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T. Yahata et al.
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Figure 5. iPAI-1 blockade increases membrane type-1 metalloprotease- dependent cellular motility. (A) Schema for experiments. (B) Representative flow cytometric profiles and mean fluores- cent intensity (MFI) (n=6-9) for iPAI-1, MT1-MMP, CD44, and VLA-4 expres- sions in freshly isolated bone marrow (BM) BCR/ABL-GFP+ LSK cells of saline or PAI-1 inhibitor treated mice. (C) Percentages of migrated BCR/ABL- GFP+Lin−c-kit+ immature chronic myeloid leukemia (CML) cells in trans-Matrigel
B migration assay (n=6 each). Data repre- sent means ± standard deviation. Statistical significance was determined by Mann-Whitney unpaired t-test. P<0.001, by a Kruskal-Wallis test. (D) Representative pictures of the BM cavi- ty of vehicle- or TM5614-treated mice. BM sections were stained with anti- transforming growth factor-β (TGF-β) (purple), anti-c-kit (red) and anti-lineage markers (white) antibodies. Red arrow- heads indicate TGF-β-expressing nich- es. Blue arrow heads indicate BCR/ABL- GFP+Lin–c-kit+ CML cells. Bars represent 100 mm. Graph indicates percentages of TGF-β-expressing niches closely con- tact to immature CML cells. More than 50 in random fields on a slide were counted for two independent experi- ments (n=4 each). Each dot represents % of contact cells in the one field. Statistical significance was determined by Mann-Whitney unpaired t-test. P<0.001, by a Kruskal-Wallis test. BCR: breakpoint cluster region; ABL: Abelson kinase: GFP: green fluorescent protein; LSK: Lineage (Lin)Sca1c-Kit L; MT1- MNP: membrane type-1 metallopro-
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tease; Ab: antibody.
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haematologica | 2021; 106(2)