SOCS3在小鼠肝臟纖維化進(jìn)展和逆轉(zhuǎn)過程中對(duì)巨噬細(xì)胞極化的影響
發(fā)布時(shí)間：2018-10-31 13:47
【摘要】：近些年,肝臟疾病的發(fā)病率在逐年增高,眾多損傷因素如酒精、病毒、化學(xué)物質(zhì)等存在下,肝臟將發(fā)生不同程度的損傷,如病毒性肝炎、酒精肝、肝纖維化、肝硬化、肝癌等肝臟疾病的發(fā)生。而肝臟作為人體重要的代謝器官,其自身器官不同程度的損傷直接影響著機(jī)體其他疾病的發(fā)生發(fā)展。因此,研究肝臟疾病的發(fā)生發(fā)展和尋找其治愈肝臟疾病的方法方式已刻不容緩。Kupffer細(xì)胞(KCs)是存在于肝臟竇狀隙內(nèi)的一類非實(shí)質(zhì)細(xì)胞,屬于定居肝臟內(nèi)的固有細(xì)胞,KCs即能夠非特異的吞噬和清除血液中的細(xì)菌、外源性異物等抗原性物質(zhì),還具有特異性的免疫應(yīng)答、抗腫瘤免疫反應(yīng)、內(nèi)毒素解毒、抗感染、調(diào)節(jié)微循環(huán)及物質(zhì)代謝等方面的作用。在肝臟不同的病理生理狀態(tài)下,KCs能夠極化成不同的亞型,即M1(classically activated macrophages)型和M2(alternatively activated macrophages)型,M1型巨噬細(xì)胞主要能夠促進(jìn)炎癥的發(fā)生發(fā)展,分泌產(chǎn)生TNF-α(tumor necrosis factor alpha)、IL-1(interleukin 1)、IL-6(interleukin 6)等促炎細(xì)胞因子;M2型巨噬細(xì)胞其主要功能是抑制炎癥的發(fā)生發(fā)展和對(duì)組織的修復(fù)產(chǎn)生作用,通過合成和分泌抗炎細(xì)胞因子IL-10(interleukin 10)以及通過Arg-1(arginase 1)將精氨酸分解成聚胺,來參與合成和穩(wěn)定細(xì)胞外基質(zhì)。巨噬細(xì)胞通過M1型和M2型的動(dòng)態(tài)調(diào)控來影響肝臟的病理生理狀況,因此,通過上述現(xiàn)象來調(diào)控肝臟巨細(xì)胞的極化從而來達(dá)到肝臟疾病的恢復(fù)這一研究思路將為肝臟疾病的治療尋找新的靶點(diǎn)和策略。已有研究表明SOCS3(Suppressor of cytokine signaling 3)參與肝臟疾病的發(fā)生發(fā)展,通過影響多種細(xì)胞因子的產(chǎn)生和信號(hào)傳導(dǎo)進(jìn)行負(fù)性調(diào)控,如通過免疫調(diào)控來影響炎癥因子和趨化因子進(jìn)而去調(diào)控炎癥反應(yīng),而肝臟疾病的發(fā)生發(fā)展有巨噬細(xì)胞的參與,巨噬細(xì)胞與炎癥的發(fā)生發(fā)展密切相關(guān),設(shè)想是否SOCS3參與肝纖維化的發(fā)生發(fā)展,此外,另有報(bào)道顯示,肝纖維化在去除損傷因素后能夠發(fā)生逆轉(zhuǎn)[1],進(jìn)而,通過調(diào)控SOCS3蛋白的表達(dá)去影響肝纖維化的逆轉(zhuǎn)過程和肝臟巨噬細(xì)胞極化分型過程從而探索其對(duì)肝纖維化的影響。且課題組前期研究發(fā)現(xiàn)SOCS3在動(dòng)物肝纖維化和逆轉(zhuǎn)過程中存在著表達(dá)的變化,且在肝臟巨噬細(xì)胞中表達(dá),但SOCS3在肝纖維化中的調(diào)控作用,尤其是對(duì)肝巨噬細(xì)胞極化分型相關(guān)作用尚未報(bào)道,于是本實(shí)驗(yàn)室重點(diǎn)研究通過調(diào)控SOCS3的表達(dá)觀察對(duì)KCs極化分型的影響進(jìn)而為肝纖維化的防治提供新的思路。主要研究?jī)?nèi)容概括如下:1.SOCS3在小鼠肝臟纖維化進(jìn)展和逆轉(zhuǎn)過程中的表達(dá)。取肝臟組織,利用免疫熒光雙染和免疫組織化學(xué)的方法觀察SOCS3在肝臟組織的的表達(dá)情況。結(jié)果發(fā)現(xiàn):SOCS3在小鼠肝臟巨噬細(xì)胞中表達(dá),并且,隨著小鼠肝纖維化的形成,SOCS3的表達(dá)逐漸增多,隨著小鼠肝纖維化的逆轉(zhuǎn)的進(jìn)行,SOCS3的表達(dá)逐漸減少。2.小鼠肝纖維化進(jìn)展和逆轉(zhuǎn)過程中肝臟巨噬細(xì)胞表型的變化。通過原位灌流技術(shù)提取不同時(shí)期小鼠肝臟巨噬細(xì)胞,利用流式細(xì)胞技術(shù)對(duì)提取的肝臟巨噬細(xì)胞進(jìn)行分型研究。結(jié)果發(fā)現(xiàn):隨著小鼠肝纖維化的形成,CD86標(biāo)記的M1型巨噬細(xì)胞表達(dá)逐漸增多,隨著小鼠肝纖維化的逆轉(zhuǎn),CD206標(biāo)記的M2型巨噬細(xì)胞逐漸增多。3.SOCS3在體外RAW264.7細(xì)胞中通過刺激因子作用在巨噬細(xì)胞分型中的表達(dá)。體外通過RAW264.7細(xì)胞通過LPS和IFN-γ刺激形成M1型,IL-4刺激形成M2型,觀察SOCS3在不同分型中的表達(dá)。結(jié)果發(fā)現(xiàn):SOCS3相對(duì)于M2型,在M1型表達(dá)量顯著增高。4.干擾SOCS3的表達(dá)對(duì)RAW264.7細(xì)胞極化分型的影響。LipofectamineTM2000轉(zhuǎn)染SOCS3-si RNA到RAW264.7細(xì)胞內(nèi),可使LPS(Lipopolysaccharides)和IFN-γ(Interferonγ)刺激形成M1型表達(dá)顯著增多,LipofectamineTM2000轉(zhuǎn)LV3-SOCS3-mus-1225到RAW264.7細(xì)胞內(nèi),可使IL-4(interleukin 4)刺激形成M2型表達(dá)顯著增多。
[Abstract]:In recent years, the incidence of liver disease is increasing year by year. In the presence of many damage factors such as alcohol, virus, chemical substance and so on, the liver will have different degrees of damage, such as viral hepatitis, alcohol liver, liver fibrosis, liver cirrhosis, liver cancer and other liver diseases. Liver, as an important metabolic organ of human body, directly affects the development of other diseases in the body. Therefore, it is urgent to study the development of liver diseases and to find ways to cure liver diseases. Kruper cell (KCs) is a kind of non-essential cell which exists in the liver allogenic gap, and belongs to the