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. 2008 Dec 23;41(2):207–216. doi: 10.1165/rcmb.2008-0209OC

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Figure 5. — Dietary zinc supplementation mitigates the effects of chronic alcohol ingestion on the expression of the tight junction occludin and ZO-1 in the alveolar epithelium. Alveolar epithelial cells were freshly isolated from control-fed rats, alcohol-fed rats, and alcohol-fed rats whose diets were supplemented with zinc acetate (500 mg/L). Gene expression of the tight junction transmembrane protein, occludin, and of the tight junction scaffolding protein, ZO-1, were determined by real-time PCR. The average relative gene expression normalized to 18S was determined by the comparative method (2^−ΔΔCt), with the target gene expression in cells from control-fed rats set as 1 in A and from alcohol-fed rats set as 1 in B. A shows the relative gene expression of occludin and ZO-1 in cells from control-fed rats (open bars) versus alcohol-fed rats (solid bars), and B shows the relative gene expression of occludin and ZO-1 in cells from alcohol-fed rats (solid bars) versus cells from alcohol-fed rats supplemented with dietary zinc (shaded bars). Each value represents the mean ± SEM of three to four separate isolated cell preparations. In parallel, freshly isolated alveolar epithelial cells from alcohol-fed rats and alcohol-fed rats supplemented with dietary zinc were cultured for 8 days to form monolayers and then occludin and ZO-1 protein localization were assessed qualitatively by fluorescent immunocytochemistry as described in Materials and Methods. Representative images are shown in C. Epithelial monolayers from alcohol-fed rats showed decreased overall staining for both occludin and ZO-1, with the most striking abnormality being discontinuous staining in the cell membranes. In contrast, epithelial monolayers from alcohol-fed rats supplemented with dietary zinc had more intense and continuous staining of occludin and ZO-1 in cell membranes. (Arrows in the occludin images are provided to highlight these findings.) (A) *P < 0.05 compared with control-fed rats; (B) *P < 0.05 compared with alcohol-fed rats.

Figure 5. — Dietary zinc supplementation mitigates the effects of chronic alcohol ingestion on the expression of the tight junction occludin and ZO-1 in the alveolar epithelium. Alveolar epithelial cells were freshly isolated from control-fed rats, alcohol-fed rats, and alcohol-fed rats whose diets were supplemented with zinc acetate (500 mg/L). Gene expression of the tight junction transmembrane protein, occludin, and of the tight junction scaffolding protein, ZO-1, were determined by real-time PCR. The average relative gene expression normalized to 18S was determined by the comparative method (2^−ΔΔCt), with the target gene expression in cells from control-fed rats set as 1 in A and from alcohol-fed rats set as 1 in B. A shows the relative gene expression of occludin and ZO-1 in cells from control-fed rats (open bars) versus alcohol-fed rats (solid bars), and B shows the relative gene expression of occludin and ZO-1 in cells from alcohol-fed rats (solid bars) versus cells from alcohol-fed rats supplemented with dietary zinc (shaded bars). Each value represents the mean ± SEM of three to four separate isolated cell preparations. In parallel, freshly isolated alveolar epithelial cells from alcohol-fed rats and alcohol-fed rats supplemented with dietary zinc were cultured for 8 days to form monolayers and then occludin and ZO-1 protein localization were assessed qualitatively by fluorescent immunocytochemistry as described in Materials and Methods. Representative images are shown in C. Epithelial monolayers from alcohol-fed rats showed decreased overall staining for both occludin and ZO-1, with the most striking abnormality being discontinuous staining in the cell membranes. In contrast, epithelial monolayers from alcohol-fed rats supplemented with dietary zinc had more intense and continuous staining of occludin and ZO-1 in cell membranes. (Arrows in the occludin images are provided to highlight these findings.) (A) *P < 0.05 compared with control-fed rats; (B) *P < 0.05 compared with alcohol-fed rats.