Mechanism of HIV protein induced modulation of mesenchymal stem cell osteogenic differentiation.
BACKGROUND
A high incidence of decreased bone mineral density (BMD) has been associated with HIV infection. Normal skeletal homeostasis is controlled, at least in part, by the maturation and activity of mature osteoblasts. Previous studies by our group have demonstrated the ability of HIV proteins to perturb osteoblast function, and the degree of osteogenesis in differentiating mesenchymal stem cells (MSCs). This study attempts to further dissect the dynamics of this effect.


METHODS
MSCs were cultured under both osteogenic (cultured in commercially available differentiation media) and quiescent (cultured in basal medium) conditions. Both cell populations were exposed to HIV p55-gag and HIV rev (100 ng/ml). Time points were taken at 3, 6, 9, and 15 days for osteogenic conditions, while quiescent cells were treated for 1 week. Cell function (alkaline phosphatase [ALP] activity, calcium deposition, and lipid levels) and the activity of the key MSC transcription factors, RUNX-2 and PPARgamma were determined post-exposure. Also, in cells cultured in differentiating conditions, cellular levels of connective tissue growth factor (CTGF) were analysed using whole cell ELISA, while BMP-2 secretion was also examined.


RESULTS
In differentiating MSCs, exposure to HIV proteins caused significant changes in both the timing and magnitude of key osteogenic events and signals. Treatment with REV increased the overall rate of mineralization, and induced earlier increases in CTGF levels, RUNX-2 activity and BMP-2 secretion, than those observed in the normal course of differntiation. In contrast, p55-gag reduced the overall level of osteogenesis, and reduced BMP-2 secretion, RUNX-2 activity, CTGF levels and ALP activity at many of the timepoints examined. Finally, in cells cultured in basal conditions, treatment with HIV proteins did not in and of itself induce a significant degree of differentiation over the time period examined.


CONCLUSIONS
These data demonstrate that the effect of HIV proteins on bone is dependent on the differentiation status of the cells that they are in contact with. The effect on bone cell signalling provides insights into the mechanism of HIV induced decreases in bone mineral density.
Cotter
Eoin J
EJ
Clinical Research Center, School of Medicine and Medical Sciences, University College Dublin, Catherine Mcauley Centre, Nelson St,, Dublin 7, Ireland. ecotter.genome@mater.ie
Ip
Herbert Shi Ming
HS
Powderly
William G
WG
Doran
Peter P
PP
D000469:Alkaline Phosphatase / Q000378:metabolism; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D055513:Connective Tissue Growth Factor; D050659:Core Binding Factor Alpha 1 Subunit / Q000378:metabolism; D006801:Humans; D017874:Immediate-Early Proteins / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D050356:Lipid Metabolism; D059630:Mesenchymal Stem Cells / Q000201:enzymology / Q000378:metabolism*; D010006:Osteoblasts / Q000201:enzymology / Q000378:metabolism*; D010012:Osteogenesis; D015398:Signal Transduction; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000378:metabolism; D054301:gag Gene Products, Human Immunodeficiency Virus / Q000378:metabolism*; D054321:rev Gene Products, Human Immunodeficiency Virus / Q000378:metabolism*
C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; C526317:CCN2 protein, human; D050659:Core Binding Factor Alpha 1 Subunit; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; C493709:RUNX2 protein, human; D016212:Transforming Growth Factor beta; D054301:gag Gene Products, Human Immunodeficiency Virus; D054321:rev Gene Products, Human Immunodeficiency Virus; C514860:rev protein, Human Immunodeficiency Virus-1; D055513:Connective Tissue Growth Factor; D000469:Alkaline Phosphatase; D002118:Calcium