Inactivation of Lkb1 in postnatal chondrocytes leads to epiphyseal growth-plate abnormalities and promotes enchondroma-like formation.

Liver serine-threonine kinase B1 (LKB1) is a tumor suppressor that has been linked to many types of tumors. However, the role of LKB1 in cartilaginous tumorigenesis is still poorly understood. In this study, we find that cartilage-specific, tamoxifen-inducible Lkb1 knockout results in multiple enchondroma-like lesions adjacent to the disorganized growth plates. We showed that chondrocytes retain an immature status caused by loss of Lkb1, which may lead to the dramatic expansion of growth-plate cartilage and the formation of enchondroma-like lesions. Additionally, increased mammalian target of rapamycin complex 1 (mTORC1) activity is observed in the Lkb1 conditional knockout (cKO) chondrocytes, and rapamycin (mTORC1 inhibitor) treatment significantly alleviates the expansion of growth-plate cartilage and eliminates the enchondroma-like lesions in Lkb1 cKO mice. Thus, our findings indicate that loss of Lkb1 leads to the expansion of chondrocytes and the formation of enchondroma-like lesions during postnatal cartilage development, and that the up-regulated mTORC1-signaling pathway is implicated in this process. Our findings suggest that modulation of LKB1 and related signaling is a potential therapy in cartilaginous tumorigenesis.-Zhou, S., Li, Y., Qiao, L., Ge, Y., Huang, X., Gao, X., Ju, H., Wang, W., Zhang, J., Yan, J., Teng, H., Jiang, Q. Inactivation of Lkb1 in postnatal chondrocytes leads to epiphyseal growth-plate abnormalities and promotes enchondroma-like formation.

Metabolic profiling of serum in patients with cartilage tumours using 1 H-NMR spectroscopy: A pilot study.

Cartilage-forming lesions include tumours that can vary in severity from benign enchondromas to high-grade malignant chondrosarcomas. Chondrosarcoma is the second most frequent malignant bone tumour, accounting for 20-30% of all malignant bone neoplasms. Surgery is the standard treatment for cartilage tumours (CTs); however, their incidental diagnosis and the difficult differentiation of low-grade lesions like chondrosarcoma grade I from benign entities like enchondroma are challenges for clinical management. In this sense, the search for circulating biomarkers for early detection and prognosis is an ongoing interest. Targeted metabolomics is a powerful tool that can propose potential biomarkers in biological fluids as well as help to discover disturbed metabolic pathways to reveal tumour pathogenesis. In this context, the aim of this study was to investigate the 1 H nuclear magnetic resonance metabolomic serum profile of patients with CTs contrasted with healthy controls. Forty-one metabolites were identified and quantified; the multivariate statistical methods principal component analysis and partial least squares discriminant analysis reveal a clear separation of the CT group, that is, the differential metabolites that were involved in two main metabolic pathways: the taurine and hypotaurine metabolism and synthesis and degradation of ketone bodies. Our results represent preliminary work for emergent serum-based diagnostics or prognostic methods for patients with chondrogenic tumours.

Paragangliomas in Carney-Stratakis Syndrome.

Carney-Stratakis Syndrome (CSS) comprises of paragangliomas (PGLs) and gastrointestinal stromal tumors (GISTs). Several of its features overlap with Carney Triad (CT) - PGLs, GISTs, and pulmonary chondromas. CSS has autosomal dominant inheritance, incomplete penetrance, and greater relative frequency of PGL over GISTs. The PGLs in CSS are multicentric and GISTs are multifocal in all the patients, suggesting an inherited susceptibility and associating the two manifestations. In this review, we highlight the clinical, pathological, and molecular characteristics of CSS, along with its diagnostic and therapeutic implications.

MTNR1B loss promotes chordoma recurrence by abrogating melatonin-mediated ß-catenin signaling repression.

