Fig 1: Effects of radiation and Ac-SDKP therapy on the mRNA and protein expression of tight junction molecules: Post-radiation rat heart tissue mRNA was analyzed by qPCR for measuring expression of common tight junction genes JAM-2 and claudin-1. Radiation exposure significantly reduced gene expression of JAM-2 (a) and Claudin-1 (e) in rat cardiac tissues (*, p = 0.003 compared to controls), which was restored by Ac-SDKP treatment (†, p = 0.03 compared to rad). N = 8–10 each group. b-d: Representative immunofluorescence images showing expression of intercellular cell adhesion proteins JAM-2 (b-d) and claudin-1 (f-h) into the rat coronary vessels. The immunofluorescence shows a well-defined TJP expression including, JAM-2 and claudin-1 (red stain). DAPI stained nuclei are shown in blue. c & g represent tissue sections from radiation exposed rats. Radiation exposed group showed patchy loss of TJP expression (arrows). Ac-SDKP reconstituted JAM-2 and claudin-1 expressions (d & h). Rad, radiation. Scale bar: 50 µm, magnification × 400
Fig 2: The importance of miR-330-3p/JAM2 axis for the mesenchymal identity of ovarian cancer in vivo.(A, D, G, J, and M) A total of 5 × 106 ID8 cells with respective treatment were subcutaneously injected into C57 mice together with plasma cell–derived exosomes and control (n = 6 for each group). Growth curve was plotted. (B, E, H, K, and N) Tumor weight and volume of ID8 cells in each group were measured at indicated time. (C, F, I, L, and O) In vivo bioluminescent imaging of tumor growth in each group was performed in mice 30 days after injection (n = 3 to 5 for each group). Data are shown as mean ± SEM. In (A), (D), (G), (J), and (M), P value out of two-way repeated measures analysis of variance (ANOVA). Otherwise, statistical significance was determined by a two-tailed, unpaired Student’s t test. *P < 0.05; **P < 0.01. ***P < 0.001. DMSO, dimethyl sulfoxide; sh, small hairpin.
Fig 3: miR-330-3p targets junctional adhesion molecule 2 for the maintenance of mesenchymal identity of ovarian cancer.(A) Venn diagram showing the most possible up-regulated genes targeted by plasma cell exosome–containing mir-330-3p. (B) Univariate regression analysis of the six overlapped target genes associated with ovarian cancer patient survival (Bonome dataset, n = 182). (C) mRNA level of JAM2 in COV318 and OVCAR-3 cells with respective treatment (n = 3 to 4 for each group). (D) Western blotting analysis of JAM2 protein levels in COV318 and OVCAR-3 cells with respective treatment (n = 3 for each group). (E) mRNA levels of JAM2 and EMT markers in miR-330-3p mimic–transfected or control COV318 and OVCAR-3 cells (n = 3 to 4 for each group). (F) Western blotting analysis of JAM2 and EMT markers in miR-330-3p mimic–transfected or control COV318 and OVCAR-3 cells (n = 3 for each group). (G) Wound healing analysis to assess the migration ability of COV318 and OVCAR-3 cells with respective treatment (n = 3 to 5 for each group). (H) Western blotting analysis of JAM2 and EMT markers in COV318 and OVCAR-3 cells with respective treatment (n = 3 for each group). si, small interfering. (I) mRNA levels of JAM2 in COV318 and OVCAR-3 cells with respective treatment (n = 3 to 4 for each group). (J) Transwell chamber analysis to assess the migration ability of COV318 and OVCAR-3 cells with respective treatment (n = 3 to 5 for each group). (K) Wound healing analysis to assess the migration ability of COV318 and OVCAR-3 cells with respective treatment (n = 3 to 5 for each group). Data are shown as mean ± SEM. All statistical significance was determined by a two-tailed, unpaired Student’s t test. *P < 0.05; **P < 0.01; ***P < 0.001.
Fig 4: Clinical significance of miR-330-3p/JAM2 axis in the cross-talk between plasma cells and ovarian cancer cells.(A) Unsupervised classification of plasma gene signature shows the optimal classification using two clusters (left), as supported by the gap statistic (right). (B) Heatmap for unsupervised hierarchical clustering of the Tothill dataset using plasma cell gene signature as classifiers. (C) Survival difference between the plasma cell–high subgroup and the plasma cell–low subgroup in the Tothill dataset. OS, overall survival. (D) Survival difference between the plasma cell–high subgroup and the plasma cell–low subgroup in all patients and the mesenchymal-subtype patients in the Mateescu dataset. (E) Differential progression-free survival (PFS) between plasma cell exosome–specific miRNA signature–high group and plasma cell exosome–specific miRNA signature–low group of patients with ovarian cancer in OC133 and OC179 datasets. (F) Differential overall survival and progression survival between miR-330-3p–high group and miR-330-3p–low group of patients with ovarian cancer in the OC179 dataset. (G) Expression levels of miR-330-3p in patients with ovarian cancer with relapse and without relapse. (H) Expression levels of miR-330-3p in patients with ovarian cancer with distinct differentiation grades. (I) Differential overall survival between JAM2-high group and JAM2-low group of patients with ovarian cancer in the Tothill, Bonome, and TCGA datasets. Data are shown as mean ± SEM. In (C), (D), (E), (F), and (I), survival difference was calculated by log-rank test; in (G) and (H), P value was calculated using the two-tailed Student’s t test. **P < 0.01.
Supplier Page from Abcam for Anti-Junctional Adhesion Molecule 2/JAM-B antibody