Hyperplastic Germinal Centers Research Articles

Histopathological studies of visceralized Leishmania (Leishmania) amazonensis in mice experimentally infected

BALB/c, C57BL/6, and DBA/2 mice were subcutaneously infected in the left footpad by injecting 104 Leishmania (Leishmania) amazonensis amastigotes. Mice were sacrificed 20, 30, 40, 60 and 90 days post-infection. Fragments of liver, kidney, spleen, skin, and draining lymph node were collected for histological examination. Light microscopy showed that at 20 days after infection BALB/c mice presented discrete inflammatory infiltrates in the skin made up of eosinophils, lymphocytes, and rare parasitized macrophages. Ninety days post-infection, the dermis showed necrotic tissue, large numbers of mononuclear cells and vacuolated macrophages filled with amastigotes. Forty days post-infection, the draining lymph nodes showed hyperplastic germinal centers, increase of high endothelial venules and apoptosis in germinal center cells. After the first 3 months post-infection, the involvement of spleen, kidney and liver was discreet, being characterized by multifocal inflammatory infiltrates. Eight months after infection, the animals presented metastatic lesions in the contralateral footpad and nose. In deep dermis, there was remarkable proliferation of fibroblasts associated with collagen fibers. The liver showed multifocal granulomas and mononuclear infiltrate around the blood vessels, but no parasites were observed, except in one animal. In some mice there were immature cells of the hematopoietic lineage. Both BALB/c and C57BL/6 mice presented osteonecrosis, which is characterized by pycnotic osteocytes and empty lacunae at the point of inoculation and subsequently, replacement of this tissue by fibrous connective tissue and colonization of the bone marrow. A diffuse inflammatory process composed of mononuclear cells and rare parasites were seen in the kidneys. In one mouse, bone marrow cells were observed in the renal medulla along with where free amastigotes. DBA/2 mice developed a mild infection and they did not visceralize. In conclusion, our data demonstrates that in susceptible mice L. (L.) amazonensis, a causative agent of tegumentary leishmaniasis, causes pathological changes similar to those produced by Leishmania (L.) infantum in both humans and canids.

Histopathological studies of visceralized Leishmania (Leishmania) amazonensis in mice experimentally infected

BALB/c, C57BL/6, and DBA/2 mice were subcutaneously infected in the left footpad by injecting 10 4 Leishmania ( Leishmania) amazonensis amastigotes. Mice were sacrificed 20, 30, 40, 60 and 90 days post-infection. Fragments of liver, kidney, spleen, skin, and draining lymph node were collected for histological examination. Light microscopy showed that at 20 days after infection BALB/c mice presented discrete inflammatory infiltrates in the skin made up of eosinophils, lymphocytes, and rare parasitized macrophages. Ninety days post-infection, the dermis showed necrotic tissue, large numbers of mononuclear cells and vacuolated macrophages filled with amastigotes. Forty days post-infection, the draining lymph nodes showed hyperplastic germinal centers, increase of high endothelial venules and apoptosis in germinal center cells. After the first 3 months post-infection, the involvement of spleen, kidney and liver was discreet, being characterized by multifocal inflammatory infiltrates. Eight months after infection, the animals presented metastatic lesions in the contralateral footpad and nose. In deep dermis, there was remarkable proliferation of fibroblasts associated with collagen fibers. The liver showed multifocal granulomas and mononuclear infiltrate around the blood vessels, but no parasites were observed, except in one animal. In some mice there were immature cells of the hematopoietic lineage. Both BALB/c and C57BL/6 mice presented osteonecrosis, which is characterized by pycnotic osteocytes and empty lacunae at the point of inoculation and subsequently, replacement of this tissue by fibrous connective tissue and colonization of the bone marrow. A diffuse inflammatory process composed of mononuclear cells and rare parasites were seen in the kidneys. In one mouse, bone marrow cells were observed in the renal medulla along with where free amastigotes. DBA/2 mice developed a mild infection and they did not visceralize. In conclusion, our data demonstrates that in susceptible mice L. ( L.) amazonensis, a causative agent of tegumentary leishmaniasis, causes pathological changes similar to those produced by Leishmania ( L.) infantum in both humans and canids.