Chordoma is an extremely rare malignant bone tumor with a high rate of relapse. While cancer stem cells (CSCs) are closely associated with tumor recurrence, which depend on its capacity to self-renew and induce chemo-/radioresistance, whether and how CSCs participate in chordoma recurrence remains unclear. The current study found that tumor cells in recurrent chordoma displayed more dedifferentiated CSC-like properties than those in corresponding primary tumor tissues. Meanwhile, MTNR1B deletion along with melatonin receptor 1B (MTNR1B) down-regulation was observed in recurrent chordoma. Further investigation revealed that activation of Gαi2 by MTNR1B upon melatonin stimulation could inhibit SRC kinase activity via recruiting CSK and SRC, increasing SRC Y530 phosphorylation, and decreasing SRC Y419 phosphorylation. This subsequently suppressed ß-catenin signaling and stemness via decreasing ß-catenin p-Y86/Y333/Y654. However, MTNR1B loss in chordoma mediated increased CSC properties, chemoresistance, and tumor progression by releasing melatonin's repression of ß-catenin signaling. Clinically, MTNR1B deletion was found to correlate with patients' survival. Together, our study establishes a novel convergence between melatonin and ß-catenin signaling pathways and reveals the significance of this cross talk in chordoma recurrence. Besides, we propose that MTNR1B is a potential biomarker for prediction of chordoma prognosis and selection of treatment options, and chordoma patients might benefit from targeting MTNR1B/Gαi2/SRC/ß-catenin axis.

Spontaneous chondroma formation in CD2-Cre-driven Erk-deficient mice.

Lineage-specific Cre Tg mice are widely used to delineate the functions of genes in a tissue-specific manner. Several T-cell-specific promoter cassettes have been developed; however, the activities of those promoters in non-T cells have not been investigated extensively. Here, we report that CD2-Cre-mediated deletion of Erk proteins by generating CD2-Cre × Erk1-/-Erk2flox/flox (Erk∆CD2-Cre) mice results in abnormal cartilage hyperplasia. Histological analysis revealed that this abnormality is caused by aberrant hyperplasia of chondrocytes. The presence of Erk-deficient T cells is not required for this chondroma formation, as it was similarly observed in the absence of T cells in a CD3ε-deficient background. In addition, adoptive transfer of bone marrow cells from Erk∆CD2-Cre mice to wild-type recipients did not cause chondroma formation, suggesting that Erk-deficient non-immune cells are responsible for this abnormality. By tracing Cre-expressed tissues using a ROSA26-STOP-RFP allele, we found that the chondroma emitted RFP fluorescence, indicating that functional Cre is expressed in hyperplastic chondrocytes in Erk∆CD2-Cre mice. Furthermore, RFP+ chondrocytes were also found in an Erk-sufficient background, albeit without aberrant growth. These results suggest that unexpected expression of CD2-driven Cre in chondrocytes generates Erk-deficient chondrocytes, resulting in hyperplastic cartilage formation. Recently, two independent reports showed that CD4-Cre-mediated Ras-Erk signaling ablation led to similar abnormal cartilage formation (Guittard, G., Gallardo, D. L., Li, W. et al. 2017. Unexpected cartilage phenotype in CD4-Cre-conditional SOS-deficient mice. Front. Immunol. 8:343; Wehenkel, M., Corr, M., Guy, C. S. et al. 2017. Extracellular signal-regulated kinase signaling in CD4-expressing cells inhibits osteochondromas. Front. Immunol. 8:482). Together with these reports, our study suggests that an unexpected link exists between T-like cell and chondrocyte lineages during ontogeny.

FGFR3 Deficiency Causes Multiple Chondroma-like Lesions by Upregulating Hedgehog Signaling.

