Ensemble noise and current relaxation analysis of K+ current in single isolated salivary acinar cells from rat

The K⁺ channel in rat parotid gland acinar cells were investigated by ensemble current noise analysis in single isolated cells employing the giga-seal whole cell current recording mode. Sets of 20–40 identical de- and hyperpolarization voltage steps were applied and the resultant current records were processed by computer to obtain the mean and the variance of the current. The time-course of the mean current could be fitted by the sum of two exponentials, suggesting a 3-state model. The simplest plausible hypothesis is a model with one open and two closed states. Assuming this model, the relationship between the variance (₂) and the mean current (I) could be fitted by the function ₂/I=i–I/N. The estimated single channeli/V-relations were similar to those taken from single channel current recordings, and the size of the population of channels per cell (N) was 76±26 (n=12). The validity of the model was tested by a successful simulation of the time-course of the variance.     

Congenital malformation of the feet with low body height

Among 75 members of a Danish family, 12 were found with a syndrome not previously described. Clinically, the syndrome consists of low body height and rigid flat feet, with weight-bearing pain in the feet. Radiologically, the deformation of the feet is a medial synostosis between the talus and the calcaneus combined with ankle joint dysplasia. The cause of the syndrome is most probably an autosomal dominant gene with complete penetrance. No linkage was found of the gene to 18 marker genes.     

Circulating immune complexes in ulcerative colitis.--II. Correlation with serum protein concentrations and complement conversion products

Several serum proteins were quantified in twenty-two patients with active ulcerative colitis, and the findings were related to disease activity and occurrence of circulating immune complexes (IC). Conversion of C3 was significantly more frequent in the IC-positive group (eight patients) as compared to the IC-negative group (fourteen patients). Factor B was demonstrable in fifteen out of the twenty-two patients and seven out of the eight IC-positive patients had detectable levels of factor B. There was no difference between the IC-positive and the IC-negative group as regards serum concentrations of the complement factors C3, C4 and factor B, or serum orosomucoid, albumin, IgM and IgG. In contrast, the serum IgA levels tended to be reduced in the IC-positive group. C3 and factor B were significantly elevated in four patients with severe disease activity. In addition, C3, factor B and C4 concentrations showed a positive correlation to the serum orosomucoid levels. The serum concentrations of orosomucoid and albumin were inversely correlated to each other.     

Protein kinase C activation modulates tumour necrosis factor-alpha priming of human neutrophils for zymosan-induced leukotriene B4 release.

Neutrophil (PMN) activation by the yeast component zymosan involves the complement receptor type 3 (CD11b/CD18). Recombinant human tumour necrosis factor-alpha (rhTNF-alpha) augmented the zymosan-stimulated leukotriene B4 (LTB4) release from PMN, reaching a fourfold increase at 10(-9) M. Co-incubation of PMN with 10(-9) M rhTNF-alpha and staurosporine resulted in a further dose-dependent increase, which became significantly greater than a purely additive effect at a staurosporine concentration of 10 nM. This synergy was maintained at all doses of staurosporine tested. In addition, doses of phorbol 12-myristate 13-acetate (PMA) that do not activate protein kinase C (PKC) (below 10(-9) M) also augmented the zymosan-stimulated release of LTB4. However, doses of PMA above 10(-9) M progressively inhibited the response to levels below that of zymosan alone. Staurosporine at 50 nM completely prevented, and 10(-9) M rhTNF-alpha partially but significantly (P less than 0.02 at 10(-8) M PMA, P less than 0.01 at 10(-7) M PMA) reversed, this high-dose PMA inhibition. PKC activation thus opposes the priming effect of rhTNF-alpha on neutrophils, while PKC inhibition may enhance the ability of rhTNF-alpha to prime PMN for zymosan activation. The combined effect of rhTNF-alpha and staurosporine suggests an intracellular synergy rather than simply a direct action due to increased zymosan receptor expression. Thus there appear to be mechanisms whereby the responses of neutrophils may be augmented without activating PKC. Indeed, kinase activation may even exert a degree of feedback control that is antagonized by rhTNF-alpha treatment.     

Differences in cytokine secretion by intestinal mononuclear cells, peripheral blood monocytes and alveolar macrophages from HIV-infected patients.

Mononuclear cells of the lamina propria (LpMNC), isolated from endoscopically taken biopsies of the large bowel from AIDS patients, were analysed for their ability to secrete tumour necrosis factor-alpha (TNF-alpha), IL-1 beta and IL-6. Stimulation of LpMNC from normal controls with pokeweed mitogen (PWM) led to a time- and dose-dependent enhancement of TNF-alpha, IL-1 beta and IL-6 secretion. In contrast, PWM stimulation of LpMNC from AIDS patients resulted in only a small increase in TNF-alpha release. Constitutive secretion of IL-1 beta and IL-6 in these patients was already increased to the concentration range of stimulated cells from normal controls and could not be further increased, probably due to maximal in vivo stimulation. Secretion of TNF-alpha, IL-1 beta and IL-6 by peripheral blood monocytes (PBM) and alveolar macrophages from AIDS patients was elevated with or without stimulation compared with normal controls. Obviously, the regulation of TNF-alpha secretion is dependent on the microenvironment. Since it is known that interferon-gamma (IFN-gamma) may induce the production of TNF-alpha, the secretion of this cytokine was examined. Release of IFN-gamma was constitutively and under stimulation lowered in LpMNC from AIDS patients compared with normal controls. Addition of IFN-gamma to LpMNC did not result in enhanced TNF-alpha secretion. Our data indicate a defective function of intestinal mononuclear cells in AIDS patients as shown by the diminished TNF-alpha secretion.     

