Kumiko Shimoyama

Activation of the Interferon Pathway in Peripheral Blood of Patients with Sjögren’s Syndrome

Objective. DNA microarray analysis and quantitative real-time polymerase chain reaction (PCR) were performed to identify key target genes in peripheral blood from patients with Sjögren’s syndrome (SS). Methods. DNA microarray analysis was performed in 19 patients with SS (all women) and 10 healthy controls (5 men and 5 women) using a low-density DNA microarray system with 778 genes. For confirmation, the expression of upregulated genes was analyzed by quantitative real-time PCR in another 37 SS patients (35 women and 2 men) and 9 healthy controls (8 women and 1 man). Relationships between gene signatures and various clinical measures, such as disease duration, symptoms and signs, complications, immunological findings, and salivary and lacrimal functions, were analyzed. Results. Interferon-α (IFN-α)-inducible protein 27 (IFI27) showed the most significant difference between SS patients and controls in the microarray screening. We performed quantitative RT-PCR for IFI27. IFI27 gene expression level was increased in patients with SS compared with controls (p < 0.01) by real-time PCR, supporting our observations from the microarray data. The level of IFI27 was significantly correlated with serum IgG levels (r = 0.462, p < 0.01) and ß2-microglobulin (r = 0.385, p < 0.05), soluble interleukin 2 receptor (r = 0.473, p < 0.01), erythrocyte sedimentation rate (r = 0.333, p < 0.05), and antinuclear antibody titer (speckled pattern; r = 0.445, p < 0.01). Conclusion. Our results suggest that upregulation of IFN-inducible genes in SS patients is a systemic phenomenon, and IFN may play an important role in the pathogenesis of SS. The expression level of IFI27 could be an effective and specific biomarker associated with SS.

Granulocytic sarcoma of megakaryoblastic differentiation in the lymph nodes terminating as acute megakaryoblastic leukemia in a case of chronic idiopathic myelofibrosis persisting for 16 years

Abstract: A 43‐yr‐old Japanese woman presented with mild anemia, leukocytosis and splenomegaly in May 1984. Splenomegaly and anemia gradually progressed. Sixteen years later, in October 2000, she developed inguinal lymphadenopathy. Biopsy of the lymph node revealed infiltration of blasts, megakaryocytes, fibroblasts and myeloid cells. Large blasts with basophilic cytoplasm with cytoplasmic projections appeared in the peripheral blood. These blasts were negative in peroxidase stain, positive in acid phosphatase and weakly positive in periodic acid‐Schiff stain. Immunohistochemical staining with monoclonal antibodies revealed that these blasts were positive with anti‐CD41 (glycoprotein IIb/IIIa) and negative with other monoclonal antibodies. So diagnosis of granulocytic sarcoma in megakaryoblastic transformation from chronic idiopathic myelofibrosis was made. A cytogenetic study revealed that bone marrow cells were 46,XX del(13)(q?) initially and additional abnormalities including der(5,5,11)(q11;q13)ins(5;?)(q11;?) were found when she developed megakaryoblastic transformation. Granulocytic sarcoma of megakaryoblastic transformation from chronic idiopathic myelofibrosis is a rare event. Immunophenotyping with monoclonal antibody for CD41(glycoprotein IIb/IIIa) confirmed the diagnosis.

Epstein-barr virus-associated composite lymphoma composed of peripheral t-cell lymphoma and an anaplastic variant of a diffuse large b-cell type of non-hodgkin’s lymphoma and strongly expressing P53 protein

We report a case of composite lymphoma consisting of peripheral T-cell lymphoma and an anaplastic variant of diffuse large B-cell lymphoma (DLBCL) and associated with Epstein-Barr virus (EBV) infection and strong p53 expression. A 65-year-old Japanese woman developed fever and generalized lymphadenopathy.A biopsy of the cervical node revealed the morphology of malignant lymphoma with 2 kinds of lymphoma coexisting in 1 lymph node. One lymphoma type consisted of immunoblastic large cells with the T-cell marker phenotype CD3+, CD45RO/UCHL-1+, CD20/L26-, CD79-, CD10-, CD30-, and CD15-; the other type consisted of large cells with abundant cytoplasm and pleomorphic nuclei with the marker phenotype CD79+, CD20/ L26+, CD45RO/UCHL-1-, CD3-, CD10-, CD30+, NPM/ALK-, and CD15-. Therefore, the diagnosis was composite lymphoma of peripheral T-cell lymphoma and an anaplastic variant of DLBCL, stage IVB, because the patient had bone marrow involvement with peripheral T-cell lymphoma. The biopsy led to findings of latent type II EBV-associated lymphoma in both the peripheral T-cell lymphoma and the anaplastic variant of DLBCL as the result of positive signals for EBV small RNAs by in situ hybridization, positive immunostaining results for EBV latent membrane protein 1 antibody, and negative immunostaining results for EBV nuclear antigen 2. Immunostaining of the mass with p53 antibody also yielded positive results for both types of lymphoma cells. This case suggests that the immunocompromised state of this patient with EBV-related peripheral T-cell lymphoma allowed the emergence of an EBV-related anaplastic variant of DLBCL and suggests a close relationship between p53 expression and latent EBV infection.

