Engineering adipose tissue from uncultured human adipose stromal vascular fraction on collagen matrix and gelatin sponge scaffolds.
Adipogenic potential was evaluated in uncultured stromal vascular fraction (SVF) loaded onto porous 3D collagen matrix and gelatin sponge scaffolds with predefined shapes. The SVF was isolated from 16 freshly lipectomized fat. Mean cell number was 6.0±4.68×10(7) cells/mL, and mean cell viability was 72%. Flow cytometric analysis revealed adipose-derived stromal cells (CD31(-), CD34(-/+), CD45(-), CD90(+), CD105(-), CD146(-), and CD166(+)) in the SVF. Three hours after harvest of fat, 200  μL of isolated SVF was loaded onto an experimental scaffold (4  mg in weight) and cultured in Dulbecco's modified Eagle's medium. Examination of the construct under an inverted light microscope and a scanning electron microscope demonstrated adequate seeding and active proliferation of the SVF cells on pore surfaces. Cells grew to varying sizes in clusters or in strands. On day 28, histologic study of the constructs by H&E staining revealed viable adipocytes in the microstructure. Positive Oil-Red O stain confirmed lipid accumulation in mature adipocytes. The presence of human adipocytes was further assessed by the presence of genomic DNA detected by GAPDH gene and by the mRNA expression of peroxisome proliferator activated receptor-γ, and lipoprotein lipase. The results demonstrated that new adipose tissue can be regenerated by seeding freshly isolated, uncultured SVF on 3D porous collagen matrix and gelatin sponge scaffolds.
Division of Plastic Surgery, Department of Surgery, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan. sidalin@kmu.edu.tw
Sin-Daw
SD
Lin
Shu-Hung
SH
Huang
Yun-Nan
YN
Lin
Sheng-Hua
SH
Wu
Hsueh-Wei
HW
Chang
Tsai-Ming
TM
Lin
Chee-Yin
CY
Chai
Chung-Sheng
CS
Lai
D017667:Adipocytes / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D000273:Adipose Tissue / Q000098:blood supply* / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D000328:Adult; D000818:Animals; D001391:Azo Compounds / Q000378:metabolism; D001808:Blood Vessels / Q000166:cytology*; D002452:Cell Count; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003094:Collagen / Q000494:pharmacology* / Q000648:ultrastructure; D005109:Extracellular Matrix / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D005260:Female; D005434:Flow Cytometry; D005780:Gelatin / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D008875:Middle Aged; D009928:Organ Specificity / Q000187:drug effects; D016062:Porosity / Q000187:drug effects; D012038:Regeneration / Q000187:drug effects; D017154:Stromal Cells / Q000166:cytology / Q000187:drug effects; D034421:Sus scrofa; D023822:Tissue Engineering / Q000379:methods*; D054457:Tissue Scaffolds / Q000737:chemistry*
D001391:Azo Compounds; D005780:Gelatin; D003094:Collagen; C011049:oil red O