Phenotypic Modulation of the Stromal Reticular Network in Normal and Neoplastic Lymph Nodes: Tissue Transglutaminase Reveals Coordinate Regulation of Multiple Cell Types

The lymph node paracortex is composed of a network of fibroblastic reticular cells (FRC) and reticular fibers linking sinuses to blood vessels. Using immunostaining for the inducible enzyme/adhesion molecule tissue transglutaminase (TG), we demonstrate coordinate regulation of multiple stromal cell types of this reticular network including FRC, endothelial cells and sinus lining cells. Tissue transglutaminase is expressed at low levels in the paracortex around primary follicles but is markedly up-regulated in stromal cells around hyperplastic germinal centers and then down-regulated around regressing follicles. In cultured FRC, TG is induced 5- to 20-fold and becomes colocalized with matrix proteins by treatment with interleukin-4, but not by other cytokines that are commonly increased in lymph upon immune activation. Coordinate TG up-regulation is observed surrounding nodules of follicular lymphoma (14 of 15 cases) and in the FRC and endothelium of classical Hodgkin's disease, two tumor types that also showed an activated paracortical phenotype. Small lymphocytic lymphoma showed minimal TG staining, with other lymphoma types showing patterns suggesting differential TG regulation. Using TG as a stromal marker, we have identified differential modulation of the phenotype of the lymph node reticular network that parallels change in the B-cell compartment.

Involvement of DNase gamma in apoptotic DNA fragmentation in histiocytic necrotizing lymphadenitis.

DNA fragmentation induced by endonucleases is a hallmark of apoptotic process. One of the endonucleases responsible for apoptotic DNA fragmentation has been identified as DNase gamma. It has been suggested that massive nuclear debris observed in histiocytic necrotizing lymphadenitis (HNL, Kikuchi disease) results mainly from apoptosis of T-lymphocytes rather than necrosis. To identify the role of DNase gamma in apoptotic foci of HNL paraffin embedded tissue sections of HNL and lymphadenopathy with hyperplastic germinal centers and paracortex in the neck lymph nodes were immunohistochemically stained for DNase gamma and the results compared with the reactivity of TdT-mediated dUTP nick end labeling (TUNEL). Immunoreactivity for DNase gamma was detected in fragmented and condensed nuclei found abundantly in HNL as well as TUNEL reactivity. Moreover, the ratio of DNase gamma immunoreactivity to TUNEL reactivity, which represents probability of apoptosis relevant to DNase gamma, was higher in foci of nuclear debris in HNL than in the hyperplastic paracortex in lymphadenopathy including T-cell apoptosis. Our findings suggest that apoptosis detected in HNL depends more specifically upon DNase gamma than apoptosis in the hyperplastic paracortex of lymphadenopathy, and that the dysregulation of DNase gamma activation is involved in apoptosis in HNL.

Angioimmunoblastic T-cell lymphoma with hyperplastic germinal centers: a clinicopathological and immunohistochemical study of 10 cases.

An absence of germinal centers is one of the histological characteristics of angioimmunoblastic T-cell lymphoma (AITL). We report here 10 unusual cases of AITL with hyperplastic germinal centers. The clinical presentation of each patient was characterized by generalized lymphadenopathy, constitutional symptoms and polyclonal hypergammaglobulinemia. The initial biopsy findings of each patient were similar and were characterized by hyperplastic germinal centers with ill-defined borders and a proliferation of high endothelial venules (HEV). In the paracortical area there was a mixed infiltrate including irregularly shaped clusters or small nests of clear cells in all cases. Moreover, the clear cells invaded the lymphoid follicles, resulting in expansion of the germinal centers, except for one case. Immunohistochemistry revealed that the tumor cells, including clear cells, were CD4-expressing T cells. Some of the atypical lymphocytes were also Bcl-6-positive. A majority of the follicular dendritic cell networks showed a normal/reactive or an expanded/disrupted pattern in all cases. Moreover, three lesions possessed a few large irregularly shaped proliferations of follicular dendritic cells around the HEV Four cases progressed to AITL within a few years. The present 10 cases probably represent an early stage of AITL preceding follicular dendritic cell hyperplasia. Detection of clear cells, Bcl-6-positive atypical T lymphocytes, and foci of irregularly shaped proliferation of follicular dendritic cells appears to be critical for early diagnosis and treatment of AITL with hyperplastic follicles.

