Bacterial superantigen-treated intestinal epithelial cells upregulate heat shock proteins 25 and 72 and are resistant to oxidant cytotoxicity

While the pathological events evoked by infection are commonly described, effective host responses to bacteria and their products should primarily be protective. Heat shock protein (Hsp) expression is upregulated by many stimuli and serves to maintain intracellular protein integrity. The ability of the prototypic superantigen, Staphylococcus aureus enterotoxin B (SEB) to induce Hsps was investigated with BALB/c mice and by in vitro addition to the murine small intestinal epithelial cell line MSIE. SEB-treated (5 or 100 microg intraperitoneally) mice revealed increased Hsp25 and Hsp72, but not Hsc73, in jejunal lymphocytes and epithelial cells. A similar Hsp response to SEB occurred in MSIE cells and was preceded by activation of the ERK1/2 and p38 mitogen-activated protein kinases but not the SAPK/JNK pathway; pharmacological inhibition of ERK1/2, but not p38, significantly reduced SEB-induced Hsps. Moreover, SEB-treated MSIE cells were protected against oxidant-induced cytotoxicity (measured by 51Cr release) and F-actin depolymerization. Thus, SEB exposure results in a rapid induction of the Hsp25 and Hsp72 in intestinal epithelial cells, both directly and through lymphocyte activation, and we suggest that this event is important in protecting the gut from damage by Staphylococcus infection or in the reparatory process and may be a generalized response to lumen-derived bacterial toxins.     

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.     

Transgenic rabbits with increased VEGF expression develop hemangiomas in the liver: a new model for Kasabach-Merritt syndrome

Clinical studies have provided ample evidence that high (either systemic or local) levels of vascular endothelial growth factor (VEGF) are associated with several pathophysiological disorders, including hemangiomas. To investigate whether elevated VEGF expression could directly affect these disorders, we created a transgenic (Tg) rabbit model with increased hepatic expression of the human VEGF(165) transgene under the control of the human alpha-antitrypsin promoter. Tg rabbits exhibited marked hepatomegaly, with livers 2.5-fold heavier than those of control rabbits. Histological analysis revealed that the livers of Tg rabbits showed prominent dilation of the sinusoids and formed various-sized blood vessel networks, a feature of diffuse hemangiomas. Immunohistochemical staining revealed that the hepatocytes produced VEGF(165), whereas plasma VEGF(165) was not detected. Furthermore, Tg rabbits suffered from hemolytic anemia, thrombocytopenia and splenomegaly, which was associated with marked extramedullary hematopoiesis. The manifestations of Tg rabbits mimic many of the features of hemangiomatous disorders in humans such as the Kasabach-Merritt syndrome, and therefore this model may be potentially useful for the study of the pathogenesis and complications of hemangiomas as well as the investigation of angiogenesis inhibitors.     

Three novel mutations of the fibrillin-1 gene and ten single nucleotide polymorphisms of the fibrillin-3 gene in Marfan syndrome patients

Marfan syndrome (MFS) is an autosomal dominant disorder of the extracellular matrix. Allelic variations in the gene for fibrillin-1 (FBN1) have been shown to cause MFS. To date, over 550 mutations have been identified in patients with MFS and related connective tissue diseases. However, about a half of MFS cases do not possess mutations in the FBN1 gene. These findings raise the possibility that variants located in other genes cause or modify MFS. To explore this possibility, firstly we analyzed FBN1 allelic variants in 12 Japanese patients with MFS, and secondly we analyzed fibrillin-3 gene (FBN3) in patients without FBN1 mutations using conformation sensitive gel electrophoresis (CSGE) and direct sequencing analysis. We identified three novel FBN1 mutations and ten FBN3 single nucleotide polymorphisms (SNPs). In this report, we could not detect a responsible mutation of the FBN3 gene for MFS. Although the number of the cases in this report is small, at least these results suggest that disease-causing mutations in exon regions of the FBN3 gene are very rare in MFS.Nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession numbers: AB177797, AB177798, AB177799, AB177800, AB177801, AB177802, AB177803     

Inhibition of MAP kinase activity moderates changes in expression and function of Cx32 but not claudin-1 during DNA synthesis in primary cultures of rat hepatocytes

The signal transduction pathways and activation of the MAP kinase or PI3 kinase signaling cascade regulate a variety of cellular processes, including proliferation and differentiation in hepatocytes. To elucidate the mechanisms of signal transmission required for the regulation of gap and tight junctions during DNA synthesis in rat hepatocytes, we determined changes of expression and function of gap and tight junctions of cells grown in primary culture, using inhibitors of signaling pathways for MAP kinase (PD98059) and PI3 kinase (LY294002). During the stimulation of DNA synthesis induced by epidermal growth factor (EGF), immunoreactivity and mRNAs of gap junction protein Cx32 and of tight junction protein claudin-1 markedly decreased with reduction of gap junctional intercellular communication (GJIC) and the fence function of tight junctions. In Western blots, whole-cell lysate of claudin-1 protein decreased and phosphorylated Cx32 protein in the insoluble fraction of Triton X-100 increased during the stimulation of DNA synthesis. During reinhibition of DNA synthesis, the changes of Cx32 and claudin-1 returned to control levels, as did both functions. In treatment with the inhibitors before DNA synthesis, PD98059 inhibited the changes of expression and function of Cx32, but not claudin-1, without inhibition of cell growth, whereas LY294002 completely inhibited cell growth. These findings indicate that the PI3 kinase pathway rather than the MAP kinase pathway plays an important role for EGF-induced proliferation of rat hepatocytes, and that changes of Cx32 in hepatocytes during the stimulation of DNA synthesis may be in part controlled through MAP kinase. Furthermore, Cx32, but not claudin-1, protein may be a target of activated MAP kinase in hepatocytes.     

