Journal of Steroid Biochemistry and Molecular Biology最新文献

Atherosclerosis, a leading cause of cardiovascular disease, is driven by the accumulation of oxidized low-density lipoproteins (oxLDL) in arterial walls. 7-ketocholesterol (7KC), a major oxysterol found in oxLDL and atherosclerotic plaques, triggers multiple cell injuries including loss of lysosomal integrity, oxidative stress, apoptosis, and impaired autophagy in vascular cells. Bis(monoacylglycero)phosphate (BMP), also known as lysobisphosphatidic acid, is a unique phospholipid concentrated in the endolysosomal compartment, known to regulate vesicle dynamics, lysosomal enzyme activities, intracellular cholesterol trafficking and its oxidative metabolism. Using a validated model of BMP enrichment in murine RAW 264.7 macrophages, we investigated whether BMP could exert protective activity against 7KC-induced damages. Our findings revealed that BMP enrichment provides comprehensive protection against 7KC at the cellular level by preserving cell viability, morphology, and neutral lipid balance. Mechanistically, BMP enrichment prevented apoptosis by maintaining mitochondrial integrity and blocking caspase activation. This was demonstrated by normalized BAX/BCL2 ratios, preserved pro-Caspase-3 levels, and reduced PARP cleavage. Remarkably, BMP enrichment also restored autophagic flux, thereby preventing the pathological accumulation of LC3-II and p62 that characterizes autophagy dysfunction. Enhanced colocalization between LC3 and BMP suggests direct functional interactions in the stress response. Gene expression analysis confirmed that BMP enrichment normalized the transcriptional dysregulation of key autophagy regulators, including Sqstm1, Becn1, and Pink1. Taken together, these results suggest that BMP is an endogenous protective factor that counteracts 7KC-induced cellular damage at multiple steps by regulating cell death and autophagy pathways in a coordinated manner.

Historically, it was thought that primary steroids released from endocrine glands exert their hormonal effects through corresponding receptors in peripheral tissues, and that their metabolism then inactivates them, followed by excretion. However, the metabolism of primary steroids is not just a way of inactivating and excreting them, but generates a variety of metabolites with different biological properties. In this review, we outline how various active steroid metabolites were discovered, describe some of the ways they are generated, and how they can in a non-classical way act on receptors or alter the activity of steroid metabolizing enzymes, thereby indirectly affecting receptor activities. Examples include the 5α-reduced ring-A metabolites of 11-deoxycorticosterone (DOC) and progesterone that are formed in the brain, act as neurosteroids and exert effects through the GABA-A membrane receptor. Another example is 11-ketoprogesterone that potently activates mineralocorticoid receptors (MR), but not glucocorticoid receptors (GR), and is more potent than its 11β-hydroxylated form, in contrast to glucocorticoids. Moreover, we discuss the microbiome as important source of bioactive metabolites, exemplified by the 11β-hydroxylated 5α-reduced ring-A corticosteroid and progesterone metabolites that were shown as potent 11β-hydroxysteroid dehydrogenase 2 (11β-HSD2) inhibitors. 11β-HSD2 inhibition results in cortisol-induced MR activation, sodium retention and hypertension. Furthermore, microbial 17,20-desmolase activity can convert glucocorticoids to androgens, potentially influencing diseases and therapeutic outcomes. There are still many knowledge gaps regarding bioactive steroid metabolites. Identifying additional bioactive steroid metabolites and characterizing their genomic and non-genomic effects should help uncovering their cell-specific functions and contributions to the maintenance of homeostatic regulation.

In this article we summarise the current mass spectrometry methods used for analysis of 7-ketocholesterol and related sterols, focusing on the advantages of the different methods with emphasis on pre-analytical and analytical precautions to avoid artefactual ex vivo formation of these oxysterols.

