OBJECTIVETo investigate the significance of detecting TCR gene clonal rearrangement in the diagnosis of mycosis fungoides (MF) and to optimize the primers used for detecting the TCR gene clonal rearrangement with PCR in paraffin embedded tissues of MF.

METHODSNineteen cases of MF were enrolled into the study. A panel of 10 antibodies were used for immunophenotypic analysis and polymerase chain reaction for TCR-γ and TCR-β gene rearrangement detection in this study.

RESULTSTCR gene clonal rearrangements were detected in all 19 cases, in which 84.2% cases (16/19) had TCR-&gamma; gene clonal rearrangements. The positive rates of the primers T(VG)/T(JX), V(2-5)/V(8-12)/JGT(1) and BIOMED-2-TCR-&gamma; were 47.4%, 78.9% and 31.6%, respectively. The positive rate of V(2-5)/V(8-12)/JGT(1) was statistically significantly higher than that of T(VG)/T(JX) and BIOMED-2-TCR-&gamma; (P < 0.05). No TCR gene clonal rearrangement was detected using the primers V(&gamma;11)/V(&gamma;101)/J&gamma;12 and V(&gamma;11)/V(&gamma;101)/J(p12). TCR-β gene clonal rearrangement was detected in 31.6% (6/19) cases.

CONCLUSIONSTCR gene clonal rearrangement analysis is a useful tool in the diagnosis of MF and TCR-&gamma; gene is a good target gene for the detection. The primers T(VG)/T(JX), V(2-5)/V(8-12)/JGT(1) and BIOMED-2-TCR-&gamma; can be used in clinicopathologic detection for TCR gene clonal rearrangement and V(2-5)/V(8-12)/JGT(1) may be the first choice.

Adolescent ; Adult ; Aged ; Antigens, CD7 ; metabolism ; Base Sequence ; CD2 Antigens ; metabolism ; CD3 Complex ; metabolism ; CD4 Antigens ; metabolism ; Child ; Child, Preschool ; Female ; Gene Rearrangement, beta-Chain T-Cell Antigen Receptor ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Humans ; Leukocyte Common Antigens ; metabolism ; Male ; Middle Aged ; Molecular Sequence Data ; Mycosis Fungoides ; diagnosis ; genetics ; metabolism ; pathology ; Paraffin Embedding ; Receptors, Antigen, T-Cell, alpha-beta ; genetics ; Receptors, Antigen, T-Cell, gamma-delta ; genetics ; Skin Neoplasms ; diagnosis ; genetics ; metabolism ; pathology ; Young Adult

OBJECTIVETo evaluate the practical values of PCR detectable T-cell receptor (TCR) gene rearrangement in paraffin embedded tissue samples in the diagnosis of T-cell malignancies using BIOMED-2 PCR multiplex tubes TCRgamma(A+B).

METHODSTraditional phenol-chloroform method was used to extract DNA from 55 cases of archival paraffin embedded tissues samples of T-cell malignancies and the DNA quality was evaluated by PCR-based amplification of housekeeping gene beta-globin. The selected BIOMED-2 PCR multiplex tubes TCRgamma(A+B) were used to detect TCR gene rearrangement and comparison with the results of universal TCR primers (T(VG)/T(JX)) was performed.

RESULTSPositive detection rates by the BIOMED-2 multiplex tubes TCRgamma(A+B) and the universal primers (T(VG)/T(JX)) were 76.4% and 60.0%, respectively. There were not statistical difference between the methods (P > 0.05).

CONCLUSIONBIOMED-2 multiplex tubes TCRgamma(A+B) is suitable for detection of clonal rearrangements of TCR genes in current archival paraffin embedded tissue samples of T-cell malignancies.

