Microarray analysis of gene expression by microdissected epidermis and dermis in mycosis fungoides and adult T-cell leukemia/lymphoma.

The characteristic histopathological feature of mycosis fungoides (MF) and adult T-cell leukemia/lymphoma (ATLL) is epidermotropism. To identify the mechanism for epidermotropism of lymphoma cells, total RNAs were obtained from skin biopsies of epidermis and dermis of MF and ATLL patients by means of laser capture microdissection, and used for subsequent complementary DNA (cDNA) microarray experiments. This procedure has made it possible for us to observe and evaluate the regional environment of MF and ATLL. Hierarchical cluster analysis revealed that the cDNAs could be clearly differentiated into MF and ATLL. CCL27 was expressed in the dermis generated from keratinocytes, CCR4/CCR6/CCR7/CCR10/cutaneous lymphocyte-associated antigen (CLA) lymphoma cells in the dermis, and CCL21 in the extracellular matrix (stroma). Lymphotoxin (LT) ß and CCL21 expression was significantly higher and that of CCR10 relatively for MF, while CCR4 and CLA expression was relatively higher for ATLL. In the epithelium, keratinocytes expressed CCL20/CCL27, and lymphoma cells CCR4/CCR6/CCR10, while CCR4, CCR6, CCL20 and CCL27 expression was relatively higher for ATLL than MF. The dermis of MF, but not that of ATLL, showed correlation between CCR7 and CCL21. These findings support the suggestion that chemokines and chemokine receptors are involved in the pathogenesis of MF and ATLL, indicate that cutaneous homing seems to be different for MF and ATLL, and point to the possibility that cutaneous T-cell lymphomas originate in regulatory T cells, especially in the case of ATLL.

Interaction of plectin and intermediate filaments.

BACKGROUND: Plectin, a member of the plakin family proteins, is a high molecular weight protein that is ubiquitously expressed. It acts as a cytolinker for the three major components of the cyotoskeleton, namely actin microfilaments, microtubules and intermediate filaments. OBJECTIVE: The aim of our experiments was to identify new binding sites for intermediate filaments on plectin and to specify these sites. METHODS: We introduced truncated forms of plectin into several cell lines and observe interaction between plectin and intermediate filaments. RESULTS: We found that a linker region in the COOH-terminal end of plectin was required for the association of the protein with intermediate filaments. In addition, we also demonstrated that a serine residue at position 4645 of plectin may have a role on binding of plectin to intermediate filaments. CONCLUSION: A linker region in the COOH-terminal end and serine residue at position 4645 may be important for the binding of plectin to intermediate filaments.

Clinicopathological features and prognostic significance of CXCL12 in blastic plasmacytoid dendritic cell neoplasm.

BACKGROUND: Blastic plasmacytoid dendritic cell neoplasm (BPDC) is a rare hematologic neoplasm, which almost always involves the skin and shows poor prognosis. OBJECTIVE: The aim of our study was to enhance BPDC diagnosis and indications for prognosis. METHODS: This study involved 26 patients with BPDC. To investigate the histogenesis of BPDC, we reviewed the clinical features and stained markers of various hematopoietic lineages, chemokines, and their receptors. RESULTS: Bone-marrow infiltration was detected in 13 of the 19 cases examined and leukemic changes in 18. Complete remission was achieved in 14 cases, but more than half of the patients showed recurrence within a short time, and 14 patients died of the disease after 1 to 25 months (mean 8.5 months). Positivity for CD123 was detected in 18 of 24 cases and for T-cell leukemia 1 in 18 of 22 cases. Of the chemokines and their receptors, 8 of 15 skin biopsy specimens proved to be positive for CXCL12. Leukemic change subsequent to skin lesions occurred in 7 of 8 CXCL12-positive cases (87.5%) and in 3 of 6 CXCL12-negative cases (50%). Seven of the 8 CXCL12-positive patients (87.5%) and two of the 6 CXCL12-negative patients (33.3%) have died, whereas one of 8 CXCL12-positive patients (12.5%) and 4 of 6 CXCL12-negative patients (66.7%) remain alive. LIMITATIONS: The number of patients was limited. CONCLUSIONS: We speculate that the presence of CXCL12-positive cells in the skin may be associated with leukemic change and a poor prognosis.

Frequent expression of gp46 in adult T-cell leukemia/lymphoma: an immunohistochemical study of 40 cases.