intrinsic cell in the liver of the sedentary liver. KCs is an antigenic substance capable of non-specific phagocytizing and clearing bacteria in blood, exogenous foreign matter and the like, and also has specific immune response and anti-tumor immune response. Endotoxins detoxification, anti-infection, regulation of microcirculation and substance metabolism. In different pathophysiological states of the liver, KCs can be polarized into different subtypes, M1 (classically activated macimages) and M2 (alternate activated macimages), and M1-type macrophages can promote the development of inflammation, and produce pro-inflammatory cytokines such as TNF-CoV (tubetrosis factor alpha), IL-1 (interleukin1) and IL-6 (interleukin 6); The main function of M2-type macrophages is to inhibit the development of inflammation and to play a role in the repair of tissues, and to participate in the synthesis and stabilization of extracellular matrix by synthesizing and secreting anti-inflammatory cytokines IL-10 (interleukin 10) and by decomposing arginine into polyamines by Arg-1 (arginiase 1). Macrophages regulate the pathological and physiological condition of liver by the dynamic regulation of M1 and M2. Therefore, it can regulate the polarization of the giant cell of the liver by the phenomenon, so as to achieve the restoration of liver disease. Previous studies have shown that SOCS3 (Suppressor of Cytokine signaling 3) is involved in the development of liver diseases, and negatively regulates the production and signaling of various cytokines, such as by regulating the immune regulation to affect the inflammatory factors and chemokine, and then to regulate the inflammatory response. The development of liver disease has the involvement of macrophages. Macrophages are closely related to the development of inflammation. It is envisaged that SOCS3 is involved in the development of liver fibrosis. In addition, another report shows that liver fibrosis can be reversed after removing the damage factors[1], and further, The effect of SOCS3 protein on hepatic fibrosis was studied by regulating the expression of SOCS3 protein to affect the reverse course of hepatic fibrosis and the polarization classification of liver macrophages. The results showed that SOCS3 was expressed in liver fibrosis and liver fibrosis, but the regulation of SOCS3 in liver fibrosis, especially the effect of SOCS3 in hepatic fibrosis, was not reported. Therefore, this laboratory focused on the influence of the expression of SOCS3 on the polarization typing of KCs, thus providing a new idea for the prevention and treatment of hepatic fibrosis. The main contents of this study were as follows: 1. SOCS3 was expressed in the progression and reversal of liver fibrosis in mice. Liver tissue was taken to observe the expression of SOCS3 in liver tissue by immunofluorescent double staining and immunohistochemistry. The results showed that SOCS3 was expressed in mouse liver macrophages, and the expression of SOCS3 increased with the formation of liver fibrosis in mice. The expression of SOCS3 decreased gradually with the reversal of liver fibrosis in mice. Changes of liver macrophage phenotype during the progression and reversal of liver fibrosis in mice. Rat liver macrophages were extracted by in situ perfusion technique, and the extracted liver macrophages were classified by flow cytometry. Results: With the formation of liver fibrosis in mice, the expression of CD86-labeled M1-type macrophages was gradually increased. With the reversal of liver fibrosis in mice, the M2-type macrophages labeled with CD206 were gradually increased. 