發(fā)布時(shí)間：2019-07-05 05:14

【摘要】：目的：探討表達(dá)IDO的KC在體內(nèi)外對同種異體T淋巴細(xì)胞增殖的抑制和凋亡作用。 方法：聯(lián)合鏈酶蛋白酶和膠原酶在體灌注和離體消化的方法分離小鼠KC，實(shí)時(shí)熒光定量PCR檢測經(jīng)IFN-γ處理或未處理的KC表達(dá)IDOmRNA和FasLmRNA的情況。高壓液相色譜儀分析IDO對色氨酸的分解作用以了解其活性。~3H嵌入增殖試驗(yàn)檢測KC對同種異體T淋巴細(xì)胞增殖的抑制情況。流式細(xì)胞儀分析KC對同種異體T淋巴細(xì)胞的細(xì)胞周期和凋亡作用。構(gòu)建BABL／c→C57BL／6的皮膚移植模型，分為KC(未處理)、KC(IFN-γ,處理)、KC(1—MT)及生理鹽水組，除了生理鹽水組外，，于移植術(shù)前14、7、2天給受體輸入供體KC，同時(shí)1—MT組于移植術(shù)后0～7天經(jīng)腹腔注射1—MT(0.9mg／kg／d)，觀察各組別對皮膚移植物存活時(shí)間的影響。于移植術(shù)后第7天每組各取1只行小鼠皮瓣HE染色和TUNEL以檢測淋巴細(xì)胞浸潤和凋亡情況。Kaplan-Meier對數(shù)秩檢驗(yàn)對各組進(jìn)行生存分析。 結(jié)果：實(shí)時(shí)熒光定量PCR提示KC并不組成性表達(dá)IDO，但是經(jīng)IFN-γ處理后，IDO和FasL在6h后表達(dá)量相對較高。IDO能降低培養(yǎng)基中色氨酸的濃度，升高其代謝產(chǎn)物犬尿氨酸濃度。表達(dá)IDO的KC能明顯抑制同種異體T淋巴細(xì)胞的增殖，但1-MT和抗FasL抗體能部分阻斷其增殖抑制作用，并且存在劑量依賴關(guān)系。表達(dá)IDO和FasL的KC能阻止同種異體T淋巴細(xì)胞于G1中期，誘導(dǎo)其凋亡。輸入表達(dá)IDO和FasL的
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圖片說明：培養(yǎng)24h后KC的形態(tài)X100
[Abstract]:Aim: to investigate the inhibitory effect of KC expressing IDO on the proliferation and apoptosis of allogenic T lymphocytes in vitro and in vivo. Methods: the expression of IDOmRNA and FasLmRNA in mouse KC, treated or untreated KC was detected by real-time fluorescence quantitative PCR combined with streptozyme protease and collagenase in vivo perfusion and in vitro digestion. The decomposition effect of IDO on tryptophan was analyzed by high pressure liquid chromatography (HPLC) to understand its activity. The inhibitory effect of KC on the proliferation of allogenic T lymphocytes was detected by ~ 3 H embedding proliferation test. The cell cycle and apoptosis of allogenic T lymphocytes were analyzed by flow cytometry. The skin transplantation model of BABL/c 鈮

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