Effects of K+ channel agonists cromakalim and pinacidil on rat basilar artery smooth muscle cells are mediated by Ca(++)-activated K+ channels

Whole-cell and cell-free inside-out patch-clamp recording techniques were used to examine the actions of potassium channel openers pinacidil and cromakalim in enzymatically isolated smooth muscle cells of rat basilar artery. Delayed rectifier and calcium-dependent potassium currents were identified from the whole-cell recordings. Only the calcium-dependent potassium current was increased by cromakalim and pinacidil. Recordings from inside-out membrane patches revealed a large conductance voltage- and calcium-dependent potassium channel, which was blocked by charybdotoxin but unaffected by ATP less than 10 mM. Cromakalim and pinacidil increased the open probability of this channel. On the basis of these results, we suggest that such drugs, acting on cerebral arterial smooth muscle cell potassium channels, may be of some benefit in the treatment of cerebral vasospasm following subarachnoid hemorrhage.     

Expression of an autoprocessing CAT-HIV-1 proteinase fusion protein: purification to homogeneity of the release 99 residue proteinase

The 99 residue human immunodeficiency virus type 1 proteinase has been expressed in Escherichia coli as part of an autocleaving fusion protein. Expression of the fusion protein is toxic to the host cells, however yields of the released proteinase have been improved by optimising induction nad harvest times to increase culture biomass, and decrease degradation of the proteinase. Soluble proteinase was extracted from these cells by a simple and highly efficient three step process. N-terminal sequence analysis confirms that the enzyme preparation is highly pure and correctly autoprocessed. The proteinase cleaves peptide substrate IGCTLNFPISPIETV between F and P at pH 6.0 with a Km of 310 microM and a Kcat of 14s-1. The enzyme is sensitive to its ionic environment, showing stimulation of activity at high salt concentrations, and shows a pH optimising 5.5.     

Blood group, secretor status and susceptibility to bacterial meningitis

Abstract Epidemiological evidence is summarized for associations of ABO blood group and secretor status with susceptibility to invasive disease due to capsulate organisms responsible for the majority of bacterial meningitis. Host-parasite interactions that might underly these findings are proposed and evidence to support or refute them provided.     

CD4-Immunoglobulin G2 Protects Hu-PBL-SCID Mice against Challenge by Primary Human Immunodeficiency Virus Type 1 Isolates

CD4-immunoglobulin G2 (IgG2) is a fusion protein comprising human IgG2 in which the Fv portions of both heavy and light chains have been replaced by the V1 and V2 domains of human CD4. Previous studies found that CD4-IgG2 potently neutralizes a broad range of primary human immunodeficiency virus type 1 (HIV-1) isolates in vitro and ex vivo. The current report demonstrates that CD4-IgG2 protects against infection by primary isolates of HIV-1 in vivo, using the hu-PBL-SCID mouse model. Passive administration of 10 mg of CD4-IgG2 per kg of body weight protected all animals against subsequent challenge with 10 mouse infectious doses of the laboratory-adapted T-cell-tropic isolate HIV-1_LAI, while 50 mg of CD4-IgG2 per kg protected four of five mice against the primary isolates HIV-1_JR-CSF and HIV-1_AD6. In contrast, a polyclonal HIV-1 Ig fraction exhibited partial protection against HIV-1_LAI at 150 mg/kg but no significant protection against the primary HIV-1 isolates. The results demonstrate that CD4-IgG2 effectively neutralizes primary HIV-1 isolates in vivo and can prevent the initiation of infection by these viruses.     

Infection with respiratory syncytial virus enhances expression of native receptors for non-pilate Neisseria meningitidis on HEp-2 cells

Respiratory virus infections have been suggested to be predisposing factors for meningococcal disease. Respiratory syncytial virus (RSV) affects young children in the age range at greatest risk of disease caused by Neisseria meningitidis. It has been previously shown that glycoprotein G expressed on the surface of RSV-infected HEp-2 cells (a human epithelial cell line) contributed to higher levels of binding of meningococci compared with uninfected cells. The aim of the present study was to examine the effect of RSV infection on expression of surface molecules native to HEp-2 cells and their role in bacterial binding. Flow cytometry and fluorescence microscopy were used to assess bacterial binding and expression of host cell antigens. Some molecules analysed in this study have not been reported previously on epithelial cells. RSV infection significantly enhanced the expression of CD15 (P < 0.05), CD14 (P < 0.001) and CD18 (P < 0.01), and the latter two contributed to increased binding of meningococci to cells but not the Gram-positive Streptococcus pneumoniae.     

