Methyl helicterilate ameliorates alcohol-induced hepatic fibrosis by modulating TGF-β1/Smads pathway and mitochondria-dependent pathway
Authors: Shimei Tan
Xing Lin
Jinlan Nie
Facheng Bai
Yuanyuan Wei
Xiaolin Zhang
Jinbin Wei
Shujuan Wen
Journal: International Immunopharmacology (2019)
Discipline: PHARMACOLOGY & TOXICOLOGY
Keywords: Alcohol; Helicteres angustifolia; Hepatic fibrosis; Hepatic stellate cells; Methyl helicterilate
Affiliations: Guangxi Medical University, Nanning 530021, China., Guangxi Medical University, Nanning 530021, China. Electronic address: .

Abstract:  This study aimed to investigate the effect and underlying mechanism of Methyl helicterilate from Helicteres angustifolia (MHHA) on alcohol-induced hepatic fibrosis. The results showed that MHHA treatment markedly alleviated alcohol-induced liver injury and notably reduced collagen deposition in liver tissue. It significantly enhanced the activity of alcohol dehydrogenase and aldehyde dehydrogenase. Moreover, MHHA treatment markedly decreased the content of inflammatory cytokines, alleviated collagen accumulation, and inhibited the expression of TGF-β1 and Smad2/3 in liver tissue. The experiments in cells showed that MHHA significantly inhibited HSC activation by blocking TGF-β1/Smads signaling pathway. Additionally, it notably induced HSC apoptosis by modulating the mitochondria-dependent pathway. The present study demonstrates that MHHA treatment significantly ameliorates alcoholic hepatic fibrosis and the underlying mechanism may be ascribed to the inhibition of the TGF-β1/Smads pathway and regulation of the mitochondria-mediated apoptotic pathway. 

Retracted: No

PubPeer comments:
 Fig 3A1, the normal control image looks like having appeared in three other papers: Huang, et al (2019) Fig 1B1 and Huang, et al (2017) Fig 2A also as  normal control Lin, et al (2016) Fig 1A2 (when flipped upside-down) as a didymin control, supposedly different from the normal control. Each (re-)appearance of likely the same image involves cropping and resizing, and in the Lin, et al (2016) paper, a vertical flip was additionally applied. ![file] , 
 Fig 3A2, the MHHA control image looks like having appeared in three other papers: Huang, et al (2017) Fig 2A as  OVD control Lin, et al (2016) Fig 1A1 as the normal control. Huang, et al (2016) Fig 5A as the normal control Each (re-)appearance of likely the same image involves cropping and resizing ![file] , 
 Shared by another reader. Several images in Figure 4B appear to have common features, sometimes flipped or rotated. Iu2019m not sure the features in the cyan boxes are a match, but... *** ![file] ![file] ![file] With caption ![file] , 
 #3 Fig. 4A.  MHHA reduced alcohol-induced collagen production. (A) ... Collagen deposition was assessed using Masson s trichrome .... A1 or B1: the normal control group; A2 or B2: the MHHA control group; A3 or B3: the alcohol model group; A4-A6 or B4-B5: the MHHA-treated groups (**16.72**, 33.45 and *66.90 mg kg*). ![file] Fig 1B2 from [Bai et al (2018)] .  GEI significantly alleviated liver injury and collagen accumulation induced by PS in rats (n = 10). ... (B) Hepatic collagen was observed by Masson s trichrome staining (200u00d7); u201cB1u201d to u201cB6u201d represent the normal control, **model**, *PDTC*, low-, medium-, and high-dose of GEI groups, respectively ![] , 
 #2 ![] There is a modified version in [Bai et al (2017)] . ![] , 
 Fig 3A3. An area is repeated three times (with different rotations). ![file] , 
 Fig 3A4, color-reversed and superimposed upon 3A3, to dramatise the large areas of the images that are the same (and cancel out). ![] , 
 Fig. 3.  MHHA ameliorated alcohol-induced liver injury. (A) Histological changes were observed using H E staining (200u00d7); A1: normal control, A2: MHHA control, **A3: alcohol-model group**, A4-A6: MHHA-treated groups (**16.72, 33.45** and 66.90 mg kg) . ![] Fig. 1 from [Huang et al (2019)] .  MH ameliorated PS-induced liver injury. (A) The appearance of the liver samples. A1 to A6 represented the normal control group, MH control group, **PS model group, 16.72, 33.45** and 66.90 mg kg MH-treated groups, respectively . ![file] Three pairs of liver images are the same. Was the fibrosis (untreated, or MH-treated) the result of porcine serum or alcohol? , 
