Study of The Potential Antifibrotic Effects of Forskolin in An Experimental Model of Induced- Liver Fibrosis in Rats

Abstract

The current study was conducted to investigate the potential antifibrotic effect of forskolin as well as the different mechanisms underlying this postulated effect. This could be established by measuring its effect on oxidative stress, NF-κB and inflammation,TGF-β1 as well as its effect on Hh pathway signaling. First, an acute preliminary study was designed to assess the most effective hepatoprotective dose of forskolin. In this study, rats were divided into 5 groups: • Group 1: served as the control group and received vehicles only. • Group 2: served as the hepatotoxic model received a single i.p. dose of CCl4 (1ml /kg) • Group 3: treated with 5 mg/kg i.p. forskolin for 7 days followed by a single i.p. dose of CCl4 (1ml/kg). • Group 4:treated with 10 mg/kg i.p. forskolin for 7 days followed by a single i.p. dose of CCl4 (1ml/kg). • Group 5:treated with 20 mg/kg i.p. forskolin for 7 days followed by a single i.p. dose of CCl4 (1ml/kg). This study revealed that 10 mg/kg forskolin is the most effective hepatoprotective dose since it significantly decreased ALT and AST levels compared to the CCl4group and the other doses of forskolin. Also, upon histopathological examination, this dose significantly reduced the extensive hepatocellular damage and inhibited necrosis caused by CCl4. Second, a chronic study was designed to investigate the potential antifibrotic effect of forskolin. In this study, rats were divided into 4 groups and treated for 6 weeks as follows: • Group A:served as the normal control, receiving vehicles only. • Group B: served as the fibrotic model, receiving CCl4 (0.5 ml/kg, 1:1 mixture with corn oil, i.p.) twice weekly. • Group C: was given both CCl4 (0.5 ml/kg, 1:1 mixture with corn oil, i.p.) twice weekly and forskolin (10 mg/kg, i.p., dissolved in a DMSO/saline solution (1:20)) five times per week. • Group D: was given forskolin alone (10 mg/kg, i.p., dissolved in a DMSO/saline solution (1:20)) five times per week.

 The following parameters were assessed: 2. Hepatotoxicity markers by: • Determination of liver index. • Measuring serum levels of ALT, AST, TC, TG, total bilirubin, and albumin colorimetrically. 3. Histopathology (H&E staining) 4. Fibrosis markers by: • Measuring liver content of TGF-β1 using ELISA kit. • Measuring α-SMA expression in liver tissue by immunohistochemical staining. • Measuring collagen by: c) Determining liver hydroxyproline content colorimetrically. d) Detection of collagen fibers using Masson’s trichrome stain. 5. Oxidative stress markers by: • Measuring liver content of GSH and MDA colorimetrically. 6. Inflammatory markers by: • Measuring NF-κBexpression in liver tissue by immunohistochemical staining. • Measuring liver content of TNF-α using ELISA kit. • Measuring COX-2 expression in liver tissue by immunohistochemical staining. 7. Hedgehog markers: • Measuring the gene expression of PTCH-1, SMO, and GLI-2 in liver by qRT-PCR.