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Lineage allocation of the marrow mesenchymal stem cells (MSCs) to osteoblasts and adipocytes is dependent on both Wnt signaling and PPARγ2 activity. Activation of PPARγ2, an essential regulator of energy metabolism and insulin sensitivity, stimulates adipocyte and suppresses osteoblast differentiation and bone formation, and correlates with decreased bone mass and increased fracture rate. In contrast, activation of Wnt signaling promotes osteoblast differentiation, augments bone accrual and reduces total body fat. This study examined the cross-talk between PPARγ2 and β-catenin, a key mediator of canonical Wnt signaling, on MSC lineage determination. Rosiglitazone-activated PPARγ2 induced rapid proteolytic degradation of β-catenin, which was prevented by either inhibiting glycogen synthase kinase 3 beta (GSK3β) activity, or blocking pro-adipocytic activity of PPARγ2 using selective antagonist GW9662 or mutation within PPARγ2 protein. Stabilization of β-catenin suppressed PPARγ2 pro-adipocytic but not anti-osteoblastic activity. Moreover, β-catenin stabilization decreased PPARγ2-mediated insulin signaling as measured by insulin receptor and FoxO1 gene expression, and protein levels of phosphorylated Akt (pAkt). Cellular knockdown of β-catenin with siRNA increased expression of adipocyte but did not affect osteoblast gene markers. Interestingly, the expression of Wnt10b was suppressed by anti-osteoblastic, but not by pro-adipocytic activity of PPARγ2. Moreover, β-catenin stabilization in the presence of activated PPARγ2 did not restore Wnt10b expression indicating a dominant role of PPARγ2 in negative regulation of pro-osteoblastic activity of Wnt signaling. In conclusion, β-catenin and PPARγ2 are in cross-talk which results in sequestration of pro-adipocytic and insulin sensitizing activity. The anti-osteoblastic activity of PPARγ2 is independent of this interaction.

Rosiglitazone (Rosi), a member of the thiazolidinedione class of drugs used to treat type 2 diabetes, activates the adipocyte-specific transcription factor peroxisome proliferator-activated receptor gamma (PPARgamma). This activation causes bone loss in animals and humans, at least in part due to suppression of osteoblast differentiation from marrow mesenchymal stem cells (MSC). In order to identify mechanisms by which PPARgamma2 suppresses osteoblastogenesis and promotes adipogenesis in MSC, we have analyzed the PPARgamma2 transcriptome in response to Rosi. A total of 4,252 transcriptional changes resulted when Rosi (1 microM) was applied to the U-33 marrow stromal cell line stably transfected with PPARgamma2 (U-33/gamma2) as compared to non-induced U-33/gamma2 cells. Differences between U-33/gamma2 and U-33 cells stably transfected with empty vector (U-33/c) comprised 7,928 transcriptional changes, independent of Rosi. Cell type-, time- and treatment-specific gene clustering uncovered distinct patterns of PPARgamma2 transcriptional control of MSC lineage commitment. The earliest changes accompanying Rosi activation of PPARgamma2 included effects on Wnt, TGFbeta/BMP and G-protein signaling activities, as well as sustained induction of adipocyte-specific gene expression and lipid metabolism. While suppression of osteoblast phenotype is initiated by a diminished expression of osteoblast-specific signaling pathways, induction of the adipocyte phenotype is initiated by adipocyte-specific transcriptional regulators. This indicates that distinct mechanisms govern the repression of osteogenesis and the stimulation of adipogenesis. The co-expression patterns found here indicate that PPARgamma2 has a dominant role in controlling osteoblast differentiation and suggests numerous gene-gene interactions that could lead to the identification of a "master" regulatory scheme directing this process.

Peroxisome proliferator activated receptor gamma (PPARγ) controls both glucose metabolism and an allocation of marrow mesenchymal stem cells (MSCs) toward osteoblast and adipocyte lineages. Its activity is determined by interaction with a ligand which directs posttranscriptional modifications of PPARγ protein including dephosphorylation of Ser112 and Ser273, which results in acquiring of pro-adipocytic and insulin-sensitizing activities, respectively. PPARγ full agonist TZD rosiglitazone (ROSI) decreases phosphorylation of both Ser112 and Ser273 and its prolonged use causes bone loss in part due to diversion of MSCs differentiation from osteoblastic toward adipocytic lineage. Telmisartan (TEL), an anti-hypertensive drug from the class of angiotensin receptor blockers, also acts as a partial PPARγ agonist with insulin-sensitizing and a weak pro-adipocytic activity. TEL decreased S273pPPARγ and did not affect S112pPPARγ levels in a model of marrow MSC differentiation, U-33/γ2 cells. In contrast to ROSI, TEL did not affect osteoblast phenotype and actively blocked ROSI-induced anti-osteoblastic activity and dephosphorylation of S112pPPARγ. The effect of TEL on bone was tested side-by-side with ROSI. In contrast to ROSI, TEL administration did not affect bone mass and bone biomechanical properties measured by micro-indentation method and did not induce fat accumulation in bone, and it partially protected from ROSI-induced bone loss. In addition, TEL induced “browning” of epididymal white adipose tissue marked by increased expression of UCP1, FoxC2, Wnt10b and IGFBP2 and increased overall energy expenditure. These studies point to the complexity of mechanisms by which PPARγ acquires anti-osteoblastic and pro-adipocytic activities and suggest an importance of Ser112 phosphorylation status as being a part of the mechanism regulating this process. These studies showed that TEL acts as a full PPARγ agonist for insulin-sensitizing activity and as a partial agonist/partial antagonist for pro-adipocytic and anti-osteoblastic activities. They also suggest a relationship between PPARγ fat “browning” activity and a lack of anti-osteoblastic activity. [ABSTRACT FROM AUTHOR]