Adult T-cell leukemia/lymphoma (ATLL) is a lymphoproliferative disease caused by human T-cell lymphotropic virus type 1 (HTLV-1) infection. HTLV-1 is spread by cell-to-cell transmission via the gp46-197 region, from Asp197 to Leu216, in the envelope protein gp46. A correlation exists between the prevalence and titer of the antibody recognizing the gp46-197 region (anti-gp46-197 antibody) and the severity of ATLL. In the present study, immunohistochemical staining was performed on samples of paraffin embedded lymph nodes of three different histological types of ATLL (anaplastic large cell type, n = 10; pleomorphic type, n = 10; and Hodgkin's-like type, n = 10) from 30 cases and 10 cases of HTLV-associated lymphadenitis. Of the three ATLL subtypes, gp46 expression was highest in the anaplastic large cell type, followed by the pleomorphic type and Hodgkin's-like type (mean: 53.4%, 34.9% and 16.0%, respectively; P= 0.0003). In HTLV-1 associated lymphadenitis cases, gp46 positive cells were rarely seen (4.0%). These results suggest that gp46-197 immunohistochemical staining can be a useful histological indicator for prediction of the aggressiveness of ATLL and prognosis for ATLL patients.

Regression of rectal mucosa-associated lymphoid tissue (MALT) lymphoma after antibiotic treatments.

Only a few reports have described regression of rectal mucosa-associated lymphoid tissue (MALT) lymphoma after antibiotic treatment are generally found to be successful for gastric tumors. We examined eight rectal MALT lymphomas treated with antibiotic treatments to determine whether they regressed after treatment. We also discuss the relationship between rectal MALT lymphomas and MALT1 gene genetic abnormalities. Eight patients who had undergone antibiotic treatments were followed up with colonoscopy after initiation of the treatment. In five of the eight cases (63%) endoscopic examination showed that the rectal tumor had disappeared, which was confirmed histologically. Polymerase chain reaction for immunoglobulin heavy chain identified a monoclonal band in seven of eight cases (88%). Of the eight cases analyzed with fluorescence in situ hybridization (FISH) for MALT1 translocation, two demonstrated MALT1 gene genetic abnormality. These cases tended to be resistant to antibiotic treatment. Investigation and analysis of a large number of rectal MALT lymphomas are needed to establish suitable standards for antibiotic treatment.

Ratio of M2 macrophage expression is closely associated with poor prognosis for Angioimmunoblastic T-cell lymphoma (AITL).

Angioimmunoblastic T-cell lymphoma (AITL) is a peripheral T-cell lymphoma characterized by systemic disease with polymorphous infiltrate including macrophages. Although many studies of tumor-associated macrophage (TAM) populations in various malignant tumors have been published, only a few have dealt with activation of macrophage phenotypes such as M1 and M2 in tumor tissue. Because M2 macrophages highly express CD163, we suspected that CD163 may be a useful marker for identification of activation of macrophage phenotypes in AITL. We performed a retrospective study of immunohistochemical expression using two markers for macrophages [CD68 (PG-M1), CD163] and of the correlation of these expressions with overall survival of 42 AITL patients. The number of CD68-positive cells in AITL tissues did not correlate with overall survival (P= 0.59), whereas the number of CD163-positive cells and overall survival correlated to some extent (P= 0.08). Meanwhile, a higher ratio of CD163-positive to CD68-positive cells in AITL significantly correlated with worse overall survival (P= 0.036). Considering that this ratio reflects the proportion of macrophages polarized to the M2 phenotype, our findings indicate that activation of macrophages towards the M2 phenotype correlates with worse prognosis. Our findings indicate that the ratio of M2 macrophages expressed may be a useful marker for prognosis of AITL.

Mantle cell lymphoma shows three morphological evolutions of classical, intermediate, and aggressive forms, which occur in parallel with increased labeling index of cyclin D1 and Ki-67.

