Clinicopathologic Assessment of Ocular Adnexal Lymphoproliferative Lesions at a Tertiary Eye Hospital in Iran.

BACKGROUND: The most common type of ocular lymphoma is non-Hodgkin lymphoma (NHL), categorized into two groups: indolent (slow growing) and aggressive (rapid growing). Differentiating benign reactive lymphoid hyperplasia (RLH) from malignant ocular adnexal lymphoma (OAL) is challenging. Histopathology, immunohistochemistry (IHC) and ow cytometry have been used as diagnostic tools in such cases. MATERIALS AND METHODS: In this retrospective case series, from 2002 to 2013 at Farabi Eye Center, 110 patients with ocular lymphoproliferative disease were enrolled. Prevalence, anatomical locations, mean age at diagnosis and the nal diagnosis of the disease with IHC were assessed. Comparison between previous pathologic diagnoses and results of IHC was made. Immunoglobulin light chains and B-cell and T-cell markers and other immuno-phenotyping markers including CD20, CD3, CD5, CD23, CD10, CYCLIND1 and BCL2 were evaluated to determine the most accurate diagnosis. The lymphomas were categorized based on revised European-American lymphoma (REAL) classi cation. RESULTS: Mean age±SD (years) of the patients was 55.6 ±19.3 and 61% were male. Patients with follicular lymphoma, large B-cell lymphoma or chronic lymphocytic leukemia/small cell lymphoma (CLL/SLL) tended to be older. Nine patients with previous diagnoses of low grade B-cell lymphoma were re-evaluated by IHC and the new diagnoses were as follows: extranodal marginal zone lymphoma(EMZL) (n=1), SLL(n=1), mantle cell lymphoma (MCL) (n=3), reactive lymphoid hyperplasia RLH (n=2). Two cases were excluded due to poor blocks. Flow cytometry reports in these seven patients revealed SLL with positive CD5 and CD23, MCL with positive CD5 and CyclinD1 and negative CD23, EMZL with negative CD5,CD23 and CD10. One RLH patient was negative for Kappa/Lambda and positive for CD3 and CD20 and the other was positive for all of the light chains, CD3 and CD20. Orbit (49.1%), conjunctiva (16.1%) and lacrimal glands (16.1%) were the most common sites of involvement. CONCLUSIONS: Accurate pathological classi cation of lesions is crucial to determine proper therapeutic approaches. This can be achieved through precise histologic and IHC analyses by expert pathologists.

[Assessment of BIOMED-2 assays for detection of clonal Ig gene rearrangements in mature B-cell lymphomas].

OBJECTIVE: To evaluate the efficiency of the BIOMED-2 PCR assay and its implication in the diagnosis of mature B-cell non-Hodgkin's lymphomas. METHODS: Clinical, 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. RESULTS: Adequate 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. CONCLUSION: BIOMED-2 assay is highly sensitive and specific for the detection of clonal B cell gene rearrangement using routine paraffin embedded formalin fixed specimens.

The importance of accurate pathological assessment of lymph node involvement in colorectal cancer.

This review presents an up-to-date analysis of the importance of accurate pathological lymph node staging in colorectal cancer. Lymph node staging is reliant on the technique of the surgeon and the pathologist as well as methods employed in the histopathology laboratory, and is vital for determining appropriate therapy. The significance of micrometastatic nodal disease is evaluated and new techniques for pathological evaluation are discussed. Recommendations for evaluation of lymph node status in colorectal cancer are provided based on current scientific evidence, and standardization of pathological dissection and laboratory handling is advocated.

Clonality assessment of lymphoproliferative disorders by multiparameter flow cytometry of paraffin-embedded tissue: an additional diagnostic tool in surgical pathology.

A major drawback of immunohistochemical detection of monoclonality in B-cell lymphoproliferative disorders is the lack of contrast between surface-immunoglobulin staining and extracellular immunoglobulin staining. To bypass this drawback, immunophenotyping of single-cell suspensions by flow cytometry is commonly used. Although the expression of immunoglobulin light chain subtype can be quantified rapidly and reliably, the technique is hampered by the requirement of fresh unfixed material. We applied a recently developed technique for the isolation of single cells from formalin-fixed, paraffin-embedded material to measure clonality in B-cell lymphoproliferative disorders (lymphoid tissue (n = 10) and non-Hodgkin's B-cell lymphoma (n = 10). Immunocytochemistry indicated that common cell surface markers as well as the immunoglobulin light chains could be detected in the cell suspensions derived from archival material. In addition, the technique also allowed combined high-resolution DNA flow cytometric analysis. To investigate the effect of formalin fixation on cross-linking of extracellular immunoglobulins to lymphocytes, a double-immunostaining experiment for both light chain immunoglobulins (kappa and lambda) was performed. This experiment showed that this cross-linking was minimal (less than 2%). All cases of reactive lymphoid hyperplasia were DNA diploid and showed a polyclonal expression of immunoglobulin light chains. In contrast, in 9 of 10 non-Hodgkin's B-cell lymphomas, monoclonality was established on the basis of light chain expression, whereas only 6 of 9 cases were conclusive by immunohistochemistry. Four of the 9 cases were DNA aneuploid. One case did not show light chain expression at all by both techniques. However, this case could be classified as malignant by flow cytometric analysis because of the DNA-aneuploid nature of the B-cell subpopulation. The average S-phase fraction (SPF) of the B cells in the reactive lymphoid tissues was 3.5%. The mean SPF values for B cells in DNA-diploid cases of lymphomas was 3.0%, whereas the mean SPF of B cells in DNA-aneuploid cases was 6.1%. The presented technique is superior to immunohistochemistry for the detection of monoclonality in B-cell lymphoproliferative disorders and therefore provides a powerful tool to support the diagnosis of malignant lymphoma in routinely processed archival samples of lymphoid tissues.

Confocal microscopy assessment of lymphoid tissues with follicular hyperplasia from patients infected with human immunodeficiency virus type 1.

OBJECTIVE: To characterize human immunodeficiency virus (HIV) infection of lymphoid tissues during follicular hyperplasia. METHODS: We examined 10 tonsil/adenoid, 3 parotid lymphoepithelial cyst, and 7 lymph node specimens that had been surgically removed from 13 patients infected with HIV-1. Characteristics of productive HIV-1 infection were assessed using immunocytochemistry for HIV-1 p24. Cellular colocalization was determined with the aid of a confocal microscope using double immunofluorescent staining for HIV-1 p24 and cell-specific markers. RESULTS: All specimens showed follicular hyperplasia. Using confocal microscopy with three-dimensional reconstruction, HIV-1 p24 was seen to be "intimately" colocalized with CD21 within the germinal centers. While lymphoid follicles were generally hyperplastic, only a subset of these follicles contained HIV-1 p24. Occasional HIV-1-expressing mononuclear cells identified outside follicles stained for CD68 or CD3. CONCLUSIONS: The differential involvement of hyperplastic follicles by HIV-1 within individual lymphoid tissues and the intimate colocalization of HIV-1 p24 and CD21 suggest that infected follicular dendritic cells may be an important reservoir of HIV-1 during follicular hyperplasia.