Robo4 inhibits gamma radiation-induced permeability of a murine microvascular endothelial cell by regulating the junctions.

BACKGROUND: Hematopoietic stem cell transplantation involves irradiation preconditioning which causes bone marrow endothelial cell dysfunction. While much emphasis is on the reconstitution of hematopoietic stem cells in the bone marrow microenvironment, endothelial cell preservation is indispensable to overcome the preconditioning damages. This study aims to ascertain the role of Roundabout 4 (Robo4) in regulating irradiation-induced damage to the endothelium. METHODS: Microvascular endothelial cells were treated with γ-radiation to establish an endothelial cell injury model. Robo4 expression in the endothelial cells was manipulated employing lentiviral-mediated RNAi and gene overexpression technology before irradiation treatment. The permeability of endothelial cells was measured using qPCR, immunocytochemistry, and immunoblotting to analyze the effect on the expression and distribution of junctional molecules, adherens junctions, tight junctions, and gap junctions. Using Transwell endothelial monolayer staining, FITC-Dextran permeability, and gap junction-mediated intercellular communication (GJIC) assays, we determined the changes in endothelial functions after Robo4 gene manipulation and irradiation. Moreover, we measured the proportion of CD31 expression in endothelial cells by flow cytometry. We analyzed variations between two or multiple groups using Student's t-tests and ANOVA. RESULTS: Ionizing radiation upregulates Robo4 expression but disrupts endothelial junctional molecules. Robo4 deletion causes further degradation of endothelial junctions hence increasing the permeability of the endothelial cell monolayer. Robo4 knockdown in microvascular endothelial cells increases the degradation and delocalization of ZO-1, PECAM-1, occludin, and claudin-5 molecules after irradiation. Conversely, connexin 43 expression increases after silencing Robo4 in endothelial cells to induce permeability but are readily destroyed when exposed to 10 Gy of gamma radiation. Also, Robo4 knockdown enhances Y731-VE-cadherin phosphorylation leading to the depletion and destabilization of VE-cadherin at the endothelial junctions following irradiation. However, Robo4 overexpression mitigates irradiation-induced degradation of tight junctional proteins and stabilizes claudin-5 and ZO-1 distribution. Finally, the enhanced expression of Robo4 ameliorates the irradiation-induced depletion of VE-cadherin and connexin 43, improves the integrity of microvascular endothelial cell junctions, and decreases permeability. CONCLUSION: This study reveals that Robo4 maintains microvascular integrity after radiation preconditioning treatment by regulating endothelial permeability and protecting endothelial functions. Our results also provided a potential mechanism to repair the bone marrow vascular niche after irradiation by modulating Robo4 expression.

Low-Dose X-Ray Increases Paracellular Permeability of Human Renal Glomerular Endothelial Cells.

Objective: Glomerular endothelium functions as a filtration barrier of metabolites in the kidney. Although X-ray irradiation modulated the permeability of the vascular endothelium, the response of human renal glomerular endothelial cells (HRGECs) to low-dose X-ray irradiation has not been investigated. We evaluated the impacts of low-dose X-ray irradiation on HRGECs and revealed the underlying mechanism. Methods: HRGECs were exposed to X-ray with doses of 0, 0.1, 0.5, 1.0, and 2.0 Gy. The proliferation, viability, and apoptosis of HRGECs were examined by MTT assay, trypan blue staining assay, and TUNEL staining, respectively. The paracellular permeability was assessed by paracellular permeability assay. The expression of VE-cadherin was investigated via immunofluorescence assay. Western blot and qRT-PCR detected the expression levels of VE-cadherin and CLDN5. Besides, the expression levels of pVE-cadherin (pY658), TGF-ß, TGF-ßRI, Src, p-Src, Smad2, p-Smad2, Smad3, p-Smad3, SNAIL, SLUG, and apoptosis-related proteins were tested by Western blot. Results: The proliferation, viability, and apoptosis of HRGECs were not affected by low-dose (<2.0 Gy) X-ray irradiation. X-ray irradiation dose-dependently reduced the level of VE-cadherin, and VE-cadherin and CLDN5 levels were reduced with X-ray irradiation. The levels of pY658, p-Src, p-Smad2, and p-Smad3 were upregulated with the increase in X-ray dose. Besides, the paracellular permeability of HRGECs was increased by even low-dose (<2.0 Gy) X-ray irradiation. Therefore, low-dose X-ray irradiation reduced the cumulative content of VE-cadherin and increased the level of pY658 via activation of the TGF-ß signaling pathway. Conclusion: Even though low-dose X-ray exposure had no impact on proliferation, viability, and apoptosis of HRGECs, it increased the paracellular permeability by deterioration and downregulation of VE-cadherin through stimulating the TGF-ß signaling pathway. This study built the framework for kidney response to low-dose irradiation exposure.

