Synergistic effect of a novel oxymatrine-baicalin combination against hepatitis B virus replication, a smooth muscle actin expression and typeⅠcollagen synthesis in vitro

AIM： To study the effect of oxymatrine-baicalin combination （OB） against HBV replication in 2.2.15 cells and α smooth muscle actin （ α SMA） expression, type I, collagen synthesis in HSC-T6 cells. METHODS： The 2.2.15 cells and HSC-T6 cells were cultured and treated respectively. HBsAg and HBeAg in the culture supernatants were detected by ELISA and HBV DNA levels were determined by fluorescence quantitative PCR. Total RNA was extracted from HSC-T6 cells and reverse transcribed into cDNA. The cDNAs were amplified by PCR and the quantities were expressed in proportion to β actin. The total cellular proteins extracted from HSC-T6 cells were separated by electrophoresis. Resolved proteins were electrophoretically transferred to nitrocellulose membrane. Protein bands were revealed and the quantities were corrected by β actin. RESULTS： In the 2.2.15 cell culture system, the inhibitory rate against secretion of HBsAg and HBeAg in the OB group was significantly stronger than that in the oxymatrine group （HBsAg, P = 0.043; HBeAg, P = 0.026; respectively）; HBV DNA level in the OB group was significantly lower than that in the oxymatrine group （P = 0.041）. In HSC-T6 cells the mRNA and protein expression levels of α SMA in the OB group were significantly lower as compared with those in the oxymatrine group （mRNA, P = 0.013; protein, P = 0.042; respectively）; The mRNA and protein expression levels of type I collagen in the OB group were significantly lower as compared with those in the oxymatrine group （mRNA, P 〈 0.01; protein, P 〈 0.01; respectively）.CONCLUSION： OB combination has a better effect against HBV replication in 2.2.15 cells and is more effective against α SMA expression and type I collagen synthesis in HSC-T6 cells than oxymatrine in vitro.

Expression analysis of a-smooth muscle actin and tenascin-C in the periodontal ligament under orthodontic loading or in vitro culture

α-smooth muscle actin （α-SMA） and tenascin-C are stress-induced phenotypic features of myofibroblasts. The expression levels of these two proteins closely correlate with the extracellular mechanical microenvironment. We investigated how the expression of α-SMA and tenascin-C was altered in the periodontal ligament （PDL） under orthodontic loading to indirectly reveal the intrinsic mechanical microenvironment in the PDL. In this study, we demonstrated the synergistic effects of transforming growth factor-β1 （TGF-β1） and mechanical tensile or compressive stress on myofibroblast differentiation from human periodontal ligament cells （hPDLCs）. The hPDLCs under higher tensile or compressive stress significantly increased their levels of α-SMA and tenascin-C compared with those under lower tensile or compressive stress. A similar trend was observed in the tension and compression areas of the PDL under continuous light or heavy orthodontic load in rats. During the time-course analysis of expression, we observed that an increase in α-SMA levels was matched by an increase in tenascin-C levels in the PDL under orthodontic load in vivo. The time-dependent variation of α-SMA and tenascin-C expression in the PDL may indicate the time-dependent variation of intrinsic stress under constant extrinsic loading.

Expression of alpha smooth muscle actin in living donor liver transplant recipients

Recently, there have been reports from liver biopsies that showed the progression of liver fibrosis in liver transplant patients after the cessation of immunosuppression. Herein, we focused on activated hepatic stellate cells expressing alpha smooth muscle actin (α-SMA) to understand the correlation between immunosuppressant medication and liver fibrosis. The study enrolled two pediatric patients who underwent living donor liver transplantation and ceased immunosuppressant therapy. The number of α-SMA-positive cells in the specimens obtained by liver biopsy from these two patients showed a three-fold increase compared with the number from four transplanted pediatric patients who were continuing immunosuppressant therapy. In addition, the α-SMA-positive area evaluated using the WinRooF image processing software program continued to increase over time in three adult transplanted patients with liver fibrosis, and the α-SMA-positive area was increasing even during the pre-fibrotic stage in these adult cases, according to a retrospective review. Therefore, α-SMA could be a useful marker for the detection of early stage fibrosis.

