Paraffin-embedded tissue samples from 30 cases of non-Hodgkin's lymphoma(NHL) and 10 of reactive hyperplasia, were processed for interphase cytogenetic chromosomal study. We performed non-fluorescent in situ hybridization(NFISH) using the enzymatic method with digoxigenin-labeled DNA centromeric probes for chromosome 7,12,18 and X, and a painting probe for chromosome 18. Chromosomal aberrations were observed in 27(90%) out of 30 cases of NHL. The most commonly observed numerical aberration was extracopy of X chromosome. There were some characteristic aberrations corresponding to each grade and group of NHL by International Working Formulation: In low grade NHL(9 cases), a third were associated with extracopy of chromosome 12, and disomy X was frequently found in small lymphocytic lymphoma(75%). With intermediate grade(16 cases), tetraploidy(25%), translocation of chromosome 18(25%), and extracopy of chromosome 18(19%) were characteristically associated. These results suggest that interphase NFISH is an easily performable method in retrograde cytogenetic study of archival materials. Some specifically correlated chromosomal aberrations corresponding to the histopathologic grades and groups could provide us more valuable information for determining pathologic diagnosis and assessing the clinical outcome of NHL.
*Chromosome Aberrations ; Human ; Immunophenotyping ; *In Situ Hybridization ; Interphase ; Lymphoma, Non-Hodgkin/*genetics/pathology ; Paraffin Embedding ; Pseudolymphoma/genetics/pathology

OBJECTIVEwe aimed to investigate the mutation and expression of BRAF gene in mature T/NK cell lymphoma tissues and cell lines, explore the correlation between gene alterations and clinicopathological features and clinical outcomes of mature T/NK cell lymphoma.

METHODSFirstly, we detected common mutant sites of BRAF (locus 1 799 mutation in exon 15 and loci 463, 465 and 468 mutation in exon 11) in lymphoma Jurkat, Hut-78 and YTS cell lines, normal peripheral blood lymphocytes, different types of mature T/NK cell lymphoma and reactive hyperplasia lymph nodes by direct sequencing. Then we measured the expression of BRAF in Jurkat, Hut-78, YTS cells and normal peripheral blood lymphocytes by real time-PCR and Western-blot detection. We also used immunohistochemistry (IHC) to detect the expression of BRAF in mature T/NK cell lymphoma tissues and reactive hyperplasia lymph nodes, and to analyze the correlation between the expression of BRAF and clinocopathological features and clinical outcomes.

RESULTSWe did not find common BRAF mutation in mature T/NK cell lymphoma tissues and cell lines, and the relatively expression of BRAF gene mRNA in normal peripheral blood lymphocytes, YTS, Hut-78 and Jurkat cells were 1.000, 5.207±0.013, 8.412±0.615 and 36.720±1.797, respectively, and protein expressions were 0.051±0.003, 0.102±0.013, 0.113±0.017 and 0.304±0.010, respectively, and the expression of BRAF in peripheral T cell lymphoma Jurkat cells was significantly higher than that of Hut-78, YTS cells and normal lymphocytes (P<0.05). Only 6 of 58 peripheral T cell lymphomas (10.3%) had positive BRAF expression, and were the subgroups of peripheral T cell lymphoma-unspecified type. The statistical data did not show any correlation between positive expression of BRAF and gender, age, clinical stage, location, lactate dehydrogenase in the 21 cases of peripheral T cell lymphoma-unspecified type (P<0.05), but the positive rate of BRAF in the effective treatment group (8.3%) was significantly lower than that of the invalid group (55.6%, P<0.05).

CONCLUSIONThe expression of BRAF gene may become a marker of malignant biological characteristics and clinical therapeutic target of peripheral T cell lymphoma.

Exons ; Humans ; Immunohistochemistry ; Killer Cells, Natural ; Lymphoma, T-Cell, Peripheral ; genetics ; metabolism ; pathology ; Proto-Oncogene Proteins B-raf ; genetics ; metabolism ; Pseudolymphoma ; genetics ; metabolism ; RNA, Messenger ; metabolism

OBJECTIVETo evaluate the diagnostic role of nuclear expression of bcl-10 protein in extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT) type.

