Structural Features of Heparan Sulfate from Multiple Osteochondromas and Chondrosarcomas.

Multiple osteochondromas (MO) is a hereditary disorder associated with benign cartilaginous tumors, known to be characterized by absence or highly reduced amount of heparan sulfate (HS) in the extracellular matrix of growth plate cartilage, which alters proper signaling networks leading to improper bone growth. Although recent studies demonstrated accumulation of HS in the cytoplasm of MO chondrocytes, nothing is known on the structural alterations which prevent HS from undergoing its physiologic pathway. In this work, osteochondroma (OC), peripheral chondrosarcoma, and healthy cartilaginous human samples were processed following a procedure previously set up to structurally characterize and compare HS from pathologic and physiologic conditions, and to examine the phenotypic differences that arise in the presence of either exostosin 1 or 2 (EXT1 or EXT2) mutations. Our data suggest that HS chains from OCs are prevalently below 10 kDa and slightly more sulfated than healthy ones, whereas HS chains from peripheral chondrosarcomas (PCSs) are mostly higher than 10 kDa and remarkably more sulfated than all the other samples. Although deeper investigation is still necessary, the approach here applied pointed out, for the first time, structural differences among OC, PCS, and healthy HS chains extracted from human cartilaginous excisions, and could help in understanding how the structural features of HS are modulated in the presence of pathological situations also involving different tissues.

Decreased EXT expression and intracellular accumulation of heparan sulphate proteoglycan in osteochondromas and peripheral chondrosarcomas.

Mutational inactivation of EXT1 or EXT2 is the cause of hereditary multiple osteochondromas. These genes function in heparan sulphate proteoglycan (HSPG) biosynthesis in the Golgi apparatus. Loss of heterozygosity of the EXT1 locus at 8q24 is frequently found in solitary osteochondromas, whereas somatic mutations are rarely found. We investigated the expression of EXT1 and EXT2 (quantitative RT-PCR) and of different HSPGs (immunohistochemistry) in solitary and hereditary osteochondromas and in cases with malignant progression to secondary peripheral chondrosarcoma, in relation to possible mutations and promoter methylation. The mutation status of patients with multiple osteochondromas correlated with decreased EXT1 or EXT2 expression found in their resected tumours. We could not show somatic point mutations or promoter hypermethylation in 17 solitary tumours; however, EXT1 expression was decreased in 15 cases, whereas EXT2 was not. Intracellular accumulation of syndecan-2 and heparan sulphate-bearing isoforms of CD44 (CD44v3) was found in most tumours, which concentrated in the Golgi apparatus as shown by confocal microscopy. This contrasted with the extracellular expression found in normal growth plates. In conclusion, mutational inactivation of either EXT1 or EXT2 leads to loss of mRNA expression of the corresponding gene. We hypothesize that loss of EXT expression disrupts the function of the EXT1/2 complex in HSPG biosynthesis, resulting in the intracellular accumulation of HSPG core proteins that we found in these tumours.

Soft tissue osteochondroma: case report and immunohistochemistry for parathyroid hormone-related protein.

Surface lesions of bone usually present little diagnostic dilemma because the majority are conventional osteochondromas. Other surface bone lesions include periosteal chondroma, periosteal chondrosarcoma, and parosteal osteosarcoma. Mineralized soft tissue lesions such as myositis ossificans, synovial chondroma, and synovial sarcoma may present in a similar fashion when they occur in a juxtaarticular position. The soft tissue osteochondroma or paraarticular osteochondroma may simulate some of these more aggressive tumors, and its recognition is important to avoid overtreatment. A case of an 11-year-old male with a soft tissue osteochondroma is reported to illustrate the characteristic radiographic and histological features of this rare entity. No prior reports have examined soft tissue osteochondroma for expression of parathyroid hormone related protein, an established cartilage tumor proliferative mitogen.

Localization of matrix proteins of hard tissue in osteochondromas.

We examined 10 cases of osteochondroma by means of histopathological, histochemical and immunohistochemical techniques. The surface of the masses was covered with a cartilage tissue showing positive immunohistochemical reaction for collagen type 2, and the deep region was composed of spongy bone, showing positive immunohistochemical reaction for collagen type 1 and osteocalcin. Between the cartilage and spongy bone, which is a metaphysis-like region, a chondroidal pattern appeared in the matrix of hypertrophy cartilage. In these regions, both type 1 (and osteocalcin) and 2 collagens were immunohistochemically detected. Although there is still no direct evidence, we believe that the cells involved in so-called "chondroid bone" temporally express cartilage phenotypes and then transform directly into bone-forming cells that survive in the chondroid bone until the tissue is resorbed and remodeled. Our examination results suggest that bone formation in osteochondromas, at least in some regions, occurs through transchondroid bone formation.

Involvement of BMP-2 signaling in a cartilage cap in osteochondroma.

This study describes the distributions of bone morphogenetic protein (BMP)-2 as well as mRNAs for BMP receptor type IB (BMPRIB). collagen types II (Col II) and III (Col III) in a growing "cartilage cap" of osteochondroma. In situ hybridization and immunohistochemical study were performed using histological sections obtained during surgery. BMP-2 was detected in mesenchymal cells in the outer fibrous layer and chondrocytes in the inner cartilaginous matrix, positive for Col III and Col II, respectively. BMPRIB mRNA was distributed in chondrocytes. This is the first study to provide observational evidence of the involvement of BMP-2 signaling in the pathogenesis of cartilage cap of osteochondroma. and suggests the role of BMP-2 in the growth of cartilage cap in osteochondroma.

