PubMed Health⌕ Search

Biomedical subjects

G Jackowski

Publications and source records attributed to G Jackowski.

At least 37 records · Page 2Linked to original sources
PubMed1989

Myocardial salvage with trolox and ascorbic acid for an acute evolving infarction.

Both Trolox (a water-soluble analogue of alpha-tocopherol) and ascorbic acid were more effective than superoxide dismutase or catalase in protecting myocyte cell cultures from free radical attack (induced by hypoxanthine and xanthine oxidase). In a canine model of two hours of left anterior descending coronary artery occlusion followed by four hours of reperfusion, Trolox and ascorbic acid reduced the area of infarction within the area at risk. The Trolox group received 500 mL of deoxygenated saline solution containing 2.0 g of Trolox, 3.0 g of ascorbic acid, and 18 mg of EDTA (ethylenediaminetetraacetic acid) infused into the ascending aorta 30 seconds before and four minutes after reperfusion. Saline controls received 500 mL of deoxygenated saline solution containing 18 mg of EDTA. The angioplasty group had unmodified reperfusion by simple release of the occlusion. The area at risk and the area infarcted were estimated with Evans blue and triphenyl tetrazolium hydrochloride stains, respectively. The ratio of the area infarcted to the area at risk was significantly lower with Trolox (angioplasty, 30.4% +/- 5.1%; saline, 20.8% +/- 2.9%; and Trolox, 8.7% +/- 4.0%; p less than 0.01). In summary, the antioxidants Trolox and ascorbic acid effectively reduced myocardial necrosis after ischemia.

Animals↗
PubMed1989

Effect of oxygen tension on the anti-oxidant enzyme activities of tetralogy of Fallot ventricular myocytes.

Since the chronically cyanotic tetralogy of Fallot (TOF) myocardium is more sensitive to reperfusion injury after cardiac surgery than the adult myocardium, we decided to study the regulation of myocardial superoxide dismutase (SOD), catalase and glutathione peroxidase by oxygen tension. TOF myocytes were cultured at a Po2 of 150 mmHg for 30 days to establish the culture. The cells were then cultured at Po2 of 150 and 40 mmHg and the myocyte antioxidant enzymes measured at days 3, 7, 14 and 21. On day 21 the myocytes cultured at Po2 of 40 mmHg were then cultured at 150 mmHg and SOD and catalase activities measured on days 28 and 35. Although there were no differences in the rates of incorporation of 35S-methionine into the myocytes at either Po2 on these days, the myocytes scavenger enzyme levels were significantly higher by day 14 when cultured at a Po2 of 150 mmHg than at a Po2 of 40 mmHg. With the increase in oxygen tension from 40 to 150 mmHg, SOD and catalase activities increased significantly by day 35. The myocytes cultured at Po2 40 mmHg were more sensitive by day 7 to an hypoxanthine-xanthine oxidase generated free radical injury than the Po2 150 mmHg cultured cells. The regulation of these enzyme activities by oxygen tension and the increased sensitivity to free radical injury of the myocytes cultured at an oxygen tension of 40 mmHg provide putative evidence that the chronically cyanotic myocardium may be less well protected than the normally perfused myocardium against oxygen-mediated free radical injury and be at higher risk for cardiovascular surgery.

Catalase↗
PubMed1988

Qualitative and quantitative differences in protein synthesis comparing fetal lamb ductus arteriosus endothelium and smooth muscle with cells from adjacent vascular sites.

During late gestation, intimal cushions form in the ductus arteriosus (DA) and these cause the vessel to close when it constricts in the postnatal period. The formation of intimal cushions suggests highly specialized functions of DA endothelial and smooth muscle cells. To investigate these properties, we established, from fetal lambs on Day 138 of a 148-day term gestation, primary cell cultures of DA endothelium and smooth muscle and compared them to cells derived from the adjacent pulmonary artery and aorta. Purity of the endothelial cell cultures from each vascular site was assessed by the contact inhibited "cobblestone" monolayer phenotype, by positive immunofluorescence for factor VIII and by angiotensin converting enzyme activity. Purity of smooth muscle cell cultures at each vascular site was assessed by the "hills and valleys" phenotype and by positive immunofluorescence with a smooth muscle actin specific monoclonal antibody. Endothelial and smooth muscle cells had different growth curves, ultrastructural features, and protein profiles on single and two-dimensional SDS-polyacrylamide gel electrophoresis (PAGE), but vascular sites were similar. To further determine whether differences related to DA origin were indeed present, endothelial and smooth muscle cells from all three vascular sites were incubated with the radiolabeled amino acids [14C]leucine, [14C]proline, and [14C]valine and the proteins in both the cells and the conditioned medium were analyzed by autoradiography after SDS-PAGE. A dense band corresponding to a 42-kDa protein was observed in valine-labeled DA endothelial cells and conditioned medium and a 52-kDa protein was observed in the conditioned medium of leucine-labeled DA smooth muscle cells only. Further isolation and characterization of these endothelial and smooth muscle proteins will be necessary to determine whether they are related to the mechanism of intimal cushion formation in the late gestation DA or are present abnormally in association with the intimal proliferation observed in pulmonary and systemic vascular disease.