Inactivation of Aspergillus species in real water matrices using medium pressure mercury lamps.

The aim of this work is to understand the inactivation efficiency of medium pressure mercury lamps, measured in terms of growth inhibition as well as cell death, damage and response, using three strains from three different Aspergillus species (A. fumigatus, A. niger and, A. terreus) spiked in filtered surface water. A complete characterization of the effect of the treatment on each strain of the fungal species was assessed considering spores' morphology, cell wall integrity and enzymatic activity, the formation of pyrimidine dimers in the DNA and proteome analysis. Results showed that, when subjected to medium pressure mercury lamps, A. niger is the most resistant to inactivation, that both A. fumigatus and A. niger suffer more morphological changes and present a higher number of damaged spores and A. terreus presented more dead spores. DNA damages detected in A. niger were able to be repaired to some extent, under both light and dark conditions. Finally, proteome analysis showed that the UV radiation treatment triggered different types of stress response, including cell wall reorganization and DNA repair in A. fumigatus and A. terreus, and oxidative stress responses like the increase in production of citric acid and itaconic acid in A. niger and A. terreus, respectively.

Enhanced permeability and photoprotective potential of optimized p-coumaric acid-phospholipid complex loaded gel against UVA mediated oxidative stress.

Photo-oxidative skin damage is mainly caused by the UV-A radiation of the sun. Synthetic sunscreens used to counter this acts mostly on the superficial skin layer and possess serious side effects. P-coumaric acid (PCA) is a UV-A protective plant phenolic having quick diffusion and distribution in superficial skin layers limiting its application as herbal sunscreen. The present study was designed to formulate an optimized phospholipid complex of PCA (PCAPC) through response surface methodology to enhance its skin permeation to deeper skin layers providing protection against photo-oxidative stress. PCAPC was characterized by FT-IR, DTA, PXRD, TEM, zeta potential etc. PCAPC was then incorporated into a gel formulation (PCAPC-GE) to facilitate its transdermal delivery. Physicochemical properties of the gel were assessed by pH, homogeneity, rheology, spreadability etc. In-vitro SPF and UVA-PF of the gel was evaluated and compared with conventional gel (PCA-GE). Ex-vivo skin permeation flux, permeability coefficient, skin deposition and dermatokinetic analysis were carried out to measure the rate and level of skin permeation. This was accompanied by in-vivo evaluation of PCAPC-GE and PCA-GE in the experimental rat model by measuring the various oxidative stress markers such as superoxide dismutase, catalase etc. PCAPC-GE provided high SPF and UVA-PF value compared to PCA-GE. The physicochemical parameters were suitable for transdermal application. PCAPC-GE enhanced the permeation rate of PCA by almost 6 fold compared to PCA-GE. Besides, a significant reduction of UV-A induced oxidative stress biomarkers were observed for PCAPC-GE. Thus, the PCAPC-GE may be an effective alternative of synthetic sunscreens due to its enhanced permeation and protection against UVA-induced oxidative stress.

Ultrasound-mediated disruption of the blood tumor barrier for improved therapeutic delivery.

