中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2012年
10期
1927-1929
,共3页
树突状细胞%小鼠甲胎蛋白%4-1BB配体%生物学特性
樹突狀細胞%小鼠甲胎蛋白%4-1BB配體%生物學特性
수돌상세포%소서갑태단백%4-1BB배체%생물학특성
Dendritic cells%Mouse alpha-fetoprotein%4-1BBL gene%Biology characteristics
目的 观察感染小鼠甲胎蛋白(mAFP)和4-1BB配体(m4-1BBL)融合基因对小鼠树突状细胞(DC)生物学特性的影响.方法 构建含mAFP和m4-1BBL的融合基因腺病毒Ad-mAFP-内部核糖体起始位点(IRES)-m4-1BBL,感染H22肝癌细胞,Western blot法检测mAFP和4-1BBL蛋白的表达；诱导培养DC,并将其分为实验组、载体对照组和阴性对照组,以上各组感染后观察DC形态学变化；流式细胞仪检测各组细胞表型；混合淋巴细胞反应(MLR)检测各组DC的刺激活性以及对T细胞分泌干扰素(IFN)-γ的影响.结果 成功构建融合基因重组腺病毒Ad-mAFP-IRES-m4-1BBL并表达于H22细胞；体外诱导培养获得典型的DC；流式细胞仪检测各组DC感染后的细胞表型,实验组DC细胞CD80[(60.7±8.7)％]、CD86[(67.0±4.6)％]表达率显著增高于其余两组(P＜0.05),刺激T细胞增殖能力(0.66±0.05)亦显著增强(P＜0.05),刺激T细胞分泌IFN-γ[(1280.7 ±68.2) ng/L]水平更高(P＜0.05).结论 成功构建含mAFP及4-1BBL融合基因腺病毒疫苗,其感染DC后刺激T细胞增殖和分泌IFN-γ的能力增强.
目的 觀察感染小鼠甲胎蛋白(mAFP)和4-1BB配體(m4-1BBL)融閤基因對小鼠樹突狀細胞(DC)生物學特性的影響.方法 構建含mAFP和m4-1BBL的融閤基因腺病毒Ad-mAFP-內部覈糖體起始位點(IRES)-m4-1BBL,感染H22肝癌細胞,Western blot法檢測mAFP和4-1BBL蛋白的錶達；誘導培養DC,併將其分為實驗組、載體對照組和陰性對照組,以上各組感染後觀察DC形態學變化；流式細胞儀檢測各組細胞錶型；混閤淋巴細胞反應(MLR)檢測各組DC的刺激活性以及對T細胞分泌榦擾素(IFN)-γ的影響.結果 成功構建融閤基因重組腺病毒Ad-mAFP-IRES-m4-1BBL併錶達于H22細胞；體外誘導培養穫得典型的DC；流式細胞儀檢測各組DC感染後的細胞錶型,實驗組DC細胞CD80[(60.7±8.7)％]、CD86[(67.0±4.6)％]錶達率顯著增高于其餘兩組(P＜0.05),刺激T細胞增殖能力(0.66±0.05)亦顯著增彊(P＜0.05),刺激T細胞分泌IFN-γ[(1280.7 ±68.2) ng/L]水平更高(P＜0.05).結論 成功構建含mAFP及4-1BBL融閤基因腺病毒疫苗,其感染DC後刺激T細胞增殖和分泌IFN-γ的能力增彊.
목적 관찰감염소서갑태단백(mAFP)화4-1BB배체(m4-1BBL)융합기인대소서수돌상세포(DC)생물학특성적영향.방법 구건함mAFP화m4-1BBL적융합기인선병독Ad-mAFP-내부핵당체기시위점(IRES)-m4-1BBL,감염H22간암세포,Western blot법검측mAFP화4-1BBL단백적표체；유도배양DC,병장기분위실험조、재체대조조화음성대조조,이상각조감염후관찰DC형태학변화；류식세포의검측각조세포표형；혼합림파세포반응(MLR)검측각조DC적자격활성이급대T세포분비간우소(IFN)-γ적영향.결과 성공구건융합기인중조선병독Ad-mAFP-IRES-m4-1BBL병표체우H22세포；체외유도배양획득전형적DC；류식세포의검측각조DC감염후적세포표형,실험조DC세포CD80[(60.7±8.7)％]、CD86[(67.0±4.6)％]표체솔현저증고우기여량조(P＜0.05),자격T세포증식능력(0.66±0.05)역현저증강(P＜0.05),자격T세포분비IFN-γ[(1280.7 ±68.2) ng/L]수평경고(P＜0.05).결론 성공구건함mAFP급4-1BBL융합기인선병독역묘,기감염DC후자격T세포증식화분비IFN-γ적능력증강.
Objective To investigate the biological characteristics of the dendritic cells transduced with truncated mouse alpha-fetoprotein(mAFP) and 4-1BBL recombinant adenoviruses.Methods Construction with mAFP and m4-1BBL coexpression gene recombinant adenovirus,H22 cells infected by recombinant adenovirus,and then mAFP and 4-1BBL expression detected by Western blotting.C57BL/6j murine bone marrow derived dendritic cells inducted.The dendritic cells (DCs) will be divided into experimental group which was adding with recombinant adenoviruses,negative Ad-eGFP group which was adding with blank adenovirus vectors and negative control group which was adding with normal saline,and then the morphological changes of DCs were observed by phase contrast microscope,flow cytometry was used to detective the phenotype of DCs,mixed lymphocyte reaction (MLR) was used to observe the DCs stimulating activity on T cell proliferation and secretion of cytokines.Results Recombinant adenovirus was successfully constructed and can be expressed in H22 cells.Through the induction and culture in vitro,the mature DCs with typical figure can be found under the microscope,the DCs cell phenotype CD80,CD86 has significant difference between the experimental group and the other two groups (P＜ 0.05),the proliferation ability of DCs stimulated T cell was enhanced in the experimental group (P ＜ 0.05),the DCs stimulated T cells secrete more interferon (IFN)-γ cytokines (P ＜ 0.05).Conclusion The recombinant adenovirus vaccine with fusion gene of mAFP and 4-1BBL was successfully constructed which transfected can increase the DC capacity to stimulate T cell proliferation and secretion of IFN-γ.