Primary dysmenorrhea (PD) is a common gynecological disorder characterized by uterine hypercontraction and inflammation. This study employed an integrated strategy combining serum metabolomics, network pharmacology, and experimental validation to investigate the therapeutic potential of different extracts of Inonotus hispidus. In a rat PD model induced by estradiol benzoate and oxytocin, petroleum ether extracts of I. hispidus (MP) significantly alleviated writhing responses, attenuated uterine tissue injury, and corrected key biochemical imbalances, including the pivotal PGF₂α/PGE₂ ratio and levels of inflammatory cytokines (TNF-α, IL-6, IL-1β). Serum metabolomics analysis revealed that the therapeutic effect of MP was fundamentally linked to the systemic rectification of dysregulated arachidonic acid (AA) metabolism. Network pharmacology and subsequent experimental validation identified that MP concurrently modulates several interconnected signaling pathways. Crucially, MP activated PPARγ by promoting its dephosphorylation, which in turn potently suppressed the NLRP3 inflammasome. Concurrently, it regulated the PI3K/Akt/NF-κB survival-inflammatory axis and inhibited the RhoA/ROCK-mediated contractile pathway, reducing phosphorylation of downstream effectors MYPT1 and MLC. In vitro, the medicated serum (MP-S), containing systemically absorbed bioactive compounds, demonstrated superior and more comprehensive cytoprotection against oxytocin-induced oxidative stress, mitochondrial dysfunction, and apoptosis in rat uterine smooth muscle cells (RUSMCs) than the single compound ergosterone, underscoring the principle of multi-component synergy. Collectively, this work delineates a multi-target mechanism for MP against PD, involving the correction of AA metabolic dysregulation, activation of the PPARγ-NLRP3 axis to suppress inflammation, and inhibition of pathways driving smooth muscle hypercontraction, providing a solid scientific foundation for its development as a modern, multi-targeted therapeutic agent.

Interleukin-2 (IL-2) receptor γ (IL2RG) is present in the Leydig cell lineage, but its functional role remains elusive. To investigate how IL2RG impacts Leydig cell development from stem cells and elucidate the underlying mechanisms, we used a rat model of ethane dimethanesulfonate (EDS)-induced cell depletion in Wild-type (WT) and Il2rg knockout (KO) mice. WT and KO male rats received intratesticular IL-2 injections (1 and 10 ng/testis) from day 7 to day 21 post-EDS. Stem Leydig cells around tubules and isolated progenitor Leydig cells were cultured with IL-2 (1 and 10 ng/mL) with/without STAT3 inhibitor. Hormone levels, cell counts, gene/protein expression, and signaling pathways were analyzed. The results revealed that IL-2 significantly reduced serum testosterone levels in WT rats but had no effects in KO rats, without altering luteinizing hormone and follicle-stimulating hormone levels. Furthermore, IL-2 lowered Leydig cell numbers and the labeling index of PCNA in CYP11A1⁺ Leydig cells at 10 ng/testis, and downregulated the expression of SCARB1, HSD3B1, CYP17A1, SRD5A1, and NR5A1, as well as their corresponding mRNA levels in WT rats with no effect observed in KO rats. IL-2 also inhibited the incorporation of 5-ethynyl-2'-deoxyuridine (EdU) into stem Leydig cells in WT rats and [³H] thymidine incorporation into progenitor Leydig cells, as well as their differentiation. Importantly, the effects of IL-2 were reversed by the STAT3 inhibitor. Signaling pathway analysis showed that IL-2 exerted effects via increasing JAK1, STAT5, and STAT3 phosphorylation. This study provides insights into the inhibitory effects of IL2RG on Leydig cell development and highlights the involvement of the JAK-STAT pathway, offering potential targets for further exploration in the context of Leydig cell biology and male reproductive health.