The blood-brain barrier (BBB) is a major anatomical and physiological barrier limiting the passage of drugs into brain. Central nervous system tumors can impair the BBB by changing the tumor microenvironment leading to the formation of a leaky barrier, known as the blood-tumor barrier (BTB). Despite the change in integrity, the BTB remains effective in preventing delivery of chemotherapy into brain tumors. Focused ultrasound is a unique noninvasive technique that can transiently disrupt the BBB and increase accumulation of drugs within targeted areas of the brain. Herein, we summarize the current understanding of different types of targeted ultrasound mediated BBB/BTB disruption techniques. We also discuss influence of the tumor microenvironment on BBB opening, as well as the role of immunological response following disruption. Lastly, we highlight the gaps between evaluation of the parameters governing opening of the BBB/BTB. A deeper understanding of physical opening of the BBB/BTB and the biological effects following disruption can potentially enhance treatment strategies for patients with brain tumors.

Focused Ultrasound and Microbubble Treatment Increases Delivery of Transferrin Receptor-Targeting Liposomes to the Brain.

The blood-brain barrier (BBB) is a major obstacle to treating several brain disorders. Focused ultrasound (FUS) in combination with intravascular microbubbles increases BBB permeability by opening tight junctions, creating endothelial cell openings, improving endocytosis and increasing transcytosis. Here we investigated whether combining FUS and microbubbles with transferrin receptor-targeting liposomes would result in enhanced delivery to the brain of post-natal rats compared with liposomes lacking the BBB-targeting moiety. For all animals, increased BBB permeability was observed after FUS treatment. A 40% increase in accumulation of transferrin receptor-targeting liposomes was observed in the FUS-treated hemisphere, whereas the isotype immunoglobulin G liposomes showed no increased accumulation. Confocal laser scanning microscopy of brain sections revealed that both types of liposomes were mainly observed in endothelial cells in the FUS-treated hemisphere. The results demonstrate that FUS and microbubble treatment combined with BBB-targeting liposomes could be a promising approach to enhance drug delivery to the brain.

Use of gamma irradiation technology for modification of bacterial cellulose nanocrystals/chitosan nanocomposite film.

The objective of this work was to investigate the influence of different gamma ray dosages (5, 10, and 10 kGy) on the structural, mechanical, surface and barrier properties of chitosan (Ch) based nanocomposite film. The results showed gamma irradiation caused an increase in the surface hydrophobicity, water vapor permeability and sensitivity of films to water and also, yellowness and opacity of films increased, simultaneously. By increasing the irradiation doses up to 10 kGy, the mechanical properties of Ch/BCNC film was significantly enhanced. As observed by FTIR spectra, no change occurred in the chemical functional groups of the films during irradiation. XRD studies confirmed that crystallinity of films was increased after irradiation. The nanocomposite film irradiated by 10 kGy had the highest thermal stability. In conclusion, gamma radiation can be considered as a safe method for sterilization of foods and modification of Ch/BCNC film properties.

Musashi expression in intestinal stem cells attenuates radiation-induced decline in intestinal permeability and survival in Drosophila.

Exposure to genotoxic stress by environmental agents or treatments, such as radiation therapy, can diminish healthspan and accelerate aging. We have developed a Drosophila melanogaster model to study the molecular effects of radiation-induced damage and repair. Utilizing a quantitative intestinal permeability assay, we performed an unbiased GWAS screen (using 156 strains from the Drosophila Genetic Reference Panel) to search for natural genetic variants that regulate radiation-induced gut permeability in adult D. melanogaster. From this screen, we identified an RNA binding protein, Musashi (msi), as one of the possible genes associated with changes in intestinal permeability upon radiation. The overexpression of msi promoted intestinal stem cell proliferation, which increased survival after irradiation and rescued radiation-induced intestinal permeability. In summary, we have established D. melanogaster as an expedient model system to study the effects of radiation-induced damage to the intestine in adults and have identified msi as a potential therapeutic target.

LPAR2 receptor activation attenuates radiation-induced disruption of apical junctional complexes and mucosal barrier dysfunction in mouse colon.

