Comparative studies in induction of micronuclei by three genetically recombinant and urinary human erythropoietins

Comparative studies on the induction of micronucleated erythrocytesby three recombinant human erythropoietins (rhEPOs) such asepoetin     

Immunosuppressive dose of azathioprine inhibits replication of human cytomegalovirus in vitro

Summary Azathioprine (Aza) was found to have anti-human cytomegalovirus (HCMV) activity in vitro at concentrations used for immunosuppression therapy. The dose of Aza for 50% plaque reduction was 0.592µg/ml for HCMV in human embryonic lung (HEL) cells, but those of Aza for 50% plaque reduction for herpes simplex virus (HSV) and varicella-zoster virus were more than 20µg/ml. The dose of Aza for 50% reduction of the HCMV yield in infected cells was 0.25µg/ml, while that for 50% reduction of the HSV yield in infected cells was more than 50µg/ml. The dose of Aza for 50% growth inhibition of HEL cells was 30µg/ml, and 50.7 and 120 times greater than the doses for 50% reduction of the plaque formation and the yield of HCMV, respectively. Thus Aza was found to have a strong anti-HCMV activity at concentrations used for immunosuppression. When HCMV infected cells were treated with cyclosporine (CsA: 0.2µg/ml) and prednisolone (Pred: 0.3µg/ml) simultaneously with Aza, the doses of Aza for 50% reduction of plaque formation and the yield of HCMV were 0.73 and 0.32µg/ml, respectively. Thus an inhibitory effect of Aza was also observed in HCMV-infected cells treated with CsA and Pred at their concentrations used for immunosuppression. Maintenance of an anti-HCMV dose of Aza in combination with CsA and Pred might establish not only satisfactory immunosuppression but also suppression of HCMV infection in transplant recipients.     

An HLA-binding-motif-aided peptide epitope library: A novel library design for the screening of HLA-DR4-restricted antigenic peptides recognized by CD4+T cells

Susceptibility to a series of autoimmune diseases is strongly associated with particular HLA class II alleles. Identification of T cell clones and antigenic epitopes bound by HLA class II molecules involved in autoimmune diseases is critical to understanding the etiology of these HLA class II-associated diseases. However, establishment of T cell clones in autoimmune diseases is difficult because the antigenic peptides are unknown. Peptide library methods which include all possible peptide sequences offer a potentially powerful tool for the detection of cross-reactive antigenic peptides recognized by T cells. Here, we reduced the number of peptides per mixture by utilizing the known binding motifs of peptides for the HLA-DRB1*0405 molecule and evaluated the effectiveness of this library design. Each library mixture evoked a strong proliferative response in the unprimed peripheral blood lymphocytes (PBL) from HLA-DRB1*0405-positive donors but little or no response in the PBL from HLA-DRB1*0405-negative donors. The library also detected antigenic peptides that activated three antigen-specific T cell lines restricted by HLA-DRB1*0405, with different specificities. The motif-based approach thus presents a powerful method for monitoring T cells in large, heterogeneous T cell populations and is useful for the identification of the mimic peptide epitopes of T cell lines and clones. Received: October 3, 1997 / Accepted: October 23, 1997     

Identification of an HLA-DQ6-derived peptide recognized by mouse MHC class I H-2Db-restricted CD8+ T cells in HLA-DQ6 transgenic mice

Summary CD8⁺ T cells from C57BL/6(B6) mice show cytotoxicity to B cell blasts prepared from syngeneic transgenic mice expressing HLA-DQ6 molecules in a mouse MHC class I H-2D^b restricted manner. Although these results suggest that CD8⁺ T cells recognize peptides derived from DQ6 molecule bound to H-2D^b on target cells, no direct evidence so far has been obtained. To clarify this, we synthesized 23 peptides corresponding to DQ6α orβ chain and carrying the motifs of D^b-binding peptides, and examined their capacity to induce cytotoxicity in the CD8⁺ T cell line. We show here that DQA1-2, one of these peptides, induced cytotoxicity of the CD8⁺ T cells when this peptide was pulsed to H-2D^b expressing target cells, as efficiently as HLA-DQ6 expressing target cells did. Thus, our results suggest that DQA1-2 can be naturally processed from DQ6 molecules and recognized by the CD8⁺ T cells in the context of H-2D^b molecules. These results suggest that allogeneic HLA class II molecules are involved in the rejection not only as the ligand for T cell receptor of alloreactive CD4⁺ T cells but also as self-peptides bound to HLA class I molecules recognized by CD8⁺ T cells.     

Extrarenal Wilms' tumor in the adult patient. A case report and review of the world literature

A case of an extrarenal Wilms' tumor arising in the retroperitoneal region of a 49-year-old male is reported. A review of the world literature indicates that the incidence of the tumor arising in the extrarenal region is extremely rare. A total of 14 cases have previously been reported, but the number of cases that occurred in adult patients is only 2. The clinical and pathologic features are briefly discussed.     

