Biology subjects

Botre, F.

Publications and source records attributed to Botre, F..

5 recordsLinked to original sources

bioRxiv2023

EPAS1 Attenuates Atherosclerosis Initiation at Disturbed Flow Sites through Endothelial Fatty Acid Uptake

BackgroundAtherosclerotic plaques form unevenly due to disturbed blood flow, causing localized endothelial cell (EC) dysfunction. Obesity exacerbates this process, but the underlying molecular mechanisms are unclear. The transcription factor EPAS1 (HIF2A) has regulatory roles in endothelium, but its involvement in atherosclerosis remains unexplored. This study investigates the potential interplay between EPAS1, obesity, and atherosclerosis. MethodsResponses to shear stress were analysed using cultured porcine aortic EC exposed to flow in vitro coupled with metabolic and molecular analyses, and by en face immunostaining of murine aortic EC exposed to disturbed flow in vivo. Obesity and dyslipidemia were induced in mice via exposure to high-fat diet or through Leptin gene deletion. The role of Epas1 in atherosclerosis was evaluated by inducible endothelial Epas1 deletion, followed by hypercholesterolemia induction (AAV-PCSK9; high-fat diet). ResultsEn face staining revealed EPAS1 enrichment at sites of disturbed blood flow that are prone to atherosclerosis initiation. Obese mice exhibited substantial reduction in endothelial EPAS1 expression, correlating with hyperlipidaemia. Sulforaphane, a compound with known atheroprotective effects, restored EPAS1 expression and concurrently reduced plasma triglyceride levels in obese mice. Consistently, triglyceride derivatives (free fatty acids) suppressed EPAS1 in cultured EC by upregulating the negative regulator PHD3. Clinical observations revealed that reduced plasma EPAS1 correlated with increased endothelial PHD3 in obese individuals. Functionally, endothelial EPAS1 deletion increased lesion formation in hypercholesterolemic mice, indicating an atheroprotective function. Mechanistic insights revealed that EPAS1 protects arteries by maintaining endothelial proliferation by positively regulating CD36 and LIPG expression to increase fatty acid beta-oxidation. ConclusionsEndothelial EPAS1 attenuates atherosclerosis at sites of disturbed flow by maintaining EC proliferative via fatty acid uptake and metabolism. This endothelial repair pathway is inhibited in obesity, suggesting a novel triglyceride-PHD3 modulation pathway suppressing EPAS1 expression. These findings have implications for therapeutic strategies addressing vascular dysfunction in obesity.

physiology↗

bioRxiv2023

Quinoa as Functional Food? Urinary elimination of ecdysterone after consumption of quinoa alone and in combination with spinach

The phytosteroid ecdysterone is included on the monitoring list of the World Anti-Doping Agency. Therefore, the consumption of food rich in ecdysterone is the focus of a lively debate. Thus, urinary excretion of ecdysterone and its metabolites in humans was investigated following quinoa consumption alone and in combination with spinach. After intake of both preparations, ecdysterone and two metabolites were excreted in urine. Maximum concentrations of ecdysterone ranged from 0.44-5.50{micro}g/mL after quinoa and 0.34- 4.09{micro}g/mL after quinoa with spinach. The total urinary excreted amount as parent drug plus metabolites was 2.60(1.09)% following quinoa and 1.71(0.86)% after combination. Significant differences were found in total urinary excreted amounts of ecdysterone, 14-deoxy-ecdysterone, and 14-deoxy-poststerone. In conclusion, only small proportions of ecdysterone from quinoa and the combination with spinach were excreted in urine. The results indicate that both, quinoa and spinach, are poor sources of ecdysterone.

pharmacology and toxicology↗

bioRxiv2022

Variability of the urinary and blood steroid profiles in healthy women

The steroidal module of the Athlete Biological Passport (ABP) targets the use of exogenous androgenous anabolic steroids (EAAS) in elite sport by monitoring urinary steroid profiles. Urine and blood samples were collected weekly during two consecutive OCP cycles (8 weeks) in 15 physically active women to investigate the low urinary steroid concentrations and putative confounding effect of OCP. In urine, testosterone (T) and/or epitestosterone (E) were below the limit of quantification of 1 ng/mL in 62% of the samples. Biomarkers variability ranged between 31% and 41%, with a significantly lesser variability for ratios (with the exception of T/E (41%)): 20% for androsterone/etiocholanolone (p < 0.001) and 25% for 5-androstane-3,17{beta}-diol/5{beta}-androstane-3,17{beta}-diol (p < 0.001). In serum, variability for testosterone (T; 24%), androstenedione (A4; 23%), dihydrotestosterone (DHT; 19%) and T/A4 (16%) was significantly lower than urinary biomarkers (p < 0.001). Urinary A/Etio increased by > 18% after the first two weeks (p < 0.05) following blood loss. In contrast, T (0.98 nmol/L during the first week), and T/A4 (0.34 the first week) decreased significantly by more than 25% and 17% (p<0.05), respectively in the following weeks. Our results outline steroidal variations during the OCP cycle highlighting exogenous hormonal preparations as confounder for steroid concentrations in blood. Low steroid levels in urine samples have a clear detrimental impact on the subsequent interpretation of steroidal variations for the ABP. With a greater analytical sensitivity and lesser variability for steroids in serum vs. urine in healthy active women, serum represents a complementary matrix to urine in the ABP steroidal module.