DNA Primers ; DNA, Neoplasm ; analysis ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Humans ; Lymphoma, T-Cell ; genetics ; metabolism ; pathology ; Paraffin Embedding ; Polymerase Chain Reaction ; methods ; beta-Globins ; metabolism

INTRODUCTIONPolymerase chain reaction (PCR)-based molecular techniques are useful adjunctive tools in the diagnosis of cutaneous T-cell lymphomas (CTCL). This study compares the sensitivity of PCR analysis of the T-cell receptor-gamma (TCR-gamma) gene rearrangements using conventional polyacrylamide gel electrophoresis (PCR-PAGE) and fluorescent capillary electrophoresis (PCR-FCE).

MATERIALS AND METHODSA total of 22 paraffin blocks were analysed using PCR-PAGE and PCR-FCE. There were 17 cases of mycosis fungoides (MF), 4 cases of non-MF CTCL and 1 case of lymphoblastic leukaemia.

RESULTSComplete agreement was obtained between PCR-PAGE and PCR-FCE in 19 of the 22 cases, giving a concordance rate of 86.4%. PCR-FCE had a higher sensitivity of 77.3%, compared to 63.6% for PCR-PAGE, allowing the detection of 3 additional cases of clonal T-cell rearrangements, which had equivocal or polyclonal bands on PAGE. Two of these 3 cases were in erythrodermic MF patients. PCR-FCE also allowed the detection of matching clones in serial specimens taken from different sites and at different time intervals in patients with MF. However, matching clones from different specimens can be achieved qualitatively in PCR-PAGE by running and comparing these on the same polyacrylamide gel block.

CONCLUSIONSBoth PCR-PAGE and PCR-FCE are useful in detecting T-cell clones in CTCL, with both methods being comparable in sensitivity and showing a high concordance rate of 86.4%. PCR-FCE has the added advantage of exhibiting semiquantitative properties, which may be important in early or erythrodermic MF cases, but the requirement for sophisticated and costly machinery limits its availability to high-capacity laboratories. The well-established PCR-PAGE method is a suitable alternative in routine clinical applications.

Base Sequence ; Electrophoresis, Agar Gel ; Electrophoresis, Capillary ; methods ; Electrophoresis, Polyacrylamide Gel ; Fluorescence ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Humans ; Lymphoma, T-Cell ; diagnosis ; Mycosis Fungoides ; Polymerase Chain Reaction ; methods ; Sensitivity and Specificity

Mycosis fungoides (MF), the most common type of cutaneous T-cell lymphoma, has various unspecific clinical and histological characteristics. Its early diagnosis is challenging. The application of T-cell receptor (TCR) gene clonal rearrangement to the diagnosis of MF has been widely studied. In this study, we used polymerase chain reaction (PCR) to investigate the diagnostic significance of detecting TCR-&gamma; and -β gene clonal rearrangement in the early diagnosis of mycosis fungoides. PCR for TCR-&gamma; and TCR-β gene rearrangement was performed on 19 patients with suspected early MF, 6 with typical MF, and 6 with chronic dermatitis. Of the 19 patients with suspected early MF, 13 had TCR-&gamma; gene clonal rearrangement, whereas none had TCR-β gene clonal rearrangement. All patients with typical MF had TCR gene clonal rearrangement, in which 4 showed TCR-&gamma; clonal rearrangement, 1 showed TCR-β gene clonal rearrangements, and 1 showed both. No patients with chronic dermatitis had TCR gene clonal rearrangement. These results indicate that TCR gene clonal rearrangement analysis is a useful tool in diagnosing early MF. TCR-&gamma; gene is recommended to the routine analysis, whereas TCR-β gene has potential in combination toward intractable cases.
Adolescent ; Adult ; Aged ; Base Sequence ; genetics ; DNA, Neoplasm ; genetics ; Early Detection of Cancer ; methods ; Female ; Gene Rearrangement, beta-Chain T-Cell Antigen Receptor ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Humans ; Male ; Middle Aged ; Mycosis Fungoides ; diagnosis ; genetics ; Polymerase Chain Reaction ; Skin Neoplasms ; diagnosis ; genetics ; Young Adult

OBJECTIVETo investigate the lineage of the malignant cells in malignant histiocytosis.

