bims-supasi 2023-05-07 papers

bims-supasi

Biomed News

on Sulfation pathways and signalling

Issue of 2023‒05‒07
thirteen papers selected by
Jonathan Wolf Mueller
University of Birmingham

Interactions of heparin with key glycoproteins of human respiratory syncytial virus.
High levels of exfoliated fragments following glycocalyx destruction in hemorrhagic fever with the renal syndrome are associated with mortality risk.
A 3-O-sulfated heparan sulfate dodecasaccharide (12-mer) suppresses thromboinflammation and attenuates early organ injury following trauma and hemorrhagic shock.
Dehydroepiandrosterone sulfate levels at 7 years old and cardio-metabolic factors at 10 and 13 years old - the generation XXI birth cohort.
Pteropine orthoreoviruses use cell surface heparan sulfate as an attachment receptor.
Characterization of Bile Acid Sulfate Conjugates as Substrates of Human Organic Anion Transporting Polypeptides.
Amino acid variation at VP1-145 of enterovirus A71 determines the viral infectivity and receptor usage in a primary human intestinal model.
Effects of resveratrol on inflammation and oxidative stress induced by the uremic toxin indoxyl sulfate in Murine macrophage-like RAW 264.7.
Electrostatic interaction between sulfated polysaccharides and oligopeptides from viral envelope glycoproteins using surface plasmon resonance.
Metabolomic Profiling to Identify Early Urinary Biomarkers and Metabolic Pathway Alterations in Autosomal Dominant Polycystic Kidney Disease.
Estimated ovulatory years prior to menopause and postmenopausal endogenous hormone levels.
Validation of an isotope dilution mass spectrometry (IDMS) measurement procedure for the reliable quantification of steroid hormones in waters.
Evaluation of the selectivity of several OATP1B biomarkers using relative activity factor method.

Front Mol Biosci. 2023 ;10 1151174

Interactions of heparin with key glycoproteins of human respiratory syncytial virus.

Deling Shi, Peng He, Yuefan Song, Robert J Linhardt, Jonathan S Dordick, Lianli Chi, Fuming Zhang.

  Introduction: The unexpected surge of respiratory syncytial virus (RSV) cases following pandemic phase of COVID-19 has drawn much public attention. Drawing on the latest antiviral research, revisiting this heightened annual outbreak of respiratory disease could lead to new treatments. The ability of sulfated polysaccharides to compete for a variety of viruses binding to cell surface heparan sulfate, suggests several drugs that might have therapeutic potential for targeting RSV-glycosaminoglycan interactions. Methods: In the current study, the binding affinity and kinetics of two RSV glycoproteins (RSV-G protein and RSV-F protein) to heparin were investigated by surface plasmon resonance. Furthermore, solution competition studies using heparin oligosaccharides of different lengths indicated that the binding of RSV-G protein to heparin is size-dependent, whereas RSV-F protein did not show any chain length preference. Results and discussion: The two RSV glycoproteins have slightly different preferences for heparin sulfation patterns, but the N-sulfo group in heparin was most critical for the binding of heparin to both RSV-G protein and RSV-F protein. Finally, pentosan polysulfate and mucopolysaccharide polysulfate were evaluated for their inhibition of the RSV-G protein and RSV-F protein-heparin interaction, and both highly negative compounds showed strong inhibition.

Keywords:  glycoproteins; heparin; human respiratory syncytial virus; mucopolysaccharide polysulfate; pentosan polysulfate

DOI:  https://doi.org/10.3389/fmolb.2023.1151174
Front Med (Lausanne). 2023 ;10 1096353

High levels of exfoliated fragments following glycocalyx destruction in hemorrhagic fever with the renal syndrome are associated with mortality risk.

Hong Du, Haifeng Hu, Jing Li, Xiaoyan Wang, Hong Jiang, Jianqi Lian, Ying Zhang, Pingzhong Wang.

