Association of Hydroxy (Steroid) Dehydrogenase 11b1 Gene with Type 2 Diabetes Mellitus in Egyptian Population

Background Type 2 diabetes mellitus is a heterogeneous group of metabolic disorders caused by the interaction between genetic predisposition and environmental factors. The incidence of type 2 diabetes mellitus has increased over the last decades with more than 170 million individuals suffering from type 2 diabetes mellitus all over the world which imposes a greater economic impact on individuals, families and health systems. Though the genetic susceptibility to type 2 diabetes mellitus is polygenic, single nucleotide polymorphisms (SNP) at Hydroxy steroid dehydrogenase (HSD11B1) gene have been strongly associated with type 2 diabetes risk in various populations and ethnic groups. Objectives The aim of this study is to assess the association of Hydroxy-Steroid-Dehydrogenase-11B1 gene (rs846910) with type 2 diabetes mellitus in a sample of Egyptian population. Patients and Methods In the present study, we included 60 diabetic obese patients and 40 age and sex-matched controls. The mean age of the included patients was 50.26 ±9.1 year; while the majority of them were females (70%). All the participating patients were subjected to detailed history and the following investigations: fasting blood sugar, post prandial sugar,HbA1c, and lipid profile. Detection of gene polymorphism by real time PCR was performed for all subjects in the study. Results The study showed that homozygous GG genotype was more prelevant than GA genotype. No significant difference between GG and GA in terms of fasting and postprandial sugar and in terms of lipid profile: cholesterol (p = 0.642), TG (p = 0.808), LDL (p = 0.238), and HDL (p = 0.945).It showed no statistically significant difference between cases and controls in terms of HSD11B1 polymorphism (rs846910). The regression analysis showed that the HSD11B1 polymorphism did not significantly increase the risk of diabetes mellitus (OR 1.724, 95% CI [0.601 – 4.94]). Conclusion In conclusion, multiple further studies involving other single nucleotide polymorphism of 11 beta hydroxysteroid dehydrogenase gene as well as other genes involved in type 2 diabetes mellitus and obesity need to be done.

In vitro differentiation of theca cells from ovarian cells isolated from postmenopausal women

STUDY QUESTION Can human theca cells (TCs) be differentiated in vitro? SUMMARY ANSWER It is possible to differentiate human TCs in vitro using a medium supplemented with growth factors and hormones. WHAT IS KNOWN ALREADY There are very few studies on the origin of TCs in mammalian ovaries. Precursor TCs have been described in neonatal mice ovaries, which can differentiate into TCs under the influence of factors from oocytes and granulosa cells (GCs). On the other hand, studies in large animal models have reported that stromal cells (SCs) isolated from the cortical ovarian layer can also differentiate into TCs. STUDY DESIGN, SIZE, DURATION After obtaining informed consent, ovarian biopsies were taken from eight menopausal women (53–74 years of age) undergoing laparoscopic surgery for gynecologic disease not related to the ovaries. SCs were isolated from the ovarian cortex and in vitro cultured for 8 days in basic medium (BM) (G1), enriched with growth factors, FSH and LH in plastic (G2) or collagen substrate without (G3) or with (G4) a GC line. PARTICIPANTS/MATERIALS, SETTING, METHODS To confirm TC differentiation, relative mRNA levels for LH receptor (Lhr), steroidogenic acute regulatory protein (Star), cholesterol side-chain cleavage enzyme (Cyp11a1), cytochrome P450 17A1 (Cyp17a1), hydroxy-delta-5-steroid dehydrogenase, 3 beta- and steroid delta-isomerase 1 (Hsd3b1) and hydroxy-delta-5-steroid dehydrogenase, 3 beta- and steroid delta-isomerase 2 (Hsd3b2) were assessed. Immunohistochemistry was also performed for their protein detection and a specific marker was identified for TCs (aminopeptidase-N, CD13), as were markers for theca and small luteal cells (dipeptidyl peptidase IV (CD26) and Notch homolog 1, translocation-associated (NOTCH1)). Finally, we analyzed cell ultrastructure before (Day 0) and after in vitro culture (Day 8), and dehydroepiandrosterone (DHEA) and progesterone levels in the medium using transmission electron microscopy (TEM) and ELISA, respectively. MAIN RESULTS AND THE ROLE OF CHANCE Results obtained from qPCR showed a significant increase (P < 0.05) in mRNA levels of Lhr in F2 (floating cells in G2) and G4, Cyp17a1 in G1 and F1 (floating cells in G1) and Hsd3b2 in G1, G2, G3 and G4. Immunohistochemistry confirmed expression of each enzyme involved in the steroidogenic pathway at the protein stage. However, apart from G1, all other groups exhibited a significant (P < 0.05) rise in the number of CD13-positive cells. There was also a significant increase (P < 0.05) in NOTCH1-positive cells in G3 and G4. Ultrastructure analyses by TEM showed a distinct difference between groups and also versus Day 0. A linear trend with time revealed a significant gain (q < 0.001) in DHEA concentrations in the medium during the culture period in G1, G2, G3 and G4. It also demonstrated a statistical increase (q < 0.001) in G2, G3 and G4 groups, but G1 remained the same throughout culture in terms of progesterone levels. LARGE SCALE DATA N/A LIMITATIONS, REASONS FOR CAUTION Shorter periods of in vitro culture (e.g. 2, 4 and 6 days) could have led to increased concentrations of differentiated TCs in G2, G3 and G4. In addition, a group of cells cultured in BM and accompanied by COV434 cells would be necessary to understand their role in the differentiation process. Finally, while our results demonstrate that TCs can be differentiated in vitro from cells isolated from the cortical layer of postmenopausal ovaries, we do not know if these cells are differentiated from a subpopulation of precursor TCs present in ovarian cortex or ovarian SCs in general. It is therefore necessary to identify specific markers for precursor TCs in human ovaries to understand the origin of these cells. WIDER IMPLICATIONS OF THE FINDINGS This is a promising step toward understanding TC ontogenesis in the human ovary. Moreover, in vitro-generated human TCs can be used for studies on drug screening, as well as to understand TC-associated pathologies, such as androgen-secreting tumors and polycystic ovary syndrome. STUDY FUNDING/COMPETING INTEREST(S) This study was supported by grants from the Fonds National de la Recherche Scientifique de Belgique (FNRS) (C.A.A. is an FRS-FNRS Research Associate; grant MIS #F4535 16 awarded to C.A.A.; grant 5/4/150/5 awarded to M.M.D.; grant ASP-RE314 awarded to P.A.) and Foundation Against Cancer (grant 2018-042 awarded to A.C.). The authors declare no competing interests.

