Analysis of stromal cell subsets in the adventitia form non-aneurysmal and aneurysmal aortas.

<p>(<b>A</b>) The gating strategy to analyze all nucleated cells (Vybrant⁺) and particularly stromal cell subsets from an aneurysmal artery by 10-color flow cytometry is shown. This strategy allows the simultaneous definition of residual aSMA^hi smooth muscle cells (SMC), CD44⁺gp38^lo/− leukocytes [macrophages (MΦ) and lymphocytes (Lympho) defined on the forward (FSC-A) and side (SSC-A) scatter], among non-leukocyte cells, CD21⁺CD44⁺ “LTo-like” cells, CD31⁺CD44⁻pg38⁻ blood endothelial cells (BEC), CD31⁺gp38⁺CD21^lo lymphatic endothelial cells (LEC), CD31⁺gp38⁺CD21^hi follicular dendritic cells (FDC), CD31⁻CD44⁺ fibroblastic reticular cells (FRC), and CD31⁻CD44⁻ double negative cells (DN). The lower left dot plot (CD31 vs pg38) shows the overlay of all stromal cell subsets excluding SMC. (<b>B</b>) Flow cytometric dot plots showing the expression of Vybrant (Vybrant^hi cells being undergoing division) and gp38 (upper panels), VCAM1 and MadCAM1 (middle panels) or ICAM1 and MadCAM1 (lower panels) for cell subsets defined in (<b>A</b>). The percentage of cells in each quadrant is indicated. Vybrant-positive events are highlighted and displayed as thick black dots in the middle and lower VCAM1/ICAM1 vs MadCAM1 dot plots. (<b>C</b>) Comparison of the percentage among total live cells of cell subsets defined in (<b>A</b>) in the adventitia from two non-aneurysmal (white filled symbols) and three aneurysmal (white filled symbols) abdominal aortas.