Much of the work conducted on adult stem cells has focused on mesenchymal stem cells (MSCs) found within the bone marrow stroma. Adipose tissue, like bone marrow, is derived from the embryonic mesenchyme and contains a stroma that is easily isolated. Preliminary studies have recently identified a putative stem cell population within the adipose stromal compartment. This cell population, termed processed lipoaspirate (PLA) cells, can be isolated from human lipoaspirates and, like MSCs, differentiate toward the osteogenic, adipogenic, myogenic, and chondrogenic lineages. To confirm whether adipose tissue contains stem cells, the PLA population and multiple clonal isolates were analyzed using several molecular and biochemical approaches. PLA cells expressed multiple CD marker antigens similar to those observed on MSCs. Mesodermal lineage induction of PLA cells and clones resulted in the expression of multiple lineage-specific genes and proteins. Furthermore, biochemical analysis also confirmed lineage-specific activity. In addition to mesodermal capacity, PLA cells and clones differentiated into putative neurogenic cells, exhibiting a neuronal-like morphology and expressing several proteins consistent with the neuronal phenotype. Finally, PLA cells exhibited unique characteristics distinct from those seen in MSCs, including differences in CD marker profile and gene expression.

Our laboratory has recently characterized a population of cells from adipose tissue, termed processed lipoaspirate (PLA) cells, which have multi-lineage potential similar to bone-marrow-derived mesenchymal stem cells (MSCs). This study is the first comparison of PLA cells and MSCs isolated from the same patient. No significant differences were observed for yield of adherent stromal cells, growth kinetics, cell senescence, multi-lineage differentiation capacity, and gene transduction efficiency. Adipose tissue is an abundant and easily procured source of PLA cells, which have a potential like MSCs for use in tissue-engineering applications and as gene delivery vehicles.

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Tissue engineering offers considerable promise in the repair or replacement of diseased and/or damaged tissues. The cellular component of this regenerative approach will play a key role in bringing these tissue engineered constructs from the laboratory bench to the clinical bedside. However, the ideal source of cells still remains unclear and may differ depending upon the application. Current research for many applications is focused on the use of adult stem cells. The properties of adult stem cells that make them well-suited for regenerative medicine are (1) ease of harvest for autologous transplantation, (2) high proliferation rates for ex vivo expansion and (3) multilineage differentiation capacity. This review will highlight the use of adipose tissue as a reservoir of adult stem cells and draw conclusions based upon comparisons with bone marrow stromal cells.

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Human processed lipoaspirate (PLA) cells are multipotent stem cells, capable of differentiating into multiple mesenchymal lineages (bone, cartilage, fat, and muscle). To date, differentiation to nonmesodermal fates has not been reported. This study demonstrates that PLA cells can be induced to differentiate into early neural progenitors, which are of an ectodermal origin. Undifferentiated cultures of human PLA cells expressed markers characteristic of neural cells such as neuron-specific enolase (NSE), vimentin, and neuron-specific nuclear protein (NeuN). After 2 weeks of treatment of PLA cells with isobutylmethylxanthine, indomethacin, and insulin, about 20 to 25 percent of the cells differentiated into cells with typical neural morphologic characteristics, accompanied by increased expression of NSE, vimentin, and the nerve-growth factor receptor trk-A. However, induced PLA cells did not express the mature neuronal marker, MAP, or the mature astrocyte marker, GFAP. It was also found that neurally induced PLA cells displayed a delayed-rectifier type K+ current (an early developmental ion channel) concomitantly with morphologic changes and increased expression of neural-specific markers. The authors concluded that human PLA cells might have the potential to differentiate in vitro into cells that represent early progenitors of neurons and/or glia.

