Dysregulation of upstream and downstream transforming growth factor-β transcripts in livers of children with biliary atresia and fibrogenic gene signatures

Abstract

INTRODUCTION Our previous work demonstrated that the transforming-growth factor (TGF) β pathway plays a central role in the liver fibrosis associated with experimental biliary atresia (BA). To confirm these findings in humans, we performed an in silico analysis of publicly available microarray data from liver specimens from children with BA, with the hypothesis that the TGF β pathway would be dysregulated. METHODS We analyzed publicly available liver gene expression microarray data from 47 infants with BA. We re-analyzed the microarray image files and clinical data to compare gene expression differences between the fibrogenic and inflammatory cohorts identified in the initial study. Targets from the microarray analysis were confirmed using the animal model of BA. RESULTS Analysis of variance (ANOVA) detected 6903 transcripts (2822 distinct genes) differentially regulated between groups (p<0.01; fold change >1.2). We used a targeted approach to identified a subgroup of 24 TGF β-related transcripts. Expressions for procollagen transcripts were increased in the fibrogenic group (1.2 fold to 1.4 fold); expression of matrix metalloproteinase (MMP)-7 was similarly increased 2-fold, while MMP-9 and plasminogen activator inhibitor-1 were decreased 2-fold and 3-fold respectively. Integrins β5 (1.18 fold) and β8 (1.84 fold) also demonstrated increased expression in the fibrogenic group. Increased expression of β5 (3-fold) and β8 (5-fold) as well as Smad-3 (4-fold) and Smad interacting protein (SIP)-1 (3.5 fold) mRNA were confirmed in experimental BA. Phosphorylated Smad 3 protein in the experimental group was also nearly twice that of the control group, further implicating the TGF-β pathway. CONCLUSION Gene transcripts for known upstream and downstream TGF-β mediators are differentially expressed in liver specimens from children with BA and a fibrogenic gene signature. The same integrins that were dysregulated in the human specimens were also found to be upregulated in our animal BA model, as were other intermediaries in the TGF-β pathway. Further investigation into whether these mediators may be attractive targets for future therapy in children with BA is warranted.

Increased MMP‐7 expression in biliary epithelium and serum underpins native liver fibrosis after successful portoenterostomy in biliary atresia

“…In rhesus rotavirus infected rats, an animal model of BA, liver expression of MMP‐7 is increased . In several previous studies, hepatic gene expression of MMP‐7 has been shown to be upregulated both at the time of PE and LTx, implicating its potential role in liver fibrogenesis in BA . It should be emphasized that at these extreme stages of the disease the majority of patients are severely cholestatic.…”

“…MMP-7 has been shown to be upregulated in liver and lung fibrosis, and studies with a selective MMP-7 inhibitor isofraxidin have been promising in counteracting progression of fibrosis [14][15][16][17]. Previous studies have shown significant alterations in hepatic expression and serum levels of MMPs and their inhibitors in BA patients at the time of PE and LTx [15,[18][19][20][21][22][23][24][25]. Whether the altered liver expression occurs secondary to cholestasis remains unknown, while little is known about expression of MMPs and TIMPs after successful PE and clearance of jaundice.…”

αv integrins: key regulators of tissue fibrosis

“…Interestingly, dysregulation of MMP14 has also been reported in both human and animal models of biliary atresia (Iordanskaia et al 2013). This suggests that the activation of TGFβ in the setting of biliary atresia may in part be mediated through both increased αvβ8 expression and altered expression of MMP14.…”

Targeting Extracellular Cyclophilins Ameliorates Disease Progression in Experimental Biliary Atresia

“…Liver samples were homogenized according to the manufacturer's instructions in lysis buffer using homogenizer FastPrep-24 (MP Biomedicals). Subsequent manipulations were performed as described earlier (19). The concentration of murine CypA in plasma was determined using ELISA kits from MyBioSource following the manufacturer's instructions.…”