Most cartilaginous tumors are formed during skeletal development in locations adjacent to growth plates, suggesting that they arise from disordered endochondral bone growth. Fibroblast growth factor receptor (FGFR)3 signaling plays essential roles in this process; however, the role of FGFR3 in cartilaginous tumorigenesis is not known. In this study, we found that postnatal chondrocyte-specific Fgfr3 deletion induced multiple chondroma-like lesions, including enchondromas and osteochondromas, adjacent to disordered growth plates. The lesions showed decreased extracellular signal-regulated kinase (ERK) activity and increased Indian hedgehog (IHH) expression. The same was observed in Fgfr3-deficient primary chondrocytes, in which treatment with a mitogen-activated protein kinase (MEK) inhibitor increased Ihh expression. Importantly, treatment with an inhibitor of IHH signaling reduced the occurrence of chondroma-like lesions in Fgfr3-deficient mice. This is the first study reporting that the loss of Fgfr3 function leads to the formation of chondroma-like lesions via downregulation of MEK/ERK signaling and upregulation of IHH, suggesting that FGFR3 has a tumor suppressor-like function in chondrogenesis.

Use of MicroRNA biomarkers to distinguish enchondroma from low-grade chondrosarcoma.

Establishing a definitive diagnosis between benign enchondroma versus low-grade chondrosarcoma presents a potential challenge to both clinicians and pathologists. microRNAs (small non-coding RNAs) have proven to be effective biomarkers for the identification of tumors and tumor progression. We present analysis, both array and quantitative PCR, that shows consistently and substantially increased expression of two microRNAs, miRs-181a and -138, in low-grade chondrosarcomas compared with enchondromas. The data suggest these microRNAs would provide an analytical distinction between the chondrosarcoma and benign neoplasms that can be performed in formalin-fixed paraffin-embedded specimens. Together with recent publications, these data indicate that miRs-181a and -138 also play a role in tumor development and homeostasis and may provide new targets for the development of much needed therapeutic intervention.

Identification of novel biomarker candidates for immunohistochemical diagnosis to distinguish low-grade chondrosarcoma from enchondroma.

Chondrosarcoma is the third most common primary bone cancer, requiring surgical resection. However, differentiation of low-grade chondrosarcoma (grade 1) from enchondroma that is benign and only requires regular follow-up is one of the most frequent diagnostic dilemmas facing orthopedic oncologists in clinical management. Although multiple techniques are applied to make the distinction, immunohistochemistry is an important ancillary technique, especially when a histopathological stain of specimen must be obtained in order to guarantee an accurate confirmation. Currently, no adequate immunohistochemical diagnostic protein biomarkers are available to distinguish low-grade chondrosarcoma from enchondroma. To discover novel protein biomarker candidates, an LC-MS/MS approach was applied to directly compare formalin-fixed, paraffin-embedded low-grade chondrosarcoma with enchondroma tissue samples. The proteomics analysis revealed 17 protein biomarker candidates. A principle was developed to prioritize the candidates using category and ranking. An algorithm, prioritization index of biomarker candidates for immunohistochemistry on tissue specimens, was developed to rank the candidates inside each category. Using the proteomics data and bioinformatics results, the prioritization index of biomarker candidates for immunohistochemistry on tissue revealed periostin as a top candidate. Immunohistochemical staining of periostin in 23 low-grade chondrosarcoma and 31 enchondroma tissue specimens disclosed 87% specificity and 70% sensitivity.

Loss of ß-catenin induces multifocal periosteal chondroma-like masses in mice.

Osteochondromas and enchondromas are the most common tumors affecting the skeleton. Osteochondromas can occur as multiple lesions, such as those in patients with hereditary multiple exostoses. Unexpectedly, while studying the role of ß-catenin in cartilage development, we found that its conditional deletion induces ectopic chondroma-like cartilage formation in mice. Postnatal ablation of ß-catenin in cartilage induced lateral outgrowth of the growth plate within 2 weeks after ablation. The chondroma-like masses were present in the flanking periosteum by 5 weeks and persisted for more than 6 months after ß-catenin ablation. These long-lasting ectopic masses rarely contained apoptotic cells. In good correlation, transplants of ß-catenin-deficient chondrocytes into athymic mice persisted for a longer period of time and resisted replacement by bone compared to control wild-type chondrocytes. In contrast, a ß-catenin signaling stimulator increased cell death in control chondrocytes. Immunohistochemical analysis revealed that the amount of detectable ß-catenin in cartilage cells of osteochondromas obtained from hereditary multiple exostoses patients was much lower than that in hypertrophic chondrocytes in normal human growth plates. The findings in our study indicate that loss of ß-catenin expression in chondrocytes induces periosteal chondroma-like masses and may be linked to, and cause, the persistence of cartilage caps in osteochondromas.

