Minimally invasive laser Doppler flowmetry is suitable for serial bone perfusion measurements in mice
Minimally invasive laser Doppler flowmetry is suitable for serial bone perfusion measurements in mice
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DOI：
10.1016/j.bonr.2019.100231
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发表时间：
2019-12-01
期刊：
影响因子：
2.5
通讯作者：
Cole, Jacqueline H.
中科院分区：
文献类型：
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作者：
Hanne, Nicholas J.;Easter, Elizabeth D.;Cole, Jacqueline H.
在体内激光多普勒血流仪（LDF）先前已被用来量化血液灌注准确地在一个单一的时间点在小鼠胫骨干骺端。然而，该手术导致覆盖骨的软组织的实质性破坏，并在手术后数周内引起明显的局部炎症，阻碍了同一小鼠的连续测量。在这项研究中，我们测试了一种微创技术，用LDF测量胫骨的灌注，并确定它可以连续用于同一只小鼠，而不会引起炎症或步态紊乱的迹象。20只14周龄的C57 Bl/6 J小鼠被平均分为每天进行跑步机运动或保持久坐的组。在这些活动组中，将小鼠均匀地细分为每周一次或在研究终点仅一次接受LDF测量的组。用LDF测量右侧胫骨干骺端前内侧区域的骨灌注。每周测量白细胞介素6的血清浓度、切口部位伤口面积和步态中的肢体间协调性，持续四周。在任何时间点，每周一次或仅终点LDF组的运动组和久坐组之间的胫骨灌注没有显著差异。相对于第二周和第四周的测量值，每周LDF组第三周的灌注显著增加。股动脉结扎导致胫骨灌注持续快速减少，证实LDF对胫骨血供变化敏感。每周一次LDF程序不会对步态产生不利影响，因为在每个时间点，每周一次LDF组和仅终点LDF组在跑步机运动期间的肢体间协调相似。切口部位的图像显示伤口在一周内闭合，每周一次组和仅终点组之间的血清白细胞介素6浓度没有显著差异。总之，这些发现表明，我们的微创LDF技术适用于骨内血液灌注的系列体内测量，而不会诱导局部炎症或对小鼠的步态产生负面影响。
In vivo laser Doppler flowmetry (LDF) has previously been used to quantify blood perfusion accurately at a single timepoint in the murine tibial metaphysis. However, this procedure entailed substantial disruption to soft tissues overlying the bone and caused notable localized inflammation for several weeks after the procedure, impeding serial measurements in the same mouse. In this study, we tested a less invasive technique to measure perfusion in the tibia with LDF and determined that it can be used serially in the same mouse without causing signs of inflammation or gait perturbations. Twenty 14-week-old C57Bl/6J mice were evenly divided into groups that either had daily treadmill exercise or remained sedentary. Within these activity groups, mice were evenly subdivided into groups that received LDF measurements either weekly or only once at the study endpoint. Bone perfusion was measured with LDF in the anteromedial region of the right tibial metaphysis. Serum concentrations of interleukin 6, incision site wound area, and interlimb coordination during gait were measured weekly for four weeks. Tibial perfusion did not differ significantly between exercise and sedentary groups within the weekly or endpoint-only LDF groups at any timepoint. Perfusion was significantly increased in the third week in the weekly LDF group relative to measurements in the second and fourth weeks. Ligation of the femoral artery caused consistent, rapid reductions in tibial perfusion, validating that LDF is sensitive to changes in tibial blood supply. Weekly LDF procedures did not adversely affect gait, as interlimb coordination during treadmill locomotion was similar between weekly and endpoint-only LDF groups at every timepoint. Images of the incision site show wound closure within one week, and serum concentrations of interleukin 6 were not significantly different between weekly and endpoint-only groups. Together, these findings demonstrate that our minimally invasive LDF technique is suitable for serial in vivo measurements of intraosseous blood perfusion without inducing localized inflammation or negatively affecting gait patterns in mice.