Summary： The main pathogenesis of saphenous vein graft neointimal hyperplasia after coronary artery bypass grafting （CABG） is inflammation-caused migration and proliferation of vascular smooth muscle cells （VSMCs...Summary： The main pathogenesis of saphenous vein graft neointimal hyperplasia after coronary artery bypass grafting （CABG） is inflammation-caused migration and proliferation of vascular smooth muscle cells （VSMCs）. Janus kinase 2/signal transducer and activators of transcription 3 （JAK2/STAT3） path- way is an important signaling pathway through which VSMCs phenotype conversion occurs. Suppressor of cytokine signaling 3 （SOCS3） is the classic negative feedback inhibitor of JAK2/STAT3 pathway. Growing studies show that SOCS3 plays an important anti-inflammatory role in numerous autoimmune diseases, inflammatory diseases and inflammation-related tumors. However, the effect and mechanism of SOCS3 on vein graft disease is unclear. The purpose of this study was to investigate the effects of SOCS3 on the inflammation, migration and proliferation of VSMCs in vitro and the mechanism. The small interference RNA plasmid targeting rat SOCS3 （SiRNA-rSOCS3） and the recombinant adenovirus vector carrying rat SOCS3 gene （pYrAd-rSOCS3） were constructed, and the empty plamid （SiRNA-control） and vector （pYrAd-GFP） only carrying GFP reported gene were constructed as control. The rat VSMCs were cultured. There were two large groups of A （SOCS3 up-regulated）： control group, IL-6/IFN-γ group, IL-6/IFN-γ＋pYrAd-rSOCS3 group, IL-6/IFN-γ＋pYrAd-GFP group; and B （SOCS3 down-regulated）： control group, IL-6/IFN-γ group, IL-6/IFN-γ＋SiRNA-rSOCS3 group and IL-6/IFN -T＋SiRNA-control group. The pYrAd-rSOCS3 and SiRNA-rSOCS3 were transfected into VSMCs in- duced by IL-6/IFN-γ. After 24 h, real-time reverse transcription polymerase chain reaction （RT-PCR） and Western blotting were used to detect the mRNA and protein expression of SOCS3, STAT3 （only by Western blotting）, P-STAT3 （only by Western blotting）, IL-1β, IL-6, TNF-α, MCP-1 and ICAM-1. The MTT, Transwell assay and flow cytometry were used to examine VSMCs proliferation, migration and cell cycle progression, respectively. As compared with control group, the mRNA and protein expression of SOCS3, STAT3, P-STAT3, IL-1β, IL-6, TNF-α, MCP-1 and ICAM-1 was significantly up-regulated in VSMCs stimulated by IL-6/IFN-γ. However, in VSMCs transfected with pYrAd-rSOCS3 before stimulation with IL-6/IFN-γ, the expression of SOCS3 mRNA and protein was further up-regulated, and that of STAT3, P-STAT3, IL-1β, IL-6, TNF-α, MCP-1 and ICAM-1 was significantly down-regulated as compared with IL-6/IFN-γ group and IL-6/IFN-γ＋pYrAd-GFP group. The expression of those re- lated-cytokines in IL-6/IFN-γ＋SiRNA-rSOCS3 group was markedly increased as compared with IL-6/IFN-γ group and IL-6/IFN-γ＋SiRNA-control group. The absorbance （A） values, the number of cells migrating to the lower chamber, and percentage of cells in the G2/M＋S phase were increased in VSMCs stimulated by IL-6/IFN-γ. In VSMCs incubated with pYrAd-rSOCS3 or SiRNA-rSOCS3 be- fore IL-6/IFN-γ stimulation, the A values, the number of cells migrating to the lower chamber, and the percentage of cells in the G2/M＋S phase were significantly decreased, and increased respectively. These results imply that IL-6/IFN-γ, strong inflammatory stimulators, can promote transformation of VSMCs phenotype form a quiescent contractile state to a synthetic state by activating JAK2/STAT3 pathway. Over-expresssed SOCS3 might inhibit pro-inflammatory effect, migration and growth of VSMCs by blocking STAT3 activation and phosphorylation. These data in vitro confirm that SOCS3 may play a negatively regulatory role in development and progression of vein graft failure. These conclusions can provide a novel strategy for clinical treatment of vein graft diseases and a new theoretic clue for related drug development.展开更多
