deconjugation reaction.

Keywords:  6-hydroxymelatonin; LC–tandem mass spectrometry; deconjugation; melatonin; urine

DOI:  https://doi.org/10.1248/cpb.c21-00982
Pediatr Res. 2022 May 05.

Human milk glycosaminoglycans inhibit cytomegalovirus and respiratory syncytial virus infectivity by impairing cell binding.

Rachele Francese, Manuela Donalisio, Massimo Rittà, Federica Capitani, Veronica Mantovani, Francesca Maccari, Paola Tonetto, Guido E Moro, Enrico Bertino, Nicola Volpi, David Lembo.

BACKGROUND: The antiviral role of glycosaminoglycans in human milk (HM-GAGs) has been poorly investigated. They are highly sulfated polysaccharides, which were proposed to act as decoy receptors according to their structure. The aim of this study is to evaluate the antiviral potential and the mechanism of action of total and individual HM-GAGs against three pediatric clinically relevant viruses: respiratory syncytial virus (RSV), cytomegalovirus (HCMV), and rotavirus.
METHODS: HM-GAGs were isolated from HM and a library of individual GAGs, structurally related to HM-GAGs, was prepared. The antiviral activity of HM-GAGs and the impact of thermal treatment were investigated in vitro by specific antiviral assays.
RESULTS: We demonstrated that HM-GAGs are endowed with anti-HCMV and anti-RSV activity and that they act by altering virus attachment to cell. We clarified the contribution of individual HM-GAGs, showing a specific structure-related activity. We did not observe any alteration of HM-GAG antiviral activity after thermal treatment.
CONCLUSIONS: We showed that HM-GAGs contribute to the overall antiviral activity of HM, likely exerting a synergic action with other HM antiviral agents. HM-GAGs can now be added to the list of endogenous factors that may reduce breast-milk-acquired HCMV symptomatic infections and protecting infants from respiratory tract infections by RSV.
IMPACT: HM-GAGs have been poorly investigated for their antiviral action so far. We demonstrated that HM-GAGs are endowed with significant anti-HCMV and anti-RSV activity and that they are able to alter virus binding to the cell. The contribution of individual HM-GAGs is mainly exerted by the FMHep and is not based on a simple charge interaction between the virus and sulfate groups but involves a specific GAG structural configuration. Our results contribute to identifying the multiple factors synergically acting in mediating HM antiviral properties and to clarifying their specific mechanism of action.

DOI: https://doi.org/10.1038/s41390-022-02091-y
Biol Pharm Bull. 2022 ;45(5): 576-582

The Effects of Sacran, a Sulfated Polysaccharide, on Gut Microbiota Using Chronic Kidney Disease Model Rats.

Miwa Goto, Yusei Kobira, Shinichiro Kaneko, Hidetoshi Arima, Akihiro Michihara, Kazuo Azuma, Taishi Higashi, Keiichi Motoyama, Hiroshi Watanabe, Toru Maruyama, Daisuke Kadowaki, Masaki Otagiri, Daisuke Iohara, Fumitoshi Hirayama, Makoto Anraku.

  The aim of this study was to investigate the beneficial effects of sacran, a sulfated polysaccharide, on renal damage and intestinal microflora, in 5/6 nephrectomy rats as a model for chronic kidney disease (CKD). 5/6 Nephrectomy rats were divided into sacran treated and non-treated groups and examined for lethality after 4 weeks. The 5/6 nephrectomy rats were also divided into three groups: sacran treated, non-treated and AST-120 treated groups, and treated orally in a concentration-dependent manner for 4 weeks. Renal function was estimated by biochemical and histopathological analyses. Metagenomic analysis of feces from each group after 4 weeks was also performed and changes in intestinal microflora were compared. The administration of sacran to CKD rats at ≥19 mg/d increased their survival. In addition, the sacran-treated group improved CKD-related parameters in a concentration-dependent manner, and the inhibitory effect of 40 mg/d of sacran was comparable to that of AST-120. The changes in the intestinal microflora of the sacran treated group were positively correlated with an increase in the number of Lactobacillus species, which are known to be rich in beneficial bacteria, and the increment of this beneficial bacteria was negatively correlated with the concentration of indoxyl sulfate, a uremic toxin, in plasma. These results strongly suggest that the oral administration of sacran could contribute to the stabilization of intestinal microflora in CKD rats and to the reduction of oxidative stress as well as the inhibition of progression of CKD.

Keywords:  adsorption; anti-oxidant; chronic kidney disease; gut microbiota; sacran

DOI:  https://doi.org/10.1248/bpb.b21-00897
RSC Adv. 2021 Oct 25. 11(55): 34788-34794

Investigation of the individual human sulfatome in plasma and urine samples reveals an age-dependency.