Autoimmune disease-associated lymphadenopathy with histological appearance of T-zone dysplasia with hyperplastic follicles. A clinicopathological analysis of nine cases.

Autoimmune disease-associated lymphadenopathy shows marked histopathological and clinical diversity. We describe the clinicopathological and immunohistochemical findings of nine cases of autoimmune disease-associated lymphadenopathy, which posed a serious differential diagnostic problem regarding T-zone dysplasia with hyperplastic follicles. There were two males and seven females aged 25 to 65 years (median 37 years). The patients had multicentric lymphadenopathy in association with clinical and laboratory findings suggestive of an "autoimmune disease". Four patients were diagnosed to have systemic lupus erythematosus (SLE), and the remaining five patients had antiphospholipid antibody syndrome and Sjogren's syndrome (SS), rheumatoid arthritis (RA), chronic thyroiditis, RA and SS, and SLE and SS, respectively. None of the nine patients developed malignant lymphomas during the follow-up periods from 44 to 225 months (median 103 months). The lesions were characterized by paracortical hyperplasia with prominent vascular proliferation and many lymphoid follicles with germinal centers. The paracortical area usually contained numerous small T-lymphocytes without cytological atypia, accompanied by a variable number of plasma cells, B-immunoblasts, and histiocytes. Polymerase chain reaction analysis demonstrated no clonal rearrangement of the T-cell receptor chain gene in four cases examined, although immunoglobulin heavy chain rearrangement was detected in only one case. These findings suggest that autoimmune disease-associated lymphadenopathy, especially SLE, shares the histological features with T-zone dysplasia with hyperplastic follicles. The nine cases presented here should be differentiated from T-zone lymphoma with follicles and angioimmunoblastic lymphoma with hyperplastic germinal centers. To avoid overdiagnosis and overtreatment, we emphasize the need to turn attention to these clinical and laboratory findings as well as to the morphological features.

Bcl-6 expression in reactive follicular hyperplasia, follicular lymphoma, and angioimmunoblastic T-cell lymphoma with hyperplastic germinal centers: Heterogeneity of intrafollicular T-cells and their altered distribution in the pathogenesis of angioimmunoblastic T-cell lymphoma

