Department of Vascular and Endovascular Surgery, University Hospital Heidelberg, University of Heidelberg, Im Neuenheimer Feld 110, 69120, Heidelberg, Germany.
Present Address: Department of Vascular Surgery, Luzerner Kantonsspital, Spitalstrasse, 6000, Luzern 16, Switzerland.
Mol Med. 2020 Sep 15;26(1):87. doi: 10.1186/s10020-020-00212-z.
Phenotypic transformation of vascular smooth muscle cells is a key element in vascular remodeling and aortic aneurysm growth. Previously, deletion of several inflammasome components decreased formation of aortic aneurysm (AA) in the Angiotensin II (AngII) -induced mouse model. We hypothesized that the inflammasome sensor Absent in melanoma 2 (Aim2) might affect the phenotype of vascular smooth muscle cells (VSMC), thereby reducing AA formation.
Aim2-/- mice and wild-type (WT) C57Bl/6 J mice were used as an animal model. VSMC were isolated from 6 months old mice and grown in vitro. Young (passage 3-5) and senescent (passage 7-12) cells were analyzed in vitro for calcification in mineralization medium by Alizarin Red S staining. Expression of calcification and inflammatory markers were studied by real-time RT-PCR and Western blotting, release of cytokines was determined by ELISA. To induce AA, osmotic mini-pumps loaded with AngII (1500 ng/kg bodyweight/min) were implanted for 28 days in male mice at 6 months of age.
Compared with VSMC from WT mice, VSMC isolated from Aim2-/- mice were larger, less viable, and underwent stronger calcification in mineralization medium, along with induction of Bmp4 and repression of Tnfsf11/Rankl gene expression. In addition, Aim2 deficiency was associated with reduced inflammasome gene expression and release of Interleukin-6. Using the mouse model of AngII induced AA, Aim2 deficiency reduced AA incidence to 48.4% (15/31) in Aim2-/- mice versus 76.5% (13/17) in WT mice. In contrast to Aim2-/- mice, AA from WT mice expressed significantly increased levels of alpha-smooth muscle actin/Acta2, indicating tissue remodeling. Reduced cell proliferation in Aim2-/- mice was indicated by significantly increased p16ink4a/Cdkn2a expression in untreated and AngII-infused aortas, and by significantly lower amounts of proliferating (Ki67 positive) VSMC in AngII-infused Aim2-/- mice.
Our results suggest a role for Aim2 in regulating VSMC proliferation and transition to an osteoblast-like or osteoclast-like phenotype, thereby modulating the response of VSMC in aortic remodeling and AA formation.
血管平滑肌细胞的表型转化是血管重塑和主动脉瘤生长的关键因素。先前的研究表明，几种炎性小体成分的缺失可减少血管紧张素 II（AngII）诱导的小鼠模型中主动脉瘤（AA）的形成。我们假设无黑色素瘤 2（Aim2）炎性小体传感器可能会影响血管平滑肌细胞（VSMC）的表型，从而减少 AA 的形成。
使用 Aim2-/- 小鼠和野生型（WT）C57Bl/6 J 小鼠作为动物模型。从 6 个月大的小鼠中分离 VSMC 并在体外培养。通过茜素红 S 染色在矿化培养基中分析年轻（传代 3-5 代）和衰老（传代 7-12 代）细胞的钙化情况。通过实时 RT-PCR 和 Western 印迹研究钙化和炎症标志物的表达，通过 ELISA 测定细胞因子的释放。为了诱导 AA，将装载 AngII（1500ng/kg 体重/分钟）的渗透微型泵植入 6 个月大雄性小鼠体内 28 天。
与 WT 小鼠的 VSMC 相比，Aim2-/- 小鼠的 VSMC 体积更大、活力更低，在矿化培养基中钙化更强，同时诱导 Bmp4 基因表达和抑制 Tnfsf11/Rankl 基因表达。此外，Aim2 缺乏与炎性小体基因表达和白细胞介素-6 的释放减少有关。在 AngII 诱导的 AA 小鼠模型中，与 WT 小鼠相比，Aim2-/- 小鼠的 AA 发生率降低至 48.4%（15/31），而 WT 小鼠的发生率为 76.5%（13/17）。与 Aim2-/- 小鼠相比，WT 小鼠的 AA 表达明显增加的 alpha-平滑肌肌动蛋白/Acta2，表明组织重塑。在未处理和 AngII 输注的主动脉中，p16ink4a/Cdkn2a 表达的显著增加表明 Aim2-/- 小鼠的细胞增殖减少，在 AngII 输注的 Aim2-/- 小鼠中增殖（Ki67 阳性）的 VSMC 数量也显著减少。
我们的结果表明 Aim2 可调节 VSMC 的增殖和向成骨样或破骨样表型的转化，从而调节 VSMC 在主动脉重塑和 AA 形成中的反应。
