Mismatched effects of receptor interacting protein kinase-3 on hepatic steatosis and inflammation in nonalcoholic fatty liver disease

AIM To validate the effects of receptor interacting protein kinase-3(RIP3) deletion in non-alcoholic fatty liver disease(NAFLD) and to clarify the mechanism of action.METHODS Wild-type(WT) and RIP3 knockout(KO) mice werefed normal chow and high fat(HF) diets for 12 wk. The body weight was assessed once weekly. After 12 wk, the liver and serum samples were extracted. The liver tissue expression levels of RIP3, microsomal triglyceride transfer protein, protein disulfide isomerase, apolipoprotein-B, X-box binding protein-1, sterol regulatory element-binding protein-1c, fatty acid synthase, cluster of differentiation-36, diglyceride acyltransferase, peroxisome proliferator-activated receptor alpha, tumor necrosis factor-alpha(TNF-α), and interleukin-6 were assessed. Oleic acid treated primary hepatocytes from WT and RIP3 KO mice were stained with Nile red. The expression of inflammatory cytokines, including chemokine(C-X-C motif) ligand(CXCL) 1, CXCL2, and TNF-α, in monocytes was evaluated.RESULTS RIP3 KO HF diet fed mice showed a significant gain in body weight, and liver weight, liver to body weight ratio, and liver triglycerides were increased in HF diet fed RIP3 KO mice compared to HF diet fed WT mice. RIP3 KO primary hepatocytes also had increased intracellular fat droplets compared to WT primary hepatocytes after oleic acid treatment. RIP3 overexpression decreased hepatic fat content. Quantitative real-time polymerase chain reaction analysis showed that the expression of very-low-density lipoproteins secretion markers(microsomal triglyceride transfer protein, protein disulfide isomerase, and apolipoprotein-B) was significantly suppressed in RIP3 KO mice. The overall NAFLD Activity Score was the same between WT and RIP3 KO mice; however, RIP3 KO mice had increased fatty change and decreased lobular inflammation compared to WT mice. Inflammatory signals(CXCL1/2, TNF-α, and interleukin-6) increased after lipopolysaccharide and pancaspase inhibitor(necroptotic condition) treatment in monocytes. Neutrophil chemokines(CXCL1, and CXCL2) were decreased, and TNF-α was increased after RIP3 inhibitor treatment in monocytes.CONCLUSION RIP3 deletion exacerbates steatosis, and partially inhibits inflammation in the HF diet induced NAFLD model.

Preclinical Long-term Magnetic Resonance Imaging Study of Silymarin Liver-protective Effects

Background and Aims:Efficacy evaluations with preclini-cal magnetic resonance imaging(MRI)are uncommon,but MRI in the preclinical phase of drug development provides information that is useful for longitudinal monitoring.The study aim was to monitor the protective effectiveness of silymarin with multiparameter MRI and biomarkers in a thioacetamide(TAA)-induced model of liver injury in rats.Correlation analysis was conducted to assess compare the monitoring of liver function by MRI and biomarkers.Meth-ods:TAA was injected three times a week for 8 weeks to generate a disease model(TAA group).In the TAA and sily-marin-treated(TAA-SY)groups,silymarin was administered three times weekly from week 4.MR images were acquired at 0,2,4,6,and 8 weeks in the control,TAA,and TAA-SY groups.Results:The area under the curve to maximum time(AUCtmax)and T_(2)*values of the TAA group decreased over the study period,but the serological markers of liver abnormality increased significantly more than those in the control group.In the TAA-SY group,MRI and serological biomarkers indicated attenuation of liver function as in the TAA group.However,pattern changes were observed from week 6 to comparable levels in the control group with si-lymarin treatment.Negative correlations between either AUCtmax or T_(2)*values and the serological biomarkers were observed.Conclusions:Silymarin had hepatoprotective effects on TAA-induced liver injury and demonstrated the usefulness of multiparametric MRI to evaluate efficacy in preclinical studies of liver drug development.