Antisense expression of PKCalpha improved sensitivity of SGC7901/VCR cells to doxorubicin.
OBJECTIVE
To explore whether antisense blocking of protein kinase C alpha (PKCalpha) would reverse multi-drug resistance (MDR) in the vincristine (VCR)-resistant human gastric cancer cell line SGC7901/VCR.


METHODS
SGC7901/VCR cells expressing antisense PKCalpha, SGC7901/VCR/aPKC, were established by transfection with a recombinant plasmid reversely inserted with PKCalpha cDNA. Empty vector (PCI-neo)-transfected cell clones, SGC7901/VCR/neo, served as the control. Western blot method was used to detect PKCalpha content in SGC7901, SGC7901/VCR, SGC7901/VCR/neo and SGC7901/VCR/aPKC cells, using PKCalpha-specific antibody. The sensitivity of SGC7901, SGC7901/VCR, SGC7901/VCR/neo and SGC7901/VCR/aPKC cells to doxorubicin (DOX) in vitro was determined by MTT assay. The uptake of DOX in these cells was detected with fluorescence spectrophotometer.


RESULTS
Western blot analysis showed that the PKCalpha protein level was about 8.7-fold higher in SGC7901/VCR cells than that in SGC7901 cells, whereas the protein expression of PKCalpha was reduced by 78% in SGC7901/VCR/aPKC cells when compared with the SGC7901/VCR cells. SGC7901/VCR/aPKC cells had a 4.2-fold increase in DOX cytotoxicity, accompanied by a 1.7-fold increase of DOX accumulation in comparison with SGC7901/VCR cells.


CONCLUSIONS
PKCalpha positively regulates MDR in SGC7901 cells, and inhibition of PKCalpha can partially attenuate MDR in human gastric cancer cells.
Wu
Da-Long
DL
Department of Pharmacology, School of Medicine, Jiaxing College, Jiaxing 314001, Zhejiang Province, China. wudalong66@sina.com
Sui
Feng-Ying
FY
Du
Cheng
C
Zhang
Cheng-Wen
CW
Hui
Bin
B
Xu
Shui-Ling
SL
Lu
Huan-Zhang
HZ
Song
Guo-Jie
GJ
D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D018076:DNA, Complementary / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics; D004317:Doxorubicin / Q000493:pharmacokinetics / Q000494:pharmacology*; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D006801:Humans; D016376:Oligonucleotides, Antisense; D051571:Protein Kinase C-alpha / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D015183:Restriction Mapping; D013274:Stomach Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology; D014750:Vincristine / Q000494:pharmacology
D018076:DNA, Complementary; D004273:DNA, Neoplasm; D016376:Oligonucleotides, Antisense; D014750:Vincristine; D004317:Doxorubicin; D051571:Protein Kinase C-alpha