[Recombinant adeno-associated virus mediated RNA interference of angiogenin expression inhibits cell growth of human lung adenocarcinoma].
OBJECTIVE
To study the inhibition of angiogenin (ANG) expression in human lung squamous cancer cell strain-A549 through adeno-associated virus (AAV)-mediated RNA-interference, and therefore to observe its effect on the growth of cancer cells and tumor formation.


METHODS
Recombinant AAV expressing H1-promoter-induced small-interference- RNA (siRNA) targeting ANG (AAV-shANG) was constructed, and then transfected into A549 cells. A549 cells and cells transfected with AAV-Null were used as the control groups. The effects of the reduced expression of ANG by RNAi from AAV-shANG on the growth, formation, reproduction, apoptosis, and microvessel-density of the carcinoma were observed.


RESULTS
In vitro experiment showed that AAV-shANG was constructed successfully, There was an significant decrease in the expression of ANG protein 72 h after transfection, compared with the normal A459 cells and AAV-Null cells (P < 0.01). Cell cycle analysis showed that the proliferation index (PI) of normal A549 cells, AAV-Null cells and AAVshANG cells were 0.32 +/- 0.29, 0.35 +/- 0.38 and 0.31 +/- 0.43, respectively. There was no statistic difference in the PIs among the 3 groups (P > 0.05). In vivo experiment using thymus-defect mice showed that, there was an remarkable reduction in the mass and volume of tumors in AAV-shANG transfected group, compared to the control groups. Microvessel-density was 9.4 +/- 1.5, 9.8 +/- 2.1 and 5.7 +/- 1.9, respectively in the 3 groups, a statistic difference among the AAV-shANG-transfected group, the normal A549 group and the AAV-Null transfected group. The percentages of apoptotic cells in each group were (7.7 +/- 3.1)%, (8.5 +/- 5.4)%, (17.1 +/- 8.6)%, respectively, the experimental group being higher than those of the control groups. Positive rates of PCNA were (84.8 +/- 9.7)%, (85.8 +/- 9.8)%, and (70.4 +/- 10.1)%, respectively, the AAV-shANG transfected cancer cells showing a lower PCNA index than the control groups.


CONCLUSIONS
AAV-mediated expression of siRNA could reduce the expression of ANG in cancer cells, significantly enough to inhibit cell proliferation, promote cell apoptosis and inhibit tumor growth.
Li
Bai-Ling
BL
Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai 200433, China.
Zhang
Guan-Xin
GX
Hou
Xiao-Lei
XL
Tan
Meng-Wei
MW
Yuan
Yang
Y
Liu
Xiao-Hong
XH
Gong
De-Jun
DJ
Huang
Sheng-Dong
SD
D000230:Adenocarcinoma / Q000378:metabolism / Q000473:pathology*; D000818:Animals; D017209:Apoptosis; D002453:Cell Cycle / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D000229:Dependovirus / Q000235:genetics; D006801:Humans; D008175:Lung Neoplasms / Q000378:metabolism / Q000473:pathology*; D051379:Mice; D008819:Mice, Nude; D034622:RNA Interference; D011995:Recombination, Genetic; D012259:Ribonuclease, Pancreatic / Q000378:metabolism*; D014162:Transfection
C046366:angiogenin; D012259:Ribonuclease, Pancreatic