Table I.
Effect of cholesterol depletion on mast cell sterol content and viability
| Time of cholesterol depletion (h) |
DHCR KO
|
WT
|
||||
|---|---|---|---|---|---|---|
| Cholesterol (μg/mg of protein) |
DHC (μg/mg of protein) |
Viability (%) |
Cholesterol (μg/mg of protein) |
Desmosterol (μg/mg of protein) |
Viability (%) |
|
| 0 | 3.6 ± 0.3 (10) | 0.3 ± 0.1 (10) | 96.4 ± 0.3 (6) | 3.9 ± 0.3 (10) | nd | 96.9 ± 0.3 (6) |
| 48 | 2.6 ± 0.2 (3) | 0.5 ± 0.1 (3) | 95.1 ± 0.1 (3) | 2.7 ± 0.2 (3) | 0.6 ± 0.3 (3) | 96.8 ± 0.2 (3) |
| 72 | 2.2 ± 0.3 (10) | 0.9 ± 0.4 (10) | 93.6 ± 0.3 (6) | 2.9 ± 0.3 (10) | 1.3 ± 0.5 (3) | 96.9 ± 0.1 (6) |
| 96 | 1.9 ± 0.1 (3) | 1.1 ± 0.2 (3) | 89.7 ± 0.6 (3) | 3.3 ± 0.1 (3) | nd | 95.2 ± 0.3 (3) |
| 120 | 2.4 ± 0.2 (4) | 1.1 ± 0.1 (4) | 89.3 ± 0.8 (3) | 3.4 ± 0.1 (5) | nd | 95.0 ± 0.2 (3) |
Cells were depleted of cholesterol by incubation for the indicated times in medium containing LPDS. Cell viability was monitored by trypan blue exclusion and propidium iodide staining. Content of cholesterol, DHC, and desmosterol was determined by GC/mass spectrometry analysis of extracted sterols. DHC was also measured in cholesterol-depleted WT cells but was nondetectable (sensitivity of assay is pg/mg of protein, n = 5). Total sterols did not differ significantly among genotypes before or after cholesterol depletion (0 vs. 72 h). Data shown are the mean ± SEM for replicate numbers as indicated in parenthesis. nd, not detected.