Yu Xin, Zhang Hai-feng, Wang Jin-zhi, Xie Yu-feng, Yang Ji-cheng, Miao Jing-cheng
Cell and Molecular Biology Institute, College of Medicine, Soochow University, Suzhou 215123, China.
Zhonghua Xue Ye Xue Za Zhi. 2007 Jun;28(6):396-400.
To observe the effect of recombinant adenovirus Ad-ING4 on K562 cells.
Human ING4 recombinant transfer vector pAdTrack-CMV-ING4 was constructed by enzyme digest and ligation of human ING4 gene which was obtained through site specific point mutation of mouse ING4. The vector was co-transduced into BJ5183 E. coli with pAdEasy-1. The new recombinant adenovirus vector pAdEasy-1-pAdTrack-CMV-hING4 was transfected into QBI-293A cells. To obtain the ING4 recombined adenovirus (Ad-ING4). Ad-ING4 was used to infect K562 cells. The effect on K562 cells of ING4 was tested by LSCM FCM and immunohistochemistry.
Human ING4 recombinant adenovirus vector was constructed successfully, and high titre ING4 recombinant adenovirus (Ad-ING4) was obtained. ING4 can down-regulate the expression of bcl-2 and up-regulate expression of bax. The apoptosis of K562 cells induced by ING4 was proved by LSCM FCM and immunohistochemistry. The apoptosis rate was 19.7% (after 72h), which displayed significant difference compared with that of control groups (P < 0.01).
Ad-ING4 can inhibit the growth of K562 cells and induce the cells apoptosis. The human ING4 recombinant adenoviral vector constructed might provide an approach to the target therapy of tumors.
观察重组腺病毒Ad-ING4对K562细胞的作用。
通过对经小鼠ING4定点突变获得的人ING4基因进行酶切和连接，构建人ING4重组转移载体pAdTrack-CMV-ING4。将该载体与pAdEasy-1共转导至BJ5183大肠杆菌中。将新的重组腺病毒载体pAdEasy-1-pAdTrack-CMV-hING4转染至QBI-293A细胞中，以获得ING4重组腺病毒（Ad-ING4）。用Ad-ING4感染K562细胞。通过激光扫描共聚焦显微镜（LSCM）、流式细胞术（FCM）和免疫组织化学检测ING4对K562细胞的作用。
成功构建了人ING4重组腺病毒载体，并获得了高滴度的ING4重组腺病毒（Ad-ING4）。ING4可下调bcl-2的表达并上调bax的表达。通过LSCM、FCM和免疫组织化学证实了ING4诱导K562细胞凋亡。凋亡率为19.7%（72小时后），与对照组相比差异有统计学意义（P<0.01）。
Ad-ING4可抑制K562细胞的生长并诱导细胞凋亡。构建的人ING4重组腺病毒载体可能为肿瘤的靶向治疗提供一种途径。
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