Fatty acids in human colonic mucosa

Wilson, Gillian Mary (1991) Fatty acids in human colonic mucosa. University of Southampton, Doctoral Thesis.

Abstract

Human colonic mucosa used throughout these studies was macroscopi-cally normal and obtained from patients undergoing resection for carcinoma. The relative sizes of individual phospholipids in the mucosa were estimated after lipid extraction of tissue, TLC and phosphorus determination. Phosphatidylcholine (44.1 ±8.9%) and phosphatidylscrinc/inositol (30.1 ±8.9%wcre significantly larger classes than either phosphatidyl-ethanolamine (13.2 ± 4.0%) or sphingomyclin (12.7 ± 4.5%) (mean ± S.D.). Phosphatidylcholine accounted for a significantly higher percentage of lipid phosphorus than phosphatidylserinc/inositol, p< 0.01.

The fatty acids present in a total lipid extract and in the separate phospholipids were estimated using TLC and gas liquid chromatography. In total extracts arachidonic acid made up 6.2% of fatty acids present. Expressed in terms of the phospholipid unit, phosphatidyl-cholinc and phosphatidyl-serine/inositol contained greater amounts of arachidonic acid 0.5 ±0.5 nmol AA/nmol phosphorus (mean ± S.D.) than sphingomyclin (0.2 ± 0.3), p< 0.05. The AA content of phosphatidylethanolaminc (0.4 ± 0.3 nmol AA/nmol phosphorus) did not differ from other phospholipids.

During incubation of fresh but not previously frozen mucosa with 14C-AA (0.3/iCi) approximately 6% became incorporated into the tissue. Of this, 36% was associated with neutral lipids, 15% with phosphatidylcholinc, 10% with phosphatidylethanolaminc, 10% with phosphatidylscrinc/inositol and 28% co-chromatographed on TLC with 'free' arachidonic acid.

Arachidonate release was time-dependent and varied between phospholipid classes and neutral lipids. 'Free' arachidonatc was resistant to depletion in 4 and 18 hour incubations.

In a 30 minute incubation of mucosa pro-labelled with 14C-AA and then incubated with 25 units PLC, there was a significant release of 14C-AA from the phosphatidylcholine class whereas there was a significant increase in that associated with the neutral lipid class.

Using gel-filtration and SDS-Pagc techniques a proportion of the UC-AA in the 'free' arachidonic acid pool was found to be protein bound. The most abundant protein present was human serum albumin; lesser amounts of other proteins such as fatty acid-binding protein were also detected.

In preliminary experiments no clear change in phospholipid mass (nmol phosphorus/mg protein) could be detected in up to 18 hour incubation. Preliminary work in developing techniques was also carried out on the sub-classes of phosphatidylcholine present.

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