Preventive and therapeutic effects of oral tolerance in a murine model of asthma

Allergic asthma is an inflammatory disease of the airways, and Th2 cells secreting IL-4 and IL-5 play a pivotal role in its pathogenesis. We have previously demonstrated that oral tolerance can be induced and maintained more profoundly in a Th2-related immune response, and that an ongoing immune response can be suppressed by the oral administration of antigen combined with an appropriate feeding regimen. In the present study, we examined the preventive and therapeutic effects of the oral administration of allergen on a Th2-mediated immune disorder using a murine model of asthma. Our results show that the development of asthma can be blocked completely by orally administering allergen. Airway hyperreactivity, allergen-specific IgE production, Th2-derived cytokines, allergen-induced T cell proliferation and the infiltration of inflammatory effector cells into the lung were prevented by such oral administration. To assess the therapeutic effects of oral administration on the progression of asthma, we tested the effects of oral tolerance in an established asthma model, and found that a multiple high dose-feeding regimen was effective at suppressing the progression of mild asthma. In the high dose-feeding group, the number of eosinophils in bronchoalveolar lavage fluid was reduced and airway reactivity also decreased. However, this was insufficient to reduce airway reactivity and eosinophilia in bronchoalveolar lavage fluid in cases of severe asthma. These results demonstrate that allergic asthma may be ameliorated by feeding allergen; there is hope that these results will provide a new immunotherapeutic strategy for allergic asthma.     

Regulation of phosphatidylinositol kinases by arachidonic acid in rat submandibular gland cells

Phosphoinositide kinases were characterized in membrane extracts of rat submandibular gland cells. Both phosphatidylinositol (PI) 4-kinase and phosphatidylinositol-4-phosphate (PI(4)P) 5-kinase phosphorylated endogenous substrates in reactions that were linear for up to 5 min, were activated by Mg²⁺ and showed maximal activity around neutral pH. PI 4-kinase was stimulated by Triton X-100 at an optimal concentration of 0.22%, but the detergent had an inhibitory effect on PI(4)P 5-kinase. Arachidonic acid (AA), at concentrations greater than 100 M, inhibited the activity of both enzymes in a dose-dependent manner. The inhibitory effect was replicated by other unsaturated fatty acids, but not by a saturated fatty acid of the sn-20 series. The nature of AA inhibition of the kinases was examined in enzyme kinetic studies with exogenous phosphoinositide and adenosine 5-triphosphate (ATP) substrates. Lineweaver-Burk plots of PI 4-kinase activity showed that AA had no effect on the apparent K _m for either PI or ATP, but that the fatty acid significantly reduced V _max (PI) from 331 to 177 pmol.mg^–1.min^–1 and V _max (ATP) from 173 to 59 pmol.mg^–1.min^–1. This inhibitory action was consistent for PI(4)P 5-kinase kinetics, where again, AA did not alter apparent K _m values, but lowered V _max for both PI(4)P and ATP by around 50%. Since the combination of a reduced V _max and an unchanged K _m value indicates noncompetitive enzyme inhibition, it is proposed that AA regulates phosphoinositide cycle activity in submandibular gland cells by acting as a noncompetitive inhibitor of PI 4-kinase and PI(4)P 5-kinase.     

Brain tumor in the first year of life: a single institute study.

Brain tumors in infants present special diagnostic and therapeutic challenges. To figure out the clinical features, pathological classification of the tumors and the treatment outcome of infantile brain tumors, 458 children (age<16) with brain tumors were reviewed retrospectively. Among them 21 cases (4.6%) were diagnosed during the first 12 months of life. Two tumors were definitely of congenital origin. The majority of infants with brain tumors presented with increased intracranial pressure. Fourteen tumors were located at the supratentorial area. Sixteen cases had neuroepithelial tumors; astrocytoma (optic pathway), supratentorial primitive neuroectodermal tumor (PNET) and medulloblastoma were found in three cases each. There were two treatment-related mortalities. Compared with the outcomes in older children, the treatment outcome was poorer in medulloblastoma and the optic pathway glioma which showed a higher growth potential. Because of the limited application of postoperative adjuvant therapy, radical surgical removal played a more important role in this age group. The prognosis of patients in whom the tumors could not be totally removed, largely depended on the pathological malignancy of the tumors. Though the treatment outcome was not always dismal, immaturity of the brain, higher growth potential, perioperative risks, limitations in adjuvant therapy, and pessimistic attitude on the part of parents made management more challenging.     

Inhibition of interleukin-8 expression by dexamethasone in human cultured airway epithelial cells.

