IL-15 up-regulates iNOS expression and NO production by gingival epithelial cells

To investigate the biological activity of epithelial cells in view of host defense, we analyzed the mRNA expression of inducible NOS (iNOS) as well as NO production by human gingival epithelial cells (HGEC) stimulated with IL-15. RT-PCR analysis revealed that HGEC expressed IL-15 receptor alpha-chain mRNA. In addition, stimulation with IL-15 enhanced iNOS expression by HGEC through an increase of both mRNA and protein levels. Moreover, IL-15 up-regulated the production of NO(2)(-)/NO(3)(-), a NO-derived stable end product, from HGEC. The enhanced NO production by IL-15 was inhibited by AMT, an iNOS-specific inhibitor. These results suggest that IL-15 is a potent regulator of iNOS expression by HGEC and involved in innate immunity in the mucosal epithelium.     

Ca2+ Channels as Targets of Neurological Disease: Lambert–Eaton Syndrome and Other Ca2+ Channelopathies

In the nervous system, voltage-gated Ca2+ channels regulate numerous processes critical to neuronal function including secretion of neurotransmitters, initiation of action potentials in dendritic regions of some neurons, growth cone elongation, and gene expression. Because of the critical role which Ca2+ channels play in signaling processes within the nervous system, disruption of their function will lead to profound disturbances in neuronal function. Voltage-gated Ca2+ channels are the targets of several relatively rare neurological or neuromuscular diseases resulting from spontaneously-occurring mutations in genes encoding for parts of the channel proteins, or from autoimmune attack on the channel protein responses. Mutations in CACNAIA, which encodes for the alpha1A subunit of P/Q-type Ca2+ channels, lead to symptoms seen in familial hemiplegic migraine, episodic ataxia type 2, and spinocerebellar ataxia type 6. Conversely, autoimmune attack on Ca2+ channels at motor axon terminals causes peripheral cholinergic nerve dysfunction observed in Lambert-Eaton Myasthenic Syndrome (LEMS), the best studied of the disorders targeting voltage-gated Ca2+ channels. LEMS is characterized by decreased evoked quantal release of acetylcholine (ACh) and disruption of the presynaptic active zones, the sites at which ACh is thought to be released. LEMS is generally believed to be due to circulating antibodies directed specifically at the Ca2+ channels located at or near the active zone of motor nerve terminals (P/Q-type) and hence involved in the release of ACh. However, other presynaptic proteins have also been postulated to be targets of the autoantibodies. LEMS has a high degree of coincidence (approximately 60%) with small cell lung cancer; the remaining 40% of patients with LEMS have no detectable tumor. Diagnosis of LEMS relies on characteristic patterns of electromyographic changes; these changes are observable at neuromuscular junctions of muscle biopsies from patients with LEMS. In the majority of LEMS patients, those having detectable tumor, the disease is thought to occur as a result of immune response directed initially against voltage-gated Ca2+ channels found on the lung tumor cells. In these patients, effective treatment of the underlying tumor generally causes marked improvement of the symptoms of LEMS as well. Animal models of LEMS can be generated by chronic administration of plasma, serum or immunoglobulin G to mice. These models have helped dramatically in our understanding of the pathogenesis of LEMS. This "passive transfer" model mimics the electrophysiological and ultrastructural findings seen in muscle biopsies of patients with LEMS. In this model, we have shown that the reduction in amplitude of Ca2+ currents through P/Q-type channels is followed by "unmasking" of an L-type Ca2+ current not normally found at the motor nerve terminal which participates in release of ACh from terminals of mice treated with plasma from patients with LEMS. It is unclear what mechanisms underlie the development of this novel L-type Ca2+ current involved in release of ACh at motor nerve terminals during passive transfer of LEMS.     

Helicobacter pylori Infection and Gastric Autoimmune Diseases: Is There a Link?