BACKGROUND:While suppressor of cytokine signaling 3(SOCS3) plays a crucial role in suppressing dysplasia and tumorigenesis,it also offers a typical instance of DNA methylation in the regulation of gene transcription,s...BACKGROUND:While suppressor of cytokine signaling 3(SOCS3) plays a crucial role in suppressing dysplasia and tumorigenesis,it also offers a typical instance of DNA methylation in the regulation of gene transcription,since the promoter region of the SOCS3 gene is rich in CpG islands(CGIs).During liver regeneration initiated by partial hepatectomy,SOCS3 acts as a suppressor to balance the acute-phase response and terminate the regeneration.This study aimed to determine whether the variation of SOCS3 expression throughout liver regeneration is also regulated by its DNA methylation.METHODS:We established a 70% partial hepatectomy mouse model and the animals were sacrificed at indicated times to assess the SOCS3 expression.We performed bisulfite sequencing PCR and DNA sequencing to investigate the detailed cytosine methylation in the SOCS3 gene.RESULTS:Within the promoter sequence,58 CGIs were identified and 30 were found variously methylated before or after operation;however,methylation remained at a very low level.No evidence indicated that the total methylation level or the methylation of any CpG site regularly changed throughout liver regeneration.CONCLUSION:DNA methylation or demethylation seems to be a relatively stable modification of cytosine,but not a dynamic and reversible process to regulate gene transcription in daily and acute pathophysiological events.展开更多
Summary： Recently, suppressor of cytokine signaling-3 （SOCS3） has been shown to be an inducible endogenous negative regulator of Janus kinase/signal transducers and activators of transcription （JAK/STAT） pathway ...Summary： Recently, suppressor of cytokine signaling-3 （SOCS3） has been shown to be an inducible endogenous negative regulator of Janus kinase/signal transducers and activators of transcription （JAK/STAT） pathway which is relevant in inflammatory response, while its functions in acute liver failure and HBV-induced acute-on-chronic liver failure （HBV-ACLF） have not been fully elucidated. In this study, we explored the role of SOCS3 in the development of mouse hepatitis virus strain 3 （MHV-3）-induced acute liver failure and its expression in liver and peripheral blood mononuclear cells （PBMCs） of patients with HBV-ACLF. Inflammation-related gene expression was detected by real-time PCR, immtmohistochemistry and Western blotting. The correlation between SOCS3 level and liver injury was studied. Our results showed that the SOCS3 expression was significantly elevated in both the liver tissue and PBMCs from patients with HBV-ACLF compared to mild chronic hepatitis B （CHB）. Moreover, a time course study showed that SOCS3 level was increased remarkably in the liver of BALB/cJ mice at 72 h post-infection. Pro-inflammatory cytokines, interleukin （IL）-1 β, IL-6, and tumor necrosis factor （TNF）-α, were also increased significantly at 72 h post-infection. There was a close correlation between hepatic SOCS3 level and IL-6, and the severity of liver injury defined by alanine aminotransferase （ALT） and aspartate aminotransferase （AST） levels, respectively. These data suggested that SOCS3 may play a pivotal role in the pathogenesis of MHV-3-induced acute liver failure and HBV-ACLF.展开更多
