Functional and molecular characterization of an anion exchanger in airway serous epithelial cells

Serous cells secreteCl and HCO₃ and play an importantrole in airway function. Recent studies suggest that aCl/HCO₃ anion exchanger (AE) maycontribute to Cl secretion by airway epithelial cells.However, the molecular identity, the cellular location, and thecontribution of AEs to Cl secretion in serous epithelialcells in tracheal submucosal glands are unknown. The goal of thepresent study was to determine the molecular identity, the cellularlocation, and the role of AEs in the function of serous epithelialcells. To this end, Calu-3 cells, a human airway cell line with aserous-cell phenotype, were studied by RT-PCR, immunoblot, andelectrophysiological analysis to examine the role of AEs inCl secretion. In addition, the subcellular location of AEproteins was examined by immunofluorescence microscopy. Calu-3 cellsexpressed mRNA and protein for AE2 as determined by RT-PCR and Westernblot analysis, respectively. Immunofluorescence microscopy identified AE2 in the basolateral membrane of Calu-3 cells in culture and rattracheal serous cells in situ. InCl/HCO₃/Na⁺-containingmedia, the 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate(CPT-cAMP)-stimulated short-circuit anion current (I_sc) was reduced by basolateral but not byapical application of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid(50 µM) and 4,4'-dinitrostilbene-2,2'-disulfonic acid [DNDS (500 µM)], inhibitors of AEs. In the absence of Na⁺ in thebath solutions, to eliminate the contributions of the Na⁺/HCO₃ andNa⁺/K⁺/2Cl cotransporters toI_sc, CPT-cAMP stimulated a small DNDS-sensitive I_sc. Taken together with previous studies, theseobservations suggest that a small component of cAMP-stimulatedI_sc across serous cells may be referable toCl secretion and that uptake of Cl acrossthe basolateral membrane may be mediated by AE2.

     

Low serum insulin-like growth factor-1 in patients with erectile dysfunction

Objective

Endothelial dysfunction and microvascular damage play a crurical role in the pathogenesis of erectile dysfunction (ED). Insulin-like growth factor-1 (IGF-1) is one of the growth factors that have a wide range of biologic effects. IGF-1 is an important mediator of cell growth, differentiation and transformation in various tissues. The purpose of the current study was to determine the association between IGF-1 levels and ED.

Materials and methods

All men were evaluated for ED and divided into two groups: 80 patients suffering from ED for >?1 year and 80 subjects without ED were enrolled as a control group in this study. Diagnosis of ED was based on the International Index of Erectile Function Score-5. IGF-1 levels were measured in serum by an automated chemiluminescence immunoassay. The relationship between IGF-1 levels and ED scores in patients was statistically evaluated.

Results

The mean age of patients in ED group was 60.4?±?11.3 years and 55.4?±?9.6 in control group. The plasma IGF-1 levels were significantly lower in ED than in control group (96.5?±?38.3 and 132.5?±?53.3 ng/ mL, respectively, P?<?0.001). The IGF-1 levels were positively correlated with ED score (r?=?0.623, P?<?0.01).

Conclusion

In this study serum IGF-1 levels were found to be associated with endothelial dysfunction that predicts ED. Serum IGF-1 level appears to be a specific predictor of ED, and it might be used in early prediction of ED in male population.

     

Genetic polymorphism of human plasminogen.

Using isoelectric focusing (IEF) in polyacrylamide gel of neuraminidase-treated serum or plasma samples and immunofixation or caseinolytic overlay after urokinase activation of gels, a common genetic polymorphism in human plasminogen has been delineated. Two alleles PLGN*A and PLGN*B, were observed with gene frequencies in whites of .69 and .30; in Orientals of .96 and .03; and in blacks of .80 and .18. Several rare alleles were also found. The distribution of phenotypes fits the Hardy-Weinberg equilibrium. Inheritance is autosomal codominant and fits the expectations of Mendelian inheritance. There is fetal synthesis, but no transplacental passage of plasminogen in either direction.     

The third component of complement: covalent attachment of a radioactive sugar to the labile binding site of C3 via the alternative pathway

A complement- (C) fixing particle consisting of agarose beads to which 5-thioglucose was attached by a --S--S-- bond (agarose-thioglucose) was employed to investigate the mechanism of attachment of C3 to surfaces. When whole serum containing [125I] C3 was incubated with agarose-thioglucose, labeled C3b was taken up in a form that was not removed by 2 M NaCl but was released by 10 mM dithiothreitol. Deposition of DTT-releasable C3b was dependent upon the alternative pathway of C activation. Gel electrophoresis of DTT-releasable C3b from similar experiments performed with unlabeled serum and agarose-[3H]thioglucose showed that the liberated C3b contained a molecule of radioactive thioglucose attached to the alpha'-chain by a covalent bond that was stable to mercaptoethanol. We propose that the thioglucose-alpha' chain bond was formed during the course of C activation by a reaction between the "labile binding site" of newly released C3b and the (then) particle-bound sugar. This formulation implies that the reaction by which C3b attaches to 5-thioglucose in this system is the reaction responsible for opsonization by C3b, and that the C3b-linked sugar represents a marker for the labile binding site. Incubation of the particle-bound C3b in serum resulted in the cleavage of the covalently linked alpha'-chain to several smaller polypeptides, the major cleavage product having a m.w. of 70,000.     

Evidence for differing modes of interaction of acriflavine with ultraviolet-induced lesions in an Hcr + bacterial strain

Summary In buffer suspensions of UV-irradiated Escherichia coli B/r WP2 Hcr⁺ (auxotrophic for tryptophan) acriflavine binds to DNA, but this treatment has little effect on killing and results in the appearance of fewer prototrophs on tryptophan-supplemented minimal agar. If plates contain a broth supplement, however, the buffer-acriflavine treatment greatly increases the yield of UV-induced prototrophs; but this increase does not depend on complete binding of acriflavine to the DNA as a whole, since it is observed with contact times too short for this to occur (as short as 20 seconds). The incorporation of acriflavine in both kinds of plating medium increases the yields of prototrophs. The maximum yield is observed when irradiated bacteria are exposed to acriflavine in buffer before they are plated on medium containing both acriflavine and a broth supplement. Thus post-irradiation effects of acriflavine cannot be accounted for in terms of a single mechanism of action. Our results support the suggestion that phenomena classed together as mutation frequency decline may not represent a single specific repair system.