Interleukin-8 (IL-8) is a neutrophil chemotactic factor expressed in many cell types, including human airway epithelial cells (HAEC). Inhaled corticosteroids are now used increasingly early in the treatment of airway inflammation such as in asthma, and directly interact with HAEC at relatively high concentrations. We have investigated the effect of dexamethasone on IL-8 expression in primary cultured HAEC obtained from transplantation donors. Northern blot analysis was used to measure IL-8 mRNA levels in HAEC, and radioimmunoassay was used to measure IL-8 protein in culture supernatant fluids. We demonstrated that IL-8 was expressed by primary cultured HAEC and that this was enhanced by IL-1 beta and tumour necrosis factor-alpha stimulation, but not by IL-6 or lipopolysaccharide. Dexamethasone suppressed IL-8 mRNA expression and protein synthesis dose-dependently in both resting and stimulated HAEC. The half-life of IL-8 mRNA determined in the presence of actinomycin D was less than 1 hr, and dexamethasone preincubation had no effect on mRNA stability. These results support the view that HAEC may play an important role in the pathogenesis of airway inflammatory diseases, and that glucocorticosteroids may exert their anti-inflammatory effects by blocking IL-8 gene expression and generation in these cells.     

Identification of vaccine candidate peptides in the NcSRS2 surface protein of Neospora caninum by using CD4+ cytotoxic T lymphocytes and gamma interferon-secreting T lymphocytes of infected holstein cattle

Previously, our laboratory showed that Holstein cattle experimentally infected with Neospora caninum develop parasite-specific CD4+ cytotoxic T lymphocytes (CTL) that lyse infected, autologous target cells through a perforin-granzyme pathway. To identify specific parasite antigens inducing bovine CTL and helper T-lymphocyte responses for vaccine development against bovine neosporosis, the tachyzoite major surface proteins NcSAG1 and NcSRS2 were targeted. In whole tachyzoite antigen-expanded bovine T-lymphocyte lines, recombinant NcSRS2 induced potent memory CD4+- and CD8+-T-lymphocyte activation, as indicated by proliferation and gamma interferon (IFN-gamma) secretion, while recombinant NcSAG1 induced a minimal memory response. Subsequently, T-lymphocyte epitope-bearing peptides of NcSRS2 were mapped by using overlapping peptides covering the entire NcSRS2 sequence. Four experimentally infected cattle with six different major histocompatibility complex (MHC) class II haplotypes were the source of immune cells used to identify NcSRS2 peptides presented by Holstein MHC haplotypes. NcSRS2 peptides were mapped by using IFN-gamma secretion by rNcSRS2-stimulated, short-term T-lymphocyte cell lines, IFN-gamma enzyme-linked immunospot (ELISPOT) assay with peripheral blood mononuclear cells, and 51Cr release cytotoxicity assay of rNcSRS2-stimulated effector cells. Four N. caninum-infected Holstein cattle developed NcSRS2 peptide-specific T lymphocytes detected ex vivo in peripheral blood by IFN-gamma ELISPOT and in vitro by measuring T-lymphocyte IFN-gamma production and cytotoxicity. An immunodominant region of NcSRS2 spanning amino acids 133 to 155 was recognized by CD4+ T lymphocytes from the four cattle. These findings support investigation of subunit N. caninum vaccines incorporating NcSRS2 gene sequences or peptides for induction of NcSRS2 peptide-specific CTL and IFN-gamma-secreting T lymphocytes in cattle with varied MHC genotypes.     

Voltage-dependent sodium and calcium currents in acutely isolated adult rat trigeminal root ganglion neurons

Voltage-dependent sodium (INa) and calcium (ICa) currents in small (<30 microM) neurons from adult rat trigeminal root ganglia were characterized with a standard whole cell patch-clamp technique. Two types of INa showing different sensitivity to tetrodotoxin (TTX) were recorded, which showed marked differences in their activating and inactivating time courses. The activation and the steady-state inactivation kinetics of TTX-resistant INa were more depolarized by about +20 and +30 mV, respectively, than those of TTX-sensitive INa. Voltage-dependent ICa was recorded under the condition that suppressed sodium and potassium currents with 10 mM Ca2+ as a charge carrier. Depolarizing step pulses from a holding potential of -80 mV evoked two distinct inward ICa, low-voltage activated (LVA) and high-voltage activated (HVA) ICa. LVA ICa was first observed at -60 to -50 mV and reached a peak at about -30 mV. Amiloride (0.5 mM) suppressed approximately 60% of the LVA ICa, whereas approximately 10% of HVA ICa was inhibited by the same concentration of the amiloride. LVA ICa was far less affected by the presence of external Cd2+ or the replacement of Ca2+ by 10 Ba2+ than HVA ICa. The omega-conotoxin GVIA (omega-CgTx), an N-type ICa blocker, suppressed approximately 65% of the whole cell HVA ICa at the concentration of 1 microM. The omega-CgTx-resistant HVA ICa was sensitive to nifedipine (10 microM), a dihydropyridine (DHP) calcium channel antagonist, which produced an additional blockade by approximately 25% of the drug-free control ( approximately 70% of the omega-CgTx-resistant ICa). The combination of 10 microM nifedipine and 1 microM omega-CgTx left approximately 13% of the drug-free control ICa unblocked. The DHP agonist S(-)-BayK8644 (5 microM) shifted the activation of the HVA ICa to more negative potentials and increased its maximal amplitude. Additionally, S(-)-BayK8644 caused the appearance of a slowed component of the tail current. These results clearly demonstrate that the presence of two types of sodium channels, TTX sensitive and resistant, and three types of calcium channels, T, L, and N type, in the small-sized adult rat trigeminal ganglion neurons.     