BRONCHIOLO-ALVEOLAR ADENOCARCINOMA WITH MULTIPLE CYSTS

An autopsy case of bronchiolo-alveolar adenocarcinoma in the lung is reported. The patient is a 70-year-old male who complained of severe cough with 500–600 ml watery sputum a day, loss of weight, and general fatigue. Autopsy revealed numerous whitish tumors in various sizes with multiple cysts in both lungs, with no metastasis being found in any other organs. Histological findings identified the tumor as a bronchiolo-alveolar adenocarcinoma originating from the lungs. Electron-microscopic findings showed that the tumor cells were covered by prominent microvilli, and contained abundant irregulary-shaped cytoplasmic vacuoles suggestive of mucin.     

Establishment of an Animal Model Using Recombinant NOD.B10.D2 Mice To Study Initial Adhesion of Oral Streptococci

An oral biofilm is a community of surface-attached microorganisms that coats the oral cavity, including the teeth, and provides a protective reservoir for oral microbial pathogens, which are the primary cause of persistent and chronic infectious diseases in patients with dry mouth or Sjögren''s syndrome (SS). The purpose of this study was to establish an animal model for studying the initial adhesion of oral streptococci that cause biofilm formation in patients with dry mouth and SS in an attempt to decrease the influence of cariogenic organisms and their substrates. In nonobese diabetogenic (NOD) mice that spontaneously develop insulin-dependent diabetes mellitus (IDDM) and SS, we replaced major histocompatibility complex (MHC) class II (A^g7 E^g7) and class I D^b with MHC class II (A^d E^d) and class I D^d from nondiabetic B10.D2 mice to produce an animal model that inhibited IDDM without affecting SS. The adhesion of oral streptococci, including Streptococcus mutans, onto tooth surfaces was then investigated and quantified in homologous recombinant N5 (NOD.B10.D2) and N9 (NOD.B10.D2) mice. We found that a higher number of oral streptococci adhered to the tooth surfaces of N5 (NOD.B10.D2) and N9 (NOD.B10.D2) mice than to those of the control C57BL/6 and B10.D2 mice. On the basis of our observation, we concluded that these mouse models might be useful as animal models of dry mouth and SS for in vivo biological studies of oral biofilm formation on the tooth surfaces.Oral streptococci are present in large numbers in dental plaque, and several types interact with the enamel salivary pellicle to form a biofilm on tooth surfaces (9, 16, 17, 21, 29). Streptococci account for approximately 20% of the total number of salivary bacteria (24), with Streptococcus salivarius being the primary organism. Further, the densities of Streptococcus mutans and Streptococcus sanguis in saliva are more than 1 × 10⁵ cells per ml. S. mutans is a pioneering organism that plays an important role in biofilm formation on tooth surfaces and is a primary causative agent of dental caries (9, 16, 21). The mechanical forces of salivary flow and tongue movement tend to dislodge and expel bacteria from tooth surfaces and the oral cavity (3, 5, 6), and their importance in controlling microbial colonization in the oral cavity has been well demonstrated in individuals with diabetes mellitus, Sjögren''s syndrome (SS), and dry mouth, who suffer from a rapid overgrowth of biofilm and rampant caries, making them highly susceptible to oral infections (1-2, 6). Thus, attempts to investigate the initial adhesion by oral streptococci, including S. mutans, in mouse models are likely to aid in the understanding and prevention of oral infectious diseases caused by the components of oral biofilm.Previous studies of S. mutans infections in the oral cavities of mice have been performed by feeding the animals diets containing sucrose in the presence of glucans (13, 15, 30, 43). Since the adherence of S. mutans to the tooth surface may depend on the balance between physical adherence and synthesis of insoluble glucans in a natural environment, that infection method may be inappropriate for investigation of natural biofilm formation associated with streptococci, including S. mutans (18, 39).The nonobese diabetogenic (NOD) mouse strain is currently the best available model for the study of insulin-dependent type 1 diabetes mellitus (IDDM) and SS (11, 31), both of which develop spontaneously and are characterized by lymphatic infiltration of the pancreas and salivary glands. Oral changes are prominent features of these diseases, which are manifested by dry mouth and hyposalivation (6, 7, 37). NOD mice are also used as an animal model for the study of oral infectious diseases associated with systemic diseases such as diabetes and SS or dry mouth.The unique major histocompatibility complex (MHC) class II genes (I-A^g7, no expression of I-E) represent dominant susceptibility factors and mediate activated T cells during the development of diabetes in NOD mice (11, 22, 25, 36, 41, 42). In the NOD model of SS, histopathological analyses of the salivary glands in MHC-congenic strains of NOD mice have indicated that the I-A^g7 region is not required for lymphocytic infiltration (26, 31). Further, replacement of the NOD MHC class I K^d region with another haplotype, MHC class I K^wm7, as well as replacement of the MHC class II A^g7 E^g7 and class I D^d regions with the corresponding region from the other MHC haplotype, has been shown to prevent diabetes (12). However, replacement with MHC class I K does not completely prevent development of insulitis. In another report, NOD mice pretreated nasally by using peptides restricted with MHC class I K^d showed a delayed onset of spontaneous IDDM, though insulitis could not be prevented by the induction of tolerance (23).In the present study, we attempted to establish an animal model for oral infectious diseases such as dental caries by focusing on replacement of the MHC class II and class I D region but not the class I K region in nondiabetic NOD mice by outcrossing B10.D2 mice (K^d, I-A^d, and D^d) with NOD mice (K^d, I-A^g7, and D^b) because the MHC class I K region in B10.D2 mice is identical with that in NOD mice (12). The present backcrossed and intercrossed NOD mice with the MHC class II and MHC class I D region replaced with that from B10.D2 mice developed SS, however, not diabetes. We then attempted to determine whether these mice would be useful as animal models for a sucrose-free study of the initial adhesion of oral streptococci on tooth surfaces in humans.     

