Effect of Fatty Acid Modification on Prostacyclin Production in Cultured Human Endothelial Cells (Arachidonate, Elcosapentaenoate)

An endothelial cell culture system was used to compare the effects of endogenous arachidonic acid (AA) and eicosapentaenoic acid (EPA) on prostacyclin synthesis. Supplemental levels of fatty acids (20-150 (mu)M) were supplied as sodium salts bound to bovine serum albumin (3:1 mole ratio) and added to M-199 with 20% fetal bovine serum. Both AA and EPA were readily incorporated into neutral and phospholipid fractions to serve as endogenous precursors for prostacyclin (PGI(,2)) synthesis. Thrombin (.5 U/ml) was used to stimulate prostacyclin synthesis. AA enrichment did not result in an increased PGI(,2) synthesis while EPA enrichment decreased PGI(,2) synthesis as measured by radioimmunoassay to 6-keto-PGF(,1(alpha)). Radiolabelled fatty acid eicosanoid products were separated by reverse phase HPLC. Unconverted fatty acid was the major product released upon thrombin stimulation in both cases. ('3)H-AA supplemented cells also released PGI(,2) and HETE in lesser amounts while ('14)C-EPA supplemented cells released only minimal amounts of PGI(,3) and HEPE. The relative percent incorporation of radiolabelled ('3)H-AA and ('14)C-EPA into cellular lipids was determined after two-dimensional thin-layer chromatography. Neutral lipid contained the vast majority of radiolabel ((TURN)50%). Of the phospholipids, phosphatidylcholine contained the greatest percentage of radiolabel. Phosphatidylinositol and phosphatidylethanolamine contained intermediate percentages while phosphatidylserine and sphingomyelin contained the least. In a dual label supplementation regimen (150 (mu)M ('14)C-EPA and 16 (mu)M ('3)H-AA), phosphatidylinositol appeared to be readily saturated with fatty acid at an early stage. This may be an important regulatory aspect for prostaglandin synthesis.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Food Science

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2015

Author dc:creator

• Basgall, Edward Joseph

• Health Sciences, Nutrition

Language dc:language

eng

Identifier

(UMI)AAI8409859

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/77444