Mário S P Correia, Bhawana Thapa, Miroslav Vujasinovic, J-Matthias Löhr, Daniel Globisch.

Metabolic microbiome interaction with the human host has been linked to human physiology and disease development. The elucidation of this interspecies metabolite exchange will lead to identification of beneficial metabolites and disease modulators. Their discovery and quantitative analysis requires the development of specific tools and analysis of specific compound classes. Sulfated metabolites are considered a readout for the co-metabolism of the microbiome and their host. This compound class is part of the human phase II clearance process of xenobiotics and is the main focus in drug or doping metabolism and also includes dietary components and microbiome-derived compounds. Here, we report the targeted analysis of sulfated metabolites in plasma and urine samples in the same individuals to identify the core sulfatome and similarities between these two sample types. This analysis of 27 individuals led to the identification of the core sulfatome of 41 metabolites in plasma and urine samples as well as an age effect for 15 metabolites in both sample types.

DOI: https://doi.org/10.1039/d1ra05994g
Acta Crystallogr D Struct Biol. 2022 May 01. 78(Pt 5): 669-682

Structural insights into choline-O-sulfatase reveal the molecular determinants for ligand binding.

Jose Antonio Gavira, Ana Cámara-Artigas, Jose Luis Neira, Jesús M Torres de Pinedo, Pilar Sánchez, Esperanza Ortega, Sergio Martinez-Rodríguez.

  Choline-O-sulfatase (COSe; EC 3.1.6.6) is a member of the alkaline phosphatase (AP) superfamily, and its natural function is to hydrolyze choline-O-sulfate into choline and sulfate. Despite its natural function, the major interest in this enzyme resides in the landmark catalytic/substrate promiscuity of sulfatases, which has led to attention in the biotechnological field due to their potential in protein engineering. In this work, an in-depth structural analysis of wild-type Sinorhizobium (Ensifer) meliloti COSe (SmeCOSe) and its C54S active-site mutant is reported. The binding mode of this AP superfamily member to both products of the reaction (sulfate and choline) and to a substrate-like compound are shown for the first time. The structures further confirm the importance of the C-terminal extension of the enzyme in becoming part of the active site and participating in enzyme activity through dynamic intra-subunit and inter-subunit hydrogen bonds (Asn146A-Asp500B-Asn498B). These residues act as the `gatekeeper' responsible for the open/closed conformations of the enzyme, in addition to assisting in ligand binding through the rearrangement of Leu499 (with a movement of approximately 5 Å). Trp129 and His145 clamp the quaternary ammonium moiety of choline and also connect the catalytic cleft to the C-terminus of an adjacent protomer. The structural information reported here contrasts with the proposed role of conformational dynamics in promoting the enzymatic catalytic proficiency of an enzyme.

Keywords:  alkaline phosphatases; choline; conformational gating; promiscuity; sulfatases

DOI:  https://doi.org/10.1107/S2059798322003709
Ther Drug Monit. 2022 Apr 30.

Short-term high-fat diet alters acetaminophen metabolism in healthy individuals.

Roos Achterbergh, Laureen A Lammers, Heinz-Josef Klümpen, Ron A A Mathôt, Johannes A Romijn.

BACKGROUND: Acetaminophen is metabolized through a non-toxic sulfation and glucuronidation pathway and toxic oxidation pathway (via CYP2E1 and CYP1A2). A short-term high-fat diet induces alterations in the steatotic liver and may alter hepatic drug enzyme activity. In the case of acetaminophen, these alterations may result in an increased risk of hepatotoxicity. Therefore, this study was conducted to assess the effect of a three-day hypercaloric high-fat diet on the plasma levels of acetaminophen metabolites.
METHODS: Nine healthy subjects participated in this randomized, cross over intervention study. The subjects consumed a regular diet or a regular diet supplemented with 500 mL cream (1700 kcal) for 3 days and then fasted overnight. After ingesting 1000 mg acetaminophen, the plasma concentration of acetaminophen (APAP) and its metabolites (acetaminophen glucuronide, acetaminophen sulfate, 3-cysteinyl-acetaminophen and 3-(N-acetyl-L-cystein-S-yl)-acetaminophen, and 3-methoxy-acetaminophen were measured.
RESULTS: The three-day high-fat diet increased the extrapolated area under the concentration-time curve from 0 to infinity (AUC 0-inf) of APAP-Cys by ∼20% (p = 0.02) and that from 0 to 8 h (AUC 0-8) of APAP-Cys-NAC by ∼39% (p = 0.01). The three-day high-fat diet did not alter the pharmacokinetic parameters of the parent compound acetaminophen and other metabolites.
CONCLUSIONS: A short-term hypercaloric high-fat diet increases the plasma levels of the APAP metabolites formed by the oxidation pathway, which may increase the risk of hepatotoxicity.