Suppression of hepatic steatosis in non-alcoholic steatohepatitis model by modified Xiaoyao San formula:Evidence,mechanisms and perspective

In this letter,we comment on a recent publication by Mei et al,in the World Journal of Hepatology,investigating the hepatoprotective effects of the modified Xiaoyao San(MXS)formula in a male rat model of non-alcoholic steatohepatitis(NASH).The authors found that MXS treatment mitigated hepatic steatosis and inflam-mation in the NASH model,as evidenced by the reduction in lipid droplets(LDs),fibrosis markers and lipogenic factors.Interestingly,these hepatoprotective effects were associated with androgen upregulation(based on metabolomics analysis of male steroid hormone metabolites),adenosine 5’-monophosphate-activated protein kinase(AMPK)activation,and restoration of phosphatase and tensin homolog(PTEN)expression.However,the authors did not clearly discuss the relationships between MXS-induced hepatic steatosis reduction in the NASH model,and androgen upregulation,AMPK activation,and restoration of PTEN expression.This editorial emphasizes the reported mechanisms and explains how they act or interact with each other to reduce hepatic steatosis and inflammation in the NASH model.As a perspective,we propose additional mechanisms(such as autophagy/lipophagy activation in hepatocytes)for the clearance of LDs and suppression of hepatic steatosis by MXS in the NASH model.A proper understanding of the mechanisms of MXS-induced reduction of hepatic steatosis might help in the treatment of NASH and related diseases.

Multinucleated giant cells of bladder mucosa are modified telocytes:Diagnostic and immunohistochemistry algorithm and relation to PDL1 expression score

BACKGROUND Multinucleated giant cells(MGCs)in bladder carcinomas are poorly studied.AIM To describe the function,morphogenesis,and origin of mononuclear and MGCs in urothelial carcinoma(UC)of the bladder in Bulgarian and French patients.METHODS Urothelial bladder carcinomas(n=104)from 2016-2020 were analyzed retrospectively using immunohistochemical(IHC)and histochemical stain examination.Giant cells in the bladder stroma were found in 35.6%of cases,more often in highgrades.RESULTS We confirm that MGCs in the mucosa in UC of the bladder were positive for both mesenchymal and myofibroblast markers(vimentin,smooth muscle actin,Desmin,and CD34)and the macrophage marker CD68.Furthermore,IHC studies revealed the following profile of these cells:Positive for p16;negative for epithelial(CK AE1/AE3 and GATA-3),vascular(CD31),neural(PS100 and CKIT),cambial,blastic(CD34-blasts and C-KIT),and immune markers(IG G,immunoglobulin G4,and PD-L1);no proliferative activity,possess no specific immune function,and cannot be used to calculate the Combined Positive Score scale.CONCLUSION In conclusion,the giant stromal cells in non-tumor and tumor bladder can be used as a characteristic and relatively constant,although nonspecific,histological marker for chronic bladder damage,reflecting the chronic irritation or inflammation.Likewise,according to the morphological and IHC of the mono-and multinucleated giant cells in the bladder,they are most likely represent telocytes capable of adapting their morphology to the pathology of the organ.

Emodin protects rat liver from CCl_4-induced fibrogenesis via inhibition of hepatic stellate cells activation

