Showing papers on "Pregnenolone published in 2022"

Desorption Electrospray Ionization Mass Spectrometry Assay for Label‐Free Characterization of SULT2B1b Enzyme Kinetics

Abstract: The sulfotransferase (SULT) 2B1b, which catalyzes the sulfonation of 3β‐hydroxysteroids, has been identified as a potential target for prostate cancer treatment. However, a major limitation for SULT2B1b‐targeted drug discovery is the lack of robust assays compatible with high‐throughput screening and inconsistency in reported kinetic data. For this reason, we developed a novel label‐free assay based on high‐throughput (>1 Hz) desorption electrospray ionization mass spectrometry (DESI‐MS) for the direct quantitation of the sulfoconjugated product (CV<10 %; <1 ng analyte). The performance of this DESI‐based assay was compared against a new fluorometric coupled‐enzyme method that we also developed. Both methodologies provided consistent kinetic data for the reaction of SULT2B1b with its major substrates, indicating the affinity trend pregnenolone>DHEA>cholesterol, for both the phospho‐mimetic and wild‐type SULT2B1b forms. The novel DESI‐MS assay developed here is likely generalizable to other drug discovery efforts and is particularly promising for identification of SULT2B1b inhibitors with potential as prostate cancer therapeutics.

A preliminary study on the relationship between environmental endocrine disruptors and precocious puberty in girls

Abstract: Abstract Objectives To explore the associations of environmental endocrine disruptors on precocious puberty in girls. Methods This was a case-control study in which 30 girls with precocious puberty and 46 age- and race-matched prepubertal females were enrolled. The concentrations of 10 environment endocrine disruptors (bisphenol A, bisphenol B, butylparaben, propylparaben, ethvlparaben, methylparaben, mono-butyl phthalate, mono-2-ethylhexyl phthalate, monoethyl phthalate, and monomethyl phthalate) in urine and 10 steroid hormones (dihydrotestosterone, corticosterone, hydrocortisone, 11-deoxycortisol, 17α-hydroxy progesterone, 4-androstene-3,17-dione, estrone, deoxycorticosterone, pregnenolone, and dehydroepiandrosterone) in serum were detected with the liquid chromatography-mass spectrometry (LC-MS). Results According to the Mann–Whitney U test, urinary levels of bisphenol A, monobutyl phthalate, and monomethyl phthalate were significantly higher in the precocious group than in the prepubertal group, and blood levels of hydrocortisone, 11-deoxycortisol, corticosterone, deoxycorticosterone, and pregnenolone were significantly lower in the precocious group than in the prepubertal group (p<0.05, VIP>1). Conclusions Our findings confirm the association between phthalate exposure and the incidence of precocious puberty in girls. Control and reduction of children exposure to phthalate esters should be considered as a health priority.

Cross‐talk between neurosteroid and endocannabinoid systems in cannabis addiction

Abstract: Steroids and endocannabinoids are part of two modulatory systems and some evidence has shown their interconnections in several functions. Homeostasis is a common steady‐state described in the body, which is settled by regulatory systems to counterbalance deregulated or allostatic set points towards an equilibrium. This regulation is of primary significance in the central nervous system for maintaining neuronal plasticity and preventing brain‐related disorders. In this context, the recent discovery of the shutdown of the endocannabinoid system (ECS) overload by the neurosteroid pregnenolone has highlighted new endogenous mechanisms of ECS regulation related to cannabis‐induced intoxication. These mechanisms involve a regulatory loop mediated by overactivation of the central type‐1 cannabinoid receptor (CB1R), which triggers the production of its own regulator, pregnenolone. Therefore, this highlights a new process of regulation of steroidogenesis in the brain. Pregnenolone, long considered an inactive precursor of neurosteroids, can then act as an endogenous negative allosteric modulator of CB1R. The present review aims to shed light on a new framework for the role of ECS in the addictive characteristics of cannabis with the novel endogenous mechanism of ECS involving the neurosteroid pregnenolone. In addition, this new endogenous regulatory loop could provide a relevant therapeutic model in the current context of increasing recreational and medical use of cannabis.

