PMC

Cell viability testing. Equisetum arvense L. extract was tested across a range of concentrations up to 5000 μg/ml in HUVECs. The viability graphs did not show significant changes in cell viability even at high doses (5000 μg/ml) (mean values ± standard deviation, n = 3).

Malondialdehyde levels evaluated in endothelial cells exposed to hyperosmotic stress and pretreated with two doses of Equisetum arvense L. extract. (a) The MDA levels after exposure to hyperosmotic stress and pretreatment for 24 h with high doses of Equisetum arvense L. extract, caffeic acid, and cathechin. (b) MDA levels after exposure to hypertonic conditions and low doses of Equisetum arvense L. extract, caffeic acid, and cathechin. The statistical significance of the difference between the treated and control groups was evaluated with two-way ANOVA, followed by Dunnett's multiple test; ^###p < 0.001, treated versus untreated cells in hypertonic medium; ^∗∗∗p < 0.001, hypertonic medium versus the parameters in normotonic conditions.

The activities of caspase-3 and caspase-8 evaluated in endothelial cells exposed to hyperosmotic stress and pretreated with two doses of Equisetum arvense L. extract. (a) The caspase-3 activity in endothelial cells after exposure to hyperosmotic stress and pretreatment for 24 h with high doses of Equisetum arvense L. extract, caffeic acid, and cathechin. (b) The caspase-3 activity in endothelial cells after exposure to hypertonic conditions and low doses of Equisetum arvense L. extract, caffeic acid, and cathechin. (c) The caspase-8 activity in endothelial cells after exposure to hyperosmotic stress and pretreatment for 24 h with high doses of Equisetum arvense L. extract, caffeic acid, and cathechin. (d) The caspase-8 activity in endothelial cells after exposure to hypertonic conditions and low doses of Equisetum arvense L. extract, caffeic acid, and cathechin. The statistical significance of the difference between treated and control groups was evaluated with two-way ANOVA, followed by Dunnett's multiple test, ^#p < 0.05, ^##p < 0.01, and ^###p < 0.001, treated versus untreated cells in hypertonic medium; ^∗p < 0.05 and ^∗∗p < 0.01, hypertonic medium versus the parameters in normotonic conditions.

Proinflammatory cytokines evaluated in endothelial cells exposed to hyperosmotic stress and pretreated with two doses of Equisetum arvense L. extract. (a) The IL-6 secretion in cell lysates after exposure to hyperosmotic stress and pretreatment for 24 h with high doses of Equisetum arvense L. extract, caffeic acid, and cathechin. (b) IL-6 level in cell lysates after exposure to hypertonic conditions and low doses of the three substances tested. (c, d) IL-6 concentrations in medium after exposure to normotonic and hypertonic conditions and treatment with the three substances tested. (e) TNF-α level in cell lysates in hypertonic conditions and pretreatment with high doses of Equisetum arvense L. extract, caffeic acid, and cathechin. (f) TNF-α secretion in cell lysates in hypertonic conditions and pretreatment with low doses of Equisetum arvense L. extract, caffeic acid, and cathechin. The statistical significance of the difference between treated and control groups was evaluated with two-way ANOVA, followed by Dunnett's Multiple test; ^###p < 0.001, treated versus untreated cells in hypertonic medium; ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001, hypertonic medium versus the parameters in normotonic conditions.

The expressions of NF-κB, pNF-κB, IκB, and pIκB in endothelial cells exposed to hyperosmotic stress and pretreated with low dose of Equisetum arvense L., caffeic acid, and cathechin. (a) Comparative Western blot images showing expressions of NF-κB, pNF-κB, IκB, and pIκB in HUVECs exposed to normotonic and hypertonic conditions and treated with the three compounds. (b) Image analysis of Western blot bands was done by densitometry; results were normalised to GAPDH. Western blot images: 1 = cells in normotonic medium, 2 = cells in normotonic medium treated with Equisetum arvense L., 3 = cells in normotonic medium treated with caffeic acid, 4 = cells in normotonic medium treated with cathechin, 5 = cells in hypertonic medium, 6 = cells in hypertonic medium treated with Equisetum arvense L., 7 = cells in hypertonic medium treated with caffeic acid, and 8 = cells in hypertonic medium treated with cathechin. Graphical representation of the quantitative Western blot results for HUVECs in the two conditions. Two-way ANOVA, followed by Dunnett's multiple test, was used to evaluate the statistical significance of differences in the mean values of the measured parameters. Each bar represents mean ± standard deviation (n = 3), ^#p < 0.05, ^##p < 0.01, and ^###p < 0.001, treated versus untreated cells in hypertonic medium; ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001, hypertonic medium versus the parameters in normotonic conditions.