AIM： To investigate the role of emodin in protecting the liver against fibrogenesis caused by carbon tetrachloride （CCh） in rats and to further explore the underlying mechanisms. METHODS： Rat models of experimental hepatic fibrosis were established by injection with CCh; the treated rats received emodin via oral administration at a dosage of 20 mg/kg twice a week at the same time. Rats injected with olive oil served as a normal group. Histopathological changes were observed by hematoxylin and eosin staining. The activities of alanine aminotransferase （ALT） and aspartate aminotransferase （AST） in serum and hepatic hydroxyproline content were assayed by biochemical analyses. The mRNA and protein relevant to hepatic stellate cell （HSC） activation in the liver were assessed using real-time reverse transcription-polymerase chain reaction （RT-PCR）, immunohistochernistry, western blotting and enzymelinked immunosorbent assay.RESULTS： The degree of hepatic fibrosis increased markedly in the CCh group compared to the normal group （P 〈 0.01）, and decreased markedly in the emodin group compared to the CCI4 group according to METAVIR scale （P 〈 0.01） compared with those in the normal control group （51.02 ± 10.64 IU/L and 132.28 ± 18.14 IU/L）. The activities of serum ALT and AST were significantly higher in rats injected with CCh （289.25 ± 68.84 IU/L and 423.89 ± 35.67 IU/L, both P 〈 0.05）. The activities of serum ALT and AST were significantly reduced by administration of emodin （176.34 ± 47.29 IU/L and 226.1 ± 44.52 IU/L, both P 〈 0.05）. Compared with the normal controls （54.53 ± 13.46 mg/g）, hepatic hydroxyproline content was significantly higher in rats injected with CCI4 （120.27 ± 28.47 mg/g, P 〈 0.05）. Hepatic hydroxyproline content was significantly reduced in the rats treated with emodin at 20 mg/kg （71.25 ± 17.02 mg/g, P 〈 0.05）. Emodin significantly protected the liver from injury by reducing serum AST and ALT activities and reducing hepatic hydroxyproline content. The mRNA levels of transforming growth factor-β1 （TGF-β1）, Smad4 and α-SMA in liver tissues were significantly down-regulated in SD rats that received emodin treatment. Furthermore, significant down-regulation of serum TGF-β1 protein levels and protein expression of Smad4 and α-SMA in liver tissues was also observed in the rats. Emodin inhibited HSC activation by reducing the abundance of TGF-β1 and Smad4. CONCLUSION： Emodin protects the rat liver from CCI4-induced fibrogenesis by inhibiting HSC activation. Emodin might be a therapeutic antifibrotic agent for the treatment of hepatic fibrosis.

Effects of interleukin-10 on activation and apoptosis of hepatic stellate cells in fibrotic rat liver

AIM： TO study the effects of interleukin-10 （IL-10） on the expression of o-smooth muscle actin （α-SMA）, nuclear factor-κB（NF-κB） and Fas/Fas ligand （FasL） in hepatic stellate cells of experimental rats with hepatic fibrosis. METHODS： Sixty clean SD rats were randomly divided into control group （group N）, liver fibrotic group （group C） and IL-10 treatment group （group I）. Control group received intraperitoneal injection of saline （2ml·kg^-1）, twice a week. Fibrotic group was injected intraperitoneally with 50% carbon tetrachloride （CCh） （2 ml·kg^-1）, twice a week. IL-10 treatment group was given IL-10 at a dose of 4 pg·kg^-1 20 minutes before CCl4 administration from the third week. Hepatic stellate cells （HSCs） were isolated from these rats at the seventh and eleventh weeks during the course of liver fibrosis, respectively. The expression of α-SMA and NF-κB in HSCs was measured by S-P immunohistochemistry. The expression of Fas and FasL mRNA was measured by RT-PCR. Furthermore, liver tissues were harvested from three groups at the same time. RESULTS： The CCh- induced experimental rat hepatic fibrosis model was established successfully. The purity of extracted hepatic stellate cells was about 95% and the yield of hepatic stellate cells was 1.2-2.3×10^6/g liver tissue averagely. The positive expression of α-SMA and NF-κB was 36.5% and 28.5% respectively in group N. The positive levels of α-SMA and NF-κB were increased significantly in group C compared to group N （P〈0.01）. The positive signals decreased significantly （P〈0.05） in group I. In the 11^th week, the HSCs of group I became round with visible pyknotic nuclei. The expression of NF-κB in group C was significantly increased in a timedependentmanner （P〈0.01）, but there was no difference in the α-SMA expression （P〉0.05）. The mRNA of Fas and FasL in group C was significantly increased in a timedependent manner compared to that in control group. After treated with IL-10, the expression level of Fas and FasL was higher in group I than in group C. CONCLUSION： The positive expression of α-SMA and NF-κB in hepatic stellate cells is decreased by ectogenic IL-10 in liver fibrosis induced by CCh. The expression of Fas and FasL is increased in the course of liver fibrosis, and is further increased by IL-10. IL-10 could inhibit the activation of HSCs and cause apoptosis of activated HSCs.

