Physiopathological role of the enzymatic complex 5α-reductase and 3α/β-hydroxysteroid oxidoreductase in the generation of progesterone and testosterone neuroactive metabolites

The enzymatic complex 5\u3b1-reductase (5\u3b1-R) and 3\u3b1/3\u3b2-hydroxysteroid oxidoreductase (HSOR) is expressed in the nervous system, where it transforms progesterone (PROG) and testosterone (T) into neuroactive metabolites. These metabolites regulate myelination, brain maturation, neurotransmission, reproductive behavior and the stress response. The expression of 5\u3b1-R and 3\u3b1-HSOR and the levels of PROG and T reduced metabolites show regional and sex differences in the nervous system and are affected by changing physiological conditions as well as by neurodegenerative and psychiatric disorders. A decrease in their nervous tissue levels may negatively impact the course and outcome of some pathological events. However, in other pathological conditions their increased levels may have a negative impact. Thus, the use of synthetic analogues of these steroids or 5\u3b1-R modulation have been proposed as therapeutic approaches for several nervous system pathologies. However, further research is needed to fully understand the consequences of these manipulations, in particular with 5\u3b1-R inhibitors

Sex differences in steroid levels and steroidogenesis in the nervous system : Physiopathological role

The nervous system, in addition to be a target for steroid hormones, is the source of a variety of neuroactive steroids, which are synthesized and metabolized by neurons and glial cells. Recent evidence indicates that the expression of neurosteroidogenic proteins and enzymes and the levels of neuroactive steroids are different in the nervous system of males and females. We here summarized the state of the art of neuroactive steroids, particularly taking in consideration sex differences occurring in the synthesis and levels of these molecules. In addition, we discuss the consequences of sex differences in neurosteroidogenesis for the function of the nervous system under healthy and pathological conditions and the implications of neuroactive steroids and neurosteroidogenesis for the development of sex-specific therapeutic interventions

Altered methylation pattern of the SRD5A2 gene in the cerebrospinal fluid of post-finasteride patients: A pilot study

Context: Post-finasteride syndrome (PFS) occurs in patients with androgenic alopecia after suspension of the finasteride treatment, leading to a large variety of persistent side effects. Despite the severity of the clinical picture, the mechanism underlying the PFS symptoms onset and persistence is still unclear. Objective: To study whether epigenetic modifications occur in PFS patients. Methods: Retrospective analysis of a multicentric, prospective, longitudinal, case\u2013control clinical trial, enrolling 16 PFS patients, compared to 20 age-matched healthy men. Main outcomes were methylation pattern of SRD5A1 and SRD5A2 promoters and concentration of 11 neuroactive steroids, measured by liquid chromatography-tandem mass spectrometry, in blood and cerebrospinal fluid (CSF) samples. Results: SRD5A1 and SRD5A2 methylation analysis was performed in all blood samples (n = 16 PFS patients and n = 20 controls), in 16 CSF samples from PFS patients and in 13 CSF samples from controls. The SRD5A2 promoter was more frequently methylated in CSF of PFS patients compared to controls (56.3 vs 7.7%). No promoter methylation was detected in blood samples in both groups. No methylation occurred in the SRD5A1 promoter of both groups. Unmethylated controls compared to unmethylated SRD5A2 patients showed higher pregnenolone, dihydrotestosterone and dihydroprogesterone, together with lower testosterone CSF levels. Andrological and neurological assessments did not differ between methylated and unmethylated subjects. Conclusions: For the first time, we demonstrate a tissue-specific methylation pattern of SRD5A2 promoter in PFS patients. Although we cannot conclude whether this pattern is prenatally established or induced by finasteride treatment, it could represent an important mechanism of neuroactive steroid levels and behavioural disturbances previously described in PFS

