Basal and inducible Osterix expression reflect equine mesenchymal progenitor cell osteogenic capacity

The two most commonly used methods to analyze data from real-time, quantitative PCR experiments are absolute quantification and relative quantification. Absolute quantification determines the input copy number, usually by relating the PCR signal to a standard curve. Relative quantification relates the PCR signal of the target transcript in a treatment group to that of another sample such as an untreated control. The 2(-Delta Delta C(T)) method is a convenient way to analyze the relative changes in gene expression from real-time quantitative PCR experiments. The purpose of this report is to present the derivation, assumptions, and applications of the 2(-Delta Delta C(T)) method. In addition, we present the derivation and applications of two variations of the 2(-Delta Delta C(T)) method that may be useful in the analysis of real-time, quantitative PCR data. Copyright 2001 Elsevier Science (USA). Show more

Human mesenchymal stem cells are thought to be multipotent cells, which are present in adult marrow, that can replicate as undifferentiated cells and that have the potential to differentiate to lineages of mesenchymal tissues, including bone, cartilage, fat, tendon, muscle, and marrow stroma. Cells that have the characteristics of human mesenchymal stem cells were isolated from marrow aspirates of volunteer donors. These cells displayed a stable phenotype and remained as a monolayer in vitro. These adult stem cells could be induced to differentiate exclusively into the adipocytic, chondrocytic, or osteocytic lineages. Individual stem cells were identified that, when expanded to colonies, retained their multilineage potential. Show more

Future cell-based therapies such as tissue engineering will benefit from a source of autologous pluripotent stem cells. For mesodermal tissue engineering, one such source of cells is the bone marrow stroma. The bone marrow compartment contains several cell populations, including mesenchymal stem cells (MSCs) that are capable of differentiating into adipogenic, osteogenic, chondrogenic, and myogenic cells. However, autologous bone marrow procurement has potential limitations. An alternate source of autologous adult stem cells that is obtainable in large quantities, under local anesthesia, with minimal discomfort would be advantageous. In this study, we determined if a population of stem cells could be isolated from human adipose tissue. Human adipose tissue, obtained by suction-assisted lipectomy (i.e., liposuction), was processed to obtain a fibroblast-like population of cells or a processed lipoaspirate (PLA). These PLA cells can be maintained in vitro for extended periods with stable population doubling and low levels of senescence. Immunofluorescence and flow cytometry show that the majority of PLA cells are of mesodermal or mesenchymal origin with low levels of contaminating pericytes, endothelial cells, and smooth muscle cells. Finally, PLA cells differentiate in vitro into adipogenic, chondrogenic, myogenic, and osteogenic cells in the presence of lineage-specific induction factors. In conclusion, the data support the hypothesis that a human lipoaspirate contains multipotent cells and may represent an alternative stem cell source to bone marrow-derived MSCs. Show more