Figure 2: Acetyl-CoA abundance is elevated in KRAS mutant acinar cells and inhibition of acetyllysine reading or cholesterol synthesis impairs acinar-to-ductal metaplasia.

In all panels, pancreatic acinar cells were harvested from 6–8-week-old wild-type (Pdx1-Cre; Cre) or (Pdx1-Cre;Kras^G12D; KC) mice. A, LC-MS quantification of acetyl-CoA in isolated acinar cells (n=3 mice, each group). B, isolated acinar cells (n=4 mice, each group) were cultured for 8 hours in the presence of the indicated ¹³C-labelled nutrient and acetyl-CoA labeling determined by LC-MS. C, mRNA expression of indicated genes in acinar cells after 48 hours culture in Matrigel, measured by qPCR (n=3 mice, each group). Mean value of each Cre column is equaled to 1, and data are normalized accordingly. Dashed blue line shows 1. D, schematic representation of acetyl-CoA and HMG-CoA labeling pattern from indicated, uniformly-labeled carbon sources. Compartmentalization also illustrated. E, HMG-CoA isotopologues after labeling of acinar cells (n=4 mice, each group) with 0.5 μM ¹³C-leucine for 8 hours. F, M+2 HMG-CoA in acinar cells labeled as in panel B. G, morphology of KC acinar cells embedded in collagen, treated with TGFα after 96 hours in the presence of the indicated inhibitors (n=3 mice, each group) Representative images shown. Scale bar, 50 μm. H, blinded quantification of ductal structures. Relative to Figure 2G. 35–50 images within 3 biological replicates were evaluated. I, collagen-embedded KC acinar cells after 96 hours treatment with atorvastatin, +/− mevalonate or cholesterol (n=3 mice, each group). Scale bar, 50 μm. J, blinded quantification of ductal structures. Relative to Figure 2E. 35–50 images within 3 biological replicates were evaluated. For all panels, columns show mean, +/− SD (*, p<0.05; **, p<0.01; ***, p<0.001).