N. Valsangkar1, X. Jin3, Z. Zhang3, T. A. Zimmers2,3, L. G. Koniaris2,3 2Indiana University School Of Medicine,Surgery,Indianapolis, IN, USA 3Thomas Jefferson University,Surgery,Philadelphia, PA, USA
Introduction: Fatty liver of any degree is associated with propensity to liver failure in mice and patients. Normal, young lean mice tolerate 70% hepatectomy well, with 100% survival and complete recovery of liver mass by 7 days. In contrast, 70% hepatectomy in diet-induced obese mice is characterized by increased liver injury and hepatocyte necroapoptosis, delayed mitoses and slower recovery of liver mass. The genetically obese, leptin receptor mutant lepr-db/db mouse displays high mortality after 70% hepatectomy, due to increased injury and defective proliferation. Others have shown this is associated with reduced expression of EGFR. Here we sought to correct liver regeneration in lepr-db/db mice by enhancing EGFR expression .
Methods: Genetically obese Lepr-db/db and age-, strain- and sex-matched wild-type mice were subjected to sham surgery or 70% hepatectomy. Introduction of EGFR or control plasmids was conferred by hydrodynamic injection of naked plasmid DNA. Survival and regeneration were assayed by serum liver enzymes, histology, Western blotting analysis and recovery of liver mass.
Results: Kaplan-Meier survival analysis of survival after 70% hepatectomy revealed significantly greater mortality in lepr-db/db mice (50% survival) versus wild-type controls (100% survival, p>0.01). Among survivors, lepr-db/db mice demonstrated increased serum AST, ALT, and bilirubin (indicating cell death and liver dysfunction) versus wild-type mice. Histologic examination of lepr-db/db livers revaled areas of necrosis and hemorrhage, vacuolar change, and mitotic figures. Recovery of liver mass was delayed. After hepatectomy, lepr-db/db mice failed to increase EGFR expression as observed in the wild-type mice and furthermore showed little phosphorylated Y1068-EGFR. At 6h post-hepatectomy, mouse liver extracts in db/db mice showed increased total hepatic protein carbonylation, indicating increased oxidative stress, and at 48h, reduced expression of pAkt, CyclinD1, and PCNA, demonstrating impaired hypertrophy and proliferation. Delivery of EGFR via transgenesis through hydrodynamic injection in db/db mice did not improve survival or regeneration.
Conclusion: Injury and impaired proliferation are very early events in the abnormal liver regeneration observed in db/db mice. This mechanism likely contributes to mitochondrial damage, impaired liver function, increased hepatocyte death and impaired proliferation in db/db mice. Regeneration in genetically obese mice cannot be corrected by EGFR replenishment alone.