Primary Follicles Research Articles

Effect of cod liver oil supplementation in commercial feed on reproductive organs weight and ovarian follicles size of local rabbits

This study aimed to determine the effect of cod liver oil supplementation in commercial feed on reproductive organs weight and ovarian follicles size of local rabbits. The experimental design used was Completely Randomized Design with four feed treatments, i.e. commercial feed without supplementation of cod liver oil (CLO) as control (P0), commercial feed supplemented by 3% CLO (P1), 4.5% CLO (P2), and 6% CLO (P3). Each treatment consisted of ten rabbits as replication and treatment were given to rabbits from aged 4 to 6 months. The parameters observed were the weight of the reproductive organs, namely the ovaries, fallopian tubes, uterus and the size of the ovarian follicles, namely primary, secondary, tertiary and preovulatory follicles of local rabbits. The results showed that various levels of cod liver oil supplementation in commercial feed differed significantly (P<0.05) to the weight of the reproductive organs and the size of the ovarian follicles of local rabbits. It can be concluded that cod liver oil supplementation at 4.5% level in commercial feed capable increase the reproductive organs weight and ovarian follicles size of local rabbits.

Primera caracterización de folículos preantrales de ovario de alpaca adulta

It is a fact that while even basic reproductive information on alpacas is unavailable, the normal ovarian reserve of this species in comparison to other species is also unidentified. In this study, the ovarian preantral follicles in healthy adult alpacas were characterized in order to establish a general model to in vitro studies. Ten ovaries were collected from five adult alpacas. The ovarian cortex samples were fixed with paraformaldehyde and histological analysis was done. Normal and degenerated follicles percentages were determined. The normal follicles were measured and classified in primordial, transitional, primary and secondary stages. Most of the preantral follicles present in the ovarian cortex of alpacas were primordial and transitional stages; primary (6.10%) and secondary (0.37%) follicles were rarely found. The primary and secondary follicles were larger in diameter when compared with the primordial and transitional follicles. The largest oocyte diameter was recorded in the secondary follicles (P < 0.05). This study serves to establish a biological model for future reproduction studies in Alpacas or as possible biological model for studies of folliculogenesis in humans.

Differentially Expressed lncRNAs After the Activation of Primordial Follicles in Mouse

The activation of primordial follicles is critical to ovarian follicle development, which directly influences female fertility and reproductive life span. Several studies have suggested a role for long noncoding RNAs (lncRNAs) in ovarian function. However, the precise involvement of lncRNAs in the initiation of primordial follicles is still unknown.Here, an in vitro culture model was used to investigate the roles of lncRNAs in primordial follicle activation.We found that primordial follicles in day 3 mouse ovaries were activated after culturing for 8 days in vitro, as indicated by ovarian morphology changes, increases in primary follicle number, and downregulation of mammalian Sterile 20-like kinase messenger RNA (mRNA) and upregulation of growth differentiation factor 9 mRNA. We next examined lncRNA expression profiles by RNA sequencing at the transcriptome level and found that among 60 078 lncRNAs, 6541 lncRNA were upregulated and 2135 lncRNA were downregulated in 3-day ovaries cultured for 8 days in vitro compared with ovaries from day 3 mice. We also found that 4171 mRNAs were upregulated and 1795 were downregulated in the cultured ovaries. Gene ontology and pathway analyses showed that the functions of differentially expressed lncRNA targets and mRNAs were closely linked with many processes and pathways related to ovary development, including cell proliferation and differentiation, developmental processes, and other signaling transduction pathways. Additionally, many novel identified lncRNAs showed inducible expression, suggesting that these lncRNAs may be good candidates for investigating mouse primordial follicle activation. This study provides a foundation for further exploring lncRNA-related mechanisms in the initiation of mouse primordial follicles.

Effect of caloric restriction and rapamycin on ovarian aging in mice.

