Lixumistat (IM156)

IM156 (HL156A) treatment of RPMCs inhibits HG-induced myofibroblast transdifferentiation and markers of epithelial-mesenchymal transition (EMT). Moreover, IM156 (HL156A) ameliorates HG-induced transforming growth factor-β1, Smad3, Snail, and fibronectin expression in the RPMCs via AMPK upregulation.^[3] IM156 treatment decreases complex I-dependent NADH oxidation in a significant, dose-dependent manner.^[2]

RPMCs are seeded at a density of 1 × 10⁵ cells/well on a six-well plate. RPMCs are grown to 70% confluence and serum-starved under NG conditions for 24 h. The specific AMPKα1 small interfering (si)RNA, AMPKα2 siRNA, or control siRNA are transfected into the cells using a siRNA transfection reagent according to the manufacturer’s procedure. AMPKα1 siRNA, AMPKα2 siRNA, or control siRNA are incubated at final concentration of 20 μM using siRNA dilution buffer for 30 min at RT. The cells are washed twice with 2 ml of siRNA transfection medium, and 0.8 ml siRNA transfection medium and 200 μl siRNA transfection reagent complex are added to the well, covering the entire layer. The contents of the plate are gently mixed by swirling to ensure that the entire cell layer is immersed in the solution. Then, the cells are incubated for 5 h at 37°C in a 5% CO2 incubator. After that, 1 ml of normal growth medium is added. Next, the cells are incubated for an additional 24 h. Then, the media are replaced with fresh completed media and treated with NG or HG in the presence or absence of IM156 (HL156A) for 24 h.

In vivo treatment of IM156 exacerbated the memory differentiation of virus-specific CD8+ T cells results in an increase in short-lived effector cells but decrease in memory precursor effector cells. Thus, IM156 treatment impaires the function of virus-specific memory CD8+ T cells, indicating that excessive AMPK activation weakens memory T cell differentiation, thereby suppressing recall immune responses.^[1]