Experimental study on the best concentration of SVFs for promoting survival rate of fat graft / 中华整形外科杂志

<p><b>OBJECTIVE</b>To investigate the effect of different concentrations of human adipose stromal vascular fraction cells (SVFs) on the survival rate of fat transplantation.</p><p><b>METHODS</b>0.3 ml fat tissue, derived and refined from clinical liposuction patients, was mixed with different concentrations of SVFs as 5 x 10(5)/ml in Group A, or 1 x 10(6)/ml in Group B, or 2 x 10(6)/ml in Group C, or completely medium in control group D. Then the mixture was injected randomly under the back skin of 6 nude mice. The transplanted fat tissue in four groups was harvested at 3 months after implantation. Wet weight of fat grafts was measured for macroscopic aspects. After HE staining, blood vessel density, viable adipocytes and fibrous proliferation were counted respectively for histological evaluation.</p><p><b>RESULTS</b>The wet weight of fat grafts in group B (81.670 +/- 7.528) mg was significantly higher than that in group A, C, D [(60.000 +/- 6.325) mg, (68.330 +/- 7.528) mg, (48.330 +/- 7.528) mg, respectively, P < 0.05)], but the difference between group A and group C was not statistically significant (P > 0.05). The grafts in group A, B and C had significantly higher blood vessel density than those in the control group D, whereas blood vessel density was the highest in group B (P < 0.05) and there was no significant difference between group A and C (P > 0.05). Compared with group A, C and D, histological analysis revealed that the fat grafts in group B was consisted predominantly of adipose tissue with less fat necrosis and fibrosis (P < 0.05). However, fibrosis counts were significant lower in group A, B and C than those in group D (P < 0.05), and there was no significant difference between group A and C (P > 0.05).</p><p><b>CONCLUSIONS</b>The human isolated SVFs has the advantages to improve the survival rate of fat transplantation, and the magnitude of 1 x 10(6)/ml is more practical and safe, indicating a wide clinical application in the future.</p>

The cellular plasticity of human adipocytes / 中华整形外科杂志

<p><b>OBJECTIVE</b>To explore the dedifferentiation phenomenon of human mature adipocytes cultured in vitro and to discuss the possibility of using dedifferentiation adipocytes (DA) as seed cells.</p><p><b>METHODS</b>Mature adipocytes and ASCs were harvested from human fat aspirates. Mature adipocytes were cultured and induced to DA by ceiling adherent culture method. Cell morphology were observed during the whole process. Viabilities of DA and ASCs were compared by MTT chromatometry and cell growth curves were drawn based on it. Cell surface markers of DA and ASCs were detected by flow cytometry. The adipogenic, osteogenic and chondrogenic ability of DA and ASCs were assessed by oil red O staining, alizarin bordeaux staining and alcian blue staining, respectively.</p><p><b>RESULTS</b>Human mature adipocytes can dedifferentiate into fibroblast-shaped DA. MTT chromatometry assay demonstrated that DA and ASCs both had strong reproductive activity, with no significant difference between them. Flow cytometry assay demonstrated that both DA and ASCs expressed HLA-ABC, CD29 and CD44, while didn't express CD45, CD34 and CD106. After two weeks of adipogenic differentiation, lipid droplets could be displayed by oil red O staining in both DA and ASCs. After two weeks of osteogenic differentiation, calcium salts mineralization in DA and ASCs could be detected by alizarin bordeaux staining. After two weeks of chondrogenic differentiation, matrix of cartilage cells in DA and ASCs could be detected by alcian blue staining.</p><p><b>CONCLUSIONS</b>Mature adipocytes can be dedifferentiated into DA in vitro. DA has strong reproductive activity, as well as osteogenic, chondrogenic ability and strong adipogenic ability. It expresses some of the stem cell-related cell surface proteins and is a promising seed cell for adipose tissue engineering.</p>

Preliminary comparison study of adipogenic differentiation capacity between dedifferentiated adipocytes cells and adipose-derived stem cells in vivo / 中华整形外科杂志