Background: The Bcl-6 gene product, a nuclear phosphoprotein, is expressed independently of Bcl-6 gene rearrangement. In lymph nodes, expression of Bcl-6 protein is restricted to germinal center (GC) B-cells and 10% to 15% of CD3 CD4 + intrafollicular T cells. Interfollicular cells are negative for Bcl-6 protein, except for rare CD3 + CD4 + T cells. Recently, we reported cases of angioimmunoblastic T-cell lymphoma (AITL) with hyperplastic GCs (AITL/GC), and observed that borders of enlarged GCs were ill defined, with features suggestive of an outward migration of GC cells to surrounding interfollicular zones. This prompted a study of follicular borders with Bcl-6 staining in reactive follicular hyperplasias and follicular lymphomas to compare with AITL/GC. Materials and Methods: Formalin-fixed paraffin sections were used for immunostaining of Bcl-6. Six cases of AITL/GC, 12 nonspecific reactive follicular hyperplasia (FH), 7 HIV adenopathy, 10 follicular lymphoma (FL), and 8 typical AITL (ie, AITL without GC) were studied. Double staining for Bcl-6/CD20, Bcl-6/CD3, and Bcl-6/CD57 was performed in selected cases. Results: In FH and HIV adenopathy, staining for Bcl-6 revealed densely populated GCs with well-defined and regular GC borders, whereas Bcl-6 + cells were rare in the interfollicular areas. An occasional GC with an ill-defined border was invariably surrounded by a broad mantle zone; those with indistinct mantle zones had well-defined, regular borders. In FL, follicles were densely populated, and their borders were irregular, with some Bcl-6 + cells in the interfollicular zones. In AITL/GC, GCs were less dense, GC borders were ill defined and irregular, and the number of interfollicular Bcl-6 + cells was markedly increased. Double staining revealed that these interfollicular Bcl-6 + cells in AITL/GC were Bcl-6 +/CD3 +/CD20 −/CD57 − T cells. Moreover, CD3 + intrafollicular T cells were depleted in AITL/GC, whereas they were abundant in FH. Intrafollicular CD57 + cells did not stain for Bcl-6, and were also depleted in AITL/GC. In typical AITL, some neoplastic cells were positive for Bcl-6, showing variable degrees of staining. Conclusions: (1) GCs of AITL/GC differed from those of other reactive follicular hyperplasias and follicular lymphomas, and staining for Bcl-6 was useful to discern them. (2) Intrafollicular CD3 + T cells, many of which were also positive for Bcl-6, were markedly depleted in AITL/GC, with increased interfollicular Bcl-6 +/CD3 + cells, suggesting an outward migration of intrafollicular T cells in this condition. (3) Interfollicular Bcl-6 +/CD3 + cells in AITL/GC were too numerous to be accounted for by migration alone, suggesting local proliferation. (4) Intrafollicular CD57 + cells were negative for Bcl-6, indicating heterogeneity of the intrafollicular T-cell population. (5) Some neoplastic cells in AITL stained for Bcl-6, suggesting up-regulation of Bcl-6 expression in this tumor.

Activated cytotoxic lymphocytes in lymph nodes from human immunodeficiency virus (HIV) 1-infected patients: a light and electronmicroscopic study.

Persistent generalized lymphadenopathy often develops during HIV-infection. It is characterized by follicular hyperplasia which progresses over time to follicular involution and finally lymphocyte depletion. To determine whether activated cytotoxic T cells (CD8+) are present in the hyperplastic germinal centres, light and electronmicroscopic immunogold labelling with monoclonal antibodies were used to localize two cytotoxic molecules, perforin and TIA-1. Perforin and TIA-1-positive cells were detected in the follicles and paracortex of lymph nodes from HIV-infected patients, whereas labelling was seen only in cells of the paracortex in the hyperplastic lymph nodes from HIV-negative patients. Cytotoxic granules, staining positive for perforin and TIA-1, were identified by transmission electronmicroscopy, often in proximity to follicular dendritic cells within the hyperplastic germinal centres of only HIV-positive patients. These cytotoxic cells may play a role in the follicular dendritic cell loss and concomitant follicular involution that occur during the evolution of HIV disease.

Paracortical Nodular T-Cell Lymphoma

Peripheral T-cell lymphomas (PTCLs) are regarded as diffuse proliferations. We describe an unusual paracortical nodular growth pattern in four nodal PTCLs that were initially interpreted as atypical lymphoid hyperplasia in three patients and small B-cell lymphoma with plasmacytic differentiation in a fourth. The nodules were vague to easily discernible and produced minimal to partial architectural distortion. Sinuses were often open, and scattered cortical lymphoid follicles with atretic to hyperplastic germinal centers were present. Clusters of tumor cells abutted some follicles in all cases, and in one case they exhibited focal T-zone expansion. Hypervascularity was not prominent, but a few nodules surrounded epithelioid venules, imparting an angiofollicular appearance. The nodules were composed primarily of small lymphocytes with irregular nuclei admixed with scattered large transformed cells, both cell types having clear cytoplasm. Paraffin immunoperoxidase showed that the nodules were composed of T cells. Dendritic cell networks were present only in follicular centers. Southern blot analysis found T-cell receptor gene rearrangements and a germline immunoglobulin gene configuration in all four nodes. These paracortical clear cell nodules of clonal T cells may be a special type of PTCL. Alternatively, they may represent early foci of lymphoma or they may be a subgroup of T-zone lymphoma. Paracortical nodular PTCL must be differentiated from atypical hyperplastic lesions and some B-cell lymphomas.