Ectomesenchymal chondromyxoid tumor: histopathologic and immunohistochemical study of two cases without a chondroid component.

Ectomesenchymal chondromyxoid tumor (ECT) is a rare benign neoplasm usually affecting the anterior dorsum of the tongue. Histopathologically, it is formed by spindle, round and/or polygonal cells embedded in a chondromyxoid matrix. Immunohistochemical positivity for vimentin, S-100 protein, glial fibrillary acid protein and neuron-specific enolase are helpful to confirm the diagnosis. There are 42 cases of ECT of the tongue reported in the English language literature, three of them showing no chondroid matrix. We describe two additional cases of ECT lacking the chondroid component, exhibiting areas of reticulated myxoid and cellular pattern. Considering the microscopical features, ECT can be classified in classic and 'chondroid-free' variants, the latter including the reticulated myxoid and cellular patterns. It is important to consider that the cellular ECT usually exhibits predominance of an infiltrative atypical cellular component that may mimic a malignant tumor.

Distinct Roles of Glutamine Metabolism in Benign and Malignant Cartilage Tumors With IDH Mutations.

Enchondromas and chondrosarcomas are common cartilage neoplasms that are either benign or malignant, respectively. The majority of these tumors harbor mutations in either IDH1 or IDH2. Glutamine metabolism has been implicated as a critical regulator of tumors with IDH mutations. Using genetic and pharmacological approaches, we demonstrated that glutaminase-mediated glutamine metabolism played distinct roles in enchondromas and chondrosarcomas with IDH1 or IDH2 mutations. Glutamine affected cell differentiation and viability in these tumors differently through different downstream metabolites. During murine enchondroma-like lesion development, glutamine-derived α-ketoglutarate promoted hypertrophic chondrocyte differentiation and regulated chondrocyte proliferation. Deletion of glutaminase in chondrocytes with Idh1 mutation increased the number and size of enchondroma-like lesions. In contrast, pharmacological inhibition of glutaminase in chondrosarcoma xenografts reduced overall tumor burden partially because glutamine-derived non-essential amino acids played an important role in preventing cell apoptosis. This study demonstrates that glutamine metabolism plays different roles in tumor initiation and cancer maintenance. Supplementation of α-ketoglutarate and inhibiting GLS may provide a therapeutic approach to suppress enchondroma and chondrosarcoma tumor growth, respectively. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

SDHC Methylation Pattern in Patients With Carney Triad.

Carney triad is a multitumor syndrome affecting almost exclusively young women in a nonfamilial setting, which manifests by multifocal gastric gastrointestinal stromal tumors, paragangliomas, and pulmonary chondroma. The Carney triad-associated tumors are characterized by a deficiency of the mitochondrial succinate dehydrogenase enzymatic complex. Recently, it has been observed that the deficiency results from epigenetic silencing of the SDHC gene by its promoter hypermethylation. To elucidate anatomic distribution of SDHC promoter methylation in Carney triad patients and thus to shed some light on the possible natural development of this epigenetic change, both neoplastic and available non-neoplastic tissues of 3 patients with Carney triad were tested for hypermethylation at the SDHC promoter site. SDHC promoter hypermethylation was proven in all tumors studied. Lack of SDHC epigenetic silencing in the non-neoplastic lymphoid and duodenal tissue (ie, tissues not involved in the development of Carney triad-associated tumors) together with the finding of SDHC promoter hypermethylation in the non-neoplastic gastric wall favors the hypothesis of postzygotic somatic mosaicism as the biological background of Carney triad; it also offers an explanation of the multifocality of gastrointestinal stromal tumors of the stomach occurring in this scenario as well. However, the precise mechanism responsible for the peculiar organ-specific distribution of Carney triad-associated tumors is still unknown.

