Nonselective cation channels activated by the stimulation of muscarinic receptors in mammalian gastric smooth muscle.

Muscarinic receptors play key roles in the control of gastrointestinal smooth muscle activity. However, specific physiological functions of each subtype remain to be determined. Single cell RT-PCR experiments showed that all five subtypes of muscarinic receptors were present in circular smooth muscle cells of the guinea-pig gastric antrum. Nonselective cation channels (NSCC) activated by ACh or CCh are coupled to pertussis toxin (PTX)-sensitive Go protein through m4 subtype as well as m2 and m3 subtypes in guinea-pig stomach. CCh-activated currents (I(CCh)), especially the steady-state I-V relationship of I(CCh) showed a chracteristic U-shaped curve; reversal potential of around 0 mV and inward rectification at around +15 mV and a negative slope conductance at negative potential range. Under physiological conditions, the measured single channel conductance of NSCC was approximately 25 pS. The single channel conductance was modulated by external monovalent and divalent cations including Na+, Cs+, Li+, and Ca2+ through changing both the open probability and unitary conductance. Through the NSCC, Ca2+ can move into the cell from extracellular solution as well as Na+. Calculated fractional Ca2+ current of I(CCh) (f(Ca)) was around 1% at the 2 mM [Ca2+]o and at the 4 mM [Ca2+]o, f(Ca) was 2.3%. Quinidine blocked I(CCh) potently in a reversible manner; IC50 was 0.25 microM. There were two kinds of I(CCh) modulations through Ca(2+)-dependent pathways in guinea-pig gastric smooth muscle cells; 1) Facilitation of I(CCh) via Ca2+/CaM-dependent MLCK pathway, 2) Desensitization of I(CCh) via Ca(2+)-dependent PKC pathway. In the mouse stomach, all seven types of TRPC mRNA were detected with RT-PCR. On the basis of electrophysiological, pharmacological, and molecular biological experiments, we reported the mTRPC5 as a candidate for the NSCC activated by muscarinic stimulation in mouse stomach.     

AC/A ratios in myopic and emmetropic Hong Kong children and the effect of timolol§

Purpose: Caucasian children with myopia have elevated response accommodative vergence to accommodation (AC/A) ratios. The purpose of this study was twofold: to determine if response AC/A ratios vary with refractive error and with myopic progression rate in Hong Kong Chinese children, and to determine the effect of beta‐adrenergic antagonism with topical timolol application on AC/A ratios. Methods: Thirty children aged eight to 12 years participated in the study. All refractive errors were corrected with spectacle lenses. Accommodative responses were measured using a Shin‐Nippon autorefractor and concurrent changes in vergence were assessed using a vertical prism and a Howell‐Dwyer card at three metres and 0.33 metre. Accommodative demand was altered using plus or minus two dioptre lenses and lens‐ and distance‐induced response AC/A ratios were calculated. Measurements were repeated 30 minutes after the instillation of topical timolol maleate (0.5 per cent). Results: AC/A ratios appeared higher in progressing myopic children but the difference was not statistically significant. Timolol application reduced accommodative convergence (AC) in the stable myopes (reduction = ‐3 ± 1.14^A) but not in the emmetropes (0.69 ± 0.9^P) or progressing myopes (0.16 ± 0.43^A) and this difference between refractive groups was statistically s ignificant (F^2,27= 3.766; P= 0.036). However, timolol did not produce a significant change in the accommodative response to positive or negative lenses or response AC/A ratios. Conclusions: We did not find that AC/A ratios in myopic Chinese children were elevated and therefore, it is unlikely that elevated AC/A ratios are responsible for the high levels of myopia that occur in Hong Kong. The finding that timolol reduced AC in the stable myopes suggests that the autonomic control of accommodative convergence in these children may be different from that in emmetropic children and those with progressing myopia.     

Activation of guinea pig eosinophil respiratory burst by leukotriene B4: role of protein kinase C

Leukotriene B4 (LTB4) and the protein kinase C activator, 4-beta-phorbol dibutyrate (PDBu), both induced a pronounced and concentration-dependent stimulation of hydrogen peroxide (H2O2) generation by purified guinea pig peritoneal eosinophils in the concentration range 1 nM-1 microM. The LTB4 response was inhibited competitively by the specific LTB4 receptor antagonist, U-75302, with a KB of 25 nM, while the concentration-response curves for both stimuli were shifted rightwards (3.8-fold and 2.8-fold for LTB4 and PDBu, respectively) by the competitive protein kinase C inhibitor, 1-O-hexadecyl-2-O-methylglycerol at a concentration of 300 microM. LTB4 appears, therefore, to induce respiratory burst in eosinophils via a receptor-mediated mechanism involving protein kinase C.     

Thrombus of the ascending aorta: a rare cause of myocardial infarction.

We present two cases of a thrombus in the ascending aorta causing an acute myocardial infarction (AMI) and review the 10 other cases previously reported in the literature. This life-threatening condition appears to be more common in female smokers in their fifth decade. Suspicion should be raised in individuals at low risk for atherosclerotic disease with coronary angiographic findings not in keeping with the clinical presentation. The diagnosis may be obtained by transesophageal echocardiography, and we generally recommend surgical thrombectomy.     

Expression of transforming growth factor-beta1 and transforming growth factor-beta receptors in hepatocellular carcinoma and dysplastic nodules.

