Microbial Steroid Production Technologies: Current Trends and Prospects

This Special Issue aims to collect articles and reviews on new methodologies, research, and achievements in the field of steroid microbial biotechnologies [...]

Effect of base media, FSH and anti-Müllerian hormone (AMH) alone or in combination on the growth of pig preantral follicles in vitro

To compare the efficiency of North Carolina State University medium 23 (NCSU23) and Alpha Minimum Essential Medium (α-MEM) as a base medium, and to evaluate the effects of Anti-Müllerian Hormone (AMH) alone or in combination with Follicle Stimulating Hormone (FSH) on the in vitro development and steroid production of isolated porcine preantral follicles. Porcine secondary follicles were cultured in NCSU23 or α-MEM media for 4 days. Once α-MEM was determined to be the optimal culture medium, secondary follicles were cultured in α-MEM alone or supplemented with FSH (1.5 ng/mL), AMH (50 ng/mL) or the combination of the two hormones. Follicle development was evaluated by measuring follicular growth, morphology and hormone production. There was no difference between the media NCSU23 and α-MEM in terms of follicle survival and growth (P > 0.05). However, at day 2, the antrum formation rate tended to be (P < 0.074) higher in α-MEM than NCSU23. At day 4 of culture, the estradiol and progesterone secretion were higher in α-MEM than NCSU23 (P < 0.01), while the opposite was observed for testosterone (P < 0.01). The addition of AMH and/or FSH did not affect follicular survival and growth. Nevertheless, the secretion of estradiol and progesterone induced by FSH was reduced with AMH (P < 0.01). α-MEM is a more effective base medium than NCSU23 for the in vitro follicular development of pig preantral follicles and AMH reduces the steroid production induced by FSH.

Stress-Hormone Dynamics and Working Memory in Healthy Women Who Use Oral Contraceptives Versus Non-Users

BackgroundWomen who use oral contraceptives (OCs) may have a higher risk of developing a depression, which is associated with both vulnerability to stress and cognitive dysfunction. OCs disrupt the hypothalamic-pituitary-gonadal (HPG) axis by suppressing endogenous sex steroid production including estradiol. The HPG axis and the hypothalamic-pituitary-adrenal (HPA) axis are known to interact, possibly through modulations driven by estradiol. OCs may affect HPA regulation capacity, i.e., disturb cortisol dynamics such as the cortisol awakening response (CAR), and influence cognition such as working memory (WM). We hypothesize that OC use is associated with blunted cortisol dynamics and impaired WM performance relative to non-users.MethodsData from 78 healthy women in the reproductive age were available from the CIMBI database. We evaluated if CAR and WM differed between OC users (n=25) and non-users (n=53) and if the level of estradiol modulated the OC use effect on CAR or WM in generalized least square models.ResultsWe found that OC users had a blunted CAR (p= 0.006) corresponding to a 61% reduction relative to non-users; however, no estradiol-BY-OC use interaction effect was observed on CAR. Also, OC users had higher cortisol levels at awakening compared to non-users (p = 0.03). We observed no effect of OC use or an estradiol-BY-OC use interaction effect on WM. Also, within the OC user group, neither CAR nor WM was associated with suppressed estradiol. CAR was not associated with WM.ConclusionHealthy women who use OCs have blunted cortisol dynamics relative to non-users. However, we could not detect OC use effects on working memory in our sample size. We speculate that disrupted cortisol dynamics may be important for the emergence of depressive symptoms in OC users.

Regulation of uterine function during estrous cycle, anestrus phase and pregnancy by steroids in red deer (Cervus elaphus L.)

Steroid synthesis and production in ruminant uterus is not obvious, especially in seasonally reproduced. We compared steroid production by investigating enzymes involved in red deer uterine steroid metabolism in reproductive seasons. Blood and uteri (endometrium and myometrium) were collected post mortem from hinds on 4th day (N = 8), 13th day of the cycle (N = 8), anestrus (N = 8) and pregnancy (N = 8). The expression of cytochrome P450 aromatase (P450), 3 -beta-hydroxysteroid dehydrogenase (3β-HSD), 17 -beta-hydroxysteroid dehydrogenase (17β-HSD), aldo–keto reductase family 1 C1 (AKR1C1), estrogen receptor alpha (ERα), and progesterone receptors (PRs), were analyzed using real-time-PCR and Western Blotting. Plasma samples were assayed for 17-beta-estradiol (E2), progesterone (P4), luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone (T4) concentrations by EIA. Hinds at the beginning of the estrous cycle, mainly in endometrium, were characterized by a high mRNA expression of 3β-HSD, AKR1C1, PRs and ERα, contrary to the expression in myometrium during pregnancy (P < 0.05). For P4, E2, and FSH, concentration was the highest during the 13th day of the estrous cycle (P < 0.05). Uterine steroid production and output in hinds as a representative seasonally reproduced ruminant occurred mainly during the estrous cycle and sustained in anestrus.

