Histopathological study on the PTHrP-induced incisor lesions in rats

Parathyroid hormone related peptide (PTHrP) was discovered as a causative factor of humoral hypercalcemia of malignancy (HHM). The present study elucidates the histopathological characters of incisor lesions in the HHM rat model. Nude rats were implanted with PTHrP-expressing tumor (LC-6) cells, maintained for 12 weeks, after which the mandibular incisors were collected. Incisor fractures were observed grossly. Microscopically, hypercalcified dentin, dentin niche with osteodentin, and thinning of dentin were observed. Hypercalcified dentin was observed as a basophilic line of calcified dentin without associated odontoblastic changes, whereas dentin niche and thinning of dentin occurred with osteodentin and loss of cell height, respectively. In contrast with hypercalcified dentin, which was distributed throughout the dentin, dentin niche and thinning of dentin were localized to the labial area of the apical and middle region, and to the labial and lingual areas of the middle and incisal region, respectively. These results suggest that hypercalcemia affected the entire calcification process resulting in hypercalcified dentin, and that high PTHrP concentrations affected selective populations of odontoblasts resulting in formation of dentin niche and thinning of dentin. The localization of dentin niche and thinning of dentin also suggest that PTHrP may also be involved odontoblastic development in the rat.     

Roles of the Maltese cross form in the development of parasitemia and protection against Babesia microti infection in mice

Babesia microti, a hemoprotozoan parasite of rodents, is also important as a zoonotic agent of human babesiosis. The Maltese cross form, which consists of four masses in an erythrocyte, is characteristic of the developmental stage of B. microti. Monoclonal antibody (MAb) 2-1E, which specifically recognizes the Maltese cross form of B. microti, has been described previously. In the present study, we examined the roles of the Maltese cross form during the infectious course of B. microti in mice. The number of the Maltese cross form increased in the peripheral blood of infected mice prior to the peak of parasitemia. With confocal laser scanning microscopy, MAb 2-1E was found to be reactive with the ring form, with the parasites undergoing transformation to the Maltese cross form and subsequent division, and also with extracellular merozoites. Furthermore, the Maltese cross form-related antigen (MRA) gene was isolated from a B. microti cDNA library by immunoscreening with MAb 2-1E, and the nucleotide sequence was determined. Genomic analyses indicated that the MRA gene exists as a single-copy gene in B. microti. Immunization of mice with recombinant MRA induced significant protective immunity against B. microti infection. These findings indicate that the Maltese cross form plays important roles in both the development of parasitemia and the protective response against the infection.     

Breast milk macrophages spontaneously produce granulocyte-macrophage colony-stimulating factor and differentiate into dendritic cells in the presence of exogenous interleukin-4 alone

Peripheral blood monocytes extravasate and differentiate into tissue macrophages to mediate effective local defence, but how tissue-specific stimuli and environments may influence their functions remains unknown. Here, we found that peripheral blood monocytes gained the ability to produce granulocyte-macrophage colony-stimulating factor (GM-CSF) upon exposure to breast milk and differentiated into CD1+ dendritic cells (DCs) in the presence of exogenous interleukin-4 (IL-4) alone. This in vitro observation appeared physiologically relevant since macrophages that were freshly isolated from breast milk were also found to produce GM-CSF spontaneously. Furthermore, in contrast to peripheral blood monocytes that differentiated into DCs only in the presence of both exogenous GM-CSF and IL-4, differentiation of breast milk macrophages into DCs was induced by incubation with exogenous IL-4 alone. These IL-4-stimulated breast milk macrophages were efficient in stimulating T cells, suggesting their potential role in mediating T-cell-dependent immune responses in situ. On the other hand, unexpected expression of DC-SIGN, a DC-specific receptor for human immunodeficiency virus (HIV), even in unstimulated breast milk macrophages, may favour HIV infection, resulting in an increased risk of mother-to-infant vertical transmission of the virus via breast milk. Thus, tissue-specific development of macrophages is often linked to effective local immunity, but may potentially provide an opportunity for a pathogen to spread and transmit.     

Virulence of Streptococcus mutans: comparison of the effects of a coupling sugar and sucrose on certain metabolic activities and cariogenicity.

A coupling sugar preparation (sucrose-free [CSSF]), which contains a mixture of sugars, oligosaccharides, and oligosaccharides terminated at the reducing end by sucrose, served as a substrate for growth and acid production by Streptococcus mutans 6715. However, CSSF was a poor substrate for cellular aggregation, glucosyltransferase activity, plaque formation, and adherence of cells to glass surfaces. In the presence of sucrose, CSSF inhibited glucosyltransfer activity and adherence of cells. The substitution of CSSF for sucrose in a rat diet significantly reduced caries score. Furthermore, rats fed diets containing sucrose and CSSF had significantly fewer carious lesions than did rats fed a sucrose diet.     