 Fig 3A3 and 3A4 again: ![] For comparison with Fig 11 from [Wei et al (2013)] : ![] , 
 #4 Fig 4A again. ![file] Fig 1 from [Lin et al (2016)] ![file] Red boxes enclose A3 ( alcohol model group ) and B5 ( CCl4 + didymin + locostatin (CDL) group ) respectively. , 
 Fic 7C.  MHHA inhibited HSC proliferation and activation. ... (C) Protein expression of u03b1-SMA was determined with Western blot assay. Band 1u20136 represented the normal group, TGF-u03b21 group, LY364947 group, and MHHA treatment groups (3, 6 and 12 u03bcmol L). ![file] For comparison: Fig 4 from [Wei et al (2019)] .  The expression of Colu2010I ... was detected by immunohistochemical staining (u00d7400) and Western blot analysis. u201c1u201d to u201c6u201d represent the normal control group, 20 ng mL PDGFu2010BB group, 100 u03bcM PD98059 group, 3.75, 7.5 or 15 u03bcM MHu2010treated groups, respectively . Below: Fig 7F from [Huang et al (2019)] .  MH significantly inhibited the TGF-u03b21 Smads and ERK pathways after miR-21 overexpression in HSC-T6 cells....(F) ...the phosphorylation of p-Smad2 3 and p-ERK were detected by Western blotting; band 1 to 6 represented the normal control group, lentivirus-NC group, lentivirus-up group, 3, 6 and 12 u03bcmol l MH-treated groups, respectively . ![] , 
 Fig 8B:  MHHA significantly induced HSC apoptosis. (A-B) Cell apoptosis was assessed by AO EB staining (200u00d7) and flow cytometry, respectively; the arrow indicates hepatocyte apoptosis. A1 to A6 (or B1 to B6) represented the normal group, TGF-u03b21 group, LY364947 group, and MHHA treatment groups (3, 6 and 12 u03bcmol L), respectively. ![file] For comparison, Fig 6 from [Huang et al (2019)] : ![file] , 
 Fig 11A B.  MHHA blocked the TGF-u03b21 Smads pathway. (A) and (B) The expression of TGF-u03b21 and Smad2 3 was observed by Immunohistochemical staining. The expression of TGF-u03b21 and Smad2 3 was marked with brown staining as the arrow pointed. The expression of TGF-u03b21was mainly found in the ECM, while Smad2 3 was observed both in cytoplasm and nucleus. A1u2013A6 (or B1u2013B6) represented the normal group, TGF-u03b21 group, LY364947 group, and MHHA treatment groups (3, 6 and 12 u03bcmol L), respectively. ![file] I have concerns. A3 (including rotated version for clarity). ![file] B2: ![file] B6: ![file] , 
 Corrigendum [March 2021] . ----------------------------------- The authors regret the incorrect publication of Figs. 3A, 4, 7, 8B and 11 in the original article. The correct figures are presented below: ![] Fig. 3. MHHA ameliorated alcohol-induced liver injury. (A) Histological changes were observed using H E staining (200u00d7); A1: normal control, A2: MHHA control, A3: alcohol-model group, A4-A6: MHHA-treated groups (16.72, 33.45 and 66.90 mg kg). #P 0.05 VS. normal control group and *P 0.05 VS. model group. ![] Fig. 4. MHHA reduced alcohol-induced collagen production. (A) and (B) Collagen deposition was assessed using Masson s trichrome and Sirius red staining (200u00d7); The arrow means collagen deposition. A1 or B1: the normal control group; A2 or B2: the MHHA control group; A3 or B3: the alcohol model group; A4-A6 or B4-B5: the MHHA-treated groups (16.72, 33.45 and 66.90 mg kg). #P 0.05 VS. normal control group and *P 0.05 VS. model group. ![] Fig. 7. MHHA inhibited HSC proliferation and activation. (C) Protein expression of u03b1-SMA was determined with Western blot assay. Band 1u20136 represented the normal group, TGF-u03b21 group, LY364947 group, and MHHA treatment groups (3, 6 and 12 u03bcmol L). #P 0.05 VS. normal control cells and *P 0.05 VS. TGF-u03b21 group. ![] Fig. 8. MHHA significantly induced HSC apoptosis. (A-B) Cell apoptosis was assessed by AO EB staining (200u00d7) and flow cytometry, respectively; the arrow indicates hepatocyte apoptosis. A1 to A6 (or B1 to B6) represented the normal group, TGF-u03b21 group, LY364947 group, and MHHA treatment groups (3, 6 and 12 u03bcmol L), respectively. #P 0.05 VS. normal control group and *P 0.05 VS. model group. ![] Fig. 11. MHHA blocked the TGF-u03b21 Smads pathway. (A) and (B) The expression of TGF-u03b21 and Smad2 3 was observed by Immunohistochemical staining. The expression of TGF-u03b21 and Smad2 3 was marked with brown staining as the arrow