本文選題：熱休克預處理　切入點：HSF1　出處：《中南大學》2009年碩士論文

【摘要】： 細胞遷移發(fā)生于胚胎形成、機體免疫、急慢性炎癥及腫瘤等多種生理及病理過程中,是炎癥過程中必不可少的環(huán)節(jié)之一。炎癥發(fā)生時,白細胞可以通過血管壁滲出到血管外并在局部組織中聚集,這個過程稱為白細胞浸潤。單核/巨噬細胞通常是到達浸潤部位的主要白細胞之一。白細胞浸潤是一個極為復雜的連續(xù)過程,主要由白細胞的三個生物學特性決定:黏附作用、趨化作用、變形作用。其中趨化作用是白細胞參與炎癥反應的重要前驅反應。 熱休克反應(HSR)是機體最重要的保護機制之一。熱休克因子1(HSF1)通過誘導熱休克蛋白(HSPs)的表達,在細胞內源性保護中發(fā)揮重要作用。近年國內外研究發(fā)現,HSF1及HSPs也參與了對小鼠胚胎成纖維細胞及人胚胎腎細胞遷移的調節(jié)過程。這為我們研究HSR及HSF1調控細胞遷移提供了重要信息。然而,HSR是否可以調控炎癥中重要細胞之一的巨噬細胞的遷移?HSF1是否及如何參與該調控過程?目前尚無相關研究。 為探討熱休克預處理對脂多糖(LPS)所致巨噬細胞遷移及黏附的影響,本研究首先采用腹腔注射大腸桿菌LPS的小鼠模型,通過加熱使小鼠肛溫升至42℃并維持15min制備熱休克預處理模型,采用支氣管肺泡灌洗觀察熱休克預處理對LPS所致小鼠肺泡白細胞滲出的影響;并采用LPS體外刺激的小鼠腹腔巨噬細胞及RAW264.7巨噬細胞株為模型,應用趨化實驗、劃痕愈合實驗及黏附實驗觀察熱休克預處理(42℃,1h)對LPS所致巨噬細胞遷移的影響;為探討HSF1在熱休克預處理對LPS所致巨噬細胞遷移的影響中是否發(fā)揮作用,采用HSF1基因敲除小鼠制備內毒素血癥模型,應用免疫組化技術觀察HSF1基因敲除小鼠肺、肝組織形態(tài)學變化及巨噬細胞浸潤情況;采用HSF1敲除小鼠腹腔巨噬細胞,應用趨化實驗觀察HSF1敲除對LPS所致巨噬細胞遷移的影響;并采用HSF1過表達的RAW264.7巨噬細胞株,應用趨化實驗觀察HSF1過表達對LPS所致巨噬細胞遷移的影響。 結果發(fā)現:(1).LPS(12mg/kg,2h)腹腔注射促進小鼠血清中TNF-α、IL-6、IL-10的釋放;LPS(600ng/ml,2h)促進RAW264.7巨噬細胞株TNF-α的釋放。(2).腹腔注射LPS(12mg/kg,48h)導致小鼠肺泡灌洗液中白細胞總數及巨噬細胞數均明顯增加,熱休克預處理抑制了LPS所致小鼠白細胞及巨噬細胞向肺泡的遷移;采用體外趨化實驗檢測LPS(600ng/ml,24h)刺激RAW264.7巨噬細胞株及小鼠腹腔巨噬細胞,發(fā)現兩種細胞的遷移均明顯增加,而熱休克預處理能抑制LPS所致的巨噬細胞遷移;LPS處理(600ng/ml,24h)促進兩種巨噬細胞向劃痕部位的遷移而熱休克預處理可抑制LPS對巨噬細胞遷移的促進作用;LPS處理(600ng/ml,1h)可使上述兩種巨噬細胞與細胞外基質的黏附增加,而熱休克預處理可抑制LPS對巨噬細胞黏附的促進作用。(3).HSF1基因敲除廢除了熱休克預處理對LPS所致的小鼠肺、肝組織巨噬細胞浸潤及組織損傷的保護作用,也廢除了熱休克預處理對LPS所致小鼠腹腔巨噬細胞遷移的抑制作用。(4).HSF1基因過表達無需進行熱休克預處理即能抑制LPS所致的RAW264.7巨噬細胞遷移,而熱休克處理并不能進一步增強這種抑制作用。 綜上所述,LPS在整體小鼠和離體巨噬細胞模型均能促進巨噬細胞的遷移;熱休克預處理能抑制LPS所致的小鼠肺、肝組織損傷和巨噬細胞浸潤以及LPS所致離體巨噬細胞的遷移;HSF1參與了熱休克預處理對LPS所致巨噬細胞遷移的抑制作用。
[Abstract]:cell migration occurs in many physiological and pathological processes , such as embryonic development , organism immunity , acute and chronic inflammation and tumor , and is one of the necessary links in the process of inflammation .



Heat shock factor ( HSR ) is one of the most important protective mechanisms of HSR , which plays an important role in the endogenous protection of human embryonic kidney cells by inducing the expression of heat shock protein ( HSPs ) . In recent years , it has been found that HSFs and HSPs also participate in the regulation of the migration of mouse embryonic fibroblasts and human embryonic kidney cells .



In order to investigate the effects of heat shock pretreatment on the migration and adhesion of LPS - induced macrophages in mice , the effects of heat shock pretreatment on LPS - induced macrophage migration were studied by means of heat shock pretreatment .



The results showed that : ( 1 ) LPS ( 12 mg / kg , 2h ) could promote the release of TNF - 偽 , IL - 6 and IL - 10 in serum of mice . LPS ( 600ng / ml , 2h ) promoted the release of TNF - 偽 in RAW264.7 macrophage . LPS ( 600ng / ml , 24h ) stimulated the migration of macrophages and macrophages . LPS treatment ( 600ng / ml , 24h ) promoted the migration of two kinds of macrophages to scratch sites . LPS treatment ( 600ng / ml , 1h ) promoted the adhesion of two kinds of macrophages to the extracellular matrix . The effect of heat shock pretreatment on LPS - induced infiltration and tissue injury of mouse lung and liver tissue was abolished by HSFI , and the inhibitory effect of heat shock pretreatment on peritoneal macrophage migration in mice induced by LPS was also abolished . It is not necessary to perform heat shock pretreatment to inhibit the migration of RAW264.7 macrophages due to LPS , but the heat shock treatment can not further enhance the inhibition .



In conclusion , LPS can promote the migration of macrophages in both the whole mouse and the ex vivo macrophage model . Heat shock pretreatment can inhibit the migration of LPS - induced lung , liver tissue injury and macrophage infiltration and LPS - induced dissociation of macrophages .

【學位授予單位】：中南大學
【學位級別】：碩士
【學位授予年份】：2009
【分類號】：R363

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相關期刊論文 前1條

1 呂運成;王佐;危當恒;姜志勝;萬煒;李國華;童中藝;王貴學;;基質細胞衍生因子1α—CXCR4趨化THP-1細胞遷移及氧化型低密度脂蛋白的影響[J];中國動脈硬化雜志;2007年01期

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