Actins↗
PubMed1985

Differences in cardiac myosin light chain LC1 among human, monkey and sheep.

The atrial and ventricular myosin light chains of human, monkey and sheep hearts were compared by dodecylsulfate polyacrylamide gel electrophoresis. The atrial light chain 2 and ventricular light chain 2 are similar among these mammals. However, the atrial light chain 1 of monkey has different electrophoretic mobility from those of human and sheep. The monkey ventricular light chain 1 has same mobility as that of sheep but different from that of human.

Adult↗
PubMed1985

Cellular regulation of poly(ADP) ribosylation of proteins. I. Comparison of hepatocytes, cultured cells and liver nuclei and the influence of varying concentrations of NAD.

The in vitro rates (vinit) of poly(ADP-ribose) polymerase of permeabilized rat hepatocytes and of nuclei, isolated from hepatocytes, did not differ significantly. Incubation beyond 3 min resulted in diminished poly(ADP) ribosylation in hepatocytes compared with nuclei, coinciding with high rates of plasma membrane-associated NAD-glycohydrolase. Cultured cells (Drosophila Kc cells, gliosarcoma 9L, human fibroblasts and mouse spleen lymphocytes) exhibit variations of NAD-glycohydrolase and poly(ADP-ribose) polymerase activities and the assessment of poly(ADP-ribose) polymerase activity in permeabilized cells requires simultaneous assay of NAD-glycohydrolase. In rat liver nuclei during 10 min incubation with 500 microM NAD, 40% of NAD is consumed, 10% ADP-ribose is bound to proteins, and 20% ADP-ribose, 5% AMP and 2.7% adenosine are liberated. As determined by solvent partitioning (Jackowski, G & Kun, E, J biol chem 258 (1983) 12587) [1], the phenol-soluble protein-ADP-ribose fraction represents largely mono(ADP)-ribose protein adducts, whereas the H2O-soluble phase contains poly(ADP)-ribosylated proteins. The quantity of ADP-ribose protein adducts, the chain length of oligomers and the nature of apparent acceptor proteins in liver nuclei vary significantly with the concentration of NAD as substrate. At 500 microM NAD concentration the quantity of ADP-ribose containing adducts was in the nmol per mg DNA range, the polymers are long chains and the acceptor proteins predominantly non-histone proteins. At 0.1 microM NAD as substrate pmol quantities of monomeric ADP-ribose adducts per mg DNA were formed and the main acceptors were sharply discernable on the basis of molecular mass as histones, high mobility non-histone proteins, two protein groups of a mass of 66 and 44 kD respectively, and the poly(ADP-ribose) polymerase enzyme protein of 119 kD mass. Whereas products in the presence of 0.1 microM NAD may indicate acceptors of highest reactivity, protein adducts formed in the presence of 500 microM NAD resemble a pattern found in vivo.

ADP Ribose Transferases↗
PubMed1984

Evidence for the macromolecular basis of regulation of heart hypertrophy.

In vivo treatment of rats with triiodothyronine (0.3 micrograms per g body wt for four consecutive days) increases both poly (ADP-ribose) polymerase activity and DNA synthesis in myocardial nuclei obtained from 18-21-days-old rats. The same T3-treatment in 30-33-days-old rats inhibits poly(ADP-ribose) polymerase activity and simultaneously increases RNA syntheses in myocardial nuclei. A correlation was observed between the degree of inhibition of poly(ADP-ribose) polymerase and ventricular enlargement in triiodothyronine treated animals. RNA synthesis in isolated myocardial nuclei was inhibited by in vitro polyADP-ribosylation only when myocardial nuclei were obtained from triiodothyronine treated animals. In vitro poly ADP-ribosylated proteins were isolated from myocardial nuclei by the SDS-phenol extraction. More than 90% of the protein-polyADP-ribose adducts partitioned into the aqueous phase behaving as if they were nucleic acids. Treatment with triiodothyronine significantly, diminished polyADP-ribosylation of three specific groups of polyADP-ribosylated non-histone chromatin proteins corresponding to 130 kDa, 90-80 kDa and 80-65 kDa.