METHODSOne hundred and forty cases of MALT lymphoma were collected from Cancer Hospital of Fudan University (including 38 cases from stomach, 35 cases from ocular adnexa, 16 cases from intestine, 15 cases from skin, 15 cases from salivary gland, 14 cases from lung, 3 cases from thyroid and 4 cases from other sites). Ten cases of reactive follicular hyperplasia of tonsil, 5 cases of reactive lymphoid hyperplasia of orbit and 143 cases of non-Hodgkin's lymphoma other than MALT lymphoma (including 20 cases of NK/T cell lymphoma, 20 cases of follicular lymphomas, 20 cases of anaplastic large cell lymphomas, 20 cases of nodal diffuse large cell B-cell lymphoma (DLBCL), 10 cases of gastric diffuse large B-cell lymphoma, 13 cases of nodal marginal zone B-cell lymphoma, 12 cases of mantle cell lymphoma, 11 cases of splenic marginal zone B-cell lymphoma, 6 cases of angioimmunoblastic T-cell lymphoma, 6 cases of peripheral T-cell lymphoma, not otherwise specified, 3 cases of small lymphocytic lymphoma, 1 case of lymphoplasmacytic lymphoma and 1 case of plasmacytoma were used as controls. Immunohistochemical study for bcl-10, as well as dual staining with CD20, was performed by EnVision method in paraffin sections.

RESULTSIn reactive follicular hyperplasia of tonsil, bcl-10 was moderately or strongly expressed in the cytoplasm of germinal center B cells, while the mantle cells were negative and the marginal zone cells and paracortical T cells showed weak staining. In the 5 cases of reactive lymphoid hyperplasia of orbit, 2 were bcl-10-negative and the remaining 3 expressed bcl-10 in the cytoplasm of germinal center B cells. As for non-MALT lymphomas, 3 gastric DLBCL showed nuclear expression. The remaining cases showed variable cytoplasmic staining. In some cases of lymphoma, bcl-10 was expressed in tumor cells but not in reactive lymphoid cells. On the other hand, 92.1% (129/140) of MALT lymphoma were bcl-10 positive. Among those cases, 54.3% (76/140) showed cytoplasmic positivity and 37.9% (53/140) showed nuclear positivity. The nuclear positivity rate of bcl-10 in different anatomic sites was different. The staining was most intense in MALT lymphoma of ocular adnexa. Dual staining with CD20 showed that the bcl-10-positive cells were also CD20-positive, though the number of bcl-10-positive cells were less than that of CD20-positive cells.

CONCLUSIONSBcl-10 expression in lymphoid hyperplasia is a universal phenomenon. Cytoplasmic expression of bcl-10 is seen in many different kinds of non-Hodgkin's lymphoma and reactive lymphoid conditions. In some cases of lymphoma, bcl-10 is expressed in tumor cells but not in reactive lymphoid cells, suggesting a possible role of abnormal bcl-10 expression in tumorgenesis. Nuclear expression of bcl-10 is seen mainly in MALT lymphoma, especially when occurring in ocular adnexa and lung. This is in contrast to loss of bcl-10 expression in residual germinal center cells.

Adaptor Proteins, Signal Transducing ; genetics ; Antigens, CD20 ; immunology ; B-Cell CLL-Lymphoma 10 Protein ; Cell Nucleus ; genetics ; Cytoplasm ; genetics ; Gene Expression Regulation, Neoplastic ; Humans ; Lymphocytes ; pathology ; Lymphoma, B-Cell, Marginal Zone ; genetics ; immunology ; pathology ; Palatine Tonsil ; pathology ; Pseudolymphoma ; genetics

OBJECTIVETo investigate bcl-6 protein expression and gene rearrangement patterns in diffuse large B-cell lymphoma (DLBCL) and their clinicopathologic significance.

METHODSImmunohistochemical studies for bcl-6 and CD10 proteins were performed on 51 cases of DLBCL paraffin-embedded tissues (including 22 nodal samples and 29 extranodal samples) and 10 cases of reactive lymphoid hyperplasia (RLH) paraffin-embedded tissues. Interphase fluorescence in-situ hybridization (FISH) with dual color breakapart probe was also used to identify rearrangement of bcl-6 gene in 32 cases of nodal DLBCL tissues (including 22 paraffin-embedded samples and 10 fresh samples) and 5 cases of RLH paraffin-embedded tissues.

