In Vitro Study of the Incorporation and Transport of Nonesterified Fatty Acids into the Phospholipids of the Red Blood Cell Membranes of Cystic Fibrosis Patients

Abstract

SummaryThe in vitro incorporation and transport of plasma nonesterified fatty acids into phospholipids of red cell membranes have been studied in cystic fibrosis and healthy children.Red blood cells were labeled in vitro by an active "acyltransferase"-dependent incorporation of radioactively labeled nonesterified fatty acids. [3HI-Palmitic and ['4CI-linoleic acid, bound to albumin, have been studied simultaneously because it has been shown before that the concentration of palmitic acid increases and the concentration of linoleic acid decreases both in the plasma nonesterified fatty acid fraction and in the various phospholipids of the erythrocyte membranes of cystic fibrosis patients.The labeled cells were reincubated in autologous serum and the radioactivity present in the serum lipids and in the major phospholipid fractions of the erythrocyte membranes was measured.A general conclusion is that the in vitro turnover of labeled palmitic and linoleic acids in the phospholipids of the erythrocyte membranes is higher for cystic fibrosis patients than for healthy children. No difference is detectable between the in vitro behaviour of ['4CI-linoleic versus [3HI-palmitic acid in cystic fibrosis patients compared with healthy children.clusion of that study (8) is that the fatty acid composition of the plasma cholesterol esters of CF patients without pancreatic insufficiency is significantly different from the results obtained for agematched controls (8). Furthermore, it was found that the fatty acid patterns of the various phospholipid fractions of the RBC membranes are abnormal in CF patients with pancreatic insufficiency (24). A positive correlation was observed between the fatty acid changes of both plasma and RBC phospholipid fractions (Rogiers, V., unpublished data).One of the mechanisms, playing a major role in the continuous in vivo renewal of RBC lipids consists of the active incorporation of plasma NEFA's into the phospholipids of the RBC membrane (28, 29). Consequently it is possible that this process could be partly responsible for the abnormalities observed in the RBC membranes of CF patients. On the other hand, abnormal fatty acid patterns intrinsic to the C F membranes could support the hypothesis of a possible defect in fatty acid metabolism.In order to obtain more information concerning the active incorporation process of plasma NEFA's into the phospholipids of the RBC membranes of CF patients, a number of incorporation and reincubation experiments with labeled NEFA's have been carried out. RBCs of healthy and CF children were incubated with [14C1-linoleic and [3H1-palmitic acid bound to albumin. The labeled RBC'S were reincubated in their autologous serum.

Abnormal Fatty Acid Turnover in the Phospholipids of the Red Blood Cell Membranes of Cystic Fibrosis Patients (in Vitro Study)

“…Materials used for the analysis of the individual phospholipid fractions of the RBC (15,16), for the in vitro experiments (20) and for the radioactivity measurements (20) were described previously.…”

“…Indeed, it wasshown that abnormal FA patterns can occur in both CF patients with and without pancreatic insufficiency (17,18). Furthermore, it was found that the in vitro turnover of FA in the phospholipids of the erythrocyte membranes is significantly higher for CF patients than for healthy children (20). This conclusion has been drawn from in vitro experiments carried out with RBC, radioactively labeled with ['4C]linoleic and pH]palmitic acid, of CF patients with pancreatic insufficiency and of healthy children with their own serum (20).…”

“…The RBC were resuspended for 5 min in their own serum in order to restore the normal surface pool of nonradioactive FA. After removing the serum, the radioactively labeled RBC were incubated separately with autologous and homologous serum with additives as described (20). At various time intervals, aliquots were taken and centrifuged, and the RBC were washed and stored with dithionite at -20·e.…”

“…The methodology of the in vitro experiments has been described previously in detail (20). In short, blood was centrifuged and the packed RBC were washed.…”

“…Furthermore, it was found that the in vitro turnover of FA in the phospholipids of the erythrocyte membranes is significantly higher for CF patients than for healthy children (20). This conclusion has been drawn from in vitro experiments carried out with RBC, radioactively labeled with ['4C]linoleic and pH]palmitic acid, of CF patients with pancreatic insufficiency and of healthy children with their own serum (20). This increased in vitro turnover of the FA in the RBC of CF patients can be intrinsic to the membrane but may also be caused by extrinsic serum factors.…”