Animals↗
PubMed1983

The effect of in vivo treatment with triiodothyronine on the in vitro synthesis of protein-poly(ADP)-ribose adducts by isolated cardiocyte nuclei and the separation of poly(ADP)-ribosylated proteins by phenol extraction and electrophoresis.

In vivo treatment of rats with triiodothyronine (30 micrograms/100 g of body weight for 4 consecutive days) inhibited poly(ADP)-ribose polymerase activity of cardiocyte nuclei, but low enzymatic activity of nuclei of noncardiocyte origin remained unaffected. RNA synthesis in cardiocyte nuclei isolated from triiodothyronine-treated rats was augmented. A positive correlation was observed between the degree of inhibition of poly(ADP)-ribose polymerase and cardiac ventricular enlargement in triiodothyronine-treated animals. RNA synthesis in isolated cardiocyte nuclei was inhibited by in vitro poly(ADP)-ribosylation only when cardiocyte nuclei were obtained from triiodothyronine-treated animals. In vitro poly(ADP)-ribosylated proteins were isolated from cardiocyte nuclei by solvent partitioning between phenol and aqueous phases. About 90% of the protein-poly(ADP)-ribose adducts partitioned into the aqueous fraction, and the chain length of polymers in this phase was between medium (n = 4-9) and long (n greater than 32), whereas the phenol phase contained protein-oligomer and monomer adducts. Not only the chain length of oligomers but the nature of modified proteins appeared to participate in determining the partitioning of polymer-protein adducts, and different proteins were separated from the two phases by gel electrophoresis. More than 90% of protein-polymer adducts formed by cardiocyte nuclei were not extracted by 0.25 N HCl, indicating prevalence of nonhistone proteins as polymer acceptors. Gel electrophoresis and near quantitative recovery of adducts in a gel system that protected from degradation of adducts to free polymers confirmed the predominance of nonhistone proteins as main acceptors and demonstrated an artifact of autoradiography that seemed to indicate histone H1 as a significant acceptor. Treatment with triiodothyronine diminished poly(ADP)-ribosylation of certain groups of proteins more than others, implying some degree of selectivity of action of the hormone. Catabolism of the polymer in vitro was not affected by triiodothyronine treatment.

Animals↗
PubMed1983

Changes in non-histone nuclear proteins during postnatal myocardial development.

Myocardial-cell nuclei isolated from ventricles of rats between 4 days and 15 months of age showed a progressive loss of DNA-synthetic activity in vitro. Correlated with this loss is the differential appearance of three major groups of non-histone nuclear proteins. Group a, with pI5.7-6.5 and Mr 67000, appeared in detectable amounts in rats more than 10 days of age, whereas group b, consisting of proteins with pI6.8-7.2 and Mr 45000, appeared in rats older than 33 days. The third group (group c), with pI6.5-8.5 and Mr 32000-42000, was present throughout postnatal development, but the amount of these proteins appeared to be greater in the 10-day-old rat than in the 15-month-old rat. All groups of these specific proteins were localized within the nucleus and did not reflect cytoplasmic constituents. Furthermore, the amino acid compositions of these polypeptides were also analysed and compared with those of actin and tubulin.

Aging↗
PubMed1983

Nonenzymatic separation of myocardial cell nuclei from whole heart tissue.

A simple and rapid nonenzymatic method has been developed to isolate myocardial cell nuclei from whole heart tissue. This method consists of a controlled disruption of cells followed by isopycnic gradient centrifugation. We have reviewed and compared our method to others more lengthy and laborious. By using a number of criteria, such as morphometric measurements, chemical composition, functional studies, specific nuclear protein markers, and mathematical analysis, we show that the nonenzymatic digestion method provides a most useful technique for the study of the biochemistry of the myocardial cell nucleus.

Aging↗
PubMed1981

Quantitative isolation of oligo- and polyadenosine-diphosphoribosylated proteins by affinity chromatography from livers of normal and dimethylnitrosamine-treated Syrian hamsters. In vivo and in vitro metabolism of the homopolymer.

