No abstract

Hepatic stellate cell (HSC) activation is a pivotal event in initiation and progression of hepatic fibrosis and a major contributor to collagen deposition driven by transforming growth factor beta (TGFβ). microRNAs (miRs), small non-coding RNAs modulating mRNA and protein expression, have emerged as key regulatory molecules in chronic liver disease. We investigated differentially expressed miRs in quiescent and activated HSCs to identify novel regulators of profibrotic TGFβ signaling. miR microarray analysis was performed on quiescent and activated rat HSCs. Members of the miR-17-92 cluster (19a, 19b, 92a) were significantly down-regulated in activated HSCs. Since miR 19b showed the highest fold-change of the cluster members, activated HSCs were transfected with miR 19b mimic or negative control and TGFβ signaling and HSC activation assessed. miR 19b expression was determined in fibrotic rat and human liver specimens. miR 19b mimic negatively regulated TGFβ signaling components demonstrated by decreased TGFβ receptor II (TGFβRII) and SMAD3 expression. Computational prediction of miR 19b binding to the 3’UTR of TGFβRII was validated by luciferase reporter assay. Inhibition of TGFβ signaling by miR 19b was confirmed by decreased expression of type I collagen and by blocking TGFβ-induced expression of α1(I) and α2(I) procollagen mRNAs. miR 19b blunted the activated HSC phenotype by morphological assessment and decreased αSMA expression. Additionally, miR 19b expression was markedly diminished in fibrotic rat liver compared to normal liver; similarly, miR 19b expression was markedly down-regulated in fibrotic compared to normal human livers. CONCLUSIONS miR 19b is a novel regulator of TGFβ signaling in HSCs suggesting a potential therapeutic approach for hepatic fibrosis.

The age-related decline in female fertility has been attributed to a variety of causes including progressive oocyte depletion, meiotic irregularities and mitochondrial dysfunction. However, additional factors could potentially be involved. To explore this possibility, comprehensive analysis of gene expression in human oocytes, discarded following IVF procedures and segregated by age, was undertaken using microarray methods. These findings indicate that the expression of oocyte genes, in a variety of major functional categories including cell cycle regulation, cytoskeletal structure, energy pathways, transcription control, and stress responses, are influenced by maternal age. These results are corroborated by a complementary extensive study using mouse oocytes.

The spindle assembly checkpoint modulates the timing of anaphase initiation in response to the improper alignment of chromosomes at the metaphase plate. If defects are detected, a signal is transduced to halt further progression of the cell cycle until correct bipolar attachment to the spindle is achieved. The mitotic arrest deficient (MAD2) and budding uninhibited by benomyl (BUB1) genes encode conserved kinetochore-associated proteins believed to be components of the checkpoint regulatory pathway. A failure in this surveillance system could lead to genomic instability that may underlie the increased incidence of aneuploidy in the gametes of older women. To explore this possibility, the concentrations of these transcripts in human oocytes at various stages of maturation were determined by real-time rapid cycle fluorescent reverse transcription-polymerase chain reaction (RT-PCR). The results obtained following quantitative analysis suggest that these messages degrade as oocytes age. Potentially, this may impair checkpoint function in older oocytes and may be a contributing factor in age-related aneuploidy.