Phosphatidylcholine synthesis‐related enzyme activities of bovine brain microvessels exhibit susceptibility to peroxidation

Abstract

In microvessels isolated from bovine brain, microsomal enzyme activities involved in phosphatidylcholine biosynthesis and degradation were determined. The microvessels possessed acyl-CoA:l-acyl-sn-glycero-3-phosphoeholine (AT) and glycerophosphocholine phosphodiesterase (GroPChoPDE) activity at a higher level compared with bovine and rat brain or rat liver microsomes whereas they expressed CTP:phosphocholine cytidylyltransferase (CT) and choline phosphotransferase (CPT) activity at a lower level. Each enzyme has been characterized in terms of response to inldbitors or activators revealing properties very similar to those in brain and liver microsomes. In the homogenate prepared from t-butylhydroperoxide-treated microvessels (10 min exposure to 10 ~ up to 1 mM concentrations), AT and CPT activities exhibited a significant dose-dependent inhibition. In contrast, GroPChoPDE activity was unaffected. CT was inhibited only at 1 mM concentration. Short treatment of microvessels with Fe 2÷ (20 ~Vl)-ascorbate (0.25 mM) or 100 ~VI linoleate hydroperoxide did not have any effect on the activity of the four enzymes. Strong inhibition of all enzymes was noted when the linoleate hydroperoxide system was fortified by Fe 2÷ ions (100 ttM). AT inactivation was also found when oxidized low density lipoprotein was preincubated with microvessels. On the other hand, oxidized LDL left unchanged CPT and GroPChoPDE activities whereas it promoted a slight stimulation of cytidylyltransferase activity. Overall, the results suggest a link between oxygen radical generation and the perturbation of the microvessel membrane structure in which the four enzymes are incorporated, coupled to a direct sulfhydryl protein modification.

Astrocytes enhance radical defence in capillary endothelial cells constituting the blood‐brain barrier

“…Reactive oxygen species, i.e. O 2 − , OH, H 2 O 2 , ONOO − , cause lipid peroxidation [2] resulting in membrane disturbances [3,4] and can open the blood‐brain barrier [5,6]. Reactive oxygen species are continuously generated in the vascular system [7] and in EC [5], which is intensified during cerebral ischemia/reperfusion [5] or Alzheimer's disease [8,9].…”

Characterization of glycerophosphocholine phosphodiesterase activity and phosphatidylcholine biosynthesis in cultured retinal microcapillary pericytes. Effect of adenosine and endothelin‐1

“…Enzyme assays. Microsomal glycerophosphorylcholine phosphodiesterase (EC 3.1.4.2) was measured as described previously (2,18). The inhibitors to which the enzyme was exposed were EDTA and L-α-glycero-3-phosphate, under concentrations described in Table 2.…”

“…and 50 µM lyso-PtdCho as substrates, as previously described (17). CTP:phosphocholine cytidylyltransferase (CT) (EC 2.7.7.15) (2). Confluent cells grown on 60-mm dishes were stimulated or incubated with agonists.…”