Carbon plasma immersion ion implantation of nickel-titanium shape memory alloys

Nickel-titanium (NiTi) shape memory alloys possess super-elasticity in addition to the well-known shape memory effect and are potentially suitable for orthopedic implants. However, a critical concern is the release of harmful Ni ions from the implants into the living tissues. We propose to enhance the corrosion resistance and other surface and biological properties of NiTi using carbon plasma immersion ion implantation and deposition (PIII&D). Our corrosion and simulated body fluid tests indicate that either an ion-mixed amorphous carbon coating fabricated by PIII&D or direct carbon PIII can drastically improve the corrosion resistance and block the out-diffusion of Ni from the materials. Our tribological tests show that the treated surfaces are mechanically more superior and cytotoxicity tests reveal that both sets of plasma-treated samples favor adhesion and proliferation of osteoblasts.     

Changes of interleukin expression correlate with Helicobacter pylori infection and lymph node metastases in gastric carcinoma.

Helicobacter pylori (HP) infection induces expression of IL-8 and IL-10 in benign gastric epithelium. This study compared the expression of cytokines in CD4+ and CD8+ lymphocyte subsets of peripheral blood lymphocytes (PBL), benign mucosal lymphocytes (ML), and tumor infiltrative lymphocytes (TIL) as well as in the benign and malignant epithelial cells of the same patient, with respect to the presence of HP infection, lymph node metastases, and tumor histologic type. The mRNA of the cytokines was measured by a semiquantitative RT-PCR method. The levels were ranked and compared using the Wilcoxon sign-ranked test. Compared with CD8+ ML, the CD8+ TIL expresses higher levels of IL-6 and IL-8 but lower level of IL-4 in patients with lymph node metastases. In patients with HP infection, expression of IL-8 and IL-10 was higher in the gastric carcinoma cells than in the benign epithelial cells while expression of IL-6 and IL-8 were higher in CD8+ TIL than CD8+ ML. Overexpression of IL-8 in HP associated gastric carcinomas suggested that they might have arisen from HP-infected epithelial cells.     

Synergistic antimicrobial effect of cefotaxime and minocycline on proinflammatory cytokine levels in a murine model of Vibrio vulnificus infection.

BACKGROUND AND PURPOSE: Vibrio vulnificus causes primary bacteremia and necrotizing wound infection, leading to high morbidity and mortality in humans. This study aimed to evaluate the antimicrobial effect of cefotaxime and minocycline on proinflammatory cytokine levels in a murine model of V. vulnificus infection. METHODS: We investigated the dynamics of proinflammatory cytokines and their modulation by antimicrobial agents using a murine model of V. vulnificus infection. The change in cytokine levels was followed over a time course to identify the antimicrobial activity of the drugs against V. vulnificus. BALB/c female mice were challenged with an intraperitoneal infection using a clinical invasive isolate of Vv05191, and their cytokine levels were assayed over various time points. RESULTS: Serum levels of tumor necrosis factor-alpha, interleukin (IL)-1 beta, and IL-6 post-infection were found to be inoculum dose-dependent and positively correlated to the subsequent fatality rate in the infected mice. With an inoculum of 6.6 x 10(6) colony-forming units and intraperitoneal administration of cefotaxime, minocycline, or both, the serum and peritoneal fluid cytokine levels increased and then declined gradually. Comparison of the 3 antimicrobial regimens revealed that the magnitude of reduction in cytokine levels was greatest in mice treated with cefotaxime-minocycline combination. Moreover, the peritoneal fluid cytokine level in the combination group was significantly lower than that in the groups treated with minocycline or cefotaxime alone. CONCLUSIONS: The current results support the superiority of the combination therapy in treating invasive V. vulnificus infections.