266 Effects of Inflammatory States on Ovarian Biology and Oocyte Competence

Female fertility is dependent on estradiol concentrations which regulate a multitude of ovarian functions including follicle development and oocyte maturation leading to ovulation of a viable oocyte. Estradiol biosynthesis is regulated by coordinated actions of follicle-stimulating hormone and intra-ovarian control mechanisms including the co-transcription factor beta-catenin. Beta-catenin is a multi-faceted protein recognized for its role in granulosa cell steroid production and is shown to be modulated by lipopolysaccharide (LPS), the endotoxin responsible for stimulation of the immune system in infections caused by Gram-negative bacteria. Beef heifers treated with subacute concentrations of LPS during a synchronized follicular wave demonstrate a decline in serum estradiol concentrations 50 h after CIDR withdrawal, corresponding with dominant follicle maturation and preceding ovulation. The endotoxin exposure also resulted in increased LPS concentration and E2:P4 ratios in follicular fluid suggesting that low dose LPS modulates the intra-follicular hormonal milieu. Additionally, in a granulosa cell line, LPS treatment decreased mRNA expression of aromatase and beta-catenin. These data indicate that LPS alters E2 synthesis by modulating beta-catenin and subsequent steroidogenic enzyme expression. To further explore the contribution of naturally occurring LPS exposure on follicular steroid production and developing oocytes, bovine ovary pairs were collected from local abattoirs. Oocytes were aspirated from small follicles and matured in vitro to evaluate meiotic events related to nuclear maturation and spindle morphology. Small follicles from ovarian pairs were separated by the detectable LPS concentrations into high and low LPS groups. Oocytes matured from low LPS follicles demonstrated an increase in the percent of abnormal maturation events. Data indicate that induced or naturally occurring low doses of LPS can alter circulating and follicular estradiol concentrations impairing oocyte maturation. Perturbation to local ovarian signaling cascades from subclinical inflammatory disease may be an unappreciated factor altering fertility and leading to decreased cow retention.

Sirt1 and Nrf2: Regulation of Leydig cell oxidant/antioxidant intracellular environment and steroid formation

Previous studies reported that, with aging, Leydig cell intracellular antioxidants are reduced in concentration and intracellular ROS levels increase, suggesting that oxidant/antioxidant imbalance may contribute to the reduced testosterone production that characterizes the aging cells. As yet, little is known about how the Leydig cell oxidant/antioxidant environment is regulated. Sirt1, an enzyme that deacetylates transcription factors, and the transcription factor Nrf2, have been shown to be associated with cellular response to oxidative stress. We hypothesized that Sirt1 and/or Nrf2 might be involved in regulating the oxidant/antioxidant environment of Leydig cells, and therefore testosterone production. We found that Sirt1 and Nrf2 are present in the Leydig cells of Brown Norway rats, though reduced in aged cells. In MA-10 cells in which Sirt1 or Nrf2 were suppressed by nicotinamide (NAM) or ML385, respectively, or in which siRNAs were used for knockdown of Sirt1 or Nrf2, increased ROS levels and decreased progesterone production occurred. In rat Leydig cells, inhibition of Sirt1 by culturing the cells with NAM resulted in increased ROS and reduced testosterone production, and subsequent removal of NAM from the culture medium resulted in increased testosterone production. Activation of rat Leydig cells Sirt1 with honokiol or of Nrf2 with sulforaphane resulted in the maintenance of testosterone production despite the exposure of the cells to oxidizing agent. These results, taken together, suggest that Sirt1 and Nrf2 are involved in maintaining the Leydig cell oxidant/antioxidant environment, and thus in maintaining steroid production.

Regulation of Uterine Function During Estrous Cycle, Anestrus Phase and Pregnancy By Steroids in Red Deer (Cervus Elaphus L.)

Steroid synthesis and production in ruminant uterus is not obvious, especially in seasonally reproduced. We compared steroid production by investigating enzymes involved in red deer uterine steroid metabolism in reproductive seasons. Blood and uteri (endometrium and myometrium) were collected post mortem from hinds on 4th day (N = 8), 13th day of the cycle (N = 8), anestrus (N = 8) and pregnancy (N = 8). The expression of cytochrome P450 aromatase (P450), 3 -beta-hydroxysteroid dehydrogenase (3β-HSD), 17 -beta-hydroxysteroid dehydrogenase (17β-HSD), aldo-keto reductase family 1 C1 (AKR1C1), estrogen receptor alpha (ERα), and progesterone receptors (PRs), were analyzed using real-time-PCR and Western Blotting. Plasma samples were assayed for 17-beta-estradiol (E2), progesterone (P4), luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone (T4) concentrations by EIA. Hinds at the beginning of the estrous cycle, mainly in endometrium, were characterized by a high mRNA expression of 3β-HSD, AKR1C1, PRs and ERα, contrary to the expression in myometrium during pregnancy (P ˂ 0.05). For P4, E2, and FSH, concentration was the highest during the 13th day of the estrous cycle (P ˂ 0.05). Uterine steroid production and output in hinds as a representative seasonally reproduced ruminant occurred mainly during the estrous cycle and sustained in anestrus.