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The Journal of Endocrinology Jun 2024Cells actively engaged in de novo steroidogenesis rely on an expansive intracellular network to efficiently transport cholesterol. The final link in the transport chain...
Cells actively engaged in de novo steroidogenesis rely on an expansive intracellular network to efficiently transport cholesterol. The final link in the transport chain is STARD1, which transfers cholesterol to the enzyme complex that initiates steroidogenesis. However, the regulation of ovarian STARD1 is not fully characterized and even less is known for upstream cytosolic cholesterol transporters STARD4 and STARD6. Here, we identified both STARD4 and STARD6 mRNAs in the human ovary but only detected STARD4 protein since the primary STARD6 transcript turned out to be a splice variant. Corpora lutea contained the highest levels of STARD4 and STARD1 mRNA and STARD1 protein, while STARD4 protein was uniformly distributed across ovarian tissues. Cyclic AMP analog (8Br-cAMP) and phorbol ester (PMA) individually increased STARD1 and STARD4 mRNA along with STARD1 protein and its phosphoform in cultured primary human luteinized granulosa cells (hGC). STARD6 transcripts and STARD4 protein were unresponsive to these stimuli. Combining lower doses of PMA and 8Br-cAMP blunted the 8Br-cAMP stimulation of STARD1 protein. Increasing cholesterol levels by blocking its conversion to steroid with aminoglutethimide or by adding LDL reduced the STARD4 mRNA response to stimuli. Sterol depletion reduced the STARD1 mRNA and protein response to PMA. These data support a possible role for STARD4, but not STARD6, in supplying cholesterol for steroidogenesis in the ovary. We demonstrate for the first time how cAMP, PMA and sterol pathways separately and combined differentially regulate STARD4, STARD6 and STARD1 mRNA levels, and STARD1 and STARD4 protein in human primary ovarian cells.
PubMed: 38829257
DOI: 10.1530/JOE-23-0385 -
Journal of Ovarian Research Feb 2024PAQR7 plays a key role in cell apoptosis as a progesterone membrane receptor. The physiological mechanism of PAQR7 in ovarian function and its anti-apoptotic action in...
PURPOSE
PAQR7 plays a key role in cell apoptosis as a progesterone membrane receptor. The physiological mechanism of PAQR7 in ovarian function and its anti-apoptotic action in mammals remain poorly understood.
METHODS
We first added 0.2 µM aminoglutethimide (AG), an inhibitor of endogenous progesterone (P4) secretion, and transfected siPAQR7 co-incubated with P4 in human KGN cells to identify granulosa cell apoptosis, respectively. Additionally, we used Paqr7 knockout (PAQR7 KO) mice to assess the role of PAQR7 in the ovary.
RESULTS
The PAQR7 deficiency significantly increased apoptosis of KGN cells, and this significant difference disappeared following P4 supplementation. The Paqr7 female mice showed a prolonged estrous cycle, reduced follicular growth, increased the number of atresia follicles, and decreased the concentrations of E2 and AMH. The litters, litter sizes, and spontaneous ovulation in the Paqr7 mice were significantly decreased compared with the Paqr7 mice. In addition, we also found low expression of PAQR7 in GCs from human follicular fluids of patients diagnosed with decreased ovarian reserve (DOR) and ovaries of mice with a DOR-like phenotype, respectively.
CONCLUSIONS
The present study has identified that PAQR7 is involved in mouse ovarian function and fertilization potential. One possible mechanism is mediating the anti-apoptotic effect of P4 on GC apoptosis via the BCL-2/BAX/CASPASE-3 signaling pathway. The mechanism underlying the effect of PAQR7 on ovarian development and aging remains to be identified.
Topics: Animals; Female; Humans; Mice; Apoptosis; Granulosa Cells; Ovarian Follicle; Ovary; Progesterone; Receptors, Progesterone; Receptors, G-Protein-Coupled
PubMed: 38317224
DOI: 10.1186/s13048-024-01348-w -
IScience Feb 2024Somatic mutations contribute to cancer development by altering the activity of enhancers. In the study, a total of 135 mutation-driven enhancers, which displayed...
