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Thymosin ?4 inhibits PDGF-BB induced activation, proliferation, and migration of human hepatic stellate cells via its actin-binding domain.

ABSTRACT: OBJECTIVES:Hepatic stellate cells (HSC) trans-differentiation is central to the development of liver fibrosis, marked by the expression of pro-fibrogenic genes and the proliferation and migration of activated HSC. Therefore, preventing and/or reverting the activation, proliferation, and migration of HSC may lead to new therapies for treating fibrosis/cirrhosis. Thymosin ?4 (T?4) inhibits PDGF-BB-induced fibrogenesis, proliferation and migration of HSC by blocking Akt phosphorylation. Here, we utilized T?4-derived peptides: amino-terminal-Ac-SDKPDMAEIEKFDKS (1-15aa) and actin-binding-LKKTETQ (17-23aa) to investigate the molecular mechanisms in the anti-fibrogenic actions of T?4. METHODS:We used RT-PCR, Western blot, and proliferation and migration assays in early passages of human HSC cultures treated with PDGF-BB and/or T?4 peptides. RESULTS:We showed that 17-23aa but not 1-15aa inhibited PDGF-BB-dependent up-regulation of PDGF? receptor, ?-SMA, and collagen 1. It also blunted the phosphorylation of Akt at T 308 and S473, resulting in the inhibition of phosphorylation of PRAS40, and HSC proliferation and migration. Interestingly, 1-15aa blocked Akt phosphorylation at S473, but not T308 by inhibiting mTOR phosphorylation, thus, it did not have any effect on HSC proliferation and migration. CONCLUSION:These findings suggest that while 1-15aa has a minor effect on Akt phosphorylation, the anti-fibrogenic actions of T?4 are exerted via 17-23aa.

SUBMITTER: Shah R

PROVIDER: S-EPMC6748868 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature

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Thymosin β4 inhibits PDGF-BB induced activation, proliferation, and migration of human hepatic stellate cells via its actin-binding domain.

Shah Ruchi R Reyes-Gordillo Karina K Rojkind Marcos M

Expert opinion on biological therapy 20180701 sup1

<h4>Objectives</h4>Hepatic stellate cells (HSC) trans-differentiation is central to the development of liver fibrosis, marked by the expression of pro-fibrogenic genes and the proliferation and migration of activated HSC. Therefore, preventing and/or reverting the activation, proliferation, and migration of HSC may lead to new therapies for treating fibrosis/cirrhosis. Thymosin β4 (Tβ4) inhibits PDGF-BB-induced fibrogenesis, proliferation and migration of HSC by blocking Akt phosphorylation. Her ...[more]

PMID: 30063851

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Project description:The purpose of this study was to explore the effect of miR-34c on PDGF-BB-induced HAVSMCs phenotypic transformation and proliferation via PDGFR-β/SIRT1 pathway, so as to find a new method for early diagnosis and treatment of cardiovascular disease. HA-VSMCs were treated with platelet-derived growth factor-BB (PDGF-BB) at 0 h, 12 h, 24 h, 48 h or 36 h to explore the optimal time for phenotypic transformation of VSMCs. And then, PDGF-BB-induced HA-VSMCs were transfected with miR-34c mimics/mimics NC and pcDNA3.1-PDGFR-β/pcDNA3.1-NC to observe cell biological behaviour. CCK8 was used to detect cell proliferation activity. Transwell chamber assay was used to detect cell invasion. Early apoptosis was analyzed by flow cytometry. The expression of α-SMA and Smemb was detected by immunofluorescence staining. The expressions of PDGFR-β, IRF9, Acetyl-NF-κB/p65, Acetyl-p53 and CyclinD1 were analyzed by Western blot analysis. The expression of miR-34a, miR-34b and miR-34c was detected by RT-PCR, and the targeting relationship between miR-34c and PDGFR-β was detected by luciferase reporting assay. The results indicated the proliferation and migration of PDGF-BB-induced HA-VSMCs significantly increased, and apoptosis significantly decreased. Besides, α-SMA decreased significantly, while Smemb increased significantly. Furthermore, expressions of PDGFR-β, IRF9, Acetyl-NF-κB/p65, Acetyl-p53 and CyclinD1 increased significantly, and SIRT1 decreased significantly. Experimental results showed that, miR-34c mimics significantly inhibited cell proliferation and migration, and promoted cell apoptosis, and miR-34c inhibitor had the opposite effects. MiR-34c mimics significantly increased α-SMA expression and decreased Smemb expression, while the opposite effects were reflected after transfection with miR-34c inhibitor. Moreover, miR-34c mimics significantly decreased the expressions of PDGFR-β, IRF9, Acetyl-NF-κB/p65, Acetyl-p53 and CyclinD1, and significantly increased the expression of SIRT1, while miR-34c inhibitor had the opposite effects. Luciferase assay confirmed that PDGFR-β was a potential target of miR-34c. Subsequently, PDGF-BB-induced HA-VSMCs were co-transfected with miR-34c mimics and pcDNA3.1-PDGFR-β. The results indicated that PDGFR-β reversed the biological function of miR-34c mimic. The results revealed the potential application value of miR-34c as a marker molecule of phenotypic transformation, providing a potential target for improving phenotypic transformation.

Dataset Information

Thymosin ?4 inhibits PDGF-BB induced activation, proliferation, and migration of human hepatic stellate cells via its actin-binding domain.

Publications

Thymosin β4 inhibits PDGF-BB induced activation, proliferation, and migration of human hepatic stellate cells via its actin-binding domain.

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