Differential genome-wide gene expression profiling of bovine largest and second-largest follicles: identification of genes associated with growth of dominant follicles

<p>Abstract</p> <p>Background</p> <p>Bovine follicular development is regulated by numerous molecular mechanisms and biological pathways. In this study, we tried to identify differentially expressed genes between largest (F1) and second-largest follicles (F2), and classify them by global gene expression profiling using a combination of microarray and quantitative real-time PCR (QPCR) analysis. The follicular status of F1 and F2 were further evaluated in terms of healthy and atretic conditions by investigating mRNA localization of identified genes.</p> <p>Methods</p> <p>Global gene expression profiles of F1 (10.7 +/- 0.7 mm) and F2 (7.8 +/- 0.2 mm) were analyzed by hierarchical cluster analysis and expression profiles of 16 representative genes were confirmed by QPCR analysis. In addition, localization of six identified transcripts was investigated in healthy and atretic follicles using in situ hybridization. The healthy or atretic condition of examined follicles was classified by progesterone and estradiol concentrations in follicular fluid.</p> <p>Results</p> <p>Hierarchical cluster analysis of microarray data classified the follicles into two clusters. Cluster A was composed of only F2 and was characterized by high expression of 31 genes including IGFBP5, whereas cluster B contained only F1 and predominantly expressed 45 genes including CYP19 and FSHR. QPCR analysis confirmed AMH, CYP19, FSHR, GPX3, PlGF, PLA2G1B, SCD and TRB2 were greater in F1 than F2, while CCL2, GADD45A, IGFBP5, PLAUR, SELP, SPP1, TIMP1 and TSP2 were greater in F2 than in F1. In situ hybridization showed that AMH and CYP19 were detected in granulosa cells (GC) of healthy as well as atretic follicles. PlGF was localized in GC and in the theca layer (TL) of healthy follicles. IGFBP5 was detected in both GC and TL of atretic follicles. GADD45A and TSP2 were localized in both GC and TL of atretic follicles, whereas healthy follicles expressed them only in GC.</p> <p>Conclusion</p> <p>We demonstrated that global gene expression profiling of F1 and F2 clearly reflected a difference in their follicular status. Expression of stage-specific genes in follicles may be closely associated with their growth or atresia. Several genes identified in this study will provide intriguing candidates for the determination of follicular growth.</p

Quantitative analysis of bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9) gene expression in calf and adult bovine ovaries

<p>Abstract</p> <p>Background</p> <p>It has been reported that calf oocytes are less developmentally competent than oocytes obtained from adult cows. Bone morphogenetic protein 15 (BMP15) and growth and differentiation factor 9 (GDF9) play critical roles in folliculogenesis, follicular development and ovulation in mammalian ovaries. In the present study, we attempted to compare the expression patterns of <it>BMP15 </it>and <it>GDF9 </it>in the cells of calf and cow ovaries to determine a relationship between the level of these genes and the low developmental competence of calf oocytes.</p> <p>Methods</p> <p>Bovine tissues were collected from 9-11 months-old calves and from 4-6 years-old cows. We characterized the gene expression of BMP15 and GDF9 in calf and adult bovine oocytes and cumulus cells using quantitative real-time reverse transcriptase polymerase chain reaction (QPCR) and <it>in situ </it>hybridization. Immunohistochemical analysis was also performed.</p> <p>Results</p> <p>The expression of <it>BMP15 </it>and <it>GDF9 </it>in cumulus cells of adult ovaries was significantly higher than that in calf ovaries, as revealed by QPCR. <it>GDF9 </it>expression in the oocytes of calf ovaries was significantly higher than in those of the adult ovaries. In contrast, <it>BMP15 </it>expression in the oocytes of calf and adult ovaries was not significantly different. The localization of gene expression and protein were ascertained by histochemistry.</p> <p>Conclusions</p> <p>Our result showed for the first time BMP15 and GDF9 expression in bovine cumulus cells. <it>BMP15 </it>and <it>GDF9 </it>mRNA expression in oocytes and cumulus cells was different in calves and cows.</p

Hitomi (ASTRO-H) X-ray Astronomy Satellite

The Hitomi (ASTRO-H) mission is the sixth Japanese x-ray astronomy satellite developed by a large international collaboration, including Japan, USA, Canada, and Europe. The mission aimed to provide the highest energy resolution ever achieved at E  >  2  keV, using a microcalorimeter instrument, and to cover a wide energy range spanning four decades in energy from soft x-rays to gamma rays. After a successful launch on February 17, 2016, the spacecraft lost its function on March 26, 2016, but the commissioning phase for about a month provided valuable information on the onboard instruments and the spacecraft system, including astrophysical results obtained from first light observations. The paper describes the Hitomi (ASTRO-H) mission, its capabilities, the initial operation, and the instruments/spacecraft performances confirmed during the commissioning operations for about a month

Differential gene expression of serine protease inhibitors in bovine ovarian follicle: possible involvement in follicular growth and atresia

Abstract Background SERPINs (serine protease inhibitors) regulate proteases involving fibrinolysis, coagulation, inflammation, cell mobility, cellular differentiation and apoptosis. This study aimed to investigate differentially expressed genes of members of the SERPIN superfamily between healthy and atretic follicles using a combination of microarray and quantitative real-time PCR (QPCR) analysis. In addition, we further determined mRNA and protein localization of identified SERPINs in estradiol (E2)-active and E2-inactive follicles by in situ hybridization and immunohistochemistry. Methods We performed microarray analysis of healthy (10.7 +/- 0.7 mm) and atretic (7.8 +/- 0.2 mm) follicles using a custom-made bovine oligonucleotide microarray to screen differentially expressed genes encoding SERPIN superfamily members between groups. The expression profiles of six identified SERPIN genes were further confirmed by QPCR analysis. In addition, mRNA and protein localization of four SERPINs was investigated in E2-active and E2-inactive follicles using in situ hybridization and immunohistochemistry. Results We have identified 11 SERPIN genes expressed in healthy and atretic follicles by microarray analysis. QPCR analysis confirmed that mRNA expression of four SERPINs (SERPINA5, SERPINB6, SERPINE2 and SERPINF2) was greater in healthy than in atretic follicles, while two SERPINs (SERPINE1 and SERPING1) had greater expression in atretic than in healthy follicles. In situ hybridization showed that SERPINA5, SERPINB6 and SERPINF2 mRNA were localized in GCs of E2-active follicles and weakly expressed in GCs of E2-inactive follicles. SERPING1 mRNA was localized in both GCs and the theca layer (TL) of E2-inactive follicles and a weak hybridization signal was also detected in both GCs and TL of E2-active follicles. Immunohistochemistry showed that SERPINA5, SERPINB6 and SERPINF2 were detected in GCs of E2-active and E2-inactive follicles. SERPING1 protein was localized in both GCs and the TL of E2-active and E2-inactive follicles. Conclusions Our results demonstrate a characteristic expression of SERPIN superfamily member genes in bovine healthy and atretic follicles. The cell-type-and stage-specific expression of SERPINs may be associated with bovine follicular growth and atresia.</p