METHODSPolymerase chain reaction with two groups of common primers for TCR-gamma gene was used to analyze the malignant cells of 28 autopsied cases of malignant histiocytosis.

RESULTSMonoclonal TCR-gamma gene rearrangements were detected in 12 out of the 28 samples (43%).

CONCLUSIONMost cases diagnosed as malignant histiocytosis in Southwest China seems to be peripheral T-cell lymphomas.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; CD56 Antigen ; analysis ; Child ; Child, Preschool ; Female ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Histiocytosis ; genetics ; immunology ; Humans ; Infant ; Male ; Middle Aged ; Polymerase Chain Reaction

CD3 Complex ; metabolism ; CD5 Antigens ; metabolism ; Diagnosis, Differential ; Female ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Humans ; Leukocyte Common Antigens ; metabolism ; Leukosialin ; metabolism ; Lymphoma, T-Cell, Cutaneous ; metabolism ; pathology ; Middle Aged ; Skin Diseases ; pathology

OBJECTIVETo investigate the roles of different clinicopathological features and expression of EBV genome in prognosis of intestinal T-cell lymphoma (ITCL).

METHODSPolymerase chain reaction for TCR-gamma gene rearrangement, in situ hybridization for EBER1/2 and immunohistochemical staining for CD4, CD8, CD45RO, CD56, TIA-1 were investigated and all patients followed-up. The LMP-1 expression was determined in forty-two ITCLs cases. The relationship between clinical data, different clinicopathological features, expression of EBV genome and prognosis were analyzed by SPSS10.0 program.

RESULTS(1) All 42 cases of ITCL had an extremely poor prognosis with a median survival of 3.0 months, of which the one year survival rate and two year survival rate being 30% and 22% respectively. (2) The patients without TCR-gamma gene rearrangements showed poorer prognosis than those with TCR-gamma gene rearrangements, and the patients who received operation and chemotherapy showed better prognosis than those who only received operation (P < 0.05). (3) No significant prognostic factor for ITCLs was determined.

CONCLUSIONThe special clinicopathological features of ITCL could be due to the cytotoxic function and the role of EBV infection in the pathogenesis of ITCL.

Adolescent ; Adult ; Child ; Female ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Humans ; Intestinal Neoplasms ; mortality ; pathology ; therapy ; Lymphoma, T-Cell ; mortality ; pathology ; therapy ; Male ; Middle Aged ; Prognosis ; Survival Rate

OBJECTIVETo study the NK/T-cell lymphoma, search for a more efficacious and simpler method and establish a standard guideline for distinguishing the NK-like T-cell lymphoma from the NK-cell lymphoma.

METHODSThirty-four NK or T-cell lymphomas from the upper aerodigestive tract (n = 22), skin (n = 2), gastrointestinal (GI) tract (n = 2), lymph nodes (n = 7), and other sites (n = 1) were studied. Immunophenotype was analyzed by immunohistochemistry. In situ hybridization with EBER 1/2 RNA probes was performed. T-cell receptor (TCR)-beta and -gamma gene rearrangement was analyzed by polymerase chain reaction (PCR).

RESULTSEighteen cases were positive for CD(56) and 16 for TIA-1 in 34 lymphomas cases. All tumor cells in the skin cases were positive for Ki-67. Epstein-Barr virus (EBV) mRNA was detected in 12 upper aerodigestive tumors including 9 of 12 nasal and 3 extranasal tumors. EBER was also detected in 1 of 2 skin lymphomas and both of the 2 GI lymphomas. Clonal TCR-beta and -gamma gene rearrangement was detected in 2 of 22 upper aerodigestive, all of the skin and GI lymphomas, and 6 of 9 nodal and other site lymphomas.

