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Interaction of lipopolysaccharide with a mammalian lyso-phosphatidate acyltransferase (LPAAT) transfected into E. coli, and effect of lisofylline on LPAAT transfected into mammalian cells.
1. Lipid A and LPS stimulate LPAAT activity (and hence unsaturated PA formation) in RMC membranes and whole cells. 2. This correlates with cell phenotypic and membrane changes associated with small G proteins. 3. Unsaturated PA and Lipid A have similar effects on cells when given exogenously. 4. Human LPAAT-alpha and -beta isoforms were cloned and transfected into E. coli, demonstrating the ability to restore PA synthesis and reduce lyso-PA accumulation in plsC strains (LPAAT deficient mutants), as well as restoring growth at high temperatures. 5. LPAAT transfection into E. coli plsC (JC201) strains results in an increase in LPS content, suggesting stimulation of LPS synthesis. 6. LPAAT transfection into human A549 lung epithelial carcinoma and endothelial ECV304 cells results in increased cytokine mRNA transcription at baseline, and a significant increase in stimulated cytokine mRNA transcription. In addition, LPAAT transfection also results in increased cytokine release in response to IL-1 beta. 7. LSF, which reduces rodent deaths in sepsis models, reduces unsaturated acyl incorporation into PA in monoblastic cell lines, and reduces serum FFA increase in human sepsis, also reduces unsaturated acyl incorporation into PA in ECV304 cells. LPAAT-alpha transfection increases linolenate incorporation into PA at the expense of linoleate incorporation, which is reversed by LSF. LPAAT-beta increases both linoleate and linolenate incorporation into PA, which is also reduced by LSF. We conclude that LPAAT and PA remodeling may play a role in diffuse renal toxicity in sepsis due to induction of cellular phenotype changes associated with PA induction by Lipid A and/or LPS. Two human isoforms of LPAAT have been cloned, and apparently address C18 unsaturated acyl chains somewhat selectively. LSF causes functional reduction in LPAAT activity in transfected systems. This does not yet imply a direct effect of LSF on LPAAT. LPAAT and LPS may interact in the membrane in a not-yet-understood manner.
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