| BACKGROUND AND OBJECTIVE:Chronic non-healing wound is a multifactorial complication of diabetes that results specifically as a consequence of impaired angiogenesis and blocked formation of granulation tissue,and currently lacks in effective treatments.While a stem cell-based therapy may provide a novel treatment to augment diabetic wound healing,inferior cell survival at the diabetic skin wound is one of the key causes that are responsible for the low efficacy of the stem cell therapy.It is still a critical challenge delivering ADSCs to diabetic wound sites while maintaining a high cell survival rate.Delivering stem cells within a suitable biomaterial may ensure sustained viability and functionality of stem cells and thus may improve their transplantation efficiency.Hydrogels that can mimic a 3D cell microenvironment may be useful biomaterials for this purpose.Pluronic F127 is an synthetic and biocompatible hydrogel which has been approved for use in humans by FDA.A unique characteristic of this hydrogel is its thermosensitivity,which enables it to easily encapsulate cells and favors high numbers of cells adhering at the defect site.However,little work has been yet reported on the application of ADSCs encapsulated in Pluronic F127 hydrogel for diabetic wound healing.In the present study,we aimed to optimize the augmentation of diabetic wound healing by topical application of allogeneic non-diabetic ADSCs seeded in Pluronic F127 hydrogel scaffold in a diabetic rat model.We investigated whether topical application of ADSCs encapsulated in Pluronic F127 hydrogel could speed up wound healing in a diabetic rat cutaneous injury model.METHODS:ADSCs were isolated from healthy SD rats by collagenase digestion and the cell phenotype was identified by flow cytometry.The model of T1DM rats was established.MTT was used to compare the proliferation of ADSCs in diabetic rats and normal rats.CCK-8 was used to detect the effect of different concentrations of Pluronic F127 on the proliferation of ADSCs.Two 9-mm diameter circular impressions were made by placement of a punch biopsy instrument at the same level,with a 1.5 cm interval.Rats were randomly treated with one of four treatment groups:ADSCs/Gel group(100ul Pluronic F127 gel solution containing 1x106 ADSCs);ADSCs alone group(100ul 1x106 ADSCs suspension solution);Gel alone group(100ul Pluronic F127 solution alone);and the control group(100ul PBS)(n = six rats and twelve wounds per group).To better understand the mechanisms for improved healing,we studied the treatment effects on changes in angiogenesis,cellular proliferation and the formation of new granulation tissue.RESULTS:ADSCs were successfully isolated and purified,and MTT results showed that normal ADSCs had better proliferative ability than ADSCs.CCK-8 shows that 20%Pluronic F127 is more suitable for cell proliferation than 30%and has a good biodegradability.The wound healing rate was(97.8 ± 1.8)%at 14 days after operation,which was significantly higher than that in other groups(P = 0.000).ADSCs/Gel group has a better histological score.MVD in ADSCs/Gel group was 21.0±2.4,which was significantly higher than that in control group(P<0.05).The number of microvessels in ADSCs/Gel group and ADSCs group was 21.0 ± 2.4,while ADSCs group was 15.7 ± 1.4(P = 0.000).The cells seeded in the hydrogel enhanced angiogenesis(CD31 marker)and promoted the cell proliferation(Ki67 marker)at the wound site,and significantly accelerated wound closure,which was accompanied by facilitated regeneration of granulation tissue.Consistently,levels of the mRNA expression of key angiogenesis growth factor VEGF and key wound healing growth factor TGFβ-1 were also up-regulated in the cell-treated wounds when compared with untreated wounds.(P<0.05).CONCLUSIONS:Topically transplanting allogeneic ADSCs may help to enhance angiogenesis in places where the microcirculation is poor and the hydrogel supports the delivery and engraftment of stem cells to the wound. |
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