Significant association between nonsyndromic oral clefts and arylhydrocarbon receptor nuclear translocator (ARNT)

The etiology of nonsyndromic oral clefts (cleft lip, cleft palate, or cleft lip and palate) is still controversial, but is considered to involve both genetic and environmental factors. One of suspected environmental factors is 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) found in tobacco, herbicides, contaminated soil, and food. TCDD administered during organogenesis in mice causes a high incidence of CP in fetuses. There is ample evidence that aryl hydrocarbon receptor (AHR), AHR nuclear translocator (ARNT), and cytochrome P450 1A1 (CYP1A1) are involved in TCDD metabolism. We assessed whether there is any association in the Japanese population of nonsyndromic oral clefts with single nucleotide polymorphisms (SNPs) in the AHR, ARNT, and CYP1A1 genes using transmission disequilibrium test (TDT) and case-control study. We identified and investigated three SNPs in ARNT; 567G/C (V189V), IVS12-19T/G, and 2117C/T (P706L). Two amino acid substitutions, R554L in AHR and I462V in CYP1A1, were also investigated. In the TDT, the C allele of ARNT 567G/C was preferentially transmitted to patients (P = 0.033). When a haplotype consisting of 567G/C and IVS12-19T/G in ARNT was considered, the preferential transmission of the CT (567C-IVS12-19T) haplotype was observed (P = 0.0012). In a case-control study, a significant association of IVS12-19T/G in ARNT was observed (P = 0.021). The SNPs studied in AHR and CYP1A1 were not associated with the disease. Our results suggest that ARNT is involved in the development of nonsyndromic oral clefts in the Japanese population.     

Momentary changes in the cardiovascular autonomic system during mental loading in patients with panic disorder: a new physiological index "rho(max)"

BACKGROUND: Although panic disorder (PD) is suggestive of autonomic nervous system dysfunction, especially in the cardiovascular autonomic system (CAS), the results in many previous studies are still controversial. Using a new physiological index which could well reflect emotional reaction to visual stimuli (Yoshizawa, M., Sugita, N., Tanaka, A., Abe, K., Yambe, T., Nitta, S., 2001. Quantiatative Physioligical Evaluation of Three Dimensional Images. The Seventh International Conference on Virtual Systems and Multumedia, 25-27.), we studied momentary changes in the CAS in patients with PD during audiovisual stimulation (AS) as mental loading. METHODS: During AS, exposed to a video of imaginary experiences such as driving a motor vehicle or diving into the sea, blood pressure (BP) and heart rate (HR) were measured in 12 remitted patients with PD and 19 age- and sex-matched normal controls (NC). We used the maximum cross-correlation coefficient (rho(max)) from the BP to the HR, whose frequency components were limited to around 0.1 Hz. RESULTS: The rho(max) was an available index which could detect the momentary changes in the CAS during AS in both groups. The two-way ANOVA disclosed significant group and time effects on the rho(max). The momentary response to emotional stimuli in the PD patients was slower than that in the NC subjects. LIMITATIONS: Antidepressants have a potential impact on the autonomic variables in this study. CONCLUSIONS: These findings suggest that there may be a dysfunction of the CAS in remitted PD patients and that the dysfunction may be one of the trait markers of PD. To confirm these findings, however, further studies with a large sample size are required.     

Prostaglandin E1-initiated immune regulation during human mixed lymphocyte reaction

Prostaglandin E1 (PGE1) has therapeutic value for transplantations due to its microvascular activity. Interleukin (IL)-18, which is elevated in plasma during the acute rejection after organ transplantation, elicits the expression of intercellular adhesion molecule (ICAM)-1, B7.1, B7.2, CD40, and CD40 ligand (CD40L) on monocytes as well as the production of interferon (IFN)-gamma and IL-12 and proliferation of T-cells during the human mixed lymphocyte reaction (MLR) in an in vitro model of acute rejection. In contrast, PGE1 inhibits all the adhesion molecule expression, cytokine production and T-cell proliferation in the presence of IL-18. The effects of PGE1 depend on stimulation of the IP/EP2/EP4-receptor, and thus, PGE1 might have therapeutic potential for treating acute rejection due to its immune regulatory effect.     

Modification of immune complexes deposited in glomeruli in tissue sections treated with sulfonized gamma-globulin.

The modification of immune complexes (IC) deposited in renal tissues obtained from the patients with membranous nephropathy (MN), membranoproliferative glomerulonephritis (MGPN), systemic lupus erythematosus (SLE), IgA nephropathy, and rabbits with chronic serum sickness was investigated by an immunofluorescent technique. Following treatment with sulfonized gamma-globulin (S-GG), IgG deposited in glomeruli disappeared. However, the deposition of other immunoglobulins, complement and protein A, and the fixation of guinea pig complement were not influenced in spite of the disappearance of IgG deposits. In addition, deposition of antigen was demonstrated following treatment with S-GG in experimental animals. By elution with 0.02 M citrate buffer at pH 3.2, the intensity of these deposits markedly decreased or disappeared completely. The data suggests that the phenomena of IgG-dispersion from IC following the treatment with either S-GG or acid buffer are essentially different. The exact mechanisms of selective IgG-dispersion by the treatment with S-GG are unclear, but an antiglobulin activity (rheumatoid factor or anti-idiotypic antibody) of deposited IC may play an important role.