Compartmentation of high-energy phosphates in resting and beating heart cells

The subcellular distribution of ATP, ADP, creatine phosphate and creatine has been analyzed by fast detergent fractionation of isolated frog heart cells. Digitonin fractionation (0.5 mg/ml, 10 s at 2 degrees C in 20 mM 4-morpholinepropanesulfonic acid/3 mM EDTA/230 mM mannitol medium) was used to separate mitochondria and myofilaments from cytosol. To separate myofilaments from the other cellular compartments. Triton X-100 was used (2%, 15 s in the same medium as digitonin). For either resting or beating cells the total cellular contents of ATP, ADP, creatine phosphate and creatine was similar, nevertheless the O2 consumption was 6-times higher. The compartmentation of these metabolites was also identical. Myofilaments contain 1.1 nmol ADP per mg total cellular proteins. In the cytosolic compartment the metabolite concentrations, all measured in nmol per mg total cellular proteins, were: ATP, 13; ADP, 0.25-0.05; creatine phosphate, 18.5 and creatine, 14. This indicated that the reaction catalyzed by creatine kinase was in a state of (or near) equilibrium.     

Effects of antimicrotubular agents in cAMP production and in steroidogenic response of isolated rat Leydig cells

In dispersed rat Leydig cells, colchicine was found to stimulate basal cAMP production and testosterone secretion in a dose and time-dependent manner, but to a lesser extent than LH. However, these drugs are unable to stimulate adenylate cyclase activity in plasma membranes isolated from these cells. The amount of testosterone secreted at 150 min under the influence of colchicine and LH added simultaneously was not different from the amount produced during stimulation by LH alone. It is only after exposure of the cells for 1 hr to colchicine that the accumulation of cAMP in response to LH was inhibited; furthermore, both intracellular and medium testosterone accumulation in response to the hormone were reduced. Similar effects were observed with two other alkaloids, vinblastine and podophyllotoxin. The three drugs also inhibited the stimulation of testosterone secretion by 8-Br-cAMP or choleratoxin. These studies suggest that the state of microtubule polymerization and/or tubulin can influence the process of steroidogenesis in rat Leydig cells.     

Decomposition of three halophytes in different habitats of an eastern scheldt salt marsh

Summary A 20.5-month study was undertaken to determine detrital processing of the halophytesSpartina anglica, Elytrigia pungens, andHalimione portulacoides in three different habitats of an estuarine salt marsh in the South-West Netherlands. Decomposition was measured using litter-bags of three different mesh sizes to partition the effects of different faunal groups on decomposition. From April 1980 through October 1981 litter-bags were sampled regulary from a creek, the upper marsh, and from a plant-debris belt on the higher marsh. Dry weights and nutritive values were measured and animals were counted. Mainly rates of loss are reported here. Zonal differences were significant. At first, decomposition in the creek was most rapid. After two months the processes in the creek slowed down because of the trapping of silt by the bags, which probably simulated the natural course of the decomposition process in the water. Decomposition on the marsh followed the most regular pattern, while in the plant-debris belt the pattern was very irregular. Population dynamics of microfaunal organisms supported these findings. In the plant-debris belts loss rates seem to be higher than on the marsh, because of the influence of detritivorous macrofaunal organisms. The loss rates of the three plant species differed significantly.Halimione decomposed fastest, especially in the beginning, and in the plant-debris habitat. On the upper marsh and in the plant-debris belt the loss rates ofSpartina seem to be a little higher than those ofElytrigia.Communication No. 233, Delta Institute for Hydrobiological Research, Yerseke, The Netherlands.     