RESULTS(1) The rates of bcl-6 protein expression in nodal DLBCL, extranodal DLBCL and RLH were 72.7% (16/22), 75.9% (22/29) and 100.0% (10/10) respectively. The rates of CD10 expression were 40.9% (9/22), 41.4% (12/29) and 100.0% (10/10) respectively. All lymphoma samples which expressed CD10 also showed co-expression of bcl-6 protein. (2) The co-expression of bcl-6 and CD10 was observed in 40.9% (9/22) nodal DLBCL and 41.4% (12/29) extranodal DLBCL. Low clinical stage (stage I and II) was more frequently observed in cases with co-expression of bcl-6 and CD10 (P < 0.05). (3) The rates of bcl-6 gene rearrangement in nodal DLBCL was 28.1% (9/32), with 27.3% (6/22) in paraffin-embedded tissues and 30.0% (3/10) in fresh tissues. There was no statistically significant difference found between the two groups (P > 0.05). Bcl-6 gene rearrangement was not found in all the 5 cases of RLH, and there was a significant difference between RLH and DLBCL (P < 0.05).

CONCLUSIONSThe rate of bcl-6 protein expression is high in DLBCL cases, and the detection of bcl-6 and CD10 protein co-expression may help in the diagnosis and differential diagnosis of DLBCL. Those DLBCL cases with co-expression of bcl-6 and CD10 may also have a better prognostic implication. On the other hand, bcl-6 gene rearrangement can be identified by interphase FISH with dual color breakapart probe in both paraffin-embedded and fresh lymphoma tissues.

Diagnosis, Differential ; Female ; Gene Rearrangement ; Humans ; In Situ Hybridization, Fluorescence ; Lymphoma, B-Cell ; genetics ; metabolism ; pathology ; Lymphoma, Large B-Cell, Diffuse ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; Neoplasm Staging ; Neprilysin ; metabolism ; Proto-Oncogene Proteins c-bcl-6 ; genetics ; metabolism ; Pseudolymphoma ; genetics

OBJECTIVETo evaluate the efficiency of the BIOMED-2 PCR assay and its implication in the diagnosis of mature B-cell non-Hodgkin's lymphomas.

METHODSClinical, morphological and immunohistochemical features of 72 cases of non-Hodgkin's lymphomas were studied, including 25 reactive lymphoid hyperplasia, 37 diffuse large B cell lymphomas (DLBCL) and 35 extranodal marginal zone lymphomas of mucosa associated lymphoid tissues (MALT lymphoma and in addition, 25 cases of reactive lymphoid hyperplasia were used as the controls). DNA was exacted from the paraffin embedded formalin fixed tissue blocks and the quality of DNA was assessed using the BIOMED-2 specimen control reaction. Adequate samples were then analyzed by BIOMED-2 for immunoglobulin heavy and kappa light chain rearrangements.

RESULTSAdequate DNA was obtained in 83 of 97 samples, including 60 mature B cell lymphomas and 23 reactive lymphoid hyperplasia. Clonal B-cell gene rearrangements were detected in 57 of 60 (95%) lymphomas. In contrast, clonal Ig gene rearrangements were not detected in any of the 23 cases of reactive lymphoid hyperplasia.

CONCLUSIONBIOMED-2 assay is highly sensitive and specific for the detection of clonal B cell gene rearrangement using routine paraffin embedded formalin fixed specimens.

Antigens, CD20 ; metabolism ; CD79 Antigens ; metabolism ; DNA, Neoplasm ; genetics ; Gene Rearrangement, B-Lymphocyte ; genetics ; Gene Rearrangement, B-Lymphocyte, Heavy Chain ; genetics ; Gene Rearrangement, B-Lymphocyte, Light Chain ; genetics ; Genes, Immunoglobulin ; Humans ; Immunophenotyping ; Lymphoma, B-Cell ; genetics ; immunology ; pathology ; Lymphoma, B-Cell, Marginal Zone ; genetics ; immunology ; pathology ; Lymphoma, Large B-Cell, Diffuse ; genetics ; immunology ; pathology ; Paraffin Embedding ; Pseudolymphoma ; genetics ; immunology ; pathology ; Sensitivity and Specificity

OBJECTIVETo study the expression profile of microRNAs (miRNAs) in peripheral T cell lymphoma, not otherwise specified (PTCL-NOS) and to explore the underlying molecular characteristics.

