Dataset Information

Rap1 regulates hepatic stellate cell migration through the modulation of RhoA activity in response to TGF??1.

ABSTRACT: Although the migration of hepatic stellate cells (HSCs) is important for hepatic fibrosis, the regulation of this migration is poorly understood. Notably, transforming growth factor (TGF)??1 induces monocyte migration to sites of injury or inflammation during the early phase, but inhibits cell migration during the late phase. In the present study, the role of transforming protein RhoA signaling in TGF??1?induced HSC migration was investigated. TGF??1 was found to increase the protein and mRNA levels of smooth muscle actin and collagen type I in HSC?T6 cells. The level of RhoA?GTP in TGF??1?stimulated cells was significantly higher than that in control cells. Furthermore, the phosphorylation of cofilin and formation of filamentous actin (F?actin) were more marked in TGF??1?stimulated cells than in control cells. Additionally, TGF??1 induced the activation of nuclear factor??B, and the expression of extracellular matrix proteins and several cytokines in HSC?T6 cells. The active form of Rap1 (Rap1 V12) suppressed RhoA?GTP levels, whereas the dominant?negative form of Rap1 (Rap1 N17) augmented RhoA?GTP levels. Therefore, the data confirmed that Rap1 regulated the activation of RhoA in TGF??1?stimulated HSC?T6 cells. These findings suggest that TGF??1 regulates Rap1, resulting in the suppression of RhoA, activation of and formation of F?actin during the migration of HSCs.

SUBMITTER: Moon MY

PROVIDER: S-EPMC6605627 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Rap1 regulates hepatic stellate cell migration through the modulation of RhoA activity in response to TGF‑β1.

Moon Mi-Young MY Kim Hee-Jun HJ Kim Mo-Jong MJ Uhm Sunho S Park Ji-Won JW Suk Ki-Tae KT Park Jae-Bong JB Kim Dong-Jun DJ Kim Sung-Eun SE

International journal of molecular medicine 20190530 2

Although the migration of hepatic stellate cells (HSCs) is important for hepatic fibrosis, the regulation of this migration is poorly understood. Notably, transforming growth factor (TGF)‑β1 induces monocyte migration to sites of injury or inflammation during the early phase, but inhibits cell migration during the late phase. In the present study, the role of transforming protein RhoA signaling in TGF‑β1‑induced HSC migration was investigated. TGF‑β1 was found to increase the protein and mRNA le ...[more]

PMID: 31173168

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Project description:Monocyte migration requires the dynamic redistribution of integrins through a regulated endo-exocytosis cycle, but the complex molecular mechanisms underlying this process have not been fully elucidated. Glia maturation factor-γ (GMFG), a novel regulator of the Arp2/3 complex, has been shown to regulate directional migration of neutrophils and T-lymphocytes. In this study, we explored the important role of GMFG in monocyte chemotaxis, adhesion, and β1-integrin turnover. We found that knockdown of GMFG in monocytes resulted in impaired chemotactic migration toward formyl-Met-Leu-Phe (fMLP) and stromal cell-derived factor 1α (SDF-1α) as well as decreased α5β1-integrin-mediated chemoattractant-stimulated adhesion. These GMFG knockdown impaired effects could be reversed by cotransfection of GFP-tagged full-length GMFG. GMFG knockdown cells reduced the cell surface and total protein levels of α5β1-integrin and increased its degradation. Importantly, we demonstrate that GMFG mediates the ubiquitination of β1-integrin through knockdown or overexpression of GMFG. Moreover, GMFG knockdown retarded the efficient recycling of β1-integrin back to the plasma membrane following normal endocytosis of α5β1-integrin, suggesting that the involvement of GMFG in maintaining α5β1-integrin stability may occur in part by preventing ubiquitin-mediated degradation and promoting β1-integrin recycling. Furthermore, we observed that GMFG interacted with syntaxin 4 (STX4) and syntaxin-binding protein 4 (STXBP4); however, only knockdown of STXBP4, but not STX4, reduced monocyte migration and decreased β1-integrin cell surface expression. Knockdown of STXBP4 also substantially inhibited β1-integrin recycling in human monocytes. These results indicate that the effects of GMFG on monocyte migration and adhesion probably occur through preventing ubiquitin-mediated proteasome degradation of α5β1-integrin and facilitating effective β1-integrin recycling back to the plasma membrane.

Project description:BackgroundIn mice, liver fibrosis is the most serious pathologic change during Schistosoma japonicum (S. japonicum) infection. Schistosomiasis is mainly characterized by schistosome egg-induced granulomatous fibrosis. Hepatic stellate cells (HSCs) are mainly responsible for the net accumulation of collagens and fibrosis formation in the liver. Activated HSCs regulated by transforming growth factor-β1 (TGF-β1)/Smad signaling have emerged as the critical regulatory pathway in hepatitis virus or carbon tetrachloride-induced liver fibrosis. However, the detailed mechanism of HSC activation in schistosome-induced liver fibrosis is poorly understood.MethodsSchistosoma japonicum-induced murine models and a control group were generated by abdominal infection with 15 (± 1) cercariae. The purity of cultured primary HSCs was evaluated by immunocytochemistry. The histopathological changes in the livers of infected mice were estimated by hematoxylin-eosin and Masson staining. Dynamic expression of pro-fibrotic molecules and microRNAs was detected by real-time quantitative PCR (RT-qPCR). Mainly members involved in the TGF-β1/Smad signaling pathway were examined via RT-qPCR and Western blot.ResultsThe egg-induced granulomatous inflammation formed at 4 weeks post-infection (wpi) and developed progressively. Alpha-smooth muscle actin (α-SMA), collagen I, collagen III, TGF-β1, Smad2, Smad3, and Smad4 showed a significant increase in mitochondrial RNA (mRNA) and protein expression compared with the control group at 7 and 9 weeks post-infection (wpi), while an opposite effect on Smad7 was observed. In addition, the mRNA expression of microRNA-21 (miRNA-21) was significantly increased at 7 wpi, and the mRNA expression of miRNA-454 was decreased starting from 4 wpi.ConclusionOur present findings revealed that HSCs regulated by the TGF-β1/Smad signaling pathway play an important role in liver fibrosis in S. japonicum-infected mice, which may provide proof of concept for liver fibrosis in schistosomiasis.

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Rap1 regulates hepatic stellate cell migration through the modulation of RhoA activity in response to TGF??1.

Publications

Rap1 regulates hepatic stellate cell migration through the modulation of RhoA activity in response to TGF‑β1.

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