Mold filling of titanium alloys in two different wedge-shaped molds

Pure titanium and titanium alloys are potential materials for the fabrication of cast dental appliances. One important factor in producing sound castings is the capacity of the metal to fill the mold. This study used a wedge-shaped mold to compare the mold filling of titanium with that of conventional dental casting alloys. The metals used were CP Ti, Ti-6Al-7Nb, Ti-6Al-4V, Ti with 1 and 4wt% Cu and ADA Type III gold alloy and an Ni-Cr alloy. The castings were cut into four pieces parallel to the triangular surface. Mold filling was evaluated as the distance between the tip of the cast wedge and theoretical tip of the triangle. The mold filling of the gold alloy was superior compared to all the metals tested, while the mold filling of the Ni-Cr alloy was the worst. There were no statistical differences at the 30 degrees marginal angle for all the cast titanium metals. At the sharper 15 degrees angle, CP Ti and Ti-6Al-7Nb was superior to both the Ti-Cu alloys. Although the mold filling of titanium was inferior compared to the gold alloy, the data justify the use of titanium for the production of dental appliances.     

Expression of BAFF-R (BR3) in normal and neoplastic lymphoid tissues characterized with a newly developed monoclonal antibody

BAFF-receptor (BAFF-R) is required for the successful maturation and survival of B-cells. We developed an anti-human BAFF-R monoclonal antibody (mAb), 8A7. The reactivity of 8A7 in normal and neoplastic tissue was examined by performing immunohistochemistry on paraffin-embedded sections. 8A7 reacted with lymphocytes in the mantle and marginal zones, but not with lymphocytes in the interfollicular area. Lymphocytes in the germinal centers were found to be negative or occasionally weakly positive for 8A7. BAFF-R expression was found only in B-cell lymphoma (44/80, positive cases/examined cases): B-lymphoblastic lymphoma 0/3, B-chronic lymphocytic leukemia/small lymphocytic lymphoma 4/4, mantle cell lymphoma 9/11, follicular lymphoma 10/14, diffuse large B-cell lymphoma (DLBCL) 11/25, marginal zone B-cell lymphoma 8/10, lymphoplasmacytic lymphoma 2/2, plasma cell myeloma 0/2, and Burkitt lymphoma 0/9, but not in T/NK cell lymphomas (0/19) or Hodgkin lymphoma (0/10). BAFF-R was expressed in most low-grade B-cell neoplasms and a small number of DLBCL, suggesting that BAFF-R may play an important role in the proliferation of neoplastic lymphoid cells. Thus, the mAb is very useful for further understanding of both healthy B-cell biology and its pathogenic neoplasms.     

Women with endometriosis have increased levels of placental growth factor in the peritoneal fluid compared with women with cystadenomas

BACKGROUND: To assess the release of placental growth factor (PlGF) into peritoneal fluid in women with and without endometriosis, we measured its concentration with reference to disease stage, the presence of red endometriotic lesions and the phase of menstrual cycle. METHODS: Surgery was scheduled in the proliferative or secretory phase of the menstrual cycle for 59 women with (n = 35) or without (n = 24) endometriosis. The latter group comprised women undergoing surgery for ovarian cystadenomas. PlGF concentrations in the peritoneal fluid were measured using an enzyme-linked immunosorbent assay. RESULTS: PlGF concentration in the peritoneal fluid was markedly elevated in the endometriosis patients (median 189 pg/ml, interquartile range 84-475 pg/ml) as compared with the controls (88 pg/ml, 41-213 pg/ml; P < 0.001), especially in women with red lesions. Significantly greater values during the secretory phase of the menstrual cycle as compared with the proliferative phase were observed in both the control (cystadenoma) group (P < 0.05) and the endometriosis group (P < 0.001). CONCLUSIONS: Our findings suggest that production of PlGF is sensitive to the cyclic changes in ovarian steroids and may contribute to the pathogenesis of endometriosis, especially that of red lesions, by promoting neovascularization.