Post-weaning epiphysiolysis causes distal femur dysplasia and foreshortened hindlimbs in fetuin-A-deficient mice.

Laura J Brylka,Bent Brachvogel,Bernd Denecke,Julia Etich,Peter Boor,Sina Köppert,Beate Kratz,Timur A Yorgan,Thorsten Schinke,Anne Babler,Willi Jahnen-Dechent,Ivan G Costa,Dengshun Miao

doi:10.1371/journal.pone.0187030

Abstract

Fetuin-A / α2-Heremans-Schmid-glycoprotein (gene name Ahsg) is a systemic inhibitor of ectopic calcification. Due to its high affinity for calcium phosphate, fetuin-A is highly abundant in mineralized bone matrix. Foreshortened femora in fetuin-A-deficient Ahsg-/- mice indicated a role for fetuin-A in bone formation. We studied early postnatal bone development in fetuin-A-deficient mice and discovered that femora from Ahsg-/- mice exhibited severely displaced distal epiphyses and deformed growth plates, similar to the human disease slipped capital femoral epiphysis (SCFE). The growth plate slippage occurred in 70% of Ahsg-/- mice of both sexes around three weeks postnatal. At this time point, mice weaned and rapidly gained weight and mobility. Epiphysis slippage never occurred in wildtype and heterozygous Ahsg+/- mice. Homozygous fetuin-A-deficient Ahsg-/- mice and, to a lesser degree, heterozygous Ahsg+/- mice showed lesions separating the proliferative zone from the hypertrophic zone of the growth plate. The hypertrophic growth plate cartilage in long bones from Ahsg-/- mice was significantly elongated and V-shaped until three weeks of age and thus prior to the slippage. Genome-wide transcriptome analysis of laser-dissected distal femoral growth plates from 13-day-old Ahsg-/- mice revealed a JAK-STAT-mediated inflammatory response including a 550-fold induction of the chemokine Cxcl9. At this stage, vascularization of the elongated growth plates was impaired, which was visualized by immunofluorescence staining. Thus, fetuin-A-deficient mice may serve as a rodent model of growth plate pathologies including SCFE and inflammatory cartilage degradation.

Highlights

Endochondral ossification is a tightly controlled developmental sequence of chondrogenic cell differentiation, proliferation and hypertrophy, followed by vascularization and cartilage mineralization
The three-dimensional visualization revealed a notable feature that had gone unnoticed in all previous studies on the bone phenotype of Ahsg-/- mice: the distal femoral epiphysis was rotated posterior, the lateral and medial condyles and epicondyles were flattened and widened, which resulted in narrowing of the intercondylar fossa (Fig 1C)
The rotation of the distal femoral epiphysis was quantified measuring the angle of the distal femoral growth plate relative to the shaft as shown in S1 Fig. This angle was significantly decreased in male Ahsg-/- mice (Fig 1E)

Summary

Introduction

Endochondral ossification is a tightly controlled developmental sequence of chondrogenic cell differentiation, proliferation and hypertrophy, followed by vascularization and cartilage mineralization. Calcified cartilage is remodeled into mineralized and vascularized bone [1]. This highly dynamic sequence of events progresses in a confined space–the growth plate– and involves cell layers connected by extracellular matrices with greatly varying mechanical properties. In order to maintain overall growth plate stability and to prevent the formation of fault lines, the progression from proliferative to hypertrophic cartilage must be tightly controlled. Remodeling of mineralized cartilage and concurrent vascularization may be critical, because both, mineralization and vascularization, greatly affect the stability of the growth plate. Fetuin-A / α2-Heremans-Schmid-glycoprotein (gene name Ahsg) is one of the most abundant non-collagen proteins in mineralized bone [6]. Fetuin-A was found to regulate mineralization in primary rat calvarial osteoblasts, calcifying vascular smooth muscle cells [9,16] and in collagen mineralization assays [17]

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