J Korean Assoc Maxillofac Plast Reconstr Surg.  2010 Jul;32(4):299-305.

Use of Human Adipose Tissue as a Source of Endothelial Cells

• Park BW
• Hah YS
• Kim JH
• Cho HY
• Jung MH
• Kim DR
• Kim UK
• Kim JR
• Jang JH
• Byun JH

Abstract

PURPOSE
Adipose tissue is located beneath the skin, around internal organs, and in the bone marrow in humans. Its main role is to store energy in the form of fat, although it also cushions and insulates the body. Adipose tissue also has the ability to dynamically expand and shrink throughout the life of an adult. Recently, it has been shown that adipose tissue contains a population of adult multipotent mesenchymal stem cells and endothelial progenitor cells that, in cell culture conditions, have extensive proliferative capacity and are able to differentiate into several lineages, including, osteogenic, chondrogenic, endothelial cells, and myogenic lineages.
MATERIALS AND METHODS
This study focused on endothelial cell culture from the adipose tissue. Adipose tissues were harvested from buccal fat pad during bilateral sagittal split ramus osteotomy for surgical correction of mandibular prognathism. The tissues were treated with 0.075% type I collagenase. The samples were neutralized with DMEM/and centrifuged for 10 min at 2,400 rpm. The pellet was treated with 3 volume of RBC lysis buffer and filtered through a 100 microm nylon cell strainer. The filtered cells were centrifuged for 10 min at 2,400 rpm. The cells were further cultured in the endothelial cell culture medium (EGM-2, Cambrex, Walkersville, Md., USA) supplemented with 10% fetal bovine serum, human EGF, human VEGF, human insulin-like growth factor-1, human FGF-beta, heparin, ascorbic acid and hydrocortisone at a density of 1 x 10(5) cells/well in a 24-well plate. Low positivity of endothelial cell markers, such as CD31 and CD146, was observed during early passage of cells.
RESULTS
Increase of CD146 positivity was observed in passage 5 to 7 adipose tissue-derived cells. However, CD44, representative mesenchymal stem cell marker, was also strongly expressed. CD146 sorted adipose tissue-derived cells was cultured using immuno-magnetic beads. Magnetic labeling with 100 microl microbeads per 108 cells was performed for 30 minutes at 4degrees C a using CD146 direct cell isolation kit. Magnetic separation was carried out and a separator under a biological hood. Aliquous of CD146+ sorted cells were evaluated for purity by flow cytometry. Sorted cells were 96.04% positivity for CD146. And then tube formation was examined. These CD146 sorted adipose tissue-derived cells formed tube-like structures on Matrigel.
CONCLUSION
These results suggest that adipose tissue-derived cells are endothelial cells. With the fabrication of the vascularized scaffold construct, novel approaches could be developed to enhance the engineered scaffold by the addition of adipose tissue-derived endothelial cells and periosteal-derived osteoblastic cells to promote bone growth.

Keyword

Adipose tissue; Endothelial cells; Adipose tissue-derived endothelial cells

MeSH Terms

Adipose Tissue
Adult
Ascorbic Acid
Bone Development
Bone Marrow
Cell Culture Techniques
Cell Separation
Collagen
Collagenases
Drug Combinations
Durapatite
Endothelial Cells
Epidermal Growth Factor
Flow Cytometry
Heparin
Humans
Hydrocortisone
Laminin
Magnetics
Magnets
Mesenchymal Stromal Cells
Microspheres
Nylons
Osteoblasts
Osteotomy, Sagittal Split Ramus
Prognathism
Proteoglycans
Skin
Stem Cells
Vascular Endothelial Growth Factor A
Ascorbic Acid
Collagen
Collagenases
Drug Combinations
Durapatite
Epidermal Growth Factor
Heparin
Hydrocortisone
Laminin
Nylons
Proteoglycans
Vascular Endothelial Growth Factor A