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Biomedical subjects

E D Robin

Publications and source records attributed to E D Robin.

At least 91 records · Page 5Linked to original sources
PubMed1981

Differences in oxygen-dependent regulation of enzymes between tumor and normal cell systems in culture.

Metabolic studies in tumor cells have indicated that bioenergetic regulatory mechanisms geared to acute changes in oxygen availability are abnormal. In the present studies we have examined bioenergetic adaptations to chronic oxygen depletion in culture maintained tumor cells in comparison to normal cell lines. Activities of two key glycolytic enzymes (pyruvate kinase (PyKI) and phosphofructokinase (PFK)) were measured in two tumor cell lines (fibrosarcoma (FS) and Hela) and two normal cell lines (rat lung fibroblasts (RLF) and WI-38) maintained in culture for up to 96 hours under aerobic (PO2 approximately 140) and hypoxic PO2 approximately 15) conditions. Exposure to low O2 tensions for 96 hours resulted in significant increases in PyKi and PFK in both RLF and WI-38, ut did not alter activities of these enzymes in either FS or HeLa cell systems. Activities of two enzymes involved in O2 metabolism (cytochrome oxidase (CyOx) and superoxide dismutase (SOD) were also measured in the two tumor cell lines and in RLF. chronic hypoxia significantly decreased the activities of CyOx and SOD in RLF cell systems but did not alter the activities of these enzymes in the tumor cells. In these studies, the tumor-derived cell lines do not demonstrate specific enzymatic responses to sustained oxygen depletion in vitro noted in normal cell systems, suggesting significant abnormalities in regulatory mechanisms geared to chronic changes in molecular O2.

Cell Line↗
PubMed1981

Glucoregulation and simulated diving in the harbor seal Phoca vitulina.

Plasma glucose, glucagon, and insulin concentrations were measured in the harbor seal, Phoca vitulina, during a 6-min dive and a 30-min recovery period. Studies were performed in the fasting state and following intravenous glucose. During diving in the fasting state, values of plasma glucose, glucagon, and insulin are not significantly different from prediving values, presumably because of the loss of perfusion to the pancreas. In the postdiving period, plasma glucagon increases significantly. The increased glucagon levels reach a peak at 6 min into the postdiving period. By 4 min postdiving, plasma glucose values increase. The hyperglycemia appears to be at least partially related to hyperglucagonemia. Despite hyperglycemia, insulin levels do not change significantly. The net effect is to increase glucose availability, particularly for non-insulin-requiring tissues (brain). Preadministration of glucose eliminates the postdiving increase in glucagon. Plasma glucose and insulin concentrations also show no significant changes during the diving and postdiving periods. These results suggest that hormonally mediated glucose conservation serves to maintain brain glucose supplies and to restore peripheral carbohydrate stores during the postdiving period.

Animals↗
PubMed1981

Regulation of intracytoplasmic pH and "apparent" intracellular pH in alveolar macrophages.

An approach for determining the permeability of cell and subcell membranes to H+/HCO3- has been applied to alveolar macrophages. The approach uses analysis of substrate ratios which are compartment specific and involved in pH-dependent reactions. By determining the changes in substrate pair ratios produced by perturbations of CO2, the effects of intracellular acidosis and alkalosis can be determined. By measuring substrate pair ratios at a constant PCO2 and variable H+/HCO3- it is possible to determine the permeability of the macrophage plasma membrane to these ions qualitatively. Measurements of intracellular lactate/pyruvate ratios showed that the alveolar macrophage plasma membrane is permeable to H+/HCO3-. Previous studies have shown that alveolar macrophages are freely permeable to Cl-. pHi was determined from measurements of pHe and Cli-/Cle- following changes of external PCO2 at a constant HCO3- concentration and following changes in pHe produced by changes of external HCO3- concentration at a constant PCO2. The data show that the relationship between pHi and Phe is: pHi = 0.973 pHe - 0.027 (r = 0.99, P less than 0.001). Thus at pHe = 7.40, macrophage intracellular pHi = 7.17. Alveolar macrophages appear to be permeable to H+/HCO3- and thus resemble erythrocytes, platelets, and lymphocytes and not brain and muscle cells.

Acid-Base Equilibrium↗
PubMed1981

Bioenergetic alterations in cultivated pulmonary artery and aortic endothelial cells exposed to normoxia and hypoxia.

