Rucci Nadia, Capulli Mattia, Piperni Sara Gemini, Cappariello Alfredo, Lau Patrick, Frings-Meuthen Petra, Heer Martina, Teti Anna
Department of Biotechnological and Applied Clinical Sciences, University of L'Aquila, L'Aquila, Italy.
J Bone Miner Res. 2015 Feb;30(2):357-68. doi: 10.1002/jbmr.2341.
Mechanical loading represents a crucial factor in the regulation of skeletal homeostasis. Its reduction causes loss of bone mass, eventually leading to osteoporosis. In a previous global transcriptome analysis performed in mouse calvarial osteoblasts subjected to simulated microgravity, the most upregulated gene compared to unit gravity condition was Lcn2, encoding the adipokine Lipocalin 2 (LCN2), whose function in bone metabolism is poorly known. To investigate the mechanoresponding properties of LCN2, we evaluated LCN2 levels in sera of healthy volunteers subjected to bed rest, and found a significant time-dependent increase of this adipokine compared to time 0. We then evaluated the in vivo LCN2 regulation in mice subjected to experimentally-induced mechanical unloading by (1) tail suspension, (2) muscle paralysis by botulin toxin A (Botox), or (3) genetically-induced muscular dystrophy (MDX mice), and observed that Lcn2 expression was upregulated in the long bones of all of them, whereas physical exercise counteracted this increase. Mechanistically, in primary osteoblasts transfected with LCN2-expression-vector (OBs-Lcn2) we observed that Runx2 and its downstream genes, Osterix and Alp, were transcriptionally downregulated, and alkaline phosphatase (ALP) activity was less prominent versus empty-vector transduced osteoblasts (OBs-empty). OBs-Lcn2 also exhibited an increase of the Rankl/Opg ratio and IL-6 mRNA, suggesting that LCN2 could link poor differentiation of osteoblasts to enhanced osteoclast stimulation. In fact, incubation of purified mouse bone marrow mononuclear cells with conditioned media from OBs-Lcn2 cultures, or their coculture with OBs-Lcn2, improved osteoclastogenesis compared to OBs-empty, whereas treatment with recombinant LCN2 had no effect. In conclusion, our data indicate that LCN2 is a novel osteoblast mechanoresponding gene and that its regulation could be central to the pathological response of the bone tissue to low mechanical forces.
机械负荷是骨骼内环境稳态调节的关键因素。负荷减少会导致骨量流失，最终引发骨质疏松症。在之前对模拟微重力条件下的小鼠颅骨成骨细胞进行的全基因组转录组分析中，与单位重力条件相比，上调最显著的基因是Lcn2，它编码脂肪因子脂质运载蛋白2（LCN2），其在骨代谢中的功能尚不清楚。为了研究LCN2的机械响应特性，我们评估了健康志愿者卧床休息时血清中的LCN2水平，发现与时间0相比，这种脂肪因子呈显著的时间依赖性增加。然后，我们通过以下方式评估了实验性诱导机械卸载小鼠体内LCN2的调节情况：（1）尾部悬吊，（2）肉毒杆菌毒素A（肉毒素）导致的肌肉麻痹，或（3）基因诱导的肌肉萎缩症（MDX小鼠），并观察到所有小鼠的长骨中Lcn2表达均上调，而体育锻炼可抵消这种增加。从机制上讲，在用LCN2表达载体转染的原代成骨细胞（OBs-Lcn2）中，我们观察到Runx2及其下游基因Osterix和Alp的转录下调，与空载体转导的成骨细胞（OBs-empty）相比，碱性磷酸酶（ALP）活性不那么显著。OBs-Lcn2的Rankl/Opg比值和IL-6 mRNA也有所增加，这表明LCN2可能将成骨细胞的分化不良与破骨细胞刺激增强联系起来。事实上，用OBs-Lcn2培养物的条件培养基孵育纯化的小鼠骨髓单核细胞，或将它们与OBs-Lcn2共培养，与OBs-empty相比，破骨细胞生成有所改善，而用重组LCN2处理则没有效果。总之，我们的数据表明LCN2是一种新的成骨细胞机械响应基因，其调节可能是骨组织对低机械力病理反应的核心。