Caloric restriction (CR) increases the preservation of the ovarian primordial follicular reserve, which can potentially delay menopause. Rapamycin also increases preservation on the ovarian reserve, with similar mechanism to CR. Therefore, the aim of our study was to evaluate the effects of rapamycin and CR on metabolism, ovarian reserve, and gene expression in mice. Thirty-six female mice were allocated into three groups: control, rapamycin-treated (4mg/kg body weight every other day), and 30% CR. Caloric restricted females had lower body weight (P < 0.05) and increased insulin sensitivity (P = 0.003), while rapamycin injection did not change body weight (P > 0.05) and induced insulin resistance (P < 0.05). Both CR and rapamycin females displayed a higher number of primordial follicles (P = 0.02 and 0.04, respectively), fewer primary, secondary, and tertiary follicles (P < 0.05) and displayed increased ovarian Foxo3a gene expression (P < 0.05). Despite the divergent metabolic effects of the CR and rapamycin treatments, females from both groups displayed a similar increase in ovarian reserve, which was associated with higher expression of ovarian Foxo3a.

THE QUALITY OF SHEEPSKIN AND HISTOLOGICAL STRUCTURE OF DORPER SHEEPSKIN

The research explores the qualitative parameters of sheepskin and histological structure of Kalmyk lambs skin and cross breeds obtained when crossing Kalmyklambs and Dorper sheepskinned lambs. Scientific and production experience was conducted at “Agrofirma Aduciya” enterprise in the Republicof Kalmykiain 2018. The authors explored sheepskin commodity properties at the histological level was conducted on the lambs after fattening at the age of 6 months. According to the area of sheep skin, the crossbred lambs of the 2nd (experimental) group surpassed the lambs od the 1st (control) group by 11.08 dm2 (or 11.8%). This difference is explained by higher fattening qualities of the weighed young stock, which have shorter length and wider width of sheepskin. This is explained by the blackness of the animal’s body after fattening. The lambs of the control group had more developed epidermis, a reticular layer and a larger skin thickness, which is characteristic of rough-wool sheep breeds. The lambs of the experimental group have more developed pilar layer, which is typical for the cattle with semi-fine and thin woolen assortments. The density of wool follicles in the crossbred lambs was 3.3% higher in comparison with the control group lambs; the density index (the ratio of secondary to primary follicles) was 12.7% higher. The lambs of the control group have 44th quality wool (39.65 mkm), it has down, crossing hair, beard hair, i.e. rough wool; the crossing lambs of the experimental group have 56th quality wool (29.25 µm), i.e. semi-fine wool. The sheepskin products with softer pile will cost higher.

Effect of transcutaneous electrical acupoint stimulation on protecting against radiotherapy- induced ovarian damage in mice

BackgroundPremature ovarian insufficiency (POI) is characterized by early loss of ovarian function that affects women before the age of 40. We aim to explore the protective effects of transcutaneous electrical acupoint stimulation (TEAS) against irradiation-induced ovarian damage in mice.MethodsC57BL6 mice were randomly divided into control and irradiation (IR) groups. Then, control group was divided into two treatment subgroups: mock TEAS treatment (control-) and TEAS treatment (control+). IR group was divided into four subgroups according to the time of treatment started: mock TEAS treatment initiated at 2 days after irradiation (IR 2D-), TEAS treatment initiated at 2 days after irradiation (IR 2D+), mock TEAS treatment initiated at 1 week after irradiation (IR 1 W-), and TEAS treatment initiated at 1 week after irradiation (IR 1 W+). The radiation model mice were exposed to single whole body X-ray irradiation (4 Gy), and the control mice received 0 Gy. TEAS stimulation (2 Hz, 1 mA, 30 min/day) was given once a day for six consecutive days per week for 2 weeks. Estrous cycle, ovarian weight, serum AMH level and follicle counts were evaluated. Then, proliferation markers, apoptotic markers and oxidative stress markers were examined.ResultsCompared with the control group, the estrous cycle was disordered, and the ovarian weight, serum AMH, and primordial, primary and secondary follicles counts decreased (all P < 0.01) in the IR 2D- and IR 1 W- groups. In the irradiation with early TEAS treatment group (IR 2D+), the estrous cycle improved, the AMH level and primordial follicular significantly increased compared to the irradiation with mock group (IR 2D-). However, there were no significant differences in the estrous cycle, AMH level and follicle counts between IR 1 W- and IR 1 W+ groups. Moreover, IR 2D+ mice reduced the expression of Bax protein and increased the levels of Bcl-2 and PCNA compared to the IR 2D- group. Furthermore, the early TEAS treated mice showed significantly lower levels of oxidative stress and number of TUNEL (+) granulosa cells than that in the IR 2D- group.ConclusionThis study is first to evaluate TEAS as a potential therapy to attenuate irradiation-induced ovarian failure through inhibiting primordial follicles loss, increasing serum AMH secretion, inducing antioxidant, and anti-apoptotic systems.

