Hemolytic anemia with impaired hexokinase activity

Keitt, Alan S.

doi:10.1172/jci106165

Cited by 52 publications

(8 citation statements)

References 31 publications

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“…The samples were continuously equilibrated with moistened gas mixtures consisting of nitrogen plus 20%, 3.3%, or 0% oxygen, and pH was maintained at 7.45 4i 0.05 by varying the CO2 concentration with a gasometric pH stat (8). Supernatant K+ concentration was determined by flame photometry.…”

Section: Methodsmentioning

confidence: 99%

Dimethyl adipimidate: a new antisickling agent.

Lubin¹,

Peña²,

Mentzer³

et al. 1975

Proc. Natl. Acad. Sci. U.S.A.

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A new approach to the prevention of sickling in vitro by use of the bifunctional crosslinking reagent, dimethyl adipimidate, is described. Prior treatment of sickle erythrocytes with dimethyl adipimidate will inhibit sickling in completely deoxygenated erythrocytes. Treated erythrocytes do not demonstrate the potassium loss and viscosity increase that usually accompany sickling. The oxygen affinity of hemoglobin in these cells is increased independently from changes in the concentration of 2,3-diphosplioglycerate. The (v/v) formaldehyde in saline for fixation. Five hundred cells were counted by described criteria (6). The viscosity of dimethyl adipimidate-treated and control cells was measured in a Wells Brookfield cone plate microviscomneter model LVT, adapted to maintain controlled atmospheric gas tension (7). After adjustment of the hematocrit to 40 i 1%, the blood was equilibrated for 1 hr with humidified 95% N2 and 5% CO2 in an IL tonometer model no. 237, then transferred in a closed system to the viscometer. The viscosity was measured at shear rates ranging from 4.5 to 90 sec' during continued exposure to nitrogen.Studies in vitro of erythrocyte metabolism and of net K + loss were conducted by incubation at 370 of thrice-washed erythrocytes in Krebs Henseleit buffer with added glucose (10 mM) in the presence of 0.1 mM ouabain. The samples were continuously equilibrated with moistened gas mixtures consisting of nitrogen plus 20%, 3.3%, or 0% oxygen, and pH was maintained at 7.45 4i 0.05 by varying the CO2 concentration with a gasometric pH stat (8). Supernatant K+ concentration was determined by flame photometry. 2,3-Diphosphoglycerate (9) concentrations in erythrocytes were determined in perchloric acid extracts.The oxygen affinity of whole blood, expressed as p50 (that oxygen tension at which hemoglobin is half-saturated), was determined by measurement of hemoglobin oxygen saturation and P02 after equilibration of the samples with gas mixtures containing 2.2%, 2.8%, 3.2%, and 3.9% oxygen plus 5% CO2 and the remainder nitrogen. The p50 was calculated by a best fit analysis of these data and corrected to a pH of 7.4 (10).Hemolysates and globin were prepared by standard methods. Gel filtration in Sephadex G-100 was performed by the method of Andrews (11). The column was calibrated with aldolase (molecular weight 158,000), ovalbumin (molecular weight 45,000), and chymotrypsinogen A (molecular weight 25,000). Hemoglobin electrophoresis was performed on cellulose acetate strips at pH 8.65 in a Beckman microphor system. Barbital buffer, 25 mMI, pH 8.0, was used for electrophoretic separation of globin chains and the cellulose acetate strips were soaked in the same buffer with 8 M urea and 5 mM 2-mercaptoethanol. Spectra were recorded with a Cary 14 recording spectrophotometer.

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Section: Methodsmentioning

confidence: 99%

Dimethyl adipimidate: a new antisickling agent.

Lubin¹,

Peña²,

Mentzer³

et al. 1975

Proc. Natl. Acad. Sci. U.S.A.

