Abstract
Prostaglandins and endogenous cannabinoid metabolites share the same lipid backbone with differing polar head groups at exactly the position through which a large molecule is attached to provide antigenicity and thus raise antisera. Hence, we hypothesized that antisera raised against prostaglandins linked to a large molecule such as BSA at the carboxyl functional group would also recognize endogenous cannabinoid metabolites and lead to highly misleading interpretations of data. We found major cross-reactivity of commercial antisera raised to prostaglandins with endocannabinoid metabolites. Furthermore, in a well-characterized cell line (WISH) or primary amnion tissue explants, endocannabinoid treatment led to increased production of endocannabinoid metabolites as opposed to primary prostaglandins. This was apparent only after separation of products by thin-layer chromatography, because they measured as prostaglandins by radioimmunoassay. These findings have major implications for our interpretation of data in situations in which these prostaglandin-like molecules are formed, and they stress the need for chromatographic or spectrometric confirmation of prostaglandin production detected by antibody-based methods.
Highlights
Prostaglandins and endogenous cannabinoid metabolites share the same lipid backbone with differing polar head groups at exactly the position through which a large molecule is attached to provide antigenicity and raise antisera
We have demonstrated for the first time that prostaglandin E2 (PGE2)-ethanolamide is a major product under conditions of evoked COX-2 activity and anandamide release; by extension, other prostamides may be formed
This finding raises the intriguing possibility that prostamide production may have been misidentified in previous studies as prostaglandin production under conditions in which anandamide may be released and COX-2 induced
Summary
Prostaglandins and endogenous cannabinoid metabolites share the same lipid backbone with differing polar head groups at exactly the position through which a large molecule is attached to provide antigenicity and raise antisera. In a well-characterized cell line (WISH) or primary amnion tissue explants, endocannabinoid treatment led to increased production of endocannabinoid metabolites as opposed to primary prostaglandins This was apparent only after separation of products by thin-layer chromatography, because they measured as prostaglandins by radioimmunoassay. Recent studies have demonstrated that anandamide and 2AG can be metabolized by cyclooxygenase-2 (COX-2) into prostaglandin-like molecules [7, 8] that consist of a prostaglandin with a polar head group, ethanolamide or glycerol, respectively. To determine the possible magnitude and significance of the problem, we investigated the relative production of these two prostamides by stimulated amnion-derived WISH cells and the production of PGE2-ethanolamide by primary human amnion explants
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