<p><b>OBJECTIVE</b>To compare the adipogenic differentiation capacity of dedifferentiated adipocytes cells (DA) and adipose-derived stem cells (ASCs) in vivo, so as to select good adipogenic seed cells for tissue engineering.</p><p><b>METHODS</b>Mature adipocytes and ASCs were isolated by means of enzymatic digestion from the liposuction aspirate. Then the DA cells were acquired by ceiling adherent culture of mature adipocytes and the 3rd passage cells were used. The DA cells and ASCs were cultured with fibrin glue in vitro respectively. The compatibility of scaffold with cells was detected by microscopy and scanning electron microscopy. The scaffold-cell composite was also labeled by DiI. The composite was injected subcutaneously on the nude mice back, respectively (DA-FG group, n = 8; ASCs-FG group, n = 8; sham FG group, n = 8). 8 weeks after implantation, the newly formed tissue was taken out for general observation and histologic study.</p><p><b>RESULTS</b>Mature adipocytes were transferred to DA cells with spindle shape, like fibroblast. The ASCs were also spindle. Three days after culture of cell-scaffold composite in vitro, the cells grew well. 8 weeks after implantation, the newly formed tissue was found under the skin both in DA-FG and ASCs-FG groups, but not in sham FG group. The newly formed tissue was mature fat tissue and originated from the seed cells. The average wet weight of the new-formed tissue was higher in DA-FG group than that in ASCs-FG group. The average fibrosis ratio was lower in DA-FG than in ASCs-FG group.</p><p><b>CONCLUSIONS</b>The tissue-engineered adipose tissue can be achieved with DA cells and ASCs as seed cells. Compared with ASCs, the new-formed fat tissue with DA has a higher wet weight and lower fibrosis ratio.</p>

Experimental study of human adipocyte dedifferentiation for adipose tissue engineering / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the dedifferentiation of mature adipocytes and the possibility of adipose tissue engineering using dedifferentiated adipocytes.</p><p><b>METHODS</b>Human adipose tissue was harvested from healthy women undergoing abdominal liposuction procedures, and mature adipocytes were isolated with enzymatic digestion and cultured by ceiling adherent culture method, using the third-passage cells for subsequent experiment. The cells were cultured in adipogenic, chondrogenic or osteogenic media, and Oil red-O staining, Alcian blue staining and Alizarin red staining were used for dedifferentiation identification. The third-passage cells labeled with DiI were mixed with fibrin glue and injected subcutaneously on one side of the nude mouse back (n=6), and fibrin glue solution without cells as control was injected on the other side (n=6). After 8 weeks of cell implantation, the specimens were harvested for general observation and histological, Oil red-O staining and fluorescent microscope analyses.</p><p><b>RESULTS</b>Mature adipocytes were round, unilocular cells. After ceiling adherent culture, the adipocytes underwent morphological changes into fibroblast-like cells indicating their dedifferentiation. The dedifferentiated adipocytes were induced for adipogenic, chondrogenic and osteogenic differentiation in specified media. Eight weeks after the cell injection in nude mice, newly formed tissue occurred which was identified as mature adipose tissue. The implants without cells were completely absorbed in the control group.</p><p><b>CONCLUSION</b>Mature adipocytes can dedifferentiate in vitro culture, and the dedifferentiated adipocytes are capable of differentiating into adipogenic, chondrogenic and osteogenic lineages. Adipose tissue engineering can be achieved in vivo using the dedifferentiated adipocytes as the seed cells.</p>

Migration of intravenously injected adipose tissue-derived stem cells in SD rats with soft tissue wound / 中华整形外科杂志

<p><b>OBJECTIVE</b>To explore the migration of intravenously injected adipose tissue-derived stem cells (ADSCs) in SD rats with soft tissue wound.</p><p><b>METHODS</b>1.8 cm x 1.8 cm full thickness skin defect and 0.5 cm in depth soft tissue defects were made on the back of 6 SD rats. ADSCs were isolated and cultured in vitro for 3 passages. 2.4 x 10(6) ADSCs were labeled with DiI and transplanted into the SD rats through tail vein. Normal skin and wound tissue samples were collected for fluorescent distribution observation 24, 48 days after injection, respectively.</p><p><b>RESULTS</b>Compared to normal skin, more fluorescent positive cells were detected in the margin and deep layer of the wound 24 days after operation. But it is accumulated within dermis and adenoid 48 days after operation.</p><p><b>CONCLUSION</b>Wound can probably induce the migration and accumulation of intravenously transplanted ADSCs.</p>

Molecular clone of adipose-derived stromal cells with high potential of adipogenic differentiation / 中华整形外科杂志

