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Gene expression profiling of preovulatory follicle in the buffalo cow: effects of increased IGF-I concentration on periovulatory events.

Rao JU, Shah KB, Puttaiah J, Rudraiah M - PLoS ONE (2011)

Bottom Line: Thus, further experiments were conducted to verify the effects of increased intrafollicular IGF-I levels on the expression of genes associated with the above mentioned processes.The results indicated that increased intrafollicular concentrations of IGF-I caused changes in expression of genes associated with steroidogenesis (StAR, SRF) and apoptosis (BCL-2, FKHR, PAWR).These results taken together suggest that onset of gonadotropin surge triggers activation of various biological pathways and that the effects of growth factors and peptides on gonadotropin actions could be examined during preovulatory follicle development.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Reproduction, Development and Genetics, Indian Institute of Science, Bangalore, India.

ABSTRACT
The preovulatory follicle in response to gonadotropin surge undergoes dramatic biochemical, and morphological changes orchestrated by expression changes in hundreds of genes. Employing well characterized bovine preovulatory follicle model, granulosa cells (GCs) and follicle wall were collected from the preovulatory follicle before, 1, 10 and 22 h post peak LH surge. Microarray analysis performed on GCs revealed that 450 and 111 genes were differentially expressed at 1 and 22 h post peak LH surge, respectively. For validation, qPCR and immunocytochemistry analyses were carried out for some of the differentially expressed genes. Expression analysis of many of these genes showed distinct expression patterns in GCs and the follicle wall. To study molecular functions and genetic networks, microarray data was analyzed using Ingenuity Pathway Analysis which revealed majority of the differentially expressed genes to cluster within processes like steroidogenesis, cell survival and cell differentiation. In the ovarian follicle, IGF-I is established to be an important regulator of the above mentioned molecular functions. Thus, further experiments were conducted to verify the effects of increased intrafollicular IGF-I levels on the expression of genes associated with the above mentioned processes. For this purpose, buffalo cows were administered with exogenous bGH to transiently increase circulating and intrafollicular concentrations of IGF-I. The results indicated that increased intrafollicular concentrations of IGF-I caused changes in expression of genes associated with steroidogenesis (StAR, SRF) and apoptosis (BCL-2, FKHR, PAWR). These results taken together suggest that onset of gonadotropin surge triggers activation of various biological pathways and that the effects of growth factors and peptides on gonadotropin actions could be examined during preovulatory follicle development.

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Expression of genes associated with steroidogenesis, cell adhesion and proteolysis in the periovulatory follicle.(A) Microarray fold expression changes at 1 and 22 h post peak LH surge in granulosa cells for few of the selected up and down regulated genes associated with specific biological process. (B) The genes selected by microarray analysis were also subjected to qPCR analysis and fold expression changes for granulosa cells and follicular wall collected at different time points are represented as bar graphs. Bar for each gene represents mean ± SEM fold expression change value at each time point (n = 3 animals). For each gene, bars with different alphabets above them are significantly different (p<0.05). More details on the analysis are provided in the results section.
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pone-0020754-g006: Expression of genes associated with steroidogenesis, cell adhesion and proteolysis in the periovulatory follicle.(A) Microarray fold expression changes at 1 and 22 h post peak LH surge in granulosa cells for few of the selected up and down regulated genes associated with specific biological process. (B) The genes selected by microarray analysis were also subjected to qPCR analysis and fold expression changes for granulosa cells and follicular wall collected at different time points are represented as bar graphs. Bar for each gene represents mean ± SEM fold expression change value at each time point (n = 3 animals). For each gene, bars with different alphabets above them are significantly different (p<0.05). More details on the analysis are provided in the results section.

Mentions: Figure 6 shows qPCR fold change in expression pattern with corresponding microarray data of selected genes associated with regulation of steroidogenesis, cell adhesion and proteolysis. Expression of StAR was shown to be up regulated after the onset of peak LH surge by both microarray and qPCR analyses, while its expression in the follicle wall did not change significantly (Fig. 6A&B). Expression of other steroidogenic genes, CYP19A1 and HSD17β showed marked changes in GCs, while their expression was not detectable in the follicle wall (Fig. 6B). Expression of CTGF, a marker of differentiation was up regulated in GCs after the occurrence of peak LH surge, whereas, its expression in the follicle wall did not appear to be regulated (Fig. 6B). Expression of PLAT, an ECM modulator showed up regulation at both 1 and 22 h time points in GCs by microarray analysis, whereas was high only at 1 and 10 h, but was low at 22 h time point in qPCR expression analysis (Fig. 6A&B). The PLAT expression in the follicle wall samples was similar to the pattern in GCs seen after the occurrence of peak LH surge i.e., up regulated at 1 and 10 h, whereas, down regulated at 22 h (Fig. 6B). Expression of PLAU, another ECM modulator was up regulated at all time points post peak LH surge in GCs by microarray analysis, but the expression was up regulated only at 22 h by qPCR analysis (Fig. 6B). In the follicle wall, the expression of PLAU was up regulated after occurrence of peak LH surge (Fig. 6B). Expression of SERPINE-1, an inhibitor of PLAT and PLAU was up regulated in GCs and follicle wall by both microarray and qPCR (except 1 h) analyses, (p<0.05; Fig. 6B). Expression of ASPN became down regulated after occurrence of peak LH surge in GCs, while in the follicle wall the expression appeared to be down regulated only at 10 h time point post peak LH surge (p<0.05, Fig. 6B).