pointed. The expression of TGF-u03b21was mainly found in the ECM, while Smad2 3 was observed both in cytoplasm and nucleus. A1u2013A6 (or B1u2013B6) represented the normal group, TGF-u03b21 group, LY364947 group, and MHHA treatment groups (3, 6 and 12 u03bcmol L), respectively. #P 0.05 VS. normal control group and *P 0.05 VS. model group. (C) mRNA levels of TGF-u03b21, Tu03b2RI, Tu03b2RII, Smad2 and Smad3 in liver tissues were analyzed by RT-PCR. (D) TGF-u03b21and Tu03b2RI II protein content and Smad2 3 phosphorylation in liver tissues were measured using Western blotting. (E) Protein expression of TGF-u03b21, Tu03b2RI and Tu03b2RII and the phosphorylation of Smad2 3 in LX-2 cells were detected by Western blotting. (F) The Smad transcriptional activity was measured using a Dual-Glou00ae Luciferase Assay System. The authors would like to apologise for any inconvenience caused. , 
 #14 I agree with the authors that it was unfortunate that the original Figures 3, 4, 7, 8 and 11 were published. I feel, though, that the Corrigendum note could be longer. It appears that the authors had prepared one Figs 3 4 7 8 11 set which is genuine and features in the correction; and a second set with the appearance of collages or stamp collections, overlapping with images used in other papers; and they used the second set by mistake. I would welcome a fuller explanation of how that second set came to exist. It would also be useful to learn more of the journal editors reasoning, when they decided to accept the replacement. They are not alone in this approach to regrettable illustrations. I compiled the following list of regret, from a range of journals but involving an overlapping roster of authors: -------------------------------------------------------  The authors regret that the wrong Figures 7 and 8 were published . [Hepatoprotective effects of total saponins isolated from Taraphochlamys affinis against carbon tetrachloride induced liver injury in rats] . QuanFang Huang, ShiJun Zhang, Li Zheng, Min He, RenBin Huang, Xing Lin, *Food and chemical toxicology* (2012). -------------------------------------------  The authors regret the incorrect publication of Figs. 3B, 3D, 4, 5B and 5C in the original article . [Antiviral activity of methyl helicterate isolated from Helicteres angustifolia (Sterculiaceae) against hepatitis B virus] . Quanfang Huang , Renbin Huang , Ling Wei , Yongxing Chen , Shujuan Lv , Chunhong Liang , Xuerong Zhang , Fujiang Yin , Hongtao Li , Lang Zhuo , Xing Lin, *Antiviral Research* (2013). ----------------------------------------------  The authors regret that the wrong Fig. 4, Fig. 6, Fig. 7 were published. [Protective effect of madecassoside against cognitive impairment induced by D-galactose in mice] . Xing Lin , Shijun Zhang , Renbin Huang , Ling Wei , Shimei Tan , Chunhong Liang , Shujuan Lv , Yongxin Chen , Shuang Liang , Yuanchun Tian , Zhongpeng Lu , Quanfang Huang, *Pharmacology Biochemistry and Behavior* (2014). --------------------------------------------------  The authors regret for the incorrect publication of Fig. 2, Fig. 5A in the original article. [Protective effect of tormentic acid from Potentilla chinensis against lipopolysaccharide D-galactosamine induced fulminant hepatic failure in mice] . Xing Lin , Shijun Zhang , Renbin Huang , Shimei Tan , Shuang Liang , Xiaoyan Wu , Lang Zhuo , Quanfang Huang, *International Immunopharmacology* (2014). ------------------------------------------------------  The authors regret that the wrong Figs. 3, 6B, 8 and 9 were published. [Didymin induces apoptosis through mitochondrial dysfunction and up-regulation of RKIP in human hepatoma cells] . Jinbin Wei , Quanfang Huang , Facheng Bai , Jun Lin , Jinlan Nie , Shengjuan Lu , Chunyuang Lu , Renbin Huang , Zhongpeng Lu , Xing Lin, *Chemico-Biological Interactions* (2017). -----------------------------------------------  The authors regret the incorrect publication of Figs. 3A, 4, 7, 8B and 11 in the original article . [Methyl helicterilate ameliorates alcohol-induced hepatic fibrosis by modulating TGF-u03b21 Smads pathway and mitochondria-dependent pathway] . Shujuan Wen , Yuanyuan Wei , Xiaolin Zhang , Facheng Bai , Shimei Tan , Jinlan Nie , Jinbin Wei, Xing Lin, *International Immunopharmacology* (2019). ,