Trolline Ameliorates Liver Fibrosis by Inhibiting the NF-κB Pathway, Promoting HSC Apoptosis and Suppressing Autophagy

Facheng Bai, Quanfang Huang, Jinlan Nie, Shengjuan Lu, Chunyuan Lu, Xunshuai Zhu, Yuxin Wang, Lang Zhuo, Zhongpeng Lu, Xing Lin
2017 Cellular Physiology and Biochemistry  
Background/Aims: Previous studies have shown that trolline possesses various forms of pharmacological activity, including antibacterial and antiviral potency. The present paper addressed the putative hepatoprotective effects of trolline. Methods: Rats received 2 ml/kg CCl 4 (mixed 1:1 in peanut oil) intragastrically twice a week for 8 weeks to induce hepatic fibrosis. The animals were then treated with trolline for additional 4 weeks. Liver pathology and collagen accumulation were observed by
more » ... were observed by hematoxylin-eosin and Masson's trichrome staining, respectively. Serum transaminase activity and collagen-related indicator level were determined by commercially available kits. NF-κB pathway activation was also examined. Moreover, the effects of trolline on hepatic stellate cell (HSC-T6) apoptosis, mitochondrial membrane potential (MMP), and autophagy were assessed. Results: Trolline significantly alleviated CCl 4 -induced liver injury and notably reduced the accumulation of collagen in liver tissues. Trolline treatment also markedly decreased inflammatory cytokines levels by inhibiting the NF-κB pathway. Trolline strongly inhibited HSC-T6 activation and notably induced cell apoptosis by modulating the Bax/Bcl-2 ratio, caspase activity, and MMP. Moreover, trolline significantly inhibited HSC-T6 autophagy, as evidenced by the decrease in the formation of autophagic vacuoles and the number of autophagosomes, by regulating the expression levles of LC3, Beclin-1, P62, Atg 5 and 7. Conclusion: Our study demonstrates that trolline ameliorates liver fibrosis, possibly by inhibiting the NF-κB pathway, promoting HSCs apoptosis and suppressing autophagy. Trolline inhibited inflammatory cytokines and NF-κB pathway The productions of TNF-α and IL-1β were significantly higher in the CCl 4 model group than those in the normal control group, while the abnormal levels of both inflammatory Fig. 3. Trolline significantly reduced inflammatory cytokine content and inhibited NF-κB activation in rats. (A) The inflammatory cytokines TNF-α and IL-1β were detected by ELISA kits. (B) Phosphorylation of IκBα and p65 was detected by Western blotting; the bands 1 to 4 represent the normal control, trolline control, CCl 4 model and trolline-treated groups, respectively. (C) The level of NF-κB mRNA was detected by RT-PCR assay. Data were expressed as the means ± SD, * P<0.05 vs. the normal control group and # P<0.05 vs. the CCl 4 model group. Fig. 4. Trolline significantly inhibited HSC activation and induced HSC apoptosis. (A) Cytotoxicity test was assessed by MTT assay. (B) The protein expression of α-SMA and collagen I in HSCs was detected using Western blotting; bands 1 to 4 represent the normal tontrol, trolline control, PDGF-BB, and trolline + PDGF-BB groups, respectively. (C) Cell apoptosis was assessed by flow cytometry. (D) The ratio of apoptosis was calculated. (E) The expression of Bax and Bcl-2 was detected by Western blotting; the bands 1 to 4 represent the normal control, 2.5, 5 and 10 μM trolline groups, respectively. (F) Caspase-3 and -8 were detected using commercially available kits. Data were expressed as the means ± SD, * P<0.05 vs. the normal control group, # P<0.05 vs. the PDGF-BB group.
doi:10.1159/000485009 pmid:29141243 fatcat:exxxzoidvfchbcfdkafbf6xvpy