Gene expression profiling of preovulatory follicle in the buffalo cow: effects of increased IGF-I concentration on periovulatory events.

Rao JU, Shah KB, Puttaiah J, Rudraiah M - PLoS ONE (2011)

Expression of genes associated with steroidogenesis, cell adhesion and proteolysis in the periovulatory follicle.(A) Microarray fold expression changes at 1 and 22 h post peak LH surge in granulosa cells for few of the selected up and down regulated genes associated with specific biological process. (B) The genes selected by microarray analysis were also subjected to qPCR analysis and fold expression changes for granulosa cells and follicular wall collected at different time points are represented as bar graphs. Bar for each gene represents mean ± SEM fold expression change value at each time point (n = 3 animals). For each gene, bars with different alphabets above them are significantly different (p<0.05). More details on the analysis are provided in the results section.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3119055&req=5

pone-0020754-g006: Expression of genes associated with steroidogenesis, cell adhesion and proteolysis in the periovulatory follicle.(A) Microarray fold expression changes at 1 and 22 h post peak LH surge in granulosa cells for few of the selected up and down regulated genes associated with specific biological process. (B) The genes selected by microarray analysis were also subjected to qPCR analysis and fold expression changes for granulosa cells and follicular wall collected at different time points are represented as bar graphs. Bar for each gene represents mean ± SEM fold expression change value at each time point (n = 3 animals). For each gene, bars with different alphabets above them are significantly different (p<0.05). More details on the analysis are provided in the results section.
Mentions: Figure 6 shows qPCR fold change in expression pattern with corresponding microarray data of selected genes associated with regulation of steroidogenesis, cell adhesion and proteolysis. Expression of StAR was shown to be up regulated after the onset of peak LH surge by both microarray and qPCR analyses, while its expression in the follicle wall did not change significantly (Fig. 6A&B). Expression of other steroidogenic genes, CYP19A1 and HSD17β showed marked changes in GCs, while their expression was not detectable in the follicle wall (Fig. 6B). Expression of CTGF, a marker of differentiation was up regulated in GCs after the occurrence of peak LH surge, whereas, its expression in the follicle wall did not appear to be regulated (Fig. 6B). Expression of PLAT, an ECM modulator showed up regulation at both 1 and 22 h time points in GCs by microarray analysis, whereas was high only at 1 and 10 h, but was low at 22 h time point in qPCR expression analysis (Fig. 6A&B). The PLAT expression in the follicle wall samples was similar to the pattern in GCs seen after the occurrence of peak LH surge i.e., up regulated at 1 and 10 h, whereas, down regulated at 22 h (Fig. 6B). Expression of PLAU, another ECM modulator was up regulated at all time points post peak LH surge in GCs by microarray analysis, but the expression was up regulated only at 22 h by qPCR analysis (Fig. 6B). In the follicle wall, the expression of PLAU was up regulated after occurrence of peak LH surge (Fig. 6B). Expression of SERPINE-1, an inhibitor of PLAT and PLAU was up regulated in GCs and follicle wall by both microarray and qPCR (except 1 h) analyses, (p<0.05; Fig. 6B). Expression of ASPN became down regulated after occurrence of peak LH surge in GCs, while in the follicle wall the expression appeared to be down regulated only at 10 h time point post peak LH surge (p<0.05, Fig. 6B).

Bottom Line: Thus, further experiments were conducted to verify the effects of increased intrafollicular IGF-I levels on the expression of genes associated with the above mentioned processes.The results indicated that increased intrafollicular concentrations of IGF-I caused changes in expression of genes associated with steroidogenesis (StAR, SRF) and apoptosis (BCL-2, FKHR, PAWR).These results taken together suggest that onset of gonadotropin surge triggers activation of various biological pathways and that the effects of growth factors and peptides on gonadotropin actions could be examined during preovulatory follicle development.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Reproduction, Development and Genetics, Indian Institute of Science, Bangalore, India.

ABSTRACT
The preovulatory follicle in response to gonadotropin surge undergoes dramatic biochemical, and morphological changes orchestrated by expression changes in hundreds of genes. Employing well characterized bovine preovulatory follicle model, granulosa cells (GCs) and follicle wall were collected from the preovulatory follicle before, 1, 10 and 22 h post peak LH surge. Microarray analysis performed on GCs revealed that 450 and 111 genes were differentially expressed at 1 and 22 h post peak LH surge, respectively. For validation, qPCR and immunocytochemistry analyses were carried out for some of the differentially expressed genes. Expression analysis of many of these genes showed distinct expression patterns in GCs and the follicle wall. To study molecular functions and genetic networks, microarray data was analyzed using Ingenuity Pathway Analysis which revealed majority of the differentially expressed genes to cluster within processes like steroidogenesis, cell survival and cell differentiation. In the ovarian follicle, IGF-I is established to be an important regulator of the above mentioned molecular functions. Thus, further experiments were conducted to verify the effects of increased intrafollicular IGF-I levels on the expression of genes associated with the above mentioned processes. For this purpose, buffalo cows were administered with exogenous bGH to transiently increase circulating and intrafollicular concentrations of IGF-I. The results indicated that increased intrafollicular concentrations of IGF-I caused changes in expression of genes associated with steroidogenesis (StAR, SRF) and apoptosis (BCL-2, FKHR, PAWR). These results taken together suggest that onset of gonadotropin surge triggers activation of various biological pathways and that the effects of growth factors and peptides on gonadotropin actions could be examined during preovulatory follicle development.

Show MeSH
Related in: MedlinePlus