Figure 3: The migration of integrin alpha-6-positive germ cell precursors towards sphingosine-1-phosphate is mediated by the G-protein–coupled receptor S1PR1 and integrin alpha-dependent binding to laminin.

(a) Migration assay of ALDH-positive cells in response to different concentrations of S1P, as indicated. No stimulant was added to control wells. Sorted cells were added to the upper chamber of a Transwell system coated with laminin, and after 2 h, migrated cells in the lower chamber were counted. Data are expressed as fold changes of numbers of migrated cells, normalized to unstimulated controls (n=6). Statistical analysis was performed using Student’s t-test. **P≤0.05 (medium significance compared with control). (b) Migration assay of ALDH-positive/Integrin-alpha-6-positive cells in the presence of 2 μm S1P with or without inhibitors of S1PR signalling. S1PR1 antagonist W146 (10 μM), PI3K-inhibitor Wortmannin (0.1 μM) or Rac1-inhibitor NSC (100 μM) were added as indicated. Data are expressed as fold changes of numbers of migrated cells, normalized to unstimulated controls. (c) Migration assay of ALDH-positive/integrin alpha-6-positive cells on uncoated plastic or laminin-coated Transwells with or without 2 μm S1P. Anti-integrin alpha-6 (5 μg ml⁻¹) or isotype control antibody was added as indicated. Data are expressed as fold changes of numbers of migrated cells, normalized to unstimulated controls on laminin. Error bars represent the standard deviation for each average (n=6) Statistical analysis was performed using Student’s t-test. **P≤0.05 (medium significance).