The "thermolabile" variant of methylenetetrahydrofolate reductase and neural tube defects: An evaluation of genetic risk and the relative importance of the genotypes of the embryo and the mother

Recent reports have implicated the "thermolabile" (T) variant of methylenetetrahydrofolate reductase (MTHFR) in the causation of folate-dependent neural tube defects (NTDs). We report herein the largest genetic study of NTD cases (n=271) and families (n=218) to date, establishing that, in Ireland, the "TT" genotype is found in 18.8% of cases versus 8.3% of controls (odds ratio 2.57; confidence interval [CI] 1.48-4.45; P=.0005). The maternal and paternal TT genotypes have intermediate frequencies of 13.8% and 11.9%, respectively, indicating that the predominant MTHFR-related genetic effect acts via the TT genotype of the developing embryo. Analysis of the 218 family triads of mother, father, and affected child with log-linear models supports this interpretation, providing significant evidence that the case TT genotype is associated with NTDs (P=.02) but no evidence of a maternal TT genotypic effect (P=. 83). The log-linear model predicted that the risk of NTDs conferred by the case TT genotype is 1.61 (CI 1.06-2.46), consistent with the paramount importance of the case TT genotype in determining risk. There is no compelling evidence for more than a modest additional risk conferred by a maternal TT genotype. These results favor a biological model of MTHFR-related NTD pathogenesis in which suboptimal maternal folate status imposes biochemical stress on the developing embryo, a stress it is ill-equipped to tolerate if it has a TT genotype.     

Smooth muscle development during postnatal growth of distal bronchioles in infant rhesus monkeys.

Development of smooth muscle in conducting airways begins early in fetal life. Whereas the pattern and regulation of smooth muscle differentiation are well-defined, the impact of airway growth on the process is not. To evaluate the transformations in organization during postnatal growth, smooth muscle bundle organization (size, abundance, and orientation) was mapped in five generations of distal airways of infant rhesus monkeys (5 days and 1, 2, 3, and 6 mo old). On the basis of direct measurement of the bronchiole proximal to the terminal bronchiole, length increased by 2-fold, diameter by 1.35-fold, and surface area by 2.8-fold between 5 days and 6 mo of age. Smooth muscle bundle size was greater in proximal bronchioles than in respiratory bronchioles and did not change with age. However, relative bundle size decreased in proportion to airway size as the airways grew. Relative bundle abundance was constant regardless of airway generation or age. The distribution of smooth muscle bundle orientation changed with age in each airway generation, and there were significant changes in the terminal and respiratory bronchioles. We conclude that smooth muscle undergoes marked organizational changes as airways grow during postnatal development.     

The effect of Mycoplasma arthritidis infection on the kinetics of colloidal carbon clearance in mice

Abstract The activity of phagocytes from A/J mice was estimated by the carbon clearance test following injection of Mycoplasma arthritidis . Phagocytic activity was significantly depressed 12 h post-infection ( P =0.001) and returned to normal values at 24 h. For animals examined 2 and 7 days post-infection, the overall phagocytic activity increased significantly ( P <10⁻⁴). Phagocytic activity gradually decreased and returned to that of the control group by the end of the fourth week. The relative weights of liver and spleen were significantly increased from the 2nd day post infection ( P =0.0028 and P =0.0014 respectively) and remained increased until the end of the experiment. The early depressive effect on phagocytic activity may be related to superantigen activity with the production of mediators such as macrophage deactivating factor. The later expansion of the macrophage population might bring about the stimulation of autoreactive clones of T and B cells and be responsible for the chronic arthritis that developed in the mycoplasma treated mice.