METHODSTwenty-one cases of PTCL-NOS were enrolled into the study. The tumor presented either as nodal (15/21) or extranodal (6/21) disease. TaqMan low density array was used to assess the expression level of 754 miRNAs in six cases of PTCL-NOS and three control cases of reactive lymphoid hyperplasia. Prediction of target genes for significant and differential expression of miRNAs was carried out using Targetscan and miRanda software. Bioinformatics tools were employed for GO-Analysis and Pathway-Analysis of target genes. The expression patterns of the three miRNAs were further analyzed in the remaining 15 cases of PTCL-NOS and 10 cases of reactive lymphoid hyperplasia, using single tube Taqman miRNA assays.

RESULTSEight miRNAs showed statistically significant difference in expression between PTCL-NOS and reactive lymphoid hyperplasia. miR-886-3p, miR-511, miR-1291, miR-572, miR-27a-3p, miR-25-3p and miR-886-5p were significantly overexpressed in PTCL-NOS while miR-182-5p was significantly underexpressed (P < 0.05). Target gene prediction showed that 1646 candidate genes involved in the pathogenesis and progression of PTCL-NOS. Further GO and Pathway-Analyses found that these genes significantly focused on 63 GO terms and 61 pathways. The results of three miRNA qRT-PCR confirmed that miR-572 and miR-1291 expression in PTCL-NOS had statistical significance.

CONCLUSIONSEight miRNAs are aberrantly expressed in PTCL-NOS, which may be involved in molecular regulation during the development of PTCL-NOS. The underlying mechanism is also related to a number of target genes and signaling pathways.

Adult ; Aged ; Aged, 80 and over ; Female ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; Humans ; Lymphoma, T-Cell, Peripheral ; genetics ; metabolism ; pathology ; Male ; MicroRNAs ; metabolism ; Middle Aged ; Pseudolymphoma ; genetics ; metabolism ; Young Adult

OBJECTIVETo evaluate the ancillary diagnostic value of IgH gene rearrangements in those B-cell lymphoproliferative disorder cases whom are difficult in making a final diagnosis.

METHODSIgH gene clonal rearrangements were retrospectively analyzed in a total of 77 diagnostically difficult B-cell lympho-proliferative patients. Standardized BIOMED-2 system IgH gene clonality assay kit targeting FR1, FR2, FR3 was used, followed by heteroduplex-polyacrylamide gel electrophoresis (PAGE) and silver nitrate staining.

RESULTSThe final diagnoses of the 77 cases were: 12 cases of reactive lymphoid hyperplasia, 20 cases of atypical lymphoid hyperplasia or suspicious lymphoma, and 45 cases of B-cell lymphoma. Detection rates of at least one positive reaction were 2/12, 11/20 (55%), 36/45 (80%) in the three groups, respectively. In B-cell lymphomas, the clonality detection rate of FR1, FR2 and FR3 was 60% (27/45), 60% (27/45) and 56% (25/45), respectively. The type distribution were: 20 marginal zone lymphomas, including 18 extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue, 7 diffuse large B-cell lymphomas, 7 follicular lymphomas, 1 mantle-cell lymphoma, 1 Burkitt's lymphoma, 4 plasma cell neoplasms and 5 unclassified B-cell lymphomas. Rearrangements of FR1, FR2 or FR3 were not detected in 9 (20%) of the B cell lymphoma cases, nevertheless, one of them had developed liver lesion later, and was confirmed finally to be B cell lymphoma. Fourteen patients of reactive lymphoid hyperplasia with positive IgH gene clonal rearrangements, and atypical lymphoid hyperplasia had follow-up history available. Four of them were diagnosed as lymphoid malignancies upon further biopsy, and in three of them, clonal IgH gene rearrangements were detected.

CONCLUSIONSB-cell lymphoproliferative disorder requiring a detection of clonal IgH gene rearrangement for making a final diagnosis. Combined detections of three IgH FR1, FR2 and FR3 rearrangements provide important ancillary diagnostic value in confirming suspected B-cell lympho-proliferative disorders. It is important to take an additional biopsy or to follow-up those patients who that have a detectable IgH gene clonal rearrangement but without apparent morphological evidence of lymphoma. For cases with a negative IgH gene rearrangements, it might be necessary to perform clonality analysis for other forms of gene rearrangements including IgH or IgK and IgL in order to further improve the detection sensitivity.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child ; Female ; Follow-Up Studies ; Gene Rearrangement, B-Lymphocyte, Heavy Chain ; Humans ; Lymphoma, B-Cell ; diagnosis ; genetics ; pathology ; Lymphoma, B-Cell, Marginal Zone ; diagnosis ; genetics ; pathology ; Lymphoma, Follicular ; diagnosis ; genetics ; pathology ; Lymphoma, Large B-Cell, Diffuse ; diagnosis ; genetics ; pathology ; Lymphoproliferative Disorders ; diagnosis ; genetics ; pathology ; Male ; Middle Aged ; Neoplasms, Plasma Cell ; diagnosis ; genetics ; pathology ; Pseudolymphoma ; diagnosis ; genetics ; pathology ; Retrospective Studies ; Young Adult

