Journal: EBioMedicine

Article Title: Endothelial damage and a thin intercellular fibrin network promote haemorrhage in acute promyelocytic leukaemia

doi: 10.1016/j.ebiom.2020.102992

Figure Lengend Snippet: APL cells adhere to ECs, causing impaired barrier function in vitro and in vivo. (A, B) The effect of APL/NB4 cells on ICAM-1 and VCAM-1 expression in HUVECs. Western blots were imaged, and the optical density was calculated using ImageJ. (C) ECs were stained with phalloidin, and RBCs were labeled with live-cell dye. Gap formation in the damaged endothelium permits RBC deposition (arrows). (D) The albumin permeability of the experimental monolayers treated with an adherence receptor antibody. (E) The permeability to RBCs upon treatment with an adherence receptor antibody. (F) The levels of sVCAM-1, sICAM-1, sE-selectin, and sTM in serum samples from the bleeding patient (open circles) and non-bleeding patient (closed circles) groups. (G) Western blotting with an anti-pVE-cadherin antibody, an anti-MLCK, and an anti-ROCK antibody for ECs treated with leukemic cells or supernatant alone. (H) Quantitation of the relative protein expression level in (G) by ImageJ (Gapdh was used as the loading control). (I) Pretreatment with MLCK inhibitor ML-7(10 −4 mol/L) or/and with ROCK inhibitor H1152 (2.5 μmol) for 1 h, then incubated with APL cells for 16 h. Permeability of treated endothelium to RBCs. (J) Expression of pVE-cadherin and MLCK in ECs treated with anti-ICAM-1 and anti-VCAM-1 antibodies, respectively, was assayed by western blotting and compared with that in the control and APL samples. (K) Quantitation of the relative protein expression level in (J) by ImageJ. The inset bar represents 20 µm in panel F. AC+APL: Asiatic acid + APL cells. The graph is presented as the mean ± SD of at least five experiments. * P <0.05 and ** P <0.01.

Article Snippet: ESM1-endocan, human soluble VCAM-1 (sVCAM-1), human soluble thrombomodulin (sTM) and human soluble ICAM-1 (sICAM-1) ELISA kits were from Aviscera Bioscience Inc. (Santa Clara, CA).

Techniques: In Vitro, In Vivo, Expressing, Western Blot, Staining, Labeling, Permeability, Quantitation Assay, Incubation

Journal: EBioMedicine

Article Title: Endothelial damage and a thin intercellular fibrin network promote haemorrhage in acute promyelocytic leukaemia

doi: 10.1016/j.ebiom.2020.102992

Figure Lengend Snippet: ATRA protects HUVEC barrier function. (A) A laser confocal microscope was used to image the changes in the junction protein VE-cadherin (green) and DAPI (blue) staining. The inset bar represents 10 µm. (B) The permeability of NB4-treated HUVECs incubated with or without ATRA (1 µmol/L) or ATO (5 µmol/L) was tested as previously described. (C) The number of leaked RBCs was measured as previously described. (D) The distribution of intercellular gap size was improved by ATRA treatment of the target HUVECs; n = 200 gaps from 10 glass coverslips. (E, F) Effect of ATRA and ATO on ICAM-1 and VCAM-1 concentrations. (G) Schematic illustration of the effect of ATRA and ATO on NB4-treated HUVECs. ATRA protects NB4-treated HUVECs, whereas ATO worsens the damage. * P <0.05 and ** P <0.01. The results are displayed as the mean ± SD from at least 6 experiments. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: ESM1-endocan, human soluble VCAM-1 (sVCAM-1), human soluble thrombomodulin (sTM) and human soluble ICAM-1 (sICAM-1) ELISA kits were from Aviscera Bioscience Inc. (Santa Clara, CA).

Techniques: Microscopy, Staining, Permeability, Incubation

Journal: EBioMedicine

Article Title: Endothelial damage and a thin intercellular fibrin network promote haemorrhage in acute promyelocytic leukaemia

doi: 10.1016/j.ebiom.2020.102992

Figure Lengend Snippet: Asiatic acid alleviates bleeding in the APL model. (A) The outline depicts the transplantation experiments. Dorplica experiments mean the back skin of the mice was cut off to observe subcutaneous bleeding. (B, C) The levels of sICAM-1 and sVCAM-1 were measured in mouse serum samples collected from the caudal vein. (D) Representative images of prepared back skins of the APL model mice with different treatments and 24 h of UV light exposure. (E) Analysis of the hemoglobin concentration in the bleeding spots. (F) The expression of PML-RARA protein in the areas of bleeding. (G) The bleeding time was tested as previously described. (H) Wound closure of APL model mice one week after injury. The white columns represent the data of mice that have not received leukemic cells, the purple columns represent the data of mice that received leukemic cells. * P <0.05, ** P <0.01, ns, nonsignificant. The results are shown as the mean ± SD of at least 3 experiments.

Article Snippet: ESM1-endocan, human soluble VCAM-1 (sVCAM-1), human soluble thrombomodulin (sTM) and human soluble ICAM-1 (sICAM-1) ELISA kits were from Aviscera Bioscience Inc. (Santa Clara, CA).

Techniques: Transplantation Assay, Concentration Assay, Expressing

Journal: EBioMedicine

Article Title: Endothelial damage and a thin intercellular fibrin network promote haemorrhage in acute promyelocytic leukaemia

doi: 10.1016/j.ebiom.2020.102992

Figure Lengend Snippet: Intercellular fibrin networks prevent erythrocyte leakage. VEGF, inflammatory cytokines, and APL blasts interact with ECs. Cellular factors activate ROCK signaling, whereas APL cells adhere to the adherence receptors ICAM-1 and VCAM-1 and trigger the MLCK pathway, resulting in stress fiber formation, EC retraction, and junction protein dissociation. The gaps among dissociated junctions are large enough to permit RBC transmigration. PS exposure on the activated ECs provides a catalytic surface for FV and FVIII, resulting in coagulation pathway activation and fibrin formation among the openings. Intercellular fibrin works to repair the integrity of the endothelium. Plasmin activated through the overexpression of t-PA from APL blasts acts to weaken the fibrin network, allowing RBCs to leak into the extravascular space.

Article Snippet: ESM1-endocan, human soluble VCAM-1 (sVCAM-1), human soluble thrombomodulin (sTM) and human soluble ICAM-1 (sICAM-1) ELISA kits were from Aviscera Bioscience Inc. (Santa Clara, CA).

Techniques: Transmigration Assay, Coagulation, Activation Assay, Over Expression