Menstrual blood derived mesenchymal stem cells combined with Bushen Tiaochong recipe improved chemotherapy-induced premature ovarian failure in mice by inhibiting GADD45b expression in the cell cycle pathway

BackgroundTo investigate the therapeutic effects of menstrual blood derived mesenchymal stem cells (MB-MSCs) combined with Bushen Tiaochong recipe (BSTCR) on epirubicin induced premature ovarian failure (POF) in mice.MethodsTwenty-four female C57BL/6 mice of 6–8 weeks were intraperitoneally injected with epirubicin to induce POF, and then they were randomized into 4 groups of 6 mice each and treated with PBS, MB-MSCs, BSTCR, and MB-MSCs combined with BSTCR, respectively. Six mice of the same age were used as controls. Vaginal smear, TUNEL and hematoxylin-eosin staining were to observe estrous cycles, ovarian cell apoptosis and follicles. Enzyme-linked immunosorbent analysis determined serum estradiol, follicle-stimulating hormone (FSH) and anti-Müllerian hormone (AMH) levels. RT-qPCR and Western Blot analysis were to determine GADD45b, CyclinB1, CDC2 and pCDC2 expressions.ResultsEpirubicin treatment resulted in a decrease in the number of primordial, primary, secondary and antral follicles, an increase in the number of atretic follicles and ovarian cell apoptosis, a decrease in estradiol and AMH levels, an increase in FSH levels, and estrous cycle arrest. However, MB-MSCs combined with BSTCR rescued epirubicin induced POF through down-regulating GADD45b and pCDC2 expressions, and up-regulating CyclinB1 and CDC2 expressions. The combined treatment showed better therapeutic efficacy than BSTCR or MB-MSCs alone.ConclusionsMB-MSCs combined with BSTCR improved the ovarian function of epirubicin induced POF mice, which might be related to the inhibition of GADD45b expression and the promotion of CyclinB1 and CDC2 expressions. The combined treatment had better therapeutic efficacy than BSTCR or MB-MSCs alone.

Hedgehog pathway inhibition causes primary follicle atresia and decreases female germline stem cell proliferation capacity or stemness