Endothelial cells function under conditions of different oxygen availability under physiologic conditions and a variety of pathologic states. We determined the effect of normal and low O2 tensions on three key bioenergetic enzymes [pyruvate kinase (ATP:pyruvate phosphotransferase, EC 2.7.1.40), phosphofructokinase (ATP:D-fructose-6-phosphate 1-phosphotransferase, EC 27.1.11) and cytochrome aa3] in culture-maintained endothelial cells derived from calf pulmonary artery and aorta. Endothelial cells derived from pulmonary artery and aorta demonstrate similar bioenergetic enzyme activities when exposed to the same PO2 in vitro. Endothelial cells exposed to hypoxia in vitro for 48-96 hr show significantly increase activities of 2 key glycolytic enzymes: pyruvate kinase, and phosphofructokinase. Freshly isolated intimal strips from calf pulmonary artery (normal PO2 = 40 torr) show significantly greater activities of PyKi than aortic intimal strips (normal PO2 = 90 torr), suggesting that a similar pattern occurs in vivo. The data suggest that both cell types have a common bioenergetic pattern which is genetically determined and that this pattern is modified by regulatory mechanisms geared to ambient O2 tension. As endothelial cells of both types are not uncommonly exposed to hypoxic conditions, these regulatory mechanisms may play an important role in maintaining vascular integrity.

Animals↗
PubMed1980

Regulation of glycolytic enzyme activity during chronic hypoxia by changes in rate-limiting enzyme content. Use of monoclonal antibodies to quantitate changes in pyruvate kinase content.

Monoclonal antibodies were prepared against pyruvate kinase (PyKi; ATP: pyruvate phosphotransferase, EC 2.7.1.40) and used to quantitate PyKi content in L2 lung cells and WI-38 fibroblasts cultivated under hypoxic and normoxic conditions. After 96 h of hypoxic cultivation, PyKi activity was significantly increased in both cell types (L2: normoxia [Po2 = 142 torr], 0.11 +/- 0.01 [SD]; hypoxia [Po2 = 14 torr], 0.25 +/- 0.04 U/microgram DNA, P < 0.01). PyKi content increased proportionately in both cell lines (L2: normoxia, 0.44 +/- 0.13; hypoxia, 0.94 +/- 0.13 microgram enzyme protein/microgram DNA). Specific activity was not significantly different after 96 h (L2: normoxia, 261 +/- 11; hypoxia, 261 +/- 14 U/mg enzyme protein). These results indicate that regulation of glycolysis during chronic hypoxia occurs at the level of enzyme content. Chronic O2 depletion leads to either an increased rate of biosynthesis or a decreased rate of biodegradation of PyKi, causing augmented glycolytic capacity. Monoclonal antibodies provide a highly specific, convenient approach to charcterizing enzymes, as well as quantitating cellular enzyme content.

Animals↗
PubMed1980

Impairment of phagocytosis by moderate hyperoxia (40 to 60 per cent oxygen) in lung macrophages.

Exposure of isolated mouse lung macrophages to 40 and 60 per cent oxygen in tissue culture for 48 hours resulted in significant depression of phagocytosis as compared to air-exposed controls. The impairment of phagocytosis was reversed when the cells were reexposed to normoxic conditions for 48 hours. The impairment of phagocytosis occurred despite significant increases in intracellular superoxide dismutase activity, an enzyme felt to play a protective role in oxygen toxicity. Exposure to 40 and 60 per cent oxygen increased the susceptibility of lung macrophages to functional impairment by 95 per cent oxygen, rather than producing tolerance. The precise biologic and clinical significance of these findings will require additional studies in integrated systems. However, these studies show unequivocal lung macrophage injury with moderate hyperoxic exposure.

Animals↗
PubMed1979

Effects of sustained oxygen depletion on tissue pyruvate kinase activities in the freshwater turtle, Pseudemys scripta elegans.

1. The effect of sustained (48 hr) oxygen depletion on tissue (brain, heart, skeletal muscle) pyruvate kinase (PyKi) activities was examined in the pond turtle (Pseudemys scripta elegans). 2. PyKi activities in skeletal muscle are significantly increased (from 322 +/- 84 to 450 +/- 95) following 48 hr of tissue hypoxia. PyKi activities in brain may already be elevated under ambient conditions but do not change following prolonged submersion (334 +/- 75 vs 325 +/- 77). Cardiac muscle PyKi is actually decreased (from 135 +/- 35 to 94 +/- 18) under anaerobic conditions. 3. The data suggest that prolonged O2 depletion may increase PyKi biosynthesis in turtle skeletal muscle (subserving enhanced glycolysis), but also demonstrate that factors other than oxygen availability are involved in the regulation of tissue PyKi activities in intact vertebrates.