  Background: The glycocalyx is a gel-like structure that covers the luminal side of vascular endothelial cells. It plays an important role in maintaining the integrity of the vascular endothelial barrier structure. However, the presence or absence of glycocalyx destruction in hemorrhagic fever with renal syndrome (HFRS) and its specific mechanism and role is still unclear.Methods: In this study, we detected the levels of exfoliated glycocalyx fragments, namely, heparan sulfate (HS), hyaluronic acid (HA), and chondroitin sulfate (CS), in HFRS patients and investigated their clinical application value on the evaluation of disease severity and prognosis prediction.
Results: The expression of exfoliated glycocalyx fragments in plasma was significantly increased during the acute stage of HFRS. The levels of HS, HA, and CS in HFRS patients during the acute stage were significantly higher than in healthy controls and convalescent stages of the same type. HS and CS during the acute stage gradually increased with the aggravation of HFRS, and both fragments showed a significant association with disease severity. In addition, exfoliated glycocalyx fragments (especially HS and CS) showed a significant correlation with conventional laboratory parameters and hospitalization days. High levels of HS and CS during the acute phase were significantly associated with patient mortality and demonstrated an obvious predictive value for the mortality risk of HFRS.
Conclusion: Glycocalyx destruction and shedding may be closely associated with endothelial hyperpermeability and microvascular leakage in HFRS. The dynamic detection of the exfoliated glycocalyx fragments may be beneficial for the evaluation of disease severity and prognosis prediction in HFRS.

Keywords:  chondroitin sulfate; glycocalyx; hemorrhagic fever with renal syndrome; heparan sulfate; hyaluronic acid

DOI:  https://doi.org/10.3389/fmed.2023.1096353
Front Immunol. 2023 ;14 1158457

A 3-O-sulfated heparan sulfate dodecasaccharide (12-mer) suppresses thromboinflammation and attenuates early organ injury following trauma and hemorrhagic shock.

Maria Del Pilar Huby Vidaurre, Baron K Osborn, Kaylie D Lowak, Michelle M McDonald, Yao-Wei W Wang, Veda Pa, Jillian R Richter, Yongmei Xu, Katelyn Arnold, Jian Liu, Jessica C Cardenas.

  Introduction: Dysregulated inflammation and coagulation are underlying mechanisms driving organ injury after trauma and hemorrhagic shock. Heparan sulfates, cell surface glycosaminoglycans abundantly expressed on the endothelial surface, regulate a variety of cellular processes. Endothelial heparan sulfate containing a rare 3-O-sulfate modification on a glucosamine residue is anticoagulant and anti-inflammatory through high-affinity antithrombin binding and sequestering of circulating damage-associated molecular pattern molecules. Our goal was to evaluate therapeutic potential of a synthetic 3-O-sulfated heparan sulfate dodecasaccharide (12-mer, or dekaparin) to attenuate thromboinflammation and prevent organ injury.Methods: Male Sprague-Dawley rats were pre-treated subcutaneously with vehicle (saline) or dekaparin (2 mg/kg) and subjected to a trauma/hemorrhagic shock model through laparotomy, gut distention, and fixed-pressure hemorrhage. Vehicle and dekaparin-treated rats were resuscitated with Lactated Ringer's solution (LR) and compared to vehicle-treated fresh-frozen-plasma-(FFP)-resuscitated rats. Serial blood samples were collected at baseline, after induction of shock, and 3 hours after fluid resuscitation to measure hemodynamic and metabolic shock indicators, inflammatory mediators, and thrombin-antithrombin complex formation. Lungs and kidneys were processed for organ injury scoring and immunohistochemical analysis to quantify presence of neutrophils.
Results: Induction of trauma and hemorrhagic shock resulted in significant increases in thrombin-antithrombin complex, inflammatory markers, and lung and kidney injury scores. Compared to vehicle, dekaparin treatment did not affect induction, severity, or recovery of shock as indicated by hemodynamics, metabolic indicators of shock (lactate and base excess), or metrics of bleeding, including overall blood loss, resuscitation volume, or hematocrit. While LR-vehicle-resuscitated rodents exhibited increased lung and kidney injury, administration of dekaparin significantly reduced organ injury scores and was similar to organ protection conferred by FFP resuscitation. This was associated with a significant reduction in neutrophil infiltration in lungs and kidneys and reduced lung fibrin deposition among dekaparin-treated rats compared to vehicle. No differences in organ injury, neutrophil infiltrates, or fibrin staining between dekaparin and FFP groups were observed. Finally, dekaparin treatment attenuated induction of thrombin-antithrombin complex and inflammatory mediators in plasma following trauma and hemorrhagic shock.
Conclusion: Anti-thromboinflammatory properties of a synthetic 3-O-sulfated heparan sulfate 12-mer, dekaparin, could provide therapeutic benefit for mitigating organ injury following major trauma and hemorrhagic shock.

