Products of the lipoxygenase pathway in human natural killer cell cytotoxicity

As earlier data suggested the importance of lipoxygenase activation for expression of human NK cell cytotoxicity, four different lipoxygenase inhibitors were tested for suppression of natural killer (NK) cell lysis. All inhibitors were found active at nontoxic concentrations with 50% inhibition at approximately 15 microM for nordihydroguaiaretic acid (NDGA). NK cell lysis could be reconstituted to NDGA-suppressed cells with leukotriene B4 (LTB4), the all-trans isomers 6-trans-LTB4 and 12-epi-6-trans-LTB4, and 20-COOH-LTB4. LTB4 reconstitution was best in the concentration range 1-100 pM and near control levels at both higher and lower concentrations. Herpesvirus Ateles-transformed killer T cells could also be inhibited by NDGA. These data indicate that lipoxygenase activity is required for human NK cell lysis and that several different LTB4-related products can restore NK activity in inhibited cells; they also suggest that the lipoxygenase pathway is present in the killer cell population.     

Inhibition by verapamil of neutrophil responses to formylmethionylleucylphenylalanine and phorbol myristate acetate. Mechanisms involving Ca2+ changes, cyclic AMP and protein kinase C

Verapamil inhibits in human neutrophils the respiratory burst, the secretion and the change of transmembrane potential induced by formylmethionylleucylphenylalanine, a Ca2+-dependent stimulus, and by phorbol myristate acetate, a Ca2+-independent stimulus. Besides the blocking of Ca2+ channels, many mechanisms are responsible for the inhibition of neutrophil responses. In fact, verapamil (i) increases the intracellular cAMP concentration, potentiates the cAMP response induced by the chemotactic peptide and induces the appearance of a cAMP response also when the stimulant is phorbol myristate acetate; (ii) causes a decrease of Ca2+ association to cell membranes, so depleting the pools of exchangeable Ca2+ and depressing the 'Ca2+ response' in terms of rise in [Ca2+]i monitored with Quin 2 and of rapid mobilization from cell membranes monitored by chlorotetracycline fluorescence change; (iii) inhibits the Ca2+-activated phospholipid-dependent protein kinase C. The data, discussed in relation to the biochemical mechanisms of the stimulus-response coupling, are compatible with the hypothesis of an involvement of the activation of protein kinase C as key step in the sequence of transduction events for the induction of many neutrophil functions.     

Cyclic nucleotide phosphodiesterases in somatic and germ cells of mouse seminiferous tubules

The distribution of phosphodiesterase forms in somatic and germ cells, and their variations during testicular development and germ cell differentiation have been investigated. Seminiferous tubules from immature mice and Sertoli cells in culture possessed two enzyme activities which were comparable to forms described for different tissues and species: (a) a calcium-calmodulin-dependent enzyme with high affinity for guanosine 3',5'-(cyclic)-monophosphate (cGMP), and (b) a calcium-calmodulin-independent enzyme with high affinity for adenosine 3',5'-(cyclic)-monophosphate (cAMP) the activity of which increased in cultured Sertoli cells after treatment with FSH or dibutyryl cAMP. Seminiferous tubules from adult animals and germ cells at the meiotic and post-meiotic stage of differentiation possessed two enzyme forms that could be distinguished from those present in somatic cells of the seminiferous tubules: (a) a calcium-calmodulin-dependent form with high affinity for both cAMP and cGMP, similar to forms described in other tissues from different species, and (b) a calcium-calmodulin-independent phosphodiesterase with high affinity for cAMP and present only in post-meiotic cells, previously identified also in germ cells of the rat.     

Construction of a cosmid library of Spirulina platensis as an approach to DNA physical mapping

Abstract A Spirulina platensis gene library has been constructed using cosmid vector pMMB34. The cosmid bank was controlled for its random gene distribution by colony hybridization. Genes were identified using either homologous or heterologous probes of genes involved in photosynthesis (large and small subunit of d -ribulose 1,5-bisphosphate carboxylase, 32 kDa thylakoid protein, α, β subunits of C-phycocyanin) and protein synthesis (elongation factors EF-Tu, EF-G).     

Respiratory mechanics in mechanically ventilated patients with respiratory failure

In 11 mechanically ventilated patients, respiratory mechanics were measured 1) during constant flow inflation and 2) following end-inflation airway occlusion, as proposed in model analysis (J. Appl. Physiol. 58: 1840-1848, 1985. During the latter part of inflation, the relationship between driving pressure and lung volume change was linear, allowing determination of static respiratory elastance (Ers) and resistance (RT). The latter represents in each patient the maximum resistance value that can obtain with the prevailing time constant inhomogeneity. Following occlusion, Ers and RT were also obtained along with RT (min) which represents a minimum, i.e., resistance value that would obtain in the absence of time constant inhomogeneity. A discrepancy between inflation and occlusion Ers and RT was found only in the three patients without positive end-expiratory pressure, and could be attributed to recruitment of lung units during inflation. In all instances Ers and RT were higher than normal. RT(min) was lower in all patients than the corresponding values of RT, indicating that resistance was frequency dependent due to time constant inequalities. Changes in inflation rate did not affect Ers, while RT increased with increasing flow.     

Studies on stimulus-response coupling in human neutrophils. II. Relationships between the effects of changes of external ionic composition on the properties of N-formylmethionylleucylphenylalanine receptors and on the respiratory and secretory responses

Studies were carried out on the mechanism responsible for the enhancement of the respiratory and secretory responses to N-formylmethionylleucylphenylalanine (fMet-Leu-Phe) exhibited by human neutrophils suspended in Na+-free, high-K+ buffered solution. The results demonstrate that: (a) the variation of Na+ concentration in the suspending solution induces in human neutrophils a marked modification of the recognition apparatus for the chemotactic peptide fMet-Leu-Phe, the lack of or low concentration of this ion increasing the number of the receptors and their specific affinity for the ligand; (b) the greater respiratory burst and secretion induced by fMet-Leu-Phe in human neutrophils suspended in Na+-free, high-K+ medium are due to the increased formation of receptor-ligand complexes at the cell membrane; (c) the greater respiratory response is partially due also to a higher efficiency of these receptor-ligand complexes. The molecular mechanism by which Na+ exerts a regulative role on the properties of the recognition apparatus for the chemotactic peptide and its possible significance are discussed.     

"In vitro" effect of CCl4, PG and EDTA on GSH levels at different time of incubation of rat liver homogenates

During CCl4-induced lipid peroxidation GSH content in total homogenate from rat liver falls very rapidly in the first 30 min. of incubation "in vitro". CCl4 does not enhance the decrease in total glutathione (TG) during the incubation time, so GSH loss is mainly due to its oxidation to GSSG. On the contrary PG and EDTA, two substances decreasing lipid peroxidation rate, are able to decrease GSH oxidation, without affecting TG content. At 25 degrees C EDTA and PG completely prevent GSH decrease at pH 7.4, while at pH 6 PG affords only a partial prevention. At 37 degrees C both compounds are able to limit GSH decrease at a large extent. Lipid peroxidation seems to have a great importance in the kinetics of GSH decrease and GSSG formation, at least "in vitro". It is noteworthy that PG which inhibits lipid peroxidation stimulated by CCl4 is also able to limit the high GSH loss observed in the homogenates incubated in the presence of halogeno-alkane.