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Introducción. El consorcio europeo BIOMED-2 se creó para determinar si una población linfoide de difícil clasificación patológica es clonal. En Colombia, la implementación de estas pruebas comenzó en el 2015 en el Instituto Nacional de Cancerología E.S.E. (Bogotá). Objetivos. Determinar el comportamiento de las pruebas de reordenamiento clonal o clonalidad linfoide. y determinar las dificultades de su uso en nuestro medio verificando su adaptación local y los resultados en una serie retrospectiva de casos y consecutiva de proliferaciones linfoides sometidas a los protocolos BIOMED-2. Materiales y métodos. A partir de las historias clínicas, se recolectaron los datos clínicos e histológicos y los resultados de los análisis de los reordenamientos en todos los casos de proliferaciones linfoides sometidas a los protocolos BIOMED-2, entre febrero de 2015 y mayo de 2019. Resultados. Se hallaron 132 casos, de los cuales 47 se clasificaron mediante los protocolos de Biomed-2 como hiperplasias linfoides reactivas, 62 como linfomas T, 19 como linfomas B y 3 como neoplasias linfoides de linaje no establecido. Solo en un caso falló la extracción de ADN. Según estos resultados, la mayor dificultad diagnóstica para el patólogo fue el análisis de los infiltrados linfoides T, la mayoría (44) de los cuales correspondía a lesiones cutáneas. Conclusiones. Las pruebas de clonalidad pueden usarse en tejidos de diversa calidad en nuestro medio como ayuda en el diagnóstico de proliferaciones linfoides de difícil clasificación. Es importante hacerlas e interpretarlas de manera multidisciplinaria y considerar cada caso por separado.
Introduction: The European BIOMED-2 consortium was created to evaluate clonality in lymphoproliferations that are difficult to diagnose. In Colombia, the implementation of these tests began in 2015 at the Instituto Nacional de Cancerología E.S.E., Bogotá. Objectives: To determine the behavior of the rearrangement tests for lymphoid clonality and the difficulties of its implementation in our field through a series of retrospective and consecutive cases of lymphoid proliferation subjected to the BIOMED-2 protocols. Materials and methods: Clinical and histological data and the results of the rearrangement analysis of all cases of lymphoid proliferation subjected to the BIOMED-2 protocols between February 2015 and May 2019 were collected from clinical histories. Results: We recovered 132 samples from which 47 corresponded to reactive lymphoid hyperplasias, 62 to T lymphomas, 19 to B lymphomas, and three to lymphoid neoplasms of unestablished lineage. Only in one case did DNA extraction fail. According to these results, the greatest diagnostic difficulty for the pathologist was the analysis of T lymphoid infiltrates, most of which (44) were skin lesions. Conclusions: Clonality tests can be used in tissues of different quality to help in the diagnosis of lymphoid proliferations that are difficult to classify. It is important to implement and interpret them in an interdisciplinary way considering each case separately.
Subject(s)
Lymphoma , Immunoglobulins , Gene Rearrangement, T-Lymphocyte , Genes, T-Cell Receptor , Electrophoresis, Polyacrylamide GelABSTRACT
Objective To assess the preferential expressions of peripheral blood T cell receptor beta chain variable region (TRBV) subfamilies in patients with psoriasis vulgaris(PV), and to estimate their role in the pathogenesis of psoriasis. Methods Thirty-three upstream primers were designed to target the human functional TRBV genes, downstream primers to target the common T cell receptor beta constant (TRBC) gene,with T cell receptor alpha constant (TRAC) gene as the internal reference. Total RNA was extracted from the peripheral blood T cells of 10 health human controls and 10 patients with PV, and transcribed into cDNA.Then, TRBV genes were amplified by real-time fluorescence quantitative PCR (RFQ-PCR) and the fluorescence intensity of each samples was detected. The expression levels of TRBV genes in the control group were used to calculate the cut-off values (mean expression levels of TRBV subfamilies in the 10 normal controls + 3 standard deviations). When the expression level of a TRBV subfamily from patients with PV was equal to or higher than the cut-off value, it was considered as the preferentially expressed TRBV subfamily. Results The threshold cycle (Ct) value varied from 21 to 24 for TRAC gene. The difference in the Ct value between TRBV subfamily genes and TRAC gene in patients with PV was 2.98 for TRBV2 gene, 3.24 for TRBV5-7 gene, 2.52 for TRBV6-6/6-9 gene, 2.04 for TRBV 12 gene, 3.56 for TRBV 24 gene, and 4.12 for TRBV 29 gene, and the expression levels of these subfamily genes were significantly higher than those in the normal controls (all P ＜ 0.05). According to the above standard, TRBV6-6/6-9, TRBV12 and TRBV29 were considered to be preferentially expressed subfamilies. Conclusions There is a preferential expression of TRBV gene subfamilies in peripheral blood of patients with psoriasis vulgaris, which may play a vital role in the abnormal T cell-mediated immune responses in psoriasis.