Keywords:  hemorrhagic shock; heparan sulfate; organ injury; thromboinflammation; traumatic injury

DOI:  https://doi.org/10.3389/fimmu.2023.1158457
J Pediatr Endocrinol Metab. 2023 May 05.

Dehydroepiandrosterone sulfate levels at 7 years old and cardio-metabolic factors at 10 and 13 years old - the generation XXI birth cohort.

Rita Santos-Silva, Manuel Fontoura, Milton Severo, Ana Cristina Santos.

  OBJECTIVES: Premature adrenarche is often linked to a cluster of endocrine-metabolic risk factors. Our objective was to explore the association of dehydroepiandrosterone sulfate (DHEAS) levels at age 7 with cardio-metabolic traits at ages 10 and 13, independently of adiposity and pubertal stage.METHODS: Longitudinal study of 603 individuals (301 girls/302 boys) from the Generation XXI birth cohort. DHEAS at age 7 was measured by immunoassay. Anthropometrics, pubertal staging, blood pressure, and metabolic outcomes were evaluated at ages 7, 10, and 13. Pearson correlations between DHEAS and cardio-metabolic traits (insulin, HOMA-IR, triglycerides, LDL-cholesterol, high-sensitivity C-reactive protein, and systolic and diastolic blood pressure) were computed. Path analysis was used to estimate the effect of DHEAS at age 7 on cardiometabolic traits at ages 10 and 13, adjusted for body mass index (BMI) z-score and Tanner stage.
RESULTS: DHEAS at age 7 correlated positively with insulin and HOMA-IR at ages 7 and 10 in both sexes, and at age 13 in girls, but not in boys. In girls, DHEAS levels at age 7 directly influenced HOMA-IR at age 13, controlling for BMI and Tanner stage. In boys, DHEAS at age 7 did not influence HOMA-IR at ages 10 and 13. DHEAS at age 7 did not influence the other cardio-metabolic outcomes analyzed.
CONCLUSIONS: DHEAS levels in mid-childhood have a positive longitudinal association with on insulin-resistance that persists, in girls, but not in boys, at least until age 13. No association was found regarding dyslipidemia, hypertension, or low-grade inflammation.

Keywords:  adrenarche; cardiovascular risk; dehydroepiandrosterone sulfate; generation XXI; metabolic syndrome

DOI:  https://doi.org/10.1515/jpem-2022-0593
Emerg Microbes Infect. 2023 May 05. 2208683

Pteropine orthoreoviruses use cell surface heparan sulfate as an attachment receptor.

Chee Wah Tan, Akshamal M Gamage, Wee Chee Yap, Leon Jia Wei Tang, Yuan Sun, Xing-Lou Yang, Alyssa Pyke, Kaw Bing Chua, Lin-Fa Wang.