Insulin-like growth factor binding protein-7 induces activation and transdifferentiation of hepatic stellate cells in vitro

AIM:To investigate the role of insulin-like growth factor binding protein-7 (IGFBP-7) in the activation and transdifferentiation of hepatic stellate cells (HSC) in vitro.METHODS:Rat HSC-T6 cells were cultured in separate dishes and treated with various concentration of transforming growth factor (TGF)-β1,IGFBP-7 or antiIGFBP-7 antibody for 24 h.The supernatant or a cytoplasm suspension was obtained from cultured HSC,followed by transfer of cells to form cell-coated dishes.Immunocytochemistry and Western blotting were used to analyze the expression of IGFBP-7 induced by TGF-β1 and the level of fibronectin,collagen and α-smooth muscle actin (SMA).The pro-apoptotic effect of antiIGFBP-7 antibody was determined by flow cytometry.RESULTS:Immunocytochemistry and Western blotting revealed that the expression of IGFBP-7 in TGF-β1 treated HSC was significantly up-regulated compared to that in the control group.In addition,fibronectin,collagen and α-SMA also showed enhanced expression in accordance with the transdifferentiation process in a dose-dependent manner to some extent.Moreover,flow cytometry suggested that anti-IGFBP-7 antibody induced apoptosis of activated HSC,which is responsible for the development of liver fibrosis,and may represent a novel pathway and target for therapeutic intervention.CONCLUSION:IGFBP-7 showed increased expression in activated HSC and played an important role in the activation and transdifferentiation process of HSC.AntiIGFBP-7 antibody may ameliorate liver fibrogenesis.

I_κB kinase-beta inhibitor attenuates hepatic fibrosis in mice

AIM: To investigate the anti-fibrosis effect of IκB kinase-beta inhibitor (IKK2 inhibitor IMD0354) in liver fibrosis. METHODS: Twenty male C57BL6 mice were divided into four groups. Five high-fat fed mice were injected with lipopolysaccharide (LPS, 10 mg/kg) intraperitoneally and five high-fat fed mice were without LPS injection to build models of liver injury, and the intervention group (five mice) was injected intraperitoneally with IKK2 inhibitor (IMD 30 mg/kg for 14 d), while the remaining five mice received a normal diet as controls. Hepatic function, pathological evaluation and liver interleukin-6 (IL-6) expression were examined. Western blotting and real-time polymerase chain reaction were used to detect the expressions of nuclear factor-κB (NF-κB), alpha-smooth muscle actin (α-SMA), tumor growth factor-beta1 (TGF-β1), tumor necrosis factor-alpha (TNF-α), typeⅠand type Ⅲ collagen proteins and mRNA. RESULTS: A mouse model of liver injury was successfully established, and IMD decreased nuclear transloca-tion of NF-κB p65 in liver cells. In the IMD-treated group, the levels of alanine aminotransferase (103 ± 9.77 μ/L vs 62.4 ± 7.90 μ/L, P < 0.05) and aminotransferase (295.8 ± 38.56 μ/L vs 212 ± 25.10 μ/L, P < 0.05) were significantly decreased when compared with the model groups. The histological changes were significantly ameliorated. After treatment, the expressions of IL-6 (681 ± 45.96 vs 77 ± 7.79, P < 0.05), TGF-β1 (Western blotting 5.65% ± 0.017% vs 2.73% ± 0.005%, P < 0.05), TNF-α (11.58% ± 0.0063% vs 8.86% ± 0.0050%, P < 0.05), typeⅠcollagen (4.49% ± 0.014% vs 1.90% ± 0.0006%, P < 0.05) and type Ⅲ collagen (3.46% ± 0.008% vs 2.29% ± 0.0035%, P < 0.05) as well as α-SMA (6.19 ± 0.0036 μ/L vs 2.16 ± 0.0023 μ/L, P < 0.05) protein and mRNA were downregulated in the IMD group compared to the fibrosis control groups (P < 0.05). CONCLUSION: IKK2 inhibitor IMD markedly improved non-alcoholic fatty liver disease in mice by lowering NF-κB activation, which could become a remedial target for liver fibrosis.

