本文關(guān)鍵詞：日本血吸蟲可溶性蟲卵抗原體外誘導(dǎo)巨噬細胞分泌TNF-α活化肝星狀細胞及黃芪總苷對其影響　出處：《安徽醫(yī)科大學》2007年碩士論文　論文類型：學位論文

  更多相關(guān)文章： 黃芪總苷 肝纖維化 血吸蟲 可溶性蟲卵抗原 巨噬細胞條件培養(yǎng)基 肝星狀細胞 腫瘤壞死因子-α

【摘要】： 血吸蟲病是一種在熱帶或亞熱帶國家和地區(qū)廣泛傳播并嚴重危害人類健康的寄生蟲病,肝纖維化是其嚴重的病變后果。纖維化的形成是通過巨噬細胞、淋巴細胞、纖維細胞和其它細胞產(chǎn)生的細胞因子動態(tài)調(diào)節(jié)的過程[1.2]。TNF-α作為重要的炎癥介質(zhì)可直接參與肝纖維化中的關(guān)鍵細胞肝星狀細胞(hepatic stellate cell, HSC)的活化,促進其增殖。黃芪總苷(astrogalosides, AST)是黃芪抗肝纖維化的主要成份,研究已證實AST具有顯著的抗炎和抗肝纖維化等作用。本文觀察日本血吸蟲可溶性蟲卵抗原(soluble egg antigen, SEA)體外刺激巨噬細胞后,TNF-α表達和分泌的改變以及SEA活化的腹腔巨噬細胞條件培養(yǎng)基(soluble egg antigen induced macrophage conditioned medium, SEA-MCM)對HSC增殖刺激膠原合成的影響,并在此基礎(chǔ)上,觀察AST對其的作用。 1 SEA誘導(dǎo)大鼠腹腔巨噬細胞活化和TNF-α表達及AST對其影響 制備SEA(0.17g·L-1),采用HE染色、免疫細胞化學染色、逆轉(zhuǎn)錄聚合酶鏈反應(yīng)(RT-PCR)技術(shù)及放射免疫法,觀察在SEA(10 mg·L-1)的刺激下,AST與大鼠腹腔巨噬細胞(peritoneal macrophage, PMФ)在體外共育3h、6h和9h后,大鼠PMФs形態(tài)改變及對TNF-α表達的影響。結(jié)果顯示:(1)SEA(10 mg·L-1)作用3h、6h和9h可明顯刺激PMΦ活化和增加TNF-α的表達(;2)AST(10,20 mg·L-1)能明顯降低SEA(10 mg·L-1)刺激的PMΦTNF-α的表達。提示SEA體外可以活化PMΦ,使其增加TNF-α的表達,AST可抑制這種TNF-α的表達。 2.SEA-MCM誘導(dǎo)HSC活化及AST對其抑制作用 大鼠腹腔注射SEA(0.17mg·ml-1,0.2ml/只),制備SEA-MCM。利用肝星狀細胞株HSC-T6細胞,采用MTT和3H-脯氨酸摻入法,觀察SEA-MCM誘導(dǎo)HSC-T6細胞增殖和膠原合成的作用及AST對其影響。結(jié)果顯示:(1)不同稀釋度的SEA-MCM(終稀釋比1:2、1:4、1:8、1:16)均可明顯促進HSC-T6細胞增殖和膠原合成;(2)AST(10和20 mg·L-1)呈濃度依賴性抑制SEA-MCM(1:4)刺激的HSC-T6細胞增殖和膠原合成。表明SEA可以通過活化的巨噬細胞促進HSC增殖和膠原合成,這種作用可被AST抑制。 3.抗TNF-α抗體對SEA-MCM誘導(dǎo)的HSC-T6細胞增殖和膠原合成影響 在SEA-MCM(1:4)中加入不同稀釋度TNF-α抗體,采用MTT、免疫印記(Western blotting)和放射免疫法檢測SEA-MCM對HSC-T6細胞增殖和膠原合成的作用。結(jié)果顯示:TNF-α抗體(1:100和1:200)均可明顯抑制SEA-MCM對HSC-T6細胞的增殖和膠原合成作用,且呈濃度依賴性趨勢。表明SEA通過巨噬細胞分泌TNF-α促進HSC的增殖和產(chǎn)生膠原可能是日本血吸蟲性肝纖維化的重要機制之一。
[Abstract]:Schistosomiasis is a parasitic disease widely spread in tropical or subtropical countries and regions and seriously endangers human health. Hepatic fibrosis is the result of its serious pathological changes. The formation of fibrosis is through macrophages. The process of dynamically regulating cytokines produced by lymphocytes, fibroblasts, and other cells. [1. TNF- 偽, as an important inflammatory medium, can be directly involved in hepatic stellate cell (HSC) hepatic stellate cell in hepatic fibrosis. Astragaloside astrogalosides (AST) is the main anti-hepatic fibrosis component of Astragalus membranaceus. It has been proved that AST has significant anti-inflammatory and anti-hepatic fibrosis effects. The soluble egg antigen of Schistosoma japonicum soluble egg antigen was observed. After stimulation of macrophages in vitro by sea. Changes in expression and secretion of TNF- 偽 and conditioned medium for SEA activated peritoneal macrophages. Soluble egg antigen induced macrophage conditioned medium. On the basis of the effects of SEA-MCM on collagen synthesis stimulated by HSC proliferation, the effects of AST on collagen synthesis were observed. 