  Pteropine orthoreoviruses (PRV) are an emerging group of fusogenic, bat-borne viruses from the orthoreovirus genus. Since the isolation of PRV from a patient with acute respiratory tract infections in 2006, the zoonotic potential of PRV has been further highlighted following subsequent isolation of PRV species from patients in Malaysia, Hong Kong and Indonesia. However, the entry mechanism of PRV is currently unknown. In this study, we investigated the role of previously identified mammalian orthoreovirus (MRV) receptors, sialic acid and junctional adhesion molecule-1, for PRV infection. However, none of these receptors played a significant role in PRV infection, suggesting PRV uses a distinct entry receptor from MRV. Given its broad-tissue tropism, we hypothesized that PRV may use a receptor that is widely expressed in all cell types, heparan sulfate (HS). Enzymatic removal of cell surface HS by heparinase treatment, and genetic ablation of HS biosynthesis genes, SLC35B2, EXT-1, NDST-1 and B3GAT3 significantly reduced infection with multiple genetically distinct PRV species. Replication kinetic of PRV3M in HS-knockout cells revealed that HS plays a crucial role in the early phase of PRV infection. Mechanistic studies demonstrated that HS is an essential host-factor for PRV attachment and internalization into cells. To our knowledge, this is the first report on the use of HS as an attachment receptor by pteropine orthoreoviruses.

Keywords:  Heparan sulfate; Pteropine orthoreovirus; attachment receptor

DOI:  https://doi.org/10.1080/22221751.2023.2208683
Mol Pharm. 2023 May 03.

Characterization of Bile Acid Sulfate Conjugates as Substrates of Human Organic Anion Transporting Polypeptides.

Christine C Orozco, Mikko Neuvonen, Yi-An Bi, Matthew A Cerny, Sumathy Mathialagan, Laurie Tylaska, Brian Rago, Chester Costales, Amanda King-Ahmad, Mikko Niemi, A David Rodrigues.

  Drug interactions involving the inhibition of hepatic organic anion transporting polypeptides (OATPs) 1B1 and OATP1B3 are considered important. Therefore, we sought to study various sulfated bile acids (BA-S) as potential clinical OATP1B1/3 biomarkers. It was determined that BA-S [e.g., glycochenodeoxycholic acid 3-O-sulfate (GCDCA-S) and glycodeoxycholic acid 3-O-sulfate (GDCA-S)] are substrates of OATP1B1, OATP1B3, and sodium-dependent taurocholic acid cotransporting polypeptide (NTCP) transfected into human embryonic kidney 293 cells, with minimal uptake evident for other solute carriers (SLCs) like OATP2B1, organic anion transporter 2, and organic cation transporter 1. It was also shown that BA-S uptake by plated human hepatocytes (PHH) was inhibited (≥96%) by a pan-SLC inhibitor (rifamycin SV), and there was greater inhibition (≥77% versus ≤12%) with rifampicin (OATP1B1/3-selective inhibitor) than a hepatitis B virus myristoylated-preS1 peptide (NTCP-selective inhibitor). Estrone 3-sulfate was also used as an OATP1B1-selective inhibitor. In this instance, greater inhibition was observed with GDCA-S (76%) than GCDCA-S (52%). The study was expanded to encompass the measurement of GCDCA-S and GDCA-S in plasma of SLCO1B1 genotyped subjects. The geometric mean GDCA-S concentration was 2.6-fold (90% confidence interval 1.6, 4.3; P = 2.1 × 10-4) and 1.3-fold (1.1, 1.7; P = 0.001) higher in individuals homozygous and heterozygous for the SLCO1B1 c.521T > C loss-of-function allele, respectively. For GCDCA-S, no significant difference was noted [1.2-fold (0.8, 1.7; P = 0.384) and 0.9-fold (0.8, 1.1; P = 0.190), respectively]. This supported the in vitro data indicating that GDCA-S is a more OATP1B1-selective substrate (versus GCDCA-S). It is concluded that GCDCA-S and GDCA-S are viable plasma-based OATP1B1/3 biomarkers, but they are both less OATP1B1-selective when compared to their corresponding 3-O-glucuronides (GCDCA-3G and GDCA-3G). Additional studies are needed to determine their utility versus more established biomarkers, such as coproporphyrin I, for assessing inhibitors with different OATP1B1 (versus OATP1B3) inhibition signatures.