Effects of pharmacological serum from normal and liver fibrotic rats on HSCs

AIM: To make drug sera of Salvia miltiorrhiza and Yigankang, both of which are Chinese herbs that activate bleeding and eliminate stasis, in normal rats and those with liver fibrosis, respectively. To investigate and compare the effects of the two different drug sera on the proliferation and activation of hepatic stellate cells (HSCs). METHODS: Some rats were induced with liver fibrosis: 40% carbon tetrachloride (CCI4) subcutaneous injection, twice a week for 9 wk. Salvia miltiorrhiza, Yigankang, colchicines and normal saline were administered into the stomachs of normal rats and those with liver fibrosis. Drug sera were extracted 5 d later. HSCs in vitro were cultivated in different drug sera for 24 h. The rates of proliferation and expression of a-smooth muscle actin (α-SMA) were detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and immunocyt-ochemistry stain, respectively. RESULTS: The drug sera from normal and liver fibrotic rats could be used to cultivate HSCs and to observe the effects of the corresponding components of herbs on HSCs. Salvia miltiorrhiza and Yigankang had better inhibitory effects on HSCs than colchicines (MTT: normal drug serum: Salvia miltiorrhiza 0.42 ±0.08, Yigankang 0.32±0.10 vs colchicines 0.45±0.12 pathological drug serum: Salvia miltiorrhiza 0.33±0.02, Yigankang 0.26±0.01 vs colchicines 0.41±0.09. P<0.05). The drug sera of Salvia miltiorrhiza, Yigankang from liver fibrotic rats had a stronger inhibitory effect than the same ones from normal rats (MTT: Salvia miltiorrhiza: normal drug serum 0.42±0.08 vs pathological drug serum 0.33±0.02. Yigankang: normal drug serum 0.32±0.10 vs pathological drug serum 0.26±0.01. P<0.05) CONCLUSION: Salvia miltiorrhiza and Yigankang could inhibit the expression of a-SMA and the proliferation of HSCs. The drug sera from normal and liver fibrotic rats had different effects on HSCs, probably due to different metabolic processes, effective components and different quantities of drug contents in drug sera from rats with different states of liver.

The ROCK pathway inhibitor Y-27632 mitigates hypoxia and oxidative stress-induced injury to retinal Müller cells

Rho kinase （ROCK） was the first downstream Rho effector found to mediate RhoA-induced actin cytoskeletal changes through effects on myosin light chain phosphorylation. There is abundant evidence that the ROCK pathway participates in the pathogenesis of retinal endothelial injury and proliferative epiretinal membrane traction. In this study, we investigated the effect of the ROCK pathway inhibitor Y-27632 on retinal Müller cells subjected to hypoxia or oxidative stress. Müller cells were subjected to hypoxia or oxidative stress by exposure to CoCl2 or H2O2. After a 24-hour treatment with Y-27632, the 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyl tetrazolium bromide assay was used to assess the survival of Müller cells. Hoechst 33258 was used to detect apoptosis, while 2′,7′-dichlorodihydrofluorescein diacetate was used to measure reactive oxygen species generation. A transwell chamber system was used to examine the migration ability of Müller cells. Western blot assay was used to detect the expression levels of α-smooth muscle actin, glutamine synthetase and vimentin. After treatment with Y-27632, Müller cells subjected to hypoxia or oxidative stress exhibited a morphology similar to control cells. Y-27632 reduced apoptosis, α-smooth muscle actin expression and reactive oxygen species generation under oxidative stress, and it reduced cell migration under hypoxia. Y-27632 also upregulated glutamine synthetase expression under hypoxia but did not impact vimentin expression. These findings suggest that Y-27632 protects Müller cells against cellular injury caused by oxidative stress and hypoxia by inhibiting the ROCK pathway.

