Revisiting the action of steroids and triterpenoids on the human sperm Ca2+ channel CatSper

ABSTRACT The sperm-specific Ca2+ channel CatSper (cation channel of sperm) is vital for male fertility. Contradictory findings have been published on the regulation of human CatSper by the endogenous steroids estradiol, testosterone and hydrocortisone, as well as the plant triterpenoids, lupeol and pristimerin. The aim of this study was to elucidate this controversy by investigating the action of these steroids and plant triterpenoids on human CatSper using population-based Ca2+-fluorimetric measurements, the specific CatSper-inhibitor RU1968 and a functional test assessing the CatSper-dependent penetration of human sperm cells into methylcellulose. Estradiol, testosterone and hydrocortisone were found to induce Ca2+-signals in human sperm cells with EC50 values in the lower μM range. By employing the specific CatSper-inhibitor RU1968, all three steroids were shown to induce Ca2+-signals through an action on CatSper, similar to progesterone. The steroids were found to dose-dependently inhibit subsequent progesterone-induced Ca2+-signals with IC50 values in the lower μM range. Additionally, the three steroids were found to significantly increase the penetration of human sperm cells into methylcellulose, similar to the effect of progesterone. The two plant triterpenoids, lupeol and pristimerin, were unable to inhibit progesterone-induced Ca2+-signals, whereas the CatSper-inhibitor RU1968 strongly inhibited progesterone-induced Ca2+-signals. In conclusion, this study supports the claim that the steroids estradiol, testosterone and hydrocortisone act agonistically on CatSper in human sperm cells, thereby mimicking the effect of progesterone, and that lupeol and pristimerin do not act as inhibitors of human CatSper.

A Simple, Low Cost, Sensitive, and Portable Electrochemical Immunochromatography Sensing Device to Measure Estrone-3-Sulfate

In livestock production, point-of-care testing (POCT) technology that enables easy on-site analysis of sex hormones is desired to improve reproductive efficiency. In this context, low-molecular-weight endogenous steroids are particularly important for perinatal management. Therefore, we attempted to use a simple method that combines electrochemical techniques with immunochromatography to measure estrone-3-sulfate (E1S), one of the low-molecular-weight endogenous steroids that is an estrogen ester. The limit of detection (LOD) for E1S achieved by electrochemical immunochromatography was 570.5 ng/mL, which was one to two orders of magnitude lower than that of small molecule compounds analyzed by other POCT techniques (Primpray et al., Anal. Chim. Acta, 2019). In addition, it was indicated by a colorimetric analysis that the sensitivity of the electrochemical immunochromatographic technique could be enhanced by improving the method of application of the antibodies on the nitrocellulose membrane and the contact between the electrochemical detector and the nitrocellulose membrane.

Influence of Pain Killers on the Urinary Anabolic Steroid Profile

Anabolic androgenic steroids (AAS) are prohibited as performance-enhancing drugs in sports. Among them, testosterone and its precursors are often referred to as “pseudoendogenous” AAS, that is, endogenous steroids that are prohibited when administered exogenously. To detect their misuse, among other methods, the World Anti-Doping Agency-accredited laboratories monitor the steroid profile (concentrations and concentration ratios of endogenous steroids, precursors and metabolites) in urine samples collected from athletes in and out of competition. Alterations in steroid profile markers are used as indicators for misuse of anabolic steroids in sports. Therefore, especially their metabolic pathways with possible interactions are crucial to elucidate. As steroid metabolism is very complex, and many enzymes are involved, certain non-prohibited drugs may influence steroid metabolite excretion. One important group of steroid-metabolizing enzymes is aldo–keto reductases (AKRs). An inhibition of them by non-steroidal anti-inflammatory drugs (NSAIDs), which are neither prohibited nor monitored, but frequently used drugs in sports, was demonstrated in vitro. Thus, this work aims to investigate the influence of NSAID intake on the urinary steroid profile. Kinetic and inhibitory studies were performed using 5α-dihydrotestosterone as substrate. The results obtained from in vitro experiments show that ibuprofen inhibits AKR1C2 and thus influences steroid biotransformation. For in vivo investigations, urine samples prior, during and postadministration of ibuprofen were analyzed using routine methods to monitor the steroid profile. Changes in markers of the steroid profile of volunteers were observed. The combination of in vitro and in vivo results suggests that monitoring of ibuprofen may be useful in doping control analysis. The presented work illustrates the importance to consider co-administration of (non-prohibited) drugs during antidoping analysis. Intake of multiple substances is likely leading to interfering effects. Divergent results in antidoping analysis may therefore be observed and misinterpretation of analytical data may occur. Similar considerations may be appropriate for other fields of forensic applications.

A method for determination of one hundred endogenous steroids in human serum by gas chromatography-tandem mass spectrometry.

Steroid profiling helps various pathologies to be rapidly diagnosed. Results from analyses investigating steroidogenic pathways may be used as a tool for uncovering pathology causations and proposals of new therapeutic approaches. The purpose of this study was to address still underutilized application of the advanced GC-MS/MS platform for the multicomponent quantification of endogenous steroids. We developed and validated a GC-MS/MS method for the quantification of 58 unconjugated steroids and 42 polar conjugates of steroids (after hydrolysis) in human blood. The present method was validated not only for blood of men and non-pregnant women but also for blood of pregnant women and for mixed umbilical cord blood. The spectrum of analytes includes common hormones operating via nuclear receptors as well as other bioactive substances like immunomodulatory and neuroactive steroids. Our present results are comparable with those from our previously published GC-MS method as well as the results of others. The present method was extended for corticoids and 17α-hydroxylated 5α/β-reduced pregnanes, which are useful for the investigation of alternative “backdoor” pathway. When comparing the analytical characteristics of the present and previous method, the first exhibit by far higher selectivity, and generally higher sensitivity and better precision particularly for 17α-hydroxysteroids.