Expressions of glutathione S-transferase alpha, mu, and pi in brains of medically intractable epileptic patients

<p>Abstract</p> <p>Background</p> <p>Glutathione S-transferases (GSTs) play an important role in metabolizing anti-epileptic drugs (AEDs) in liver. Expressions of GSTs in brain, which may result in poor efficacy of AEDs, have not been well studied. Using clinical cortex specimen from 32 intractable epileptic subjects and 8 non-epileptic controls, the present study investigated the correlation between GSTs and intractable epilepsy.</p> <p>Results</p> <p>Three different GST isoforms (α, μ, and π) were detected with immunohistochemistry. GST-α expression was not seen in any cortex specimens. Sixty three percent (63%) of control and 53% of intractible epileptic specimens showed GST-μ immunoreactivity. No significant difference in intensity of GST-μ staining was observed between these two groups. GST-π expression was found in endothelial cells and glial cells/astrocytes. Fifty percent (50%) of the control patients and 66% of the epileptic patients were GST-π positive. The grading of epileptic patients was significantly higher than that of control patients (<it>p </it>< 0.01).</p> <p>Conclusion</p> <p>High levels of GST-π in endothelial cells and glial cells/astrocyte correlate to medical intractable epilepsy, suggesting that GST-π contributes to resistance to AED treatment.</p

A Novel, Non-Apoptotic Role for Scythe/BAT3: A Functional Switch between the Pro- and Anti-Proliferative Roles of p21 during the Cell Cycle

BACKGROUND: Scythe/BAT3 is a member of the BAG protein family whose role in apoptosis has been extensively studied. However, since the developmental defects observed in Bat3-null mouse embryos cannot be explained solely by defects in apoptosis, we investigated whether BAT3 is also involved in cell-cycle progression. METHODS/PRINCIPAL FINDINGS: Using a stable-inducible Bat3-knockdown cellular system, we demonstrated that reduced BAT3 protein level causes a delay in both G1/S transition and G2/M progression. Concurrent with these changes in cell-cycle progression, we observed a reduction in the turnover and phosphorylation of the CDK inhibitor p21, which is best known as an inhibitor of DNA replication; however, phosphorylated p21 has also been shown to promote G2/M progression. Our findings indicate that in Bat3-knockdown cells, p21 continues to be synthesized during cell-cycle phases that do not normally require p21, resulting in p21 protein accumulation and a subsequent delay in cell-cycle progression. Finally, we showed that BAT3 co-localizes with p21 during the cell cycle and is required for the translocation of p21 from the cytoplasm to the nucleus during the G1/S transition and G2/M progression. CONCLUSION: Our study reveals a novel, non-apoptotic role for BAT3 in cell-cycle regulation. By maintaining a low p21 protein level during the G1/S transition, BAT3 counteracts the inhibitory effect of p21 on DNA replication and thus enables the cells to progress from G1 to S phase. Conversely, during G2/M progression, BAT3 facilitates p21 phosphorylation by cyclin A/Cdk2, an event required for G2/M progression. BAT3 modulates these pro- and anti-proliferative roles of p21 at least in part by regulating cyclin A abundance, as well as p21 translocation between the cytoplasm and the nucleus to ensure that it functions in the appropriate intracellular compartment during each phase of the cell cycle.Dissertatio

HLA-B-associated transcript 3 (Bat3/Scythe) negatively regulates Smad phosphorylation in BMP signaling

Members of the transforming growth factor-β (TGF-β) superfamily participate in numerous biological phenomena in multiple tissues, including in cell proliferation, differentiation, and migration. TGF-β superfamily proteins therefore have prominent roles in wound healing, fibrosis, bone formation, and carcinogenesis. However, the molecular mechanisms regulating these signaling pathways are not fully understood. Here, we describe the regulation of bone morphogenic protein (BMP) signaling by Bat3 (also known as Scythe or BAG6). Bat3 overexpression in murine cell lines suppresses the activity of the Id1 promoter normally induced by BMP signaling. Conversely, Bat3 inactivation enhances the induction of direct BMP target genes, such as Id1, Smad6, and Smad7. Consequently, Bat3 deficiency accelerates the differentiation of primary osteoblasts into bone, with a concomitant increase in the bone differentiation markers Runx2, Osterix, and alkaline phosphatase. Using biochemical and cell biological analyses, we show that Bat3 inactivation sustains the C-terminal phosphorylation and nuclear localization of Smad1, 5, and 8 (Smad1/5/8), thereby enhancing biological responses to BMP treatment. At the mechanistic level, we show that Bat3 interacts with the nuclear phosphatase small C-terminal domain phosphatase (SCP) 2, which terminates BMP signaling by dephosphorylating Smad1/5/8. Notably, Bat3 enhances SCP2–Smad1 interaction only when the BMP signaling pathway is activated. Our results demonstrate that Bat3 is an important regulator of BMP signaling that functions by modulating SCP2–Smad interaction

Identification of Kinases Regulating Prostate Cancer Cell Growth Using an RNAi Phenotypic Screen

As prostate cancer progresses to castration-resistant disease, there is an increase in signal transduction activity. Most castration-resistant prostate tumors continue to express the androgen receptor (AR) as well as androgen-responsive genes, despite the near absence of circulating androgen in these patients. The AR is regulated not only by its cognate steroid hormone, but also by interactions with a constellation of co-regulatory and signaling molecules. Thus, the elevated signaling activity that occurs during progression to castration resistance can affect prostate cancer cell growth either through the AR or independent of the AR. In order to identify signaling pathways that regulate prostate cancer cell growth, we screened a panel of shRNAs targeting 673 human kinases against LNCaP prostate cancer cells grown in the presence and absence of hormone. The screen identified multiple shRNA clones against known and novel gene targets that regulate prostate cancer cell growth. Based on the magnitude of effect on growth, we selected six kinases for further study: MAP3K11, DGKD, ICK, CIT, GALK2, and PSKH1. Knockdown of these kinases decreased cell growth in both androgen-dependent and castration-resistant prostate cancer cells. However, these kinases had different effects on basal or androgen-induced transcriptional activity of AR target genes. MAP3K11 knockdown most consistently altered transcription of AR target genes, suggesting that MAP3K11 affected its growth inhibitory effect by modulating the AR transcriptional program. Consistent with MAP3K11 acting on the AR, knockdown of MAP3K11 inhibited AR Ser 650 phosphorylation, further supporting stress kinase regulation of AR phosphorylation. This study demonstrates the applicability of lentiviral-based shRNA for conducting phenotypic screens and identifies MAP3K11, DGKD, ICK, CIT, GALK2, and PSKH1 as regulators of prostate cancer cell growth. The thorough evaluation of these kinase targets will pave the way for developing more effective treatments for castration-resistant prostate cancer

Development of a toxicokinetic model in rat of the cis and trans-permethrin and theirs three metabolites.

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