333. HETEROGENEITY OF GENE EXPRESSION IN BOVINE SMALL FOLLICLES

Abstract

Small antral follicles <5 mm in bovine ovaries undergo one of two fates: further growth and selection to become the dominant follicle for ovulation, or atresia. Atresia can occur before, during or after selection. As follicle grow past >5 mm there is upregulation in expression of focimatrix genes and later upregulation of the LH receptor and steroidogenic enzymes, especially aromatase, in the granulosa cells. For follicles at sizes >5 mm entering atresia the granulosa cells are the first in the follicle to die. Thus expression of genes in granulosa cells is critical to the fate of the follicle. To examine granulosa cells of small follicles we collected bovine ovaries and dissected follicles, removed part of the follicle wall for subsequent classification of health or atresia, and harvested the remaining granulosa cells for RNA isolation. Follicles examined included small follicles (<5 mm), both healthy (n = 10) and atretic (n =5), and healthy large follicles (>10 mm, n = 4). RNA was hybridized to Affymetrix GeneChip Bovine Genome Arrays and the results were analysed using Partek Genomics Suite software. The number of genes which were 2 fold differentially regulated between large and small follicles by Benjamini Hochberg post hoc test (False Discovery Rate, P < 0.05) was 2408 and between healthy and atretic small follicles was 4931. The coefficient of variation (CV; SD/mean × 100) for the expression level of each gene for each group was calculated. A gene frequency distribution indicated greater heterogeneity in expression levels in small follicles in comparison to large follicles. Furthermore, the greatest variability in genes in small follicles includes those that are either up or down regulated due to atresia or growth. We therefore conclude that variability in small follicles is a consequence of alternative fates that small follicle can undergo.