In this study we analyzed by immunohistochemistry the expression of TGF-beta1 protein and TGF-beta receptors I and II in 4 low-grade dysplastic nodules, 2 high-grade dysplastic nodules, 6 early, 22 small, and 62 advanced hepatocellular carcinomas. The expression of TGF-beta1 protein by hepatocytes was decreased in advanced hepatocellular carcinoma compared with small or early hepatocellular carcinoma(P < .05). Frequent and intense staining of TGF-beta1 protein was noted in the sinusoidal endothelium of advanced hepatocellular carcinomas despite of its decreased staining in hepatocellular carcinoma cells. Reduced expression of TGF-beta receptors I and II compared with surrounding nontumorous tissue were noted from the early hepatocellular carcinoma stage suggesting that down-regulation of TGF-beta receptors is correlated with progression from premalignant to malignant phenotype. Reduced expression of both TGF-beta1 and TGF-beta receptor II in neoplastic hepatocytes were also significantly correlated with increased tumor size and increased proliferative activity(P < .05). These findings suggest that during hepatocarcinogenesis, the inhibitory effects of TGF-beta1 protein on hepatocellular carcinoma cells is outweighed by its effects on stromal elements, which, overall, contributes indirectly to a tumor growth stimulatory environment. Also, the growth-inhibitory effects of TGF-beta1 may have been further negated by reduced TGF-beta receptors on hepatocellular carcinoma cells.     

Functional tyrosine kinase inhibitor profiling: a generally applicable method points to a novel role of platelet-derived growth factor receptor-beta in tuberous sclerosis

A ubiquitous herpesvirus that establishes life-long infection, the Epstein-Barr virus (EBV) has yielded little insight into how a single agent in general accord with its host can produce diverse pathologies ranging from oral hairy leukoplakia to nasopharyngeal carcinoma, from infectious mononucleosis to Hodgkin's disease (HD) and Burkitt's lymphoma. Its pathogenesis is further confounded by the less than total association of virus with histologically similar tumors. In other viral systems, defective (interfering) viral genomes are known to modulate outcome of infection, with either ameliorating or intensifying effects on disease processes initiated by prototype strains. To ascertain whether defective EBV genomes are present in HD, we examined paraffin-embedded tissue from 56 HD cases whose EBV status was first determined by cytohybridization for nonpolyadenylated EBV RNAs (EBERs). Using both standard polymerase chain reaction (PCR) and PCR in situ hybridization, we successfully amplified sequences that span abnormally juxtaposed BamHI W and Z fragments characteristic of defective heterogeneous (het) EBV DNA from 10 of 32 (31%) EBER-positive tumors. Of 24 EBER-negative HD, 8 yielded PCR products indicating presence of het EBV DNA. Two of these contained defective EBV in the apparent absence of the prototype virus. Of the 42 tumors analyzed for defective EBV by both PCR techniques, there was concordance of results in 38 (90%). Detection of defective EBV genomes with the potential to disrupt viral gene regulation suggests one mechanism for pathogenic diversity that may also account for loss of prototypic EBV from individual tumor cells.     

B-cell repertoire specific for an unfolded self-determinant of mouse lysozyme escape tolerance and dominantly participate in the autoantibody response

We previously found that autoantibodies against mouse lysozyme (ML) were strongly induced in normal BALB/c mice when immunized with mutant ML that has triple mutations rendering the dominant T-cell epitope of hen egg lysozyme (HEL), HEL 107-116. As T cells specific for HEL 107-116 were primed in these mice, the anti-ML immunoglobulin G (IgG) responses would be the result of collaborations between autoreactive B cells specific for ML and T cells specific for HEL 107-116. Serum IgG responses against ML were dominantly focused on the ML 14-69 region, indicating that B cells responding to the epitope escape tolerance. In the present study, we prepared several monoclonal antibodies (mAbs) specific for ML 14-69 and examined their antigen specificities in detail, to characterize the nature of the remaining B-cell repertoire specific for ML. mAbs specific for ML 14-69 interacted weakly with soluble, native ML, but the interactions were strengthened by denaturation of ML. The apparent affinity constants between these mAbs and ML showed an increase, ranging from six- to 80-fold, by denaturation of ML. Therefore, these mAbs were more specific for the denatured determinant than for the determinant in the native structure. These results indicate that a substantial number of autoreactive B cells, specific for the unfolded conformation of ML, escape tolerance and are dominantly involved in the autoantibody response to ML. Our finding provides important information to understand the naturally occurring autoreactive B-cell repertoire in normal mice.     

Detection of highly pathogenic and low pathogenic avian influenza subtype H5 (Eurasian lineage) using NASBA

Nucleic acid sequence-based amplification (NASBA) is a technique that allows the rapid amplification of specific regions of nucleic acid obtained from a diverse range of sources. It is especially suitable for amplifying RNA sequences. A NASBA technique has been developed that allows the detection of avian influenza A subtype H5 from allantoic fluid harvested from inoculated chick embryos. The amplified viral RNA is detected by electrochemiluminescence. The NASBA technique described below is rapid and specific for the identification of influenza A subtype H5 viruses of the Eurasian lineage. More importantly, it can be used to distinguish highly pathogenic and low pathogenic strains of the H5 subtype.     

Is there an effect of monocular deprivation on the proportions of X and Y cells in the cat lateral geniculate nucleus?

Summary The proportions and receptive field properties of X and Y cells in the A and A1 layers of the lateral geniculate nucleus (LGN) were studied in monocularly deprived cats. Contrary to previous reports, we found that there was no change in the relative number of Y cells in the geniculate layers driven by the deprived eye. There was also no marked change in the spatial resolution of X or Y cells driven from the deprived eye as compared to the cells driven from the normally experienced eye. In these same cats, the visual evoked potential from stimulation of the deprived eye with grating patterns was markedly reduced in amplitude. Furthermore, the cell bodies of the cells in the LGN driven by the deprived eye had shrunk. Therefore, these usual consequences of monocular deprivation are not necessarily associated with a loss of geniculate Y cells.