Association of Epstein-Barr virus with human immunodeficiency virus-negative peripheral T-cell lymphomas in Japan.

Abstract:  The association of Epstein–Barr virus (EBV) with human immunodeficiency virus‐negative T‐cell lymphoma was examined in 68 patients using the polymerase chain reaction (PCR) with DNA obtained from formalin‐fixed paraffin‐embedded tissues and an in situ hybridization technique. EBV‐encoded RNA (EBER) was detected in 43 of 68 cases (63%) of peripheral T‐cell lymphoma: in 100% (11 of 11 cases) of NK/T‐cell lymphomas, 70% (14 of 20 cases) of angioimmunoblastic T‐cell lymphomas (AILT) and 49% (18 of 37 cases) of other types of peripheral T‐cell lymphoma. A positive band was also detected at high incidence (36 of 65 cases; 55%) in a PCR analysis using primers to detect the Bam HI‐W fragment of EBV. In the immunohistochemical analysis using a monoclonal antibody to latent membrane protein 1 (LMP‐1) of EBV, one of the EBV‐encoded latent gene products, LMP‐1, was found to be expressed in 13 of 64 cases (20%), but EBNA‐2 was not expressed in all the cases examined (0 of 59 cases; 0%). The 5‐yr survival rate was 28% for peripheral T‐cell lymphomas overall, 0% for NK/T‐cell lymphomas, 38% for AILTs and 28% for other types of peripheral T‐cell lymphoma. The difference in the overall survival rate between NK/T‐cell lymphoma and non‐NK/T‐cell lymphoma was significant (P = 0.0498 by Log‐rank test). Among peripheral T‐cell lymphoma patients overall, the group severely infected with EBV (EBER‐ISH ++) had a lower 5‐yr survival rate (8%) than the group slightly (EBER‐ISH +) or not infected (38%; P = 0.0013).

Incidence of Diffuse Large B-Cell Lymphoma of Germinal Center B-Cell Origin in Whole Diffuse Large B-Cell Lymphoma : Tissue Fluorescence In Situ Hybridization Using t(14 ; 18) Compared with Immunohistochemistry

Diffuse large B-cell lymphoma (DLBCL) can be divided into prognostically important categories such as germinal center B (GCB)-like and non-GCB-like groups. The t(14;18)(q32;q21) translocation defines a unique subset of DLBCL cases with a GCB gene expression profile. Two-color fluorescence in situ hybridization (FISH) analysis was applied to detect t(14;18) (q32;q21) in the nuclei of paraffin-embedded tissue sections from 61 patients with de novo DLBCL.Nine (15%) of 61 cases had a positive pattern. Fifty-seven cases were subclassified in an immunohistochemical study with anti-CD10, anti-bcl-6, and anti-MUM1 antibodies. In this classification, 21 cases (37%) were placed in the GCB group, and 36 (63%) were placed in the non-GCB group. There was a discrepancy between t(14;18) occurrence and bcl-2 protein expression. Bcl-2 protein expression was positive in 40 (67%) of 60 cases.The expression of bcl-2 protein in the GCB and non-GCB groups was not significantly different: 15 (71%) of 21 cases in the GCB group and 24 (67%) of 36 cases in the non-GCB group tested positive.We found no difference between the FISH-positive and FISH-negative groups in overall survival time (P = .6019, log-rank test).The overall survival rates of GCB and non-GCB groups did not differ significantly by immunohistochemical classification (P = .5399, log-rank test). Overall survival was significantly longer in the group with a low International Prognostic Index (IPI) score than in the group with a high IPI score (P = .0002, log-rank test).Our results suggest that immunohistochemical study and cytogenetic study with t(14;18) FISH cannot predict the clinical outcomes of DLBCL patients.Astudy with a larger number of patients may show a difference in clinical outcomes between FISH-positive and FISH-negative groups and between GCB and non-GCB groups.

難治性関節リウマチ（RA）に対する白血球除去療法（LCAP）の臨床的有用性の検討

PURPOSE To examine therapeutic effect of leukocytapheresis (LCAP) for rheumatoid arthritis (RA) resistant to various treatments. METHOD Thirteen patients with RA (mean age : 60.8+/-11.4, male : female=5 : 8), 1 who were resistant to disease-modifying anti-rheumatic drugs (DMARDs) and biologics, or 2 who failed with those medicines because of side effects or complications. We performed LCAP, which was carried out once a week for a total of five sessions, with a throughput of about 0.1 L/kg. Before and after LCAP, we evaluated the effect of LCAP therapy. RESULT DAS28 (CRP) score was 5.70+/-1.12 before LCAP, 4.57+/-1.19 (P<0.05) just after the final LCAP and 4.83+/-1.35 (P<0.05) about 4 weeks after LCAP. DAS28 score decreased in all patients after LCAP. No serious adverse events were observed except temporary anemia. CONCLUSION LCAP therapy may be useful and safe for patients with RA resistant to conventional medication. Patients who show good clinical response by LCAP needs to be clarified.