Background. Helicobacter pylori is thought to be involved in atrophic body gastritis. We explored the prevalence of H. pylori infection in asymptomatic subjects with gastric parietal cell antibodies, as well as in patients with pernicious anemia, to evaluate a possible role of H. pylori gastric infection in gastric autoimmunity. Patients and Methods. We studied 79 consecutive asymptomatic subjects with parietal cell antibodies, 24 patients with pernicious anemia, and 66 parietal cell antibody‐negative controls. All patients underwent gastric biopsies for histology and detection of H. pylori. Red blood cell count and volume, serum levels of gastrin, pepsinogen I, iron, folic acid, vitamin B₁₂, and circulating antibodies to H. pylori and to intrinsic factor were also determined. Results. We found an atrophic body gastritis in 14 of the 79 asymptomatic subjects with parietal cell antibodies (18%) and in 2 of the 66 controls (3%) (p = .01). Mean levels of gastrin were increased (p < .0001), while those of pepsinogen were reduced (p < .001) compared with controls. H. pylori was identified at the gastric level and/or circulating anti‐H. pylori antibodies were detected in 46 parietal cell antibody‐positive subjects (58%) compared with 26 controls (39%) (p = .03). In patients with pernicious anemia we found an atrophic body gastritis in 18 of 24 cases (75%) (p < .001 vs. controls). Mean levels of gastrin were markedly increased (p < .0001) and those of pepsinogen I decreased (p < .0001) relative to controls. Only five of these patients (21%) had evidence of H. pylori infection compared with 46 of the parietal cell antibody‐positive subjects (58%) (p = .003) and 26 of the controls (39%). Considering all patients with gastric autoimmunity (i.e. with parietal cell antibodies and/or with pernicious anemia), H. pylori was found in 44 of 72 of those without atrophy (61%) but in 6 of 31 with gastric body atrophy (19%) (p < .001), indicating that H. pylori infection is greatly reduced when gastric acid secretion decreases. Conclusions. The frequent detection of H. pylori infection in subjects with early gastric autoimmunity, indicated by the presence of parietal cell antibodies, suggests that H. pylori could have a crucial role in the induction and/or the maintenance of autoimmunity at the gastric level.     

Diversity of amyloid beta protein fragment [1-40]-formed channels

1. The lipid bilayer technique was used to characterize the biophysical and pharmacological properties of several ion channels formed by incorporating amyloid beta protein fragment (AP) 1–40 into lipid membranes. Based on the conductance, kinetics, selectivity, and pharmacological properties, the following AP[1–40]-formed ion channels have been identified: (i) The AP[1–40]-formed bursting fast cation channel was characterized by (a) a single channel conductance of 63 pS (250/50 mM KCl cis/trans) at +140 mV, 17 pS (250/50 mM KCl cis/trans) at –160 mV, and the nonlinear current–voltage relationship drawn to a third-order polynomial, (b) selectivity sequence P _K > P _Na > P _Li = 1.0:0.60:0.47, (c) P_o of 0.22 at 0 mV and 0.55 at +120 mV, and (d) Zn²⁺-induced reduction in current amplitude, a typical property of a slow block mechanism. (ii) The AP[1–40]-formed spiky fast cation channel was characterized by (a) a similar kinetics to the bursting fast channel with exception for the absence of the long intraburst closures, (b) single channel conductance of 63 pS (250/50 KCl) at +140 mV 17 pS (250/50 KCl) at –160 mV, the current–voltage relationship nonlinear drawn to a third-order polynomial fit, and (c) selectivity sequence P _Rb > P _K > P _Cs > P _Na > P _Li = 1.3:1.0:0.46:0.40:0.27. (iii) The AP[1–40]-formed medium conductance channel was charcterized by (a) 275 pS (250/50 mM KCl cis/trans) at +140 mV and 19 pS (250/50 mM KCl cis/trans) at –160 mV and (b) inactivation at V_ms more negative than –120 and more positive than +120 mV. (iv) The AP[1–40]-formed inactivating large conductance channel was characterized by (a) fast and slow modes of opening to seven multilevel conductances ranging between 0–589 pS (in 250/50 mM KCl) at +140 mV and 0–704 pS (in 250/50 mM KCl) at –160 mV, (b) The fast mode which had a conductance of <250 pS was voltage dependent. The inactivation was described by a bell-shaped curve with a peak lag time of 7.2 s at +36 mV. The slow mode which had a conductance of >250 pS was also voltage dependent. The inactivation was described by a bell-shaped curve with a peak lag time of 7.0 s at –76 mV, (c) the value of P _K/P _choline for the fast mode was 3.9 and selectivity sequence P _K > P _Cs > P _Na > P _Li = 1.0:0.94:0.87:0.59. The value of P _K/P _choline for the slow mode was 2.7 and selectivity sequence P _K > P _Na > P _Li > P _Cs = 1.0:0.59:0.49:0.21, and (d) asymmetric blockade with 10 mM Zn²⁺-induced reduction in the large conductance state of the slow mode mediated via slow block mechanism. The fast mode of the large conductance channel was not affected by 10 mM Zn²⁺.2. It has been suggested that, although the bursting fast channel, the spiky fast channel and the inactivating medium conductance channel are distinct, it is possible that they are intermediate configurations of yet another configuration underlying the inactivating large conductance channel. It is proposed that this heterogeneity is one of the most common features of these positively-charged cytotoxic amyloid-formed channels reflecting these channels ability to modify multiple cellular functions.3. Furthermore, the formation of -sheet based oligomers could be an important common step in the formation of cytotoxic amyloid channels.     

