RESUMEN
OBJECTIVE: The aim of this study is to describe and review the cytological features of Kaposi sarcoma-associated herpes virus (KSHV) related entities, such as multicentric Castleman's disease (MCD), plasmablastic-lymphoma (PBL) and primary effusion lymphoma (PEL), which all may present as body cavity effusions. Serous fluid cytology of MCD and PBL has not, to our knowledge, thus far been described. Although different in nature, MCD, PBL and PEL are characterized by similar morphological features. MATERIALS AND METHODS: Body cavity effusions from four different patients with previously known or unknown KSHV-related lymphoproliferations have been examined by routine cytology, immunocytochemistry (IC) and polymerase chain reaction (PCR). RESULTS: MCD, PBL and PEL are all characterized by increased cellularity, comprising mainly lymphoid and plasmacytoid cells with variable proportions of immunoblasts. Immunocytochemistry and PCR results show the MCD to be CD138 and KSHV positive, CD30 negative, IgM, IgH and lambda restricted but IgH polyclonal. PBL was CD138 positive, kappa restricted, weakly positive with VS38 and over 80% positive with MIB 1. PEL was CD45, EMA, CD138, KSHV, p53 and CD3 positive, CD20, EBV, CD30, CD2, CD4, ALK1, epithelial and mesothelial markers negative, and PCR monoclonal B-cell expanded (Ig-kappa bands). CONCLUSION: Cytological examination of effusions in KSHV-related lymphoproliferative disorders may show similar morphological features but clonality studies and immunocytochemistry are very helpful in distinguishing between these rare benign and malignant lymphoproliferative diseases.
Asunto(s)
Líquidos Corporales/citología , Líquidos Corporales/virología , Enfermedad de Castleman/complicaciones , Enfermedad de Castleman/virología , Herpesvirus Humano 8/fisiología , Trastornos Linfoproliferativos/complicaciones , Trastornos Linfoproliferativos/virología , Humanos , Inmunohistoquímica , Linfoma de Efusión Primaria/complicaciones , Linfoma de Efusión Primaria/virología , Masculino , Persona de Mediana Edad , Derrame Pleural/complicaciones , Derrame Pleural/virologíaRESUMEN
Chordomas are radio- and chemo-resistant tumours and metastasise in as many as 40% of patients. The aim of this study was to identify potential molecular targets for the treatment of chordoma. In view of the reported association of chordoma and tuberous sclerosis complex syndrome, and the available therapeutic agents against molecules in the PI3K/AKT/TSC1/TSC2/mTOR pathway, a tissue microarray of 50 chordoma cases was analysed for expression of active molecules involved in this signalling pathway by immunohistochemistry and a selected number by western blot analysis. Chordomas were positive for p-AKT (92%), p-TSC2 (96%), p-mTOR (27%), total mTOR (75%), p-p70S6K (62%), p-RPS6 (22%), p-4E-BP1 (96%) and eIF-4E (98%). Phosphatase and tensin homologue deleted on chromosome 10 expression was lost in 16% of cases. Mutations failed to be identified in PI3KCA and RHEB1 in the 23 cases for which genomic DNA was available. Fluorescence in situ hybridisation analysis for mTOR and RPS6 loci showed that 11 of 33 and 21 of 44 tumours had loss of one copy of the respective genes, results which correlated with the loss of the relevant total proteins. Fluorescence in situ hybridisation analysis for loci containing TSC1 and TSC2 revealed that all cases analysed harboured two copies of the respective genes. On the basis of p-mTOR and or p-p70S6K expression there is evidence indicating that 65% of the chordomas studied may be responsive to mTOR inhibitors, rapamycin or its analogues, and that patients may benefit from combined therapy including drugs that inhibit AKT.
