Synthetic Low-Density Lipoprotein,a Novel Biomimetic Lipid Supplement for Serum-Free Tissue Culture

摘要

Lipid supplementation in serum-free tissue culture employs solubilization techniques to permit the addition of lipids,but these systems are potentially cytotoxic and do not present lipid in a natural form.In this research a simplified preparation method for synthetic low-density lipoprotein (sLDL) has been developed that involves micro-fluidization of a solvent lipid solution in a simple aqueous solution.This produces material with size and t,potential characteristics similar to those of native LDL.sLDL supplementation in tissue culture media provides cholesterol concentrations higher than those achieved by 10% serum supplementation and existing chemically defined lipid supplements.sLDL stimulates NSO and U937 cellular proliferation in completely serum-free media,the former in a lipid concentration dependent manner that is also related to both the receptor peptide structure employed and its concentration on the particle.The greatest NSO cellular proliferation was obtained at the highest cholesterol concentration tested (0.5 mg/mL),which was 10 times higher than the cholesterol concentration achieved by standard 10% serum supplementation.U937 cellular proliferation was influenced by variation of sLDL's fatty acid constituents with a natural mixture producing maximal effect.Cell uptake studies in NSO with fluor-escently labeled sLDL indicated that assimilation is reduced by competition from native LDL.The planktonic nature of NSO cell growth meant that cell binding and uptake experiments were difficult to conduct because of cellular aggregation.However,sLDL-induced U937 proliferation is ablated by the presence of an anti-LDL receptor antibody. The results indicate that sLDL uptake is via the LDL receptor and that sLDL can function as a lipid supplement for serum-free media capable of supplementation to cholesterol concentrations up to 0.5 mg/mL.Cellular uptake studies also suggest that sLDL will be useful for the targeting and delivery of materials to cells.sLDL therefore represents a new and promising synthetic biomimetic alternative to native LDL with multiple applications.