Objective: To study the correlation of suppressor of cytokine signaling 3 (SOCS3) expression in placenta of intrahepatic cholestasis of pregnancy (ICP) with maternal Th cell imbalance and placental trophoblast cell da...Objective: To study the correlation of suppressor of cytokine signaling 3 (SOCS3) expression in placenta of intrahepatic cholestasis of pregnancy (ICP) with maternal Th cell imbalance and placental trophoblast cell damage. Methods: Primiparae who were diagnosed with ICP in Bazhong Hospital of Traditional Chinese Medicine between June 2014 and August 2017 were selected as the ICP group, and the healthy primiparae who gave birth during the same period were selected as the control group. The placenta was collected after delivery to determine the expression of SOCS3, Th cytokines and apoptosis genes. Results: SOCS3 mRNA expression and protein expression as well as IL-4 and IL-13 protein expression in placenta of ICP group were significantly lower than those of control group whereas IFN-γ, TNF-α, IL-2, p53, Caspase-3, Fas, Caspase-8, Cyt-C and Caspase-9 protein expression were significantly higher than those of control group;IFN-γ, TNF-α, IL-2, p53, Caspase-3, Fas, Caspase-8, Cyt-C and Caspase-9 protein expression in ICP placenta with lower SOCS3 expression were significantly higher than those in ICP placenta with higher SOCS3 expression whereas IL-4 and IL-13 protein expression were significantly lower than those in ICP placenta with higher SOCS3 expression. Conclusion: The low expression of SOCS3 in the ICP placenta can aggravate the imbalance of Th cells and the damage of placental trophoblast cells induced by apoptosis.展开更多
目的观察慢性乙型肝炎患者(CHB)CISH、SOCS1和SOCS3 m RNA及CD4+T淋巴细胞亚群相关细胞因子的表达,探讨CISH、SOCS1和SOCS3在慢性乙型肝炎病毒(HBV)感染中的作用。方法选取慢性乙型肝炎患者(CHB)31例,乙肝携带者(As C)18例,正常健康者1...目的观察慢性乙型肝炎患者(CHB)CISH、SOCS1和SOCS3 m RNA及CD4+T淋巴细胞亚群相关细胞因子的表达,探讨CISH、SOCS1和SOCS3在慢性乙型肝炎病毒(HBV)感染中的作用。方法选取慢性乙型肝炎患者(CHB)31例,乙肝携带者(As C)18例,正常健康者18例作为对照,采用实时荧光定量PCR的方法检测外周血单个核细胞(PBMC)中CISH、SOCS1和SOCS3 m RNA的表达,ELISA法检测PBMC上清中IL-2、IL-4、IFN-γ、IL-10、IL-17A和TGF-β细胞因子的表达水平,并分析其相关性。结果CHB组患者CISH m RNA表达高于健康对照组(P<0.01),CHB组SOCS3 m RNA表达显著高于健康对照组(P<0.01),较As C组高(P<0.05),SOCS1 m RNA表达量在各组间比较无明显差异(P>0.05);CHB组IL-4的表达量高于As C组,CHB组IFN-γ/IL-4的比值低于健康对照组,As C组IL-4和TGF-β的表达量均高于健康对照组,差异均有统计学意义(P<0.05),CHB组IL-4、IL-17A和TGF-β的表达量均显著高于健康对照组,差异有显著统计学意义(P<0.01);CHB组患者SOCS1 m RNA表达量与CISH m RNA表达量和SOCS3 m RNA表达量之间均存在显著性关联(P=0.004,P=0.002),SOCS3 m RNA表达量与IFN-γ水平之间存在相关性(P=0.037)。结论 CISH和SOCS3基因可能参与慢性乙型肝炎疾病的发生发展,并与CD4+T淋巴细胞亚群变化之间存在一定的相关性。展开更多
目的探讨紫绀型先心病患儿右心室心肌组织中细胞信号抑制因子3(suppressor of cell signaling 3,SOCS3)基因启动子甲基化水平及其可能机制。方法选取先心病患儿34例,其中紫绀组18例,非紫绀组16例。取手术中切除的右心室流出道心肌组织...目的探讨紫绀型先心病患儿右心室心肌组织中细胞信号抑制因子3(suppressor of cell signaling 3,SOCS3)基因启动子甲基化水平及其可能机制。方法选取先心病患儿34例,其中紫绀组18例,非紫绀组16例。取手术中切除的右心室流出道心肌组织作为标本,采用甲基化特异性PCR(methylation specific PCR,MSP)及重亚硫酸盐测序(bisulfite sequencing PCR,BSP)检测心肌细胞中SOCS3启动子CpG岛甲基化程度。Western blot检测DNA甲基化转移酶3A(DNA methyltransferase,DNMT3A)、DNA甲基化转移酶3B(DNMT3B)蛋白在心肌细胞中的表达情况。结果与非紫绀组相比,紫绀组SOCS3启动子CpG岛甲基化程度较高,DNMT3A蛋白较高[(0.407±0.469)vs(0.160±0.034),P<0.05],DNMT3B蛋白无明显差异[(0.054±0.012)vs(0.052±0.093),P>0.05]。结论紫绀型先心病患儿心肌组织中,SOCS3启动子CpG岛呈高甲基化。高表达的DNMT3A蛋白很可能参与了SOCS3启动子CpG岛高甲基化。高甲基化的SOCS3启动子CpG岛,可能不是调控慢性缺氧适应心肌中SOCS3转录的主要因素。高表达的SOCS3启动子CpG岛以及高表达的DNA甲基化转移酶3A蛋白可能是心肌慢性缺氧适应的一种体现。展开更多