Asunto(s)
Cordoma/genética , Fosfatidilinositol 3-Quinasas/genética , Proteínas Proto-Oncogénicas c-akt/genética , Factores de Transcripción/genética , Proteínas Supresoras de Tumor/genética , Adulto , Anciano , Anciano de 80 o más Años , Western Blotting , Cordoma/patología , Femenino , Humanos , Inmunohistoquímica , Hibridación Fluorescente in Situ , Masculino , Diana Mecanicista del Complejo 1 de la Rapamicina , Persona de Mediana Edad , Complejos Multiproteicos , Análisis de Secuencia por Matrices de Oligonucleótidos , Fosfatidilinositol 3-Quinasas/metabolismo , Análisis por Matrices de Proteínas , Proteínas , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/metabolismo , Serina-Treonina Quinasas TOR , Factores de Transcripción/metabolismo , Esclerosis Tuberosa/tratamiento farmacológico , Esclerosis Tuberosa/genética , Esclerosis Tuberosa/patología , Proteína 1 del Complejo de la Esclerosis Tuberosa , Proteína 2 del Complejo de la Esclerosis Tuberosa , Proteínas Supresoras de Tumor/metabolismo , Adulto JovenRESUMEN
Polymerase chain reaction (PCR) assessment of clonal immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangements is an important diagnostic tool in mature B-cell neoplasms. However, lack of standardized PCR protocols resulting in a high level of false negativity has hampered comparability of data in previous clonality studies. In order to address these problems, 22 European laboratories investigated the Ig/TCR rearrangement patterns as well as t(14;18) and t(11;14) translocations of 369 B-cell malignancies belonging to five WHO-defined entities using the standardized BIOMED-2 multiplex PCR tubes accompanied by international pathology panel review. B-cell clonality was detected by combined use of the IGH and IGK multiplex PCR assays in all 260 definitive cases of B-cell chronic lymphocytic leukemia (n=56), mantle cell lymphoma (n=54), marginal zone lymphoma (n=41) and follicular lymphoma (n=109). Two of 109 cases of diffuse large B-cell lymphoma showed no detectable clonal marker. The use of these techniques to assign cell lineage should be treated with caution as additional clonal TCR gene rearrangements were frequently detected in all disease categories. Our study indicates that the BIOMED-2 multiplex PCR assays provide a powerful strategy for clonality assessment in B-cell malignancies resulting in high Ig clonality detection rates particularly when IGH and IGK strategies are combined.
Asunto(s)
Genes de Inmunoglobulinas , Leucemia de Células B/genética , Linfoma de Células B/genética , Reacción en Cadena de la Polimerasa/métodos , Cromosomas Humanos Par 11 , Cromosomas Humanos Par 14 , Cromosomas Humanos Par 18 , Reordenamiento Génico , Genotipo , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Leucemia de Células B/diagnóstico , Leucemia de Células B/inmunología , Linfoma de Células B/diagnóstico , Linfoma de Células B/inmunología , Receptores de Antígenos de Linfocitos T/genética , Translocación GenéticaRESUMEN
Indirect antigenic stimulation by H. pylori-specific T cells is implicated in the development of low-grade gastric lymphoma of mucosa-associated lymphoid tissue (MALT), however, the role of direct antigen stimulation is unknown. To study the role of direct antigen stimulation in MALT lymphomagenesis and its relationship with the pathogenesis of distinct pathological lesions, which represent different stages of the tumour progression, we cloned and sequenced the rearranged immunoglobulin (Ig) heavy chain gene in three low-grade (two from the lung, one from the stomach) and one high-grade (from the stomach) cases. In the low-grade gastric case, we studied the Ig sequence in primary as well as its disseminated and recurrent tumours. In the high-grade gastric case, we analysed the Ig sequence in tumour cell populations microdissected from the residual diffuse low-grade lesions, diffuse high-grade areas from follicles colonized by high-grade blasts. Compared with the published germline sequences, the heavy chain variable (VH) genes of three MALT lymphomas, in which the putative germline was identified, contained frequent somatic mutations, showing a much higher ratio of replacement/silent mutations in the complementarity determining regions (CDRs) than the framework regions (FRs). Ongoing mutation as indicated by intraclonal variation of the Ig sequence clearly existed in low-grade tumour including its dissemination and recurrence, but was not evident in high-grade tumour cell populations including those microdissected from independent colonized follicles. In addition, the germlines of VH genes used by the three MALT lymphomas are frequently found in autoreactive antibodies. Our results suggest that MALT lymphoma derives from postgerminal centre memory B cells, possibly autoreactive B cell clones, and that direct antigen stimulation may play an important role in the clonal expansion of low-grade MALT lymphoma.