Animals↗
PubMed1979

Bioenergetic pattern of turtle brain and resistance to profound loss of mitochondrial ATP generation.

The adaptations in the freshwater turtle that permit survival despite prolonged loss of mitochondrial ATP generation were investigated by comparing the bioenergetics of turtle brain slices with rat brain slices. Aerobic turtle brain shows no significant difference in basal levels of total ATP generation compared to rat brain; levels in turtle brain and rat brain were 18.4 +/- 2.8 (SD) and 19.4 +/- 2.2 mumol (100 mg of tissue)-1 hr-1, respectively. However, in turtle brain, a significantly greater fraction of ATP is derived from glycolysis both under aerobic and anaerobic conditions [aerobic turtle (24%) and rat (13%), P less than 0.02; anaerobic, turtle (28%) and rat (18%), P less than 0.05]. The increased glycolytic capacity is related to high levels of rate-limiting glycolytic enzymes, such as pyruvate kinase (EC 2.7.1.40). Turtle brain operates close to glycolytic capacity even under aerobic conditions, and no Pasteur effect can be demonstrated. Quantitatively, anaerobic glycolysis accounts for a maximum of 28% of basal aerobic ATP generation, suggesting that prolonged diving is also accompanied by a reduction in brain energy requirements. The adaptation subserving short-term (natural) diving is an increase in brain glycolytic capacity. The adaptation subserving prolonged diving (days to weeks) may be a reduction in the energy requirements of brain (and other cells).

Animals↗
PubMed1979

Effects of high oxygen exposure on bioenergetics in isolated type II pneumocytes.

O2-mediated alterations in cell energy metabolism may play a role in structural and functional abnormalities described in type II pneumocytes (T-II-P) following in vivo hyperoxia. Bioenergetic alterations produced by hyperoxia (95% O2) were therefore examined in a culture-maintained cell line derived from T-II-P. Exposure of cell monolayers to 95% O2 for 96 h results in a significant decrease in O2 consumption (from 0.52 +/- 0.07 to 0.30 +/- 0.08, P less than 0.01), suggesting impaired mitochondrial energy provision. In addition, there are increased rates of aerobic lactate production (from 2.89 +/- 0.52 to 3.84 +/-0.80, P less than 0.05) with loss of Pasteur effect, indicating a shift to glycolytic metabolism at relatively high PO2's. These metabolic changes are not accompanied by altered activities of critical mitochondrial (cytochrome oxidase) or glycolytic (pyruvate kinase, phosphofructokinase) enzymes. Altered cell bioenergetics following hyperoxia may this represent an important secondary mechanism leading to functional abnormalities in T-II-P.

Aerobiosis↗
PubMed1978

Bioenergetic pattern of isolated type II pneumocytes in air and during hypoxia.

The bioenergetic pattern of a cell clone derived from rat lung with ultrastructural and biochemical characteristics like those of type II pneumocytes (T-II-P), has been studied in a tissue culture system. During air cultivation, these cells have a high rate of aerobic and anaerobic glycolysis associated with high activities of two rate-limiting enzymes in glycolysis (pyruvate kinase [PyKi] and phosphofructokinase [PFK]). This is present despite the rates of oxygen consumption and activities of cytochrome oxidase (CyOx) similar to other lung cells. Presumably the high rate of aerobic glycolysis explains the substantial lactate production previously described in lung slices and in the intact perfused lung. Hypoxic cultivation results in a decrease in CyOx. Acute re-exposure to air does not restore the oxygen consumption to normal, presumably as a result of decreased mitochondrial O(2) utilization associated with decreased CyOx activity. As a result, hypoxically cultivated T-II-P cells have a decreased capacity for mitochondrial ATP generation in air as compared to air-cultivated cells. During hypoxia, aerobic and anaerobic glycolysis are further increased as well as the activities of PyKi and PFK. The high rate of glycolysis and high activities of PyKi and PFK in cultivated T-II-P appear to reflect intrinsic genetic regulation. The decreased CyOx activity and increased PyKi and PFK activities in hypoxic T-II-P appear to reflect alterations in enzyme biosynthesis/biodegradation regulated by O(2) availability.

Animals↗