CONCLUSIONMost upper aerodigestive NK/T-cell lymphomas are genotypically NK derivation, and a few belong to T lineage. However, NK-like T-cell lymphomas more frequently seen in skin and GI tract. Nodal NK-cell lymphoma are quite rare. These two kinds of lymphomas can only be diagnosed with additional immunohistochemical markers, EBER detection by ISH, TCR gene rearrangement or NK-cell receptors (NKRs) RNA detection. Detection of TCR rearrangement remains the important standard for the diagnosis of T-cell lymphoma.

Adolescent ; Adult ; Aged ; Child ; Female ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Herpesvirus 4, Human ; isolation & purification ; Humans ; Immunophenotyping ; Killer Cells, Natural ; pathology ; Lymphoma, T-Cell ; genetics ; immunology ; pathology ; Male ; Middle Aged

OBJECTIVETo improve the techniques for minimal residual disease (MRD) detection in acute lymphoblastic leukemia (ALL).

METHODSA real time quantitative PCR method was established for quantifying the clonal TCRVgammaI-Jgamma gene rearrangement in 36 ALL patients.

RESULTSThe sensitivity of the established real time quantitative PCR was at 10(-4) level. The amount of TCRVgammaI-Jgamma gene rearrangement in newly diagnosed group, complete remission (CR) group and post hematopoietic stem cell transplantation (HSCT) group was (7.38 +/- 6.65) x 10(-2), (1.02 +/- 1.08) x 10(-2) and (3.89 +/- 5.65) x 10(-3) level, respectively. and the amount in newly diagnosed group was higher than that in CR group and HSCT group (P = 0.001). The MRD level of ALL patients in CR group was higher than that in HSCT group (P = 0.022). MRD can be detected in 6 ALL patients after HSCT, 2 of them with low MRD level (< 1 x 10(-3)) survived long disease-free survival, the other 4 with high MRD level relapsed within one year.

CONCLUSIONThe established real time quantitative PCR assay is simple, rapid, sensitive and specific. Use of this assay to evaluate MRD in the remission ALL cases is helpful for prognosis prediction.

Adolescent ; Adult ; Female ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; genetics ; Humans ; Immunoglobulin Variable Region ; genetics ; Male ; Polymerase Chain Reaction ; methods ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; pathology ; therapy ; Prognosis ; Reproducibility of Results

OBJECTIVETo evaluate the clinical significance of IgH and TCR gamma gene rearrangement in plasma free DNA in patients with non-Hodgkin Lymphoma (NHL).

METHODSPlasma free DNA in 74 patients with NHL were extracted and identified by Globin gene. IgH (FR3A/VLJH), TCR gamma (TVG/TJX) clonal rearrangements were amplified by PCR and compared with results of mononuclear cell DNA and pathological biopsy sample DNA.

RESULTSPlasma free DNAs were successfully obtained from 58 cases (35 B-NHL and 23 T-NHL) of newly diagnostic, refractory and relapsed NHL out of total 74 patients (78.4%), but not found in the rest 16 patients in remission. Of 35 B-NHL cases, 31 showed IgH rearrangement (88.6%), and none with TCR gamma rearrangement; of 23 T-NHL cases, 8 showed TCR gamma rearrangement (34.8%), and 2 with IgH gene rearrangement synchronously. In comparison with the results of IgH and TCR gamma gene rearrangement in biopsy samples in 30 B-NHL cases, 26 cases in plasma free DNA (86.7%) and 24 in biopsy samples (80%) were positive (P > 0.05). In 20 T-NHL patients, 7 cases in plasma cell-free DNA (35%) and 6 cases in biopsy samples (30%) were positive (P >0.05).

CONCLUSIONSTumor-derived DNA could be detected in plasma from underlying cancer patients. For NHL patients, detecting IgH and TCR gamma gene rearrangement in plasma free DNA has the same clinical significance as in biopsy samples.

Adolescent ; Adult ; Aged ; Child ; DNA ; blood ; Female ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Humans ; Immunoglobulin Heavy Chains ; genetics ; Lymphoma, Non-Hodgkin ; blood ; genetics ; Male ; Middle Aged ; Young Adult