Asunto(s)
Antígenos/inmunología , Genes de Inmunoglobulinas/genética , Linfoma de Células B de la Zona Marginal/genética , Mutación , Animales , Secuencia de Bases , Femenino , Reordenamiento Génico de Cadena Pesada de Linfocito B , Humanos , Linfoma de Células B de la Zona Marginal/inmunología , Linfoma no Hodgkin/genética , Linfoma no Hodgkin/inmunología , Masculino , Datos de Secuencia MolecularRESUMEN
The formation of neoplastic B-cell follicles is universally accepted as diagnostic of a follicle centre cell (FCC) lymphoma. Low-grade B-cell lymphomas of mucosa-associated lymphoid tissue (MALT) are characterized by a diffuse infiltrate of cells of uncertain lineage known as "centrocyte-like" cells because of their resemblance to centrocytes (small cleaved cells). Some MALT lymphomas, however, contain numerous follicles and may even have a predominantly follicular appearance. These follicles may be reactive or show immunoglobulin (Ig) light-chain restriction, indicating their neoplastic nature. We have proposed that these neoplastic follicles are not composed of follicle centre cells but result from colonization of reactive follicles by CCL cells. In this study, the immunophenotype and genotype of 10 primary gastrointestinal lymphomas with a follicular component have been determined. One case exhibited the morphological, immunophenotypic, and genotypic features of FCC lymphoma (Ig light-chain restriction, CD10+, KB61 (CDw32)-, Jh, and bcl-2 gene rearrangement). Neoplastic follicles in the remaining nine cases, which showed the features of MALT lymphoma, were of a different phenotype (Ig light-chain restriction, CD10- KB61(CDw32)+), and these lymphomas showed Jh but not bcl-2 gene rearrangement. Taken in conjunction with the morphological features, these findings suggest that in these cases the neoplastic follicles formed as the result of colonization of previously reactive follicles by neoplastic CCL cells. Thus, not all lymphomas containing neoplastic follicles are of FCC origin. Follicular colonization, as seen in low-grade MALT lymphomas, is likely to be a recapitulation of an as yet undescribed normal immunological phenomenon that may involve marginal zone B cells.
Asunto(s)
Tejido Linfoide/patología , Linfoma de Células B/patología , Reordenamiento Génico , Humanos , Inmunohistoquímica , Linfoma de Células B/genética , Membrana Mucosa/patologíaRESUMEN
We describe four female patients with primary splenic low-grade non-Hodgkin's B-cell lymphomas with the morphology and immunophenotype of splenic marginal zone lymphocytes. The patients presented with splenomegaly, anemia, and weight loss. The bone marrow was involved in all four cases. Liver involvement was found in one patient; and in another, a CT scan revealed lymphadenopathy in the chest and abdomen. The histology of the spleen was characterized by broad concentric strands of monomorphic medium-sized lymphocytes around lymphoid follicles in one case and infiltrating follicles in two cases. Selective replacement of follicles was seen in one case. Tumor in splenic hilar lymph nodes (four cases) and liver (one case) was similar. Three patients remain well 4, 9, and 12 months, respectively, after splenectomy without further treatment. One patient who received chemotherapy died 1 year after splenectomy.
Asunto(s)
Linfoma de Células B/patología , Neoplasias del Bazo/patología , Adulto , Anciano , Southern Blotting , Sondas de ADN , ADN de Neoplasias/análisis , Femenino , Humanos , Técnicas para Inmunoenzimas , Inmunohistoquímica , Linfoma de Células B/diagnóstico , Linfoma de Células B/cirugía , Persona de Mediana Edad , Bazo/patología , Esplenectomía , Neoplasias del Bazo/diagnóstico , Neoplasias del Bazo/cirugíaRESUMEN
The distinction between reactive and neoplastic cutaneous T-cell infiltrates is difficult and requires good clinicopathologic correlation. Many cases manifest changes that are at the borderline between the two. The polymerase chain reaction (PCR) has been reported to detect monoclonality in 52-90% of cutaneous T-cell lymphomas and may be of use in the diagnosis of histologically borderline lesions. We have investigated the use of PCR in a series of borderline lesions including borderline biopsy samples from patients who subsequently developed cutaneous lymphoma. PCR amplification of T-cell receptor (TCR)-gamma chain gene was performed on formalin-fixed, paraffin-embedded tissue from 27 cases of clinically and histologically typical mycosis fungoides (MF), 22 borderline biopsy samples from 10 patients who subsequently developed MF (pre-MF), 32 clinically suspicious, histologically borderline lesions, and 31 cases of chronic dermatitis. Monoclonality was demonstrated in 16 of 27 (59%) cases of MF, 10 of 22 (50%) pre-MF biopsy samples (six of 10 patients), and six of 32 (19%) borderline biopsy samples. The same size monoclonal band was detected in pre-MF biopsy samples from six of seven patients in which a band was demonstrated in the diagnostic MF biopsy. Sequencing confirmed that the MF biopsy sample and the pre-MF biopsy sample contained the same clone. The 31 dermatitis cases gave rise to polyclonal PCR products. Monoclonality can be demonstrated using PCR in 59% of MF cases, which is comparable with other T-cell lymphomas and in up to 50% of borderline biopsy samples in patients who later develop lymphoma. Detection of T-cell monoclonality by PCR is strong evidence of an established or evolving cutaneous T-cell lymphoma.