Somatic mutations contribute to cancer development by altering the activity of enhancers. In the study, a total of 135 mutation-driven enhancers, which displayed significant chromatin accessibility changes, were identified as candidate risk factors for breast cancer (BRCA). Furthermore, we identified four mutation-driven enhancers as independent prognostic factors for BRCA subtypes. In Her2 subtype, enhancer G > C mutation was associated with poorer prognosis through influencing its potential target genes FBXW9, TRIR, and WDR83. We identified aminoglutethimide and quinpirole as candidate drugs targeting the mutated enhancer. In normal subtype, enhancer G > A mutation was associated with poorer prognosis through influencing its target genes ALOX15B, LINC00324, and MPDU1. We identified eight candidate drugs such as erastin, colforsin, and STOCK1N-35874 targeting the mutated enhancer. Our findings suggest that somatic mutations contribute to breast cancer subtype progression by altering enhancer activity, which could be potential candidates for cancer therapy.
PubMed: 38303701
DOI: 10.1016/j.isci.2024.108780 -
International Journal of Molecular... Feb 2023Hexaconazole is widely used as a fungicide for agricultural purposes. However, the endocrine-disrupting potential of hexaconazole is still under investigation. In...
Hexaconazole is widely used as a fungicide for agricultural purposes. However, the endocrine-disrupting potential of hexaconazole is still under investigation. In addition, an experimental study found that hexaconazole may disrupt the normal synthesis of steroidal hormones. The potency of hexaconazole to bind with sex hormone-binding globulin (SHBG), a plasma carrier protein that binds androgens and oestrogens, is unknown. In this study, we evaluated the efficacy of hexaconazole to bind with SHBG by molecular interaction, a molecular dynamics method. In addition, principal component analysis was performed to understand the dynamical behaviour of hexaconazole with SHBG in comparison with dihydrotestosterone and aminoglutethimide. The binding scores of hexaconazole, dihydrotestosterone, and aminoglutethimide with SHBG were found to be -7.12 kcal/mol, -11.41 kcal/mol, and -6.84 kcal/mol, respectively. With respect to stable molecular interaction, hexaconazole showed similar molecular dynamics patterns of root mean square deviation (RMSD), root mean square fluctuation (RMSF), radius of gyration (Rg), and hydrogen bonding. The solvent surface area (SASA) and principal component analysis (PCA) of hexaconazole exhibit similar patterns in comparison with dihydrotestosterone and aminoglutethimide. These results show that hexaconazole has a stable molecular interaction with SHBG, which may acquire the active site of the native ligand, resulting in significant endocrine disruption during agricultural work.
Topics: Aminoglutethimide; Androgens; Dihydrotestosterone; Sex Hormone-Binding Globulin; Triazoles
PubMed: 36835294
DOI: 10.3390/ijms24043882 -
Journal of Natural Medicines Mar 2023Methanol extract from the capitula of Coreopsis tinctoria Nutt. (Asteraceae), which is also known as a flowering tea or blooming tea "Snow Chrysanthemum," was found to...
Methanol extract from the capitula of Coreopsis tinctoria Nutt. (Asteraceae), which is also known as a flowering tea or blooming tea "Snow Chrysanthemum," was found to inhibit the enzymatic activity of aromatase. A total of 24 known isolates (1-24) were identified from the extract, including three chalcones (1-3), an aurone (4), five flavanones (5-9), four flavanols (10-13), a flavonol (14), and two biflavanones (15, 16). Among them, okanin (1, Ki = 1.6 μM), (2S)-naringenin (5, 0.90 μM), isookanin (6, 0.81 μM), (2S)-7,3',5'-trihydroxyflavaone (7, 0.13 μM), and (2S)-5,7,3',5'-tetrahydroxyflavanone (8, 0.32 μM) exhibited relatively potent competitive inhibition. Specifically, the isolates 7 and 8, having a common 3',5'-resorcinol moiety at the B ring in their flavanone skeleton, exhibited potent inhibitory activities compared to those of a clinically applied aminoglutethimide (0.84 μM) and naturally occurring flavone, chrysin (0.23 μM), which is a common non-steroidal aromatase inhibitor. Importantly, the active flavonoid constituents (1 and 5-8) did not inhibit the activity of 5α-reductase enzyme, which normally reacts with the same substrate "testosterone," thus, these compounds were suggested to be specific to aromatase.
Topics: Aromatase Inhibitors; Plant Extracts; Coreopsis; Aromatase; Chrysanthemum; Tea
PubMed: 36630026
DOI: 10.1007/s11418-022-01678-3 -
Turkish Journal of Pharmaceutical... Dec 2022Aromatase is an enzyme that catalyzes the conversion of androgens to estrogens. While inhibition of aromatase is a useful approach for treating breast cancer, it may...