Gut Steroids and Microbiota: Effect of Gonadectomy and Sex

Abstract: Sex steroids, derived mainly from gonads, can shape microbiota composition; however, the impact of gonadectomy and sex on steroid production in the gut (i.e., gut steroids), and its interaction with microbiota composition, needs to be clarified. In this study, steroid environment and gut steroidogenesis were analysed by liquid chromatography tandem mass spectrometry and expression analyses. Gut microbiota composition as branched- and short-chain fatty acids were determined by 16S rRNA gene sequence analysis and gas chromatography flame ionisation detection, respectively. Here, we first demonstrated that levels of pregnenolone (PREG), progesterone (PROG), and isoallopregnanolone (ISOALLO) were higher in the female rat colon, whereas the level of testosterone (T) was higher in males. Sexual dimorphism on gut steroidogenesis is also reported after gonadectomy. Sex, and more significantly, gonadectomy, affects microbiota composition. We noted that a number of taxa and inferred metabolic pathways were associated with gut steroids, such as positive associations between Blautia with T, dihydroprogesterone (DHP), and allopregnanolone (ALLO), whereas negative associations were noted between Roseburia and T, ALLO, PREG, ISOALLO, DHP, and PROG. In conclusion, this study highlights the novel sex-specific association between microbiota and gut steroids with possible relevance for the gut-brain axis.

Pregnenolone Attenuates the Ischemia-Induced Neurological Deficit in the Transient Middle Cerebral Artery Occlusion Model of Rats

Abstract: Neurosteroids are apparent to be connected in the cerebral ischemic injury for their potential neuroprotective effects. We previously demonstrated that progesterone induces neuroprotection via the mitochondrial cascade in the cerebral ischemic stroke of rodents. Here, we sought to investigate whether or not pregnenolone, a different neurosteroid, can protect the ischemic injury in the transient middle cerebral artery occlusion (tMCAO) rodent model. Male Wistar rats were chosen for surgery for inducing stroke using the tMCAO method. Pregnenolone (2 mg/kg b.w.) at 1 h postsurgery was administered. The neurobehavioral tests and (TTC staining) 2, 3, 5-triphenyl tetrazolium chloride staining were performed after 24 h of the surgery. The mitochondrial membrane potential and reactive oxygen species (ROS) were measured using flow cytometry. Oxygraph was used to examine mitochondrial bioenergetics. The spectrum of neurobehavioral tests and 2, 3, 5-triphenyltetrazolium chloride staining showed that pregnenolone enhanced neurological recovery. Pregnenolone therapy after a stroke lowered mitochondrial ROS following ischemia. Our data demonstrated that pregnenolone was not able to inhibit mitochondrial permeability transition pores. There was no effect on mitochondrial bioenergetics such as oxygen consumption and respiratory coupling. Overall, the findings demonstrated that pregnenolone reduced the neurological impairments via reducing mitochondria ROS but not through the inhibition of the mitochondria permeability transition pore (mtPTP).

Common variability in oestrogen-related genes and pancreatic ductal adenocarcinoma risk in women

Abstract: The incidence of pancreatic ductal adenocarcinoma (PDAC) is different among males and females. This disparity cannot be fully explained by the difference in terms of exposure to known risk factors; therefore, the lower incidence in women could be attributed to sex-specific hormones. A two-phase association study was conducted in 12,387 female subjects (5436 PDAC cases and 6951 controls) to assess the effect on risk of developing PDAC of single nucleotide polymorphisms (SNPs) in 208 genes involved in oestrogen and pregnenolone biosynthesis and oestrogen-mediated signalling. In the discovery phase 14 polymorphisms showed a statistically significant association (P < 0.05). In the replication none of the findings were validated. In addition, a gene-based analysis was performed on the 208 selected genes. Four genes (NR5A2, MED1, NCOA2 and RUNX1) were associated with PDAC risk, but only NR5A2 showed an association (P = 4.08 × 10-5) below the Bonferroni-corrected threshold of statistical significance. In conclusion, despite differences in incidence between males and females, our study did not identify an effect of common polymorphisms in the oestrogen and pregnenolone pathways in relation to PDAC susceptibility. However, we validated the previously reported association between NR5A2 gene variants and PDAC risk.