Asunto(s)
Linfoma Cutáneo de Células T/patología , Reacción en Cadena de la Polimerasa , Secuencia de Bases , Biopsia , Complejo CD3/análisis , Enfermedad Crónica , Células Clonales , Dermatitis/patología , Diagnóstico Diferencial , Epidermis/química , Epidermis/patología , Humanos , Linfoma Cutáneo de Células T/genética , Datos de Secuencia Molecular , Micosis Fungoide/patología , Estadificación de Neoplasias , Linfocitos T/patologíaRESUMEN
We describe the morphologic, immunohistologic, and genotypic characteristics of 13 cases of true histiocytic lymphomas. Six cases presented with primary gastrointestinal involvement, five with lymphadenopathy, the other sites involved being the bone marrow and the skin. The neoplastic cells displayed large abundant eosinophilic cytoplasm, occasionally vacuolated with folded or bizarre-shaped nuclei with prominent nucleoli. Mitotic figures were numerous. Multinucleated cells were common. The pattern of growth was usually diffuse and noncohesive. Spindle cell sarcoma-like areas were evident in five cases, with a prominent foam cell component in four cases. All cases expressed histiocyte-associated markers (CD68, lysozyme, alpha-1-antitrypsin), CD45 or CD45RO, and were negative for CD1a, epithelial, and B- and T-cell lineage-specific markers. Reactivity for S-100 was observed in a variable proportion of cells in 11 cases. The proliferation fraction varied from 3 to 88%. Genotypic analysis for T-cell receptor or immunoglobulin gene rearrangement demonstrated a germline configuration in all cases. We demonstrate that true histiocytic lymphoma is a rare distinctive pathologic entity that may be defined by immunohistochemical criteria and that recognition among histiocytic disorders is important for clinical and prognosis reasons.
Asunto(s)
Biomarcadores de Tumor/análisis , Neoplasias de la Médula Ósea/patología , ADN de Neoplasias/análisis , Neoplasias Gastrointestinales/patología , Enfermedades Linfáticas/patología , Linfoma de Células B Grandes Difuso/patología , Neoplasias Cutáneas/patología , Adulto , Anciano , Anciano de 80 o más Años , Antígenos de Neoplasias/análisis , Neoplasias de la Médula Ósea/química , Neoplasias de la Médula Ósea/genética , Preescolar , Cartilla de ADN/química , Femenino , Neoplasias Gastrointestinales/química , Neoplasias Gastrointestinales/genética , Genotipo , Histiocitos/patología , Humanos , Técnicas para Inmunoenzimas , Inmunofenotipificación , Enfermedades Linfáticas/genética , Enfermedades Linfáticas/metabolismo , Linfoma de Células B Grandes Difuso/química , Linfoma de Células B Grandes Difuso/genética , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Neoplasias Cutáneas/química , Neoplasias Cutáneas/genéticaRESUMEN
Low-grade mucosa-associated lymphoid tissue (MALT) type B-cell lymphomas of the salivary gland arise in a background of myoepithelial sialadenitis (MESA), usually in association with Sjögren's syndrome. The distinction between benign MESA and early lymphoma has proved difficult using histological criteria alone and the significance of B-cell monoclonality in this respect is controversial. We have used immunohistochemistry and polymerase chain reaction (PCR) amplification of immunoglobulin heavy-chain VDJ regions to assess clonality in biopsies from 45 patients with lymphoid infiltration of the parotid. Sequential biopsies spanning 3-18 years were available from seven patients, three of whom had developed disseminated nodal B-cell lymphoma. In light of previous studies, each biopsy was additionally analyzed for the presence of t(14;18) and Epstein Barr Virus (EBV) DNA using PCR. Monoclonality was detected in 34/45 cases. Comparison of histology with clonality confirmed earlier suggestions that the emergence of an identifiable population of centrocyte-like B cells around ducts or epithelial islands correlated with monoclonality. In six of seven patients with sequential biopsies PCR fragments of identical size were amplified from each biopsy, suggesting that demonstrable monoclonality in "lymphoepithelial" lymphoproliferative lesions of the salivary gland is indicative of lymphoma. No t(14;18) chromosome translocations were identified; EBV sequences were detected in three of 45 cases.