Polyadenosine- and adenosine-diphosphoribosylated proteins of hamster liver were quantitatively isolated with the aid of m-aminophenyl boronic acid glutaryl hydrazide polyacrylamide affinity resin by selective adsorption at pH 8.2 and elution at pH 4.0. Polymer-free proteins, DNA, and RNA are readily separated from adenosine-diphosphoribosylated proteins. The total quantity of proteins that is covalently modified by the homopolymer is 14.3 micrograms/mg of DNA or 37.4 micrograms/g of liver in controls and 38.7 micrograms/mg of DNA or 116 micrograms/g of liver in dimethylnitrosamine-treated hamsters. Polymer content increases from 9 to 15 nmol/mg of DNA to 42 to 118 nmol/mg of DNA following treatment with dimethylnitrosamine. Pulse labeling with [14C]ribose results in a parallel doubling in dimethylnitrosamine-treated animals of the specific activities of adenosine- diphosphoribose and NAD+ and of the [14C]ribose content of polyadenosine-diphosphoribose of chain length between 20 and 40, indicating chain elongation of pre-existing larger polymers. Two groups of proteins that are isolated as polyadenosine-diphosphoribose adducts are increased significantly after treatment with dimethylnitrosamine, one minor component of a mass between 100-112 X 10(3) daltons, and a major group exhibiting a mass of 158-162 X 10(3) daltons. Polyadenosine-diphosphoribose synthetase activity of isolated hepatic nuclei is increased by 32-37% after dimethylnitrosamine treatment, and since the change in glycohydrolase activity is negligible relative to the increase in synthetase, the augmentation of polyadenosine-diphosphoribosylated proteins can be explained by the increased synthetase of nuclei. The molecular size distribution of DNA in liver nuclei of control and dimethylnitrosamine-treated hamsters is indistinguishable.

Adenosine Diphosphate Ribose↗
PubMed1981

Age-dependent variation of rates of polyadenosine-diphosphoribose synthesis by cardiocyte nuclei and the lack of correlation of enzymatic activity with macromolecular size distribution of DNA.

Cardiocyte nuclei from neonate (5-day-old) and from adult (90-day-old) male Wistar rats were selectively isolated by a technique that eliminates interference by noncardiocyte nuclei (Jackowski, G., and Liew, C. C. (1980) Biochem. J. 188, 363-373). Polyadenosine-diphosphoribose synthetase activity of cardiocyte nuclei of neonates was ten times greater than in nuclei of adult rats, as calculated from initial velocity linear rate measurements. The molecular size of DNA extracted from cardiocyte nuclei of neonates was significantly larger (peak at 54 S) than DNA of cardiocyte nuclei of adults (peak at 33 S) as determined by alkaline sucrose density gradient ultracentrifugation (Knopf, K. W., and Weissbach, A. (1977) Biochemistry 16, 3190-3194). Comparison of the molecular sizes of DNA extracted from whole cardiac tissue with DNA isolated from cardiocyte nuclei shows that no DNA fragmentation takes place during the process of isolation of nuclei. Polyadenosine-diphosphoribose glycohydrolase activity was about 30% higher in cardiocyte nuclei of neonates than in adults, but the activity represents only 10% of the rate of the synthetase. Results demonstrate that polyadenosine-diphosphoribose metabolism in differentiated tissues is not directly related to the molecular size of DNA.

Aging↗
PubMed1980

Fractionation of rat ventricular nuclei.

Myocardial cells were isolated after treatment with collagenase (0.05%) and hyaluronidase (0.1%) by discontinuous-gradient centrifugation on 3% Ficoll. Nuclei derived from these myocardial cells were then fractionated on a discontinuous sucrose density gradient with the following steps: (I) 2.0M/2.3M, (II) 2.3M/2.4M, (III) 2.4M/2.5M, (IV) 2.5M/2.6M, and (V) 2.6M/2.85M. The myocardial nuclei were sedimented in the interfaces of gradient fractions (II) and (III). Nuclei from whole ventricles that had been treated with the enzymes before isolation sedimented into five major subsets of nuclei. These findings suggest that nuclei sedimented in the isopycnic gradient at fractions (II) and (III) are most probably derived from myocardial cells. However, this procedure is laborious and lengthy, and the recovery of myocardial-cell nuclei is low. An alternative method was developed to isolate an enriched fraction of myocardial-cell nuclei from whole ventricular tissue without exposing the tissues to enzyme digestion. These ventricular nuclei could be fractionated into five nuclear subsets by using the same discontinuous sucrose density gradient as that described above. The content of DNA, RNA and protein per nucleus for each band was determined. Although the DNA content per nucleus was constant (10pg), that of RNA varied from 1.5 to 4.5pg and that of protein from 16 to 24pg. Nuclei from each band were examined by light-microscopy: large nuclei occurred in the ligher regions whereas smaller nuclei were found in the denser regions of the gradient. From the size distribution pattern of myocardial-cell nuclei compared with that of total ventricular nuclei, it was found that nuclear subsets (II), (III), and (IV) were similar to myocardial nuclei. Electrophoretic analyses of the proteins solubilized in sodium dodecyl sulphate/phenol or Tris/EDTA/2-mercaptoethanol/phenol obtained from each nuclear subset indicate that these fractions are similar, with limited qualitative differences. These findings indicate that isolation of an enriched fraction of myocardial-cell nuclei could be achieved by discontinuous-sucrose-density-gradient centrifugation.

Animals↗