The FDA has approved the clinical use of recombinant bone morphogenetic proteins (BMPs). However, the use of recombinant BMPs in humans has required large doses of the proteins to be effective, which suggests that the delivery method of bone morphogenetic proteins needs to be optimized. Gene therapy is an alternative method to deliver such recombinant proteins, and gene transfer techniques have been tested on a variety of cell types including bone marrow cells, skin fibroblasts, peripheral blood monocytes, and muscle-derived cells. In this study, we sought to determine the ability of BMP-2-producing human adipose-derived mesenchymal stem cells to heal a critically sized femoral defect in a nude rat model. After approval by the human subjects protection committee, human adipose tissue was obtained from healthy donors. The lipoaspirate was processed as previously described (De Ugarte, D.A., et al. Cells Tissues Organs 174, 101, 2003). Cells were grown in culture and infected with a BMP-2-carrying adenovirus. Five million cells were applied to a collagen- ceramic carrier and implanted into femoral defects as previously described (Zuk, P.A., et al. Mol. Biol. 13, 4279, 2002). All animals were killed at 8 weeks. Femora were dissected out and underwent radiographic, histologic, and biomechanical analysis. Eleven of the 12 femora in the group treated with human processed lipoaspirate (HPLA) cells genetically modified to overexpress BMP-2 had healed at 8 weeks. This was assessed by radiographs, by mechanical testing, and by histology. The one femur that did not heal had a subacute infection. All eight of the femora treated with the rhBMP-2-impregnated collagen-ceramic carrier healed. No statistically significant difference was detected between these two groups. Evaluation of the control groups: group II (collagen- ceramic carrier with HPLA cells) and group III (collagen-ceramic carrier alone) showed that none of the femora had healed by 8 weeks. Our results indicate that HPLA cells genetically modified by adenoviral gene transfer to overexpress BMP-2 can induce bone formation in vivo and heal a critically sized femoral defect in an athymic rat. The HPLA cells alone did not induce significant bone formation. However, when combined with an osteoinductive factor these cells may be an effective method for enhancing bone healing and the tissue engineering of bone.

Puvpos~: We have isolated pluripotent mesenchymal progenitor cells in large numbers from liposuction aspirates (processed lipoaspirate cells or PLAs). This study examines the osteogenic potential of PLAs and bone marrow aspirate cells (BMAs), when exposed to either recombinant human bone morphogenetic protein (BMPI-2 (rh-BMP-2) or adenovirus containing BMP-2 cDNA Mrthoulc: Liposuction aspirates underwent proteolytic digestion to obtain PLAs. After exposure to exogenous rh-BMP-2 or Ad-BMP-2 for four or seven days, PLAs and BMAs were assessed by histochemistry, spectrophotometry and RT-PCR. Western blotting and ELISA confirmed BMP gene transduction. Results were compared to osteoblasts and cells in osteogenic media only. PLA-Ad-BMP-2 cells were seeded on matrices and implanted in the hind limbs of SCID mice.Results: Analysis of quantified bone precursor assays including extracellular ALP histomorphometry, intracellular ALP spectrophotometry, and calcified extracellular matrix (von Kossa) histomorphometry revealed that PLAs treated with exogenous rh-BMP-2 or transduced with a BMP-2 containing adenovirus (PLA-Ad-BMP-2) produced more bone precursors than osteoblasts ( p = 0.001). PLAs treated with exogenous rh-BMP-2 or PLA-Ad-BMP-2 also produced more bone precursors than BMAs ( p = 0.001), except for day 7 ALP histomorphometry (p = 0.343). ELISA confirmed successful BMP-2 production by both progenitor cell groups transduced with Ad-BMP-2. H&E sections from collagen I matrices seeded with PLA-Ad-BMP-2 cells confirmed bone formation at six weeks.Conclusion.r: Liposuction aspirates contain PLAs that can be transfected with the BMP-2 gene, with rapid induction into the osteoblast phenotype at a rate comparable to rh-BMP-2 and osteoblast groups. Transduced PLAs produce more bone precursors with faster onset of calcified extracellular matrix than transduced BMAs. PLAs may be an ideal source of mesenchyme-lineage stem cells for gene therapy and tissue engineering.