Although the 2008 World Health Organization classification defines two subtypes of mantle cell lymphoma (MCL), classical and aggressive, we often encounter MCL with both features in the same site. We named this feature "MCL with focal aggressive form (intermediate MCL)". In the present study, we reclassified 237 patients with cyclin D1 (CCND1)-positive MCL on the basis of the concept of intermediate MCL, and analyzed the correlation of this reclassification with immunohistochemical detection of CCND1, Ki-67, p53, p27(Kip1), and p21(WAF/Cip1). The median overall survival was 77, 31, and 18 months for classical, intermediate, and aggressive MCL, respectively, showing a statistically significant difference (P < 0.0001). The expression levels of CCND1, Ki-67, p53, and p21(WAF/Cip1) in aggressive MCL (mean 80.1 +/- 27.8%, 73.7 +/- 28.9%, 31.0 +/- 69.0%, and 10.4 +/- 24.8%, respectively) were higher than those in classical MCL (mean 58.1 +/- 36.7%, 25.2 +/- 25.5%, 6.5 +/- 24.3%, and 2.5 +/- 13.0%, respectively) and intermediate MCL (mean 75.7 +/- 31.4%, 30.8 +/- 33.3%, 21.0 +/- 57.4%, and 4.8 +/- 16.5%, respectively). Significantly different levels of Ki-67 and p21(WAF/Cip1) were only recognized between intermediate and aggressive (P < 0.05 and P < 0.0001, respectively), whereas those of CCND1 and p53 were only between classical and intermediate (P < 0.0001 and P < 0.05, respectively). There were no significant differences in p27(Kip1) among the three groups. The subsequent discriminant analysis with independent prognostic factors clearly demonstrated that the morphological evolution of MCL occurs in parallel with increased labeling index of CCND1 and Ki-67. The diagnosis of intermediate MCL thus proved to be of major significance and should enable the design of more tailored therapies.

Analysis of CXCL9 and CXCR3 expression in a case of intravascular large B-cell lymphoma.

Intravascular large B-cell lymphoma is a rare disease with multiorgan involvement that also affects the skin. Skin manifestations include purpuric to red macules, plaques, or nodules with occasional edema and tenderness. We report a 68-year-old woman with bilateral leg edema and occasional high fever. A biopsy specimen from a subcutaneous nodule showed that the blood vessels in the dermis and subcutaneous tissue were filled with irregularly shaped chromatin-rich large atypical lymphocytes positive for CD20 and bcl-2, consistent with the diagnosis of intravascular large B-cell lymphoma. In addition, immunohistochemical analysis showed expression of CXCR3 in the atypical lymphocytes; its ligand, CXCL9, was detected in blood vessels. Although limited to a single case, our study could provide a possible new clue to the pathogenesis of intravascular large B-cell lymphoma by virtue of the characteristic expression of CXCL9-CXCR3.

Hepatocellular apoptosis associated with cytotoxic T/natural killer-cell infiltration in chronic active EBV infection.

The aim of the present study was to identify the mechanism of hepatocellular apoptosis induced by EBV-infected cytotoxic T/natural killer (NK) cells in chronic active EBV infection (CAEBV). Eight patients with CAEBV were studied, and infected T-cell expansion and NK-cell expansion were detected in four patients each. Biopsy or necropsy was performed on lymph node, liver, or spleen, and each specimen was subjected to immunohistochemical double staining of CD3 plus caspase-3 with the addition of cytotoxic markers of T-cell restricted intracellular antigen-1 (TIA-1), perforin, and granzyme B, as well as EBV in situ hybridization (EBV-ISH). In the liver, some of the infiltrating CD3-positive lymphocytes stained positively for EBV-ISH and cytotoxic markers. Double staining of CD3 plus caspase-3 indicated caspase-3 positive hepatocytes with apoptotic features, accompanied by extensive infiltration of CD3-positive cells, which were directly attached to the apoptotic caspase-3 positive hepatocytes. In contrast, far fewer cells stained positive for caspase-3 in lymph node and spleen than in liver. The present findings suggest that in patients with CAEBV, cytotoxic T/NK cells may directly induce hepatocytes to undergo apoptosis more frequently than they do cells in other organs of the reticulo-endothelial system.

Downregulation of RCAS1 and upregulation of cytotoxic T cells affects synovial proliferation and apoptosis in rheumatoid arthritis.

OBJECTIVE: The main histological change in rheumatoid arthritis (RA) is the villous proliferation of synovial lining cells. This seems to be the result of the proliferation and apoptosis induced by immune balance. We studied the involvement of RCAS1 and the infiltration of cytotoxic T lymphocytes (CTL), and examined the synovium immunohistochemically to determine the involvement of proliferation and apoptosis in synovial lining cells, and their relationship with the activity of RATreg cells in the germinal center. METHODS: We used double-immunological staining of Ki-67 and caspase-3 to investigate proliferation and apoptosis. We analyzed CTL, regulatory T cells (Treg), and receptor-binding cancer antigen expressed on SiSo cells (RCAS1), recently recognized to play a role in immune evasion. Proliferation and apoptosis were more frequently encountered in synovial lining cells in RA than in those in osteoarthritis (OA) that were used as a control. RESULTS: High expression of RCAS1 was detected more frequently in the synovial lining cells of OA, but CTL infiltration into the synovium was rarely found. In RA, on the other hand, CTL were observed, while RCAS1 expression was lacking. We compared the presence of Foxp3-positive cells with the level of C-reactive protein (CRP) that served as an active inflammatory marker. Foxp3-positive cells in the germinal center and in CRP showed possible correlation in terms of the range of inflammatory states. CONCLUSION: In RA, the lack of RCAS1 is thought to induce CTL infiltration through loss of the ability to evade immune attack, thus leading to apoptosis of the synovial lining cells. In addition, Treg cells may play a role in the downregulation of activated T cells.