Determination of 19 Steroid Hormones in Human Serum and Urine Using Liquid Chromatography-Tandem Mass Spectrometry

Abstract: This paper describes a methodology for simultaneous determination of 19 steroid hormones, viz. estrone, estradiol, estriol, testosterone, 5α-dihydrotestosterone, androstenedione, androstenediol, dehydroepiandrosterone, progesterone, pregnenolone, 17α-OH-progesterone, 17α-OH-pregnenolone, cortisone, cortisol, 11-deoxycortisol, 11-deoxycorticosterone, 11-dehydrocorticosterone, aldosterone, and corticosterone, in 500-µL of urine or serum/plasma. The method was optimized using isotopically labeled internal standards and liquid-liquid extraction followed by detection using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-MS/MS). Dansylation of estrogens significantly improved their sensitivities (~11- to 23-fold) and chromatographic separation. The respective limit of detection (LOD) and limit of quantification (LOQ) of all analytes were 0.04–0.28 and 0.14–0.92 ng/mL in human urine, and 0.11–0.35 and 0.38–1.18 ng/mL in human serum/plasma. Recoveries of all analytes (except for progesterone) fortified at 10, 20, and 200 ng/mL in urine and serum were 80–120%, with standard deviations ranging from 0 to 17.3%. Repeated analysis of similarly fortified urine and serum samples yielded intra-day and inter-day variations of 0–21.7% and 0.16–11.5%, respectively. All analytes except cortisone exhibited weak matrix effects in urine and serum (−13.9–18.2%). The method was further validated through the analysis of the National Institute of Standards and Technology (NIST) plasma Standard Reference Material (SRM1950) with certified concentrations for cortisol, progesterone, and testosterone (coefficient of variation: 3–11%). The developed method was applied in the analysis of urine samples from 20 volunteers, which revealed the occurrence of 16 analytes with detection frequencies (DFs) > 80%. Furthermore, 15 analytes were found in plasma SRM1950, indicating the feasibility of our method in the analysis of steroid hormones in urine and serum/plasma. This method will facilitate analysis of steroid hormones in population-based biomonitoring studies.

Regulation of retinoid mediated StAR transcription and steroidogenesis in hippocampal neuronal cells: Implications for StAR in protecting Alzheimer's disease.

Abstract: Retinoids (vitamin A and its derivatives) play pivotal roles in diverse processes, ranging from homeostasis to neurodegeneration, which are also influenced by steroid hormones. The rate-limiting step in steroid biosynthesis is mediated by the steroidogenic acute regulatory (StAR) protein. In the present study, we demonstrate that retinoids enhanced StAR expression and pregnenolone biosynthesis, and these parameters were markedly augmented by activation of the PKA pathway in mouse hippocampal neuronal HT22 cells. Deletion and mutational analyses of the 5'-flanking regions of the StAR gene revealed the importance of a retinoic acid receptor (RAR)/retinoid X receptor (RXR)-liver X receptor (LXR) heterodimeric motif at -200/-185 bp region in retinoid responsiveness. The RAR/RXR-LXR sequence motif can bind RARα and RXRα, and retinoid regulated transcription of the StAR gene was found to be influenced by the LXR pathway, representing signaling cross-talk in hippocampal neurosteroid biosynthesis. Steroidogenesis decreases during senescence due to declines in the central nervous system and the endocrine system, and results in hormone deficiencies, inferring the need for hormonal balance for healthy aging. Loss of neuronal cells, involving accumulation of amyloid beta (Aβ) and/or phosphorylated Tau within the brain, is the pathological hallmark of Alzheimer's disease (AD). HT22 cells overexpressing either mutant APP (mAPP) or mutant Tau (mTau), conditions mimetic to AD, enhanced toxicities, and resulted in attenuation of both basal and retinoid-responsive StAR and pregnenolone levels. Co-expression of StAR with either mAPP or mTau diminished cytotoxicity, and concomitantly elevated neurosteroid biosynthesis, pointing to a protective role of StAR in AD. These findings provide insights into the molecular events by which retinoid signaling upregulates StAR and steroid levels in hippocampal neuronal cells, and StAR, by rescuing mAPP and/or mTau-induced toxicities, modulates neurosteroidogenesis and restores hormonal balance, which may have important implications in protecting AD and age-related complications and diseases.

