Genome-Wide DNA Methylation Study Identifies Significant Epigenomic Changes in Osteoarthritic Subchondral Bone and Similarity to Overlying Cartilage.

OBJECTIVE: To perform a genome-wide DNA methylation study to identify differential DNA methylation patterns in subchondral bone underlying eroded and intact cartilage from patients with hip osteoarthritis (OA) and to compare these with DNA methylation patterns in overlying cartilage. METHODS: Genome-wide DNA methylation profiling using Illumina HumanMethylation 450 arrays was performed on eroded and intact cartilage and subchondral bone from within the same joint of 12 patients undergoing hip arthroplasty. Genes with differentially methylated CpG sites were analyzed to identify shared pathways, upstream regulators, and overrepresented gene ontologies, and these patterns were compared with those of the overlying cartilage. Histopathology was graded by modified Mankin score and assessed for correlation with DNA methylation. RESULTS: We identified 7,316 differentially methylated CpG sites in subchondral bone underlying eroded cartilage, most of which (∼75%) were hypomethylated, and 1,397 sites in overlying eroded cartilage, 126 of which were shared. Samples clustered into 3 groups with distinct histopathologic scores. We observed differential DNA methylation of genes including the RNA interference-processing gene AGO2, the growth factor TGFB3, the OA suppressor NFATC1, and the epigenetic effector HDAC4. Among known susceptibility genes in OA, 32 were differentially methylated in subchondral bone, 8 were differentially methylated in cartilage, and 5 were shared. Upstream regulator analysis using differentially methylated genes in OA subchondral bone showed a strong transforming growth factor ß1 signature (P = 1 × 10(-40) ) and a tumor necrosis factor family signature (P = 3.2 × 10(-28) ), among others. CONCLUSION: Our data suggest the presence of an epigenetic phenotype associated with eroded OA subchondral bone that is similar to that of overlying eroded OA cartilage.

Primary intracranial Ewing's sarcoma with unusual features.

Pediatric primary "small round blue cell" tumors in the CNS represent several entities, some more common than others. Ewing sarcoma/peripheral primitive neuroectodermal tumor (ES/pPNET) is rare and must be distinguished from other tumors such as medulloblastoma [1, 2], atypical rhabdoid/teratoid tumor, ependymomal tumors, metastatic sarcomas, hematologic malignancies, and other mimics. Although therapy for ES/pPNET is effective, it brings severe side effects, including cardiac toxicity, making correct recognition important [3]. As small blue cell tumors look similar, diagnosis often depends on special stains, immunohistochemistry, and molecular techniques. While the combination of membranous immunohistochemical reactivity for CD99 with cytoplasmic glycogen provides effective screening, demonstration of characteristic translocations of EWSR1 (chromosome 22) or FUS (chromosome 16) by fluorescent in situ hybridization (FISH) can confirm the diagnosis. We are reporting three primary ES/pPNET of the CNS, two of which occurred in children. While the adult case demonstrates the classic histopathology, the two pediatric cases have histopathology that significantly deviates from the usual. One is suggestive of a primary sarcoma, and the other mimics an ependymoma, but all three cases are confirmed with FISH. These observations suggest that primary ES in the CNS may have histology different from the classic morphology and a high index of suspicion should be maintained in order to make the correct diagnosis. A search of the literature suggests that these tumors are most frequently seen in children and young adults. Imaging often shows a supratentorial enhancing mass that touches the leptomeninges. Survival over three years is good but long term prognosis is unknown [3, 4].

Features of intraventricular tanycytic ependymoma: report of a case and review of literature.

Tanycytic ependymoma is the rarest variant of ependymoma and occurs primarily in the spinal cord. Intracranial cases are even rarer. Only 9 ventricular and 5 subcortical tanycytic ependymoma have been reported in the literature. Amongst the 9 ventricular cases, only one tumor arose from the third ventricle. We report here another case of tanycytic ependymoma arising from the third ventricle completed with immunohistochemical, ultrastructural, and molecular pathology study. The patient was a 44 year-old male who presented with headache, nausea and visual disturbances of a few months duration. Neuroradiological findings showed a well-defined mass arising from the posterolateral wall of third ventricle. Histologically the tumor was composed of monotonous spindle cells arranged in fascicles without definitive perivascular rosettes. The tumor cells were diffusely positive for glial fibrillary acidic protein and epithelial membrane antigen, showed faint immunoreactivity for synaptophysin but were negative for neurofilament proteins and Ki-67 was less than 1%. Molecular studies showed absence of isocitrate dehydrogenase gene 1 and 2 mutation. A diagnosis of tanycytic ependymoma (TE) was made. From literature review with our current case included, intraventricular tanycytic ependymomas ranged from 1.8 to 4.0 cm. The age of patients ranged from 3.5 to 75 years with a mean age of 37.5 and a male predominance. The tumors occurred as well-defined, solitary ventricular mass without significant peritumoral edema with or without cystic changes. Histopathology and immunohistochemical profile are rather similar among different tumors. The immediate to short term outcome is excellent but long term follow up data is lacking.

