Incorporation of orthophosphate-32P into erythrocyte phospholipids in normal subjects and in patients with hereditary spherocytosis

Reed, Claude F.

doi:10.1172/jci105946

Cited by 20 publications

(3 citation statements)

References 27 publications

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“…Incorporation of glycerides, glycerol, inorganic phosphorus, or cytidine diphosphate compounds were all minor in Release of Phospholipid Fatty Acid from Human Erythrocytes 1673 loo0 comparison to these two major incorporation routes (29,30). In other studies we have shown that for the active pathway the delivery of fatty acid from the plasma to the lysophosphatide acceptors within the cell proceeds through two discrete precursor pools (10).…”

Section: Discussionmentioning

confidence: 77%

Release of phospholipid fatty acid from human erythrocytes

Shohet¹

1970

J. Clin. Invest.

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A B S T R A C T To study the catabolism of erythrocyte phospholipids, human erythrocytes were labeled with radioactive fatty acid (FA). Labeling was performed by the two separate routes which together are thought to be responsible for the majority of phosphatide renewal in the red cell: (a) passive equilibration of erythrocytes with preformed acid-labeled red cell phosphatidylcholine (PC) and (b) active, "acylase"-dependent, incorporation of free fatty acid in the presence of ATP coenzyme A and magnesium. (As measured here "acylase" = the over-all effect of fatty acid thioesterification and the action of acyl-CoA: acylglycerophosphoryl acyltransferase.) The labeled cells were then reincubated in serum and the loss of radioactivity from cells into serum was examined.The phosphatide fatty acid introduced by these two routes was incorporated into two distinct cellular phosphatide pools with little mixing. The fatty acid in passively introduced phosphatidyl choline was released primarily as the intact phosphatide, whereas actively incorporated PC fatty acid was released primarily as free FA. As expected for an exchange process, the passively introduced PC was released at the same rate as it was introduced. However, the actively incorporated FA was released from the cell at a slower rate than its incorporation into PC. This latter phenomenon was explained by the observation of transfer of PC fatty acid to phosphatidyl ethanolamine and "neutral lipid" before its release to serum. This transfer process was inhibited by previous heating of the reincubation serum.The differences in the character of the released label, in the rate of release of the label, and in the influence of the reincubation serum all indicate separate pathways for the renewal of phosphatide introduced into red cells by these two routes.Part of the material presented here has previously been published in abstract form (1969 J. Clin. Invest. 48: 77a).

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Section: Discussionmentioning

confidence: 77%

Release of phospholipid fatty acid from human erythrocytes

Shohet¹

1970

J. Clin. Invest.

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“…The membrane nature of the defect in HS was first suspected from the finding of an increased Na+ permeability in intact erythrocytes, an observation that has been shown consistently (Bertles, 1957;Jacob & Jandl, 1964;Wiley, 1972). Other membrane abnormalities have been described, such as an increased 32p turnover in the phospholipids (Reed, 1968), a reduction in the amount of membrane lipid per cell (Cooper & Jandl, 1969a) and decreased drug-induced endocytosis (Schrier et al, 1974). Despite these defects in membrane function, there have been no gross abnormalities detected with respect to membrane phospholipid distribution or fatty acid composition (de Gier et al , 1964; 0306-3275/82/020259-08$01.50/1 (© 1982 The Biochemical Society Robertson & Lands, 1964), although minor differences in the amounts of long-chain fatty acids in some phospholipid types have been claimed but not substantiated (Kuiper & Livne, 1972;Zail & Pickering, 1979).…”

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confidence: 99%

Hereditary spherocytosis of man. Altered binding of cytoskeletal components to the erythrocyte membrane