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“…A further interesting result provided by these studies is that HK deficiency can influence the glucose metabolic ability of cells other than the erythrocyte. So far we and others (Valentine et al, 1967;Keitt, 1969;Board, 1978) have been able to show that in HK deficiency, red blood cells metabolize reduced amounts of glucose but the rate of the pentose phosphate pathway is the same. Now we know that this metabolic impairment is also present in cells other than the erythrocyte and it must be further evaluated to understand fully the role of HK in glucose metabolism.…”

Section: Discussionmentioning

confidence: 81%

“…Reports on red blood cell HK deficiency have shown that surviving erythrocytes have a reduced ability to utilize glucose (Valentine et al, 1967;Keitt, 1969;Board et al, 1978) because of the haemolytic anaemia usually associated with HKdeficiency; the results probably underestimate the real glycolytic rate because of the cell age dependence of this metabolic pathway. Reduced ability to utilize glucose must also be expected in tissues containing HK type I as the predominant glucose phosphorylating enzyme.…”

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confidence: 99%

Glucose metabolism in fibroblasts from patients with erythrocyte hexokinase deficiency

Magnani

Chiarantini

Stocchi

et al. 1985

J of Inher Metab Disea

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Four different hexokinase (HK) isoenzymes are distributed in different proportions in human tissues. Fibroblasts contain HK type I as the predominant glucose phosphorylating activity, the same isoenzyme that predominates in red blood cells (RBC). We have established cell lines from two patients homozygous for RBC HK deficiency but carrying different mutations. In one case (HK-Melzo) the residual RBC enzyme shows a marked heat instability but possesses normal kinetic and regulatory properties; in the other (HK-Napoli), the enzyme is characterized by an increased Ki for glucose-1,6-diphosphate. These properties are also retained by the fibroblasts' hexokinase. Glucose utilization by cultured fibroblasts from these patients was markedly reduced in the cell lines where HK deficiency was more pronounced. However, cells with only 30% HK activity retained their full ability to utilize glucose in the hexose monophosphate pathway. This was shown to be true not only under basal conditions but also in the presence of oxidative agents such as methylene blue. Significant reduction of the ATP level was also found in HK-Melzo fibroblasts. Thus, HK deficiency is associated with reduced glucose utilization and normal hexose monophosphate shunt rates. Results previously obtained on RBC support similar conclusions.

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“…The kinetic (2) and Received for publication 9 September 1977 and in revised formn 3 April 1978. electrophoretic (3) behavior of the enzyme showed no abnormalities. Since the report of Valentine et al (2), some other variants of hexokinase deficiency related to hereditary nonspherocytic hemolytic anemia have been reported (4)(5)(6)(7). Most variants showed increased Michaelis-Menten constants for the substrates glucose and MgATP2-(5-7), and in one case a decreased heat stability was found (5).…”

Section: Introductionmentioning

confidence: 99%

Human erythrocyte hexokinase deficiency. Characterization of a mutant enzyme with abnormal regulatory properties.

Rijksen¹,

Staal²

1978

J. Clin. Invest.

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A B S T R A C T In the erythrocytes of a patient with hereditary nonspherocytic hemolytic anemia, a homozygous expression of hexokinase deficiency was detected. The mutant enzyme was characterized by normal kinetic parameters with respect to its substrates, glucose and MgATP2-, normal pH optimum, normal heat stability at 40°C, but abnormal behavior with respect to its regulation by glucose-1,6-diphosphate and inorganic phosphate, and an altered electrophoretic pattern. Interpretation of the results revealed the presence of two different hexokinases type I in normal human erythrocytes: one enzyme with a high affinity for glucose-1,6-diphosphate, the inhibition of which is regulated by inorganic phosphate; and another enzyme with a lower affinity for the inhibitor, not regulated by inorganic phosphate. The former enzyme was not detectable in the erythrocytes of the patient, whereas the presence of the latter enzyme could be demonstrated.

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Hemolytic anemia with impaired hexokinase activity

Cited by 52 publications

References 31 publications

Dimethyl adipimidate: a new antisickling agent.

Dimethyl adipimidate: a new antisickling agent.

Glucose metabolism in fibroblasts from patients with erythrocyte hexokinase deficiency

Human erythrocyte hexokinase deficiency. Characterization of a mutant enzyme with abnormal regulatory properties.

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