Mechanism of endotoxin-induced reduction in the number of beta-adrenergic receptors in dog livers: role of phospholipase A

The role of phospholipase A on the endotoxin-induced reduction in the number of beta-adrenergic receptors in dog liver plasma membranes was investigated. The results show that digestion of control liver plasma membranes with exogenous phospholipase A2 (0.2 unit/200 micrograms protein) decreased the specific binding of (-)-[3H]dihydroalprenolol by 37.3% (P less than 0.01) and reduced the number of receptor sites by 31.7% (P less than 0.05). These decreases in the specific binding and the number of beta-adrenergic receptors were completely reversible by the addition of phosphatidylcholine (0.2 mM). Endotoxin administration (2 hr postendotoxin) decreased the specific binding by 36% (P less than 0.05) and reduced the number of beta-adrenergic receptors by 33% (P less than 0.05), and these decreases were completely reversible by the addition of 0.2 mM phosphatidylcholine. Digestion of control liver membranes with exogenous phospholipase A2 decreased phosphatidylcholine and phosphatidylethanolamine levels by 50.6 and 51.2%, respectively, but increased lysophosphatidylcholine and lysophosphatidylethanolamine levels by 12- and 8.4-fold, respectively. Endotoxin administration decreased phosphatidylcholine and phosphatidylethanolamine contents by 21.4 and 23.8%, respectively, but increased lysophosphatidylcholine and lysophosphatidylethanolamine contents by 2.1- and 1.4-fold, respectively. In addition, endotoxin administration increased endogenous phospholipase A activity by 73.5%. Based on these results, it is suggested that the decreases in the specific binding and the number of beta-adrenergic receptors in dog livers during endotoxic shock are a result of phospholipase A activation.     

High Rates of Obesity and Chronic Disease Among United Methodist Clergy

We used self‐reported data from United Methodist clergy to assess the prevalence of obesity and having ever been told certain chronic disease diagnoses. Of all actively serving United Methodist clergy in North Carolina (NC) 95% (n = 1726) completed self‐report height and weight items and diagnosis questions from the Behavioral Risk Factor Surveillance Survey (BRFSS). We calculated BMI categories and diagnosis prevalence rates for the clergy and compared them to the NC population using BRFSS data. The obesity rate among clergy aged 35–64 years was 39.7%, 10.3% (95% CI = 8.5%, 12.1%) higher than their NC counterparts. Clergy also reported significantly higher rates of having ever been given diagnoses of diabetes, arthritis, high blood pressure, angina, and asthma compared to their NC peers. Health interventions that address obesity and chronic disease among clergy are urgently needed.     

COMPARISON OF ISOZYMES AMONG TYPHA SPECIES IN THE EASTERN UNITED STATES

Biochemical phenotsypes of four taxa of Typha from the eastern United States were determined by starch gel electrophoresis. The isozyme banding patterns of T. latifolia, T. angustifolia and T. domingensis are distinct and allow unambiguous species identification when morphological characters are inadequate or unsuitable. The fourth form, T. glauca, is not an F₁ hybrid, but it does appear to be intermediate between T. latifolia and T. angustifolia. The status of T. glauca and evolutionary relationships among the four forms may now be clarified by additional sampling because of the distinct and relatively invariant isozyme banding patterns which are described.     

Sphingosine kinase 1 regulates HMGB1 translocation by directly interacting with calcium/calmodulin protein kinase II-δ in sepsis-associated liver injury

Previously, we confirmed that sphingosine kinase 1 (SphK1) inhibition improves sepsis-associated liver injury. High-mobility group box 1 (HMGB1) translocation participates in the development of acute liver failure. However, little information is available on the association between SphK1 and HMGB1 translocation during sepsis-associated liver injury. In the present study, we aimed to explore the effect of SphK1 inhibition on HMGB1 translocation and the underlying mechanism during sepsis-associated liver injury. Primary Kupffer cells and hepatocytes were isolated from SD rats. The rat model of sepsis-associated liver damage was induced by intraperitoneal injection with lipopolysaccharide (LPS). We confirmed that Kupffer cells were the cells primarily secreting HMGB1 in the liver after LPS stimulation. LPS-mediated HMGB1 expression, intracellular translocation, and acetylation were dramatically decreased by SphK1 inhibition. Nuclear histone deacetyltransferase 4 (HDAC4) translocation and E1A-associated protein p300 (p300) expression regulating the acetylation of HMGB1 were also suppressed by SphK1 inhibition. HDAC4 intracellular translocation has been reported to be controlled by the phosphorylation of HDAC4. The phosphorylation of HDAC4 is modulated by CaMKII-δ. However, these changes were completely blocked by SphK1 inhibition. Additionally, by performing coimmunoprecipitation and pull-down assays, we revealed that SphK1 can directly interact with CaMKII-δ. The colocalization of SphK1 and CaMKII-δ was verified in human liver tissues with sepsis-associated liver injury. In conclusion, SphK1 inhibition diminishes HMGB1 intracellular translocation in sepsis-associated liver injury. The mechanism is associated with the direct interaction of SphK1 and CaMKII-δ.Subject terms: Hepatotoxicity, Sepsis     

