Preclinical high intensity focused ultrasound: mechanisms and applications

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。骨髓间充质干细胞是炎症区域的家园，在那里局部表达细胞粘附分子和趋化因子梯度存在。脉冲聚焦超声（pFUS）采用非连续暴露，从而降低能量沉积速率，并允许在脉冲之间进行冷却，从而最大限度地减少热效应，并强调由FUS的非热机制（即声辐射力和声空化）产生的效应。我们检查了pFUS对组织的组织学完整性几乎没有影响，也不会诱导细胞死亡后肾脏的变化。pFUS增加了几种化学引诱剂的表达，在pFUS后的第0天和第1天产生了一个短暂的分子邮编，在pFUS后的第3天恢复到基线水平。pFUS暴露诱导肌肉血管细胞粘附分子的上调。我们诱导机械转导反应，启动一个很大程度上抗炎的m2型巨噬细胞环境。我们证明了pfu处理的肾脏中局部化学引诱剂的上调导致BMSC的归巢、通透性和保留增强。我们还证明，pFUS进入肾脏后诱导的分子变化可以被布洛芬（一种环氧化酶2抑制剂）或TNF α受体结合蛋白依那西普阻断，这表明机械转导效应是通过组织中的NFkB和COX 2途径起作用的。在pFUS和干细胞输注之前使用布洛芬或依那西普都可以阻断干细胞向目标肌肉的归巢，这表明使用这种方法我们可以使用pFUS来询问药物-宿主组织相互作用及其对归巢的影响。

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 cell adhesion molecules and chemokine gradients are present. Pulsed focused ultrasound (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). We examined changes in the kidney following pFUS has little effect on the histological integrity of the tissue and does not induce cell death. pFUS increased expression of several chemoattractants creating a transient molecular zip code on days 0 and 1 following pFUS that returns to baseline levels by day 3 post-pFUS. pFUS exposures induced up-regulation of cell adhesion molecules on muscle vasculature. We induce a mechanotransductive response that initiates a largely an anti-inflammatory M2-type macrophage environment. We demonstrated local upregulation of chemoattractants in pFUS-treated kidney leads to enhance homing, permeability, and retention of BMSC. We also demonstrate that the induced molecular changes following pFUS to the kidney can be block by ibuprofen, a cyclooxygenase 2 inhibitor, or TNF alpha receptor binding protein, etanercept, indicating that the mechanotransductive effects are acting through a NFkB and COX 2 pathway in the tissue. Both ibuprofen or etanercept administered prior to pFUS and stem cell infusion block the homing of stem cells to targeted muscle which indicated that using this approach we can use pFUS to interrogate drug-host tissue interactions and their effect on homing. We also demonstrated that pFUS exposures in combination with BMSC in an acute kidney injury model induce mechanotransductive effects in the murine kidney (AKI). To examine the efficacy of pFUS-enhanced cell homing in disease, we targeted pFUS to kidneys to enhance BMSC homing after cisplatin-induced AKI. We found that pFUS enhanced BMSC homing at 1 day post-cisplatin, prior to renal functional deficits, and that enhanced homing improved outcomes of renal function, tubular cell death, and regeneration at 5 days post-cisplatin compared to BMSCs alone. After observing improved homing and AKI outcomes during early AKI, we investigated whether pFUS+BMSC therapy could rescue established AKI. BMSC administration alone at 3 days post-cisplatin, after renal functional deficits become obvious, significantly improved 7-day survival of animals. Survival was further improved using pFUS+BMSC. BMSCs, alone or with pFUS, shifted the kidney macrophage phenotype from M1 to M2. This study shows that pFUS serves as a neoadjuvant treatment to improve MSC homing to diseased organs. We have also shown that pFUS increases interferon gamma in tissue that augments the local potency of the infused BMSC that home to the tissue by stimulating release of interleukin (IL) 10 from infused cells.. We also have similar results in critical limb ischemia model in which we have been able to demonstrate that the combination of pFUS+BMSC resulted in greater reperfusion of the ischemic limb compared to animal receiving pFUS or BMSC alone. Evaluation of the tissue demonstrated pFUS pretreatment followed by BMSC infusion of the ischemic muscle increased the amount of human vascular endothelia growth factor and interleukin 10 present in the microenvironment compared to animals only receiving BMSC injections. These results indicate that pFUS preconditions the tissues that results in increased potency of the BMSC that homed to targeted tissues and has important implication for translation to clinical trials. Magnetic resonance imaging (MRI)-guided pulsed focused ultrasound (pFUS) combined with systemic infusion of ultrasound contrast agent microbubbles (MB) causes localized bloodbrain barrier (BBB) disruption as evidenced by contrast-enhanced MRI, resulted in an immediate damage-associated molecular pattern (DAMP) response including elevations in heat-shock protein 70, IL-1, IL-18, and tumor necrosis factor indicative of a sterile inflammatory response (SIR) in the parenchyma. Concurrent with DAMP presentation, significant elevations in proinflammatory, antiinflammatory, and trophic factors along with neurotrophic and neurogenesis factors were detected that lasted 24 h. The effect of pFUS+MB was due to an intravascular shock wave from stable cavitation of the MB and resulted in TUNEL+ neurons, activated microglia and astrocytes and an innate immune response that would be compatible with mild trauma or ischemia in the brain.