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Transcriptome Profiling of the Theca Interna from Bovine Ovarian Follicles during Atresia. Hatzirodos N 2014 et al.
The theca interna is a specialized stromal layer that envelops each growing ovarian follicle. It contains capillaries, fibroblasts, immune cells and the steroidogenic cells that synthesize androgens for conversion to estradiol by the neighboring granulosa cells. During reproductive life only a small number of follicles will grow to a sufficient size to ovulate, whereas the majority of follicles will undergo regression/atresia and phagocytosis by macrophages. To identify genes which are differentially regulated in the theca interna during follicular atresia, we undertook transcriptome profiling of the theca interna from healthy (n?=?10) and antral atretic (n?=?5) bovine follicles at early antral stages (<5 mm). Principal Component Analyses and hierarchical classification of the signal intensity plots for the arrays showed primary clustering into two groups, healthy and atretic. A total of 543 probe sets were differentially expressed between the atretic and healthy theca interna. Further analyses of these genes by Ingenuity Pathway Analysis and Gene Ontology Enrichment Analysis Toolkit software found most of the genes being expressed were related to cytokines, hormones and receptors as well as the cell cycle and DNA replication. Cell cycle genes which encode components of the replicating chromosome complex and mitotic spindle were down-regulated in atretic theca interna, whereas stress response and inflammation-related genes such as TP53, IKBKB and TGFB1 were up-regulated. In addition to cell cycle regulators, upstream regulators that were predicted to be inhibited included Retinoblastoma 1, E2 transcription factor 1, and hepatocyte growth factor. Our study suggests that during antral atresia of small follicles in the theca interna, arrest of cell cycle and DNA replication occurs rather than up- regulation of apoptosis-associated genes as occurs in granulosa cells.
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Characterization of the Inhibitor of KappaB Kinase (IKK) Complex in Granulosa Cell Tumors of the Ovary and Granulosa Cell Tumor-Derived Cell Lines. Jamieson S et al. Granulosa cell tumors of the ovary (GCT) are a distinct, hormonally active subset of ovarian cancers. Although it has recently been shown that ~97?% of all adult GCT harbor a novel somatic missense mutation in the FOXL2 gene, given its almost universal presence, it does not explain differences in tumor stage and/or recurrence. The nuclear factor kappaB (NF?B) transcription factor is constitutively active in two human GCT-derived cell lines, COV434 and KGN, which are useful in vitro models to investigate juvenile and adult GCT, respectively. This study aimed to determine the molecular basis and pathogenetic significance of this aberrant NF?B activity. Selective chemical inhibitors were used to target candidate components of the pathway. The constitutive activity was blocked by two independent inhibitors of I?Ba phosphorylation, suggesting that aberrant activation occurs upstream of this point. NF?B inhibition resulted in a dose-dependent decrease in cell proliferation and viability and a dose-dependent increase in apoptosis. Inhibitors of earlier components of the pathway were without effect. Two independent inhibitors of inhibitor of kappaB kinase (IKK)? a catalytic subunit of the NF?B activation complex, were unable to inhibit the constitutive activity, but surprisingly also ligand-induced activity. These findings suggest a central role for IKK? however, no mutations or altered expression of the IKK? IKKa, or IKK? genes was observed in the cell lines or in a panel of human GCT samples. This study highlights unresolved issues in understanding the pathogenesis of GCT and in the use of the COV434 and KGN cells lines as model systems.
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