The Mechanism Research Of Wound Healing In Rats Through Calcium Alginate Dressing

Objective: To detect the effect of relative proliferation rate which alginate calcium dressing on fibroblast in vitro, then evaluate the cytotoxicity of the material and the worst environment affected the proliferation of fibroblast. To obtain the fibroblast cells(L929) apoptosis induced by the calcium alginate dressing in vitro. To evaluate the effect of calcium alginate in rats wound healing process and the effect of calcium alginate on transforming growth factor-β(TGF-β)、vascular endothelial growth factor(VEGF) and interleukin-6(IL-6) in the process of wound healing. To evaluate the effect of calcium alginate in rats wound healing process.Methods: ①Tetrazolium salt colorimetry(methyl thiazolyl tetrazolium, MTT) was used to detect the absorbance of the extracted liquid of the material in different concentrations(0%,50% and 100%), then the relative proliferation rate of fibroblast in each group were calculated, and their cytotoxicity was evaluated; ②The fibroblast cells(L929) co-cultured with calcium alginate dressing for 72 h as experimental group. Terminal uridine nick-end labeling(Tunel) method was used to identify the pattern of cell death, then the apoptosis rate of fibroblast in each group were calculated under the fluorescence microscope.③The wound models on rat’s back skin were made. The calcium alginate dressings(experimental group, n=24) or classical gauze(controlled group, n=24) were used, and the healing rates at different times after operation were obtained and compared. Meanwhile HE, Masson, histological and immunohistochemical staining methods were applied to evaluate the histological change of skin tissue within wound area at different periods post-operation.Results: ①The former results showed that relative proliferation rate in each group kept up a high level, and the cytotoxicity of extracted liquid of material in each group was 0 or 1 grade. The latter results showed that the concentration under 10% had no influence on the proliferation of fibroblast, and the concentration over 20% wound affect its proliferation. ②The results showed that the apoptotic cells were appeared in both groups, and increased along with the incubation time. However, the apoptosis rate in each group showed no significant differences.③The results showed that the granulation tissue was thin and the inflammatory reaction was serious in the two groups on the 3rd postoperative day, and the granulation tissue in controlled group was thinner that in the experimental group on the 7rd postoperative day. At the 14 rd day, the granulation tissue in the experimental group had reconstructed completely, while inflammatory reaction in the controlled group has alleviated. The immunohistochemical staining results showed that there was no obvious difference in the level of VEGF and TGF-βbetween the two groups postoperative periods. However the level of IL-6 in experiment group was below that in the control group on the 3 and 7rd postoperative day, while there no obvious difference between them on the 14 rd postoperative day.④The healing rate of the wound in experiment group was accelerated than that in control group from day 3. And no significant statistical difference in the thickness of granulation tissue(1540.0±118.5μm vs1504.6±131.8μm, P>0.05) and the area of vascular in newly formed granulation tissue(11.8±0.73%vs11.3±0.67%, P>0.05) were found between two groups on the 3rd day. However the area of collagen protein in experimental group increased significantly from day 3 compared to controlled group(45.7±5.3%vs11.6±2.5%,P<0.05).And the above indicators in experiment group were higher than the control group on the 7rd postoperative day(all P<0.05), and the trend was still increased on the 14 rd postoperative day(all P<0.05).Conclusion: Alginate calcium dressing had no cytotoxicity and did not induce the apoptosis of fibroblast cells, and it possessed a good cytocompatibility, meanwhile it reduced inflammatory reaction and promoted wound healing by increasing the collagen level and accelerating the formation of granulation tissue.