OBJECTIVES
Aromatase is an enzyme that catalyzes the conversion of androgens to estrogens. While inhibition of aromatase is a useful approach for treating breast cancer, it may also have toxicological consequences due to its endocrine disrupting/modulating effect. In this study, sensitivity and performance of two assays -a cell free and a cell-based- for evaluating aromatase activity were investigated by testing known aromatase inhibitors and partial validation of the methods was performed. Advantages and disadvantages of these methods are also discussed.
MATERIALS AND METHODS
Aromatase activity was evaluated two models; direct measurement with a cell-free assay using a fluorescent substrate and recombinant human enzyme and indirect evaluation with a cell-based assay where cell proliferation was determined in estrogen receptor positive human breast cancer cells (MCF-7 BUS) in the absence of estrogen and the presence of testosterone.
RESULTS
In the cell-free direct measurement assay, reference compounds ketoconazole and aminoglutethimide have been shown to inhibit the aromatase enzyme with half-maximal inhibitory concentration (IC) values concordant with literature. In cell-based indirect measurement assay, only ketoconazole dose-dependently inhibited cell proliferation with 3.47 x 10 M IC. Inter-assay and intra-assay reproducibility of both methods was found to be within acceptable deviation levels.
CONCLUSION
Both methods can be successfully applied. However, to evaluate the potential aromatase activity of the novel compounds , it seems better to perform both the cell-based and the cell-free assays that allows low-moderate biotransformation and eliminate cytotoxicity potential, respectively.
PubMed: 36544280
DOI: 10.4274/tjps.galenos.2021.85530 -
International Journal of Molecular... Sep 2022With the aim of searching for phytochemicals with aromatase inhibitory activity, five new prenylcoumarins, mammeasins K (), L (), M (), N (), and O (), were isolated...
Phytochemicals with Chemopreventive Activity Obtained from the Thai Medicinal Plant (Miq.) T. Anders.: Isolation and Structure Determination of New Prenylcoumarins with Inhibitory Activity against Aromatase.
With the aim of searching for phytochemicals with aromatase inhibitory activity, five new prenylcoumarins, mammeasins K (), L (), M (), N (), and O (), were isolated from the methanolic extract of (Miq.) T. Anders. flowers (fam. Calophyllaceae), originating in Thailand. The stereostructures of - were elucidated based on their spectroscopic properties. Among the new compounds, (IC = 7.6 µM) and (9.1 µM) possessed relatively strong inhibitory activity against aromatase, which is a target of drugs already used in clinical practice for the treatment and prevention of estrogen-dependent breast cancer. The analysis through Lineweaver-Burk plots showed that they competitively inhibit aromatase (, i = 3.4 µM and , 2.3 µM). Additionally, the most potent coumarin constituent, mammea B/AB cyclo D (, i = 0.84 µM), had a competitive inhibitory activity equivalent to that of aminoglutethimide (0.84 µM), an aromatase inhibitor used in therapeutics.
Topics: Aminoglutethimide; Aromatase; Aromatase Inhibitors; Coumarins; Estrogens; Mammea; Phytochemicals; Plant Extracts; Plants, Medicinal; Thailand
PubMed: 36232534
DOI: 10.3390/ijms231911233 -
Medicine Sep 2022Airway neutrophilia has been associated with asthma severity and asthma exacerbations. This study attempted to identify biomarkers, pathogenesis, and therapeutic...
BACKGROUND
Airway neutrophilia has been associated with asthma severity and asthma exacerbations. This study attempted to identify biomarkers, pathogenesis, and therapeutic molecular targets for severe asthma in neutrophils using bioinformatics analysis.
METHODS
Fifteen healthy controls and 3 patients with neutrophilic severe asthma were screened from the Gene Expression Omnibus (GEO) database. Based on the analysis of differentially expressed genes (DEGs), functional and pathway enrichment analyses, gene set enrichment analysis, protein-protein interaction network construction, and analysis were performed. Moreover, small-molecule drug candidates have also been identified.
