Integrated copy number and gene expression profiling analysis of Epstein-Barr virus-positive diffuse large B-cell lymphoma.

Viral oncogenes and host immunosenescence have been suggested as causes of Epstein-Barr virus-positive diffuse large B-cell lymphoma (EBV + DLBCL) of the elderly. To investigate the molecular genetic basis of immune evasion and tumor outgrowth, we analyzed copy number alterations (CNAs) and gene expression profiles in EBV + DLBCL samples compared with EBV - DLBCL. There were relatively few genomic alterations in EBV + DLBCL compared with those detected in EBV-negative DLBCL. The most frequent CNAs (>30%) in EBV + DLBCLs were gains at 1q23.2-23.3, 1q23.3, 1q32.1, 5p15.3, 8q22.3, 8q24.1-24.2, and 9p24.1; losses at 6q27, 7q11.2, and 7q36.2-36.3 were also recurrent. A gene expression profile analysis identified the host immune response as a key molecular signature in EBV + DLBCL. Antiviral response genes, proinflammatory cytokines, and chemokines associated with the innate immune response were overexpressed, indicating the presence of a virusinduced inflammatory microenvironment. Genes associated with the B-cell receptor signaling pathway were downregulated. An integrated analysis indicated that SLAMF1 and PDL2 were key targets of the gains detected at 1q23.2-23.3 and 9p24.1. The chromosomal gain at 9p24.1 was associated with poor overall survival. Taken together, our results led to the identification of recurrent copy number alterations and distinct gene expression associated with the host immune response in EBV + DLBCL. We suggest that the upregulation of PDL2 on 9p24.1 promotes immune evasion and is associated with poor prognosis in EBV + DLBCL.

Frequent detection of BRAF(V600E) mutations in histiocytic and dendritic cell neoplasms.

AIMS: In this study, we examined BRAF mutation in a wide range of histiocytic and dendritic cell neoplasms and compared its detection rate in each disease group. METHODS AND RESULTS: A total of 129 cases of histiocytic, dendritic cell and other related lesions were reviewed from the archives of 10 hospitals in Korea. The cases consisted of histiocytic sarcoma, follicular dendritic cell (FDC) sarcoma, interdigitating cell sarcoma, Langerhans cell histiocytosis (LCH), Langerhans cell sarcoma, blastic plasmacytoid dendritic cell neoplasm, acute monocytic leukaemia M5, giant cell tumour, xanthogranuloma, inflammatory myofibroblastic tumour and Rosai-Dorfman disease. BRAF mutation analysis was performed by Sanger sequencing and PNAcqPCR. All the detected mutations of BRAF were V600E. Histiocytic sarcoma exhibited the highest rate of BRAF(V600E) (62.5%, five of eight), followed by Langerhans cell tumours (25%, seven of 28), FDC sarcoma (18.5%, five of 27) and giant cell tumour (6.7%, two of 30). The other tumours did not harbour BRAF mutations. In histiocytic sarcoma, FDC sarcoma and LCH, there were no significant differences in clinical features between tumours containing BRAF(V600E) and those with BRAF(wt) . CONCLUSIONS: BRAF(V600E) was not limited to LCH and was detected more frequently in histiocytic sarcoma. Our findings suggest that the BRAF pathway may contribute to the pathogenesis or malignant transformation of histiocytic and dendritic cell neoplasms.

Development of nano-hydroxyapatite graft with silk fibroin scaffold as a new bone substitute.

PURPOSE: This study involves a comparison between the bone regeneration of nano-hydroxyapatite (nHA), as derived from eggshells either with or without silk fibroin scaffolds, and the unfilled control in the rabbit calvarial bony defect model. MATERIALS AND METHODS: Sixteen 4-month-old New Zealand white rabbits, with a mean weight of 2.8 kg (range, 2.5-3.0 kg), were used in this experiment. After the formation of bilateral parietal bony defects (diameter, 8.0 mm), either an nHA or an nHA+silk fibroin combination (nHA+silk) was grafted. The control was unfilled defect. The bone regeneration was evaluated by micro-computed tomography (µCT) and histomorphometric analyses at 4 and 8 weeks. RESULTS: All measured variables of the µCT analysis were significantly higher in the grafted groups (nHA and nHA+silk) than in the unfilled control groups at both 4 and 8 weeks after operation (P < .05). On histomorphometric analysis, there was no significant difference between the groups at 4 weeks after operation. However, the nHA group exerted significantly higher bone regeneration (40.16% ± 8.27%) compared with the unfilled control group (25.66% ± 10.98%) or the nHA+silk group (16.62% ± 3.05%) (P < .05). CONCLUSION: The nHA from eggshells exerted better bone formation than the unfilled control group on both µCT and histomorphometric analyses. Considering the rapid healing in bony defect and easy availability, the nHA from the eggshells could prove to be a good new bone substitute.

Expression of bone morphogenic protein-4 is inversely related to prevalence of lymph node metastasis in gastric adenocarcinoma.

