Friday, September 4, 2009


Shox2 is required for chondrocyte proliferation and maturation in proximal limb skeleton.

"Mutations in the short stature homeobox gene SHOX lead to growth retardation associated with Turner, Leri-Weill dyschondrosteosis, and Langer mesomelic dysplasia syndromes, which marked the shortening of the forearms and lower legs. We report here that in contrast to the SHOX mutations in humans, Shox2 deficiency in mice leads to a virtual elimination of the stylopod in the developing limbs, while the zeugopod and autopod appear relatively normal. This phenotype is consistent with the restriction of the Shox2 expression to the proximal mesenchyme in the limb bud and later to chondrocytes associated with the forming stylopod. In the Shox2(-/-) embryo, the mesenchymal condensation for the stylopod initiates normally but the cartilaginous element subsequently fails in growth, chondrogenesis and endochondral ossification. A dramatic down-regulation of Runx2 and Runx3 could account for the lack of chondrocyte hypertrophy, while a down-regulation of Ihh expression may be responsible for a significant reduction in chondrocyte proliferation in the mutant stylopod. We further demonstrate that an enhanced and ectopic Bmp4 expression in the proximal limb of the Shox2 embryo may underlie the down-regulation of Runx2, as ectopically applied exogenous BMP4 represses Runx2 expression in the early limb bud. Moreover, we show that mouse Shox2, similar to human SHOX, can perform opposite roles on gene expression: either as a transcription activator or a repressor in different cell types. Our results establish a key role for Shox2 in regulating the growth of stylopod by controlling chondrocyte maturation via Runx2 and Runx3."

"deletions of Hoxa10, Hoxc10 and Hoxd10 {LSJL upregulates this} drastically shorten the femur, whereas loss of Hoxa11 and Hoxd11 results in a virtual loss of the radius and the ulna "

"Ectopic expression of either Meis1 or Meis2 in developing chick limbs reduces or truncates distal limb compartments"

"as limb bud elongates, Shox2 expression was confined to the mesenchyme of the proximal limb, with stronger expression remaining in the dorsal region"

Shox2 may affect Ihh expression.

The role of Shox2 in SAN development and function. states that Shox2 may have a role in Nkx3.2 repression.

Shox2 regulates progression through chondrogenesis in the mouse proximal limb.

" Prrx1{up in LSJL}-Cre-driven limb-specific deletion of the paralogous gene Shox2 results in severe rhizomelia. In this study, we show that Col2a1-Cre-driven deletion of Shox2 in developing chondrocytes also results in shortening of the stylopodial skeleton (i.e. humerus, femur) and that this rhizomelia is due to precocious chondrocyte maturation and hypertrophy. We demonstrate, using the micromass culture model system, that increased BMP activity triggers accelerated maturation and hypertrophy in Col2a1-Cre Shox2 mutant chondrocytes and we confirm in vivo that elevated transcript levels and expanded expression domains of Bmp2{up in LSJL} and 4 are associated with premature formation of the hypertrophic zone in mutant humeri. In micromass cultures of Prrx1-Cre Shox2 mutant limb cells, we find that Shox2 deletion in undifferentiated mesenchymal cells results in increased BMP activity that enhances early chondrogenesis, but is insufficient to provoke chondrocyte maturation and hypertrophy. Similarly, shRNA-mediated Shox2 knockdown in multipotent C3H10T1/2 cells and primary mouse bone marrow mesenchymal stem cells results in spontaneous chondrogenesis in the absence of chondrostimulation, but again fails to induce progression through the later stages of chondrogenic differentiation. Importantly, exogenous BMP supplementation can overcome the block to maturation and hypertrophy caused by Shox2 depletion prior to overt chondrogenesis. Thus, we provide evidence that Shox2 regulates progression through chondrogenesis at two distinct stages - the onset of early differentiation and the transition to maturation and hypertrophy."

"undifferentiated mesenchymal cells upregulate expression of the transcription factor Sox9{up} and commence chondrogenesis. Newly differentiated chondrocytes produce the characteristic cartilage extracellular matrix (ECM) components aggrecan (Acan){up} and collagen type II (Col2a1){up} and proliferate to expand the size of the nascent cartilage anlage.  Prompted by Runx2 expression, proliferating chondrocytes at the center of the template exit the cell cycle and initiate hypertrophic differentiation. Hypertrophic chondrocytes express the unique marker collagen type X (Col10a1){up} and experience a massive increase in cytoplasmic volume. Perichondrial cells and chondrocytes at the ends of the cartilage template express Pthlp, which stimulates chondrocytes to proliferate. As these proliferating growth plate chondrocytes leave the domain of PTHLP influence, they mature into prehypertrophic chondrocytes and express Ihh. It is the enormous increase in hypertrophic chondrocyte volume and the columns of rapidly proliferating chondrocytes that are responsible for driving longitudinal bone growth. Finally, hypertrophic chondrocytes express genes like Bglap, Ibsp, and Spp1 in order to mineralize their ECM before undergoing apoptosis"

"Col2a1-Cre Shox2 mutant cultures express significantly lower levels of genes involved in early chondrogenesis, like Sox9, Sox6, Acan, and Col2a1, and chondrocyte maturation, like Runx2 and Ihh, but significantly higher levels of the hypertrophic marker Col10a1"

"chondrocyte-specific Shox2 deletion significantly increased levels of Bmp2 and 4 mRNAs"

"Prrx1-Cre Shox2 mutant cultures express significantly lower levels of the tenogenic marker gene Scx{up} and the myogenic markers Myf5, Myod1, and Myog{down}"

"when embryonic chick limb bud cells are cultured in micromass, limb positional information is gradually lost unless the cells are exposed to PD patterning signals, like retinoic acid (RA), FGF8, and WNT3A"

"mutant micromasses with 10 ng/ml Noggin for the last 3 days of a 7-day culture period and found that Noggin completely blocked the internodular chondrogenesis normally observed in Prrx1-Cre Shox2 mutant cultures"<-Could Shox2 deletion induce ectopic chondrogenesis?

"Col2a1-Cre-driven conditional Shox2 deletion causes significant shortening of the humerus and femur and that this rhizomelia is caused by precocious hypertrophic differentiation of stylopodial chondrocytes"

"Shox2 knockdown induces MSC chondrogenesis without hypertrophy, as long as Shox2-deficient MSC pellets are kept BMP-free"

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