BAFF受體TACI融合蛋白的表達純化與功能研究
發(fā)布時間：2018-07-13 19:16
【摘要】：B細胞活化因子(B cell activating factor,BAFF)屬于腫瘤壞死因子超家族的一員。它是外周B細胞存活的重要因子。在動物模型中,BAFF的過表達會導(dǎo)致自身免疫病樣癥狀的發(fā)生。研究發(fā)現(xiàn),在不同類型的自身免疫性疾病患者的血清中,BAFF的含量是明顯升高的。近期研究還發(fā)現(xiàn)BAFF還與部分腫瘤疾病的發(fā)生有關(guān)。因此,BAFF的拮抗劑可能成為治療自身免疫疾病和某些腫瘤的新型藥物。 由于BAFF和受體TACI具有高親和力,可特異性識別BAFF并與之結(jié)合,因此體外構(gòu)建表達的TACI可中和體內(nèi)過表達的BAFF,抑制B細胞的增殖和活化,緩解和治療與BAFF相關(guān)的自身免疫病和B細胞腫瘤。本課題的研究為開發(fā)BAFF拮抗劑治療BAFF相關(guān)疾病提供了實驗基礎(chǔ)。 本課題研究方法:對BAFF受體TACI融合蛋白原核表達與純化,在體外研究TACI對B系淋巴瘤細胞增殖的影響,并初步分析了TACI抑制腫瘤細胞系增殖的機理。 主要研究結(jié)果: (一)TACI融合蛋白原核表達純化:本實驗室已將釣取的TACI基因片段連接到原核表達載體pET-32a上。將重組正確的質(zhì)粒轉(zhuǎn)化大腸桿菌Rosetta,獲得TACI表達工程菌。經(jīng)IPTG誘導(dǎo),融合蛋白利用鎳金屬螯合瓊脂糖凝膠親和層析柱純化。 (二)TACI抑制B系淋巴瘤細胞系細胞的增殖:某些B系淋巴瘤細胞系表達BAFF及其受體,并且通過自分泌BAFF來促進腫瘤細胞的增殖�？扇苄訲ACI蛋白能夠通過特異性阻斷BAFF抑制B系淋巴瘤細胞系的增殖,并且具有劑量依賴性。 (三)TACI抑增殖的機理:TACI能夠?qū)audi細胞阻滯在DNA合成期(即S期) ,而且具有一定的劑量依賴性。TACI對Daudi細胞增殖抑制作用,主要是通過調(diào)控細胞周期,而非凋亡達到抑增殖效果。
[Abstract]:B cell activating factor (B cell activating factor) belongs to the tumor necrosis factor superfamily. It is an important factor in the survival of peripheral B cells. Overexpression of BAFF may lead to autoimmune disease-like symptoms in animal models. Serum BAFF levels were significantly increased in patients with different types of autoimmune diseases. Recent studies have also found that BAFF is also associated with some tumor diseases. Therefore, BAFF antagonist may be a new drug for autoimmune diseases and some tumors. Because BAFF and TACI have high affinity, BAFF can be specifically recognized and combined with BAFF, so TACI can neutralize the overexpression of BAFF in vivo and inhibit the proliferation and activation of B cells. To relieve and treat BAFF associated autoimmune diseases and B cell tumors. This study provides experimental basis for the development of BAFF antagonist in the treatment of BAFF related diseases. Methods: the prokaryotic expression and purification of BAFF receptor TACI fusion protein were studied in vitro to study the effect of TACI on the proliferation of B-lineage lymphoma cells and the mechanism of TACI inhibiting the proliferation of tumor cell lines was preliminarily analyzed. The main results were as follows: (1) expression and purification of TACI fusion protein: the TACI gene fragment was ligated to the prokaryotic expression vector pET-32a in our laboratory. The recombinant plasmid was transformed into E. coli Rosetta to obtain TACI expression engineering bacteria. After induced by IPTG, the fusion protein was purified by nickel metal chelate agarose gel affinity chromatography. (2) TACI inhibits the proliferation of B-lineage lymphoma cell lines: some B-lineage lymphoma cell lines express BAFF and its receptors, and promote the proliferation of tumor cells by autocrine BAFF. Soluble TACI protein can inhibit the proliferation of B line lymphoma cell line by blocking BAFF in a dose-dependent manner. (3) Taci inhibited the proliferation of Daudi cells by blocking Daudi cells in DNA synthesis phase (S phase) in a dose-dependent manner. The inhibitory effect of TACI on the proliferation of Daudi cells was mainly achieved by regulating cell cycle, but not by apoptosis.
【學(xué)位授予單位】：天津大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2010
【分類號】：R392.1
本文編號：2120472
[Abstract]:B cell activating factor (B cell activating factor) belongs to the tumor necrosis factor superfamily. It is an important factor in the survival of peripheral B cells. Overexpression of BAFF may lead to autoimmune disease-like symptoms in animal models. Serum BAFF levels were significantly increased in patients with different types of autoimmune diseases. Recent studies have also found that BAFF is also associated with some tumor diseases. Therefore, BAFF antagonist may be a new drug for autoimmune diseases and some tumors. Because BAFF and TACI have high affinity, BAFF can be specifically recognized and combined with BAFF, so TACI can neutralize the overexpression of BAFF in vivo and inhibit the proliferation and activation of B cells. To relieve and treat BAFF associated autoimmune diseases and B cell tumors. This study provides experimental basis for the development of BAFF antagonist in the treatment of BAFF related diseases. Methods: the prokaryotic expression and purification of BAFF receptor TACI fusion protein were studied in vitro to study the effect of TACI on the proliferation of B-lineage lymphoma cells and the mechanism of TACI inhibiting the proliferation of tumor cell lines was preliminarily analyzed. The main results were as follows: (1) expression and purification of TACI fusion protein: the TACI gene fragment was ligated to the prokaryotic expression vector pET-32a in our laboratory. The recombinant plasmid was transformed into E. coli Rosetta to obtain TACI expression engineering bacteria. After induced by IPTG, the fusion protein was purified by nickel metal chelate agarose gel affinity chromatography. (2) TACI inhibits the proliferation of B-lineage lymphoma cell lines: some B-lineage lymphoma cell lines express BAFF and its receptors, and promote the proliferation of tumor cells by autocrine BAFF. Soluble TACI protein can inhibit the proliferation of B line lymphoma cell line by blocking BAFF in a dose-dependent manner. (3) Taci inhibited the proliferation of Daudi cells by blocking Daudi cells in DNA synthesis phase (S phase) in a dose-dependent manner. The inhibitory effect of TACI on the proliferation of Daudi cells was mainly achieved by regulating cell cycle, but not by apoptosis.
【學(xué)位授予單位】：天津大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2010
【分類號】：R392.1
【共引文獻】
相關(guān)期刊論文 前2條
1 于孟學(xué);曹金;李薇;;生物制劑治療系統(tǒng)性紅斑狼瘡的新進展[J];北京醫(yī)學(xué);2008年01期
2 顏克香;項蕾紅;鄭志忠;Stuart Mardim;;2010年后皮膚科的主要治療進展[J];中國中西醫(yī)結(jié)合皮膚性病學(xué)雜志;2011年01期
相關(guān)碩士學(xué)位論文 前1條
1 張小丹;TACI-Ig對ConA活化淋巴細胞的染色質(zhì)誘導(dǎo)SLE樣小鼠模型的治療作用[D];安徽醫(yī)科大學(xué);2011年
，本文編號：2120472
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