Heterotopic ossification in a patient with diffuse idiopathic skeletal hyperostosis: Input from histological findings

Abstract

A high incidence of heterotopic ossification (HO) has been reported in patients with diffuse idiopathic skeletal hyperostosis (DISH), a metabolic disease characterized by calcifications of entheses at spine and peripheral sites. We performed histological and immunohistochemical analyses in five different HO sites in a patient with DISH to study a possible mutual interaction of bone morphogenetic protein 2 (BMP-2), transforming growth factor beta (TGF-β), and decorin, crucial for bone mass increasing, matrix calcification, and endochondral bone formation. We speculated that the surgical trauma triggered HO, inducing TGF-β release at the lesion site. TGF-β recruits osteoblast precursor cells and determines the overexpression of BMP-2 in the surrounding skeletal muscle, inducing a further osteogenic differentiation, contributing to HO onset.

Figure 1.

Radiological and CT evidence of HO around the hip. A) Pre-operative X-rays image showing bridging ossification along T10, T11 and T12 vertebrae. B,C) Pre-operative CT images; IP, ilio-psoas; Mg, medius gluteus; Pir, piriformis. D) Pre-operative X-rays image showing HO at right hip. E) X-rays image taken at 1 day after surgery. e) X-rays image taken at 8 weeks after surgery.

Figure 2.

A-F) Haematoxylin & Eosin staining of HO. A) Compact bone with adjacent cartilage layer. Fibrous tissue and bone marrow are also shown; scale bar 100 μm. B) Ossification site; scale bar 50 μm. C) Muscular and fibrous tissues adjacent to bone; scale bar 100 μm. D) Compact bone; scale bar 100 μm. E) Trabecular bone; scale bar 100 μm).F) Particular of ossification site, black arrowheads indicate osteoclasts; scale bar 10 μm. G-I) Toluidine Blue staining; blue colour indicates high proteoglycan content. G) Compact bone with adjacent cartilage layer; scale bar 100 μm. H) Compact bone; scale bar 100 μm. I) Particular of compact bone; scale bar 50 μm. J-L) Safranin O staining; red colour indicates high proteoglycan content. J) Compact bone with adjacent cartilage layer; scale bar 100 μm. K) Compact bone; scale bar 100 μm. L) Particular of compact bone; scale bar 50 μm; W, woven bone; L, lamellar bone; B, bone; C, cartilage; BM, bone marrow; M, muscular tissue; F, fibrous tissue; OS, ossification site.

Figure 3.

A) Immunohistochemical analysis of BMP-2, TGF-beta and decorin in healthy and HO tissues; scale bar 50 μm; BMP-2 staining in (a) healthy lamellar bone, (b) HO lamellar and woven bone (W), and (c) representative histogram of BMP-2 expression (**p<0.01); BMP-2 expression in (d) healthy and e) HO muscular tissue, and (f ) representative histogram of BMP-2 expression (**p<0.01); TGF-beta staining in (g) healthy lamellar bone (arrowheads), (h) HO woven and lamellar bone (arrowheads), and (i) representative histogram of TGF-beta expression; decorin expression in (j) healthy lamellar bone, (k) HO woven (W) and lamellar bone, and (l) representative histogram of decorin expression (**p<0.01). B) BMP-2, TGF-beta and decorin staining in cartilage tissue in HO; a) at the ossification site (OS), BMP-2 expression in cartilage, chondrocytes and osteoclast (arrowheads) in cartilage; b) at the ossification site (OS), TGF-beta expression in cartilage and chondrocytes; c) at the ossification site (OS), decorin expression in cartilage and chondrocytes.