Gardenia jasminoides attenuates hepatocellular injury and fibrosis in bile duct-ligated rats and human hepatic stellate cells

AIM:To investigate the anti-hepatofibrotic effects of Gardenia jasminoides in liver fibrosis.METHODS:Male Sprague-Dawley rats underwent common bile duct ligation(BDL) for 14 d and were treated with Gardenia jasminoides by gavage.The ef-fects of Gardenia jasminoides on liver fibrosis and the detailed molecular mechanisms were also assessed in human hepatic stellate cells(LX-2) in vitro.RESULTS:Treatment with Gardenia jasminoides decreased serum alanine aminotransferase(BDL vs BDL + 100 mg/kg Gardenia jasminoides,146.6 ± 15 U/L vs 77 ± 6.5 U/L,P = 0.0007) and aspartate aminotransferase(BDL vs BDL + 100 mg/kg Gardenia jasminoides,188 ± 35.2 U/L vs 128 ± 19 U/L,P = 0.005) as well as hydroxyproline(BDL vs BDL + 100 mg/kg Gardenia jasminoides,438 ± 40.2 μg/g vs 228 ± 10.3 μg/g liver tissue,P = 0.004) after BDL.Furthermore,Gardenia jasminoides significantly reduced liver mRNA and/or protein expression of transforming growth factor β1(TGF-β1),collagen type?Ⅰ?(Col?Ⅰ) and α-smooth muscle actin(α-SMA).Gardenia jasminoides significantly suppressed the upregulation of TGF-β1,Col?Ⅰand α-SMA in LX-2 exposed to recombinant TGF-β1.Moreover,Gardenia jasminoides inhibited TGF-β1-induced Smad2 phosphorylation in LX-2 cells.CONCLUSION:Gardenia jasminoides exerts antifibrotic effects in the liver fibrosis and may represent a novel antifibrotic agent.

Interaction between insulin-like growth factor binding protein-related protein 1 and transforming growth factor beta 1 in primary hepatic stellate cells

BACKGROUND: We previously showed that insulin-like growth factor binding protein-related protein 1 (IGFBPrP1) is a novel mediator in liver fibrosis. Transforming growth factor beta 1 (TGF beta 1) is known as the strongest effector of liver fibrosis. Therefore, we aimed to investigate the detailed interaction between IGFBPrP1 and TGF beta 1 in primary hepatic stellate cells (HSCs). METHODS: We overexpressed TGF beta 1 or IGFBPrP1 and inhibited TGF beta 1 expression in primary HSCs for 6, 12, 24, 48, 72, and 96 hours to investigate their interaction and observe the accompanying expressions of a-smooth muscle actin (alpha-SMA), collagen I, fibronectin, and phosphorylated-mothers against decapentaplegic homolog 2/3 (p-Smad2/3). RESULTS: We found that the adenovirus vector encoding the TGF beta 1 gene (AdTGF beta 1) induced IGFBPrP1 expression while that of alpha-SMA, collagen I, fibronectin, and TGF beta 1 increased gradually. Concomitantly, AdIGFBPrP1 upregulated TGF beta 1, alpha-SMA, collagen I, fibronectin, and p-Smad2/3 in a time-dependent manner while IGFBPrP1 expression was decreased at 96 hours. Inhibition of TGF beta 1 expression reduced the IGFBPrP1-stimulated expression of alpha-SMA, collagen I, fibronectin, and p-Smad2/3. CONCLUSIONS: These findings for the first time suggest the existence of a possible mutually regulation between IGFBPrP1 and TGF beta 1, which likely accelerates liver fibrosis progression. Furthermore, IGFBPrP1 likely participates in liver fibrosis in a TGF beta 1-depedent manner, and may act as an upstream regulatory factor of TGF beta 1 in the Smad pathway.

Myofibroblastic cell activation and neovascularization predict native liver survival and development of esophageal varices in biliary atresia

AIM: To study the relation between collagen 1, &#x003b1;-smooth muscle actin (&#x003b1;-SMA) and CD34 expression and the most essential portoenterostomy (PE) outcomes.