Keywords:  biomarker; drug interaction; organic anion transporting polypeptide; sulfated bile acids

DOI:  https://doi.org/10.1021/acs.molpharmaceut.3c00040
Front Microbiol. 2023 ;14 1045587

Amino acid variation at VP1-145 of enterovirus A71 determines the viral infectivity and receptor usage in a primary human intestinal model.

Ikrame Aknouch, Inés García-Rodríguez, Francesca Paola Giugliano, Carlemi Calitz, Gerrit Koen, Hetty van Eijk, Nina Johannessson, Sjoerd Rebers, Lieke Brouwer, Vanesa Muncan, Koert J Stittelaar, Dasja Pajkrt, Katja C Wolthers, Adithya Sridhar.

  Enterovirus A71 (EV-A71) can elicit a wide variety of human diseases such as hand, foot, and mouth disease and severe or fatal neurological complications. It is not clearly understood what determines the virulence and fitness of EV-A71. It has been observed that amino acid changes in the receptor binding protein, VP1, resulting in viral binding to heparan sulfate proteoglycans (HSPGs) may be important for the ability of EV-A71 to infect neuronal tissue. In this study, we identified that the presence of glutamine, as opposed to glutamic acid, at VP1-145 is key for viral infection in a 2D human fetal intestinal model, consistent with previous findings in an airway organoid model. Moreover, pre-treatment of EV-A71 particles with low molecular weight heparin to block HSPG-binding significantly reduced the infectivity of two clinical EV-A71 isolates and viral mutants carrying glutamine at VP1-145. Our data indicates that mutations in VP1 leading to HSPG-binding enhances viral replication in the human gut. These mutations resulting in increased production of viral particles at the primary replication site could lead to a higher risk of subsequent neuroinfection.Importance: With the near eradication of polio worldwide, polio-like illness (as is increasingly caused by EV-A71 infections) is of emerging concern. EV-A71 is indeed the most neurotropic enterovirus that poses a major threat globally to public health and specifically in infants and young children. Our findings will contribute to the understanding of the virulence and the pathogenicity of this virus. Further, our data also supports the identification of potential therapeutic targets against severe EV-A71 infection especially among infants and young children. Furthermore, our work highlights the key role of HSPG-binding mutations in the disease outcome of EV-A71. Additionally, EV-A71 is not able to infect the gut (the primary replication site in humans) in traditionally used animal models. Thus, our research highlights the need for human-based models to study human viral infections.Graphical Abstract.

Keywords:  EV-A71; Transwell; VP1-145; heparin sulfate proteoglycan; human inestinal organoids; polarized epithelium

DOI:  https://doi.org/10.3389/fmicb.2023.1045587
Biochimie. 2023 May 02. pii: S0300-9084(23)00097-4. [Epub ahead of print]

Effects of resveratrol on inflammation and oxidative stress induced by the uremic toxin indoxyl sulfate in Murine macrophage-like RAW 264.7.

Livia Alvarenga, Juliana F Saldanha, Milena B Stockler-Pinto, Denis Fouque, Christophe O Soulage, Denise Mafra.

  Indoxyl sulfate (IS) is a uremic toxin produced by the gut microbiota that commonly accumulates in patients with chronic kidney disease (CKD) and can be harmful. Resveratrol is a polyphenol with properties that attenuate oxidative stress and inflammation. This study aims to evaluate the effect of resveratrol against the damage caused by IS in RAW 264.7 murine macrophages. Cells were treated with 0, 250, 500 and 1000 μmol/L of IS, in the presence of 50 μmol/L of resveratrol. The mRNA and protein expressions of erythroid-related nuclear factor 2 (Nrf2) and nuclear factor kappa-B (NF-κB) were measured using rt-PCR and western blot analysis, respectively. Malondialdehyde (MDA) and reactive oxygen species (ROS) levels were also analyzed. As a result, it was demonstrated that resveratrol induces the activation of the Nrf2 pathway that enhances cytoprotective response. IS upregulated the NF-κB expression and downregulated the Nrf2 expression. In contrast, resveratrol treatment significantly reduced the MDA and ROS production and inhibited the IS-induced expression of NF-κB in macrophage-like RAW 264.7. In conclusion, resveratrol can mitigate inflammation and oxidative stress caused by uremic toxins produced by the gut microbiota, such as IS.

