CAJ | 학술논문

背景:对于抗原递呈细胞树突状细胞及其分泌的细胞因子白细胞介素10在气道高反应性和炎症中的作用国内外文献报道较少.目的:构建小鼠白细胞介素10腺病毒重组体Ad-mIL-10,获得小鼠白细胞介素10基因修饰的树突状细胞,以期为下一步基因治疗的动物实验打下基础.方法:通过人工合成获得小鼠白细胞介素10基因,根据白细胞介素10基因序列及腺病毒载体的多克隆位点,合成包括酶切位点的基因序列,连接到pMD18-T载体并测序鉴定.用基因工程的方法将小鼠白细胞介素10基因克隆到BDAdeno-X~(TM)腺病毒载体,于人胚肾293细胞中包装、扩增病毒并测定白细胞介素10蛋白表达,转染到体外培养的小鼠骨髓来源的树突状细胞.结果与结论:成功构建了小鼠Ad-mIL-10重组腺病毒载体并高包装、扩增成功,测定高表达白细胞介素10蛋白,体外成功培养小鼠骨髓来源的小鼠树突状细胞并顺利转染Ad-mIL-10.提示用基因工程方法构建小鼠Ad-mIL-10重组腺病毒载体并转染小鼠骨髓来源的树突状细胞是可行的,可为进行相关基因治疗的可能性提供更充足的理论依据.
배경:대우항원체정세포수돌상세포급기분비적세포인자백세포개소10재기도고반응성화염증중적작용국내외문헌보도교소.목적:구건소서백세포개소10선병독중조체Ad-mIL-10,획득소서백세포개소10기인수식적수돌상세포,이기위하일보기인치료적동물실험타하기출.방법:통과인공합성획득소서백세포개소10기인,근거백세포개소10기인서렬급선병독재체적다극륭위점,합성포괄매절위점적기인서렬,련접도pMD18-T재체병측서감정.용기인공정적방법장소서백세포개소10기인극륭도BDAdeno-X~(TM)선병독재체,우인배신293세포중포장、확증병독병측정백세포개소10단백표체,전염도체외배양적소서골수래원적수돌상세포.결과여결론:성공구건료소서Ad-mIL-10중조선병독재체병고포장、확증성공,측정고표체백세포개소10단백,체외성공배양소서골수래원적소서수돌상세포병순리전염Ad-mIL-10.제시용기인공정방법구건소서Ad-mIL-10중조선병독재체병전염소서골수래원적수돌상세포시가행적,가위진행상관기인치료적가능성제공경충족적이론의거.
BACKGROUND: Few reports concern the effects of dendritic cells-a kind of antigen presenting cells, and interleukin-10 (IL-10) on airway hyperreactivity or inflammation. OBJECTIVE: To construct mice IL-10 recombinant adenovirus vector Ad-mIL-10 to acquire the dendritic cells modified by mIL-10, which can provide a foundation for the further study. METHODS: Mouse IL-10 (mIL-10) gene comprise of enzyme cutting spot was synthesized according to the mIL-10 gene sequence and multiclone spot of adenovirus vector, connected to pMD18-T vector and sequenced. MIL-10 was subcloned to BD Adeno-X~(TM) vector, packed and augmented in HEK 293 cells, following determine the protein expression, and the vector was transfected to mice bone marrow-derived dendritic cells. RESULTS AND CONCLUSION: Recombinant adenovirus vector Ad-mIL-10 was successfully synthesized, packed and augmented, which could highly express protein IL-10. Bone marrow-derived dendritic cells were successfully cultured and transduced in vitro. It suggested that it is feasible to transfect mice dendritic cells by Ad-mIL-10 adenovirus vector. The study can provide more sufficient theoretic evidence for the possibility of correlative gene therapy.