Alejandra Jiménez Escobar is completing her Bachelor of Arts Degree in Biology at UNAM, Mexico City. For her independent research, she is studying the changes in fibroblasts focal adhesions depending on the substrate stiffness in LANSBioDyT, Mexico City. This summer, she is working in Rebecca Wells’ lab on a project to determine the nuclear and cytoskeletal mechanical stress generated by short chain fatty acids. Post-graduation, Alejandra will apply for graduate school to continue mechanobiology research.
Nuclear and cytoskeletal mechanical stress generated by short chain fatty acids in HuH7 cells
Hepatocellular carcinoma (HCC) predominantly occurs in patients with cirrhotic liver and is associated with increased matrix stiffness. Yet, in patients with non-alcoholic fatty liver disease (NAFLD), HCC can occur without increased liver stiffness. In NAFLD, hepatocytes are characterized by lipid droplets that occupy most of their cytoplasmic space. Our group has hypothesized that accumulation of intracellular lipid may generate nuclear and cytoskeletal mechanical stress, either through the physical presence of lipid droplets or the disruption of cellular mechanosensing. As lipid accumulation is impacted by fatty acid composition, here we investigated the effect of three different short chain fatty acids (SCFAs) on mechanosensing in a HCC cell line. HuH7 cells were cultured in PAA hydrogels of 500 Pa (normal liver stiffness), 10 kPA (cirrhotic liver stiffness) and glass and they were treated with different SCFAs (acetate, propanoate, and butyrate). IImmunofluorescence staining was completed to mark actin fibers (rhodamine-phalloidin), lipid droplets (Bodipy) and the nucleus (DAPI). Somewhat surprisingly, we found that treatment with SCFAs does not induce lipid droplet formation in HuH7 cells; however, normal mechanosensing appears disrupted. Specifically, acetate treatment increases cell spread area on all stiffness substrates over BSA control. Cell area then linearly decreases as carbons are added to the fatty acid chain . Additionally, SCFA treatment decreases actin intensity. YAP nuclear intensity and localization increases with stiffness for all treatment groups, yet we also find that SCFAs may impact YAP localization on stiff substrates. Hence, we propose that SCFAs alter the mechanosensitivity of HuH7 cells.