High-grade osteosarcoma is characterized by extensive genetic instability, thereby hampering the identification of causative gene mutations and understanding of the underlying pathological processes. It lacks a benign precursor lesion and reports on associations with hereditary predisposition or germline mutations are uncommon, despite the early age of onset. Here we demonstrate a novel comprehensive approach for the study of premalignant stages of osteosarcoma development in a murine mesenchymal stem cell (MSC) system that formed osteosarcomas upon grafting. By parallel functional and phenotypic analysis of normal MSCs, transformed MSCs and derived osteosarcoma cells, we provide substantial evidence for a MSC origin of osteosarcoma. In a stepwise approach, using COBRA-FISH karyotyping and array CGH in different passages of MSCs, we identified aneuploidization, translocations and homozygous loss of the cdkn2 region as the key mediators of MSC malignant transformation. We then identified CDKN2A/p16 protein expression in 88 osteosarcoma patients as a sensitive prognostic marker, thereby bridging the murine MSCs model to human osteosarcoma. Moreover, occasional reports in patients mention osteosarcoma formation following bone marrow transplantation for an unrelated malignancy. Our findings suggest a possible hazard for the clinical use of MSCs; however, they also offer new opportunities to study early genetic events in osteosarcoma genesis and, more importantly, to modulate these events and record the effect on tumour progression. This could be instrumental for the identification of novel therapeutic strategies, since the success of the current therapies has reached a plateau phase.

The Ewing sarcoma breakpoint region 1 (EWSR1; also known as EWS) represents one of the most commonly involved genes in sarcoma translocations. In fact, it is involved in a broad variety of mesenchymal lesions which includes Ewing's sarcoma/peripheral neuroectodermal tumor, desmoplastic small round cell tumor,clear cell sarcoma, angiomatoid fibrous histiocytoma, extraskeletal myxoid chondrosarcoma, and a subset of myxoid liposarcoma. The fusion products between EWSR1 and partners usually results in fusion of the N-terminal transcription-activating domain of EWSR1 and the C-terminal DNA-binding domain of the fusion partner, eventually generating novel transcription factors. EWSR1 rearrangement can be visualized by the means of fluorescence in situ hybridization (FISH). As soft tissue sarcomas represent a diagnostically challenging group, FISH analysis is an extremely useful confirmatory diagnostic tool. However, as in most instances a split-apart approach is used, the results of molecular genetics must be evaluated in context with morphology.

Mesenchymal stem cells (MSCs) are multipotent cells capable of differentiation into several mesodermal lineages. These cells have been isolated from various tissues, such as adult bone marrow, placenta, and fetal tissues. The comparative potential of these cells originating from different tissues to differentiate into the chondrogenic lineage is still not fully defined. The aim of our study was to investigate the chondrogenic potential of MSCs isolated from different sources. MSCs from fetal and adult tissues were phenotypically characterized and examined for their differentiation capacity, based on morphological criteria and expression of extracellular matrix components. Our results show that both fetal and adult MSCs have chondrogenic potential under appropriate conditions. The capacity of bone marrow-derived MSCs to differentiate into chondrocytes was reduced on passaging of cells. MSCs of bone marrow origin, either fetal or adult, exhibit a better chondrogenesis than fetal lung- and placenta-derived MSCs, as demonstrated by the appearance of typical morphological features of cartilage, the intensity of toluidine blue staining, and the expression of collagen type II, IX, and X after culture under chondrogenic conditions. As MSCs represent an attractive tool for cartilage tissue repair strategies, our data suggest that bone marrow should be considered the preferred MSC source for these therapeutic approaches.

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Primary cilia are specialized cell surface projections found on most cell types. Involved in several signaling pathways, primary cilia have been reported to modulate cell and tissue organization. Although they have been implicated in regulating cartilage and bone growth, little is known about the organization of primary cilia in the growth plate cartilage and osteochondroma. Osteochondromas are bone tumors formed along the growth plate, and they are caused by mutations in EXT1 or EXT2 genes. In this study, we show the organization of primary cilia within and between the zones of the growth plate and osteochondroma. Using confocal and electron microscopy, we found that in both tissues, primary cilia have a similar formation but a distinct organization. The shortest ciliary length is associated with the proliferative state of the cells, as confirmed by Ki-67 immunostaining. Primary cilia organization in the growth plate showed that nonpolarized chondrocytes (resting zone) are becoming polarized (proliferating and hypertrophic zones), orienting the primary cilia parallel to the longitudinal axis of the bone. The alignment of primary cilia forms one virtual axis that crosses the center of the columns of chondrocytes reflecting the polarity axis of the growth plate. We also show that primary cilia in osteochondromas are found randomly located on the cell surface. Strikingly, the growth plate-like polarity was retained in sub-populations of osteochondroma cells that were organized into small columns. Based on this, we propose the existence of a mixture ('mosaic') of normal lining (EXT þ /À or EXT wt/wt ) and EXT À/À cells in the cartilaginous cap of osteochondromas.