Preclinical high intensity focused ultrasound: mechanisms and applications

脉冲FUS （pFUS）采用非连续暴露，降低能量沉积速率，并允许在脉冲之间进行冷却，从而最大限度地减少热效应，并强调FUS的非热机制（即声辐射力和声空化）产生的效应。然而，对于这些暴露对组织的FUS影响知之甚少，特别是关于细胞促归巢因子（生长因子、细胞因子、细胞粘附分子）。我们检查了pFUS或cFUS后小鼠腘绳肌的变化，并证明pFUS与cFUS不同，对肌肉的组织学完整性几乎没有影响，也不会诱导细胞死亡。在pFUS或cFUS暴露后3天和8天观察巨噬细胞的浸润。pFUS增加了几种细胞因子的表达（如il -1 α、il -1 β、TNFalpha、INFgamma、mip -1 α、MCP-1、GMCSF），在pFUS后的第0天和第1天产生局部细胞因子梯度，并在pFUS后的第3天恢复到基线水平。pFUS暴露诱导其他信号分子（如VEGF、FGF、PlGF、HGF、SDF-1alpha）和细胞粘附分子（如ICAM-1和VCAM-1）对肌肉血管的上调。pFUS后观察到的肌肉分子变化可以通过增加病理区域的归巢来用于靶向细胞治疗。骨髓基质细胞（BMSCs）在疾病治疗中显示出巨大的希望，但其治疗效果往往受到系统给药细胞归巢效率低的限制，这导致病理部位的细胞数量较少。BMSC是炎症区域的家，在那里局部表达整合素和趋化因子梯度存在。我们证明，pFUS暴露强调超声-组织相互作用的机械效应，可诱导小鼠肾脏中化学引诱剂（即细胞因子、整合素和生长因子）的局部和短暂升高。ppfus诱导的细胞因子上调发生在治疗后约1天，并在第3天恢复到对侧肾脏水平。这个细胞因子表达显著增加的窗口伴随着其他营养因子和整合素的局部增加，这些因子和整合素已被证明可以促进BMSC归巢。在pFUS治疗后，单肾静脉给予骨髓间充质干细胞，与对侧肾脏相比，在治疗肾脏中观察到骨髓间充质干细胞的归巢、通透性和保留性增强。组织学分析显示，在治疗后第1天和第3天，治疗后肾脏小管周围区域的BMSC增加了8倍。此外，在给予BMSC后，ppfus处理肾脏的细胞因子水平与对照组相似，表明BMSC调节了细胞因子水平。这些结果表明，pFUS可以通过建立局部化学引诱剂梯度和增加细胞粘附分子的表达来增强细胞对治疗组织的趋向性，从而潜在地改善基于细胞的治疗方法，作为一种无创的靶向归巢方式。

Pulsed FUS (pFUS) employs non-continuous exposures, that lower the rate of energy deposition and allow cooling to occur between pulses, thereby minimizing thermal effects and emphasizing the effects created by non-thermal mechanisms of FUS (i.e. acoustic radiation forces and acoustic cavitation). However, little is understood about the FUS effects these exposures have on tissue, especially with regard to cellular pro-homing factors (growth factors, cytokines, cell adhesion molecules). We examined changes in mice hamstring muscle following pFUS or cFUS and demonstrate that pFUS, unlike cFUS, has little effect on the histological integrity of muscle and does not induce cell death. Infiltration of macrophages was observed 3 and 8 days following pFUS or cFUS exposures. pFUS increased expression of several cytokines (e.g. IL-1alpha;, IL-1beta;, TNFalpha;, INFgamma;, MIP-1alpha;, MCP-1, GMCSF) creating a local cytokine gradient on days 0 and 1 following pFUS that returns to baseline levels by day 3 post-pFUS. pFUS exposures induced up-regulation of other signaling molecules (e.g. VEGF, FGF, PlGF, HGF, SDF-1alpha;) and cell adhesion molecules (e.g. ICAM-1 and VCAM-1) on muscle vasculature. The observed molecular changes in muscle following pFUS may be utilized to target cellular therapies, by increasing homing to areas of pathology. Bone marrow stromal cells (BMSCs) have shown significant promise in the treatment of disease, but their therapeutic efficacy is often limited by inefficient homing of systemically administered cells, which results in low number of cells accumulating at sites of pathology. BMSC home to areas of inflammation where local expression of integrins and chemokine gradients is present. We demonstrated that pFUS exposures that emphasize the mechanical effects of ultrasound-tissue interactions induced local and transient elevations of chemoattractants (i.e., cytokines, integrins, and growth factors) in the murine kidney. pFUS-induced upregulation of cytokines occurred through approximately 1 day post-treatment and returned to contralateral kidney levels by day 3. This window of significant increases in cytokine expression was accompanied by local increases of other trophic factors and integrins that have been shown to promote BMSC homing. When BMSCs were intravenously administered following pFUS treatment to a single kidney, enhanced homing, permeability, and retention of BMSC was observed in the treated kidney versus the contralateral kidney. Histological analysis revealed up to eight times more BMSC in the peritubular regions of the treated kidneys on days 1 and 3 post-treatment. Furthermore, cytokine levels in pFUS-treated kidneys following BMSC administration were found to be similar to controls, suggesting modulation of cytokine levels by BMSC. These results suggest that pFUS could potentially improve cell-based therapies as a noninvasive modality to target homing by establishing local chemoattractant gradients and increasing expression of cell adhesion molecules to enhance tropism of cells toward treated tissues.