Hideki Sudo1, Akio Minami. 1. Department of Advanced Medicine for Spine and Spinal Cord Disorders, Hokkaido University Graduate School of Medicine, Sapporo, Hokkaido, Japan. hidekisudo@yahoo.co.jp
Abstract
OBJECTIVE: Although the etiology of intervertebral disc degeneration is poorly understood, one possible approach for its regulation is apoptosis inhibition. This study was undertaken to investigate the antiapoptotic effects of caspase 3 in intervertebral disc degeneration in rabbits. METHODS: We investigated the effects of caspase 3 small interfering RNA (siRNA) on rabbit nucleus pulposus cells in a serum-starved medium. The effects of direct injection of Alexa Fluor 555-labeled caspase 3 siRNA into the intervertebral disc were also determined in vivo using the rabbit anular needle puncture model. RESULTS: Rabbit nucleus pulposus cells transfected with caspase 3 siRNA showed a significant decrease in serum-starved apoptotic cells. After local injection of caspase 3 siRNA into intervertebral discs, red fluorescence was observed in the nucleus pulposus upon treatment with Alexa Fluor 555-labeled caspase 3 siRNA. Caspase 3 messenger RNA and protein were down-regulated in the caspase 3 siRNA group. Magnetic resonance imaging and histologic evaluation showed that degenerative changes were significantly suppressed in the caspase 3 siRNA group 4 and 8 weeks after injection. Quantification of TUNEL staining showed that the caspase 3 siRNA group had significantly fewer apoptotic nucleus pulposus cells than the control siRNA group. CONCLUSION: Our findings indicate that caspase 3 knockdown in rabbit intervertebral disc cells is effective in preventing apoptotic cell death, thus regulating intervertebral disc degeneration.
OBJECTIVE: Although the etiology of intervertebral disc degeneration is poorly understood, one possible approach for its regulation is apoptosis inhibition. This study was undertaken to investigate the antiapoptotic effects of caspase 3 in intervertebral disc degeneration in rabbits. METHODS: We investigated the effects of caspase 3 small interfering RNA (siRNA) on rabbit nucleus pulposus cells in a serum-starved medium. The effects of direct injection of Alexa Fluor 555-labeled caspase 3 siRNA into the intervertebral disc were also determined in vivo using the rabbit anular needle puncture model. RESULTS:Rabbit nucleus pulposus cells transfected with caspase 3 siRNA showed a significant decrease in serum-starved apoptotic cells. After local injection of caspase 3 siRNA into intervertebral discs, red fluorescence was observed in the nucleus pulposus upon treatment with Alexa Fluor 555-labeled caspase 3 siRNA. Caspase 3 messenger RNA and protein were down-regulated in the caspase 3 siRNA group. Magnetic resonance imaging and histologic evaluation showed that degenerative changes were significantly suppressed in the caspase 3 siRNA group 4 and 8 weeks after injection. Quantification of TUNEL staining showed that the caspase 3 siRNA group had significantly fewer apoptotic nucleus pulposus cells than the control siRNA group. CONCLUSION: Our findings indicate that caspase 3 knockdown in rabbit intervertebral disc cells is effective in preventing apoptotic cell death, thus regulating intervertebral disc degeneration.
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