P-selectin and IL-8 cooperatively trigger β2 integrin extension and hybrid-domain swing-out in human neutrophils
| . | Inhibitor . | |||
|---|---|---|---|---|
| None . | AS-605240 . | GGTI-298 . | AS-605240 + GGTI-298 . | |
| β2 integrin, total (mAb IB4) | ||||
| Stimulation | ||||
| Control | 155 ± 25 | 165 ± 18 | 150 ± 22 | 170 ± 25 |
| P-sel | 160 ± 20 | 148 ± 20 | 140 ± 25 | 150 ± 30 |
| IL-8 (0.1 nM) | 150 ± 22 | 170 ± 25 | 155 ± 20 | 180 ± 25 |
| IL-8 (1 nM) | 200 ± 35 | 190 ± 15 | 200 ± 18 | 210 ± 40 |
| P-sel/IL-8 (0.1 nM) | 185 ± 30 | 185 ± 18 | 178 ± 20 | 200 ± 30 |
| P-sel/IL-8 (1 nM) | 210 ± 35 | 200 ± 40 | 190 ± 35 | 220 ± 50 |
| β2 integrin, extended (mAb KIM127) | ||||
| Stimulation | ||||
| Control | 8 ± 2 | 11 ± 3 | 10 ± 2 | 12 ± 5 |
| P-sel | 45 ± 8* | 38 ± 10 | 40 ± 7 | 42 ± 5 |
| IL-8 (0.1 nM) | 10 ± 2 | 13 ± 4 | 15 ± 8 | 12 ± 5 |
| IL-8 (1 nM) | 120 ± 18* | 78 ± 10† | 70 ± 10† | 30 ± 10†‡ |
| P-sel/IL-8 (0.1 nM) | 90 ± 10§ | 68 ± 8† | 62 ± 5† | 48 ± 5†‡ |
| P-sel/IL-8 (1 nM) | 165 ± 22§ | 110 ± 15† | 105 ± 10† | 65 ± 15†‡ |
| β2 integrin, extended with hybrid-domain swing-out (mAb MEM148) | ||||
| Stimulation | ||||
| Control | 15 ± 5 | 17 ± 5 | 20 ± 8 | 20 ± 5 |
| P-sel | 18 ± 5 | 20 ± 4 | 15 ± 5 | 22 ± 5 |
| IL-8 (0.1 nM) | 20 ± 5 | 16 ± 5 | 22 ± 6 | 20 ± 5 |
| IL-8 (1 nM) | 178 ± 20* | 88 ± 10† | 75 ± 10† | 25 ± 5†‡ |
| P-sel/IL-8 (0.1 nM) | 65 ± 8§ | 35 ± 6† | 40 ± 5† | 17 ± 10†‡ |
| P-sel/IL-8 (1 nM) | 220 ± 25§ | 145 ± 30† | 115 ± 20† | 35 ± 10†‡ |
| . | Inhibitor . | |||
|---|---|---|---|---|
| None . | AS-605240 . | GGTI-298 . | AS-605240 + GGTI-298 . | |
| β2 integrin, total (mAb IB4) | ||||
| Stimulation | ||||
| Control | 155 ± 25 | 165 ± 18 | 150 ± 22 | 170 ± 25 |
| P-sel | 160 ± 20 | 148 ± 20 | 140 ± 25 | 150 ± 30 |
| IL-8 (0.1 nM) | 150 ± 22 | 170 ± 25 | 155 ± 20 | 180 ± 25 |
| IL-8 (1 nM) | 200 ± 35 | 190 ± 15 | 200 ± 18 | 210 ± 40 |
| P-sel/IL-8 (0.1 nM) | 185 ± 30 | 185 ± 18 | 178 ± 20 | 200 ± 30 |
| P-sel/IL-8 (1 nM) | 210 ± 35 | 200 ± 40 | 190 ± 35 | 220 ± 50 |
| β2 integrin, extended (mAb KIM127) | ||||
| Stimulation | ||||
| Control | 8 ± 2 | 11 ± 3 | 10 ± 2 | 12 ± 5 |
| P-sel | 45 ± 8* | 38 ± 10 | 40 ± 7 | 42 ± 5 |
| IL-8 (0.1 nM) | 10 ± 2 | 13 ± 4 | 15 ± 8 | 12 ± 5 |
| IL-8 (1 nM) | 120 ± 18* | 78 ± 10† | 70 ± 10† | 30 ± 10†‡ |
| P-sel/IL-8 (0.1 nM) | 90 ± 10§ | 68 ± 8† | 62 ± 5† | 48 ± 5†‡ |
| P-sel/IL-8 (1 nM) | 165 ± 22§ | 110 ± 15† | 105 ± 10† | 65 ± 15†‡ |
| β2 integrin, extended with hybrid-domain swing-out (mAb MEM148) | ||||
| Stimulation | ||||
| Control | 15 ± 5 | 17 ± 5 | 20 ± 8 | 20 ± 5 |
| P-sel | 18 ± 5 | 20 ± 4 | 15 ± 5 | 22 ± 5 |
| IL-8 (0.1 nM) | 20 ± 5 | 16 ± 5 | 22 ± 6 | 20 ± 5 |
| IL-8 (1 nM) | 178 ± 20* | 88 ± 10† | 75 ± 10† | 25 ± 5†‡ |
| P-sel/IL-8 (0.1 nM) | 65 ± 8§ | 35 ± 6† | 40 ± 5† | 17 ± 10†‡ |
| P-sel/IL-8 (1 nM) | 220 ± 25§ | 145 ± 30† | 115 ± 20† | 35 ± 10†‡ |
The data represent the average mean fluorescence intensity (MFI) ± SD, n = 3. AS-605240 is a PI3Kγ inhibitor. GGTI-298 is a Rap1 inhibitor. P < .05 for each comparison.
P-sel, P-selectin.
Compared with control group.
Compared with non–inhibitor-treated group.
Compared with single inhibitor–treated group in each stimulation group.
Compared with any single stimulation group.