A Method For Increasing Revascularization After Grainy Fat Transplantation By Using An Adipose-Derived Stem Cells In Combination With Platelet-rich Plasma Transplantation Therapy

Objective: To examine whether the rabbit adipose-derived stem cells (ADSCs)either alone or in combination with the platelet-rich plasma (PRP) can improve therevascularization after grainy fat transplantation.Methods:1. Adapted the type Ⅰ collagenase digestion method to isolate andculture the ADSCs on cervicodorsal area fat pads of three-month old New Zealandrabbits.2. Induced the third generation ADSCs into adipose cells or osteoblasts, whichwere identified by the oil red O or alizarin red vital staining method to enable themultiplex differentiation potential of the ADSCs.3. Exsanguinated hearts of the New Zealand rabbits, adapted the secondarycentrifugation on the exsanguinated blood to obtain the PRP, and then recorded thenumber of platelets.4. Detect, used the MTT method, the PRP, plasma and a control group at differentpoints of time (the24th,48th and72th hour).5. Randomly numbered28athymic mice, randomly divided them into four groupseach comprising seven mice. Marked five points on the back of each mouse: upper left,lower left, upper right, lower right, and the center (spacing between any two points isgreater than2cm), and randomly numbered the above five points. Transplanted thefollowing five group grafts into the subcutaneous tissues of the28athymic mice at theabove five points, respectively, where the five group grafts were:①lipochondrias+ADSCs+PRP+spongia gelatinosas;②lipochondrias+ADSCs+saline+spongia gelatinosas;③lipochondrias+PRP+saline+spongia gelatinosas;④lipochondrias+whole plasma+saline+spongia gelatinosas;⑤lipochondrias+saline+spongiagelatinosas, where the lipochondrias and spongia gelatinosas were prepared in advance.6. Procured the above five group grafts from the subcutaneous tissue of theathymic mice on the7th,14th,30th and90th day, respectively, and paraffinized theminto paraffin sections. Observed the paraffin sections and counted the number ofcapillaries using the HE staining method, and the immunohistochemical method.Results:1. The ADSCs grew in the form of the collagenoblasts, there was noobvious variation in the continuous19generations cells. Adipose cells were inductedafter two weeks, the oil red O attaining results were positive. The osteoblasta wereinducted after three weeks, results of the alizarin red staining were positive. The resultsof the control group staining were negative.2. The number of the whole blood platelets was:(330.2±148.8)×109/L, the numberof the PRP platelets was:(1398.3±888.5)×109/L, the number of the PRP platelets was4.23times of that of the whole blood platelet.3. The results of the MTT method showed that the PRP speeded up theproliferation of the ADSCs which was statistically different from that of the plasma andthe control group (p≤0.05).4. The ADSCs in combination with the PRP transplantations therapy producedsignificantly more capillaries than other transplantation therapies, and subjected to lessfat necrosis and fibrosis as revealed by the histological analysis. Also the ADSCs andthe PRP in alone could bring a similar effect which was however not as good as thecombination thereof.Conclusions: The ADSCs in combination with the PRP can boost therevascularization after grainy fat transplantation.