The ilvIH operon of Escherichia coli K-12. Identification of the gene products and recognition of the translational start by polypeptide microsequencing

The ilvI and ilvH gene products were identified physically by electrophoretic analysis of in vivo-labelled polypeptides produced in minicells from plasmids carrying the wild-type ilvIH operon of Escherichia coli K-12 and derivatives of it. An analysis of the distribution of methionine residues in the amino-terminal portion of micro-quantities of the ilvI product eluted from gel showed that the translational start of the ilvI gene is the promoter-proximal one of three putative methionine codons predicted from the DNA sequence.     

Characterisation of Trypanosoma cruzi populations by dna polymorphism of the cruzipain gene detected by single-stranded dna conformation polymorphism (SSCP) and direct sequencing

Fifty fresh isolates of Trypanosoma cruzi from Triatoma dimidiata vectors and 31 from patients with Chagas disease were analysed for DNA polymorphisms within the 432-bp core region of the cruzipain gene which encodes the active site of cathepsin L-like cystein proteinase. The cruzipain gene showed signs of polymorphism consisting of four different DNA sequences in Central and South American isolates of T. cruzi. The PCR fragments of Guatemalan isolates could be divided into three groups, Groups 1, 2 and 3, based on different patterns of single-stranded DNA conformation polymorphism. All of the strains isolated from Brazil, Chile, and Paraguay, except for the CL strain, showed a Group 4 pattern. Two to four isolates from each group were analysed by cloning and sequencing. A silent mutation occurred between Groups 1 and 2, and five nucleotides and two aa substitutions were detected between Groups 1 and 3. The DNA sequence of Group 4 contained five nucleotides and one aa substitution from Group 1. All of the DNA sequences corresponded well with the single-stranded DNA conformation polymorphism. The Group 1 isolates, the majority in the Guatemalan population (70/81, 86.4%), were isolated from both triatomines and humans, but Group 3 were isolated only from humans. Moreover, the Group 2 isolates were detected only in triatomine vectors (9/50; 18%), but never in humans (0/32, P<0.05) suggesting that this group has an independent life-cycle in sylvatic animals and is maintained by reservoir hosts other than humans.     

Evolution of green lacewings (Neuroptera: Chrysopidae): a molecular supermatrix approach

We present a time‐calibrated phylogeny of the charismatic green lacewings (Neuroptera: Chrysopidae). Previous phylogenetic studies on the family using DNA sequences have suffered from sparse taxon sampling and/or limited amounts of data. Here we combine all available previously published DNA sequence data and add to it new DNA sequences generated for this study. We analysed these data in a supermatrix using Bayesian and maximum likelihood methods and provide a phylogenetic hypothesis for the family that recovers strong support for the monophyly of all subfamilies and resolves relationships among a large proportion of chrysopine genera. Chrysopinae tribes Leucochrysini and Belonopterygini were recovered as monophyletic sister clades, while the species‐rich tribe Chrysopini was rendered paraphyletic by Ankylopterygini. Relationships among the subfamilies were resolved, although with relatively low statistical support, and the topology varied based on the method of analysis. Greatest support was found for Apochrysinae as sister to Nothochrysinae and Chrysopinae, which is in contrast to traditional concepts that place Nothochrysinae as sister to the rest of the family. Divergence estimates suggest that the stem groups to the various subfamilies diverged during the Triassic‐Jurassic, and that stem groups of the chrysopine tribes diverged during the Cretaceous.