Significance: Disruption of the blood–brain barrier (BBB) for drug delivery to the central nervous system (CNS) is achieved clinically through intracarotid artery hyperosmotic mannitol (ICAHM) infusions. However, ICAHM is often viewed as a simple drug delivery mechanism, and biological implications have been largely overlooked. This study demonstrates that BBB disruption by ICAHM generates a transient sterile neuroinflammatory response marked by increased production of cytokines, chemokines, and trophic factors leading activation of astrocytes, microglia, and macrophages. Additionally, unilateral ICAHM generates plasma increases in inflammatory molecules with evidence of sterile inflammation on the untreated contralateral hemisphere. These biological responses to ICAHM suggest its potential for immune modulation in several CNS pathologies.

Continuous focused ultrasound (cFUS) has been widely used for thermal ablation of tissues, relying on continuous exposures to generate temperatures necessary to induce coagulative necrosis. 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 effects created by non-thermal mechanisms of FUS (i.e., acoustic radiation forces and acoustic cavitation). pFUS has shown promise for a variety of applications including drug and nanoparticle delivery; however, little is understood about the effects these exposures have on tissue, especially with regard to cellular pro-homing factors (growth factors, cytokines, and cell adhesion molecules). We examined changes in murine 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-1α, IL-1β, TNFα, INFγ, MIP-1α, MCP-1, and GMCSF) creating a local cytokine gradient on days 0 and 1 post-pFUS that returns to baseline levels by day 3 post-pFUS. pFUS exposures induced upregulation of other signaling molecules (e.g., VEGF, FGF, PlGF, HGF, and SDF-1α) 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.

Chlorotoxin (CTX) is a 36-amino-acid disulfide-containing peptide derived from the venom of the scorpion Leiurus quinquestriatus. CTX alters physiology in numerous ways. It interacts with voltage gated chloride channels, Annexin-2, and matrix metalloproteinase-2 (MMP-2). CTX-based bioconjugates have been widely subjected to phase I/II clinical trials and have shown substantial promise. Many studies have demonstrated that CTX preferentially binds to neuroectodermal tumors, such as glioblastoma, without cross-reactivity to normal brain cells. With its ability to penetrate the blood-brain-barrier (BBB) and its tyrosine residue allows covalent conjugation with functional moieties, CTX is an attractive platform to explore development of diagnostic and therapeutic agents for gliomas. In this review, we outline CTX structure and its molecular targets, summarize molecular variations of CTX developed for glioma imaging, and discuss future trends and perspectives for CTX conjugates as a theranostic agent.

Pulsed focused ultrasound (pFUS) technology is being developed for clinical neuro/immune modulation and regenerative medicine. Biological signal transduction of pFUS forces can require mechanosensitive or voltage-gated plasma membrane ion channels. Previous studies suggested pFUS is capable of activating either channel type, but their mechanistic relationship remains ambiguous. We demonstrated pFUS bioeffects increased mesenchymal stem cell tropism (MSC) by altering molecular microenvironments through cyclooxygenase-2 (COX2)-dependent pathways. This study explored specific relationships between mechanosensitive and voltage-gated Ca2+ channels (VGCC) to initiate pFUS bioeffects that increase stem cell tropism. Methods: Murine kidneys and hamstring were given pFUS (1.15 or 1.125 MHz; 4MPa peak rarefactional pressure) under ultrasound or magnetic resonance imaging guidance. Cavitation and tissue displacement were measure by hydrophone and ultrasound radiofrequency data, respectively. Elastic modeling was performed from displacement measurements. COX2 expression and MSC tropism were evaluated in the presence of pharmacological ion channel inhibitors or in transient-receptor-potential-channel-1 (TRPC1)-deficient mice. Immunohistochemistry and co-immunoprecipitation examined physical channel relationships. Fluorescent ionophore imaging of cultured C2C12 muscle cells or TCMK1 kidney cells probed physiological interactions. Results: pFUS induced tissue deformations resulting in kPa-scale forces suggesting mechanical activation of pFUS-induced bioeffects. Inhibiting VGCC or TRPC1 in vivo blocked pFUS-induced COX2 upregulation and MSC tropism to kidneys and muscle. A TRPC1/VGCC complex was observed in plasma membranes. VGCC or TRPC1 suppression blocked pFUS-induced Ca2+ transients in TCMK1 and C2C12 cells. Additionally, Ca2+ transients were blocked by reducing transmembrane Na+ potentials and observed Na+ transients were diminished by genetic TRPC1 suppression. Conclusion: This study suggests that pFUS acoustic radiation forces mechanically activate a Na+-containing TRPC1 current upstream of VGCC rather than directly opening VGCC. The electrogenic function of TRPC1 provides potential mechanistic insight into other pFUS techniques for physiological modulation and optimization strategies for clinical implementation.

Animal studies have shown that mesenchymal stromal cell (MSC) infusions improve acute kidney injury (AKI) outcomes when administered early after ischemic/reperfusion injury or within 24 hours after cisplatin administration. These findings have spurred several human clinical trials to prevent AKI. However, no specific therapy effectively treats clinically obvious AKI or rescues renal function once advanced injury is established. We investigated if noninvasive image-guided pulsed focused ultrasound (pFUS) could alter the kidney microenvironment to enhance homing of subsequently infused MSC. To examine the efficacy of pFUS-enhanced cell homing in disease, we targeted pFUS to kidneys to enhance MSC homing after cisplatin-induced AKI. We found that pFUS enhanced MSC 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 MSC alone. We then investigated whether pFUS+MSC therapy could rescue established AKI. MSC alone at 3 days post-cisplatin, after renal functional deficits were obvious, significantly improved 7-day survival of animals. Survival was further improved by pFUS and MSC. pFUS prior to MSC injections increased IL-10 production by MSC that homed to kidneys and generated an anti-inflammatory immune cell profile in treated kidneys. This study shows pFUS is a neoadjuvant approach to improve MSC homing to diseased organs. pFUS with MSC better prevents AKI than MSC alone and allows rescue therapy in established AKI, which currently has no meaningful therapeutic options. Stem Cells 2015;33:1241–1253

Stem cells are promising therapeutics for cardiovascular diseases, and i.v. injection is the most desirable route of administration clinically. Subsequent homing of exogenous stem cells to pathological loci is frequently required for therapeutic efficacy and is mediated by chemoattractants (cell adhesion molecules, cytokines, and growth factors). Homing processes are inefficient and depend on short‐lived pathological inflammation that limits the window of opportunity for cell injections. Noninvasive pulsed focused ultrasound (pFUS), which emphasizes mechanical ultrasound–tissue interactions, can be precisely targeted in the body and is a promising approach to target and maximize stem cell delivery by stimulating chemoattractant expression in pFUS‐treated tissue prior to cell infusions. We demonstrate that pFUS is nondestructive to murine skeletal muscle tissue (no necrosis, hemorrhage, or muscle stem cell activation) and initiates a largely M2‐type macrophage response. We also demonstrate that local upregulation of chemoattractants in pFUS‐treated skeletal muscle leads to enhance homing, permeability, and retention of human mesenchymal stem cells (MSC) and human endothelial precursor cells (EPC). Furthermore, the magnitude of MSC or EPC homing was increased when pFUS treatments and cell infusions were repeated daily. This study demonstrates that pFUS defines transient “molecular zip codes” of elevated chemoattractants in targeted muscle tissue, which effectively provides spatiotemporal control and tunability of the homing process for multiple stem cell types. pFUS is a clinically translatable modality that may ultimately improve homing efficiency and flexibility of cell therapies for cardiovascular diseases. Stem Cells 2013;31:2551–2560