Keywords:  Indoxyl sulfate; Inflammation; Oxidative stress; Resveratrol

DOI:  https://doi.org/10.1016/j.biochi.2023.05.001
Carbohydr Res. 2023 Apr 19. pii: S0008-6215(23)00077-0. [Epub ahead of print]528 108815

Electrostatic interaction between sulfated polysaccharides and oligopeptides from viral envelope glycoproteins using surface plasmon resonance.

Tungalag Battulga, Ayiguli Dagaerbieke, Chaolumen Bai, Daisuke Asai, Takuro Koshikawa, Hiromu Takemura, Kensuke Miyazaki, Takashi Yoshida.

  Biotinylated oligopeptides from the envelope glycoproteins of dengue fever virus, influenza A and B viruses, and human immunodeficiency virus (HIV) were synthesized and their interaction with curdlan and dextran sulfates was investigated using surface plasmon resonance to evaluate the antiviral mechanisms of sulfated polysaccharides. More than two clusters consisting of basic amino acids in the oligopeptides from dengue fever virus, strongly interacted with the sulfated polysaccharides elucidated by the association- and dissociation-rate constants. Interactions decreased with the decreasing molecular weights of the sulfated polysaccharides. Although oligopeptides from influenza A virus potently interacted with the sulfated polysaccharides, no interaction was detected on a B/Hong Kong virus oligopeptide bearing few basic amino acids. For the C terminus and V3 region short and long oligopeptides from HIV gp120, the interaction was enhanced by the number of clustered basic amino acids and was inhibited by acidic and bulky amino acids.

Keywords:  Basic amino acid; Electrostatic interaction; Oligopeptides; SPR; Sulfated polysaccharides

DOI:  https://doi.org/10.1016/j.carres.2023.108815
Am J Physiol Renal Physiol. 2023 May 04.

Metabolomic Profiling to Identify Early Urinary Biomarkers and Metabolic Pathway Alterations in Autosomal Dominant Polycystic Kidney Disease.

Eden A Houske, Matthew G Glimm, Annika R Bergstrom, Sally K Slipher, Hope D Welhaven, Mark C Greenwood, Greta M Linse, Ronald K June, Alan S L Yu, Darren P Wallace, Alyssa K Hahn.

  Autosomal dominant polycystic kidney disease (ADPKD) is characterized by the formation of numerous fluid-filled cysts that lead to progressive loss of functional nephrons. Currently, there is an unmet need for diagnostic and prognostic indicators of early-stage ADPKD. Metabolites were extracted from the urine of early-stage ADPKD patients (n=48) and age- and sex-matched normal controls (n=47) and analyzed by liquid chromatography-mass spectrometry. Orthogonal partial least squares-discriminant analysis was employed to generate a global metabolomic profile of early ADPKD for the identification of metabolic pathway alterations and discriminatory metabolites as candidate diagnostic and prognostic biomarkers. The global metabolomic profile exhibited alterations in metabolism of steroids, fatty acids, pyruvate, amino acids, and the urea cycle. A panel of 46 metabolite features were identified as candidate diagnostic biomarkers. Notable putative identities of candidate diagnostic biomarkers for early detection include creatinine, cAMP, dCMP, various androgens, betaine aldehyde, phosphoric acid, choline, 18-hydroxycorticosterone, and cortisol. Metabolic pathways associated with variable rates of disease progression included metabolism of steroids, vitamin D3, fatty acids, amino acids, sialic acid, the pentose phosphate pathway, the tricarboxylic acid cycle, and chondroitin sulfate and heparin sulfate degradation. A panel of 41 metabolite features were identified as candidate prognostic biomarkers. Notable putative identities of candidate prognostic biomarkers include ethanolamine, C20:4 anandamide phosphate, progesterone, various androgens, betaine aldehyde, inflammatory lipids, and choline. Our exploratory data support metabolic reprogramming in early ADPKD and demonstrate the ability of mass spectrometry-based global metabolomic profiling to detect metabolic pathway alterations as new therapeutic targets and biomarkers of ADPKD.