BackgroundFollicle depletion is one of the causes of premature ovarian failure (POF) and primary ovarian insufficiency (POI). Hence, maintenance of a certain number of female germline stem cells (FGSCs) is optimal to produce oocytes and replenish the primordial follicle pool. The mechanism that regulates proliferation or stemness of FGSCs could contribute to restoring ovarian function, but it remains uncharacterized in postnatal mammalian ovaries. This study aims to investigate the mechanism by which inhibiting the activity of the hedgehog (Hh) signaling pathway regulates follicle development and FGSC proliferation.Methods and resultsTo understand the role of the Hh pathway in ovarian aging, we measured Hh signaling activity at different reproductive ages and the correlation between them in physiological and pathological mice. Furthermore, we evaluated the follicle number and development and the changes in FGSC proliferation or stemness after blocking the Hh pathway in vitro and in vivo. In addition, we aimed to explain one of the mechanisms for the FGSC phenotype changes induced by treatment with the Hh pathway-specific inhibitor GANT61 via oxidative stress and apoptosis. The results show that the activity of Hh signaling is decreased in the ovaries in physiological aging and POF models, which is consistent with the trend of expression levels of the germline stem cell markers Mvh and Oct4. In vitro, blocking the Hh pathway causes follicular developmental disorders and depletes ovarian germ cells and FGSCs after treating ovaries with GANT61. The proliferation or stemness of cultured primary FGSCs is reduced when Hh activity is blocked. Our results show that the antioxidative enzyme level and the ratio of Bcl-2/Bax decrease, the expression level of caspase 3 increases, the mitochondrial membrane potential is abnormal, and ROS accumulate in this system.ConclusionsWe observed that the inhibition of the Hh signaling pathway with GANT61 could reduce primordial follicle number and decrease FGSC reproductive capacity or stemness through oxidative damage and apoptosis.

Ovarian follicle resilience in mice orally dosed with methoxychlor: Are reproductive impacts possible in mammals as ecological receptors?

Abstract Mammals are routinely evaluated with chemical exposure modeling in ecological risk assessments for contaminated terrestrial properties, such as sites managed under the U.S. EPA’s Superfund program. Despite modeling outcomes frequently indicating that site mammals are experiencing severe reproductive impacts, there are virtually no known instances of site mammal populations being either unhealthful or reduced in size. This observation has triggered the development of the one-time patented and recently ASTM International-certified Rodent Sperm Analysis method that directly assesses male reproductive health, and thereby the overall health of mammals that occur at sites. Just as experimentation has led to an understanding of the degree of sperm count reduction, sperm motility reduction, and increase in morphologically altered sperm that need occur for reproduction to be impacted, the degree of ovarian follicle count reduction to similarly impact reproductive success should be able to be determined. Thus, adult female CD-1 mice were orally exposed to vehicle control or methoxychlor at 32 and 64 mg/kg/day for 30 days. Pursuant to resting periods of 30 and 60 days post-dosing, half of the treated animals were euthanized and their ovaries were collected and subjected to histological evaluations of follicle numbers and health. The other half of the animals were mated to proven breeder males to enable evaluation of breeding success. At 30 days post-dosing, primary and pre-antral follicle numbers were increased at both methoxychlor doses compared to controls. Further, the number of atretic follicles nearly doubled at both methoxychlor doses compared to controls. In contrast, at 60 days post-dosing, follicle numbers did not differ between methoxychlor treatment and control groups. These results were corroborated by 60-day mated and non-mated mice displaying an increase in serum anti-Mullerian hormone level, a recognized marker for the quantitative aspect of ovarian reserve. At both 30 day and 60 days post-dosing, pup weights and litter sizes in methoxychlor exposed animals were the same as those of controls. Collectively, these data indicate that oral exposure to methoxychlor did not increase follicle atresia or reduce follicle numbers enough to compromise female fertility. These outcomes may be indicators of ovarian follicle resiliency to chemical exposure, a capability that could benefit mammals in the wild. Ovarian follicle resiliency could, in part, explain the recurrent discovery at contaminated terrestrial sites, of rodent populations that are identical (in terms of size, sex ratio, and age distribution) to those of nearby (non-contaminated) reference locations.