Hill

Sawyer

Howlett

et al. 1982

Biochemical Journal

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Human erythrocytes possess a lattice work of extrinsic proteins on the inner face of the membrane (;cytoskeleton') that maintains the shape and deformability of the cell. The major proteins of the cytoskeleton are spectrin and actin, which are attached to the membrane by protein bands 2.1 (;ankyrin') and 4.1. The interactions of spectrin/actin with erythrocyte membranes from normal subjects and from patients with hereditary spherocytosis (HS) have been studied by using an air-driven ultracentrifuge, which can rapidly separate membranes from soluble proteins (150000g for 30s). The total amount of spectrin/actin in HS and normal ghosts is similar. However, the rate of dissociation of spectrin and actin from HS erythrocyte membranes at low ionic strength is significantly lower than that observed for normal membranes. Spectrin and actin isolated from either HS or normal membranes re-associated in a similar manner to spectrin/actin-depleted vesicles prepared from normal cells. Scatchard analysis showed an average binding capacity of 278mug/mg of membrane protein. However, spectrin/actin-depleted vesicles prepared from HS cells bound significantly less spectrin/actin prepared from either the normal or abnormal cells (average binding capacity 158mug/mg of membrane protein). The defect was defined further by studying the cytoskeleton obtained by Triton X-100 extraction of membranes. Under conditions of low ionic strength cytoskeletons prepared from HS membranes dissociated more slowly than those prepared from normal membranes, and only 80% of the protein from HS cytoskeletons could be solubilized after 180min compared with 100% for normal cytoskeletons. The difference between HS and normal membranes, which persists in isolated cytoskeletons, suggests that alterations in either the primary structure or the degree of phosphorylation of protein bands 2.1 or 4.1 may be central to the molecular basis of hereditary spherocytosis.

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“…Ces resultats ont 6tB confirm& par Reed [7] demontrant une incorporation intense de [32P]orthophosphate dans les acides phosphatidiques, plus faible dans les phosphatidyls&ines, celle-ci Atant cependant accrue dans les hematies de malades atteints de sph6rocytose hereditaire et en rapport avec une augmentation du transfert du sodium.…”

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Métabolisme phospholipidique des hématies nucléées

Soula

Souillard

Gard

et al. 1971

European Journal of Biochemistry

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The in vitro incorporation of [32P]orthophosphate into chicken-erythrocyte phospholipids has been studied by means of one-and two-dimensional thin-layer chromatography as a timerelated function: from 10 min to 6 h.Labelled phosphatidic acid appears after 10 min and represents about 90°/, of the total labelled phospholipids. Sphingomyelin turnover is as low as in mammal erythrocytes whereas glycerophospholipids' turnover is high. The specific activity of phosphatidylserine is ten times higher than that of phosphatidylethanolamine ; phosphstidylserine decarboxylase seems to he absent. In contrast with the metabolism of non-nucleated erythrocytes, the [32P]orthophosphate is incorporated into phosphatidylglycerol, the turnover of which is important.These data should agree with a de novo biosynthesis of phospholipids in nucleated erythrocytes, but many studies on mammal erythrocytes have demonstrated that there are essentially exchange reactions of phospholipids with plasma phospholipids and eventual acylation of plasma lysophospholipids. Par contre les phospholipides des hematies nucl66es ont 6th Btudihs seulement sur le plan analytique [8,9]; leur turnover n'a jamais BtB envisag6. L'absence de travaux dans ce domaine et les d86rences de composition phospholipidique entre les h6maties de mammifhres et les hematies nuclebes des oiseaux, nous ont incites Q entreprendre 1'Btude de l'incorporation in vitro de [32P]orthophosphate dans les globules rouges de poulet. MATfCRIEL ET MkTHODES Isolement et incubation des hkmatiesLe sang est pr6lev6 sur hhparine 8, des poulets de race White Leghorn, Bges de 2 Q 3 mois, Q la veine alaire.Aprks centrifugation Q 2500 tourslmin pendant 15 min, le plasma est rejet6 et les globules rouges laves A trois reprises au s6rum physiologique.

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Incorporation of orthophosphate-32P into erythrocyte phospholipids in normal subjects and in patients with hereditary spherocytosis

Cited by 20 publications

References 27 publications

Release of phospholipid fatty acid from human erythrocytes

Release of phospholipid fatty acid from human erythrocytes

Hereditary spherocytosis of man. Altered binding of cytoskeletal components to the erythrocyte membrane

Métabolisme phospholipidique des hématies nucléées

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