Progesterone Elevation and Preventive Strategies to Avoid Implantation Failure

Despite the wide utilization of gonadotropin-releasing hormone analogs, progesterone elevation (P4E) in the late follicular phase occurs in 5 to 30% of all ovarian stimulation (OS) cycles. Although the detrimental effect of P4E on pregnancy rates in fresh in vitro fertilization cycles is valid in all subsets of cases, higher levels of P4 and a longer duration of P4E may be needed in patients with a hyper-ovarian response in order for a negative impact on pregnancy rates to occur. Available preclinical and clinical data suggest that aggressive OS with high doses of follicle-stimulating hormone might increase 3β-hydroxy steroid dehydrogenase and 17β-hydroxy steroid dehydrogenase enzyme activity in human granulosa cells, which leads to high P4 production and hence a higher amount of leakage to the systemic circulation due to a lack of 17α-hydroxylase enzyme expression in human species. High P4 concentrations appear to alter gene expression in the endometrium; however, caution is necessary regarding its potential effect on oocyte/embryo quality with respect to the role of inherent follicular disruption in some women. In terms of the mechanism of overproduction in P4 synthesis, the main preventive strategy should be avoiding aggressive stimulation. Unfortunately, there is lack of large-scale randomized controlled trials for other approaches, including deferred embryo transfer in the thaw cycle. Since there is a significant inter-assay variability for P4 measurement, it may be wise to recommend that every center should define their own P4E and the level needed for harm to occur based on their own assays and datasets before deciding the best approach.

Nad(P) Dependent Steroid Dehydrogenase-Like, Involved In Cholesterol Biosynthesis, Regulates Proliferation And Metastasis In Breast Cancer

Background Cholesterol biosynthesis pathway is a common upregulated pathway in breast cancer. NSDHL (NAD(P) Dependent Steroid Dehydrogenase-Like), a gene which is involved in cholesterol biosynthesis has not been elucidated for its role in breast cancer. Methods After knock-down of NSDHL in human breast cancer cell lines, cell proliferation assay, cell cycle analysis, 3D culture, clonogenic assay, migration and invasion assay, and wound healing assay were performed. We tested erlotinib as a target drug for NSDHL. Xenograft was developed with MDA-MB-231 transfected with NSDHL shRNA in NOD/SCID mice. Public gene expression databases were used for the analysis of prognostic role of NSDHL in breast cancer Results Both NSDHL mRNA and protein levels were higher in a Luminal (MCF7) and triple negative cells (MDA-MB-231, BT-20) than normal breast cancer cell (MCF10A). Silencing of NSDHL by siRNA inhibited cell cycle and proliferation, and also inhibited migration and invasion in MDA-MB-231 and BT-20 cells (p< 0.05). The sensitivity to erlotinib was significantly decreased in siNSDHL treated cells, especially in MDA-MB-231 (p<0.01). Additionally, we found that NSDHL regulated total cholesterol levels in breast cancer cells. In vivo xenograft model, NSDHL knock down in the cancer cell decreased the tumor growth and metastasis. Clinically, higher NSDHL expression in the tumor of 3951 breast cancer patients was associated with significantly lower recurrence-free survival. Conclusions In conclusion, we showed that NSDHL might have a role for the progression of breast cancer. Further studies are needed to clarify that NSDHL can be a therapeutic target in breast cancer.

Antispermatogenic Effect of Piper betel Leaf Stalk Extract with Reference to Kinetic Studies of 17β-hydroxy Steroid Dehydrogenase Enzyme in Testes of Albino Rats

Background: In the development and maintenance of male reproductive function and fertility, steroidogenesis plays a key role. Aims: The aim of the present study was to investigate the effect of the betel leaf stalk extract on 17β- hydroxy steroid dehydrogenase activity levels. Methods: The observed elevation in testicular cholesterol levels may be due to decreased androgen production, which resulted in impaired spermatogenesis. The decreased steroidogenic enzyme 17β- HSD activity represents decreased androgen production by the extract administration. Reduction in enzyme active site density and Km value revealed that there was a reduction in enzyme-substrate affinities and rate of E-S complex breakdown in the administered rat testes. Results & Conclusion: The administration of betel leaf stalk extract resulted in decreased enzyme content probably through impaired synthesis.