Fig. 1.

Deleterious effects of next generation tobacco and nicotine products on primary human endothelial cells. (A) Representative pictures of wound healing after 8 h including degree of wound healing in % are shown. Direction of flow was perpendicular to the wound in the cell layer, n ≥ 8. Evaluation of endothelial cell viability under (B) high laminar flow conditions (30 dyn/cm²) or (C) low laminar flow conditions (1 dyn/cm²) using CellTiter-Glo Luminescent Cell Viability Assay. HUVEC monolayers were stimulated with aqueous extracts of the indicated tobacco products in a nicotine concentration range from 0.56 to 4.96 μg/ml for 24 h, n ≥ 3. (D) Evaluation of area under the curve data of endothelial wound healing capability under combined stimulation with aqueous extracts of different tobacco products (3R4F, HTP, e-cig and nic) and high laminar flow, n ≥ 8. (E) Regulation of endothelial nitric oxide (NO) release in response to flow and AqE stimulation. NO release was determined by Griess assay, n ≥ 4. (F) mRNA expression of eNOS in HUVEC after exposure to high laminar flow in combination with aqueous extracts for 8 h, n ≥ 4. (G) eNOS phosphorylation status after 8 h of AqE treatment under static and high laminar flow conditions, n ≥ 4. max = maximum nicotine concentration (Table 2). Data are shown as mean ± SD. Statistics: One-Way ANOVA, *p < 0.05 vs. static control, #p < 0.05 vs. time-matched control (0 μg/ml nicotine, 1 respectively 30 dyn/cm²), §p < 0.05.