Calcium- and integrin-binding protein 1 regulates megakaryocyte ploidy, adhesion, and migration

Supplemental materials for: Kostyak et al

Files in this Data Supplement:

Figure S1. Cib1^−∕− mice have more leukocytes than WT controls, while megakaryocyte morphology is unchanged (JPG, 463 KB) -
(A) Wright-Giemsa-stained blood smears from WT and Cib1^−∕− mice. Leukocytes are indicated by horizontal arrows. Scale bars are 100 µm. Images were captured using a Zeiss Axiophot at 100× magnification. (B) Wright-Giemsa-stained bone marrow touch preps. Scale bars are 100 µm. Images were captured using a Zeiss Axiophot at 200× magnification.
Figure S2. Platelet production and clearance are unaltered upon Cib1 deletion (JPG, 585 KB) -
(A) Representative histograms of platelets from WT and Cib1^−∕− mice labeled with sulfo-NHS-biotin and thiazole orange 1, 24, 48, and 72 hours after injection. Reticulated platelets appear in Q1 and Q2, while biotinylated platelets appear in Q2 and Q4. (B) Representative histograms of thiazole orange (TO) labeled platelets from WT and Cib1^−∕− mice.
Figure S3. cMpl expression on WT and Cib1^−∕− mouse platelets and BM cells (JPG, 332 KB) -
Western blot of lysates of WT and Cib1^−∕− mouse platelets (A) and BM cells (B) probed for cMpl expression using anti-cMpl. Blot was reprobed with anti-tubulin to ensure equal loading. Flow cytometric analysis of WT and Cib1^−∕− mouse platelets (C) and BM cells (D) for surface expression of cMpl. Quantitation of the mean fluorescence intensities is shown underneath the corresponding histograms (n=4).
Figure S4. Highly pure polyploid cells were obtained via a discontinuous BSA gradient (JPG, 498 KB) -
(A) Microscopic images of the MK and mononuclear cell fractions of BM cells separated by BSA-gradient at the same magnifications. (B) Flow cytometry histograms showing the ploidy distribution of BM cell fractions obtained following BSA gradient separation compared to mononuclear cells from the same separation.