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Additional Supporting Information may be found in the online version of this article.

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HEP_25613_sm_SuppInfo.doc48KSupporting Information
HEP_25613_sm_SuppFigS1.TIF5146KSupporting Information Figure 1. Immunohistochemistry of quiescent and activated markers of HSCs. Liver sections from two models of hepatic fibrosis [BDL; top panels, Ethanol/LPS (ELPS); bottom panels]. Quiescent and activated HSCs were detected by GFAP and αSMA staining, respectively (brown chromagen) and counterstained with methyl green.
HEP_25613_sm_SuppFigS2.TIF300KSupporting Information Figure 2. Efficient transfection of miR 19b in activated HSCs. Representative qRT-PCR analysis of miR 19b expression in HSCs transfected with Lipofectamine 2000 alone (LIPO), negative control (SCR) or mature miR 19b (25-75nM) for 24 h. Expression was normalized to 4.5S RNA. Data are presented as mean ± SE. *Differs from SCR, p<0.05.
HEP_25613_sm_SuppFigS3.TIF292KSupporting Information Figure 3. TGFβRII is downregulated by miR 19b in human HSCs. qRT-PCR analysis of TGFβRII gene expression following 24 h of miR 19b transfection in LX-2 cells normalized to SNORD44. Data are presented as mean ± SE. *Differs from SCR, p<0.05.
HEP_25613_sm_SuppFigS4.TIF276KSupporting Information Figure 4. miR 19b does not affect SMAD4 gene expression in activated HSCs. qRT-PCR analysis of SMAD4 gene expression following 24 h of miR 19b transfection studies as normalized to levels of β-actin (n=3).
HEP_25613_sm_SuppFigS5.TIF253KSupporting Information Figure 5. Collagen secretion is inhibited by miR 19b in activated HSCs. Quantitative densitometry of immunoblot analyses of secreted type I collagen protein expression following 48 h of miR 19b (75 nM) transfection in HSCs (day 6). Culture medium was harvested following 48 h and proteins concentrated using Nanosep tubes (Pall Corporation; Ann Arbor, MI). Concentrated protein (10 μg) was used for standard immunoblotting analysis. Quantitative densitometry of secreted type I collagen was performed and normalized to total protein (n=3). Data are presented as mean ± SE.
HEP_25613_sm_SuppFigS6.TIF415KSupporting Information Figure 6. miR 19b decreases TGFβ1 expression in activated HSCs. qRT-PCR analysis of TGFβ1 gene expression following (A) 24 h of miR 19b transfection. (B) qRT-PCR analysis of TGFβ1 gene expression following 48 h of miR 19b transfection with or without recombinant human TGFβ (5 ng/ml). Expression was normalized to β-actin (n=3). Data are presented as mean ± SE. *Differs from SCR, #Differs from TGFβ, p<0.05.
HEP_25613_sm_SuppFigS7.TIF744KSupporting Information Figure 7. miR 19b blunts the activated HSC phenotype. Morphological assessment of activated HSCs (day 6) following 48 h of transfection with negative control (SCR) or miR 19b (75 nM).
HEP_25613_sm_SuppFigS8.TIF5360KSupporting Information Figure 8. Down-regulation of miR 19b in a rodent model of hepatic fibrosis. Representative light micrographs (20X with 40X insets) of sham (normal) and BDL (fibrotic) tissue sections following in situ hybridization with double DIG labeled LNA miR 19b probes. miR 19b expression levels are marked by dark blue chromagen staining (denoted by black arrows).
HEP_25613_sm_SuppFigS9.TIF310KSupporting Information Figure 9. miR 19b expression in primary liver cells. Cells were harvested and miR 19b expression was determined in quiescent (n=7), activated (day 14, n=6) HSCs and primary hepatocytes as assessed by qRT-PCR. Data are presented as mean ± SE. *Differs from quiescent, p<0.05.
HEP_25613_sm_SuppFigS10.TIF1706KSupporting Information Figure 10. Immunofluorescent images of decreased miR 19b expression in a rodent model of hepatic fibrosis. Representative immunofluorescent images of sham (normal) and BDL (fibrotic) tissue sections following in situ hybridization with double DIG labeled LNA miR 19b probes. Quiescent HSCs were detected by GFAP (green fluorescence) and miR 19b expression levels are marked by red fluorescence. Nuclear staining is observed by DAPI (blue) staining.
HEP_25613_sm_SuppFigS11.TIF1687KSupporting Information Figure 11. In situ hybridization of miR 19b in a mild fibrotic liver model. Representative light micrographs (20×) of liver tissue sections. miR 19b expression levels are marked by dark blue chromagen staining. Liver tissue harvested from controls and rats fed ethanol with bi-weekly injections of LPS 15.

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