physiology↗

bioRxiv2021

Controlled Administration of Dehydrochloromethyltestosterone in Humans: Urinary Excretion and Long-Term Detection of Metabolites for Anti-Doping Purpose

Dehydrochloromethyltestosterone (DHCMT) is an anabolic-androgenic steroid that was developed by Jenapharm in the 1960s and was marketed as Oral Turinabol(R). It is prohibited in sports at all times; nevertheless, there are several findings by anti-doping laboratories every year. New long-term metabolites have been proposed in 2011/12, which resulted in adverse analytical findings in retests of the Olympic games of 2008 and 2012. However, no controlled administration trial monitoring these long-term metabolites was reported until now. In this study, DHCMT (5 mg, p.o.) was administered to five healthy male volunteers and their urine samples were collected for a total of 60 days. The unconjugated and the glucuronidated fraction were analyzed separately by gas chromatography coupled to tandem mass spectrometry. The formation of the described long-term metabolites was verified, and their excretion monitored in detail. Due to interindividual differences there were several varieties in the excretion profiles among the volunteers. The metabolite M3, which has a fully reduced A-ring and modified D-ring structure, was identified by comparison with reference material as 4-chloro-17{beta}-hydroxymethyl-17-methyl-18-nor-5-androstan-13-en-3-ol. It was found to be suitable as long-term marker for the intake of DHCMT in four of the volunteers. In one of the volunteers, it was detectable for 45 days after single oral dose administration. However, in two of the volunteers M5 (already published as long-term metabolite in the 1990s) showed longer detection windows. In one volunteer M3 was undetectable but another metabolite, M2, was found as the longest detectable metabolite. The last sample clearly identified as positive was collected between 9.9 and 44.9 days. Furthermore, the metabolite epiM4 (partially reduced A-ring and a modified D-ring structure which is epimerized in position 17 compared to M3) was identified in the urine of all volunteers with the help of chemically synthesized reference as 4-chloro-17-hydroxymethyl-17{beta}-methyl-18-nor-androsta-4,13-dien-3{beta}-ol. It may serve as additional confirmatory metabolite. It is highly recommended to screen for all known metabolites in both fractions, glucuronidated and unconjugated, to improve identification of cheating athletes. This study also offers some deeper insights into the metabolism of DHCMT and of 17-methyl steroids in general.

pharmacology and toxicology↗

bioRxiv2019

Targeting the Administration of Ecdysterone in Doping Control Samples

PurposeThe phytosteroid ecdysterone was recently reported to enhance performance in sports and may thus be considered as a substance of relevance in anti-doping control. To trace back an administration of ecdysterone from urine samples analytical properties have been investigated to assess its integration into initial testing procedures (ITP) in doping control laboratories.\n\nMethodsAnalytical properties of ecdysterone were evaluated using GC-QTOF-MS and LC-QTOF-MS. Its metabolism and elimination in human were studied using urines collected after administration.\n\nResultsThe detectability of ecdysterone by GC-MS (after derivatization) and/or LC-MS(/MS) has been demonstrated and sample preparation methods were evaluated. Dilute-and-inject for LC-MS(/MS) or SPE using Oasis HLB for GC-MS or LC-MS were found most suitable, while liquid-liquid extraction was hampered by the high polarity of ecdysteroids.\n\nMost abundantly, ecdysterone was detected in the post administration urines as parent compound besides the metabolite desoxy-ecdysterone. Additionally desoxy-poststerone was tentatively assigned as minor metabolite, however further investigations are needed.\n\nConclusionAn administration of ecdysterone can be targeted using existing procedures of anti-doping laboratories. Ecdysterone and desoxy-ecdysterone appeared as suitable candidates for integration in ITP. Using dilute-and-inject a detection of the parent compound was possible for more than two days after the administration of a single dose of ~50 mg.

pharmacology and toxicology↗