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Biomedical subjects

K A Sukalski

Publications and source records attributed to K A Sukalski.

14 recordsLinked to original sources
PubMed2001

Endocrine physiology in a patient-centered learning curriculum.

The medical curriculum at the University of North Dakota School of Medicine and Health Sciences has recently been redesigned into a problem-based/traditional hybrid model that utilizes an integrated organ systems-based approach to teach basic and clinical sciences. The number of lecture hours in general has been greatly reduced, and, in particular, lecture hours in physiology have been reduced by 65%. Students learn basic science in small groups led by a faculty facilitator, and students are responsible for a great deal of their own teaching and learning. The curriculum is centered around patient cases and is called patient-centered learning (PCL). The curriculum includes traditional lectures and laboratories supporting faculty-generated learning objectives. Endocrine physiology is taught in year one, utilizing four weeks of patient cases that emphasize normal structure and function of endocrine systems. Endocrine physiology is revisited in year two, which is primarily focused on pathobiology. The PCL curriculum, with emphasis on the endocrine component, is described in detail along with key portions of an endocrine case.

Education, Medical↗
PubMed1999

Protein and lipid oxidation of banked human erythrocytes: role of glutathione.

In banked human erythrocytes (RBCs), biochemical and functional changes are accompanied with vesiculation and reduced in vivo survival. We hypothesized that some of these changes might have resulted from oxidative modification of membrane lipids, proteins, or both as a result of atrophy of the antioxidant defense system(s). In banked RBCs, we observed a time-dependent increase in protein clustering, especially band 3; carbonyl modification of band 4.1; and malondialdehyde, a lipid peroxidation product. Examination of the antioxidative defense system showed a time-dependent decline in glutathione (GSH) concentration and glutathione-peroxidase (GSH-PX) activity, with a concomitant increase in extracellular GSH, cysteine, and homocysteine, and unchanged catalase activity. When subjected to acute oxidant stress by exposure to ferric/ascorbic acid or tert-butylhydroperoxide (tert-BHT), catalase activity showed a steeper decline compared with GSH-PX. The results demonstrate that GSH and GSH-PX appear to provide the primary antioxidant defense in stored RBCs, and their decline, concurrent with an increase in oxidative modifications of membrane lipids and proteins, may destabilize the membrane skeleton, thereby compromising RBC survival.

Antioxidants↗
PubMed1997

In vivo oxidative modification of erythrocyte membrane proteins in copper deficiency.

Oxidative stress has been postulated to contribute to the pathology associated with dietary copper deficiency. In vivo, erythrocytes are probable targets of oxidative damage because they are exposed to high concentrations of oxygen and contain heme iron that can autoxidize, which results in the formation of superoxide anions. Activity of the important antioxidant enzyme, copper, zinc superoxide dismutase, decreases markedly in erythrocytes during copper deficiency. The effect of dietary copper deficiency on indicators of oxidative stress was examined in erythrocyte membranes of rats maintained on a purified copper-deficient diet for 35 days after weaning. Erythrocytes were separated into young and old populations on a Percoll gradient prior to membrane isolation and quantification of lipid peroxides and protein carbonyls. Protein carbonyls, determined by Western blot immunoassay, were detected predominantly in both the alpha and beta chains of spectrin. Alpha and beta subunits of spectrin in erythrocyte membranes from copper-deficient rats contained higher amounts of carbonyls than controls, regardless of the population of erythrocytes studied. This study suggests that spectrin may be a specific target for oxidative damage when erythrocyte copper, zinc superoxide dismutase activity is reduced by copper deficiency.

Animals↗
PubMed1993

Hysteresis at near-physiologic substrate concentrations underlies apparent sigmoid kinetics of the glucose-6-phosphatase system.

Although Canfield and Arion (J. Biol. Chem. 263, 7458-7460 (1990)) have described the kinetics as hyperbolic, Traxinger and Nordlie (J. Biol. Chem. 262, 10015-10019 (1987)) reported sigmoid kinetics in the glucose-6-phosphatase system of intact microsomes at near-physiologic glucose-6-P concentrations. We show here that apparent sigmoidal kinetics, most clearly seen as sharp upward inflections in Hanes plots as substrate concentration approaches zero, are a consequence of the hysteretic lag in product formation during the first minutes of incubation of the enzyme with low concentrations of substrate. The appearance of sigmoidicity, observed when reaction velocities are calculated from changes in Pi concentration between 0 and 6 min of incubation, is not present when velocity is determined from slopes of [product]-time plots after linearity is achieved. The Km,glucose-6-P value, 0.86 mM, based on these hysteresis-corrected velocity values determined with intact microsomes from normal, control rats at low substrate concentrations, approached the upper limit of physiologic hepatic glucose-6-P concentrations. This suggests that glucose-6-phosphatase activity may be regulated by factors other than substrate concentrations alone. We propose that the hysteretic behavior, not sigmoid kinetics of the glucose-6-phosphatase enzyme system, may be a prime regulatory feature.

Animals↗
PubMed1993

Human microsomal glucose-6-phosphatase system.