Asunto(s)
Herpesvirus Humano 4/aislamiento & purificación , Linfoma de Células B de la Zona Marginal/etiología , Neoplasias de la Parótida/etiología , Sialadenitis/complicaciones , Translocación Genética , Adulto , Anciano , Anciano de 80 o más Años , Linfocitos B/patología , Secuencia de Bases , Cromosomas Humanos Par 14 , Cromosomas Humanos Par 18 , Células Clonales , Femenino , Humanos , Inmunohistoquímica , Linfoma de Células B de la Zona Marginal/genética , Linfoma de Células B de la Zona Marginal/inmunología , Linfoma de Células B de la Zona Marginal/virología , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Neoplasias de la Parótida/genética , Neoplasias de la Parótida/inmunología , Neoplasias de la Parótida/virología , Reacción en Cadena de la Polimerasa , Sialadenitis/genética , Sialadenitis/inmunología , Sialadenitis/virologíaRESUMEN
We report a case of a composite lymphoma comprising both mande cell lymphoma and a plasmacytoma. The two components were morphologically and immunohistochemically quite distinct. These properties, coupled with both direct and indirect molecular genetic evidence, suggest that these were two separate tumours occurring together by chance or by unknown oncogenic mechanisms, rather than clonally linked lymphomas.
Asunto(s)
Biomarcadores/análisis , Linfoma no Hodgkin/complicaciones , Linfoma no Hodgkin/patología , Plasmacitoma/complicaciones , Plasmacitoma/patología , Biopsia , Ciclina D1/análisis , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas kappa de Inmunoglobulina/análisis , Cadenas lambda de Inmunoglobulina/análisis , Inmunohistoquímica , Inmunofenotipificación , Ganglios Linfáticos/química , Ganglios Linfáticos/patología , Linfoma no Hodgkin/genética , Masculino , Persona de Mediana Edad , Plasmacitoma/química , Plasmacitoma/genética , Reacción en Cadena de la PolimerasaRESUMEN
Primary lymphoma is an uncommon tumor in the breast. Review of the literature shows two distinct clinicopathologic groups. One, which affects young women, is frequently bilateral, is often associated with pregnancy, and is a Burkitt-type lymphoma. The second group affects older women, is usually B-cell non-Hodgkins-type lymphoma, and presents clinical features identical to carcinoma of the breast. Three recent studies have suggested that up to half of the cases in the latter group belong to the category of lymphomas arising from the mucosa-associated lymphoid tissues. We have identified nine cases of primary lymphoma from the files of Guys Hospital Clinical Oncology Breast Unit in the 16-year period from 1974 to 1990. The clinical features have been reviewed and the tumors have been evaluated both on a morphologic and an immunohistochemical basis, and seven of nine of the cases have been screened for t[14;18] translocation using the polymerase chain reaction. All the tumors occurred in women older than 50 years and who presented with features of mammary carcinoma. One tumor was true histiocytic lymphoma; the remaining eight cases were B-cell lymphomas. Seven of the latter cases were high-grade B-cell lymphomas and one was a true follicular lymphoma. None of our cases showed the features of lymphoma arising in mucosa-associated lymphoid tissue.
Asunto(s)
Neoplasias de la Mama/epidemiología , Linfoma de Células B/epidemiología , Linfoma de Células B Grandes Difuso/epidemiología , Anciano , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Cromosomas Humanos Par 14 , Cromosomas Humanos Par 18 , ADN de Neoplasias/genética , Femenino , Estudios de Seguimiento , Humanos , Inmunohistoquímica , Incidencia , Antígenos Comunes de Leucocito/análisis , Linfoma de Células B/genética , Linfoma de Células B/patología , Linfoma de Células B Grandes Difuso/genética , Linfoma de Células B Grandes Difuso/patología , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Translocación Genética , Reino Unido/epidemiologíaRESUMEN
A case of primary low-grade pulmonary lymphoma is described. The histologic features conformed to those laid down by Saltzstein for a diagnosis of "pseudolymphoma." However, the immunocytochemical and molecular investigations confirmed the tumor to be a low-grade B-cell lymphoma of mucosa-associated lymphoid tissue (MALT). Cytogenetic studies revealed an abnormal karyotype with a translocation t(1;14). This is the first reported case of an abnormal karyotype in a case of a lymphoma of MALT.
