Effects Of Neural Stem Cells On Neural Regeneration In Skin Wounds Of The Rats

PART ONETHE CULTURE AND IDENTIFICATION OF NSCs WEREINDUCED THROUNGH SD RATS’ BMSCs BY AFGFObjective: To culture in vitro and purify rat bone marrow mesenchy-mal (BMSCs), and select the cell in accordance with morphology. Theacidic fibroblast growth factor (aFGF) was used to induce rat bone marrowmesenchymal stem cells (BMSCs) to differentiate into neural stem cells(NSCs), and to identify the cells before and after differentiation.Methods: Rat BMSCs were cultured by the whole bone marrowadherent culture method. The original generation of BMSCs were selectedby the different adherent time, were purified through subculture and change.The cell morphological characteristics were observed under the invertedphase contrast microscope and identified by BMSCs marker. aFGF (80ng/ml) was used to induce BMSCs to differentiate into NSCs. The cellmorphological characteristics were observed under the inverted phasecontrast microscope. The expression of the Nestin (NSE) was detected by the immunofluorescence after induced6hours.Results: The cultured cells showed adherent growth and long spindleappearance. With the best effect dose80ng/ml completed the subsequentinduced differentiation experiments. After being induced by80ng/ml aFGFfor6hours, some BMSCs cells contracted to become oval and extendelongated protrusions. Immunofluorescence showed a positive expressionof NSE.Conclusion: The whole bone marrow adherent culture method cancultivate highly purified rat BMSCs with stem cells’ characteristics.80ng/ml aFGF can induce BMSCs to differentiate into NSCs. PART TWOEFFECTS OF NEURAL STEM CELLS ON NEURALREGENERATION IN SKIN WOUNDS OF THE RATSObjective: After neural stem cells (NSCs) was differentiated from bonemarrow mesenchymal stem cells (BMSCs), it was implanted to skin wounds and the study is to observe the effects of NSCs on neural regeneration in skinwounds.Methods: SD rat BMSCs were isolated and cultured by the wholebone marrow adherent culture method，and was induced by aFGF todifferentiate into NSCs, and was marked with DAPI.81SD rats were madeback full-thickness defect skin wounds. Each group had9SD rats. NSCswere randomly divided into the local wound injection (group A), the tailvein injection (group B), and NSCs with plasma graft implantation (groupC). BMSCs were divided into the local wound injection (group D), the tailvein injection of (group E), BMSCs with plasma graft implantation (groupF), and plasma graft implantation (group G). Tail veins were injected withnormal saline (group H), and local wounds were injected with normalsaline (group I). The expression of protein gene product9.5(PGP9.5) inwound skin after transplant treatment at the4st，5nd，7thweek was detectedby immunofluorescence and RT-PCR in order to reflect the skin nerveregeneration.Results: The difference of the fluorescence density value of PGP9.5in wound healing at4th，5th，7thweek had statistical significance (F=427.03,P=0.000). There were differences in stem cells transplant treatment tospecific expression of nerve repair; the use of data analysis was repeatedmeasures analysis of variance design information in the packet on，the difference was statistically significant (F=130.44, P=0.000). Theinteraction of stem cell transplantation results in terms of time and groupwas statistically significant (F=3.543, P=0.001). Nerve repair wassignificantly after local injection of NSCs transplantation compared withintravenous injection of NSCs transplantation (P=0.001). Nerve repairwas significantly after intravenous injection of NSCs transplantedcompared with NSCs transplanted outside (P=0.000), and nerve repair wassignificantly after local injection of NSCs transplanted compared with localinjection of BMSCs transplantation (P=0.000), nerve repair wassignificantly after local injection of BMSCs transplantation compared withsaline local injection of transplantation (P=0.000), there was statisticallysignificant (P=0.014) between the local injection of local externalaccessories plasma graft transplantation and saline, there was no statisticalsignificance between the local injection of saline and transplantedintravenously transplantation (P=0.446). The detection of PGP9.5withRT-PCR further illustrates the above resultsConclusion: Differentiating from BMSCs induced by aFGF, NSCscan promote the neural regeneration of wound when transplanted into skinwounds. NSCs has better regeneration effects compared to BMSCs underthe same transplantation method. Local injection is the most effectivepattern in promoting neural regeneration of wound. PART THREESTUDY ON NEURA STEM CELLS IN ACCELERATINGWOUND HEALING IN RATSObjective：To observe the effects of neural stem cells (NSCs) afterimplantation in full-thickness skin wound in rats.Methods：Rat BMSCs were isolated and cultured，and aFGF was usedto induce BMSCs to differentiate into NSCs, and marked with DAPI. The27SD rats were made back skin wounds. The NSCs were randomlydivided into the local wound injection (group A). BMSCs were divided intothe local wound injection (group B), local wounds were injected withnormal saline (group C). Wound healing time was recorded. The woundhealing was evaluated on the3rd,7thand14thday postoperatively. Tissuesamples at the wound site were collected for VG and HE staining and toreflect the repair of skin wound inflammation and tissue collagen fibers.Results：After the artificial skin transplantation, the wounds in group Ahealed well, accompanied by slight local inflammatory response withoutbleeding, empyema or necrosis. The wound healing time in group Asignificantly shortened compared with group B and group C (P=0.000).The healing rate of NSCs group was significantly higher than that ofBMSCs group, and the healing rate of BMSCs group was higher than thatof the saline control group (P=0.000). Conclusion：As seed cells, neural stem cell can be used to repair skinwounds and shorten wound healing time when locally injected into skinwound.