BCL6 gene amplification/3q27 gain is associated with unique clinicopathological characteristics among follicular lymphoma without BCL2 gene translocation.

Although approximately 10-20% cases of follicular lymphoma lack BCL2 gene rearrangement, there are few reports having described the alternative genetic aberrations and their association about clinicopathological features. In this study, analysis by Fluorescence in situ hybridization of BCL6 gene aberrations in 100 follicular lymphoma cases without IGH/BCL2 rearrangement resulted in the identification of four subgroups. Group I: BCL6 gene rearrangement (n=41); Group II: BCL6 gene amplification/3q27 gain (n=30); Group III: the absence of both (n=23); and Group IV: the presence of both (n=6). Group II showed higher grade morphology (Grade 3a/b: 93%), higher bcl2 and MUM1 expression (73 and 57%, respectively), and more frequent combination with BCL2 gene amplification/18q21 gain (90%) than the other groups. BCL6 gene aberration, especially amplification/3q27 gain, indicates the presence of certain morphological and phenotypical findings in follicular lymphoma cases without IGH/BCL2 rearrangement.

The relationship of FOXP3 expression and clinicopathological characteristics in adult T-cell leukemia/lymphoma.

Adult T-cell leukemia/lymphoma is an aggressive malignant disease associated with regulatory T cells as discussed in some recent reports. We analyzed the expression of FOXP3, a key molecule of regulatory T cells, in adult T-cell leukemia/lymphoma and its association with clinicopathological features. Of 169 adult T-cell leukemia/lymphoma cases examined, 60 (36%) showed FOXP3 expression in lymphoma cells. Morphologically, 22 cases were classified as anaplastic large cell variant and 147 as pleomorphic cell variant. Only 1 (5%) of the anaplastic large cell variant cases and 59/147 (40%) of the pleomorphic cell variant cases expressed FOXP3. Epstein-Barr virus-infected cells were significantly more frequently found in FOXP3(+) cases (23/60; 38%) than in FOXP3(-) cases (12/109; 11%) (P<0.0001). Cytogenetic analysis showed that FOXP3(+) cases had simpler chromosomal abnormalities than FOXP3(-) cases. Clinically, FOXP3(+) and FOXP3(-) cases did not differ significantly in age distribution, clinical stage, lactate dehydrogenase and calcium in serum and overall survival. However, 8 of 34 FOXP3(+) cases suffered a severe infectious state, an indication of immunosuppression, while only 2 of 62 FOXP3(-) cases did so (P<0.005). FOXP3 expression in adult T-cell leukemia/lymphoma thus reflects morphological features and is clinically and pathologically associated with an immunosuppressive state.

Distribution of malignant lymphoma in Japan: analysis of 2260 cases, 2001-2006.

The World Health Organization classification was used to conduct an analysis of geographic, age, sex, and lesion primarily biopsied/resected distribution of 2260 lymphoid neoplasms diagnosed during 2001-2006 throughout Japan. B-cell neoplasms accounted for 65% of all lymphoid neoplasms, T/natural killer (T/NK)-cell neoplasms for 25% and Hodgkin lymphoma for 7%. The most common type was diffuse large B-cell lymphoma (DLBCL, 33%), followed by follicular lymphoma (18%), and adult T-cell leukemia/lymphoma (ATLL, 10%). The high rate of 18% for follicular lymphoma was similar to that in Western countries (11-33%). T/NK-cell neoplasms accounted for a higher percentage of lymphoid neoplasms in Kyushu (30%) and Okinawa (38%) compared with other areas of Japan (18-20%). Among T/NK-cell neoplasms, ATLL was the most common type in Okinawa (54%) and Kyushu (59%). Extranodal NK/T cell lymphoma was the second most common type of T/NK-cell neoplasms in Okinawa (15%). This epidemiological study shows that the distribution patterns of malignant lymphoma differ especially in Kyushu and Okinawa, the endemic area of human T-cell leukemia/lymphoma virus type 1.