Epstein-Barr virus-associated anaplastic large cell variant of diffuse large B-cell-type non-Hodgkin's lymphoma with concurrent p53 protein expression.

In the new World Health Organization (WHO) classification of malignant lymphoma, anaplastic large cell lymphoma of B-cell phenotype is classified either as the anaplastic large cell variant of diffuse large B-cell lymphoma or as Hodgkin’s lymphoma. A 71-year-old Japanese man developed fever and generalized lymphadenopathy. Biopsy of the right axillary node revealed morphology of malignant lymphoma in which large cells with abundant cytoplasm and pleomorphic nuclei were scattered among small lymphocytes. Immunostaining with various monoclonal antibodies revealed the large cells to be CD79+, CD20/L26+, CD45RO/UCHL-1-, CD3-, CDl0-, CD30+, NPM/ALK-, EMA-, CD15-, and bcl-2-. Amplification of the J region of the immunoglobulin heavy chain by polymerase chain reaction revealed a single rearranged band. Therefore the diagnosis of anaplastic large cell variant of diffuse large B-cell lymphoma, stage IIIB, was made from the standpoint of the new WHO classification of malignant lymphoma. Biopsy led to findings of Epstein-Barr virus (EBV)-associated lymphoma with positive in situ hybridization results for EBV small RNAs, positive results of immunostaining with EBV latent membrane 1 antibody, and negative results of immunostaining with Epstein-Barr nuclear antigen 2. Results of immunostaining of the mass with p53 antibody also were positive for lymphoma cells. The findings in this case may suggest a close relationship between p53 expression and latent EBV infection.

リウマチ膠原病診療における抗環状シトルリン化ペプチド抗体（抗CCP抗体）の臨床的有用性の検討

OBJECTIVE To examine clinical significance of anti-cyclic citrullinated peptide antibody (anti-CCP antibody) in RA. METHODS Hundred fifteen patients with polyarthralgia (89 females, 26 males) were recruited, and subjected for the study. We studied anti-CCP antibody, ESR, CRP, IgM-RF, IgG-RF, RAPA, MMP-3, CARF, C1q-IC, Stage, Class, Joint score, Sharp score, KL-6, SP-D, chest CT. RESULTS Anti-CCP antibody test had high specificity (93.5%). In RA with positive anti-CCP antibody, Sharp score (10.9+/-22.4) was higher than those with negative anti-CCP (1.7+/-1.8), and may serve as a prognostic marker of joint destruction (P<0.05). Anti-CCP antibody in RA with interstitial pneumonia is higher (84.5+/-36.4 U/mL) than those without interstitial pneumonia (52.6+/-44.7 U/mL) (P<0.05). CONCLUSION Anti-CCP antibody is useful for diagnosis of RA, and could be a specific marker of joint destruction. Further investigation is necessary to clarify the relation of anti-CCP antibody with organ involvement and activity of RA.

Immunophenotypic Analysis of Various Blastic Crises in Chronic Myeloid Leukemia: Correlations between CD7 Expression and Response to Chemotherapy

Chronic myeloid leukemia (CML), a stem cell disorder characterized by the presence of the Philadelphia chromosome (Ph), invariably terminates in blast crisis. Various studies have illustrated that all hematopoietic cell lineages may be involved in the CML blast crisis and also that both hybrid transformation and lineage switches can occur [1]. We therefore performed immunophenotypic marker analysis of 14 patients in CML blast crisis. CD7 was expressed in all patients in myeloid crisis and in 1 T-lymphoid crisis patient, as well as in 1 patient in biphenotypic crisis and 1 in megakaryoblast crisis. The mean duration of blast crisis was 11.2 months. The survival rate of CD7+ patients was lower than that of CD7– patients (P = .0046 by log-rank test). This result may suggest a poorer response to chemotherapy in CD7+ patients. Among our 42 patients with CML diagnosed from 1979 to 2002 from positive results for t(9; 22)(q34;q11) and fluorescence in situ hybridization analysis for bcr and abl genes, 14 patients developed blast crises of various cell lineages. These 14 patients (8 male and 6 female), aged 29 to 76 years at the time of blast crisis, were studied. Peripheral blood and bone marrow mononuclear cells were obtained by Ficoll-Conray density gradient centrifugation. Cytospin samples from the peripheral blood and bone marrow were tested with a panel of monoclonal antibodies using the alkaline phosphatase and monoclonal anti–alkaline phosphatase (APAAP) method. Flow cytometric analysis was performed according to the method previously described [2]. Immunophenotypic Analysis of Various Blastic Crises in Chronic Myeloid Leukemia: Correlations between CD7 Expression and Response to Chemotherapy