Lipolysis inhibition improves the survival of fat grafts through ameliorating lipotoxicity and inflammation.

Fat grafting is a promising technique for correcting soft tissue abnormalities, but oil cyst formation and graft fibrosis frequently impede the therapeutic benefit of fat grafting. The lipolysis of released oil droplets after grafting may make the inflammation and fibrosis in the grafts worse; therefore, by regulating adipose triglyceride lipase (ATGL) via Atglistatin (ATG) and Forskolin (FSK), we investigated the impact of lipolysis on fat grafts in this study. After being removed from the mice and chopped into small pieces, the subcutaneous fat from wild-type C57BL/6J mice was placed in three different solutions for two hours: serum-free cell culture medium, culture medium+FSK (50 µM), and culture medium+ATG (100 µM). Following centrifugation to remove water and free oil droplets, 0.3 mL of the fat particles per mouse was subcutaneously injected into the back of mice. Additionally, the subcutaneous fat grafting area was immediately injected with PBS (control group), ATG (30 mg/kg), and FSK (15 mg/kg) following fat transplantation. Detailed cellular events after grafting were investigated by histological staining, real-time polymerase chain reaction, immunohistochemistry/immunofluorescent staining, and quantification. Two weeks after grafting, grafts treated with ATG showed lower expression of ATGL and decreased mRNA levels of TNFα and IL-6. In contrast, grafts treated with ATG showed elevated expression levels of IL-4 and IL-13 compared to the control grafts. In addition, fewer apoptotic cells and oil cysts were observed in ATG grafts. Meanwhile, a higher CD206+/CD68+ ratio of macrophages and more CD31+ vascular endothelial cells existed in the 2-month ATG grafts. In comparison to the control, ATG treatment improved the volume retention of grafts, and decreased graft fibrosis and oil cyst formation. By preventing oil droplet lipolysis, pharmacological suppression of ATGL shielded adipocytes from lipotoxicity following grafting. Additionally, ATG ameliorated the apoptosis and inflammation brought on by adipocyte death and oil droplet lipolysis in grafted fat. These all indicate that lipolysis inhibition improved transplanted fat survival and decreased the development of oil cysts and graft fibrosis, offering a potential postoperative pharmacological intervention for bettering fat grafting.

Composite Microparticles of Fat Graft and GFR Matrigel Improved Volume Retention by Promoting Cell Migration and Vessel Regeneration.

BACKGROUND: The retention volume of autologous fat grafts decreases after transplantation due to limited nutrition infiltration and insufficient blood supply. Structural fat grafts and the 3M (multipoint, multitunnel, and multilayer) injection technique have been considered to improve the survival of grafts; however, it is difficult for surgeons to practice in the clinic because grafts tend to gather into a cluster, especially in large volume fat grafting. Therefore, we hypothesize that prefabricated microparticle fat grafts (PFMG) may improve the retention rate. METHODS: The C57BL/6 mouse fat particles were embedded in growth factor-reduced (GFR)-Matrigel to detect cell migration by immunofluorescence staining in vitro. PFMG was prepared by mixing mouse fat particles and GFR Matrigel in a 1:1 volume ratio and injected subcutaneously into C57BL/6 mice. Fat particles mixed with PBS in equal volume served as control group. The grafts were harvested at 1, 4, 8, and 12 weeks after sacrifice. The retention rate of grafts at each time point was measured, and the structural alterations were detected by SEM. Fat necrosis and blood vessel density were evaluated by histological analysis. RESULTS: CD34+ cells are migrated from the PFMG and formed a tree-like tubular network in the in vitro study. The retention rate was higher in the PFMG group than in the control group at week 12 (38% vs. 30%, p < 0.05). After transplantation, the dissociated structure of fat particles was maintained in PFMG by SEM analysis. Histological analysis of PFMG confirmed less fat necrosis and more blood vessel density in the PFMG group at the early stage than in the control group. The GFR Matrigel was displaced by adipose tissue with time. CONCLUSIONS: In this study, we developed a novel fat grafting method, PFMG that dispersed fat grafts and maintained the structure after transplantation. High volume retention volume of PFMG was achieved by promoting cell migration and vessel regeneration. NO LEVEL ASSIGNED: This journal requires that authors assign a level of evidence to each submission to which Evidence-Based Medicine rankings are applicable. This excludes Review Articles, Book Reviews, and manuscripts that concern Basic Science, Animal Studies, Cadaver Studies, and Experimental Studies. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Preoperative Short-term High-Carbohydrate Diet Provides More High-Quality Transplantable Fat and Improves the Outcome of Fat Grafts in Mice.

