Influence of co-down-regulation of caspase-3 and caspase-7 by siRNAs on sodium butyrate-induced apoptotic cell death of Chinese hamster ovary cells producing thrombopoietin.
Previously, the expression of caspase-3 siRNA could not effectively inhibit sodium butyrate (NaBu)-induced apoptotic cell death of recombinant Chinese hamster ovary (rCHO) cells producing human thrombopoietin (hTPO). Caspase-3 siRNA expressing cells appeared to compensate for the lack of caspase-3 by increasing active caspase-7 levels. For the successful inhibition of NaBu-induced apoptosis of rCHO cells, both caspase-3 and caspase-7 were down-regulated using the siRNA expression vector system. Co-down-regulation of caspase-3 and caspase-7 increased cell viability and extended culture longevity in serum-free culture in the presence or absence of 1mM NaBu addition. In the cultures with 1mM NaBu addition, the maximum hTPO concentration in rCHO cells with down-regulation of both caspases was approximately 55% higher than that in rCHO cells without down-regulation of caspases and approximately 16% higher than rCHO cells with down-regulation of only caspase-3. However, in the culture with 3mM NaBu, this strategy could not dramatically enhance the culture longevity and hTPO production, compared to Bcl-2 overexpression. The different result in hTPO production between down-regulation of caspases and Bcl-2 overexpression may be because the down-regulation of caspase-3 and caspase-7, unlike Bcl-2 overexpression, could not maintain mitochondrial membrane potential in the presence of 3mM NaBu. Taken together, co-down-regulation of caspase-3 and caspase-7 is effective in regard to extension of culture longevity and enhancement of hTPO production in a serum-free culture in the presence or absence of 1mM NaBu addition.
Sung
Yun Hee
YH
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 373-1 Kusong-Dong, Yusong-Gu, Daejon 305-701, Republic of Korea.
Lee
Jae Seong
JS
Park
Soon Hye
SH
Koo
Jane
J
Lee
Gyun Min
GM
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001562:Benzimidazoles / Q000378:metabolism; D020148:Butyric Acid / Q000494:pharmacology*; D016466:CHO Cells; D002232:Carbocyanines / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism*; D053179:Caspase 7 / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D003412:Cricetulus; D016895:Culture Media, Serum-Free; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects; D005456:Fluorescent Dyes / Q000378:metabolism; D008564:Membrane Potentials / Q000502:physiology; D008928:Mitochondria / Q000502:physiology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis; D034741:RNA, Small Interfering / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D013926:Thrombopoietin / Q000096:biosynthesis* / Q000235:genetics; D014162:Transfection
D001562:Benzimidazoles; D002232:Carbocyanines; D016895:Culture Media, Serum-Free; D005456:Fluorescent Dyes; D019253:Proto-Oncogene Proteins c-bcl-2; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D020148:Butyric Acid; C068624:5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine; D013926:Thrombopoietin; D053148:Caspase 3; D053179:Caspase 7