DOI: https://doi.org/10.1097/FTD.0000000000000993
Biochemistry. 2022 May 06.

The N-Terminus of Human Sulfotransferase 2B1b─a Sterol-Sensing Allosteric Site.

Ian Cook, Thomas S Leyh.

Among human cytosolic sulfotransferases, SULT2B1b is highly specific for oxysterols─oxidized cholesterol derivatives, including nuclear-receptor ligands causally linked to skin and neurodegerative diseases, cancer and atherosclerosis. Sulfonation of signaling oxysterols redirects their receptor-binding functions, and controlling these functions is expected to prove valuable in disease prevention and treatment. SULT2B1b is distinct among the human SULT2 isoforms by virtue of its atypically long N-terminus, which extends 15 residues beyond the next longest N-terminus in the family. Here, in silico studies are used to predict that the N-terminal extension forms an allosteric pocket and to identify potential allosteres. One such allostere, quercetin, is used to confirm the existence of the pocket and to demonstrate that allostere binding inhibits turnover. The structure of the pocket is obtained by positioning quercetin on the enzyme, using spin-label-triangulation NMR, followed by NMR distance-constrained molecular dynamics docking. The model is confirmed using a combination of site-directed mutagenesis and initial-rate studies. Stopped-flow ligand-binding studies demonstrate that inhibition is achieved by stabilizing the closed form of the enzyme active-site cap, which encapsulates the nucleotide, slowing its release. Finally, endogenous oxysterols are shown to bind to the site in a highly selective fashion─one of the two immediate biosynthetic precursors of cholesterol (7-dehydrocholesterol) is an inhibitor, while the other (24-dehydrocholesterol) is not. These findings provide insights into the allosteric dialogue in which SULT2B1b participates in in vivo and establishes a template against which to develop isoform-specific inhibitors to control SULT2B1b biology.

DOI: https://doi.org/10.1021/acs.biochem.1c00740
J Neural Transm (Vienna). 2022 May 02.

Differential genetic associations and expression of PAPST1/SLC35B2 in bipolar disorder and schizophrenia.

Akihito Uezato, Daisuke Jitoku, Dai Shimazu, Naoki Yamamoto, Akeo Kurumaji, Yoshimi Iwayama, Tomoko Toyota, Takeo Yoshikawa, Vahram Haroutunian, Eduard Bentea, Jarek Meller, Courtney R Sullivan, James H Meador-Woodruff, Robert E McCullumsmith, Toru Nishikawa.

  Lithium's inhibitory effect on enzymes involved in sulfation process, such as inhibition of 3'(2')-phosphoadenosine 5'-phosphate (PAP) phosphatase, is a possible mechanism of its therapeutic effect for bipolar disorder (BD). 3'-Phosphoadenosine 5'-phosphosulfate (PAPS) is translocated from cytosol to Golgi lumen by PAPS transporter 1 (PAPST1/SLC35B2), where it acts as a sulfa donor. Since SLC35B2 was previously recognized as a molecule that facilitates the release of D-serine, a co-agonist of N-methyl-D-aspartate type glutamate receptor, altered function of SLC35B2 might be associated with the pathophysiology of BD and schizophrenia (SCZ). We performed genetic association analyses of the SLC35B2 gene using Japanese cohorts with 366 BD cases and 370 controls and 2012 SCZ cases and 2170 controls. We then investigated expression of SLC35B2 mRNA in postmortem brains by QPCR using a Caucasian cohort with 33 BD and 34 SCZ cases and 34 controls and by in situ hybridization using a Caucasian cohort with 37 SCZ and 29 controls. We found significant associations between three SNPs (rs575034, rs1875324, and rs3832441) and BD, and significantly reduced SLC35B2 mRNA expression in postmortem dorsolateral prefrontal cortex (DLPFC) of BD. Moreover, we observed normalized SLC35B2 mRNA expression in BD subgroups who were medicated with lithium. While there was a significant association of SLC35B2 with SCZ (SNP rs2233437), its expression was not changed in SCZ. These findings indicate that SLC35B2 might be differentially involved in the pathophysiology of BD and SCZ by influencing the sulfation process and/or glutamate system in the central nervous system.

Keywords:  Bipolar disorder; Brain mRNA expression; Genetic association; Lithium; PAPST1/SLC35B2; Schizophrenia

DOI:  https://doi.org/10.1007/s00702-022-02503-7