BACKGROUND: Patients with a low BMI may have inadequate high-quality adipose tissue for transplantation. The influence of high-energy diets on adipose tissue and graft retention remains unknown. OBJECTIVES: The authors explored inguinal fat pad alternation in mice fed on a short-time high-fat diet (HFD) or a high-carbohydrate diet (HCD) preoperatively and the morphological and histological differences after transplantation. METHODS: Mice were fed HFD (60% kcal from fat, 20% from carbohydrate), HCD (9.3% kcal from fat, 80.1% from carbohydrate), or normal (12% kcal from fat, 67% kcal from carbohydrate) diets for 2 or 4 weeks. Histological analyses were carried out following hematoxylin and eosin staining as well as CD34 and proliferating cell nuclear antigen immunostaining. The uncoupling protein-1 expression was determined by western blotting. Fat pads from each group were grafted into the dorsal region of the recipient mice, and morphological and histological changes were determined 4, 8, and 12 weeks posttransplantation. Vascular endothelial growth factor-α and platelet-derived growth factor-α expression were determined using quantitative polymerase chain reaction. RESULTS: The inguinal fat pad volume increased in the HFD and HCD groups. The presence of multilocular adipocytes in inguinal fat of HCD-fed mice, combined with the increased uncoupling protein-1 content, suggested adipocyte browning. HCD grafts showed higher volume retention and reduced oil cyst formation, possibly attributed to better angiogenesis and adipogenesis. CONCLUSIONS: HCD enlarged adipose tissue and improved graft survival rates, which may be due to the browning of fat before grafting and enhanced angiogenesis after grafting.

A Short-Term High-Fat Diet Improved the Survival of Fat Grafts in Mice by Promoting Macrophage Infiltration and Angiogenesis.

Background: Adipose tissue is an ideal filler material that is widely used for soft tissue defects. But the low survival rate and complications associated with such grafts pose a serious challenge, which limits their clinical application. Adipose tissue is a metabolic diet-responsive tissue; however, the influence of diets on fat grafting remains ambiguous. Methods: We extracted inguinal fat pads from C57/BL6 male mice, and transplanted them into the dorsal region of recipient mice (0.3 ml). Post-fat-grafting, mice (n = 54) were randomized into three groups, namely normal diet (ND), high carbohydrate diet (HC), and high-fat diet (HF). Structural changes were assessed by histological staining. Lipolysis activity and vascular regeneration of grafts on day 30 were analyzed using real-time polymerase chain reaction, immunofluorescence, and western blotting. Results: The grafts of mice on HC and HF diets exhibited significantly fewer oil cysts and larger volume retention (0.18 ± 0.01, 0.21 ± 0.01, and 0.25 ± 0.01 ml, for ND, HC, and HF group, respectively, p < 0.05) on day 90. In comparison, grafts for the mice belonging to the HF groups exhibited higher expression of lipolysis-related genes, including adipose triglyceride lipase (ATGL), hormone-sensitive lipase (HSL), and carnitine palmitoyltransferase 1 (CPT1), on day 30. Furthermore, increased infiltration of macrophages (F4/80+) and the higher expression of angiogenesis genes were reported in the HF groups. Conclusion: Altogether, the administration of short-term HF diet remarkably enhanced angiogenesis and improved the quality of fat grafts, which was characterized by fewer oil cysts and higher long-term volume retention. The possible mechanisms may be due to the increased macrophage infiltration, and the promoted angiogenesis in HF grafts.