PTHR1 mutations associated with Ollier disease result in receptor loss of function.

PTHR1-signaling pathway is critical for the regulation of endochondral ossification. Thus, abnormalities in genes belonging to this pathway could potentially participate in the pathogenesis of Ollier disease/Maffucci syndrome, two developmental disorders defined by the presence of multiple enchondromas. In agreement, a functionally deleterious mutation in PTHR1 (p.R150C) was identified in enchondromas from two of six unrelated patients with enchondromatosis. However, neither the p.R150C mutation (26 tumors) nor any other mutation in the PTHR1 gene (11 patients) could be identified in another study. To further define the role of PTHR1-signaling pathway in Ollier disease and Maffucci syndrome, we analyzed the coding sequences of four genes (PTHR1, IHH, PTHrP and GNAS1) in leucocyte and/or tumor DNA from 61 and 23 patients affected with Ollier disease or Maffucci syndrome, respectively. We identified three previously undescribed missense mutations in PTHR1 in patients with Ollier disease at the heterozygous state. Two mutations (p.G121E, p.A122T) were present only in enchondromas, and one (p.R255H) in both enchondroma and leukocyte DNA. Assessment of receptor function demonstrated that these three mutations impair PTHR1 function by reducing either the affinity of the receptor for PTH or the receptor expression at the cell surface. These mutations were not found in DNA from 222 controls. Including our data, PTHR1 functionally deleterious mutations have now been identified in five out 31 enchondromas from Ollier patients. These findings provide further support for the idea that heterozygous mutations in PTHR1 that impair receptor function participate in the pathogenesis of Ollier disease in some patients.

Increased expression of matrilin-3 not only in osteoarthritic articular cartilage but also in cartilage-forming tumors, and down-regulation of SOX9 via epidermal growth factor domain 1-dependent signaling.

OBJECTIVE: To identify regulators of the cartilaginous phenotype, on the basis of their differential expression in human conventional chondrogenic tumors compared with articular cartilage. METHODS: Differential proteomics analysis revealed matrilin-3 (MATN3) as a candidate regulator of the cartilaginous phenotype. Its capacity to modulate gene expression was investigated in human HCS-2/8 chondrosarcoma cells and transfected chondrocytes, using cell culture fractionation, reverse transcription-polymerase chain reaction, and Western blot analyses. RESULTS: Increased expression of the cartilage-specific matrix protein MATN3 was specifically observed in enchondromas and conventional chondrosarcomas. A substantial fraction of MATN3 was found in cytoplasmic structures of tumor cells, as demonstrated by immunohistochemistry. Analyses of intracellular MATN3 revealed that it corresponded to an imperfectly maturated MATN3 polypeptide, both in HCS-2/8 human chondrosarcoma cells and in transfected human chondrocytes. Moderately increased expression of MATN3 resulted in its intracellular retention. Antibody-mediated blockade of soluble, extracellular MATN3 in HCS-2/8 cell cultures resulted in increased expression of MATN3 and the chondrogenic transcription factor SOX9. Conversely, increased ectopic expression of MATN3 resulted in decreased expression of MATN3 and SOX9 in primary chondrocytes, while a mutant MATN3 lacking its first epidermal growth factor (EGF)-like domain failed to down-regulate SOX9. CONCLUSION: Aberrant expression and processing of MATN3 are hallmarks of conventional cartilaginous neoplasms. A particular step in the maturation of MATN3 limits its processing through the secretion machinery, resulting in its intracellular accumulation upon increased expression. Soluble, secreted MATN3, however, down-regulates SOX9 at the messenger RNA and protein levels. The first EGF-like domain of MATN3 is a critical determinant of its regulatory activity toward SOX9. These activities of MATN3 suggest that its increased expression in osteoarthritis might contribute to the degeneration of articular cartilage.

Hypoxia-inducible factor 1-alpha and vascular endothelial growth factor in cartilage tumors.