1 SEA induced activation and expression of TNF- 偽 in rat peritoneal macrophages and effects of AST on it SEA(0.17g 路L -1 was prepared by HE staining, immunocytochemical staining, reverse transcriptase polymerase chain reaction (RT PCR) and radioimmunoassay. The co-culture of SEA(10 with peritoneal macrophage (PM) in vitro for 3 h was observed. After 6 and 9 hours, the morphologic changes of PM and its effect on the expression of TNF- 偽 were observed in rats. The results showed that 10 mg 路L -1 of SEAA (10 mg 路L ~ (-1)) was treated with 10 mg 路L ~ (-1) of TNF- 偽 for 3 h. The activation of PM 桅 and the expression of TNF- 偽 were significantly stimulated at 6h and 9h. 2the expression of PM 桅 TNF- 偽 stimulated by SEA(10 mg 路L ~ (-1) was significantly decreased by SEA (20 mg 路L ~ (-1)), suggesting that SEA could activate PM 桅 in vitro. By increasing the expression of TNF- 偽, AST could inhibit the expression of TNF- 偽. 2. SEA-MCM induces HSC activation and AST inhibits it. SEA-MCM was prepared by intraperitoneal injection of SEA(0.17mg 路ml-1 (0.2 ml / rat). Hepatic stellate cell line HSC-T6 cells were used. MTT and 3H-proline incorporation were used. The effects of SEA-MCM on the proliferation and collagen synthesis of HSC-T6 cells and the effects of AST on them were observed. The results showed that the final dilution ratio of SEA-MCMs with different dilution was 1: 2. 1: 4 + 1: 1: 1: 16) could significantly promote the proliferation and collagen synthesis of HSC-T6 cells. AST 10 and 20 mg 路L -1) inhibited SEA-MCM 1: 4 in a concentration-dependent manner. It is suggested that SEA can promote HSC proliferation and collagen synthesis through activated macrophages. This effect can be inhibited by AST. 3. Effects of anti-TNF- 偽 antibody on proliferation and collagen synthesis of HSC-T6 cells induced by SEA-MCM TNF- 偽 antibody with different dilution was added to SEA-MCM-1: 4) and MTT was used. Western blotting and radioimmunoassay were used to detect the effects of SEA-MCM on the proliferation and collagen synthesis of HSC-T6 cells. 1: 100 and 1: 200) could significantly inhibit the proliferation and collagen synthesis of HSC-T6 cells induced by SEA-MCM. The results indicated that SEA could promote the proliferation of HSC and produce collagen through macrophage secreting TNF- 偽, which might be one of the important mechanisms of schistosomiasis hepatic fibrosis.
【學位授予單位】：安徽醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2007
【分類號】：R285.5;R392

【引證文獻】

相關(guān)期刊論文 前1條

1 田新雁;姜北;;中藥黃芪防治寄生蟲病研究進展[J];醫(yī)學綜述;2013年07期

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本文編號：1397495