文摘Summary： The main pathogenesis of saphenous vein graft neointimal hyperplasia after coronary artery bypass grafting （CABG） is inflammation-caused migration and proliferation of vascular smooth muscle cells （VSMCs）. Janus kinase 2/signal transducer and activators of transcription 3 （JAK2/STAT3） path- way is an important signaling pathway through which VSMCs phenotype conversion occurs. Suppressor of cytokine signaling 3 （SOCS3） is the classic negative feedback inhibitor of JAK2/STAT3 pathway. Growing studies show that SOCS3 plays an important anti-inflammatory role in numerous autoimmune diseases, inflammatory diseases and inflammation-related tumors. However, the effect and mechanism of SOCS3 on vein graft disease is unclear. The purpose of this study was to investigate the effects of SOCS3 on the inflammation, migration and proliferation of VSMCs in vitro and the mechanism. The small interference RNA plasmid targeting rat SOCS3 （SiRNA-rSOCS3） and the recombinant adenovirus vector carrying rat SOCS3 gene （pYrAd-rSOCS3） were constructed, and the empty plamid （SiRNA-control） and vector （pYrAd-GFP） only carrying GFP reported gene were constructed as control. The rat VSMCs were cultured. There were two large groups of A （SOCS3 up-regulated）： control group, IL-6/IFN-γ group, IL-6/IFN-γ＋pYrAd-rSOCS3 group, IL-6/IFN-γ＋pYrAd-GFP group; and B （SOCS3 down-regulated）： control group, IL-6/IFN-γ group, IL-6/IFN-γ＋SiRNA-rSOCS3 group and IL-6/IFN -T＋SiRNA-control group. The pYrAd-rSOCS3 and SiRNA-rSOCS3 were transfected into VSMCs in- duced by IL-6/IFN-γ. After 24 h, real-time reverse transcription polymerase chain reaction （RT-PCR） and Western blotting were used to detect the mRNA and protein expression of SOCS3, STAT3 （only by Western blotting）, P-STAT3 （only by Western blotting）, IL-1β, IL-6, TNF-α, MCP-1 and ICAM-1. The MTT, Transwell assay and flow cytometry were used to examine VSMCs proliferation, migration and cell cycle progression, respectively. As compared with control group, the mRNA and protein expression of SOCS3, STAT3, P-STAT3, IL-1β, IL-6, TNF-α, MCP-1 and ICAM-1 was significantly up-regulated in VSMCs stimulated by IL-6/IFN-γ. However, in VSMCs transfected with pYrAd-rSOCS3 before stimulation with IL-6/IFN-γ, the expression of SOCS3 mRNA and protein was further up-regulated, and that of STAT3, P-STAT3, IL-1β, IL-6, TNF-α, MCP-1 and ICAM-1 was significantly down-regulated as compared with IL-6/IFN-γ group and IL-6/IFN-γ＋pYrAd-GFP group. The expression of those re- lated-cytokines in IL-6/IFN-γ＋SiRNA-rSOCS3 group was markedly increased as compared with IL-6/IFN-γ group and IL-6/IFN-γ＋SiRNA-control group. The absorbance （A） values, the number of cells migrating to the lower chamber, and percentage of cells in the G2/M＋S phase were increased in VSMCs stimulated by IL-6/IFN-γ. In VSMCs incubated with pYrAd-rSOCS3 or SiRNA-rSOCS3 be- fore IL-6/IFN-γ stimulation, the A values, the number of cells migrating to the lower chamber, and the percentage of cells in the G2/M＋S phase were significantly decreased, and increased respectively. These results imply that IL-6/IFN-γ, strong inflammatory stimulators, can promote transformation of VSMCs phenotype form a quiescent contractile state to a synthetic state by activating JAK2/STAT3 pathway. Over-expresssed SOCS3 might inhibit pro-inflammatory effect, migration and growth of VSMCs by blocking STAT3 activation and phosphorylation. These data in vitro confirm that SOCS3 may play a negatively regulatory role in development and progression of vein graft failure. These conclusions can provide a novel strategy for clinical treatment of vein graft diseases and a new theoretic clue for related drug development.