Dose optimisation of PTEN inhibitor, bpV (HOpic), and SCF for the in-vitro activation of sheep primordial follicles

The in-vitro development of primordial follicles is critical for improving mammalian fertility and wildlife conservation. This study aimed to optimise the effective doses of bpV (HOpic) and stem cell factor (SCF) for the in-vitro activation of sheep primordial follicles. To do this, sheep ovarian cortex was treated with bpV (1.5, 15, and 150 μM) and SCF (50 and 100 ng/ml). Follicular count indicated that 15 μM bpV and 100 ng/ml SCF significantly increased normal primary follicles compared to other groups (p < 0.05). Also, a significant downregulation of P53 and PTEN, as well as the increased expression of PI3K was observed. The in-vitro maturation was more pronounced when the fragmented tissues were co-treated with selected doses of bpV and SCF. In conclusion, the combination of 15 μM bpV and 100 ng/ml SCF was the most effective treatment strategy for the activation and survival of primordial follicles in sheep ovarian fragments.

Divergent immunohistochemical expression of CD21 and CD23 by follicular dendritic cells with increasing grade of follicular lymphoma

BackgroundUltrastructural and immunohistochemical differences have been described in FDCs of primary and secondary follicles, illustrating the highly compartmentalized structure of lymph follicles. Differences in FDC immunophenotype in different grades of FL may reflect some parallelism between reactive and neoplastic conditions in terms of FDC-B cell interaction and may be used as a valuable additional tool for grading FL.MethodsA total of 60 paraffin blocks from patients with follicular lymphoma, 30 cases each of grade 1 and 3, were retrieved from our archive. Immunohistochemical analysis was carried out for CD21, CD23, cyclin A, and Ki-67.ResultsOur study demonstrates that during evaluation, six patterns of FDC distribution were distinguished. The intensity of stain for CD21 was not statistically significant in grade 1 and grade 3 FL (p = 0.340). In contrast, grade 3 FLs exhibited a significant decrease of CD23 expression by the FDCs (p < 0.001). By CD21 stain, there was no significant difference in the distribution of pattern 1 in grades 1 and 3 (p = 0.098). In contrast, in grade 3, this pattern was significantly less observed by CD23 stain (p = 0.016). The same was observed for pattern 2 for CD21 (p = 0.940) and CD23 (p = 0.010) and pattern 4 for CD21 (p = 0.305) and CD23 (p = 0.005), respectively. Distribution of pattern 5 was significantly different between grades 1 and 3 both for CD21 (p = 0.005) and CD23 (p < 0.001). Distribution of patterns 2 and 6 was not significantly different between grades 1 and 3 for CD21 and CD23. The values of cyclin A and Mib-1 were also significantly different between grades 1 and 3 (p < 0.001).ConclusionsThe observed patterns of FDCs lead us to believe that similar to reactive lymph node follicles, neoplastic follicles in FL, at least in early stages, have an organized structure. Hypothetically, with CD21, CD23, and cyclin A immunohistochemistry, the sequence of events in FL progression may be traced.

CRISPR/Cas9-mediated VDR knockout plays an essential role in the growth of dermal papilla cells through enhanced relative genes.

BackgroundHair follicles in cashmere goats are divided into primary and secondary hair follicles (HFs). HF development, which determines the morphological structure, is regulated by a large number of vital genes; however, the key functional genes and their interaction networks are still unclear. Although the vitamin D receptor (VDR) is related to cashmere goat HF formation, its precise effects are largely unknown. In the present study, we verified the functions of key genes identified in previous studies using hair dermal papilla (DP) cells as an experimental model. Furthermore, we used CRISPR/Cas9 technology to modify the VDR in DP cells to dissect the molecular mechanism underlying HF formation in cashmere goats.ResultsThe VDR expression levels in nine tissues of Shaanbei white cashmere goats differed significantly between embryonic day 60 (E60) and embryonic day 120 (E120). At E120, VDR expression was highest in the skin. At the newborn and E120 stages, the VDR protein was highly expressed in the root sheath and hair ball region of Shaanbei cashmere goats. We cloned the complete CDS of VDR in the Shaanbei white cashmere goat and constructed a VDR-deficient DP cell model by CRISPR/Cas9. Heterozygous and homozygous mutant DP cells were produced. The growth rate of mutant DP cells was significantly lower than that of wild-type DP cells (P < 0.05) and VDR mRNA levels in DP cells decreased significantly after VDR knockdown (P < 0.05). Further, the expression levels of VGF, Noggin, Lef1, and β-catenin were significantly downregulated (P < 0.05).ConclusionsOur results indicated that VDR has a vital role in DP cells, and that its effects are mediated by Wnt and BMP4 signaling.

