Microscopic analyses of DEN-induced HCC with liver cirrhosis.

<p>Pathologic findings after 16 weeks of DEN administration; hematoxylin and eosin (H&E) staining. DN, dysplastic nodule, black arrowheads; early HCC, green arrowheads; HCC, blue arrowheads; A, original magnification ×5; B, normal; C, DN; D, early HCC; E, HCC; B–E, original magnification ×200. The mean areas (F and H) and numbers (G and I) of DNs and tumors after 16 and 19 weeks of DEN administration were evaluated and compared. Values shown are means ± standard deviation (n = 5/group). *<i>P</i><0.05; **<i>P</i><0.005.</p

Primer sequences used for semi-quantitative and quantitative Real-Time RT-PCR.

<p>Primer sequences used for semi-quantitative and quantitative Real-Time RT-PCR.</p

Effects of BCAA on DEN-induced liver fibrosis.

<p>Paraffin-embedded sections were stained with Masson’s trichrome to evaluate the progression of fibrosis after 14 and 16 weeks of DEN administration (A–F, original magnification ×5 and ×50). Index of fibrosis (G). The expression of α-smooth muscle actin (SMA) protein in livers was evaluated by Western blot analysis at 16 weeks post-DEN injection (H). An anti-β-actin antibody served as the loading control. Values shown are means ± standard deviation (n = 5/group). *<i>P</i><0.05. Relative fibrosis area: rMA = (MA/FA)×100. FA, overall field area (mm²); MA, overall mask area (mm²), which is the summed areas for each detected object in each layer.</p

Effects of BCAA supplementation on angiogenesis in rats after 16 weeks of DEN administration.

<p>The relative band intensities of the angiogenesis markers are shown in the right-hand panels (A). The expression level of Tie-2 mRNA was determined by RT-PCR (B). VEGF protein expression was determined by Western blotting (C). The expression level of each gene is normalized to those of GAPDH (A and B) and β-actin (C). The lanes contain mRNA (A) and protein (C) samples from three rats per group. Values shown are means ± standard deviation. *<i>P</i><0.05.</p

Liver/body weight ratio and results of biochemical analyses according to the duration of DEN administration in each group.

<p>Values shown are means ± SD.</p>*<p>Significantly different from the 3% casein group at <i>P</i><0.05.</p>**<p>Significantly different from the 3% casein group at <i>P</i><0.005.</p>***<p>Significantly different from the 3% casein group at <i>P</i><0.0005.</p