AIM: To investigate the effects of mammalian target of rapamycin (mTOR) 
inhibition on liver regeneration and autophagy in a surgical resection model.
METHODS: C57BL/6 mice were subjected to a 70% partial hepatectomy (PH) and 
treated intraperitoneally every 24 h with a combination of the mTOR inhibitor 
rapamycin (2.5 mg/kg per day) and the steroid dexamethasone (2.0 mg/kg per day) 
in phosphate buffered saline (PBS) or with PBS alone as vehicle control. In the 
immunosuppressant group, part of the group was treated subcutaneously 4 h prior 
to and 24 h after PH with a combination of human recombinant interleukin 6 
(IL-6; 500 μg/kg per day) and hepatocyte growth factor (HGF; 100 μg/kg per day) 
in PBS. Animals were sacrificed 2, 3 or 5 d after PH and liver tissue and blood 
were collected for further analysis. Immunohistochemical staining for 
5-Bromo-2'-deoxyuridine (BrdU) was used to quantify hepatocyte proliferation. 
Western blotting was used to detect hepatic microtubule-associated protein 1 
light chain 3 (LC3)-II protein expression as a marker for autophagy. Hepatic 
gene expression levels of proliferation-, inflammation- and angiogenesis-related 
genes were examined by real-time reverse transcription-polymerase chain reaction 
and serum bilirubin and transaminase levels were analyzed at the clinical 
chemical core facility of the Erasmus MC-University Medical Center.
RESULTS: mTOR inhibition significantly suppressed regeneration, shown by 
decreased hepatocyte proliferation (2% vs 12% BrdU positive hepatocyte nuclei at 
day 2, P < 0.01; 0.8% vs 1.4% at day 5, P = 0.02) and liver weight 
reconstitution (63% vs 76% of initial total liver weight at day 3, P = 0.04), 
and furthermore increased serum transaminase levels (aspartate aminotransferase 
641 U/L vs 185 U/L at day 2, P = 0.02). Expression of the autophagy marker 
LC3-II, which was reduced during normal liver regeneration, increased after mTOR 
inhibition (46% increase at day 2, P = 0.04). Hepatic gene expression showed an 
increased inflammation-related response [tumor necrosis factor (TNF)-α 3.2-fold 
upregulation at day 2, P = 0.03; IL-1Ra 6.0-fold upregulation at day 2 and 
42.3-fold upregulation at day 5, P < 0.01] and a reduced expression of cell 
cycle progression and angiogenesis-related factors (HGF 40% reduction at day 2; 
vascular endothelial growth factor receptor 2 50% reduction at days 2 and 5; 
angiopoietin 1 60% reduction at day 2, all P ≤ 0.01). Treatment with the 
regeneration stimulating cytokine IL-6 and growth factor HGF could overcome the 
inhibitory effect on liver weight (75% of initial total liver weight at day 3, P 
= 0.02 vs immunosuppression alone and P = 0.90 vs controls) and partially 
reversed gene expression changes caused by rapamycin (TNF-α and IL-1Ra levels at 
day 2 were restored to control levels). However, no significant changes in 
hepatocyte proliferation, serum injury markers or autophagy were found.
CONCLUSION: mTOR inhibition severely impairs liver regeneration and increases 
autophagy after PH. These effects are partly reversed by stimulation of the IL-6 
and HGF pathways.