Evidence against direct transfer of the adenine nucleotides by the heart mitochondrial creatine kinase-adenine nucleotide translocase complex

Adenine nucleotide translocase inhibitors, atractyloside and carboxyatractyloside, added to respiration inhibited rat heart mitochondria had no significant effects on the apparent Km's for ATP and ADP and Vmax's of the creatine kinase reaction. The results suggest that there is no sequential, mandatory, direct transfer of the nucleotides between the translocase and the creatine kinase active site. The reaction was run in both the forward and reverse directions in media containing either 0.25 M sucrose or 0.13 M KCl. The apparent Km's, however, were found to be 2–3 times higher in the KCl medium than in sucrose, suggesting the use of the more physiological medium for meaningful kinetic studies.     

Preparation of monoclonal antibodies against salivary gland immunogens of femaleRhipicephalus evertsi evertsi

The preparation of monoclonal antibodies raised against antigens of salivary gland extracts of femaleRhipicephalus evertsi evertsi is reported. Nine hybridoma cell lines were established of which eight secreted IgG3 and one IgG1. The immune response was biased towards immunogens unique to the prefed stage of the tick salivary glands by prior cyclophosphamide suppression of the immune response against unfed tick salivary glands. Immunoblots of salivary gland antigens, separated by SDS-PAGE showed reactivity with three of the monoclonal antibodies, all of which had identical specificity for antigens unique to prefed salivary extracts. Each of these monoclonal antibodies identified two prominent bands with relative molecular masses corresponding to 23 and 46 and a third minor band with molecular mass of 70 kDa.     

Comparison between total and ultrafiltrable serum zinc as test to diagnose zinc deficiency in infants and children

Total Serum Zinc (TSZn) and albumin were determined, and low molecular weight serum Zn measured by radiochemical Ultra-Filtration (UFSZn) in healthy Dutch infants and children, and in samples obtained from those with diseases that are expected to alter TSZn. Our control TSZn values, 10.2±3.5 μmol/L, were low compared to those reported in the literature. Variation in serum albumin could not explain this: No correlation of TSZn with serum albumin was found (p>0.5). Likely explanations are the nonfasting state and the stress owing to hospital surroundings at the time of sampling. A range of other influences not registered may be active and are discussed. No significant age-dependence was found (p<0.8). Boys over 9 yr of age showed higher TSZn compared with girls of the same age (p<0.08). In a separate experiment a 17% decrease in TSZn was demonstrated by food intake (eggs). These results support the opinion that TSZn is of little value to measure Zn status. There was no discrimination in TSZn between healthy subjects and patients. Our UFSZn values, 0.28±0.13 μmol/L in the controls as well as in the patients, were correlated with TSZn and therefore not a suitable alternative for the measurement of TSZn as parameter to determine the Zn status. The UFSZn was not correlated with serum albumin (p>0.7). UFSZn values were higher in infants (p<0.01), no sex dependence was found. We conclude that TSZn as well as UFSZn are of limited clinical relevance.     

Absence of acute doxorubicin-induced dysfunction of heart mitochondrial oxidative phosphorylation and creatine kinase activities

Since reductions in cardiac high-energy phosphate content and dysfunction of mitochondrial activities have been demonstrated after doxorubicin exposure, one mechanism of doxorubicin cardiotoxicity has been thought to be an interference with mitochondrial energy metabolism. To determine whether mitochondrial dysfunction is induced by acute drug exposure, isolated rat hearts were perfused with 10(-5) M doxorubicin for 70 min followed by mitochondrial isolation. Rates of electron transport, creatine kinase activity, acceptor control, respiratory control, and ADP/O ratios were assayed and correlated to doxorubicin-induced abnormalities in left ventricular function. At doses of doxorubicin sufficient to cause a marked deterioration of left ventricular systolic pressure and a rise in end-diastolic pressure, no decreases were noted in the measured mitochondrial parameters with either glutamate plus malate or succinate as respiratory substrates. In fact, in some cases the rates of electron transport were higher in mitochondria isolated from the treated hearts. In addition, isolated heart mitochondria were directly incubated in doxorubicin at doses as high as 10(-4) M for up to 70 min at 0 and 20 degrees C and 1.5 min at 37 degrees C. Under these conditions functional impairment of mitochondrial respiration was also not detected. Therefore, it appears that acute doxorubicin cardiotoxicity cannot be related to primary mitochondrial defects in high-energy phosphate metabolism. These data lend further support to the notion that doxorubicin cardiotoxicity may be fundamentally related to changes in coronary vascular resistance and resultant damage induced by hypoperfusion.     

In vitro exchangeable erythrocytic zinc

Exchangeable erythrocytic zinc is measured by 65Zn uptake in and release from erythrocytes under standardized and near, physiological conditions: 7.6 microM zinc and 580 microM albumin in the medium. The intracellular exchangeable erythrocytic zinc pool in healthy volunteers amounts to 5 mumol zinc/L packed cells. The half-time of the exchange is 7 h, its activation energy 84 kJ/mol. The effects of the variation in temperature and the concentrations of albumin, as well as the effects of some zinc carriers, cell transport inhibitors, and stress hormones on the 65Zn uptake are measured.     

Genetic population structure of the large blue butterfly Maculinea alcon in Denmark

We have investigated the genetic population structure within and the genetic differentiation between local populations of the large blue butterfly Maculinea alcon throughout the Jutland peninsula. Samples were collected as eggs on foodplants (Gentiana pneumonanthe), and reared to 4th instar caterpillars in the laboratory. A significant excess of homozygotes was found for all the investigated allozyme loci in most of the populations. A North-South cline was observed for the allele frequencies at some of the loci and for several linkage groups. Because some of the allele frequency clines were parallel to clines in adult morphological variation, we interpret our results as evidence for the co-existence of at least two gene pools within the Danish Maculinea alcon populations. Multilocus electrophoretic data revealed highly positive but variable FST values, which under this scenario would reflect varying frequencies of the Maculinea gene pools across the local populations. The significantly positive FIS values indicate that these gene pools are at least partly reproductively isolated (Wahlund effect). The co-occurrence of several Maculinea alcon gene pools on many local sites in Jutland is of great importance for conservation of the fragmented Maculinea populations. Our results show that there is probably more Maculinea biodiversity to conserve than was previously thought, and suggests that extant populations are more fragmented and vulnerable than counts of flying adults or eggs on foodplants indicate.     

A comparative study of proteins present in sweating-sickness-inducing and non-inducing strains ofHyalomma truncatum ticks

The identification of antigens unique to the positive strain of the tickHyalomma truncatum is discussed. Cattle were separately immunized against sweating-sickness-positive and sweating-sickness-negative ticks. A study was done to demonstrate the difference in protein patterns of crude salivary-gland extracts between positive and negative ticks. Immunological differences were identified using Western blots. A 32 kDa band was found to be unique to the positive strain.