CD5 Antigens ; metabolism ; Chromosomes, Human, Pair 11 ; Chromosomes, Human, Pair 14 ; Chromosomes, Human, Pair 18 ; Cyclin D1 ; metabolism ; Diagnosis, Differential ; Humans ; Ki-67 Antigen ; metabolism ; Lymphoma, B-Cell, Marginal Zone ; genetics ; metabolism ; pathology ; Lymphoma, Follicular ; genetics ; metabolism ; pathology ; radiotherapy ; Lymphoma, Mantle-Cell ; genetics ; metabolism ; pathology ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Pseudolymphoma ; metabolism ; pathology ; Translocation, Genetic

Aged ; CD5 Antigens ; metabolism ; Chromosomes, Human, Pair 11 ; Chromosomes, Human, Pair 14 ; Cyclin D1 ; metabolism ; Diagnosis, Differential ; Female ; Humans ; Liver Neoplasms ; genetics ; metabolism ; pathology ; surgery ; Lymphoma, B-Cell ; metabolism ; pathology ; Lymphoma, Follicular ; metabolism ; pathology ; Lymphoma, Mantle-Cell ; genetics ; metabolism ; pathology ; surgery ; Pseudolymphoma ; metabolism ; pathology ; Translocation, Genetic

OBJECTIVETo study the clinicopathologic features of primary thymic extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT).

METHODSThe clinical and pathologic findings were evaluated in 3 cases of biopsy confirmed thymic MALT lymphoma. The clincopathologic features, treatment and prognosis were discussed and literatures reviewed.

RESULTSOne male and two female patients presented with asymptomatic mediastinal masses with a history of Sjögren syndrome. They were aged 36, 35 and 41 years respectively, and only one patient had B symptoms. Grossly, all three tumors were encapsulated and had multiple variable-sized cysts on cut-surface. Histopathologically, the normal thymic lobular architecture was effaced by abnormal dense lymphoid infiltration. Prominent lymphoepithelial lesions were formed by centrocyte-like cells infiltrating and expanding Hassall's corpuscles and epithelial cyst lining. All cases showed apparent plasmacytic differentiation. Immunohistochemically, the tumor cells were positive for CD20, CD79a, bcl-2 and negative for CD3, CD5, cyclin D1, CD43, CD10, bcl-6, and CD23. The plasma cells showed kappa light chain restriction. Immunoglobulin heavy chain rearrangement in three cases was confirmed by PCR. All patients were at early stage and received routine chemotherapy with or without radiotherapy after surgical removal. All patients achieved complete remission with 24, 18 and 3 months follow-up, respectively.

CONCLUSIONSPrimary thymic MALT lymphoma may be a rare distinctive lymphoma. It can be diagnosed by HE and immunohistochemical study and should be differentiated from reactive lymphoid proliferation, other types of lymphoma and mediastinal thymoma.

Adult ; Antibodies, Monoclonal, Murine-Derived ; therapeutic use ; Antigens, CD20 ; metabolism ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Cyclophosphamide ; therapeutic use ; Diagnosis, Differential ; Doxorubicin ; therapeutic use ; Female ; Follow-Up Studies ; Gene Rearrangement, B-Lymphocyte, Heavy Chain ; Humans ; Immunoglobulin Heavy Chains ; genetics ; Keratin-19 ; metabolism ; Lymphoma, B-Cell, Marginal Zone ; drug therapy ; genetics ; metabolism ; pathology ; surgery ; Male ; Prednisone ; therapeutic use ; Pseudolymphoma ; pathology ; Thymus Hyperplasia ; pathology ; Thymus Neoplasms ; drug therapy ; genetics ; metabolism ; pathology ; surgery ; Vincristine ; therapeutic use