Genome-wide DNA methylation study identifies significant epigenomic changes in osteoarthritic cartilage.

OBJECTIVE: To perform a genome-wide DNA methylation study to identify DNA methylation changes in osteoarthritic (OA) cartilage tissue. METHODS: The contribution of differentially methylated genes to OA pathogenesis was assessed by bioinformatic analysis, gene expression analysis, and histopathologic severity correlation. Genome-wide DNA methylation profiling of >485,000 methylation sites was performed on eroded and intact cartilage from within the same joint of 24 patients undergoing hip arthroplasty for OA. Genes with differentially methylated CpG sites were analyzed to identify overrepresented gene ontologies, pathways, and upstream regulators. The messenger RNA expression of a subset of differentially methylated genes was analyzed by reverse transcription-polymerase chain reaction. Histopathology was graded by modified Mankin score and correlated with DNA methylation. RESULTS: We identified 550 differentially methylated sites in OA. Most (69%) were hypomethylated and enriched among gene enhancers. We found differential methylation in genes with prior links to OA, including RUNX1, RUNX2, DLX5, FURIN, HTRA1, FGFR2, NFATC1, SNCAIP, and COL11A2. Among these, RUNX1, HTRA1, FGFR2, and COL11A2 were also differentially expressed. Furthermore, we found differential methylation in approximately one-third of known OA susceptibility genes. Among differentially methylated genes, upstream regulator analysis showed enrichment of TGFB1 (P = 4.40 × 10(-5) ) and several microRNAs including miR-128 (P = 4.48 × 10(-13) ), miR-27a (P = 4.15 × 10(-12) ), and miR-9 (P = 9.20 × 10(-10) ). Finally, we identified strong correlations between 20 CpG sites and the histologic Mankin score in OA. CONCLUSION: Our data implicate epigenetic dysregulation of a host of genes and pathways in OA, including a number of OA susceptibility genes. Furthermore, we identified correlations between CpG methylation and histologic severity in OA.

Performance validation in anatomic pathology: successful integration of a new classification system into the practice setting using the updated lung non-small cell carcinoma recommendations.

CONTEXT: The new, international, multidisciplinary classification of lung adenocarcinoma, from the International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society, presents a paradigm shift for diagnostic pathologists. OBJECTIVE: To validate our ability to apply the recommendations in reporting on non-small cell lung cancer cases. DESIGN: A test based on the new non-small cell lung cancer classification was administered to 16 pathology faculty members, senior residents, and fellows before and after major educational interventions, which included circulation of articles, electronic presentations, and live presentations by a well-known lung pathologist. Surgical and cytologic (including cell-block material) reports of lung malignancies for representative periods before and after the educational interventions were reviewed for compliance with the new guidelines. Cases were scored on a 3-point scale, with 1 indicating incorrect terminology and/or highly inappropriate stain use, 2 indicating correct diagnostic terminology with suboptimal stain use, and 3 indicating appropriate diagnosis and stain use. The actual error type was also evaluated. RESULTS: The average score on initial testing was 55%, increasing to 88% following the educational interventions (60% improvement). Of the 54 reports evaluated before intervention, participants scored 3 out of 3 points on 15 cases (28%), 2 of 3 on 31 cases (57%), and 1 of 3 on 8 cases (15%). Incorrect use of stains was noted in 23 of 54 cases (43%), incorrect terminology in 15 of 54 cases (28%), and inappropriate use of tissue, precluding possible molecular testing, in 4 out of 54 cases (7%). Of the 55 cases after intervention, participants scored 3 out of 3 points on 46 cases (84%), 2 of 3 on 8 cases (15%), and 1 of 3 on 1 case (2%). Incorrect use of stains was identified in 9 of 55 cases (16% of total reports), and inappropriate use of tissue, precluding possible molecular testing, was found in 1 of the 55 cases (2%). CONCLUSIONS: The study results demonstrated marked improvement in the pathologists' understanding and application of the new non-small cell lung cancer classification recommendations, which was sufficient to validate our use of the system in routine practice. The results also affirm the value of intensive education on, and validation of, pathologists' use of a classification or diagnostic algorithm.