Essential role for cyclic AMP and its receptor protein in Yersinia enterocolitica virulence

Insertion mutations were isolated in cya and crp of Yersinia enterocolitica, which encode adenylate cyclase and the cyclic AMP (cAMP) receptor protein (CRP). The cya and crp mutants were affected for the production of proteins exported by the Ysc, Ysa, and flagellar type III secretion systems (TTSS). Protein production by each TTSS was restored when the respective mutation was complemented by a plasmid-encoded copy of the wild-type gene. Both cya and crp mutants exhibited reduced virulence for orally infected BALB/c mice in a 50% lethal dose analysis. Examination of bacterial survival in host tissues showed that cya and crp mutants colonized Peyer's patches and, to a lesser extent, mesenteric lymph nodes. However, the mutants did not appear to disseminate to the liver and spleen of infected mice. An initial examination of the effectiveness of Y. enterocolitica cya and crp mutants to stimulate protective immunity against subsequent challenge with virulent bacteria in mice was promising. The results indicate that the cAMP-CRP regulatory system is required for Y. enterocolitica virulence.     

Frequent polymorphism of the mitochondrial DNA polymerase gamma gene (POLG) in patients with normal spermiograms and unexplained subfertility

BACKGROUND: Male fertility largely depends on the quality of sperm production, which may be affected by environmental and genetic factors. In this study, we explored a possible role of the polymerase gamma (POLG) gene polymorphism, recently reported to be associated with male infertility in some populations. METHODS: The polymorphic CAG repeat (usually 10 codons long) in the POLG gene was studied in 1298 male subjects: 429 patients with infertility/subfertility, and 869 controls (495 men from the general population with unknown fertility and 374 recent fathers). In all subjects, the POLG polymorphism was assessed in relation to their semen quality, and--in the fertile controls--with biological fecundity measured as waiting time-to-pregnancy (TTP) for the couples. In the patients lacking the common POLG allele, the outcome of the assisted reproductive techniques (ART) for the couples was evaluated. RESULTS: The absence of one (10/ not equal to 10) or both common POLG alleles (not equal to 10/not equal to 10) was more frequent among the subfertile patients than among fertile controls (P=0.021 and P=0.04 respectively). The estimated predictive value for infertility in a man homozygous for the POLG polymorphism was 15.5% (95% CI: 4.8-51%). There was a positive association with sperm concentration: 14.3% of the normospermic subfertile patients were homozygous for the absence of the common POLG allele (not equal to 10/not equal to 10), in comparison with 2.3% of unselected controls (P=0.001) and 0.9% of the fertile men (P=0.0001). No association with sperm motility, morphology and TTP was found. Spermatozoa of the three not equal to 10/not equal to 10 patients treated with IVF retained the ability to penetrate the egg, but the fertilization rate was low. Nine homozygous not equal to 10/ not equal to 10 patients were treated with ICSI, resulting in pregnancy in seven couples. CONCLUSIONS: The POLG gene polymorphism should be considered as a possible contributing factor in patients with unexplained subfertility and normal spermiograms. The oocyte penetration ability of sperm may be partially impaired in the not equal to 10/not equal to 10 patients but most of them can be successfully treated with ICSI.     

Secretory transmembrane potentials in acinar cells from the cat submandibular gland during perfusion with a chloride-free sucrose solution

Summary The cat submandibular gland was perfused with a normal NaCl Locke solution and a chloride-free sucrose solution. The numerical increase in acinar membrane potential (secretory potential) was recorded after intra-arterial injection of acetylcholine.There was no significant difference between the size of the secretory potentials recorded during perfusion with the sucrose solution [23.6 mV±1.4 (n=23)] and the size of those recorded during the control periods [20.6 mV±1.2 (n=24)].The maximal value of the membrane potential after injection of acetylcholine was higher [51.8 mV±2.4 (n=23)] during perfusion with the sucrose solution than during the control periods [44.8 mV±1.8 (n=22)].The results show that a pump transporting chloride into the acinar cells cannot be responsible for the generation of the secretory potentials. The results are best accounted for by assuming that an outward passive transport of potassium, being partly short-circuited by an inward passive sodium transport, is responsible for the change in membrane potential after stimulation with acetylcholine.Supported by the Danish State Research Foundation and Johann and Hanne Weimann's legate.