p75NTR敲除对小鼠股骨矿化发育的抑制作用

Knocking out p75 neurotrophin receptor suppresses mineralization in the femurs of mice

目的对比野生型和p75 neurotrophin receptor(p75NTR)敲除小鼠股骨骨质的改变,探讨p75NTR敲除对小鼠股骨矿化发育的影响。方法取同窝p75NTR^(+/+)(野生型)和p75NTR^(-/-)(敲除型)雄性小鼠,出生后8周龄时行股骨Micro-CT扫描,检测股骨松质骨的骨量(BV)、骨体积分数(BV/TV)、骨小梁数(Tb.N)、骨小梁厚度(Tb.Th)、骨表面积/骨量(BS/BV)、骨小梁分离度(Tb.Sp)和皮质骨的骨量(Ct.BV)、骨皮质厚度(Ct.Tb Th)、骨皮质的骨表面积/骨量(Ct.BS/BV);出生后40 d分别取同窝两种基因型雄性小鼠行钙黄绿素荧光双标实验,以检测股骨的矿化沉积速率;提取4周龄大小鼠股骨总RNA及蛋白质,RT-PCR和Western blot检测ALP、Runx2的表达。结果 Micro-CT结果显示:p75NTR^(-/-)小鼠股骨的BV、BV/TV、Tb.N、Tb.Th、Ct.BV、Ct.Tb Th明显低于同窝p75NTR~(^(+/+))小鼠(P<0.05),同时,p75NTR^(-/-)小鼠的BS/BV、Tb.Sp、Ct.BS/BV明显高于p75NTR^(+/+)小鼠(P<0.01);钙黄绿素荧光双标检测结果显示:p75NTR^(-/-)小鼠股骨的矿化速率明显低于同窝p75NTR^(+/+)小鼠(P<0.01);RT-PCR及Western blot结果显示:p75NTR~(^(-/-))小鼠的ALP、Runx2表达均明显低于p75NTR~(^(+/+))小鼠(P<0.01)。结论 p75NTR敲除对小鼠股骨的矿化发育有一定的抑制作用。

Objective To compare the quantitative parameters of the femoral bone between wild-type mice and p75 neurotrophin receptor knockout （p75NTR^-/-） mice and explore the impact of p75NTR knockout on mineralization and development of the femoral bone in mice. Mothods The left femurs of p75NTR^-/- and wild-type （p75NTR^＋/＋） male mice （8 weeks old ） were scanned with micro-CT for measurement of the cancellous bone volume （BV）, bone volume fraction （BV/TV）, number of bone trabeeula （ Tb. N ） , trabecular thickness （ Tb. Th ） , bone surface area/bone volume ratio （ BS/BV ） , trabecular separation （Tb. Sp）, cortical bone volume （Ct. BV）, cortical bone thickness （Ct. Tb. Th）, and cortical bone surface area./ bone volume ratio （Ct. BS/BV ）. The mineral apposition rate （MAR） of the femurs of the p75NTR^ -/- mice and their wild-type littermates was analyzed on day 40 after birth using calcein double fluorescent labeling. At the 4th week, the femurs were taken to detect the expression of Runx2 and ALP using real-time PCR and Western blotting. Results Micro-CT imaging data showed significantly lower BV, BV/TV, Tb. N, Tb. Th, Ct. BV and Ct. TbTh （P 〈0.05） and significantly higher BS/BV, Tb. Sp and Ct. BS/BV （P 〈0. 01 ） in p75NTR^-/- mice than in p75NTR^＋/＋ mice. Calcein double fluorescent labeling showed that the MAR of the femur was significantly lower in in p75NTR^ -/- mice （ P 〈 0.01 ）. PCR and Western blotting results showed significantly decreased expressions of Runx2 and ALP in the femurs of p75NTR^-/-mice as compared with those in p75NTR^＋/＋ mice （P 〈 0. 01 ）. Conclusion p75NTR suppresses mineralization in the femoral bone of mice. Knocking out