Arsenic-induced monocyte adhesion to VCAM-1 is prevented by antioxidant <i>in vitro</i>.

<p>U937 cells (1000 cells/ml) were pretreated for 1 h with NAC (1 mM) and then exposed to arsenic for 3h (0, 10, 50 or 200 ppb). Cells were stained with HEt and staining detected by flow cytometry (A). Alternatively, cells were fluorescently-labelled with orange tracker and incubated on VCAM-1/Fc coated plates (B). The non-adherent cells were washed away, and the adherent fluorescent cells were counted. Values are expressed as mean ± S.D., n ≥ 3. * p < 0.05: **: p < 0.01; ***: p < 0.001, compared to unexposed controls.</p

Arsenic increases adhesion molecule expression <i>in vivo</i>, which can be prevented by addition of high selenium diet.

<p>(A-B) In order to evaluate <i>in vivo</i> effects of arsenic, five-week-old male apoE^-/- mice fed normal rodent diet were exposed to arsenic (200 ppb) for 13 weeks or maintained on tap water. Carotids were stained for VCAM-1 (A) and whole blood was collected and CD29 expression was detected using flow-cytometry (B). (C-F) In order to evaluate ROS involvement in arsenic-induced atherosclerosis, five-week-old male apoE^-/- mice were exposed to arsenic (200 ppb) for 13 weeks or maintained on tap water. Mice were fed with low selenium or high selenium chow. Carotids were stained for ROS (C), or VCAM-1 (D). Blood was collected and CD29 expression was detected using flow-cytometry (E), or cells were fluorescently-labelled and incubated on VCAM-1/Fc coated plates (F). Values are expressed as mean ± S.D., n ≥ 3. * p < 0.05: **: p < 0.01; ***: p < 0.001, compared to unexposed controls.</p

Arsenic does not enhance platelet activation, platelet/monocyte interaction or platelet/neutrophil aggregates formation.

<p>C57BL/6 wild-type and apoE^-/- mouse platelets were collected and exposed to arsenic (10, 50 or 200 ppb) and/or thrombin for 5 minutes. Representative pictures of platelet spreading after control, 200 ppb arsenic or 1U thrombin are shown in A (left panels: C57BL/6; right panels: apoE^-/-). (B) P-selectin (CD62) expression was assessed by flow cytometry (up panel: C57BL/6; down panel: apoE^-/-). (C) Circulating CCL5 levels were measured in apoE^-/- male mice were left untreated or exposed to 200 ppb arsenic for 8 or 13 weeks using an immunoassay kit (multiplex bead-based) on a Bio-Plex 200 (Bio-Rad Laboratories, ON, Canada). Each sample (n = 4) was analyzed in duplicate (technical replicate). (D-E) The platelet/monocyte aggregates (CD14⁺/CD41⁺; D) and the platelet/neutrophil aggregates (Ly6G⁺/CD41⁺; E) were followed from day 14 to 28 in the circulation of mice exposed to 200 ppb arsenic. One mouse, as positive control, was treated with LPS for 18 hours before the blood collection. Values are expressed as mean ± S.D., n ≥ 3 animals.</p

Arsenic increases adhesion of mononuclear cells via increased CD29 binding to VCAM-1.

<p>U937 or human PBMC cells (1000 cells/ml) were exposed to arsenic for 72h (0, 10 or 200 ppb) (A). Cells were fluorescently-labelled and incubated on VCAM-1/Fc coated plates. The non-adherent cells were washed away, and adherent fluorescent cells were counted. In B, cellular surface CD29 antigens were detected by flow-cytometry using anti-CD29 antibody. (C-D) CD29 blocking antibody was added to <i>in vitro</i> U937 binding assays to VCAM-1/Fc (C) or to <i>ex vivo</i> organ cultures with primary mononuclear cells (D). Values are expressed as mean ± S.D., n ≥ 3. * p < 0.05: **: p < 0.01; ***: p < 0.001, compared to unexposed controls.</p

Arsenic induces monocyte adhesion to endothelial cells, with maximal binding achieved following exposure of both cell types.

<p>(A) U937 and/or HUVEC cells (1000 cells/ml) were exposed to arsenic overnight (0, 10 or 200 ppb). U937 cells were fluorescently-labelled and were incubated with HUVEC cells. The non-adhered cells were washed away, and the adherent fluorescent cells were counted. Data are expressed as relative number of U937 over total HUVEC stained cells. ** = p<0.01; *** = p<0.001 B) Representative pictures are shown (40X). C-D) Organ culture of carotid arteries and fluorescently-labelled bone marrow cells where neither, one, or both components were exposed to arsenic trioxide overnight and allowed to adhere to each other for 30 min before being washed. Adherent cells were counted. Representative pictures of 200 ppb arsenic-exposed are shown in C. D) Both components were exposed to either 10 or 200 ppb arsenic. Data represent ratio ± S.D., n ≥ 3. **: p < 0.01; ***: p < 0.001, compared to unexposed controls.</p