Positive feedback control between STIM1 and NFATc3 is required for C2C12 myoblast differentiation.

Up-regulation of STIM1-mediated store-operated Ca(2+) entry (SOCE) and Ca(2+)-dependent NFAT signaling is important for myogenic differentiation. However, the molecular mechanisms for differentiation specific up-regulation of STIM1/SOCE-mediated signaling are poorly understood. This study explored whether functional crosstalk between STIM1 and a member of NFAT transcription factor is important for C2C12 myoblast differentiation. Transient increase of NFATc3 expression was observed in the initial phase of differentiation, and the increased activity of NFATc3 isoform was correlated with up-regulation of STIM1 expression. Overexpression of NFATc3 increased STIM1 expression, SOCE activity, and myotube formation, whereas NFATc3 knockdown showed the opposite effects. Overexpression of STIM1 increased the activity and expression level of NFATc3, and enhanced myotube formation, whereas STIM1 knockdown resulted in the opposite effects. Taken together, our findings suggest that a positive feedback control between STIM1/SOCE and NFATc3 is required for efficient induction and progression of myoblast differentiation.

A critical role for AKT activation in protecting cells from ionizing radiation-induced apoptosis and the regulation of acinus gene expression.

Although AKT activation leads to the activation of various pathways related to cell survival, the roles of AKT in modulating cellular responses induced by ionizing radiation in normal human cells remain unclear. Here we show that low-dose radiation of 0.05Gy did not affect cell death, but high-dose radiation (> 0.2Gy) induced apoptosis through the activation of caspases and acinus cleavage. Ionizing radiation induced acinus phosphorylation via AKT activation. Thus, we examined the effect of AKT activation on radiation-induced cell death using CCD-18Lu cells transduced with a retroviral vector expressing constitutively active AKT (CA-AKT). The overexpression of CA-AKT rendered the cells resistant to ionizing radiation and prevented the proteolytic cleavage of acinus via phosphorylation. In addition, overexpression of CA-AKT resulted in the upregulation of acinus expression by activation of the NF-kappaB pathway. On the other hand, suppression of endogenous AKT expression by siRNA resulted in the reduction of acinus expression and enhanced the radiation-induced apoptosis in both CCD-18Lu and IM-9 cells. Our results suggest that AKT activation inhibits cell death during radiation-induced apoptosis through the regulation of phosphorylation and expression of acinus. The AKT/NF-kappaB/acinus pathway functions as one of the important regulatory mechanisms required for modulating ionizing radiation sensitivity.

Immunomodulatory activity of betulinic acid by producing pro-inflammatory cytokines and activation of macrophages.

Betulinic acid (BA), a pentacyclic triterpene isolated from Lycopus lucidus, has been reported to be a selective inducer of apoptosis in various human cancer and shown anti-inflammatory and immunomodulatory properties. We postulated that BA modulates the immunomodulatory properties at least two groups of protein mediators of inflammation, interleukin-1beta (IL-1beta) and the tumor necrosis factor-alpha (TNF-alpha) on the basis of the critical role of the monocytes and tissue macrophages in inflammatory and immune responses. TNF-alpha and IL-1beta were produced by BA in a dose dependent manner at concentration of 0.625 and 10 microg/mL. The production of NO associated with iNOS was inhibited when treated with LPS at the concentration of 2.5 to 20 microg/mL of BA whereas COX-2 expression was decreased at 2.5 to 20 microg/mL. These modulations of inflammatory mediators were examined in LPS-stimulated RAW 264.7 cells and peritoneal macrophages. The morphology of macrophage was also examined and enhanced surface CD 40 molecule was expressed when treated BA at 0.625 to approximately 5 microg/mL with or without LPS. Furthermore, BA (20 microg/mL) enhanced apoptosis by producing DNA ladder in the RAW 264.7 cells. Our results indicated that BA induced activation of macrophage and pro-inflammatory cytokines. This may provide a molecular basis for the ability of BA to mediate macrophage, suppress inflammation, and modulate the immune response.