基金supported by grants from the Natural Science Foundation of China (30870983 and 30971118)
文摘BACKGROUND:While suppressor of cytokine signaling 3(SOCS3) plays a crucial role in suppressing dysplasia and tumorigenesis,it also offers a typical instance of DNA methylation in the regulation of gene transcription,since the promoter region of the SOCS3 gene is rich in CpG islands(CGIs).During liver regeneration initiated by partial hepatectomy,SOCS3 acts as a suppressor to balance the acute-phase response and terminate the regeneration.This study aimed to determine whether the variation of SOCS3 expression throughout liver regeneration is also regulated by its DNA methylation.METHODS:We established a 70% partial hepatectomy mouse model and the animals were sacrificed at indicated times to assess the SOCS3 expression.We performed bisulfite sequencing PCR and DNA sequencing to investigate the detailed cytosine methylation in the SOCS3 gene.RESULTS:Within the promoter sequence,58 CGIs were identified and 30 were found variously methylated before or after operation;however,methylation remained at a very low level.No evidence indicated that the total methylation level or the methylation of any CpG site regularly changed throughout liver regeneration.CONCLUSION:DNA methylation or demethylation seems to be a relatively stable modification of cytosine,but not a dynamic and reversible process to regulate gene transcription in daily and acute pathophysiological events.
基金supported by the grants from the National Science Foundation of China Advanced Program(No.NSFC81171558,NSFC81271808 and NSFC81030007)Innovation Team Development Plan of the Ministry of Education of China[No.IRT1131(2011)]National Twelfth-Five Years Project in Science and Technology of China(No.2013ZX10002-003)
文摘Summary： Recently, suppressor of cytokine signaling-3 （SOCS3） has been shown to be an inducible endogenous negative regulator of Janus kinase/signal transducers and activators of transcription （JAK/STAT） pathway which is relevant in inflammatory response, while its functions in acute liver failure and HBV-induced acute-on-chronic liver failure （HBV-ACLF） have not been fully elucidated. In this study, we explored the role of SOCS3 in the development of mouse hepatitis virus strain 3 （MHV-3）-induced acute liver failure and its expression in liver and peripheral blood mononuclear cells （PBMCs） of patients with HBV-ACLF. Inflammation-related gene expression was detected by real-time PCR, immtmohistochemistry and Western blotting. The correlation between SOCS3 level and liver injury was studied. Our results showed that the SOCS3 expression was significantly elevated in both the liver tissue and PBMCs from patients with HBV-ACLF compared to mild chronic hepatitis B （CHB）. Moreover, a time course study showed that SOCS3 level was increased remarkably in the liver of BALB/cJ mice at 72 h post-infection. Pro-inflammatory cytokines, interleukin （IL）-1 β, IL-6, and tumor necrosis factor （TNF）-α, were also increased significantly at 72 h post-infection. There was a close correlation between hepatic SOCS3 level and IL-6, and the severity of liver injury defined by alanine aminotransferase （ALT） and aspartate aminotransferase （AST） levels, respectively. These data suggested that SOCS3 may play a pivotal role in the pathogenesis of MHV-3-induced acute liver failure and HBV-ACLF.
