Biochemistry & Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK, USA.
Nutritional Sciences, University of Oklahoma Health Sciences Center, Oklahoma City, OK, USA.
Geroscience. 2023 Dec;45(6):3241-3256. doi: 10.1007/s11357-023-00955-7. Epub 2023 Oct 4.
To study the impact of necroptosis-induced chronic inflammation on age-related diseases and aging, two knockin mouse models (Ripk3-KI and Mlkl-KI) were generated that overexpress two genes involved in necroptosis (Ripk3 or Mlkl) when crossed to Cre transgenic mice. Crossing Ripk3-KI or Mlkl-KI mice to albumin-Cre transgenic mice produced hepatocyte specific hRipk3-KI or hMlkl-KI mice, which express the two transgenes only in the liver. Ripk3 and Mlkl proteins were overexpressed 10- and fourfold, respectively, in the livers of the hRipk3-KI or hMlkl-KI mice. Treating young (2-month) hRipk3-KI or hMlkl-KI mice with carbon tetrachloride (CCl), a chemical inducer of oxidative stress, resulted in increased necroptosis (Mlkl-oligomers) and inflammation in the liver compared to control mice receiving CCl. Mlkl-oligomerization also was significantly increased in old (18-month) hRipk3-KI and hMlkl-KI mice compared to old control (Cre negative, Ripk3-KI and Mlkl-KI) mice. The increase in necroptosis was associated with an increase in inflammation, e.g., inflammatory cytokines (TNFα, IL-6) and macrophage markers (F4/80, CD68). Importantly, steatosis (triglycerides) and fibrosis (e.g., picrosirius red staining, hydroxyproline levels, and transcripts for TGFβ, Col1α1, and Col3α1) that increase with age were significantly higher in the livers of the old hRipk3-KI or hMlkl-KI mice compared to old control mice. In addition, markers of cellular senescence were significantly increased in the livers of the old hRipk3-KI and hMlkl-KI mice. Thus, the first mouse models have been developed that allow researchers to study the impact of inducing necroptosis in specific cells/tissues on chronic inflammation in aging and age-related diseases.
为了研究坏死性凋亡诱导的慢性炎症对与年龄相关的疾病和衰老的影响，生成了两种过表达两种参与坏死性凋亡的基因（Ripk3 或 Mlkl）的敲入小鼠模型（Ripk3-KI 和 Mlkl-KI），当与 Cre 转基因小鼠杂交时。将 Ripk3-KI 或 Mlkl-KI 小鼠与白蛋白-Cre 转基因小鼠杂交，产生了肝细胞特异性 hRipk3-KI 或 hMlkl-KI 小鼠，这两种转基因只在肝脏中表达。hRipk3-KI 或 hMlkl-KI 小鼠肝脏中 Ripk3 和 Mlkl 蛋白的表达分别增加了 10 倍和 4 倍。用四氯化碳（CCl）处理年轻（2 个月）的 hRipk3-KI 或 hMlkl-KI 小鼠，与接受 CCl 的对照小鼠相比，肝脏中坏死性凋亡（Mlkl-寡聚物）和炎症增加。与年老（18 个月）的对照（Cre 阴性、Ripk3-KI 和 Mlkl-KI）小鼠相比，年老的 hRipk3-KI 和 hMlkl-KI 小鼠中 Mlkl-寡聚化也显著增加。坏死性凋亡的增加与炎症的增加有关，例如，炎症细胞因子（TNFα、IL-6）和巨噬细胞标志物（F4/80、CD68）。重要的是，与年龄相关的肝脂肪变性（甘油三酯）和纤维化（例如，天狼星红染色、羟脯氨酸水平以及 TGFβ、Col1α1 和 Col3α1 的转录物）在年老的 hRipk3-KI 或 hMlkl-KI 小鼠的肝脏中显著高于年老的对照小鼠。此外，年老的 hRipk3-KI 和 hMlkl-KI 小鼠肝脏中的细胞衰老标志物显著增加。因此，已经开发出第一种小鼠模型，允许研究人员研究在特定细胞/组织中诱导坏死性凋亡对衰老和与年龄相关疾病的慢性炎症的影响。