3. SOCS3 was expressed in RAW264.7 cells in vitro by stimulating factor. The expression of SOCS3 in different types was observed by means of RAW264.7 cells stimulated by LPS and IFN-OVA to form M1 type, IL-4 stimulated to form M2 type. The results showed that the expression level of SOCS3 was significantly higher than that of M2 type. The effect of interfering SOCS3 expression on the polarization typing of RAW264.7 cells. With the transfection of SOCS3-si RNA into RAW264.7 cells by Lipofectinine TM2000, the expression of LPS (LPS) and IFN-jun (Interfering RNAs) could be increased significantly, and the expression of IL-4 (interleukin4) could be increased significantly in the cells of LV3-SOCS3-mus-1225 to RAW264.7. The expression of IL-4 (interleukin4) could be increased significantly.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R575.2
本文編號(hào)：2302422
[Abstract]:In recent years, the incidence of liver disease is increasing year by year. In the presence of many damage factors such as alcohol, virus, chemical substance and so on, the liver will have different degrees of damage, such as viral hepatitis, alcohol liver, liver fibrosis, liver cirrhosis, liver cancer and other liver diseases. Liver, as an important metabolic organ of human body, directly affects the development of other diseases in the body. Therefore, it is urgent to study the development of liver diseases and to find ways to cure liver diseases. Kruper cell (KCs) is a kind of non-essential cell which exists in the liver allogenic gap, and belongs to the intrinsic cell in the liver of the sedentary liver. KCs is an antigenic substance capable of non-specific phagocytizing and clearing bacteria in blood, exogenous foreign matter and the like, and also has specific immune response and anti-tumor immune response. Endotoxins detoxification, anti-infection, regulation of microcirculation and substance metabolism. In different pathophysiological states of the liver, KCs can be polarized into different subtypes, M1 (classically activated macimages) and M2 (alternate activated macimages), and M1-type macrophages can promote the development of inflammation, and produce pro-inflammatory cytokines such as TNF-CoV (tubetrosis factor alpha), IL-1 (interleukin1) and IL-6 (interleukin 6); The main function of M2-type macrophages is to inhibit the development of inflammation and to play a role in the repair of tissues, and to participate in the synthesis and stabilization of extracellular matrix by synthesizing and secreting anti-inflammatory cytokines IL-10 (interleukin 10) and by decomposing arginine into polyamines by Arg-1 (arginiase 1). Macrophages regulate the pathological and physiological condition of liver by the dynamic regulation of M1 and M2. Therefore, it can regulate the polarization of the giant cell of the liver by the phenomenon, so as to achieve the restoration of liver disease. Previous studies have shown that SOCS3 (Suppressor of Cytokine signaling 3) is