MicroRNA indicators of follicular steroidogenesis.

MicroRNAs (miRNAs) can provide useful biomarkers of tissue function. The aim of the present study was to determine, in bovine follicles (n = 66; diameter 4-22 mm), the relationship among several indices of steroidogenesis and levels of 15 miRNAs previously identified to be associated with follicle development. Oestradiol levels, the oestradiol : progesterone (E : P) ratio and cytochrome P450 family 19 subfamily A member 1 (CYP19A1) expression were strongly correlated with each other (ρ > 0.8) and with LH/choriogonadotropin receptor (LHCGR) expression (ρ ≥ 0.6; P < 0.01). Levels of nine different miRNAs in the follicular wall were correlated (P < 0.01) with oestradiol, the E : P ratio and CYP19A1, with miR-873 showing the strongest correlation in each case (ρ > 0.7). Analyses of follicular fluid miRNAs identified miR-202 as correlated with oestradiol, the E : P ratio and CYP19A1 (ρ > 0.5; P < 0.01). When considering all follicle end-points together, we found that using a cut-off value of E : P = 1 overestimated the number of oestrogen-inactive follicles, whereas using CYP19A1 as a classifier provided a clearer separation of follicle samples based on oestrogen activity, in agreement with the E : P ratio, LHCGR expression and levels of miR-873 and miR-202. In conclusion, we identified miR-873 and miR-202 as miRNAs whose levels in follicular tissues can be used as indicators of steroidogenic capacity in bovine. We showed that these or other gene expression parameters, in addition or alternatively to the E : P ratio, should be used to accurately classify follicles based on steroidogenic capacity.

Involvement of miRNAs in equine follicle development.

Previous evidence from in vitro studies suggests specific roles for a subset of miRNAs, including miR-21, miR-23a, miR-145, miR-503, miR-224, miR-383, miR-378, miR-132, and miR-212, in regulating ovarian follicle development. The objective of this study was to determine changes in the levels of these miRNAs in relation to follicle selection, maturation, and ovulation in the monovular equine ovary. In Experiment 1, follicular fluid was aspirated during ovulatory cycles from the dominant (DO) and largest subordinate (S) follicles of an ovulatory wave and the dominant (DA) follicle of a mid-cycle anovulatory wave (n=6 mares). Follicular fluid levels of progesterone and estradiol were lower (P<0.01) in S follicles than in DO follicles, whereas mean levels of IGF1 were lower (P<0.01) in S and DA follicles than in DO follicles. Relative to DO and DA follicles, S follicles had higher (P≤0.01) follicular fluid levels of miR-145 and miR-378. In Experiment 2, follicular fluid and granulosa cells were aspirated from dominant follicles before (DO) and 24 h after (L) administration of an ovulatory dose of hCG (n=5 mares/group). Relative to DO follicles, L follicles had higher follicular fluid levels of progesterone (P=0.05) and lower granulosa cell levels of CYP19A1 and LHCGR (P<0.005). Levels of miR-21, miR-132, miR-212, and miR-224 were increased (P<0.05) in L follicles; this was associated with reduced expression of the putative miRNA targets, PTEN, RASA1, and SMAD4. These novel results may indicate a physiological involvement of miR-21, miR-145, miR-224, miR-378, miR-132, and miR-212 in the regulation of cell survival, steroidogenesis, and differentiation during follicle selection and ovulation in the monovular ovary.

Involvement of miRNAs in ovarian follicular and luteal development.

Although much progress has been made in the genetic dissection of biological networks involved in follicular/luteal development in the mammalian ovary, the gene regulation mechanisms involved are still poorly understood. Over the last 10 years, miRNAs have emerged as master regulators of tissue growth and differentiation in animals. However, compared with other body tissues, little is still known about the functional involvement of miRNAs in the ovary. Several studies have identified miRNA populations specifically associated with the development of follicles and corpora lutea, particularly in relation to the follicular-luteal transition, and the functional involvement of some of these miRNAs has been characterised in vitro and/or in vivo. Specifically, three different miRNAs, miR-224, miR-378 and miR-383, have shown to be involved in regulating aromatase expression during follicle development. In addition, miR-21 has been identified as promoting follicular cell survival during ovulation, and pro-angiogenic miR-17-5p and let-7b were shown to be necessary for normal development of the corpus luteum. Experimental evidence for the involvement of several other miRNAs in different aspects of follicle/luteal development has also been obtained. In addition, many of these studies exemplify the challenges associated with identifying physiologically relevant targets of ovarian miRNAs. Continuous advances in this field will be considerably facilitated by progress in understanding miRNA physiology in other body systems and will eventually lead to a much better understanding of the control of follicular/luteal development. In turn, through the potential offered by miRNA diagnostics and miRNA therapeutics, this new knowledge should bring considerable benefits to reproductive medicine.

The 34,XY1,der(13) chromosome constitution with loss of Y2 is associated with unilateral testicular hypoplasia in the endangered Indian blackbuck antelope (Antilope cervicapra).

The present study is the first report of unilateral testicular hypoplasia in 3 of 15 (20%) Indian blackbuck antelopes (Antilope cervicapra). Interestingly, the condition was restricted to only the right testis in all cases. Cytogenetic analysis revealed chromosomal aneuploidy in the affected individuals which had a 34,XY(1),der(13) karyotype with loss of the acrocentric (autosomal) Y(2) and an aberrant chromosome 13. We further determined that the semen output and the circulating testosterone levels were markedly low in the males with hypoplastic testes as compared to fertile males.