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Objective To investigate T cell receptor(TCR)variable β(BV)chain usage at the maternal-fetal interface and explore the relationship between the skewed TCR BV usage and unexplained recurrent spontaneous abortion(BSA).Methods Eighteen cases with unexplained RSA,together with matched 41 women with normal pregnancies in first trimester from Renji Hospital,Shanghai Jiao Tong University were studied.A high-resolution spectrum typing analysis of complementarity-determining region 3 (CDR3)was used to detect and compare the degree and frequency of TCR BV family expression in deciduas between RSA patients and normal controls.Results(1)The expression degree of BV19(0.029±0.031 vs.0.013±0.010,P=0.038)in RSA group showed a higher usage,while BV5.2(0.040±0.035 vs.0.067±0.052,P=0.046)showed a significantly lower usage when compared with normal controls.No significant difference in the expression of the other TCR BV families between RSA and controls were observed(P>0.05).(2),TCR BV2,3,6,and 7 were the four most common BV families in deciduas of patients with RSA and normal controls,whose frequencies were all mors than 50%.In RSA group,higher frequencies of BV15 (33.3%vs.7.3%,P=0.018),BV19(38.9%vs.14.6%,P=0.049)and BV20(33.3%vs.7.3%,P=0.018)were observed;meanuhile lower frequencies of BV4(33.3%vs.65.9%,P=0.026)and BV7 (66.7%vs.92.7%.P=0.018)distributions were observed.The other TCR BV families did not display significantly different freqencies of distribution(P>0.05).Conclusions It is suggested that a significant skewed TCR BV family occurs at the maternal-fetal interface in patients who undergo abortion.The specific skewed usages of TCR BV might be associated with the susceptibility to unexplained pregnancy loss.
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Objective To investigate the expression and clonality of TCR V? subfamily T cell in 11 patients undergoing chronic hemodialysis before and after renal transplantation.Methods The CDR3 of TCR V? 24 subfamily genes were amplified in samples of peripheral blood mononuclear cells which were drawn before hemodialysis and at 20th day post-operatively. To observe the usage of TCR V? repertoire, the RT-PCR products were further labeled with fluorescent and analyzed by genescan technique for CDR3 size.Results (1) One patient was attacked by acute cellular rejection (ACR) and one suffered from delayed graft function (DGF) diagnosed by renal transplant biopsy. (2) 1-10 TCR V? subfamilies T cells could be identified before transplantation, and most of TCR V? subfamily T cells expressed as polyclonality. The most frequent expression of TCR V? genes was V?3. (3) Only 1-5 TCR V? subfamilies T cells could be detected post-operatively, most of them expressed as oligoclonality. The TCR V?3 subfamilies still were the most frequently expressed (in 9 cases). (4) There were no TCR V? subfamilies T cells before pulse of methylprednisolone, and were 5 subfamilies during pulse of methylprednisolone expressed as oligoclonality or biclonality, 2 subfamilies after ACR expressed as polyclonality. In DGF patient, there were 4 TCR V? subfamilies during DGF, and 5 following DGF.Conclusion (1) The significantly skew distribution of TCR V? subfamily T cells could be found in all patients with chronic renal failure and chronic hemodialysis. (2) Furthermore, the skewing distributions of TCR V? genes came to more significant at 20th day post-transplantation than before. (3) ACR might be closely associated with some special clones of TCR V? subfamily T cells, which subsided immediately after ACR. (4) The expression as polyclonality after ACR and DGF may indicate that the immune function partially was inclined to normalization even though immunosuppressed.
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Objective To study the early diagnosis of cutaneous T-cell lymphoma(CTCL).Methods DNA from39formalin-fixed,paraffin-embedded tissues of24cases of CTCL,2cases of dubious lymphoma,and4cases with non-specific erythroderma,was amplified by polymerase chain reaction(PCR),with special primers of TCR V?1-8,V?9,and J?1/?2to analyze T-Cell receptor?gene clonal rearrangement(TCR-GR clonal).The specimens from30patients with non-specific dermatitis were taken as negative control.Results Clonal of TCR V?1-8was demonstrated in38of39paraffin-embedded specimens.V?9-GR was shown in37of39specimens TCR?-GR was shown especially in13patients with MF of early stage,2patients with dubious CTCL,and4patients with non-specific erythroderma,as well as in6of30patients with histiologically nonspecific dermatitis which suggested that these6cases were"clonal dermatitis".Conclusion Clonal TCR?-GR may be detected in patients with early CTCL,even when the histologic findings are not unequivocally diagnostic.