7-ketocholesterol (7KCh), a cytotoxic oxysterol enriched in atherosclerotic plaques, provokes macrophage dysfunction through oxiapoptophagy, a linked process of oxidative stress, apoptosis and autophagy culminating in pro-inflammatory M1 polarization. Targeting this process could mitigate oxysterol-driven vascular inflammation. In this study, murine IC-21 macrophages were induced with 7KCh and co-exposed to fenchone, a bicyclic monoterpene with known anti-inflammatory properties. Cellular oxidative stress, apoptosis and autophagy were assessed by spectrofluorometric and cytometric assays. Expression of key mediators (iNOS, COX2, HO1, Casp3, Bcl2, LC3B, PARP1, Arg1, KLF4 and PPARγ) was quantified by RT-qPCR and western blotting. Molecular docking was used to identify interactions of fenchone with KLF4 and PPARγ. The results showed that 7KCh significantly increased ROS and NO production, disrupted mitochondrial membrane potential and induced apoptosis and autophagy in macrophages. Fenchone co-treatment counteracted these effects, restoring redox balance and membrane integrity. Molecular analyses revealed downregulation of iNOS, COX2, Casp3 and PARP1, alongside upregulation of HO1, Arg1, KLF4 and PPARγ. Docking analysis confirmed strong binding of fenchone to KLF4 and PPARγ, suggesting transcriptional regulation of macrophage polarization via the KLF4-PPARγ-Arg1 axis. These findings suggest that fenchone mitigates 7KCh-induced oxiapoptophagy and reprograms macrophages toward an anti-inflammatory M2 phenotype through modulation of KLF4/PPARγ signalling, positioning fenchone as a potential immunomodulatory candidate for combating oxysterol-mediated vascular inflammation.

Steroids represent a large family of organic compounds that, as signaling molecules, play important role in variety of physiological processes as control of metabolic pathways, inflammation processes, immune response, and growth, development and reproduction. Modifying the steroid core has allowed the creation of novel synthetic derivatives, which can offer significant benefits in medicine for treating a wide range of pathological conditions. Quite a number of natural steroids have been identified as effective inhibitors of cholinesterases, pointing them out as potential therapeutic alternatives in the application in Alzheimer's disease (AD). Nevertheless, only a limited number of synthetic steroids have so far been studied as cholinesterase inhibitors, highlighting an opportunity for the development of new therapeutic agents derived from natural steroidal frameworks. In this study, we selected a set of structurally diverse steroid hormone derivatives and evaluated in vitro their inhibitory activity against human AChE and BChE. IC₅₀ values were estimated for several of the most active compounds, with pyridine-containing hydroxy derivative 8 exhibiting affinity toward BChE (IC₅₀ 2.76 µM), similar to that for clinically used cholinesterase inhibitors. In silico analyses suggest that hydrophobic interactions with key amino-acid residues predominantly govern the binding of these compounds within the enzyme's active site.

Anabolic androgenic steroids are synthetic derivatives of testosterone that mimic its actions in various tissues. Due to its strong anabolic activity, weak androgenic effects, and resistance to hepatic metabolism, oxandrolone is one of the most commonly used anabolic steroids among female athletes. This study aimed to evaluate the anabolic effects of oxandrolone and its toxicological profile in female rats subjected to a strength training protocol. A total of 24 female Wistar rats (60 days old) were randomly assigned to receive oxandrolone (1.77 mg/kg/day) or its vehicle (corn oil) (n = 12 per group) via daily gavage for 28 days. The exercise protocol consisted of six climbs on an inclined ladder, with two climbs per workload (50 %, 75 %, and 100 % of each animal's maximum load) performed three times per week. Investigators remained blinded throughout experimentation and data analysis. Oxandrolone did not significantly affect body weight gain, relative organ and muscle mass, or muscle strength. However, it altered mean corpuscular volume, eosinophil count, and urea levels. Additionally, liver TBARS levels increased, while no changes were observed in plasma lipid peroxidation, antioxidant enzyme activity, total non-protein thiol levels, or mitochondrial respiratory chain complex activity. Histopathological analysis revealed oxandrolone-induced damage to cardiac and skeletal muscle, along with structural alterations in the spleen and adrenal gland. Given its limited effect on muscle strength, along with histopathological changes and increased liver lipoperoxidation, these findings raise concerns about oxandrolone use in healthy individuals seeking aesthetic or athletic benefits.