Asunto(s)
Neoplasias Pulmonares/patología , Linfoma de Células B/patología , Antígenos de Diferenciación/metabolismo , Antígenos de Diferenciación de Linfocitos B/metabolismo , Antígenos de Neoplasias/metabolismo , Moléculas de Adhesión Celular/metabolismo , Cromosomas Humanos Par 1 , Cromosomas Humanos Par 14 , ADN de Neoplasias/genética , Femenino , Humanos , Cadenas lambda de Inmunoglobulina/metabolismo , Inmunohistoquímica , Cariotipificación , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Linfoma de Células B/genética , Linfoma de Células B/metabolismo , Persona de Mediana Edad , Neprilisina , Receptores de Complemento/metabolismo , Receptores de Complemento 3d , Translocación Genética/genéticaRESUMEN
AIMS: To compare the sensitivity of detecting Helicobacter pylori in gastric biopsy and resection specimens using tinctorial and silver impregnation stains, immunohistochemistry and the polymerase chain reaction (PCR). METHODS: Formalin fixed, paraffin wax embedded tissue from 33 gastric biopsy specimens (26 showing chronic gastritis and seven showing low grade mucosa associated lymphoid tissue (MALT) lymphoma) together with blocks of uninvolved mucosa from gastrectomy specimens for MALT lymphoma (five cases) were studied. Consecutive sections were stained using haematoxylin and eosin, Giemsa, the Warthin-Starry silver stain, and a polyclonal antibody directed against H pylori using an immunoperoxidase technique following heat induced antigen retrieval. PCR analysis of DNA extracted from a further section was carried out using primers which amplified a 411 base pair fragment of the urease A gene. RESULTS: H pylori was detected in 14 (37%) sections stained with haematoxylin and eosin, 21 (55%) with Giemsa, 23 (61%) with Warthin-Starry, and 25 (66%) stained with the antibody. Seventeen (45%) cases were positive on PCR. Immunohistochemistry was positive in all cases in which H pylori was detected by other methods. CONCLUSION: Immunohistochemistry using an immunoperoxidase technique following heat induced antigen retrieval for detecting H pylori in gastric biopsy and resection specimens is highly sensitive and easy to use.
Asunto(s)
Gastritis/microbiología , Helicobacter pylori/aislamiento & purificación , Linfoma de Células B de la Zona Marginal/microbiología , Estómago/microbiología , Secuencia de Bases , Enfermedad Crónica , Humanos , Técnicas para Inmunoenzimas , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Coloración y EtiquetadoRESUMEN
AIMS: To evaluate the polymerase chain reaction (PCR) demonstration of clonal immunoglobulin heavy chain gene rearrangements using routinely prepared, unstained, and stained formalin fixed, paraffin wax embedded tissue samples. METHODS: Extracts from (a) fresh frozen tissue samples, (b) unstained, and (c) haematoxylin and eosin stained formalin fixed, paraffin wax embedded 5 microns tissue sections from 42 cases of low grade B cell lymphoma, all shown to be monoclonal by Southern blot analysis, were analysed using PCR. Two regions of the variable segment of the immunoglobulin heavy chain gene were amplified (framework 2 to joining region [Fr2/JH] and framework 3 to joining region [Fr3/JH]). Twelve samples of reactive lymphoid tissue were studied as controls. Products from each case were directly compared on polyacrylamide gels. RESULTS: Using both primer combinations, monoclonality was detected in 38 of 42 (90%) cases using fresh material, 37 of 42 (88%) using unstained paraffin wax embedded samples, and in 35 of 42 (83%) cases using haematoxylin and eosin stained sections. No false positive results attributable to fixation, processing, or staining were identified, although the efficiency of amplification using the Fr2/JH primers was significantly reduced. CONCLUSIONS: PCR determination of B cell clonality using paraffin wax embedded material is sufficiently sensitive and reliable for use as a routine diagnostic adjunct to conventional morphological and immunocytochemical assessment of lymphoproliferative disease.
Asunto(s)
ADN de Neoplasias/análisis , Reordenamiento Génico de Cadena Pesada de Linfocito B/genética , Linfoma de Células B/diagnóstico , Secuencia de Bases , Southern Blotting , Células Clonales , Cartilla de ADN/genética , Electroforesis en Gel de Poliacrilamida , Estudios de Evaluación como Asunto , Humanos , Datos de Secuencia Molecular , Adhesión en Parafina , Reacción en Cadena de la PolimerasaRESUMEN
AIM: To establish a simple and reliable polymerase chain reaction (PCR) methodology for random amplification of whole genomic DNA from limited histopathological samples. METHODS: Trace amounts of genomic DNA extracted from fresh tissue and individual lymphoid follicles microdissected from archival paraffin wax tissue sections were amplified using a two-phase PCR protocol with random hexamers as primers (RP-PCR). The randomly amplified DNA samples were used as templates for specific PCR amplifications. To check the fidelity of the RP-PCR, products of the specific PCR amplifications were further analysed by single stranded conformation polymorphism (SSCP) or sequencing. RESULTS: Using a minute fraction of RP-PCR template pool, multiple PCR analyses, including those for beta globin gene, p53 gene (exon 5-6, exon 7, exon 8-9 and exon 7-9), and rearranged immunoglobulin heavy chain gene fragments (VH framework 3 to JH and VH framework 2 to JH) were successfully performed. No artefactual mutations were identified in the products of these specific PCR reactions by SSCP or sequencing when compared with the products from the original DNA. CONCLUSION: This method is simple and reliable, and permits multiple genetic analyses when only a limited amount of tissue is available.
