Effect of NF-κB p65 antisense oligodeoxynucleotide on transdifferentiation of normal human lens epithelial cells induced by transforming growth factor-β2

AIM：To study the inhibition of nuclear factor kappa-B p65（NF-κB p65）antisense oligodeoxynucleotide（ASODN）on transdifferentiation of normal human lens epithelial cells induced by transforming growth factor-β2（TGF-β2）.·M ETHODS：NF-κBp65ASODNand NF-κBp65missense oligodeoxynucleotide（MSODN）were designed and synthesized.Human lens epithelial cell line（HLE B-3）cells were prepared for study and divided into 7 groups.Control group was HLE B-3 cells cultured in dulbecco’s modified eagle medium（DMEM）.T1,T2,and T3 group were HLE B-3 cells cultured in DMEM with 10 ng/m L TGF-β2 for 6h,12h,24h respectively.A＋T group was HLE B-3 cells cultured with 10 ng/m L TGF-β2for 24h after transfected by NF-κB p65 ASODN for 24h.M＋T group was HLE B-3 cells cultured with 10 ng/m L TGF-β2 for 24h after transfected by NF-κB p65 MSODN for 24h.The negative control group was HLE B-3 cells cultured with 10 ng/m L TGF-β2 for 24h after cultured with transfer agent（Hi Per Fect）for 24h.Cell morphology was observed at different time points using an inverted microscope.The expression of NF-κB p65 m RNA was detected with reverse transcription-polymerase chain reaction（RT-PCR）,and the expression ofα-smooth muscle actin（α-SMA）protein was assayed with ELISA.·RESULTS：With the TGF-β2 stimulation prolongation,the expression of NF-κB p65 m RNA and a-SMA protein increased in T1,T2,T3 groups compared with the control group,and the difference was statistically significant（〈0.05）.NF-κB p65 ASODN lowered the expression of NF-κB p65 m RNA andα-SMA protein induced by TGF-β2.NF-κB p65 MSODN and Hi Per Fect did not lower the expression of NF-κB p65 m RNA andα-SMA protein induced by TGF-β2.The difference between control group and A＋T group was not statistically significant（〉0.05）,but the difference among A＋T group and other groups was statistically significant（〈0.05）.·CONCLUSION：NF-κB p65 ASODN could lower the expression of NF-κB p65 m RNA andα-SMA protein induced by TGF-β2,and antagonized TGF-β2-induced transdifferentiation of HLE B-3.NF-κB p65ASODN could be used as a new biological therapeutic target of posterior capsular opacification.

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.

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.