The discovery of glucose-6-phosphatase (EC 3.1.3.9) and of its physiological function in releasing glucose from the liver are discussed briefly. The identification by the Coris of glucose-6-phosphatase deficiency as the underlying defect in certain cases of glycogenosis (type I glycogenosis; von Gierke disease) is described. Characteristics of the catalyst, with a focus on its multiplicity of functions and multicomponent character, are considered with an emphasis on the human liver enzyme. Pioneering studies from the author's laboratory leading to the characterization of two variants of type I glycogenosis, types Ib and Ic, are described.

Glucose-6-Phosphatase↗
PubMed1993

Decreased susceptibility of liver mitochondria from diabetic rats to oxidative damage and associated increase in alpha-tocopherol.

The susceptibility of mitochondria from liver and kidney of diabetic and normal rats to in vitro oxidative damage was assessed. Mitochondria were isolated from diabetic rats 4 weeks after streptozotocin injection and from age-matched, normal rats. Liver mitochondria from diabetic rats were less susceptible to oxidative damage (induced by Fe3+/adenosine 5'-diphosphate (ADP) xanthine/xanthine oxidase), as assessed by the formation of thiobarbituric acid reacting substances (TBARS) and sulfhydryl loss, than were mitochondria from normal rats. The decreased susceptibility of liver mitochondria from diabetic rats to oxidative damage correlated with a sevenfold increase in mitochondrial alpha-tocopherol levels. Activities of the antioxidant enzymes, glutathione reductase, glutathione peroxidase, and superoxide dismutase, were lower in liver mitochondria from diabetic compared to normal rats. Manipulation of dietary alpha-tocopherol, to counteract the increased intake of alpha-tocopherol due to diabetes-associated polyphagia, failed to lower liver mitochondrial alpha-tocopherol to the levels found in normal rats. Mitochondria from kidney of diabetic rats were equally as susceptible to in vitro oxidative damage as kidney mitochondria from normal rats. They had increased levels of superoxide dismutase and glutathione peroxidase but identical levels of alpha-tocopherol compared to mitochondria from normal rats. Dietary manipulation of alpha-tocopherol had no effect on kidney mitochondrial levels of the nutrient.

Animals↗
PubMed1991

Hysteretic behavior of the hepatic microsomal glucose-6-phosphatase system.

Carbamyl-P:glucose and PPi:glucose phosphotransferase, but not inorganic pyrophosphatase, activities of the hepatic microsomal glucose-6-phosphatase system demonstrate a time-dependent lag in product production with 1 mM phosphate substrate. Glucose-6-P phosphohydrolase shows a similar behavior with [glucose-6-P] less than or equal to 0.10 mM, but inorganic pyrophosphatase activity does not even at the 0.05 or 0.02 mM level. The hysteretic behavior is abolished when the structural integrity of the microsomes is destroyed by detergent treatment. Calculations indicate that an intramicrosomal glucose-6-P concentration of between 20 and 40 microM must be achieved, whether in response to exogenously added glucose-6-P or via intramicrosomal synthesis by carbamyl-P:glucose or PPi:glucose phosphotransferase activity, before the maximally active form of the enzyme system is achieved. It is suggested that translocase T1, the transport component of the glucose-6-phosphatase system specific for glucose-6-P, is the target for activation by these critical intramicrosomal concentrations of glucose-6-P.

Animals↗
PubMed1989

Radiation inactivation analysis of rat liver microsomal glucose-6-phosphatase.

Radiation inactivation analysis was utilized to estimate the sizes of the units catalyzing the various activities of hepatic microsomal glucose-6-phosphatase. This technique revealed that the target molecular weights for mannose-6-P phosphohydrolase, glucose-6-P phosphohydrolase, and carbamyl-P:glucose phosphotransferase activities were all about Mr 75,000. These results are consistent with the widely held view that all of these activities are catalyzed by the same protein or proteins. Certain observations indicate that the molecular organization of microsomal glucose-6-phosphatase is better described by the conformational hypothesis which envisions the enzyme as a single covalent structure rather than by the substrate transport model which requires the participation of several physically separate polypeptides. These include the findings: 1) that the target sizes for glucose-6-P phosphohydrolase and carbamyl-P:glucose phosphotransferase activities were not larger than that for mannose-6-P phosphohydrolase in intact microsomes and 2) that the target size for glucose-6-P phosphohydrolase in disrupted microsomes was not less than that observed in intact microsomes. These findings are most consistent with a model for glucose-6-phosphatase of a single polypeptide or a disulfide-linked dimer which spans the endoplasmic reticulum with the various activities of this multifunctional enzyme residing in distinct protein domains.

Animals↗
PubMed1986

N-acetylglucosamine-insensitive glucose phosphorylation in isolated hepatocytes from rats fasted for 48 h.

Glucose phosphorylation in isolated hepatocytes was studied by the release of 3H from D-[2-3H]glucose. Glucokinase activity was decreased by fasting rats for 48 h and was further reduced in cells by adding 30 mM GlcNAc, a potent competitive inhibitor. Although this treatment resulted in the loss of more than 97% of glucokinase activity in hepatocytes, glucose phosphorylation proceeded at an appreciable rate. These observations demonstrate the involvement of a high -K0.5 enzyme system in addition to glucokinase in hepatocyte glucose phosphorylation.

