Influences Of Water Jet-Assisted Liposuction (WAL) Technique On The Survival Of Fat Grafts

Background:Autologous fat grafts have been considered the ideal soft tissue filler and been wildly used in the field of plastic surgery, because they are abundant, easy to obtain and have no immunological rejection after transplantation. However, with the more and more widely application of autologous fat transplantation technique, deficiencies of autologous fat grafts for soft tissue filling are exposing gradually. Part of fat grafts could not survive after transplantation is the trickiest problem which make plastic surgeons perplexed. Necrosis and absorption of fat grafts and related complications post transplantation severely impact the effect of fat transplantation, therefore, how to promote the survival of fat grafts become a hot topic in the filed of plastic surgery.Techniques for fat graft harvest, preparation and injection may cause damages to adipocytes and affect the survival of fat grafts. In order to find the optimal techinique for fat transplantation, a large amount of experiments were carried out during the last 20 years. Water jet-assisted liposuction (WAL) technique is one of fat graft harvest techniques which is more and more popular used during in recent years. Plastic surgeons consider it easy to operate, and could harvest smaller and uniform fat particles which facilitate the injection procedure. However, only few clinical and experimental studies concerned on the application of WAL technique during fat transplantation. What are differences between fat grafts harvested via WAL technique and convetional liposuction technique? Is there any survival differece between fat grafts harvested via these two techniques? What are effects of different preparation methods on the survival of WAL fat grafts? Studies on these issues are quite important for the use of WAL technique in fat transplantation.Objective:1. Comparing the effects of WAL technique and conventional liposuction technique on the survival of fat grafts.(1) Comparing the difference between fat grafts harvested via WAL technique and conventional liposuction technique in vitro.(2) Comparing the survival difference between fat grafts harvested via WAL technique and conventional liposuction technique in vivo.2. Comparing the effect of different preparation techniques on the survival of WAL fat grafts.Methods:1. Comparing the effects of WAL technique and conventional liposuction technique on the survival of fat graftsHuman fat grafts were obtained from the waists or thighs of healthy female volunteers. Fat grafts were harvested at the corresponding area of each donor side by one plastic surgeon using WAL technique (group A) and conventional manual liposuction technique (group B) respectively.(1) In vitro study on the difference between fat grafts harvested via WAL technique and conventional liposuction technique.Two groups of fat grafts harvested from each volunteer were compared. Hematoxylin-eosin staining (HE) staning were performed to compare morphological integrity of fat grafts in each group. 3-(4,5-Dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay and glucose transport test were used to compare viability of fat grafts in each group. Flow cytometry analysis were performed to compare the ratio of adipose derived stem cells in stromal vascular fraction (SVF) cells extracted from fat grafts in each group. Real-time polymerase chain reaction (PCR) assay were used to compare the adipogenic and angiogenic ability of SVF cells extracted from each group.(2) In vivo study on the survival difference between fat grafts harvested via WAL technique and conventional liposuction technique. Fat grafts were injected subcutaneously to BALB/c nude. Specimens were harvested at 1 day,7 days,14 days, 21 days,28 days and 56 days after transplantation. Specimens in each group were weighed to compare the changes of weight after transplantation. HE staning and Perilipin immunofluorescence staining were performed to compare morphological integrity of specimens in each group. CD31 immunohistochemical staining were performed to compare angiogenesis of specimens in each group. West blot technique was used to detect apoptosis of specimens in each group.2. Comparing the effect of different preparation techniques on the survival of WAL fat graftsHuman fat grafts were obtained from the waists or thighs of healthy Chinese female volunteers. Fat grafts were harvested at the corresponding area of each donor side by one plastic surgeon using WAL technique. Fluid in fat grafts were discarded after 20 minutes sedimentation. Fat grafts from one donor side were randomly assigned to sedimentation group (group A). Fat grafts from the other side were centrifuged at 1200g for 3 minutes, liquid layer and oil layer were discarded, and assigned to centrifugation group (group B). In vitro studies:HE staning were performed to compare morphological integrity of fat grafts in each group. Glucose transport test were used to compare viability of fat grafts in each group. SVF cells extracted from fat grafts were counted and compared. Flow cytometry analysis were performed to compare the number of adipose derived stem cells in each group. In vivo studies:Fat grafts were injected subcutaneously to BALB/c nude, specimens were harvested at 1 day,7 days,28 days and 56 days after transplantation. Fat specimens in each group were weighd to compare the changes of weight after transplantation. HE staning and Perilipin immunofluorescence staining were performed to compare morphological integrity of specimens in each group. CD31 immunohistochemical staining were performed to compare angiogenesis in fat specimens in each group.Results:1. The application of WAL technique benefit the survival of fat grafts.(1) Fat graftes harvested with WAL technique (group A) have more intact adipocytes and greater viability than fat grafts harvested with conventional liposuction technique (group B) (p<0.05). SVF cells extracted from fat grafts in group A contain more CD34+CD31-CD45-adipose tissue derived stem cells, and have greater gene up-regulation level of von Willebrand factor (vWF), CD31, peroxisome proliferator-activated receptor-γ(PPAR-γ) and CCAATenhancer-binding protein a (CCBP/a) after induction culture than that of group B (p< 0.05).(2) The weight of fat grafts decreased over time after transplantation in both groups. At 14,21,28, and 56 days after transplantation, specimens from group A were significantly heavier than specimens from group B (p<0.05). Specimens from group A maintain better morphological integrity. From 7 days to 56 days after transplantation, samples from group A have more perilipin-postive viable adipocytes than specimens from group B (p<0.05). From 21 days to 56 days after transplantation, samples from group A have higher percentage of intact adipocytes (p <0.05). Less apoptosis was detected from specimens in group A, from 7 days to 56 days after transplantation, specimens from group B have greater expression of Annexin V than that of group A. Angiogenesis was earlier detected in specimens from group A, from 21 days to 56 days after transplantation, specimens from group A have more vessels than that of group B (p<0.05).2. The survival of WAL fat graft was enhanced after centrifugation.There is no difference of viability between fat grafts from sedimentation group (group A) and centrifugation group (group B) (p>0.05). Liquid and oil were effectively eliminated from fat grafts after centrifuged at 1200g for 3minutes. SVF cells and ASCs were concentrated after centrifugation, fat grafts from group B have more SVF cells and ASCs than fat grafts from group A (p<0.05). Fat grafts from group B have better survival, the weight of fat specimens from group B was greater than specimens from group A at each time point (P<0.05). Specimens from group B exhibited better morphological integrity and more vessels than specimens from group A from 28 days to 56 days after transplantation (p<0.05).Conclusion:1. The application of WAL technique benefit the survival of fat graft after transplantation. WAL technique cause less damages to adipocyte than conventional liposuction technique during harvesting procedure. WAL fat grafts have greater viability and more adipose tissue derived stem cells. After transplantation, WAL fat grafts have better weight retention, greater morphological integrity, less apoptosis and greater angiogenesis.2. The survival of WAL fat grafts was enhanced after centrifugation. There is no difference of viability between fat grafts prepared by sendimentation and fat grafts prepared by centrifugation (1200g,3 minutes) after sendimentation. Liquid and oil were effectively eliminated from WAL fat grafts after centrifuged at 1200g for 3 minutes, and the number of ASCs per volume was increased. After transplantation, centrifuged WAL fat grafts had better weight retention, greater morphological integrity and greater angiogenesis.