| Objective: To study the effect of electrical stimμlation combined with BMSCstransplantation on GAP-43and Syp expression in caudate nucleus ofintracerebral hemorrhage in rats. Methods: After routinely recovered andpassaged, GFP-labeled bone mesenchymal stem cells (BMSCs) wereinductively cμltured for six hours, then collected and reserved with theconcentration of2.5×107cells/ml. Brain hemorrhage (ICH) models in SDrats were builded by the injection of collagenase and heparin. Respectivelyevaluated by nerve injury severity score (NSS scores) after model-builded for2hours,24hours and48hours of, the rats were selected with scores exceeded9for further experiments. Three days later of successfμl model-build, ICH-ratswere divided threegroups: experimental control group (NS group), BMSCstransplantation group (BMSCs group) and BMSCs transplantation combinedwith electro-acupuncture stimμlation group (Ea-BMSCs group), by separateinjection with20μl physiological saline,20μl BMSCs and20μl BMSCs2-hour-later supplement with electro-acupuncture stimμlation in rats Baihui andDazhui. According to the treatment time, each group is respectively redividedinto five subgroups namely1-day,3-day,5-day,7-day and14-day group aftertransplantation. Then, the rats were sacrificed and standbyed after assessmentwith NSS score at the above-mentioned time points. By western blot and RT-PCR analysis, GAP-43and Syp expression were detected in caudateputamen area of ICH rats in different groups at each time point. Resμlts:(1)NSS score. The pre-transplant score: Before transplantation in a rat ICHmodeling, NSS score of model rats in different groups (namely Ea-BMSCsgroup, BMSCs group and NS group) is basically identical with no significantdifference at the same time point (P>0.05). The post-transplant score:Compared with NS group, NSS score in Ea-BMSCs group significantlydecreased in3days,5days,7days and14days after transplantation(P<0.05),while that in BMSCs group did in5days (P<0.05). Simμltaneously Ea-BMSCsgroup subject to BMSCs group, NSS score markedly descended in5days,7daysand14days (P<0.05).(2) GAP-43and Syp protein expression by Western blotanalysis. Three groups in one day after transplantation had a slight GAP-43expression, while Syp displayed no significant expression in one day andvisible expression in three days. Three groups comparison at each time point:GAP-43expression of Ea-BMSCs group gradually increased in3days,5days,7days and14days, while Syp expression enhanced in5days7days14days.Both markers showed the strongest expression in7-14days (P<0.05). GAP-43expression of BMSCs group gradually increased in3days,5days and7days,peaked (P<0.05) in seven days, afterward became weakening. Meanwhile, Sypexpression enhanced in5days,7days and14days, strengthened in7-14days.GAP-43expression of NS group gradually increased in3days,5days and7days, presented a stronger in7days, then weakened, while Syp showed visibleexpression in5days and7days, but no significant expression in14days. Three groups comparison between each other: When Ea-BMSCs group comparedwith NS group, GAP-43and Syp expression was significantly increased in5days,7days and14days (P<0.05). However, GAP-43expression ofEa-BMSCs group was markedly enhanced subject to the BMSCs group in7days and14days (P<0.05), while Syp expression increased in14days (P<0.05).(3) GAP-43and Syp mRNA expression by semi-quantitative RT-PCRanalysis. In one day after transplantation, GAP-43mRNA levels of three groupsincreased, while Syp mRNA levels lowered, but in3days Syp mRNA levelselevated. Three groups comparison at each time point: GAP-43mRNA levels ofEa-BMSCs group gradually rised in3days,5days,7days and14days, whileSyp mRNA levels were higher in5days,7days and14days. Both mRNAlevels peaked in7-14days (P<0.05). GAP-43mRNA levels of BMSCs group in3days,5days and7days increased, especially in7days were more staggering(P<0.05), followed by begining to decline. Syp mRNA showed higherexpression in5days,7days and14days, mostly higher in7-14days. GAP-43mRNA levels of NS group gradually increased in3days,5days and7days,became higher in7days, and then decreased. At the same time, Syp mRNAdisplayed visible expression in5days and7days, but no significant expressionin14days. Three groups comparison between each other: In comparison withNS group, GAP-43and Syp mRNA levels of Ea-BMSCs group significantlyincreased in5days,7days and14days (P<0.05). When compared withBMSCs group, GAP-43mRNA expression of Ea-BMSCs group in5days,7days and14days significantly increased (P<0.05), while Syp mRNA being higher in14days (P<0.05). Conclusion:(1) Electro-acupuncture stimμlationcombined with BMSCs transplantation can be effective in treating experimentalICH and superior to a single BMSCs transplantation.(2) Electro-acupuncturestimμlation is capable of significantly enhancing GAP-43and Syp expressionafter BMSCs transplantation in the treatment of rat ICH, stimμlating exogenousBMSCs and endogenous stem cell proliferation and differentiation, andpromoting damaged nerve cells regeneration and repair.(3) The therapeuticalmechanism of electro-acupuncture plus BMSCs for rat ICH may be through theupregμlation of GAP-43and Syp expression and the acceleration ofsynaptogenesis so as to establish synaptic connections and neuronal circuits,improve the nerve regeneration microenvironment, and promote BMSCsproliferation and differentiation into neurons and glial cells, μltimately inducethe damaged nerve function recovery. |
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