Acetylglucosamine↗
PubMed1986

Implications of distinct inhibitory effects of N-acetylglucosamine on glucose uptake by an isolated perfusion system incorporating erythrocytes with livers from fed and 48-hour fasted rats.

Net glucose uptake in a perfusion system including erythrocytes and isolated livers from fed rats was inhibited by N-acetylglucosamine (GlcNAc), a competitive inhibitor of glucokinase. Net glucose uptake also occurred in the system incorporating livers from 48-h fasted rats, but its inhibition by GlcNAc did not. This distinction could not be explained on the basis of a different sensitivity of glucokinase from fasted compared with fed rats to inhibition by GlcNAc. Nor could it be rationalized based on several other hepatic enzymes possibly involved in glucose utilization or production. Because erythrocytes were included in our system, other explanations were sought related to the total enzymic environment. The involvement of an indirect pathway including glycolysis of glucose to lactate in erythrocytes followed by conversion of this lactate to glucose-6-P and then glycogen in liver was considered. This pathway contributed no more than 17% to total net glucose uptake in the system incorporating livers from fed rats. This per cent contribution increased when hepatic glucokinase was reduced by fasting or through inhibition by GlcNAc. However, it was too small to explain observed overall rates of net glucose uptake. We propose that the presence of erythrocytes may also promote a greater net glucose uptake by the direct hepatic pathway. An enhanced inhibition of hepatic glucose-6-P hydrolysis by some intermediate metabolite generated in the presence of lactate infusion from erythrocytes may promote net glucose uptake independently of the mechanism of residual hepatic glucose phosphorylation. This may explain why we and others who have employed liver perfusion systems including erythrocytes have seen greater net glucose uptake than have workers using systems devoid of erythrocytes.

Acetylglucosamine↗
PubMed1983

Type Ic, a novel glycogenosis. Underlying mechanism.

The availability of a fresh, unfrozen liver biopsy specimen permitted the characterization of a unique type of glycogen storage disease. The subject, an 11-year-old female, showed the classic clinical symptoms of type I glycogenosis. However, her hepatic D-glucose-6-phosphate phosphohydrolase (EC 3.1.3.9) level as determined with detergent-activated homogenate was normal. The underlying mechanism was studied with intact microsomes from this fresh liver homogenate. Glucose-6-P phosphohydrolase was 75% latent, compared with 25% in normal controls matched for age and sex. Inorganic pyrophosphatase, PPi:glucose phosphotransferase, and carbamyl-P:glucose phosphotransferase activities of glucose 6-phosphatase were totally latent. While not observed with intact microsomes, these activities were fully manifested with detergent-disrupted microsomes. D-Glucose inhibited glucose-6-P phosphohydrolase activity of both intact and disrupted microsomes, but exogenous Pi inhibited only with the detergent-disrupted preparation. These observations are interpreted on the basis of the multicomponent glucose 6-phosphatase system of Arion et al. (Arion, W. J., Lange, A. J., Walls, H. E., and Ballas, L. M. (1980) J. Biol. Chem. 255, 10396-10406). All are consistent with a defect in T2, the putative translocase specific for Pi, PPi, and carbamyl-P. However, Pi produced endogenously from glucose-6-P hydrolysis within the microsomal lumen did not inhibit. This suggests that (i) a pathway for egress of Pi from the microsomal lumen exists independently of T2, (ii) T2 in this case works only unidirectionally, or (iii) the catalytic unit of glucose 6-phosphatase in situ has become desensitized to interactions with Pi, PPi, and carbamyl-P in this mutant model. Defects in both T1, the translocase specific for glucose-6-P, and T2 thus appear involved in this unique glycogenosis.

Child↗
PubMed1982

Stimulation by 3-mercaptopicolinate of net glucose uptake by perfused livers from diabetic rats.

Glucose uptake/production was studied as a function of varied glucose loads in isolated perfused livers from glucagon-treated alloxan-diabetic rats. Uptake of D-[U-14C]glucose was seen at all levels studied - 9.5-71 mM. In studies with unlabelled D-glucose carried out in the absence of 3-mercaptopicolinate, livers of diabetic rats showed a net production of glucose with perfusate glucose levels less than 22 mM. Above this level, these livers exhibited a time- and concentration-dependent net uptake of glucose for a period of 20-30 min. When 4 mM 3-mercaptopicolinate, which inhibited gluconeogenesis from endogenous substrates, was included in perfusates, a continuous net uptake of unlabelled glucose was observed at all levels above 4 mM. This lowering of the null-point, cross-over glucose concentration was shown to relate mechanistically to the observed reduction in steady-state hepatic glucose 6-phosphate level produced by mercaptopicolinate. The need for supplemental mechanisms of glucose utilization by high Km hepatic enzyme(s) operative in the virtual absence of insulin-dependent glucokinase also is indicated by these observations and by kinetic analysis.

Animals↗