RESULTS
Three hundred and three upregulated and 59 downregulated genes were identified. Gene ontology function enrichment analyses were primarily related to inflammatory response, immune response, leukocyte migration, neutrophil chemotaxis, mitogen-activated protein kinase cascade, Jun N-terminal kinase cascade, I-kappaB kinase/nuclear factor-κB, and MyD88-dependent toll-like receptor signaling pathway. Pathway enrichment analyses and gene set enrichment analysis were mainly involved in cytokine-cytokine receptor interaction, the TNF signaling pathway, leukocyte transendothelial migration, and the NOD-like receptor signaling pathway. Furthermore, 1 important module and 10 hub genes (CXCL8, TLR2, CXCL1, ICAM1, CXCR4, FPR2, SELL, PTEN, TREM1, and LEP) were identified in the protein-protein interaction network. Moreover, indoprofen, mimosine, STOCK1N-35874, trapidil, iloprost, aminoglutethimide, ajmaline, levobunolol, ethionamide, cefaclor, dimenhydrinate, and bethanechol are potential drugs for the treatment of neutrophil-predominant severe asthma.
CONCLUSION
This study identified potential biomarkers, pathogenesis, and therapeutic molecular targets for neutrophil-predominant severe asthma.
Topics: Ajmaline; Aminoglutethimide; Asthma; Bethanechol; Biomarkers; Cefaclor; Computational Biology; Cytokines; Dimenhydrinate; Ethionamide; Gene Expression Profiling; Humans; Iloprost; Indoprofen; JNK Mitogen-Activated Protein Kinases; Levobunolol; Mimosine; Mitogen-Activated Protein Kinases; Myeloid Differentiation Factor 88; NF-kappa B; NLR Proteins; Neutrophils; Receptors, Cytokine; Toll-Like Receptor 2; Trapidil; Triggering Receptor Expressed on Myeloid Cells-1
PubMed: 36197221
DOI: 10.1097/MD.0000000000030661 -
ACS Omega Jun 2022[This corrects the article DOI: 10.1021/acsomega.2c00011.].
Correction to "Synthesis and Characterization of an Inclusion Complex of dl-Aminoglutethimide with β-Cyclodextrin and Its Innovative Application in a Biological System: Computational and Experimental Investigations".
[This corrects the article DOI: 10.1021/acsomega.2c00011.].
PubMed: 35755346
DOI: 10.1021/acsomega.2c03057 -
The Journal of Biological Chemistry Jul 2022Neurosteroids, modulators of neuronal and glial cell functions, are synthesized in the nervous system from cholesterol. In peripheral steroidogenic tissues, cholesterol...
Neurosteroids, modulators of neuronal and glial cell functions, are synthesized in the nervous system from cholesterol. In peripheral steroidogenic tissues, cholesterol is converted to the major steroid precursor pregnenolone by the CYP11A1 enzyme. Although pregnenolone is one of the most abundant neurosteroids in the brain, expression of CYP11A1 is difficult to detect. We found that human glial cells produced pregnenolone, detectable by mass spectrometry and ELISA, despite the absence of observable immunoreactive CYP11A1 protein. Unlike testicular and adrenal cortical cells, pregnenolone production in glial cells was not inhibited by CYP11A1 inhibitors DL-aminoglutethimide and ketoconazole. Furthermore, addition of hydroxycholesterols increased pregnenolone synthesis, suggesting desmolase activity that was not blocked by DL-aminoglutethimide or ketoconazole. We explored three different possibilities for an alternative pathway for glial cell pregnenolone synthesis: (1) regulation by reactive oxygen species, (2) metabolism via a different CYP11A1 isoform, and (3) metabolism via another CYP450 enzyme. First, we found oxidants and antioxidants had no significant effects on pregnenolone synthesis, suggesting it is not regulated by reactive oxygen species. Second, overexpression of CYP11A1 isoform b did not alter synthesis, indicating use of another CYP11A1 isoform is unlikely. Finally, we show nitric oxide and iron chelators deferoxamine and deferiprone significantly inhibited pregnenolone production, indicating involvement of another CYP450 enzyme. Ultimately, knockdown of endoplasmic reticulum cofactor NADPH-cytochrome P450 reductase had no effect, while knockdown of mitochondrial CYP450 cofactor ferredoxin reductase inhibited pregnenolone production. These data suggest that pregnenolone is synthesized by a mitochondrial cytochrome P450 enzyme other than CYP11A1 in human glial cells.
Topics: Aminoglutethimide; Cholesterol; Cholesterol Side-Chain Cleavage Enzyme; Humans; Ketoconazole; Neuroglia; Neurosteroids; Pregnenolone; Reactive Oxygen Species
PubMed: 35688208
DOI: 10.1016/j.jbc.2022.102110