EFFECT OF FINASTERIDE TREATMENT IN MALE RAT: DEPRESSIVE-LIKE BEHAVIOR AND RELATED PARAMETERS

Neuroactive steroids are important physiological modulators of the nervous system as well as protective agents in different psychiatric and neurodegenerative diseases. This group of molecules includes hormonal steroids released by peripheral steroidogenic tissues and neurosteroids directly synthetized in nervous system. One key process regulating the synthesis of these molecules is the enzyme 5\u3b1-reductase (5\u3b1-R). This enzyme converts progesterone (PROG) and testosterone (T) to their 5\u3b1-reduced metabolites, dihydroprogesterone (DHP) and dihydrotestosterone (DHT), respectively. These metabolites are then further converted into other neuroactive steroids, such as tetrahydroprogesterone (THP) and isopregnanolone (ISOPREG) in the case of DHP and 5\u3b1- androstane-3\u3b1, 17\u3b2-diol (3\u3b1-diol) and 5\u3b1-androstane-3\u3b2, 17\u3b2-diol (3\u3b2-diol) in the case of DHT. In agreement, with the protective role exerted by neuroactive steroids their levels are affected in neurodegenerative as well as psychiatric disorders. In particular, persistent alteration of plasma neuroactive steroid levels associated with major depression has been recently reported in men after the suspension of the treatment for androgenetic alopecia with finasteride, an inhibitor of the enzyme 5\u3b1-R. Thus, these patients are affected by the so-called post-finasteride syndrome (PFS). Interestingly, suspension of finasteride treatment did not only lead, as expected, to an alteration of 5\u3b1-reduced metabolites of PROG and T but also to a global alteration of neuroactive steroid levels. Moreover, recent observations performed in male rats, show that this persistent alteration in neuroactive steroid levels did not only occur in plasma and CSF but also in the brain. On this basis, in the present PhD project, we have ascertained whether subchronic finasteride treatment (i.e. 20 days) and one month of its withdrawal may induce depressive-like behavior in this animal model. In addition, the effect of finasteride on other depressive-related parameters such as neurogenesis, gliosis, neuroinflammation/inflammation and gut microbiota have been analysed. At the end of finasteride treatment, we observed: 1) increased proliferation in the subgranular zone of the dentate gyrus; 2) increased number of microglia with reactive phenotype in the hilus and 3) increased mRNA levels of TNF-\u3b1. One month after finasteride withdrawal, we reported: 1) decreased proliferation in the subgranular zone; 2) decreased granule cell density in the granule cell layer; 3) increased astrogliosis in the hilus and 4) a possible (i.e., only detected by Student's t-test) increased of mRNA levels of IL-1\u3b2 in the hippocampus. Moreover, these latter changes coincide with the onset of depressive-like behavior, suggesting that long-term effects of finasteride treatment on neurogenesis and neuroinflammation may participate in the lasting effects of the drug on depressive-like behavior, which are detected even one month after discontinuation of the drug. In addition, peripheral inflammation assessment revealed, after drug treatment, a significant decrease in plasma levels of IL-1\u3b2 in finasteride-treated vs control animals. On the other hand, in agreement with literature and with the depressive-like behavior, a significant increase in plasma levels of IL-1\u3b2 in finasteride-treated vs control animals at withdrawal period was observed. These observations seem to support a role of peripheral inflammation in addition to what we reported on neuroinflammation. Finally, alteration of gut microbiota (i.e., an increase in Bacteroidetes phylum and in Prevotellaceae family at the end of the treatment and a decrease in Ruminococcaceae family, Oscillospira and Lachnospira genus at the end of the withdrawal period) was detected. It should be noted that important changes in the levels of neuroactive steroids such as PROG and DHP are detected in the hippocampus by one month after finasteride withdrawal. Since neuroactive steroids regulate neurogenesis, gliosis and neuroinflammation, and since PFS patients also show changes in neuroactive steroid levels, the effect of finasteride on depression, neuroactive steroid levels, neurogenesis and neuroinflammation/inflammation may be interrelated events. In addition, the changes here observed at the end of treatment and at withdrawal on gut microbiota may depict further possible signals involved in the so call gut microbiota-brain axis. In conclusion, finasteride treatment in male rats has long term effects on depressive-like behavior, hippocampal neurogenesis, neuroinflammation/inflammation and gut microbiota composition

Sex differences in the brain expression of steroidogenic molecules under basal conditions and after gonadectomy

The brain is a steroidogenic tissue. It expresses key molecules involved in the synthesis and metabolism of neuroactive steroids, such as steroidogenic acute regulatory protein (StAR), translocator protein 18 kDa (TSPO), cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc), 3\u3b2-hydroxysteroid dehydrogenases (3\u3b2-HSD), 5\u3b1-reductases (5\u3b1-R) and 3\u3b1-hydroxysteroid oxidoreductases (3\u3b1-HSOR). Previous studies have shown that the levels of brain steroids are different in male and female rats under basal conditions and after gonadectomy. In the present study, we assessed gene expression of key neurosteroidogenic molecules in the cerebral cortex and cerebellum of gonadally intact and gonadectomised adult male and female rats. In the cerebellum, the basal mRNA levels of StAR and 3\u3b1-HSOR were significantly higher in females than in males. By contrast, the mRNA levels of TSPO and 5\u3b1-R were significantly higher in males. In the cerebral cortex, all neurosteroidogenic molecules analysed showed similar mRNA levels in males and females. Gonadectomy increased the expression of 5\u3b1-R in the brain of both sexes, although it affected the brain expression of StAR, TSPO, P450scc and 3\u3b1-HSOR in females only and with regional differences. Although protein levels were not investigated in the present study, our findings indicate that mRNA expression of steroidogenic molecules in the adult rat brain is sexually dimorphic and presents regional specificity, both under basal conditions and after gonadectomy. Thus, local steroidogenesis may contribute to the reported sex and regional differences in the levels of brain neuroactive steroids and may be involved in the generation of sex differences in the adult brain function

