Although the RhoA/Rho kinase (RhoA/ROK) pathway has been extensively investigated, its roles and downstream signaling pathways are still not well understood in myogenic processes. Therefore, we examined the effects of RhoA/ROK on myogenic processes and their signaling molecules using H9c2 and C2C12 cells. Increases in RhoA/ROK activities and serine phosphorylation levels of insulin receptor substrate (IRS)-1 (Ser307 and Ser636/639) and IRS-2 were found in proliferating myoblasts, whereas IRS-1/2 tyrosine phosphorylation and phosphatidylinositol (PI) 3-kinase activity increased during the differentiation process. ROK strongly bound to IRS-1/2 in proliferation medium but dissociated from them in differentiation medium (DM). ROK inactivation by a ROK inhibitor, Y27632, or a dominant-negative ROK, decreased IRS-1/2 serine phosphorylation with increases in IRS-1/2 tyrosine phosphorylation and PI 3-kinase activity, which led to muscle differentiation even in proliferation medium. Inhibition of ROK also enhanced differentiation in DM. ROK activation by a constitutive active ROK blocked muscle differentiation with the increased IRS-1/2 serine phosphorylation, followed by decreases in IRS-1/2 tyrosine phosphorylation and PI 3-kinase activity in DM. Interestingly, fibroblast growth factor-2 added to DM also blocked muscle differentiation through RhoA/ROK activation. Fibroblast growth factor-2 blockage of muscle differentiation was reversed by Y27632. Collectively, these results suggest that the RhoA/ROK pathway blocks muscle differentiation by phosphorylating IRS proteins at serine residues, resulting in the decreased IRS-1/2 tyrosine phosphorylation and PI 3-kinase activity. The absence of the inhibitory effects of RhoA/ROK in DM due to low concentrations of myogenic inhibitory growth factors seems to allow IRS-1/2 tyrosine phosphorylation, which stimulates muscle differentiation via transducing normal myogenic signaling.

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PurposeImmunosuppression is a characteristic of cancer recurrence after curative resection. The neutrophil-to-lymphocyte ratio (NL ratio) in peripheral blood is associated with immune function. However, it is not clear whether the postoperative NL ratio is a predictor for cancer relapse after resection. Thus, we investigated the effectiveness of the short-term postoperative NL ratio in the prediction of disease recurrence within 5 years after stomach cancer surgery by a retrospective chart review.MethodsNinety-three patients with stomach cancer were enrolled. Significant risk factors for cancer recurrence were determined by multivariate Cox regression. Independent variables to increase the NL ratio to >7.7 by postoperative day (POD) 3 were examined by multivariate logistic regression analysis.ResultsThe 5-year risk of cancer recurrence after gastrectomy was 4.2 times higher for patients with a POD3 NL ratio of >7.7 (P = 0.005), 3.4 times higher for normal-weight patients compared with overweight patients (P = 0.008), and 20 times higher for stage III compared with stage 0 according to the tumor-node-metastasis cancer staging system (P = 0.003). The surgical duration (hours) increased the chance of high NL ratio >7.7 (odds ratio, 2.5; P = 0.006).ConclusionThe postoperative NL ratio, especially the POD3 NL ratio, predicts long-term recurrence after stomach cancer surgery.