Adhesion of RBC to HUVEC and the Formation of Lipid Peroxides
| Treatment of HUVEC . | Inhibitor . | nmole MDA/106 HUVEC/4 h . | Adherence of RBC (%) . |
|---|---|---|---|
| None | — | 8.4 ± 1.3 | — |
| EC-CM (100 μg/mL) | — | 8.7 ± 0.6 | — |
| AA RBC | — | 10.8 ± 3.2 | 0.3 ± 0.1 |
| AA RBC + E-CM | — | 11.5 ± 3.5 | 0.5 ± 0.15 |
| SS RBC | — | 30.3 ± 5.5 | 0.8 ± 0.2 |
| SS RBC + E-CM (50 μg/mL) | — | ND | 1.4 ± 0.3 |
| SS RBC + E-CM (100 μg/mL) | — | 54.2 ± 4.0 | 2.9 ± 0.4 |
| SS RBC + E-CM (200 μg/mL) | — | 56.0 ± 5.5 | 2.9 ± 0.5 |
| SS RBC + E-CM (100 μg/mL) | KYRGDS | 24.0 ± 7.4 | 0.9 ± 0.2 |
| SS RBC + E-CM (100 μg/mL) | AGDV | 51.4 ± 4.1 | 2.8 ± 0.3 |
| SS RBC + E-CM (100 μg/mL) | SOD + Catalase | 17.3 ± 3.5 | ND |
| SS RBC + E-CM (100 μg/mL) | Ab-vWF | ND | 1.2 ± 0.2 |
| Treatment of HUVEC . | Inhibitor . | nmole MDA/106 HUVEC/4 h . | Adherence of RBC (%) . |
|---|---|---|---|
| None | — | 8.4 ± 1.3 | — |
| EC-CM (100 μg/mL) | — | 8.7 ± 0.6 | — |
| AA RBC | — | 10.8 ± 3.2 | 0.3 ± 0.1 |
| AA RBC + E-CM | — | 11.5 ± 3.5 | 0.5 ± 0.15 |
| SS RBC | — | 30.3 ± 5.5 | 0.8 ± 0.2 |
| SS RBC + E-CM (50 μg/mL) | — | ND | 1.4 ± 0.3 |
| SS RBC + E-CM (100 μg/mL) | — | 54.2 ± 4.0 | 2.9 ± 0.4 |
| SS RBC + E-CM (200 μg/mL) | — | 56.0 ± 5.5 | 2.9 ± 0.5 |
| SS RBC + E-CM (100 μg/mL) | KYRGDS | 24.0 ± 7.4 | 0.9 ± 0.2 |
| SS RBC + E-CM (100 μg/mL) | AGDV | 51.4 ± 4.1 | 2.8 ± 0.3 |
| SS RBC + E-CM (100 μg/mL) | SOD + Catalase | 17.3 ± 3.5 | ND |
| SS RBC + E-CM (100 μg/mL) | Ab-vWF | ND | 1.2 ± 0.2 |
HUVEC monolayer (2 × 106 cells) were incubated with RBC (2% Hct) in the absence and presence of E-CM for 4 hours. Where indicated synthetic peptides (KYRGDS or AGDV [100 μmol/L]; free-radical scavengers (superoxide dismutase [200 U/mL]) and catalase (200 U/mL); or antibody (vWf, 5 μg/mL) were preincubated with HUVEC for 30 minutes before the addition of RBC. Nonadherent RBC were removed by aspiration and HUVEC washed three times with PBS. The amount of lipid peroxides ie, MDA formed in HUVEC was determined by reaction with TBA. Incubation of SS RBC (2% Hct) alone in buffer at 37°C for 2 or 4 hours did not show the formation of MDA, as its formation was below the limit of detection by the MDA assay used. The extent of adherence of RBC was determined by incubation of51Cr-labeled RBC with HUVEC monolayer. Results are mean ± SD of AA RBC (n = 4) and SS RBC (n = 4) of duplicate determinations.
Abbreviation: ND, not determined.