Comparative histological study of skin in Jenubi and its crossbreed cow

The aim of this study was to comparative the histological structure of skin in Jenubi cow and its Friesians crossbreed cow. Samples of skin were collected from abdominal regions of Jenubi cow and its Friesians crossbreed cow immediately after slaughter in Governorate of Misan. The results showed similarities in the structure of the normal skin layers which was consists of three layers epidermis, dermis and hypodermis. Sweet glands was more distributed in the region between papillary and reticular layer and are associated with hair follicles. The primary hair follicle was associated with Sebaceous glands which were scattered in the deeper part of the reticular layer in both Jenubi and cross breed skin. The deference’s found in the micrometrical observations in the dermal layer, these variations between breeds in epithelium thickness of sweet glands where small in Jenubi cow while the epithelium thickness of crossbreed cow was more greater. The present study it was revealed that the sweat gland diameter in the skin of Jenubi cow was lower than in crossbreed cow. The conclusion of this study revealed that the location of the papillary layer is superficial and just under the basement membrane while the reticular layer composed the deeper layer in the dermis. The type of sebaceous gland was a simple, branched or compound. The epithelium thickness of sweet glands in crossbreed cow was more greater than Jenubi cow.

Ulmus minor bark hydro-alcoholic extract ameliorates histological parameters and testosterone level in an experimental model of PCOS rats.

Polycystic ovary syndrome (PCOS) is a common and multifactorial disease associated with female factor infertility. Ulmus minor bark (UMB) is one of the medicinal plants used in Persian folklore as a fertility enhancer. In the current study, we aimed to elucidate the effect of UMB hydro-alcoholic extract on histological parameters and testosterone condition in an experimental model of PCOS rats. Thirty female rats were randomly divided into five groups: (1) control, (2) vehicle, (3) PCOS/50 mg [6 mg/kg dehydroepiandrosterone (DHEA) + 50 mg/kg UMB hydro-alcoholic extract], (4) PCOS/150 mg (6 mg/kg DHEA + 150 mg/kg UMB hydro-alcoholic extract), and (5) PCOS (6 mg/kg DHEA). All interventions were performed for 21 days. Afterwards, stereological analysis was done for determination of ovarian volume and follicle number. The serum level of testosterone was measured by ELISA kit. UMB hydro-alcoholic extract improved the total number of the corpus luteum in the treatment groups when compared to the PCOS group (p<0.05). PCOS/150 mg and PCOS/50 mg groups showed significantly lower total number of the primordial, primary, and secondary follicles as well as testosterone level compared to the PCOS group (p<0.05). The total number of antral follicles and volume of ovary did not differ significantly between groups. UMB extract may be an effective and good alternative in improving PCOS histo-logical and testosterone disturbances although further studies are warranted to confirm the safety of UMB plant in human.

Distinctive Genes Involved in Steroidogenesis Associated with Follicular Abnormal Development in Polycystic Ovary Syndrome Model

