Smad4 regulates growth plate matrix production and chondrocyte polarity.

Amanda T Whitaker,Tamara Alliston,Ellora Berthet,Diana J Laird,Andrea Cantu

doi:10.1242/bio.021436

Amanda T Whitaker, Tamara Alliston + Show 3 more

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https://doi.org/10.1242/bio.021436

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Abstract

ABSTRACTSmad4 is an intracellular effector of the TGFβ family that has been implicated in Myhre syndrome, a skeletal dysplasia characterized by short stature, brachydactyly and stiff joints. The TGFβ pathway also plays a critical role in the development, organization and proliferation of the growth plate, although the exact mechanisms remain unclear. Skeletal phenotypes in Myhre syndrome overlap with processes regulated by the TGFβ pathway, including organization and proliferation of the growth plate and polarity of the chondrocyte. We used in vitro and in vivo models of Smad4 deficiency in chondrocytes to test the hypothesis that deregulated TGFβ signaling leads to aberrant extracellular matrix production and loss of chondrocyte polarity. Specifically, we evaluated growth plate chondrocyte polarity in tibiae of Col2-Cre+/−;Smad4fl/fl mice and in chondrocyte pellet cultures. In vitro and in vivo, Smad4 deficiency decreased aggrecan expression and increased MMP13 expression. Smad4 deficiency disrupted the balance of cartilage matrix synthesis and degradation, even though the sequential expression of growth plate chondrocyte markers was intact. Chondrocytes in Smad4-deficient growth plates also showed evidence of polarity defects, with impaired proliferation and ability to undergo the characteristic changes in shape, size and orientation as they differentiated from resting to hypertrophic chondrocytes. Therefore, we show that Smad4 controls chondrocyte proliferation, orientation, and hypertrophy and is important in regulating the extracellular matrix composition of the growth plate.

Highlights

The physis, or growth plate, is comprised of precisely organized chondrocytes that confer longitudinal growth of the bone through interactions of multiple signaling pathways that cooperate to control chondrocyte shape, polarity, proliferation, differentiation, and apoptosis (Abad et al, 1999; Sasaki et al, 2001)
Chondrocyte-intrinsic ablation of Smad4 disrupts the balance of extracellular matrix synthesis and degradation To evaluate the chondrocyte-intrinsic role of Smad4, excision was induced from primary chondrocytes of Smad4fl/fl mice by adenoviral infection with Cre recombinase (Ad-Cre) in vitro
We examined mRNA expression of aggrecan and matrix metallopeptidase 13 (MMP13) to determine if the Smad4-dependent defect in extracellular matrix (ECM) composition corresponds to an imbalance in the expression of anabolic and catabolic genes, as it does in Smad3-deficient chondrocytes (Chen et al, 2012)

Summary

Introduction

The physis, or growth plate, is comprised of precisely organized chondrocytes that confer longitudinal growth of the bone through interactions of multiple signaling pathways that cooperate to control chondrocyte shape, polarity, proliferation, differentiation, and apoptosis (Abad et al, 1999; Sasaki et al, 2001). Received 24 August 2016; Accepted 30 January 2017 these cellular events, either by trauma or genetic mutation, leads to physeal defects that can result in skeletal deformities and abnormal limb growth. In one such example, dysregulation of Smad, a common intracellular effector of all transforming growth factor β (TGFβ) family members, is responsible for human Myhre syndrome and characterized by short stature, brachydactyly, and joint stiffness (Caputo et al, 2012; Le Goff et al, 2012; Caputo et al, 2014). The cellular basis of physeal defects in Smad4-deficient mice and skeletal deformities in Myhre syndrome remains unclear

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