Effect of electro-acupuncture on the expression of heat shock protein-70 gene in rat spinal cords following spinal cord injury

BACKGROUND:It is generally believed that the mechanism by which heat shock protein-70(HSP70) protects cells is related to its effectiveness in maintaining the normal stereochemical structure of intracellular proteins,and in participating in the process of cell apoptosis.Whether electro-acupuncture participates in HSP70 expression and produces neuroprotective effects remain unclear.OBJECTIVE:This study aimed at detecting HSP70 expression after electro-acupuncture in rats with transected spinal cord,in order to further validate the mechanism of electro-acupuncture-induced effects in the treatment of spinal cord injury.DESIGN:A controlled observational experiment.SETTING:Shanghai University of Traditional Chinese Medicine and Toho University,School of Medicine.MATERIALS:Seventy adult male Sprague-Dawley rats of SPF grade,weighing 200±20g,were provided by the Laboratory Animal Center of Shanghai University of Traditional Chinese Medicine,with permission No.SYXK(hu)2004-2005.The animals were handled in accordance with the requests from Animal Ethics Committees for guidance.A G6805-2 multiple purpose treatment machine was used (Shanghai Medical Instruments High-Tech Co.,Ltd.,Shanghai,China).METHODS:This study was carried out in the state level laboratories of Shanghai University of Traditional Chinese Medicine and Toho University,School of Medicine between January 2005 and July 2007.The rats were randomly divided into the electro-acupuncture treated group,which received electro-acupuncture treatment in addition to spinal cord surgery and the control group,which received only spinal cord surgery,with 35 rats in each group.All the rats underwent the same surgery consisting of spinal cord transection at the T10 level.If the spinal cord was completely transected and the two posterior limbs were completely paralyzed,then the surgery was considered successful and the animal was kept for further analysis and testing.After surgery,rats in the experimental group were electro-acupunctured with a G6805-2 multiple purpose treatment machine.Two needle electrodes were inserted under the T7 and T10 spinal processes,The treatment was administered once a day for 20 minutes.Rats in the control group were not given any treatment after surgery.Five rats were sacrificed separately in each group on days 1,2,3,7,14,21 and 28 after surgery.HSP70 gene expression at the site of lesion was located and quantitatively analyzed by immunohistochemistry and real-time PCR methods.Simultaneously,the spinal cord injury region and neurons were observed by HE and Kl(u)ver-Barrera stainings.MAIN OUTCOME MEASURES:①HSP70 gene expression in the spinal cord injury region.②The number of neurons in the spinal cord injury region.RESULTS:Seventy rats were involved in the final analysis.①At the end of each pre-determined block of time,HSP70 mRNA level in the spinal cord injury region of rats in the electro-acupuncture treated group was significantly higher than that in the control group (P＜0.05).HSP70 gene expression in the two groups reached peak levels on day 2 after surgery.②On days 7,14,21 and 28 after surgery,the number of neurons in the spinal cord injury region in the electro-acupuncture treated group was significantly higher than that in the control group(P＜0.05).CONCLUSION:Electro-acupuncture can effectively enhance HSP70 expression in the spinal cord injury region.HSP70 may participate in this apparent neuroprotective effect.