Objective: Polycystic ovary syndrome (PCOS), a heterogeneous endocrine disorder, affects female reproductive function, but its etiology has not been elucidated. In this study, we analyzed the differential genes related to ovarian steroid biosynthesis in patients with PCOS, to explore the mechanism of PCOS. Methods: The GSE59456 data were downloaded from the Gene Expression Omnibus database. We identified differentially expressed genes (DEGs) in ovaries between the female Sprague–Dawley rats those implanted with 5α-dehydrotestestrone (DHT) and those in control (CTL, implanted with empty capsule). Gene ontology, pathway enrichment analysis, and protein–protein interaction (PPI) network construction were subsequently performed. Results: In total, 530 upregulated DEGs and 522 downregulated DEGs were identified. The identified DEGs were mostly associated with steroid biosynthesis. In the PPI network, the module M1 was mainly related to steroid biosynthesis, and five genes (Hsd17b7, Tm7sf2, Idi1, Msmo1, and Sqle) of the module M1 were from the aforementioned group of upregulated genes. Furthermore, the 19 DEGs (Idi1, Cga, C1qb, Thy1, Gpx1, Ctss, Lpl, A2m, Cited2, Plppr4, Prkar2b, Slc44a1, Inha, Rbp4, Pla2g2a, Gata4, Fabp3, Cpa2, and Cpa1) between DHT and CTL groups were associated with the process of the transformation of primordial follicles to primary follicles. Conclusions: These DEGs, such as Hsd17b7, Tm7sf2, Idi1, Msmo1, Sqle, Rbp4, Gata4, Inha, and Cited2, may be used to elucidate the etiology of PCOS, which may provide new insights into the exploration of pathological mechanism and biomarkers for polycystic ovary.

Analysis of Expression and Functional Activity of Aromatic L-Amino Acid Decarboxylase (DDC) and Serotonin Transporter (SERT) as Potential Sources of Serotonin in Mouse Ovary.

The origin of serotonin in the ovary is the key question for understanding mechanisms of serotonergic regulation of reproductive function. We performed a study of the expression and functional activity of the serotonin transporter (SERT) and the enzyme for the synthesis of serotonin, aromatic l-amino acid decarboxylase (DDC) in mouse ovary. A pronounced peak of SERT mRNA expression occurs at the age of 14 days, but serotonin synthesis enzymes are expressed at the maximum level in the ovaries of newborn mice. SERT is detected immunohistochemically in all cellular compartments of the ovary with a maximum level of immunostaining in the oocytes of growing ovarian follicles. DDC immunolocalization, in contrast, is detected to a greater extent in primordial follicle oocytes, and decreases at the later stages of folliculogenesis. Serotonin synthesis in all cellular compartments occurs at very low levels, whereas specific serotonin uptake is clearly present, leading to a significant increase in serotonin content in the oocytes of growing primary and secondary follicles. These data indicate that the main mechanism of serotonin accumulation in mouse ovary is its uptake by the specific SERT membrane transporter, which is active in the oocytes of the growing ovarian follicles.

Metformin prevents murine ovarian aging.

A number of studies have shown that metformin can delay aging process and extend healthy lifespan in animals. However, its role in female reproductive lifespan is unclear. This study was aimed to explore the potential anti-aging effect of metformin on the ovary and its possible mechanisms. Female C57BL/6 mice of 27-week old were divided into two groups, the control group (CON) and metformin-treated group (MET). CON mice were fed ad libitum, while MET mice were fed on chows supplied with 100mg/kg metformin for half a year. Ovarian reserve and function were assessed by ovarian follicle counts, estrous cycle and sex hormones levels. The expressions of oxidized metabolites, such as 8-hydroxy-2´-deoxyguanosine (8-OHdG), 4-hydroxynonenal (4-HNE), nitrotyrosine (NTY), and ovarian aging associated proteins P16, SIRT1, p-rpS6 and Bcl2 were examined. The MET mice exhibited increased level of serum E2 hormone and higher percentage of regular estrous cycles after 6 months' feeding, compared to the CON mice. The amount of primordial and primary follicles and the expression of SIRT1 were significantly increased, but the levels of P16, 8-OHdG, 4-HNE and p-rpS6 were decreased in the MET mice. These results indicate that metformin can delay ovarian aging process, probably by inducing the expression of SIRT1 and reducing the oxidative damage.

Noise stress on female mice impair oocyte developmental potential.