Effects of transforming growth factor β2 and connective tissue growth factor on induction of epithelial mesenchymal transition and extracellular matrix synthesis in human lens epithelial cells

AIM:To Investigate the effects of transforming growth factorβ2(TGF-β2)and connective tissue growth factor(CTGF)on transdifferentiation of human lens epithelial cells(HLECs)cultured in vitro and synthesis of extracellular matrix(ECM).METHODS:HLECs were treated with TGF-β2(0,0.5,1.0,5,10μg/L)and CTGF(0,15,30,60,100μg/L)for different times(0,24,48,72h)in vitro and the expression ofα-smooth muscle actin(α-SMA),the main component of the extracellular matrix typeⅠcollagen(Col-1)and fibronectin(Fn)were measured by using real-time polymerase chain reaction(PCR)and western-blot.RESULTS:TGF-β2 and CTGF significantly increased expression ofα-SMA mRNA and protein(P【0.05,P【0.001),Fn mRNA and protein(P【0.001),Col-1 mRNA and protein(P【0.001).TGF-β2 could induce HLECs expression of CTGF mRNA and protein in dosedependent manner(P【0.05,P【0.001).TGF-β2 and CTGF could induce HLECs to expressα-SMA,Fn and Col-1 in time-dependent manner.Each time of TGF-β2and CTGF induced HELCs expression ofα-SMA,Fn,Col-1 mRNA and protein was significant increase compared with control(P【0.05,P【0.001).CONCLUSION:TGF-β2 and CTGF could induce HLECs epithelial mesenchymal transition and ECM synthesis.

Morphological and migratory alterations in retinal Müller cells during early stages of hypoxia and oxidative stress

In the present study, retinal MOiler cells were cultured in vitro and treated with hydrogen peroxide （oxidative stressor） and cobalt chloride （hypoxic injury）. Following 24 hours of culture, compensatory hypertrophy was observed and cellular apoptosis increased. Hypoxia enhanced the migration ability of retinal MOiler cells and induced the expression of a-smooth muscle actin. Oxidative stress altered the morphology of MOiler cells when compared with hypoxia treatment.

Low-power laser irradiation inhibits arecoline-induced fibrosis:an in vitro study

Oral submucous fibrosis （OSF） is a potentially malignant disorder that is characterized by a progressive fibrosis in the oral submucosa. Arecoline, an alkaloid compound of the areca nut, is reported to be a major aetiological factor in the development of OSF. Low-power laser irradiation （LPLI） has been reported to be beneficial in fibrosis prevention in different damaged organs. The aim of this study was to investigate the potential therapeutic effects of LPLI on arecoline-induced fibrosis. Arecoline- stimulated human gingival fibroblasts （HGFs） were treated with or without LPLI. The expression levels of the fibrotic marker genes alpha-smooth muscle actin （a-SMA） and connective tissue growth factor （CTGF/CCN2） were analysed by quantitative real- time reverse transcription polymerase chain reaction （RT-PCR） and western blots. In addition, the transcriptional activity of CCN2 was further determined by a reporter assay. The results indicated that arecoline increased the messenger RNA and protein expression of CCN2 and a-SMA in HGF. Interestingly, both LPLI and forskolin, an adenylyl cyclase activator, reduced the expression of arecoline-mediated fibrotic marker genes and inhibited the transcriptional activity of CCN2. Moreover, pretreatment with SQ22536, an adenylyl cyclase inhibitor, blocked LPLI＇s inhibition of the expression of arecoline-mediated fibrotic marker genes. Our data suggest that LPLI may inhibit the expression of arecoline-mediated fibrotic marker genes via the cAMP signalling pathway.

Mesenteric inflammatory myofibroblastic tumors in an elder patient with early recurrence:A case report

Inflammatory myofibroblastic tumor （IMT） of the alimentary tract often occurs in children or young adults, but may occur at any age. Symptoms are nonspecific and depend on the location of the tumor. The most often involved sites are small bowel mesentery especially the distal ileum, mesotransverse colon, or great omentum. Recurrence appears to be more frequent in the extrapulmonary lesion. Herein we demonstrate a 63-year-old male patient with mesenteric INT, with an early recurrence after his first operation. We should be aware that if the tumor is larger than 8 cm, multinodular, omental, with ill-defined margin, with pathologically atypia, or ganglion-like cells, a close surveillence after primary surgery with image study might be necessary to detect the tumor recurrence early. Tumor recurrence may be asymptomatic, and it may act like a malignant tumor with a poor prognosis.