Asunto(s)
ADN/genética , Reacción en Cadena de la Polimerasa/métodos , Secuencia de Bases , Cartilla de ADN , Electroforesis en Gel de Agar , Humanos , Tejido Linfoide/química , Datos de Secuencia Molecular , Adhesión en ParafinaRESUMEN
AIMS: To evaluate polymerase chain reaction (PCR) amplification of T cell receptor (TCR) beta and gamma chain genes as a means of demonstrating monoclonality in T cell lymphomas using histological samples; to compare the performance of PCR with Southern blot analysis. METHODS: TCR-beta, TCR-gamma and immunoglobulin heavy chain (IGH) genes were analysed using PCR in 55 cases of T cell lymphoma (28 frozen tissue and 27 paraffin wax embedded samples), diagnosed using morphological and immunohistochemical criteria. The 28 frozen samples were subjected to Southern blot analysis using TCR-beta, TCR-gamma and IGH gene probes. Twenty five B cell lymphomas and 21 non-neoplastic lymphoid tissue samples were used as controls. RESULTS: Using TCR-beta PCR, monoclonality was detected in 24 (44%) of 55 T cell lymphomas compared with 43 (78%) of 55 using TCR-gamma PCR and in 82% with both techniques. Five (9%) of 55 T cell lymphomas were IGH PCR positive. None of the non-neoplastic lymphoid control samples were PCR positive. All B cell lymphomas showed a polyclonal pattern with TCR-beta PCR while a single B cell lymphoma was positive using TCR-gamma primers. With TCR-beta PCR, a monoclonal result was seen in 12 (43%) of 28 frozen samples of T cell lymphoma, compared with 23 (82%) of 28 using Southern blot analysis. With TCR-gamma PCR, 19 (68%) of 28 frozen tissue samples were positive, compared with 26 (93%) of 28 using Southern blot analysis. A single case showed IGH rearrangement by Southern blot analysis. CONCLUSION: TCR-gamma PCR should be the method of choice for analysis of clonality in paraffin wax embedded sections of lymphoproliferative lesions, as TCR-beta PCR has a high false negative rate. Southern blot analysis remains the most successful technique when sufficient fresh tissue samples and resources are available.
Asunto(s)
Reordenamiento Génico de la Cadena beta de los Receptores de Antígenos de los Linfocitos T , Reordenamiento Génico de la Cadena gamma de los Receptores de Antígenos de los Linfocitos T , Linfoma de Células T/genética , Secuencia de Bases , Southern Blotting , Electroforesis en Gel de Poliacrilamida , Genes de Inmunoglobulinas , Humanos , Datos de Secuencia Molecular , Células Madre Neoplásicas/patología , Reacción en Cadena de la PolimerasaRESUMEN
A case of fatal Epstein-Barr virus (EBV) associated lymphoproliferative disorder is reported in an 11 month old female. Heavy infiltrates of CD20 + and EBV EBER mRNA expressing lymphoid blasts were found to cause a series of ulcers along the entire length of the gastrointestinal tract and there was an ileal perforation. Similar infiltrates were also found in lymph nodes, spleen, and liver. Although blood phenotypic analysis performed shortly before her death revealed a severe decrease in T lymphocytes, neither the patient nor other members of her family had a history of primary or secondary immunodeficiency. EBV infection is common in children. However, such a fatal infection of the virus has not apparently been described previously in infants without pre-established immunodeficiency.