PURPOSE: Bone morphogenic proteins (BMPs) are the largest subfamily of the transforming growth factor-ß superfamily. Initially characterized as factors that induce bone and cartilage formation, BMPs have been found to be critical during mesoderm formation, organogenesis, and cellular differentiation. Bone morphogenic proteins are also known to modulate the morphologic alteration, adhesion, motility, and invasion of carcinoma cells derived from several organs. However, BMP-4 expression in gastric adenocarcinoma has not yet been clarified. We conducted the present study to define the clinical significance of BMP-4 expression in gastric carcinoma. METHODS: Using immunohistochemistry, we investigated the expression of BMP-4 in normal mucosae and gastric adenocarcinoma samples from 64 patients with gastric carcinoma. RESULTS: The expression of BMP-4 was significantly higher in the adenocarcinoma than in the normal mucosae. Moreover, increased BMP-4 expression was associated with the presence of Helicobacter pylori infection. By contrast, the BMP-4 expression rate in gastric carcinoma was inversely related to the prevalence of lymph node metastasis and tumor invasiveness. CONCLUSIONS: The findings of this study suggest that BMP-4 expression may be a useful prognostic factor for predicting the outcome of patients with gastric carcinoma. Continued investigation to define the pathophysiologic mechanism underlying the role of BMP-4 in gastric carcinoma is warranted.

A combination graft of low-molecular-weight silk fibroin with Choukroun platelet-rich fibrin for rabbit calvarial defect.

OBJECTIVE: The objective of this study was to determine the capabilities of silk fibroin as a biomaterial template for bone formation when mixed with Choukroun platelet-rich fibrin (PRF) in vivo. STUDY DESIGN: Ten New Zealand white rabbits were used for this study and bilateral round shaped defects were formed in the parietal bone (diameter 9.0 mm). The silk fibroin was digested by acid and made into powder (molecular weight <1.0 kDa). The right side (experimental group) received the silk fibroin plus platelet-rich fibroin and the left side (control group) did not receive a graft. Animals were killed at 6 weeks and 12 weeks. The specimens were examined by microscopic computerized tomography (micro-CT). Subsequently, they underwent decalcification and were stained for histologic analysis. RESULTS: There was no significant difference between groups at 6 weeks after operation. In the micro-CT results, however, tissue mineral content in the experimental group at 12 weeks after operation was 132.09 +/- 4.41 and that in the control group was 126.42 +/- 6.62 (P = .011). Tissue mineral density in the experimental group was 2,088.88 +/- 648.34, and that in the control group was 2,029.72 +/- 668.22 (P = .013). The results of the histomorphometric analysis were in accordance with the micro-CT results. The total new bone was 49.86 +/- 7.49% in the control group at 12 weeks after the operation and 59.83 +/- 10.92% in the experimental group (P = .021). CONCLUSION: A combined application of Choukroun PRF with acid-digested silk fibroin showed more rapid bone healing than unfilled control.

The differential expression pattern of BMP-4 between the dentigerous cyst and the odontogenic keratocyst.

BACKGROUND: Bone morphogenic protein-4 (BMP-4) is widely expressed in oral cavity and involved in tooth morphogenesis, cellular differentiation and proliferation. The purpose of this study was to compare the difference in expression pattern of BMP-4 in odontogenic keratocysts (OKC) and dentigerous cysts (DC). METHODS: We evaluated 77 cysts, OKC (n = 34) or DC (n = 43). The average age of patients with OKC was 29.5 +/- 14.4 and that of patients with DC was 36.1 +/- 19.4. The male to female ratio was 20:14 for OKC and 27:16 for DC. Ten cases of OKC were recurrences. Expression of BMP-4 was determined by immunohistochemistry and in situ hybridization. RESULTS: The intensity scales were (-) for invisible or trace staining, (+) for visible staining, and (++) for dense, strong staining. OKCs exhibited the following staining patterns: the epithelium in 15/34 specimens and the mesenchymal cells in 17/34 specimens showed (++) stain. In contrast, the staining pattern of DC was (-) for epithelium in 37/43 specimens. The mesenchymal cells showed (-) degree staining in 30/43 specimens. The difference between the groups studied was significant (P < 0.001 in epithelium and mesenchymal cells). When recurrent and non-recurrent OKC were compared BMP-4 was expressed more intensely in the recurrent cases (P = 0.036 in epithelium). The difference in BMP-4 expression in mesenchymal cells was not significant. In situ hybridization demonstrated positive mRNA probes to BMP-4 were localized in epithelium and mesenchymal cells of OKCs and DCs. CONCLUSIONS: BMP-4 was expressed more intensely in OKC when compared with DC, and was more intensely expressed in recurrent cases.

Expression of osteoprotegerin and RANK ligand in breast cancer bone metastasis.

Bone destruction is primarily mediated by osteoclastic bone resorption, and cancer cells stimulate the formation and activation of osteoclasts next to metastatic foci. Accumulating evidences indicate that receptor activator of NF-kB ligand (RANKL) is the ultimate extracellular mediator that stimulates osteoclast differentiation into mature osteoclasts. In contrast, osteoprotegerin (OPG) inhibits osteoclast development. In order to elucidate a mechanism for cancer-induced osteoclastogenesis, cells from a human breast cancer line, MDA-MB-231, were directly co-cultured with ST2, MC3T3-E1, or with primary mouse calvarial cells. Osteoclast-like cells and tartarate resistant acid phosphatase (TRAP) activities were then quantitated. We examined these cell lines and samples from breast cancer by RT-PCR for the expressions of OPG and RANKL mRNA. Compared to controls, co-culture of MDA-MB-231 cells with stromal or osteoblastic cells induced an increase in number of osteoclasts and TRAP activities. MDA-MB-231 cells alone or breast cancer samples did not express RANKL mRNA. However, co-culture of these cancer cells with stromal or osteoblastic cells induced RANKL mRNA expression and decreased OPG mRNA expression. These experiments demonstrate that direct interactions between breast cancer and stromal or osteoblastic cells induce osteoclastogenesis in vitro through modulating RANKL expression.