Inhibitory effect of antiserum to surface antigen P50 of Babesia gibsoni on growth of parasites in severe combined immunodeficiency mice given canine red blood cells

The inhibitory effect of an antiserum to surface protein P50 of Babesia gibsoni on the growth of the parasite was determined with severe combined immunodeficiency mice given canine red blood cells. The antiserum to the recombinant P50 protein significantly inhibited the parasite growth, indicating that P50 might be a useful vaccine candidate.     

Purification of merozoites ofTheileria sergenti from infected bovine erythrocytes

A simple and rapid method for purifying merozoites ofTheileria sergenti from infected bovine erythrocytes was developed. Infected erythrocytes were lysed by the use of a cytolytic toxin produced byAeromonas hydrophila and the lysate was subjected to ultracentrifugation in a Percoll discontinuous density gradient. Pure and morphologically intact merozoites free of erythrocyte membrane were recovered from a band formed at the interface of 40% and 60% (by vol.) Percoll solutions. In this merozoite fraction, contamination of erythrocyte membrane proteins was not detected as examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.     

Role of CD4 molecule in intrathymic T-cell development.

We have investigated the role of CD4 molecules in intrathymic T-cell repertoire selection. The administration of monoclonal antibody (mAb) to CD4 in organ culture of murine foetal thymus (FTOC) completely inhibited the development of CD4+8- cells, and additional treatment with anti-class II MHC (Ia) mAb caused no further effects on this inhibition. On the other hand, when the potentially autoreactive cells in Mls-1a mice were monitored by expression of the Mls-1a-reactive V beta 6 gene product of T-cell receptor alpha beta (TcR alpha beta), the treatment with anti-CD4 resulted in the appearance of V beta 6-bearing cells to some extent, but this effect was considerably reinforced by the combinatory use of anti-Ia mAb with anti-CD4. In a model system where the bacterial superantigen staphylococcal enterotoxin B serves as self-antigen to deplete V beta 8-bearing cells in FTOC, the depletion of V beta 8+ cells was restored partially by anti-CD4 alone but completely by the combination with anti-Ia. These results suggest that CD4 is indispensable for positive selection of all CD4+8- thymocytes, whereas participation of CD4 in negative selection is only partial. It was also observed that the development of TcR alpha beta-bearing cells in the CD4-8- population was inhibited by the treatment with anti-CD4 mAb. In Mls-1a mice, V beta 6-bearing cells were developed in CD4-8+, CD4+8+, and also in CD4-8- populations after anti-CD4 mAb treatment. It is suggested that TcR alpha beta-bearing CD4-8- cells are possibly originated from CD4+ cells and undergo CD4-mediated thymic selection.     

Immunohistochemical localization of transforming growth factor alpha in chemically induced rat hepatocellular carcinomas with reference to differentiation and proliferation

Hepatocellular carcinomas (HCCs) were induced in male Fischer 344 rats with dietary 3'-methyl-4-(dimethylamino)-azobenzene treatment and were classified into solid, glandular (well- or poorly differentiated), and trabecular types. Investigation of cell proliferation kinetics and immunohistochemical localization of transforming growth factor alpha (TGF-alpha) demonstrated all solid (n = 24) and poorly differentiated glandular type (n = 6) HCCs to have TGF-alpha-positive nuclei. Nuclear staining of TGF-alpha was also observed in 13 of 28 (46%) trabecular-type HCCs, whereas 12 (43%) exhibited cytoplasmic staining, and 3 (11%) were negative. As for well-differentiated glandular HCCs, 7 of 20 (35%) were positively stained in their nucleus, another 7 (35%) demonstrated antibody binding in the cytoplasm, and 6 (30%) were negative. The order for growth rate evaluated by bromodeoxyuridine (BrdU) labeling was solid (38.22%), poorly differentiated glandular (26.82%), trabecular (7.98%), and well-differentiated glandular (2.57%) types. For trabecular HCCs with nuclear, cytoplasmic, or negative TGF reactions, values were 13.39% (n = 13), 3.61% (n = 12), and 2.01% (n = 3), respectively. Likewise, BrdU-labeling indices for the counterpart groups of well-differentiated glandular type HCCs were 4.53, 1.91, and 1.29%, respectively. The results indicate that TGF-alpha expression might be linked to histopathological differentiation and cell proliferation in rat HCCs.