COVID-19 patients with altered steroid hormone levels are more likely to have higher disease severity

Abstract: This study aims to evaluate the correlations between the severity of the disease and serum steroid levels by analyzing the serum steroid levels in COVID-19 patients with different levels of disease progression and the control group. Morning serum Aldosterone, 11-deoxycortisol, Androstenedione, 17-hydroxyprogesterone, Dihydrotestosterone (DHT), Dehydroepiandrosterone (DHEA), Corticosterone, Dehydroepiandrosterone sulfate (DHEAS), Estrone, Estradiol, Progesterone, 11-deoxycorticosterone, Cortisol, Corticosterone, Androsterone, Pregnenolone, 17-hydroxypregnenolone and 21-deoxycortisol levels were measured in 153 consecutive patients were grouped as mild, moderate, and severe based on the WHO COVID-19 disease severity classification and the control group. Steroid hormone levels were analyzed at once with a liquid chromatography-tandem mass spectrometric method (LC-MS/MS). In our study, nearly all steroids were statistically significantly higher in the patients’ group than in the control group (p < 0.001). Also, DHEA was an independent indicator of the disease severity with COVID-19 Our study reveals that the alteration in steroid hormone levels was correlated with disease severity. Also, steroid hormone levels should be followed up during COVID-19 disease management.

Pregnenolone Reduces Stress-Induced Craving, Anxiety, and Autonomic Arousal in Individuals with Cocaine Use Disorder

Abstract: Chronic cocaine use leads to adaptations in stress biology and in neuroactive steroid system. These adaptations are associated with high cocaine craving and increased relapse risk. This study tested whether potentiation of the neuroactive steroid system with the precursor pregnenolone (PREG) affects stress- and cue-induced cocaine craving, anxiety and autonomic response in individuals with cocaine use disorder (CUD). Thirty treatment-seeking individuals (21 Male, 9 Female) with CUD were randomized to placebo (PBO) or supraphysiologic PREG doses of 300 mg or 500 mg per day for 8 weeks. After 2 weeks of treatment, participants were exposed to 5-min personalized guided imagery provocation of stress, cocaine, or neutral/relaxing cues in a 3-day experiment, one condition per day on separate days, in a random, counterbalanced order. Repeated assessment of cocaine craving, anxiety, heart rate (HR), systolic (SBP) and diastolic blood pressure (DBP) were assessed on each day. PREG significantly increased pregnenolone levels compared to PBO. Both PREG doses decreased stress- and cocaine cue-induced craving and reduced both stress- and cue-induced anxiety only in the 500 mg/day group. The 500 mg/day PREG group also displayed decreased stress-induced HR, SBP and DBP. Findings indicate that pregnenolone decreases stress- and cocaine cue-provoked craving and anxiety and reduces stress-induced autonomic arousal in individuals with CUD.

Erratum: Krausova et al., “Site of Action of Brain Neurosteroid Pregnenolone Sulfate at the N-Methyl-D-Aspartate Receptor”

Abstract: In the article “Site of Action of Brain Neurosteroid Pregnenolone Sulfate at the N-Methyl-D-Aspartate Receptor,” by Barbora Hrcka Krausova, Bohdan Kysilov, Jiri Cerny, Vojtech Vyklicky, Tereza Smejkalova, Marek Ladislav, Ales Balik, Miloslav Korinek, Hana Chodounska, Eva Kudova, and Ladislav