Spontaneous Browning of White Adipose Tissue Improves Angiogenesis and Reduces Macrophage Infiltration After Fat Grafting in Mice

Background: Fat grafting is a frequently used technique; however, its survival/ regeneration mechanism is not fully understood. The browning of white adipocytes, a process initiated in response to external stimuli, is the conversion of white to beige adipocytes. The physiologic significance of the browning of adipocytes following transplantation is unclear.Methods: C57BL/6 mice received 150 mg grafts of inguinal adipose tissue, and then the transplanted fat was harvested and analyzed at different time points to assess the browning process. To verify the role of browning of adipocytes in fat grafting, the recipient mice were allocated to three groups, which were administered CL316243 or SR59230A to stimulate or suppress browning, respectively, or a control group after transplantation.Results: Browning of the grafts was present in the center of each as early as 7 days post-transplantation. The number of beige cells peaked at day 14 and then decreased gradually until they were almost absent at day 90. The activation of browning resulted in superior angiogenesis, higher expression of the pro-angiogenic molecules vascular endothelial growth factor A (VEGF-A) and fibroblast growth factor 21 (FGF21), fewer macrophages, and ultimately better graft survival (Upregulation, 59.17% ± 6.64% vs. Control, 40.33% ± 4.03%, *p < 0.05), whereas the inhibition of browning led to poor angiogenesis, lower expression of VEGF-A, increased inflammatory macrophages, and poor transplant retention at week 10 (Downregulation, 20.67% ± 3.69% vs. Control, 40.33% ± 4.03%, *p < 0.05).Conclusion: The browning of WAT following transplantation improves the survival of fat grafts by the promotion of angiogenesis and reducing macrophage

Fat Grafting for Rejuvenation and Regeneration with Stromal Vascular Fraction Gel

SUMMARY: Lipotransfer has become a powerful regenerative tool, largely because of its cellular components, the stromal vascular fraction (SVF). However, the clinical separation of cells with collagenase is strictly legislated. In 2017, Yao et al. postulated a novel fat-derived product mechanically concentrating SVF cells and an extracellular matrix (ECM) and named it stromal vascular fraction gel (SVF-gel). This review discussed the protocol of SVF-gel and its component as well as its inner structure. The histologic examination and the retention rate after the transplantation of SVF-gel were also rendered. Moreover, we summed up the rejuvenating and regenerative use of SVF-gel and introduced its possible mechanism

In vivo dedifferentiation of adult adipose cells.

Adipocytes can dedifferentiate into fibroblast-like cells in vitro and thereby acquire proliferation and multipotent capacities to participate in the repair of various organs and tissues. Whether dedifferentiation occurs under physiological or pathological conditions in vivo is unknown.A tissue expander was placed under the inguinal fat pads of rats and gradually expanded by injection of water. Samples were collected at various time points, and morphological, histological, cytological, ultrastructural, and gene expression analyses were conducted. In a separate experiment, purified green fluorescent protein+ adipocytes were transplanted into C57 mice and collected at various time points. The transplanted adipocytes were assessed by bioluminescence imaging and whole-mount staining.The expanded fat pad was obviously thinner than the untreated fat pad on the opposite side. It was also tougher in texture and with more blood vessels attached. Hematoxylin and eosin staining and transmission electron microscopy indicated there were fewer monolocular adipocytes in the expanded fat pad and the morphology of these cells was altered, most notably their lipid content was discarded. Immunohistochemistry showed that the expanded fat pad contained an increased number of proliferative cells, which may have been derived from adipocytes. Following removal of the tissue expander, many small adipocytes were observed. Bioluminescence imaging suggested that some adipocytes survived when transplanted into an ischemic-hypoxic environment. Whole-mount staining revealed that surviving adipocytes underwent a process similar to adipocyte dedifferentiation in vitro. Monolocular adipocytes became multilocular adipocytes and then fibroblast-like cells.Mature adipocytes may be able to dedifferentiate in vivo, and this may be an adipose tissue self-repair mechanism. The capacity of adipocytes to dedifferentiate into stem cell-like cells may also have a more general role in the regeneration of many tissues, notably in fat grafting

Rat fat pad expantion.

<p>Left: Rats after transplantation of the expander. Middle: The expanded fat pad became thinner (white arrows). Right: After removal of the expander, the regrowth of adipose-liked tissue (yellow arrows) was seen. Scale bar = 2 cm.</p

Histology observation of the fat pad.

<p>Normal adipocytes were uniform in size and had a normal morphology. During expansion, the growth of connect tissue (white arrow) and the reduced volume of adipocytes (yellow arrow) were observed. After removing the expander, many adipocytes of small sizes and a normal morphology emerged around the new vessels (red arrow). Scale bar = 200 μm.</p

Morphology change of GFP+ adipocytes during dedifferention.

<p>A: monolocular mature adipocytes (white arrow) with GFP signal at day 0. After 7 days ceiling culture, cells with multilocular lipid droplets were observed (yellow arrow). 10 days later, the cells had a fibroblast-like morphology with no visible fat droplets. Scale bar = 100 μm. B: The adipogenic (left), osteogenic (middle) and chondrogenic (right) differentiation of DFAT cells. Scale bar = 25 μm.</p

Fate of GFP+ adipocytes after transplantation.

<p>On Day 1 after transplantation, some GFP+ adipocytes exhibited membrane contraction (blue arrow). On Day 7, some surviving GFP+ adipocytes had multilocular lipid droplets (white arrow). On Day 21, very few adipocytes survived and those had completely discarded their lipid droplets and changed into fibroblast-like cells (red arrow). Scale bar = 100 μm.</p

The retention of GFP+ adipocytes after transplantation.

<p>On Day 1 after transplantation, there was intense green fluorescent protein fluorescence (red arrow) in the injected region. On Day 21 after transplantation, there were only a few scattered regions of fluorescence.</p

The expression level of adipogenic gene.

<p>During expansion, PPARγ, adiponectin and C/EBPα expression levels decreased gradually. The expression level of them increased significantly after expander removel (*P<0.05).</p

The proliferation and angiogenesis of the fat pad.

<p>A: The number of Ki67+ cells (white arrow) was increased during expension (day 0 left, day 14 right). Scale bar = 50 μm. B: Some of the Ki67+ cells (red arrow) were also perilipin+ (yellow arrow). Scale bar = 10 μm. C: The number of blood vessels (white arrow) did not decrease significantly, and when removing the expander, the angiogenic level increased. Scale bar = 25 μm.</p