<p><b>OBJECTIVE</b>To investigate the surface markers of adipose-derived stromal cells (ASCs) with high potential of adipogenic differentiation for increasing efficiency of adipose tissue engineering with selected seeds cells.</p><p><b>METHODS</b>ASCs were harvested from human adipose tissue by collagenase digestion. After proliferation and adipogenic induction of ASCs, the mature induced adipocytes floating in the induction medium were collected. Ceiling culture was used to culture adipocytes and then dedifferentiated adipocytes was obtained at the ceiling. The reproductive activity, adipogenic differentiation potency and expression of surface markers were compared between the dedifferentiated adipocytes and ASCs.</p><p><b>RESULTS</b>The reproductive activity between the dedifferentiation adipocytes and ASCs were similar. The potential of adipogenic differentiation of the dedifferentiated adipocytes was stronger than that of the ASCs. The expression of cell surface markers of both cells were almost the same. But the CD54 positive expression in dedifferentiated adipocytes was higher than that in ASCs.</p><p><b>CONCLUSION</b>The CD54 expression maybe closely associated with high potential of adipogenic differentiation of dedifferentiated adipocytes. CU54 maybe the specific identification of cell surface marker of ASCs with high adipogenic differentiation.</p>

Study of adipose tissue engineering with human adipose-derived stem cells and collagen type I scaffold / 中华整形外科杂志

<p><b>OBJECTIVE</b>To explore the possibility of building tissue-engineered adipose tissue and looking for a new approach for the repair of soft tissue defects.</p><p><b>METHODS</b>The cells using enzymatic digestion from human liposuction part of the lipid extract were used as adipose tissue-derived cells and labeled with DiI fluorescent marker, the induced group using I collagen scaffold material as a carrier, the induced cell were planted into left back subcutaneously in nude mice at 1 x 10(7)/ml cell density, in the uninduced group cells were not induced by any, in the same cell density and type I collagen scaffold composite inoculated in nude right mouse back skin, the blank control group I collagen scaffold gaps in nude mice inoculated subcutaneously center of the neck, each of the six mice; Remove implants after 12 weeks and judge the adipogenic capacity through general and fluorescence microscopy, wet - determination, histological detection and oil red O staining qualitative.</p><p><b>RESULTS</b>The primary source of fat cultured stem cells, similar to the fibroblast morphology, and has a strong proliferative capacity. In the role of adipose differentiation medium, it can be the mature fat cells in which cytoplasmic lipid droplets gather, oil red O staining was positive. In the induced group, newborn tissue were found in the experimental groups of nude mice and its average weight is about 0.020 g. Conventional pathological slices and oil red O staining confirmed it is mature adipose tissue, the fluorescence staining positive confirm them are exogenous. Uninduced group newborn tissue are found in the experimental groups of nude mice and its average weight is about 0.014 g. Conventional pathological slices and oil red O staining confirmed it include some mature adipose tissue, the fluorescence staining positive confirm them are exogenous. Two groups of the new wet weight with have statistical significance (P < 0.01); gaps in the control group no new organization formed.</p><p><b>CONCLUSIONS</b>The cells using enzymatic digestion from human liposuction part of the lipid extract are adipose tissue-derived cells. The cells can be as seed cells and with solid scaffold of collagen type I it can become fat tissue in vivo successfully.</p>

Effect of hypoxia on dedifferentiation of mature adipocytes: an experimental study / 南方医科大学学报

<p><b>OBJECTIVE</b>To study the morphological changes of mature adipocytes in hypoxic condition in vitro and investigate the effect of hypoxia on dedifferentiation of mature adipocytes.</p><p><b>METHODS</b>Human adipose tissue was collected from individuals undergoing liposuction procedures, from which the stem cells were isolated and induced to become mature adipocytes. After cell culture in hypoxic condition, the adipocytes were observed for morphological changes at different time points.</p><p><b>RESULTS</b>After a few days of culture in hypoxic condition, the mature adipocytes derived from the adipose tissue-derived stem cells underwent morphological changes from spherical cells into fibroblast-like cells capable of continuous cell division.</p><p><b>CONCLUSION</b>Mature adipocytes may dedifferentiate into fibroblast-like cells under appropriate conditions.</p>

Cellular compatibility of type collagen I scaffold and human adipose-derived stem cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To study the cellular compatibility of type I collagen scaffold and human adipose-derived stem cells (ADSC(S)) in order to explore appropriate scaffold materials for adipose tissue engineering.</p><p><b>METHODS</b>The morphology and function of the ADSC(S) were observed by inverted phase contrast microscope, scanning electron microscope and XTT assay when cocultured type I collagen scaffold with ADSC(S) in vitro. Cells adhesive rates were also calculated.</p><p><b>RESULT</b>ADSC(S) were able to attach, grow and proliferate well on the scaffolds.</p><p><b>CONCLUSION</b>collagen I scaffold exhibits excellent cellular compatibility and can be used as a vehicle for adipose tissue engineering.</p>