The Application Of The Leptin Gene-modified HPMSCs To The Healing Of Combined Radiation-wound Injury As A Countermeasure

With the social development as well as an gradual increase in the rhythms ofliving, more and more environmental and social psychological factors have made theincidence rate of malignant tumor higher each year(year by year), with no exceptionfor oral and maxillofacial malignant tumor. Nowadays the tissue defects after theoperation for removal of the oral and maxillofacial malignant tumor is usuallyrepaired by the skin grafting method, and radiation therapy or postoperativechemotherapy is followed after the operation as well. This not only causes the severedamage of the skin tissue, but also makes the healing process of the flap delayed oreven lead the flap’s necrosis, which is a typical “combined radiation-wound injury”and is one of the refractory wounds rather difficult to cure.Human placenta-derived mesenchymal stem cells, a kind of cells seprated fromthe placenta parenchyma, have self-renewal capacity, to which is paid attentiongradually in recent years as a new source of stem cells. Zhu Zhenwei in our lab hasbeen successfully extracted HPMSCs from human placenta..Zhu Zhenwei found thatHPMSCs can deliver target gene efficiently and is a good carrier of cell or genetherapy. The studies of Liu zhihui have shown that HPMSCs not only is applid to theanimal models of ischemic skin flap as an ideal seed cells, but also act as vetor cellsof VEGF and exert therapeutic effects. Numerous studies show that HPMSCs hasimportant applications in the field of wound healing. However, the appliance tocombined radiation-wound injury ptomoting its healing has no relevant reports inreferences from China and other country currently. Leptin is an polypeptide hormoneand secreted by white adipose tissue, which can promote the epithelial regenerationand the formation of new blood vessels, and plays an important role in the process ofpromoting wound healing. But the half-life of Leptin is very short, so it is difficult toplay persistent effect by simple topical application. In order to make Leptin playpersistent effect, we considered that whether it could be transfected into mesenchymal stem cells and get continuous expression by MSCs, so as to the leptin could playpersistent effenc on improving wound healing. The purpose of this study is to graft theleptin gene-modified HPMSCs transfected successfully by Zhu zhenwei into theradiation-wound area in the Wistar rat model to observe the wound healing of theleptin gene-modified HPMSCs in combined radiation-wound injury.Methods: Male Wistar rats is applied to the study. To build animal model if therandom skin flap on the back of the rat with the size of3.0×5.0cm and sutured in situ,and exposed it to5Gy of electron ray after three days of the surgery. The Wistar ratsis divided into three groups by random and after24hours of radiation the normalHPMSCs and the leptin gene-modified HPMSCs and the normal saline weretransplanted subcutaneously into flap with respectively. In this study, wound healingof leptin and HPMSCs in combined radiation-wound injury was assessed by generalobservation, the histological examination, immunohistochemical methods and othermeans.Results:1. There is no immunological rejection in every experimental group during14days after cell-transplantation. On the seven days after cell-transplatation, the survivalrate of flap: leptin gene-modified HPMSCs group> HPMSCs group> blank controlgroup.2. Histological examination HE staining on the fourteenth day aftercell-transplatation, we found that in group I, there are the most abundant granulationtissues, new capillaries and fibroblasts and there are less inflammatory cell infiltrationaround the capillaries and collagen arranged irregularly; in group II, there are moreabundant granulation tissues, new capillaries and fibroblasts and there are moreinflammatory cell infiltration around the capillaries and collagen arranged irregularly;in group III we could see less abundant granulation tissues, new capillaries and cellcompoonents, but a large number of inflammatory cell infiltration and hyaline fibrousconnective tissue.3、The fourteenth day after cell-transplatation, the results of the vWF factorpolyclonal antibody immunohistochemical staining showed that the vascular endothelial cells express factor vWF, and the cytoplasm stained brown. Under themicroscope in each group we could observe the new relation of microvessel density:leptin gene-modified HPMSCs group> HPMSCs group> blank control group; theresults of the leptin polyclonal antibody immunohistochemical staining showed thatthe epithelial keratinocytes and vascular endothelial cells express leptin, and thecytoplasm stained brown. Under the microscope in each group we could observe thepositive expression of leptin: leptin gene-modified HPMSCs group> HPMSCs group> blank control group.Conclusion:1. Human placental-derived mesenchymal stem cells have low immunogenicityand the application of different species does not cause immunological rejection;2. Under the role of local microenvironment, HPMSCs can differentiate intoendothelial cells, promote the formation of new capillaries, accelerate the healing ofthe combined radiation-wound injury and increase the incidence rate of the flap;3. HPMSCs can as the carrier cells of gene therapy secrete leptin continuously;4. HPMSCs and leptin has a synergistic effect in the process of the woundhealing, and whether HPMSCs with gene-modified or not could significantly increasethe incidence rate of the flap with radiation therapy, however, HPMSCs withgene-modified have more effect.