Identification of miRNAs associated with the follicular-luteal transition in the ruminant ovary.

Little is known about the involvement of microRNAs (miRNAs) in the follicular-luteal transition. The aim of this study was to identify genome-wide changes in miRNAs associated with follicular differentiation in sheep. miRNA libraries were produced from samples collected at defined stages of the ovine oestrous cycle and representing healthy growing follicles, (diameter, 4.0-5.5  mm), pre-ovulatory follicles (6.0-7.0  mm), early corpora lutea (day 3 post-oestrus) and late corpora lutea (day 9). A total of 189 miRNAs reported in sheep or other species and an additional 23 novel miRNAs were identified by sequencing these libraries. miR-21, miR-125b, let-7a and let-7b were the most abundant miRNAs overall, accounting for 40% of all miRNAs sequenced. Examination of changes in cloning frequencies across development identified nine different miRNAs whose expression decreased in association with the follicular-luteal transition and eight miRNAs whose expression increased during this transition. Expression profiles were confirmed by northern analyses, and experimentally validated targets were identified using miRTarBase. A majority of the 29 targets identified represented genes known to be actively involved in regulating follicular differentiation in vivo. Finally, luteinisation of follicular cells in vitro resulted in changes in miRNA levels that were consistent with those identified in vivo, and these changes were temporally associated with changes in the levels of putative miRNA targets in granulosa cells. In conclusion, this is the first study to characterise genome-wide miRNA profiles during different stages of follicle and luteal development. Our data identify a subset of miRNAs that are potentially important regulators of the follicular-luteal transition.

Evaluation of awareness about pharmacovigilance and adverse drug reaction monitoring in resident doctors of a tertiary care teaching hospital.

OBJECTIVE: Adverse drug reactions (ADRs) are associated with significant morbidity and mortality and have a major impact on public health. Pharmacovigilance helps in early detection of ADRs and identification of risk factors. Underreporting of ADRs can be improved by imparting knowledge regarding pharmacovigilance to healthcare professionals. This study was aimed at investigating the knowledge and attitude of resident doctors about ADR reporting and suggesting possible ways of improving ADR reporting. MATERIALS AND METHODS: This study was a cross-sectional, questionnaire-based survey conducted in a tertiary care teaching hospital. The respondents were resident doctors. Study instrument was a self-developed, pre-validated, semi-structured questionnaire consisting of open- and close-ended items. RESULTS: A total of 84 questionnaires were considered for analysis, giving a response rate of 93.33%. In all, 64.28% of the respondents were aware about pharmacovigilance, 52.38% were aware of ADR reporting system in India, 83.33% opined that only serious ADR with any medicine should be reported, and 35.72% believed that ADRs should be reported only for newly marketed agents. Although 67.85% of respondents observed an ADR, only 25% reported it; 44.04% were aware about the complete procedure of ADR reporting. General attitude of the respondents about ADR reporting was as follows: ADR reporting should be compulsory (15.19%), voluntary (41.66%), remunerated (3.57%), identity of prescriber should be concealed (21.42%), and identity of reporter should be concealed (29.7%). CONCLUSION: Increasing awareness about pharmacovigilance will be helpful in improving the status of ADR reporting. Other measures such as making ADR reporting guidelines available in the form of booklets and displaying posters can also play a useful role.

Immunomodulatory effect of Tinospora cordifolia extract in human immuno-deficiency virus positive patients.

OBJECTIVES: To assess the safety and efficacy of TCE in human immuno-deficiency virus positive patients. MATERIALS AND METHODS: Efficacy of Tinospora cordifolia extract (TCE) in HIV positive patients was assessed in randomized double blind placebo controlled trial. 68 HIV positive participants were randomly assigned to two groups to receive either TCE or placebo for six months. After clinical examination TLC, DLC, ESR, platelet count, hemoglobin and CD4 count were done. The hematological investigations were repeated at bimonthly intervals and CD4 count was repeated at the end of the study. Patients were clinically reviewed at monthly intervals for compliance, refill and ADR monitoring. The drugs were decoded at the end of the trial. RESULTS: TCE treatment caused significant reduction in eosinophil count and hemoglobin percentage. 60% patients receiving TCE and 20% on placebo reported decrease in the incidence of various symptoms associated with disease. Some of the common complaints reported by patients on TCE were anorexia, nausea, vomiting and weakness. CONCLUSION: Tinospora cordifolia extract, a plant derived immunostimulant, significantly affected the symptoms of HIV. This was validated by clinical evaluation. However not all of the objective parameters studied by us, back this up. Tinospora cordifolia could be used as an adjunct to HIV/AIDS management.

Differential erythrocyte agglutination pattern in normal and cancer patients with Synadenium grantii root (Hook f) lectin.

In the present study the property of lectin agglutination in blood on normal and different cancer patients has been observed. The purifiedSynadenium grantii root lectin was non blood group specific and its utility as a diagnostic tool in malignancy was studied. Hemagglutination (units/ml) of red blood cells of different types of cancer was compared with the normal control's red blood cells. Out of 113 total cancer patients, only a group of 29 breast cancer patients showed significant increase in titre value (P<0.05) compared to normal control.