Characterization of Beta Tubulin Isotypes During Foam Cell Formation

Abstract

AbstractFoam cells contribute to the development of a cardiovascular condition called atherosclerosis. They arise when monocytes become engorged and lipid-laden after exposure to native low-density lipoproteins (Falk, 2006). It is assumed that the cytoskeleton is responsible for the morphological changes observed during foam cell formation. Beta tubulin and alpha tubulin are proteins that dimerize and polymerize to form microtubules, which are an important component of the cytoskeleton (Joshi, 1998). Little is known regarding the changes in cytoskeletal composition, particularly that of beta tubulins, throughout foam cell induction. The purpose of this study was to elucidate the expression patterns of beta tubulin isotypes 1–4 in human THP-1 monocytes throughout foam cell formation and to determine what relationship exists between beta tubulin expression and foam cell lipid aggregation. Levels of beta tubulin 1–4 were measured by western blot and immunofluorescence throughout the stages of foam cell differentiation, and beta tubulin isotypes were manipulated by siRNA to determine the effects of diminished beta tubulin expression on foam cell formation. Regardless of isotype, beta tubulin was always present in the highest amounts in monocytes. Levels of beta tubulin-1 and -4 were significantly decreased in macrophage and foam cells relative to monocytes (p < 0.0079, p < 0.0208, respectively). Beta 3 levels also exhibited a decrease. Beta 2 levels remained low regardless of differentiation stage. The distribution of beta tubulin 1 was shown to be more spindle-like (stretching across cells), compared to beta tubulins 2, 3, and 4, which exhibited a more “clumped,” less interconnected arrangement. When expression of beta tubulin isotypes 1, 3, and 4 were reduced in monocytes, resulting foam cells appeared to have more lipid aggregates and were significantly larger (p<0.0001) when compared to the size of foam cells without siRNA following treatment with PMA + LDL. In conclusion, the distribution of beta tubulins 1, 3, and 4 changes throughout the stages of foam cell induction, and manipulation of beta tubulins altered foam cell formation. Unexpectedly, the silencing or decreasing of beta tubulin enhanced lipid aggregation. Information concerning how beta tubulin expression can effect foam cell formation may offer insight into how to reduce plaque formation in patients with atherosclerosis.