Antifibrotic effect of aloe vera in viral infection-induced hepatic periportal fibrosis

AIM:To investigate the anti-oxidative and anti-fibrotic effects of aloe vera in patients with liver fibrosis.METHODS:Aloe vera high molecular weight fractions(AHM) were processed by patented hyper-dry system in combination of freeze-dry technique with microwave and far infrared-ray radiation.Fifteen healthy volunteers as the control group and 40 patients were included.The patients were randomly subdivided into two equal groups:the conventional group was treated with placebo(starch),and AHM group was treated with 0.15 gm/d AHM,both for 12 consecutive weeks.The patients were investigated before and after treatment.Serum activity of aspartate aminotransferase(AST),alanine aminotransferase(ALT),alkaline phosphatase(ALP),hyaluronic acid(HA),transforming growth factor-β(TGF-β) and matrixmetalloproteinase-2(MMP-2) were determined.The reduced glutathione(GSH) and malondialdehyde(MDA) levels in liver were assayed and the expression of hepatic α-smooth muscle actin(α-SMA) was identified by immunohistochemistry.RESULTS:At the start of the study,the hematoxylin and eosin staining revealed fibro-proliferated bile ductules,thick fibrous septa and dense inflammatory cellular infiltration in the patients before treatment.The use of AHM for 12 wk significantly ameliorated the fibrosis,inhibited the inflammation,and resulted in minimal infiltration and minimal fibrosis compared to the conventional group.The enzyme activities of the liver(ALT,AST and ALP) were attenuated after treatment in both groups,and the decrease in the AHM group was more significant as compared with the conventional group.Similar to the AST,the MDA levels were significantly higher before treatment,and were attenuated after treatment in both groups.In contrast,the hepatic glutathione content in the patients were decreased significantly in the AHM group compared to the controls.The serum levels of the fibrosis markers(HA,TGF-β and MMP-2) were also reduced significantly after treatment.The expression of α-SMA was modified in patients before and after treatment as compared with the normal controls.In the conventional group,there was only thin and incomplete parenchymal α-SMA positive septum joining the thickened centrilobular veins,while in the AHM group,few α-SMA positive cells were present in sinusoid and lobule after treatment.CONCLUSION:Oral supplementation with AHM could be helpful in alleviating the fibrosis and inflammation of hepatic fibrosis patients.

Expression of transcription factors Slug in the lens epithelial cells undergoing epithelial-mesenchymal transition induced by connective tissue growth factor

AIMTo investigate the expression of transcription factors Slug in human lens epithelial cells (HLECs) undergoing epithelial-mesenchymal transition (EMT) induced by connective tissue growth factor (CTGF).METHODSHLECs were treated with CTGF of different concentrations (20, 50 and 100 ng/mL) or without CTGF (control) for 24h. The morphological changes of HLECs were analysed by microscopy. The expression and cellular localization of Slug was evaluated by immumo-fluorescence. Expressions of Slug, E-cadherin and alpha smooth muscle actin (&#x003b1;-SMA) were further determined by Western blot analysis.RESULTSHLECs showed spidle fibrolasts-like characteristics and loosely connected each other after CTGF treatment. The immuno-fluorescence staining indicated that Slug was localized in the nuclei and its expression was induced by CTGF. The relative expressions of Slug protein were 1.64&#x000b1;0.11, 1.96 &#x000b1;0.03, 3.12 &#x000b1;0.10, and 4.08&#x000b1;0.14, respectively, in response to control group and treatment with CTGF of 20, 50 and 100 ng/mL (F=443.86, P&#x0003c;0.01). The increased Slug protein levels were correlated well with up-expression of &#x003b1;-SMA (0.78&#x000b1;0.05, 0.85&#x000b1;0.06, 2.17&#x000b1;0.15, 2.86&#x000b1;0.10; F=449.85, P&#x0003c;0.01) and down-expression of E-cadherin (2.50&#x000b1;0.11, 1.79&#x000b1;0.26, 1.05&#x000b1;0.14, 0.63&#x000b1;0.08; F=101.55, P&#x0003c;0.01).CONCLUSIONTranscription factor Slug may be involved in EMT of HLECs induced by CTGF in vitro.