Objectives: To find out the effects of noise stress on variations in number of ovarian follicles in mice. Study Design: Randomized control trial, laboratory based. Place and Duration of Study: Anatomy department, Army Medical College Rawalpindi, it was conducted from April to May 2014. Material and Method: 20 adult mice (BALBc strain) female weighing 25-27 grams were taken for this study on day first of their estrous cycle. Two groups comprising of 10 rats each were made. Control group was given name tag of group A and was kept in animal house under normal heathy environment for one month. Noise stress of 100 decibels for 06 hours per day for one month was given to experimental group B. Mice were dissected after 1 month. Mice right ovaries were examined for size, shape, color and weight. The specimen were processed and embedded. Hematoxyline and Eosin stained slides were prepared for histological study. Follicular count (primary, secondary and tertiary follicles) was done and noted. For statistical analysis SPSS 20 was used. The tests applied was independent sample’s t test for descriptive statistics. Results: There was a significant decrease in primary, secondary/antral and graafian follicular count in the ovary of Group B (Experimental). Conclusion: The noise stress prominently affected the follicular count in ovaries of noise stress treated group.

Influence of mouse defective zona pellucida in folliculogenesis on apoptosis of granulosa cells and developmental competence of oocytes†.

Zona pellucida (ZP), which enwraps the oocyte during folliculogenesis, initially forms in the primary follicle and plays an important role in female fertility. Here, we investigated a mouse strain ("mutant mice" for short) carrying two types of ZP defects in folliculogenesis, i.e., ZP thinned (but intact) and ZP cracked, caused by targeted mutation in the Zp1 gene. Using this mutant mouse strain and wild-type mouse as control, we studied the effects of the ZP defects on the development of oocytes and granulosa cells during folliculogenesis. For each ZP defect, we examined the morphology of transzonal projections and apoptosis of granulosa cells in the corresponding growing follicles, as well as the morphology of corresponding ovulated eggs and their abilities to develop into viable individuals. Our results suggested that ZP integrity rather than thickness or porosity is crucial for preventing the ectopia of granulosa cells, maintaining adequate routine bilateral signaling between oocyte and surrounding granulosa cells, and thus for ensuring the survival of granulosa cells and the establishment of the full developmental competence of oocytes. This is the first study to elucidate the effects of different degrees of ZP defects caused by the same gene mutation, on the apoptosis of granulosa cells and developmental competence of oocytes, and to explore the potential mechanisms underlying these effects.

Meiotic Chromosomes, Synaptonemal Complexes in a Female Viviparous Lizard (Zootoca vivipara) in Prophase I of Meiosis

In the females of the viviparous lizard Zootoca vivipara (Lichtenstein, 1823) (family Lacertidae) from Northwest Russia (2n = 35: 32A (acrocentric autosomes) + Z1Z2W sex chromosomes), the ovarian lumen germinal vesicles (oocytes), as well as germinal lamina cells, were examined. Chromosome preparations were obtained using the direct method and the method of total oocyte nuclei spreading developed by Dresser and Moses. Chromosome preparations were stained with Giemsa; for visualization of synaptonemal complexes (SCs), total preparations of oocyte nuclei were stained with silver nitrate and DAPI. It was demonstrated that, during oogenesis in the female, primary follicles enter the early stages of the meiotic prophase I (stages from leptotene to diplotene, lampbrush chromosomes are formed). Here, for the first time, total oocyte spreads were obtained and studied. On the basis of light microscopic analysis of the oocyte SCs and taking into account their length in a female with 2n = 35, the SC karyotype is presented, consisting of 19 SC elements, among which 16 SC autosomal bivalents are distinguished. The remaining three SC elements, according to the authors, can be univalents of Z1Z2W sex chromosomes or one WZ1 bivalent and Z2 and B chromosome univalents. As in the SC karyotype of Z. vivipara males, in a female, specific features in the morphology of SC elements of chromosomes 5 and 6 were observed.