Figure 3
Tryptase, chymase and carboxypeptidase expression of 8 wk HuMC cultured in rhSCF alone and with 100 ng/ml LPS; Live gate analysis of CD117low (R1 region) and CD117high (R2 region) cells. A) Left-sided graph – HuMC numbers expressing tryptase, chymase, and carboxypeptidase following culture for 72 hr in 10 ng/ml rhSCF alone, and with 100 ng/ml LPS. For 10 ng/ml rhSCF alone, the mean ± SEM of percentages of protease positive HuMC are: tryptase 88.51 ± 5.30%, chymase 91.34 ± 1.62%, carboxypeptidase 91.53 ± 1.20%, tryptase/chymase 83.75 ± 4.61% and tryptase/carboxypeptidase 87.32 ± 4.90%. For 10 ng/ml rhSCF and LPS, the mean ± SEM of percentages of protease positive HuMC cells are: tryptase 92.50 ± 2.32%, chymase 96.30 ± 0.75%, carboxypeptidase 94.43 ± 4.21%, tryptase/chymase 92.33 ± 3.85%, and tryptase/carboxypeptidase 90.50 ± 2.30%. Right-sided graph – 100 ng/ml rhSCF alone, and with 100 ng/ml LPS. For 100 ng/ml rhSCF alone, the mean ± SEM of percentages of protease positive HuMC are: tryptase 94.56 ± 2.30%, chymase 93.30 ± 1.38%, carboxypeptidase 88.50 ± 3.21%, tryptase/chymase 88.70 ± 5.41%, and tryptase/carboxypeptidase 88.55 ± 3.83%. For 100 ng/ml rhSCF and LPS, the mean ± SEM of percentages of protease positive HuMC are: tryptase 92.57 ± 3.30%, chymase 95.33 ± 0.85%, carboxypeptidase 89.81 ± 3.90%, tryptase/chymase 93.63 ± 3.25%, and tryptase/carboxypeptidase 89.50 ± 5.42%. Results are shown as the mean ± SEM of 3 separate experiments performed in duplicate; B) Representative dot plots and histograms of CD117low (R1 region) and CD117high (R2 region) cells analyzed for FcεRI, CD117, tryptase, chymase and carboxypeptidase expression during culture in 10 ng/ml rhSCF alone and with 100 ng/ml LPS (as seen in Figure A, left). In the histograms, horizontal axis represents PE or PE5.5-conjugated markers analyzed; vertical axis represents total cell numbers analyzed. Cultures with 10 ng/ml rhSCF alone are shown by the shaded areas. Representative dot plots and histograms are shown of n = 3 separate experiments performed in duplicate.