Ruths Leonie, Hengge Jana, Teixeira Graciosa Q, Haffner-Luntzer Melanie, Ignatius Anita, Riegger Jana
Division for Biochemistry of Joint and Connective Tissue Diseases, Department of Orthopedics, Ulm University Medical Center, Ulm, Germany.
Institute of Orthopedic Research and Biomechanics, Ulm University Medical Center, Ulm, Germany.
Front Immunol. 2025 Jan 14;15:1470907. doi: 10.3389/fimmu.2024.1470907. eCollection 2024.
The complement system is locally activated after joint injuries and leads to the deposition of the terminal complement complex (TCC). Sublytic TCC deposition is associated with phenotypical alterations of human articular chondrocytes (hAC) and enhanced release of inflammatory cytokines. Chronic inflammation is a known driver of chondrosenescence in osteoarthritis (OA). Therefore, we investigated whether TCC deposition contributes to stress-induced premature senescence (SIPS) during aging and after cartilage injury.
Femoral condyles of male 13-week-old and 72-week-old CD59-ko (higher TCC deposition), C6-deficient (insufficient TCC formation), and C57BL/6 (WT) mice were collected to assess age-related OA. Furthermore, macroscopically intact human and porcine cartilage explants were traumatized and cultured with/without 30% human serum (HS) to activate the complement system. Explants were additionally treated with clusterin (CLU, TCC inhibitor), N-acetylcysteine (NAC, antioxidant), Sarilumab (IL-6 receptor inhibitor), STAT3-IN-1 (STAT3 inhibitor), or IL-1 receptor antagonist (IL-1RA) in order to investigate the consequences of TCC deposition. Gene and protein expression of senescence-associated markers such as CDKN1A and CDKN2A was determined.
In the murine aging model, CD59-ko mice developed after 72 weeks more severe OA compared to C6-deficient and WT mice. mRNA analysis revealed that the expression of Cdkn1a, Cdkn2a, Tp53, Il1b, and Il6 was significantly increased in the cartilage of CD59-ko mice. In human cartilage, trauma and subsequent stimulation with HS increased mRNA levels of CDKN1A, CDKN2A, and IL6, while inhibition of TCC formation by CLU reduced the expression. Antioxidative therapy with NAC had no anti-senescent effect. In porcine tissue, HS exposure and trauma had additive effects on the number of CDKN2A-positive cells, while Sarilumab, STAT-IN-1, and IL-1RA reduced CDKN2A expression by trend.
Our results demonstrate that complement activation and consequent TCC deposition is associated with chondrosenescence in age-related and trauma-induced OA. We provided evidence that the SIPS-like phenotype is more likely induced by TCC-mediated cytokine release rather than oxidative stress. Overall, targeting TCC formation could be a future approach to attenuate OA progression.
关节损伤后补体系统在局部被激活，导致末端补体复合物（TCC）沉积。亚溶解剂量的TCC沉积与人类关节软骨细胞（hAC）的表型改变及炎性细胞因子释放增加有关。慢性炎症是骨关节炎（OA）中软骨衰老的已知驱动因素。因此，我们研究了TCC沉积是否在衰老过程以及软骨损伤后导致应激诱导的早衰（SIPS）。
收集13周龄和72周龄雄性CD59基因敲除小鼠（TCC沉积更高）、C6缺陷小鼠（TCC形成不足）和C57BL/6野生型（WT）小鼠的股骨髁，以评估与年龄相关的OA。此外，将肉眼观察完整的人及猪软骨外植体进行创伤处理，并在有/无30%人血清（HS）的条件下培养以激活补体系统。外植体还分别用簇集素（CLU，TCC抑制剂）、N-乙酰半胱氨酸（NAC，抗氧化剂）、沙瑞鲁单抗（IL-6受体抑制剂）、STAT3-IN-1（STAT3抑制剂）或IL-1受体拮抗剂（IL-1RA）处理，以研究TCC沉积的后果。测定衰老相关标志物如CDKN1A和CDKN2A的基因和蛋白表达。
在小鼠衰老模型中，与C6缺陷小鼠和WT小鼠相比，72周后CD59基因敲除小鼠发生了更严重的OA。mRNA分析显示，CD59基因敲除小鼠软骨中Cdkn1a、Cdkn2a、Tp53、Il1b和Il6的表达显著增加。在人软骨中，创伤及随后用HS刺激可增加CDKN1A、CDKN2A和IL6的mRNA水平，而CLU抑制TCC形成可降低其表达。NAC抗氧化治疗无抗衰老作用。在猪组织中，HS暴露和创伤对CDKN2A阳性细胞数量有累加效应，而沙瑞鲁单抗、STAT-IN-1和IL-1RA可使CDKN2A表达呈下降趋势。
我们的结果表明，补体激活及随之的TCC沉积与年龄相关和创伤诱导的OA中的软骨衰老有关。我们提供的证据表明，类似SIPS的表型更可能是由TCC介导的细胞因子释放而非氧化应激诱导的。总体而言，靶向TCC形成可能是未来减缓OA进展的一种方法。
