Regulation of apoptosis in nucleus pulposus cells by optimized exogenous Bcl-2 overexpression.
Although the etiology of intervertebral disc degeneration is poorly understood, one possible approach to regulate the process of intervertebral disc degeneration may include the inhibition of apoptosis. We investigated the anti-apoptotic effects of bcl-2 in nucleus pulposus cells to enhance disc cell survival. Rat nucleus pulposus cells were transfected in vitro with a codon optimized rat bcl-2 gene. Forty-eight hours after transfection, cells were cultured in serum-deprived medium. After serum withdrawal, the cells were evaluated for bcl-2 protein levels and cell apoptosis. To investigate the effects of bcl-2 overexpression on the final apoptotic pathways and on basic genes important for nucleus pulposus homeostasis, mRNA levels of caspase-3, type II collagen, and aggrecan were also quantified. Nucleus pulposus cells were successfully transfected with codon optimized bcl-2 gene, which effectively reduced serum starvation-induced cell apoptosis. Overexpression of bcl-2 also reduced the mRNA expression level of caspase-3. mRNA levels of type II collagen and aggrecan were significantly higher in bcl-2 transfected groups compared to control plasmid vector groups after serum withdrawal. We firstly showed that bcl-2 overexpression in intervertebral disc cells was effective in preventing in vitro apoptotic cell death, indicating the potential advantages of this therapeutic approach in regulating disc degeneration.
Sudo
Hideki
H
Department of Advanced Medicine for Spine and Spinal Cord Disorders, Hokkaido University Graduate School of Medicine, North-15, West-7, Kita-ku, Sapporo, Hokkaido 060-8638, Japan. hidekisudo@yahoo.co.jp
Minami
Akio
A
D053674:Aggrecans / Q000096:biosynthesis; D000818:Animals; D017209:Apoptosis; D001483:Base Sequence; D053148:Caspase 3 / Q000096:biosynthesis; D024043:Collagen Type II / Q000096:biosynthesis; D005786:Gene Expression Regulation; D019254:Genes, bcl-2; D007403:Intervertebral Disc / Q000166:cytology / Q000378:metabolism*; D055959:Intervertebral Disc Degeneration / Q000517:prevention & control; D008297:Male; D008969:Molecular Sequence Data; D051381:Rats; D017207:Rats, Sprague-Dawley; D016415:Sequence Alignment
D053674:Aggrecans; D024043:Collagen Type II; D053148:Caspase 3