High glucose and advanced glycation end products induce phospholipid hydrolysis and phospholipid enzyme inhibition in bovine retinal pericytes

In the present study, we investigated the possible role of oxidative stress and the modulation of phospholipid turnover in two related models of peric yte injury, i.e., treatment with high glucose or advanced glycation end pro ducts (AGEs). Growing microcapillary pericytes from bovine retinas in cultu re were incubated, for 3 weeks, with 20-50 mM glucose or 2-20 muM AGEs, and peroxidation parameters (malondialdehyde, conjugated diene, hydroperoxide, glutathione (GSH) levels and lactate dehydrogenase (LDH) release) were eva luated. Arachidonate (AA) and choline release from membrane phospholipids w as determined in pericytes prelabeled with [1-C-14]arachidonate and [Me-H-3 ]choline, respectively, and stimulated with elevated glucose or AGEs for 30 min or 2 h. [1-C-14]arachidonate and [Me-H-3]choline incorporation into ph ospholipids, for 2 It and 3 h respectively, was also studied in conditioned and serum-starved cultures. Finally, lysates of treated and control cells were assayed for cytosolic phospholipase A(2) (cPLA(2)), acyl-CoA: 1-acyl-s n-glycero-3-phosphocholine O-acyltransferase (AT), CTP:phosphocholine cytid ylyltransferase (CT) and microsomal choline phosphotransferase (CPT) enzyme activities. We found that high glucose and AGEs caused neither significant production of reactive oxygen species nor cell toxicity or death, unlike o ther cell types. Both agents had no significant effect on the cellular ultr astructure, evaluated by light and electron microscopy, AA incorporation an d release, cytosolic phospholipase A(2) (cPLA(2)) and AT activities. Oil th e contrary, choline incorporation into phosphatidylcholine, CT and CPT acti vities were significantly reduced either by 50 mM glucose or 20 muM AGEs. S imultaneously, [Me-H-3]choline release was significantly stimulated by both agents, We conclude that prolonged treatments with high glucose or AGEs ar e not able to induce oxidative injury in bovine retinal capillary pericytes . Nevertheless, they do induce phospholipid hydrolysis and phospholipid enz yme activity inhibition. (C) 2001 Elsevier Science B.V. All rights reserved .