IgG heavy chain allotypes (Gm) in autoimmune diseases.

Serum samples from 100 patients with myasthenia gravis, 322 with Graves' disease, 113 with Hashimoto's disease, 132 with systemic lupus erythematosus (SLE), 192 with insulin-dependent juvenile diabetes mellitus, 83 with Behçet's syndrome, 73 with psoriasis vulgaris, 258 with leprosy, 112 with Duchenne progressive muscular dystrophy and 343 non-related normal controls were studied for Gm allotypes. The incidence of Gm phenotypes with Gm(2) was significantly increased in patients with myasthenia gravis. Graves' disease, Hashimoto's disease, and high in SLE patients. The Gm1,2,21 haplotype was increased in patients with myasthenia gravis (chi 2 = 34 . 08, corrected P less than 0 . 001), Hashimoto's disease (chi 2 = 12 . 39, corrected P less than 0 . 05), Graves' disease (chi 2 = 8 . 65, corrected P less than 0 . 05), and SLE (chi 2 = 6 . 41, 0 . 1 greater than corrected P greater than 0 . 05). The total chi-square for the four different Gm haplotypes was significantly increased in patients with myasthenia gravis (chi 2 = 44 . 46, corrected P less than 0 . 001), SLE (chi 2 = 20 . 70, corrected P less than 0 . 005), Hashimoto's disease (chi 2 = 17 . 03, corrected P less than 0 . 025), and Graves' disease (chi 2 = 11 . 87, corrected P less than 0 . 025). Our data suggest the presence of Gm-associated pathogenic polygenes in certain autoimmune disorders.     

Functional dissection of the cytoplasmic subregions of the interleukin-5 receptor alpha chain in growth and immunoglobulin G1 switch recombination of B cells

The interleukin-5 receptor alpha chain (IL-5Ralpha) is known to regulate the development and function of B cells and eosinophils. Although the functions of IL-5Ralpha cytoplasmic domain subregions have been studied extensively using cultured cell lines, this approach has limitations when studying the functions of distinct primary B-cell subpopulations and their responsiveness to IL-5. In the present study, we generated mice on an IL-5Ralpha null background, each expressing a mutant form of an IL-5Ralpha transgene ligated to a mu enhancer and VH promoter, either lacking the cytoplasmic DC3 region or substituting two proline residues for alanine (ApvA) in the membrane-proximal ppvp motif of the cytoplasmic domain. The ppvp motif, which mediates activation of JAK2/STAT5 and Btk, also contributes to c-fos, c-jun and c-myc expression. IL-5Ralpha null mutant mice showed impaired B-1-cell development, reduced serum immunoglobulin G3 (IgG3) and IgM, no IL-5-induced enhancement of B-cell proliferation and IL-5-induced switch recombination from the mu gene to gamma1 gene; these were not recovered following the expression of the ApvA mutant. In contrast, absence of the DC3 region affected the IL-5-induced switch recombination from the mu to the gamma1 gene and B-1-cell development, while IL-5-induced proliferation and IgM production were at levels similar to those of B cells expressing wild-type IL-5Ralpha transgene. The results clearly indicated that the ppvp motif and the DC3 region of IL-5Ralpha played distinct roles in B-cell proliferation and differentiation. Thus, this present approach offers new insights into the functions of the cytoplasmic subregions of IL-5Ralpha, in particular its carboxy-terminal region.     

Prevalence of Lyme borreliosis spirochetes in ixodid ticks of Japan, with special reference to a new potential vector, Ixodes ovatus (Acari: Ixodidae).

To determine the vectors of Lyme borreliosis in Japan, the ixodid ticks taken in Hokkaido, Fukushima, Nagano, Ehime, and Kochi Prefectures were examined individually for spirochetal infections by culturing their midgut tissues in BSK medium. The cultivable spirochetes identified as Borrelia burgdorferi were detected in Ixodes persulcatus Schulze adults (15.5% of 634), nymph (6.7% of 15) and Ixodes ovatus Neumann adult (9.6% of 188) from Hokkaido, I. ovatus adults (25.5% of 47) from Fukushima, and I. persulcatus adults (21.7% of 143) and I. ovatus adults (27.1% of 85) from Nagano. However, 72 Haemaphysalis spp. from Ehime and Kochi were free from spirochetal infection. I. persulcatus is a vector of Lyme borreliosis in the Far East, but in Japan, I. ovatus may also transmit the spirochetes to human and feral animals.     

Receptor autoradiographic evidence of specific brain natriuretic peptide binding sites in the porcine subfornical organ

Specific binding sites of brain natriuretic peptide (BNP), a newly discovered peptide in the subfornical organ (SFO) of porcine brain were investigated, following incubation of related tissue sections with 125I-BNP, then using autoradiography and an image analysis coupled with computer-assisted microdensitometry. Specific 125I-BNP binding sites were found to be localized in the SFO, an area densely labeled by 125I-alpha-rat atrial natriuretic peptide and 125I-(Sar1,Ile8)-angiotensin II. Specific 125I-BNP binding to the SFO was displaced by unlabeled BNP, with a high affinity, and was calculated to be Ka = 0.385 x 10(-9) M and Bmax = 40.1 fmol/mg using a LIGAND computer program. Acquisition of these present findings enhances our knowledge of the physiology of BNP, atrial natriuretic peptides and angiotensin II system in the SFO.     

Clinical application of antithrombogenic hydrogel with long poly(ethylene oxide) chains

Thirty clinical tests on PVC drain tubes coated with hydrophilic copolymer with long poly(ethylene oxide) chains (PEO-COAT) were carried out. Controls were non-coated PVC drain tubes. Thrombogenesis was observed in 24 out of 30 non-coated PVC drain tubes (80%) and in only 4 out of 30 PEO-COAT drain tubes (13%). PEO-COAT drain tubes significantly suppressed absorption of plasma proteins and adhesion of platelets. The excellent antithrombogenic property of this hydrophilic polymer, already suggested by in vitro and in vivo experiments, was demonstrated here clinically.     

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.     

Pioglitazone improves the phenotype and molecular defects of a targeted Pkd1 mutant

Mutations of either PKD1 or PKD2 are associated with autosomal dominant polycystic kidney disease (ADPKD). The molecular function of the gene product of PKD1, polycystin-1, in vitro has been elucidated recently, but the molecular pathological consequences of the loss of polycystin-1 in vivo have remained unclear. We have generated a mouse with a targeted deletion of exons 2-6 of Pkd1 to study the molecular defects in Pkd1 mutants. Homozygote embryos (Pkd1(-/-)) developed hydrops, cardiac conotruncal defects and renal cystogenesis. Total protein levels of beta-catenin in heart and kidney and c-MYC in heart were decreased in Pkd1(-/-) embryos. In the kidneys of Pkd1(-/-), the expression of E-cadherin and PECAM in basolateral membranes of renal tubules was attenuated, and tyrosine phosphorylation of epidermal growth factor receptor and Gab1 were constitutively enhanced when cystogenesis started on embryonic day (E) 15.5-16.5. Maternally administered pioglitazone, a thiazolidinedione compound, resolved these molecular defects of Pkd1(-/-). Treatment with pioglitazone improved survival of Pkd1(-/-) embryos and ameliorated the cardiac defects and the degree of renal cystogenesis. Long-term treatment with pioglitazone improved the endothelial function of adult Pkd1(+/-). These data indicated that molecular defects observed in Pkd1(-/-) embryos contributed to the pathogenesis of ADPKD and that thiazolidinediones had a compensatory effect on the pathway affected by the loss of polycystin-1. Pathways activated by thiazolidinediones may provide new therapeutic targets in ADPKD.