Bizarre parosteal osteochondromatous proliferation (Nora's lesion) of the foot.

A 22-year-old man presented with a growing lump on the fifth metatarsal of the right foot. Radiographically, the lesion was a calcified mass stuck on to the bone. The T2-weighted magnetic resonance images showed heterogeneity in intensity. A tumor was suspected and an excisional biopsy was done. The lesion was composed of a cartilaginous cap and bone tissue. Histological examination revealed characteristic features of bizarre parosteal osteochondromatous proliferation (BPOP), such as hypercellularity, a blue tinctorial quality in the osteocartilaginous interfaces, and a scattering of binucleated or bizarre enlarged chondrocytes. Immunohistochemically, basic fibroblast growth factor was expressed in nearly all chondrocytes within the cartilaginous cap, while vascular endothelial growth factor was expressed only in enlarged chondrocytes near the osteocartilaginous interfaces. Reverse transcription-polymerase chain reaction detected chondromodulin-I transcripts in the tissue of the cartilaginous cap. These findings indicate that the processes occurring in BPOP are similar to those occurring in endochondral ossification in the growth plate, and they support the concept that BPOP is a reparative process. BPOP is a rare tumorous lesion of the bone and is occasionally confused with other benign or malignant conditions. Thus, it is important to consider the clinical, radiographical and the gross histological features of the lesion when making a diagnosis.

Expression of tau proteins and tubulin in extraskeletal myxoid chondrosarcoma, chordoma, and other chondroid tumors.

Tau proteins are microtubule-associated proteins required for the polymerization of tubulin. The abnormal accumulation of tau proteins in neurofibrillary tangles is a well-known phenomenon and has been studied extensively. However, the role of tau protein in chondroid tissue and its neoplasia is unknown. In the present study, 2 extraskeletal myxoid chondrosarcomas (EMCs), 6 chordomas, 6 chondrosarcomas, 3 myxoid chondrosarcomas of bone, 2 osteochondromas, 6 chondroblastomas, and 2 nonneoplastic adult articular cartilages were immunostained with monoclonal antibodies against tau proteins and tubulin. The results showed that the coexpression of tau proteins and tubulin was present only in EMCs (2/2) and chordomas (4/6). Although tubulin was detected in chondroblastomas (5/6), osteochondromas (2/2), chondrosarcomas (5/6), and myxoid-chondrosarcomas of bone (3/3), tau expression was absent in these tumors. The perichondrial chondroblasts but not chondrocytes from nonneoplastic articular cartilage also localize tau and tubulin with a much weaker staining intensity. The results mainly demonstrate that there is frequent expression of tau proteins in some microtubule-rich neoplasms, such as EMC and chordoma. The different immunostaining pattern of tau proteins between chordoma and myxoid chondrosarcoma of bone may be useful in the differential diagnosis, especially when both neoplasms occur in the base of skull.

Immunolocalization of matrix metalloproteinases and tissue inhibitors of metalloproteinases in human chondrosarcomas.

We have immunohistochemically examined the localization of matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) in human chondrosarcomas (CHS) (23 cases) and benign chondroid lesions (BCL) (16 cases of osteochondromas and 11 cases of enchondromas). In CHS, all the MMPs and TIMPs examined were positive. Among them, MMP-1 was immunolocalized in more than 90% of both CHS and BCL, but positive score of MMP-1 was significantly higher in CHS than that in BCL (p < 0.01). Compared with BCL, CHS expressed MMP-3 at a low level, and more often positive in MMP-9. It is possible that chondrosarcoma might have a tendency to lose the ability to secrete MMP-3, which is a metalloproteinase that can degrade cartilage proteoglycans and is related to normal cartilage turnover. MMP-2, TIMP-1 and TIMP-2 were immunolocalized in more than 70% of the cases of both BCL and CHS, but the positive scores of these were not statistically different between the two groups. Interestingly, in several cases of CHS, both MMP-1 and MMP-9 immunostains were observed preferentially within the cells at the marginal areas of cartilaginous lobules. These findings suggest that increased expression of MMP-1 and MMP-9 and decrease in MMP-3 expression are associated with the malignant phenotype of the cartilaginour tumors.

The identity of proliferating cells in bone tumors with cartilaginous components: evaluation by double-immunohistochemical staining using proliferating cell nuclear antigen and S-100 protein.

S-100 protein (S-100) appears to be a marker for bone tumors of cartilaginous origin. Any analyses of proliferative activity in S-100-positive tumor cells, however, has not yet been presented. This study assessed the proliferative activity of those cells by means of a double-immunohistochemical staining method using proliferating cell nuclear antigen (PCNA) and S-100. The most intense reactivity for S-100 was found in the well-differentiated chondrocytes of enchondromas, osteochondromas, and osteosarcomas. On the contrary, the more immature the tumor cells were, the more intensely positive they were for PCNA. In parosteal chondrosarcoma, exceptionally, PCNA-positive as well as S-100-positive cells were abundant, suggesting that these proliferating cells produced S-100. In periosteal osteosarcoma, however, the proliferating cells labeled by PCNA revealed little reactivity for S-100. This immunohistochemical method is potentially useful to know the identity and origin of proliferating cells and may sometimes be diagnostic for bone tumors containing cartilaginous elements.