involved in the development of liver diseases, and negatively regulates the production and signaling of various cytokines, such as by regulating the immune regulation to affect the inflammatory factors and chemokine, and then to regulate the inflammatory response. The development of liver disease has the involvement of macrophages. Macrophages are closely related to the development of inflammation. It is envisaged that SOCS3 is involved in the development of liver fibrosis. In addition, another report shows that liver fibrosis can be reversed after removing the damage factors[1], and further, The effect of SOCS3 protein on hepatic fibrosis was studied by regulating the expression of SOCS3 protein to affect the reverse course of hepatic fibrosis and the polarization classification of liver macrophages. The results showed that SOCS3 was expressed in liver fibrosis and liver fibrosis, but the regulation of SOCS3 in liver fibrosis, especially the effect of SOCS3 in hepatic fibrosis, was not reported. Therefore, this laboratory focused on the influence of the expression of SOCS3 on the polarization typing of KCs, thus providing a new idea for the prevention and treatment of hepatic fibrosis. The main contents of this study were as follows: 1. SOCS3 was expressed in the progression and reversal of liver fibrosis in mice. Liver tissue was taken to observe the expression of SOCS3 in liver tissue by immunofluorescent double staining and immunohistochemistry. The results showed that SOCS3 was expressed in mouse liver macrophages, and the expression of SOCS3 increased with the formation of liver fibrosis in mice. The expression of SOCS3 decreased gradually with the reversal of liver fibrosis in mice. Changes of liver macrophage phenotype during the progression and reversal of liver fibrosis in mice. Rat liver macrophages were extracted by in situ perfusion technique, and the extracted liver macrophages were classified by flow cytometry. Results: With the formation of liver fibrosis in mice, the expression of CD86-labeled M1-type macrophages was gradually increased. With the reversal of liver fibrosis in mice, the M2-type macrophages labeled with CD206 were gradually increased. 3. SOCS3 was expressed in RAW264.7 cells in vitro by stimulating factor. The expression of SOCS3 in different types was observed by means of RAW264.7 cells stimulated by LPS and IFN-OVA to form M1 type, IL-4 stimulated to form M2 type. The results showed that the expression level of SOCS3 was significantly higher than that of M2 type. The effect of interfering SOCS3 expression on the polarization typing of RAW264.7 cells. With the transfection of SOCS3-si RNA into RAW264.7 cells by Lipofectinine TM2000, the expression of LPS (LPS) and IFN-jun (Interfering RNAs) could be increased significantly, and the expression of IL-4 (interleukin4) could be increased significantly in the cells of LV3-SOCS3-mus-1225 to RAW264.7. The expression of IL-4 (interleukin4) could be increased significantly.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R575.2
【參考文獻(xiàn)】
相關(guān)期刊論文 前2條
1 沈竹夏;段勝仲;;巨噬細(xì)胞極化在2型糖尿病和心血管疾病中的作用[J];生命科學(xué);2013年02期
2 孔德松;鄭仕中;陸茵;王愛云;;肝內(nèi)肌成纖維細(xì)胞的來源及其在肝纖維化中作用的研究[J];中國(guó)藥理學(xué)通報(bào);2011年03期
，本文編號(hào)：2302422
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