Keywords:  biomarkers; liquid chromatography-mass spectrometry; metabolomics; polycystic kidney disease; urine

DOI:  https://doi.org/10.1152/ajprenal.00301.2022
Cancer Epidemiol Biomarkers Prev. 2023 May 01. pii: EPI-23-0102. [Epub ahead of print]

Estimated ovulatory years prior to menopause and postmenopausal endogenous hormone levels.

Daniel W Cramer, Allison F Vitonis, Tianyi Huang, Amy L Shafrir, A Heather Eliassen, Robert L Barbieri, Susan E Hankinson.

Background Lifetime ovulatory years (LOY) is estimated by the difference between ages at menopause and menarche subtracting time for events interrupting ovulation. We tested whether LOY influences sex-hormone levels in postmenopausal women with at least one intact ovary not using hormones. Methods Estradiol, estrone, estrone sulfate, total testosterone, dehydroepiandrostendione sulfate (DHEAS), prolactin, and sex-hormone-binding globulin (SHBG) were measured in 1,976 postmenopausal women from the Nurses' Health Study. Associations of age, body mass index (BMI), smoking, alcohol use, and other factors on hormones were assessed by t-tests and ANOVA. Linear regression was used to assess multivariable-adjusted associations between LOY and hormones and trends in hormone levels per 5-year increases in LOY were estimated. Results Women averaged 61.4 years old, 11.0 years since menopause, with BMI of 25.8 kg/m2. 13.6% had irregular cycles, 17.5% hysterectomy, 6.4% unilateral oophorectomy, and 13.8% were current smokers. Variables associated with one or more hormone levels were included as covariates. Each 5-year increase in LOY was significantly associated with a 5.2% increase in testosterone in women with BMI<25 kg/m2 and a 7.4% increase in testosterone and 7.3% increase in estradiol in women with above-average BMI. Conclusions This is the first study to show that greater LOY is associated with higher testosterone in postmenopausal women and higher estradiol in those with elevated BMI, suggesting accumulation of functioning stromal and thecal cells from repeated ovulations and peripheral conversion of testosterone. Impact A possible explanation for why greater LOY increases risk for breast, endometrial, and ovarian cancer is offered.

DOI: https://doi.org/10.1158/1055-9965.EPI-23-0102
Anal Bioanal Chem. 2023 Apr 29.

Validation of an isotope dilution mass spectrometry (IDMS) measurement procedure for the reliable quantification of steroid hormones in waters.

Elodie Mirmont, Amandine Bœuf, Mélissa Charmel, Béatrice Lalère, Sophie Lardy-Fontan.

  Reliable data are compulsory to efficiently monitor pollutants in aquatic environments, particularly steroid hormones that can exert harmful effects at challenging analytical levels below the ng L-1. An isotope dilution two-step solid-phase extraction followed by an ultra-performance liquid chromatography separation coupled to tandem mass spectrometry (UPLC-MS/MS) detection method was validated for the quantification of 21 steroid hormones (androgens, estrogens, glucocorticoids, and progestogens) in whole waters. To achieve a realistic and robust assessment of the performances of this method, the validation procedure was conducted using several water samples representative of its intended application. These samples were characterized in terms of concentration of ionic constituents, suspended particulate matter (SPM), and dissolved organic carbon contents (DOC). For estrogens that are part of the European Water Framework Directive Watchlist (17beta-estradiol and estrone), the performances met the European requirements (decision 2015/495/EU) in terms of limit of quantification (LQ) and measurement uncertainty. For 17alpha-ethinylestradiol, the challenging LQ of 0.035 ng L-1 was reached. More generally, for 15 compounds out of 21, the accuracy, evaluated in intermediate precision conditions at concentrations ranging between 0.1 and 10 ng L-1, was found to be within a 35% tolerance. The evaluation of the measurement uncertainty was realized following the Guide to the expression of Uncertainty in Measurement. Finally, a water monitoring survey demonstrated the suitability of the method and pointed out the contamination of Belgium rivers by five estrogens (17alpha-ethinylestradiol, estriol, 17alpha-estradiol, 17beta-estradiol, and estrone) and three glucocorticoids (betamethasone, cortisol, and cortisone) which have been up to now poorly documented in European rivers.