Asunto(s)
Enfermedades Gastrointestinales/virología , Infecciones por Herpesviridae/complicaciones , Herpesvirus Humano 4/aislamiento & purificación , Trastornos Linfoproliferativos/virología , Infecciones Tumorales por Virus/complicaciones , Resultado Fatal , Femenino , Enfermedades Gastrointestinales/patología , Infecciones por Herpesviridae/patología , Humanos , Lactante , Trastornos Linfoproliferativos/patología , Infecciones Tumorales por Virus/patologíaRESUMEN
Caecal biopsy specimens from Jamaican children with the Trichuris dysentery syndrome (TDS) and age matched Jamaican controls were investigated by immunohistochemistry and by light microscopy. Biopsy specimens from all children (with TDS and controls) showed a mild to moderate increase in inflammatory cells. Except in the vicinity of the worm, where the epithelium was flattened, there was no other epithelial abnormality. Compared with controls, children with TDS had increased IgM lamina propria plasma cells and decreased intraepithelial T cells. There was also an increase in crypt epithelial cell proliferation. Lamina propria T cells (both activated and non-activated) were no more common in children with the Trichuris syndrome than controls. Epithelial cell HLA-DR and VLA-1 expression (which are increased in other colitides) were the same in both groups. Despite the presence of large worm burdens and chronic dysentery, therefore, only minor changes were seen in the caecal mucosa of children with TDS.
Asunto(s)
Ciego/inmunología , Disentería/inmunología , Tricuriasis/inmunología , Antígenos CD/análisis , Biopsia , Ciego/patología , Niño , Preescolar , Disentería/patología , Femenino , Humanos , Técnicas para Inmunoenzimas , Masculino , Células Plasmáticas/inmunología , Síndrome , Linfocitos T/inmunología , Tricuriasis/patologíaRESUMEN
The recently described B-cell lymphomas arising in mucosa-associated lymphoid tissue (MALT) form a distinct clinico-pathologic group of non-Hodgkin's lymphoma, and therefore would be expected to be characterized by a recurrent chromosomal aberration. We have analyzed the cytogenetics of 23 cases of MALT lymphomas arising in the stomach, small intestine, lung, and lacrimal gland. In each case the presence of an abnormal clonal cell population was confirmed by the identification of rearranged bands when digested tumor DNA was hybridized with a probe to the joining region of the immunoglobulin heavy chain gene. Metaphase spreads were obtained in 14 cases, of which 9 cases showed an abnormal karyotype. Although no unifying aberration was detected, rearrangements of chromosome 1p, and numerical abnormalities of chromosomes 3 and 7, may play a role in the genesis of these tumors.
Asunto(s)
Aberraciones Cromosómicas , Linfoma de Células B/genética , Cromosomas Humanos Par 1 , Cromosomas Humanos Par 3 , Cromosomas Humanos Par 7 , Mucosa Gástrica/patología , Reordenamiento Génico de Linfocito B/genética , Genes de Inmunoglobulinas , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Región de Unión de la Inmunoglobulina/genética , Tejido Linfoide/patología , Linfoma de Células B/patología , Membrana Mucosa/patologíaRESUMEN
OBJECTIVES: Sjögren's syndrome is an autoimmune disorder in which patients have a well-recognized risk of developing malignant lymphoma. Although some clinical parameters may herald the onset of lymphoma, few reliable histologic or molecular markers are available that predict progression to a malignant lymphoproliferative disorder. The purpose of this study was to identify the prevalence of immunoglobulin heavy chain monoclonality in labial gland biopsies of patients with Sjögren's syndrome and to compare this to clinical outcome. STUDY DESIGN: The polymerase chain reaction was applied to 76 sequential labial salivary gland biopsies from patients under investigation for Sjögren's syndrome. A seminested polymerase chain reaction technique was used on DNA extracted from formalin-fixed, paraffin-embedded tissue to amplify the V-D-J region of the immunoglobulin heavy chain gene. Thirty-four randomly selected labial salivary glands that showed nonspecific sialadenitis from patients without Sjögren's syndrome were used as controls. RESULTS: Monoclonality, as defined by a single band on polyacrylamide gel electrophoresis was detected in 11 cases (14.5%). Of cases that showed monoclonality, four patients were subsequently diagnosed with extrasalivary lymphoma. In each case the rearranged bands in the lip biopsy and the lymphoma were the same size. In one patient who later developed lymphoma, a monoclonal rearranged immunoglobulin band was not identified. In addition, no cases of the translocation t(14;18) were identified by polymerase chain reaction in any of the lip biopsies showing heavy chain monoclonality or in any of the extrasalivary gland lymphomas. CONCLUSIONS: These results suggest that monoclonal immunoglobulin heavy chain gene rearrangements are a relatively common finding in patients with Sjögren's syndrome and may prove to be a useful marker for predicting the progression to, and early detection of malignant lymphoma.