Expression of interleukin-6 in Castleman's disease

Although mixed forms of Castleman's disease (CD) may occur, two classically recognized forms are the angiofollicular (hyaline vascular [V]) variant and the plasma cell (P) variant. The two forms of CD differ greatly in their clinical and histopathologic manifestations. Plasma cell CD is characterized by the presence of hyperplastic germinal centers (GCs) and sheets of plasma cells in the interfollicular areas. In this study we demonstrated an abundant expression of interleukin-6 (IL-6) in most GC B cells and in the numerous immunoblastoid B cells in the mantle zone and interfollicular areas in CD-P. Patients with CD-P also have an elevated serum IL-6 level. The increased IL-6 production is responsible for the marked plasma cell infiltration in lymph nodes and bone marrow as well as for the elevated gammaglobulin level in serum. In contrast, CD-V is distinguished by the presence of atrophic GCs, which often are populated by cytologically atypical follicular dendritic reticulum (FDR) cells, as well as by sheets of T-zone plasmacytoid histiocytes and increased numbers of capillaries in the interfollicular areas. In contrast to the findings in CD-P, we did not observe significant expression of IL-6 in GC cells or in immunoblastoid cells in CD-V; this may account for the paucity of plasma cells in this form of CD. The reason for the atypical changes in FDR cells as well as the increases in T-zone plasmacytoid histiocytes and capillaries seen in CD-V are not known inasmuch as no cytokines, such as IL-1, IL-4, IL-6, IL-7, IL-8, IL-9, tumor necrosis factor-α, granulocyte-macrophage colony-stimulating factor, or granulocyte colony-stimulating factor, were detectable in tissues. It is possible that in CD-V the atypical change in FDR cells could lead to a disturbance of B-lymphocyte/FDR cell interaction and subsequently to poor development of GCs. The study clearly indicates that the histopathologic and clinical features of CD vary greatly depending on the capacity of activated B cells to produce IL-6. However, lack of IL-6 secretion by GC cells alone cannot explain the histopathologic alterations in CD-V.

Recombinant gp120 specifically enhances tumor necrosis factor-alpha production and Ig secretion in B lymphocytes from HIV-infected individuals but not from seronegative donors.

The effect of recombinant protein from the envelope (gp120) of the HIV on B lymphocytes purified from either HIV-infected individuals or healthy seronegative controls was examined. B cells from peripheral blood and lymph nodes of HIV-infected individuals spontaneously secreted TNF-alpha; this secretion was augmented by the presence of gp120, whereas B cells from healthy seronegative donors failed to secrete significant levels of TNF-alpha in the presence or absence of gp120. In a coculture system of B cells and chronically HIV-infected T cells (ACH-2), where viral expression is largely mediated by TNF-alpha, gp120 increased virus expression only if the B cells were obtained from HIV-infected individuals. The effects of gp120 on viral expression in this system were not mediated via CD4 receptor binding or FcR binding of anti gp120-gp120 immune complexes. Besides its effect on cytokine production, gp120 also stimulated Ig secretion in B cells from HIV-infected individuals, but not from normal donors. Finally, it was demonstrated by in situ hybridization that germinal centers of lymph nodes from HIV-infected individuals contain large amounts of HIV RNA that is in close proximity to germinal center B cells. These findings suggest that the hyperplastic germinal centers of lymph nodes provide an unique environment for virus expression and accumulation where gp120 stimulates B cells to secrete HIV inductive cytokines, such as IL-6 and TNF-alpha, and thereby further enhances virus expression in infected cells in a paracrine manner.