Neoangiogenesis has been demonstrated in chondrosarcoma. Anti-angiogenic therapies are being tested in clinical trials for chondrosarcomas. Studies of the underlying mechanisms have been performed almost exclusively in cell lines. We immunostained 20 samples of chondrosarcoma and 20 samples of enchondromas with antibodies against hypoxia-inducible factor 1-alpha (HIF-1-alpha) and vascular endothelial growth factor (VEGF). The immunohistochemical staining of HIF-1-alpha and VEGF were highly correlated. Enchondromas were HIF-1-alpha and VEGF negative, whereas all chondrosarcoma exhibited HIF-1-alpha and VEGF immunostaining. HIF-1-alpha/VEGF double positive cases were almost exclusively chondrosarcomas with a high tumor grade. We suggest that HIF-1-alpha is a marker of malignancy in chondrosarcomas that correlates with tumor neo-angiogenesis. Our findings also suggest that a HIF-1-alpha/VEGF angiogenic pathway may exist in chondrosarcoma in vivo as in other malignant tumors. The inclusion of novel inhibitors to HIF-1-alpha and other factors may optimize anti-angiogenic interventions in chondrosarcoma.

Intracellular cholesterol biosynthesis in enchondroma and chondrosarcoma.

Enchondroma and chondrosarcoma are the most common benign and malignant cartilaginous neoplasms. Mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) are present in the majority of these tumors. We performed RNA-seq analysis on chondrocytes from Col2a1Cre;Idh1LSL/+ animals and found that genes implied in cholesterol synthesis pathway were significantly upregulated in the mutant chondrocytes. We examined the phenotypic effect of inhibiting intracellular cholesterol biosynthesis on enchondroma formation by conditionally deleting SCAP (sterol regulatory element-binding protein cleavage-activating protein), a protein activating intracellular cholesterol synthesis, in IDH1 mutant mice. We found fewer enchondromas in animals lacking SCAP. Furthermore, in chondrosarcomas, pharmacological inhibition of intracellular cholesterol synthesis significantly reduced chondrosarcoma cell viability in vitro and suppressed tumor growth in vivo. Taken together, these data suggest that intracellular cholesterol synthesis is a potential therapeutic target for enchondromas and chondrosarcomas.

IDH1 immunohistochemistry reactivity and mosaic IDH1 or IDH2 somatic mutations in pediatric sporadic enchondroma and enchondromatosis.

Mosaic somatic mutations in the isocitrate dehydrogenase 1/2 (IDH1/2) genes have been identified in most enchondromas by targeted mutation analysis. Next-generation sequencing (NGS), that may detect even low-level mosaic mutation rates, has not previously been applied to enchondromas. Immunohistochemistry using the H09 clone is routinely used as a surrogate for the common R132H IDH1 mutation in gliomas. We compared immunohistochemistry and NGS results in a series of 13 enchondromas from 8 pediatric patients. NGS identified a heterozygous IDH mutation in all enchondromas, showing identical mutation status in patients with multiple tumors assessed, thereby confirming somatic mosaicism. A majority of the tumors harbored an IDH1 mutation (p.R132H in 3 tumors; p.R132C in 4 tumors from 2 patients; p.R132L and p.R132G in one tumor each). A p.R172S IDH2 mutation was identified in 4 enchondromas, but not in the ependymoma from one patient with Ollier disease, who further displayed a heterozygous STK11 missense mutation. IDH mutation rates varied between 14% (indicative of mutations in 28% of the cells and of intratumoral mosaicism) and 45% (tumor content was close to 100%). Cytoplasmic H09 reactivity was observed as expected in tumors with an IDH1 p.R132H mutation; cross-reactivity was seen with the p.R132L variant. This first NGS study of pediatric enchondromas confirms that IDH mutations may occur in a mosaic fashion. STK11 gene mutations may provide insights in the development of multiple cartilaginous tumors in enchondromatosis, this tumor suppressor gene having been shown in animal models to regulate both chondrocyte maturation and growth plate organization during development.

Immunohistochemical expression of estrogen receptors in chondrosarcomas and enchondromas.