文摘Objective: To study the correlation of suppressor of cytokine signaling 3 (SOCS3) expression in placenta of intrahepatic cholestasis of pregnancy (ICP) with maternal Th cell imbalance and placental trophoblast cell damage. Methods: Primiparae who were diagnosed with ICP in Bazhong Hospital of Traditional Chinese Medicine between June 2014 and August 2017 were selected as the ICP group, and the healthy primiparae who gave birth during the same period were selected as the control group. The placenta was collected after delivery to determine the expression of SOCS3, Th cytokines and apoptosis genes. Results: SOCS3 mRNA expression and protein expression as well as IL-4 and IL-13 protein expression in placenta of ICP group were significantly lower than those of control group whereas IFN-γ, TNF-α, IL-2, p53, Caspase-3, Fas, Caspase-8, Cyt-C and Caspase-9 protein expression were significantly higher than those of control group;IFN-γ, TNF-α, IL-2, p53, Caspase-3, Fas, Caspase-8, Cyt-C and Caspase-9 protein expression in ICP placenta with lower SOCS3 expression were significantly higher than those in ICP placenta with higher SOCS3 expression whereas IL-4 and IL-13 protein expression were significantly lower than those in ICP placenta with higher SOCS3 expression. Conclusion: The low expression of SOCS3 in the ICP placenta can aggravate the imbalance of Th cells and the damage of placental trophoblast cells induced by apoptosis.
文摘目的观察慢性乙型肝炎患者(CHB)CISH、SOCS1和SOCS3 m RNA及CD4+T淋巴细胞亚群相关细胞因子的表达,探讨CISH、SOCS1和SOCS3在慢性乙型肝炎病毒(HBV)感染中的作用。方法选取慢性乙型肝炎患者(CHB)31例,乙肝携带者(As C)18例,正常健康者18例作为对照,采用实时荧光定量PCR的方法检测外周血单个核细胞(PBMC)中CISH、SOCS1和SOCS3 m RNA的表达,ELISA法检测PBMC上清中IL-2、IL-4、IFN-γ、IL-10、IL-17A和TGF-β细胞因子的表达水平,并分析其相关性。结果CHB组患者CISH m RNA表达高于健康对照组(P<0.01),CHB组SOCS3 m RNA表达显著高于健康对照组(P<0.01),较As C组高(P<0.05),SOCS1 m RNA表达量在各组间比较无明显差异(P>0.05);CHB组IL-4的表达量高于As C组,CHB组IFN-γ/IL-4的比值低于健康对照组,As C组IL-4和TGF-β的表达量均高于健康对照组,差异均有统计学意义(P<0.05),CHB组IL-4、IL-17A和TGF-β的表达量均显著高于健康对照组,差异有显著统计学意义(P<0.01);CHB组患者SOCS1 m RNA表达量与CISH m RNA表达量和SOCS3 m RNA表达量之间均存在显著性关联(P=0.004,P=0.002),SOCS3 m RNA表达量与IFN-γ水平之间存在相关性(P=0.037)。结论 CISH和SOCS3基因可能参与慢性乙型肝炎疾病的发生发展,并与CD4+T淋巴细胞亚群变化之间存在一定的相关性。