Sex differences in steroid levels and steroidogenesis in the nervous system: Physiopathological role

The nervous system, in addition to be a target for steroid hormones, is the source of a variety of neuroactive steroids, which are synthesized and metabolized by neurons and glial cells. Recent evidence indicates that the expression of neurosteroidogenic proteins and enzymes and the levels of neuroactive steroids are different in the nervous system of males and females. We here summarized the state of the art of neuroactive steroids, particularly taking in consideration sex differences occurring in the synthesis and levels of these molecules. In addition, we discuss the consequences of sex differences in neurosteroidogenesis for the function of the nervous system under healthy and pathological conditions and the implications of neuroactive steroids and neurosteroidogenesis for the development of sex-specific therapeutic interventions

Sex differences in the brain expression of steroidogenic molecules under basal conditions and after gonadectomy

The brain is a steroidogenic tissue. It expresses key molecules involved in the synthesis and metabolism of neuroactive steroids, such as steroidogenic acute regulatory protein (StAR), translocator protein 18 kDa (TSPO), cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc), 3β-hydroxysteroid dehydrogenases (3β-HSD), 5α-reductases (5α-R) and 3α-hydroxysteroid oxidoreductases (3α-HSOR). Previous studies have shown that the levels of brain steroids are different in male and female rats under basal conditions and after gonadectomy. In the present study, we assessed gene expression of key neurosteroidogenic molecules in the cerebral cortex and cerebellum of gonadally intact and gonadectomised adult male and female rats. In the cerebellum, the basal mRNA levels of StAR and 3α-HSOR were significantly higher in females than in males. By contrast, the mRNA levels of TSPO and 5α-R were significantly higher in males. In the cerebral cortex, all neurosteroidogenic molecules analysed showed similar mRNA levels in males and females. Gonadectomy increased the expression of 5α-R in the brain of both sexes, although it affected the brain expression of StAR, TSPO, P450scc and 3α-HSOR in females only and with regional differences. Although protein levels were not investigated in the present study, our findings indicate that mRNA expression of steroidogenic molecules in the adult rat brain is sexually dimorphic and presents regional specificity, both under basal conditions and after gonadectomy. Thus, local steroidogenesis may contribute to the reported sex and regional differences in the levels of brain neuroactive steroids and may be involved in the generation of sex differences in the adult brain function.This research was supported by grants from MIUR Progetto Eccellenza and Intramural Grant Line‐B from Università degli Studi di Milano to Silvia Giatti. We also thank Agencia Estatal de Investigación, Spain (grant number BFU2017‐82754‐R), Centro de Investigación Biomédica en Red de Fragilidad y Envejecimiento Saludable (CIBERFES), Instituto de Salud Carlos III, Madrid, Spain, and Fondos Feder for financial support to L.M. Garcia‐Segura

Alterations of gut microbiota composition in post-finasteride patients: a pilot study

Purpose: Post-finasteride syndrome (PFS) has been reported in a subset of patients treated with finasteride (an inhibitor of the enzyme 5alpha-reductase) for androgenetic alopecia. These patients showed, despite the suspension of the treatment, a variety of persistent symptoms, like sexual dysfunction and cognitive and psychological disorders, including depression. A growing body of literature highlights the relevance of the gut microbiota-brain axis in human health and disease. For instance, alterations in gut microbiota composition have been reported in patients with major depressive disorder. Therefore, we have here analyzed the gut microbiota composition in PFS patients in comparison with a healthy cohort. Methods: Fecal microbiota of 23 PFS patients was analyzed by 16S rRNA gene sequencing and compared with that reported in ten healthy male subjects. Results: Sexual dysfunction, psychological and cognitive complaints, muscular problems, and physical alterations symptoms were reported in more than half of the PFS patients at the moment of sample collection. The quality sequence check revealed a low library depth for two fecal samples. Therefore, the gut microbiota analyses were conducted on 21 patients. The \u3b1-diversity was significantly lower in PFS group, showing a reduction of richness and diversity of gut microbiota structure. Moreover, when visualizing \u3b2-diversity, a clustering effect was found in the gut microbiota of a subset of PFS subjects, which was also characterized by a reduction in Faecalibacterium spp. and Ruminococcaceae UCG-005, while Alloprevotella and Odoribacter spp were increased compared to healthy control. Conclusion: Gut microbiota population is altered in PFS patients, suggesting that it might represent a diagnostic marker and a possible therapeutic target for this syndrome