The effect of treating seed potato tubers with benzimidazole, imidazole and phenylpyrrole fungicides on the control of rot and skin blemish diseases

Over 6 yr the effectiveness of imazalil, prochloraz and fenpiclonil, applied either alone or in a mixture, in controlling gangrene, dry rot, skin spot and silver scurf on potato tubers in store was compared with that of 2-aminobutane and thiabendazole. An assessment was also made of their efficiency in controlling the development of these diseases on the daughter tubers of seed tubers treated at harvest or before planting. Overall, 2-aminobutane was more effective in controlling gangrene (Phoma foveata) in store than the spray-applied fungicides. Deposits of imazalil, thiabendazole and fenpiclonil were greater when sprays were applied with an electrostatic sprayer than with a hydraulic sprayer. The opposite was found with the mixture of prochloraz Mn and tolclofos-methyl. More effective gangrene control was associated with the highest deposits. Fenpiclonil and the mixture of thiabendazole and imazalil were more effective in controlling dry rot (Fusarium solani var. coeruleum) than imazalil alone. The development of dry rot was, however, increased by 2-aminobutane treatment on eight out of 14 stocks. 2-aminobutane gave the greatest reduction (83%) in the severity of skin spot during storage whereas thiabendazole alone, and the mixture of thiabendazole and imazalil, gave mean reductions of 70% and 65% respectively. This mixture and fenpiclonil gave the greatest reduction in the severity of silver scurf although, in general, reductions in silver scurf with fungicide treatment were less than with skin spot. The type of sprayer used to apply a fungicide did not affect the effectiveness of the fungicides in controlling either skin spot or silver scurf on tubers in store, or on the daughter tubers. The incidence of gangrene and dry rot on daughter tubers was not reduced consistently by fungicide treatment of seed tubers of the six stocks tested. However, the severity of skin spot and silver scurf was reduced by fungicide treatments of all eight stocks but the reduction in disease was greater for skin spot than for silver scurf. All fungicides gave reductions in the severity of skin spot, and fenpiclonil and the mixture of thiabendazole and imazalil were the most effective for silver scurf. The effectiveness of the fungicides in controlling these diseases was similar for seed treated at harvest and that treated before planting.     

Conformational changes and disease - serpins, prions and Alzheimer''s

Some of the most perplexing disorders in medicine are each now known to arise from the conformational instability of an underlying protein. The consequence is a continuum of pathologies with typically a change in fold leading to ordered aggregation and tissue deposition. The serpins provide a structural prototype for these pathologies and give a perspective on the assessment of current proposals as to the conformational basis of both Alzheimer's disease and the transmissible prion encephalopathies.     

Sam68 sequestration and partial loss of function are associated with splicing alterations in FXTAS patients

Fragile X‐associated Tremor/Ataxia Syndrome (FXTAS) is a neurodegenerative disorder caused by expansion of 55–200 CGG repeats in the 5′‐UTR of the FMR1 gene. FXTAS is characterized by action tremor, gait ataxia and impaired executive cognitive functioning. It has been proposed that FXTAS is caused by titration of RNA‐binding proteins by the expanded CGG repeats. Sam68 is an RNA‐binding protein involved in alternative splicing regulation and its ablation in mouse leads to motor coordination defects. Here, we report that mRNAs containing expanded CGG repeats form large and dynamic intranuclear RNA aggregates that recruit several RNA‐binding proteins sequentially, first Sam68, then hnRNP‐G and MBNL1. Importantly, Sam68 is sequestered by expanded CGG repeats and thereby loses its splicing‐regulatory function. Consequently, Sam68‐responsive splicing is altered in FXTAS patients. Finally, we found that regulation of Sam68 tyrosine phosphorylation modulates its localization within CGG aggregates and that tautomycin prevents both Sam68 and CGG RNA aggregate formation. Overall, these data support an RNA gain‐of‐function mechanism for FXTAS neuropathology, and suggest possible target routes for treatment options.