Immunohistochemical expression of estrogen receptors alpha and beta was studied in chondrosarcomas and enchondromas and was correlated with chondrosarcoma grade, type, and dedifferentiation. Estrogen receptor alpha was studied in 37 chondrosarcomas, 10 enchondromas, and 2 extraskeletal myxoid chondrosarcomas. Estrogen receptor beta was studied in 23 chondrosarcomas, 6 enchondromas, and 2 extraskeletal myxoid chondrosarcomas. Ventana prediluted monoclonal anti-ER alpha (clone 6F11) and Biogenex prediluted polyclonal anti-ER beta were used on the Ventana ES autostainer and BenchMark XT IHC/ISH, respectively. Percent of cell staining and intensity (+, ++, or +++) was evaluated. Overall, 61% of conventional chondrosarcoma and 60% of enchondroma were positive for estrogen receptor alpha. Low-grade chondrosarcoma expressed estrogen receptor alpha more frequently than high-grade chondrosarcoma (P

Differential expression of runx2 and Indian hedgehog in cartilaginous tumors.

Runx2-Cbfa1, a Runt transcription factor, plays important roles during skeletal development. In its absence, chondrocyte hypertrophy is severely impaired and there is no vascularization of cartilage templates during skeletal development. In addition, Indian hedgehog (Ihh) signaling molecules control the space and timing of chondrocyte differentiation. Our goal was to gain a better understanding of the molecular process underlying the development of chondrosarcoma and to investigate whether there is a biological difference among variable types of chondrosarcomas. To accomplish this we collected a series of 10 enchondromas and 57 chondrosarcomas (conventional, n = 17; mesenchymal, n = 20; clear cell, n = 20), and investigated the expression of Runx2 and Ihh in these cartilaginous tumors by immunohistochemistry. Cellular and matrix-rich areas were evaluated separately. Runx2 was expressed in 100% of conventional, mesenchymal, and clear cell chondrosarcomas, and in 30% of enchondromas. Higher levels of expression of Runx2 were found in cellular areas than in matrixrich areas. Expression levels increased with increasing histological grade in conventional chondrosarcoma, suggesting involvement in tumor progression. Ihh was expressed in 100% of conventional and clear cell chondrosarcomas, especially in matrix-rich areas. Mesenchymal chondrosarcomas revealed only focal expression of Ihh in matrix-rich areas. Small cell areas were negative. Ihh was absent or focally expressed in enchondromas. These findings demonstrate that Runx2 expression is active in variable chondrosarcomas compared to enchondromas, suggesting its importance in growth and differentiation of neoplastic cartilage. Ihh expression is considered a marker of the hypertrophic stage of differentiation in these tumor cells.

Exploring the key genes and pathways in enchondromas using a gene expression microarray.

Enchondromas are the most common primary benign osseous neoplasms that occur in the medullary bone; they can undergo malignant transformation into chondrosarcoma. However, enchondromas are always undetected in patients, and the molecular mechanism is unclear. To identify key genes and pathways associated with the occurrence and development of enchondromas, we downloaded the gene expression dataset GSE22855 and obtained the differentially expressed genes (DEGs) by analyzing high-throughput gene expression in enchondromas. In total, 635 genes were identified as DEGs. Of these, 225 genes (35.43%) were up-regulated, and the remaining 410 genes (64.57%) were down-regulated. We identified the predominant gene ontology (GO) categories and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways that were significantly over-represented in the enchondromas samples compared with the control samples. Subsequently the top 10 core genes were identified from the protein-protein interaction (PPI) network. The enrichment analyses of the genes mainly involved in two significant modules showed that the DEGs were principally related to ribosomes, protein digestion and absorption, ECM-receptor interaction, focal adhesion, amoebiasis and the PI3K-Akt signaling pathway.Together, these data elucidate the molecular mechanisms underlying the occurrence and development of enchondromas and provide promising candidates for therapeutic intervention and prognostic evaluation. However, further experimental studies are needed to confirm these results.