Keywords:  Measurement accuracy; Measurement uncertainties; Steroids; Ultra-trace quantification; Whole water

DOI:  https://doi.org/10.1007/s00216-023-04698-4
Drug Metab Dispos. 2023 May 03. pii: DMD-AR-2022-000972. [Epub ahead of print]

Evaluation of the selectivity of several OATP1B biomarkers using relative activity factor method.

Grace Hoyee Chan, Robert Houle, Jinghui Zhang, Ravi Katwaru, Yang Li, Xiaoyan Chu.

  In recent years, some endogenous substrates of organic anion transporting polypeptide 1B (OATP1B) have been identified and characterized as potential biomarkers to assess OATP1B-mediated clinical drug-drug interactions (DDIs). However, quantitative determination of their selectivity to OATP1B are still limited. In this study, we developed a relative activity factor (RAF) method to determine the relative contribution of hepatic uptake transporters OATP1B1, OATP1B3, OATP2B1, and sodium-taurocholate co-transporting polypeptide (NTCP) on hepatic uptake of several OATP1B biomarkers, including coproporphyrins I (CPI), CPIII, sulfate conjugates of bile acids: glycochenodeoxycholic acid sulfate (GCDCA-S), glycodeoxycholic acid sulfate (GDCA-S), and taurochenodeoxycholic acid sulfate (TCDCA-S). RAF values for OATP1B1, OATP1B3, OATP2B1, and NTCP were determined in cryopreserved human hepatocytes and transporter transfected cells using pitavastatin, cholecystokinin, resveratrol-3-O-β-D-glucuronide, and taurocholic acid (TCA) as reference compounds, respectively. OATP1B1-specific pitavastatin uptake in hepatocytes was measured in the absence and presence of 1 µM estropipate, while NTCP-specific TCA uptake was measured in the presence of 10 µM rifampin. Our studies suggested that CPI was a more selective biomarker for OATP1B1 than CPIII, while GCDCA-S and TCDCA-S were more selective to OATP1B3. OATP1B1 and OATP1B3 equally contributed to hepatic uptake of GDCA-S. The mechanistic static model, incorporating the fraction transported (ft) of CPI/III estimated by RAF and in vivo elimination data, predicted several perpetrator interactions with CPI/III. Overall, RAF method combined with pharmacogenomic and DDI studies is a useful tool to determine the selectivity of transporter biomarkers and facilitate the selection of appropriate biomarkers for DDI evaluation. Significance Statement We developed a new RAF method to quantitatively determine the contribution of hepatic uptake transporters OATP1B1, OATP1B3, OATP2B1, and NTCP on several OATP1B biomarkers (CPI, CPIII, GCDCA-S, GDCA-S, and TCDCA-S) and evaluated their predictivity on perpetrator-biomarker interactions. Our studies suggest that RAF method is a useful tool to determine the selectivity of transporter biomarkers. This method combined with pharmacogenomic and DDI studies will facilitate mechanistic interpretation and modeling of biomarker data and the selection of appropriate biomarkers for DDI evaluation.

Keywords:  active transport; drug-drug interactions; pharmacokinetics

DOI:  https://doi.org/10.1124/dmd.122.000972