MTCBP-1 Protein Expression is Enhanced in Fat Overloaded HepG2

Author(s): Masego Tsimako Johnstone, Paul Kroon, Peter Wilce

Background: Macrovesicular steatosis, is characterised as excessive triglyceride accumulation in the liver which perpetuate cell injury through signalling cascades. Although there are several hypotheses that have been proposed in liver damage, it is unclear as to how the events occur as well identification of the key proteins that facilitate this event. Aims: HepG2 in vitro fat cell model was used to investigate the relationship between cellular triglycerides and MTCBP-1 expression. Methods: The cells were treated with linoleic acid conjugated to defatted BSA. There were two controls to the experiments, consisting of cells grown in 10% defatted BSA and normal growth medium. Triglycerides were analysed by a quantitative standard curve and nile red staining. Western blots were used for detection of MTCBP-1 in transfected and untransfected cells grown in LA-BSA and compared with control cells. The subcellular localisation of MTCBP-1 was also determined by immunofluorescence. Results: Nile red staining of HepG2 showed presence of triglyceride mainly in the cytoplasm, with higher amounts of lipid droplets found in LA-BSA treated cells. The lipid treated cells contained a significantly increased amount of triglycerides (ρ = 0.004). Lipid overloaded HepG2 cells transfected with pSG5 plasmid encoding c-myc-MTCBP-1 contained much lower triglycerides when compared with the untransfected HepG2 cells. There was an increased expression of endogenous MTCBP-1 in lipid overloaded cells. MTCBP-1 is located in the cytoplasm and nucleus of the cell. Conclusion The increased expression of MTCBP-1 during triglyceride accumulation may suggest a relationship between the protein and triglyceride synthesis. It can further be explored as a potential target in therapeutics/diagnostics. The next step is to understand the interaction of MTCBP-1 with other proteins in triglyceride synthesis and packaging.

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