The tight junction (TJ) and barrier function of colonic epithelium is highly sensitive to ionizing radiation. We evaluated the effect of lysophosphatidic acid (LPA) and its analog, Radioprotein-1, on γ-radiation-induced colonic epithelial barrier dysfunction using Caco-2 and m-ICC12 cell monolayers in vitro and mice in vivo. Mice were subjected to either total body irradiation (TBI) or partial body irradiation (PBI-BM5). Intestinal barrier function was assessed by analyzing immunofluorescence localization of TJ proteins, mucosal inulin permeability, and plasma lipopolysaccharide (LPS) levels. Oxidative stress was analyzed by measuring protein thiol oxidation and antioxidant mRNA. In Caco-2 and m-ICC12 cell monolayers, LPA attenuated radiation-induced redistribution of TJ proteins, which was blocked by a Rho-kinase inhibitor. In mice, TBI and PBI-BM5 disrupted colonic epithelial tight junction and adherens junction, increased mucosal permeability, and elevated plasma LPS; TJ disruption by TBI was more severe in Lpar2-/- mice compared to wild-type mice. RP1, administered before or after irradiation, alleviated TBI and PBI-BM5-induced TJ disruption, barrier dysfunction, and endotoxemia accompanied by protein thiol oxidation and downregulation of antioxidant gene expression, cofilin activation, and remodeling of the actin cytoskeleton. These data demonstrate that LPAR2 receptor activation prevents and mitigates γ-irradiation-induced colonic mucosal barrier dysfunction and endotoxemia.

The enigma of headaches associated with electromagnetic hyperfrequencies: Hypotheses supporting non-psychogenic algogenic processes.

Although an electrohypersensitivity (EHS) is reported in numerous studies, some authors associate hyperfrequencies (HF)-related pains with a nocebo effect while others suggest a biological effect. Therefore, we aimed to suggest hypotheses about the complex mechanisms of headaches related to HF-exposure. We crossed basic features of headaches with relevant studies (from the year 2000 up to 2018) emphasizing on the HF effects that may lead to pain genesis: neuroglial dysmetabolism, neuroinflammation, changes in cerebral blood perfusion, blood-brain barrier dysfunction and electrophysiological evidences of hyperexcitability. We privileged studies implying a sham exposure (for in vivo studies) and a specific absorption rate lower than 4 W/Kg. HF-induced headaches may involve an indirect inflammatory process (neurogenic, magnetogenic or thermogenic) as well as a direct biophysical effect (thermogenic or magnetogenic). We linked inflammatory processes to meningeal dysperfusion or primary neuroglial dysfunction triggered by non-thermal irradiation or HF-induced heating at thermal powers. In the latter case, HF-induced excitoxicity and oxidative stress probably play a crucial role. Such disorders may lead to vascular-trigeminal activation in predisposed people. Interestingly, an abnormal oxidative stress predisposition had been demonstrated in overall 80% of EHS self-reporting patients. In the case of direct effects, pain pathways' activation may be directly triggered by HF-irradiation (heating and/or transcranial HF-induced ectopic action potentials). Further research on HF-related headaches is needed.

Treatment and management strategies of onychomycosis.

Onychomycosis is one of the most prevalent and severe nail fungal infections, which is affecting a wide population across the globe. It leads to variations like nail thickening, disintegration and hardening. Oral and topical drug delivery systems are the most desirable in treating onychomycosis, but the efficacy of the results is low, resulting in a relapse rate of 25-30%. Due to systemic toxicity and various other disadvantages associated with oral therapy like gastrointestinal, hepatotoxicity, topical therapy is commonly used. Topical therapy improves patient compliance and reduces the cost of treatment. However, due to poor penetration of topical therapy across the nail plate, research is focused on different chemical, mechanical and physical methods to improve drug delivery. Penetration enhancers like Thioglycolic acid, Hydroxypropyl-ß-cyclodextrin (HP-ß-CD), Sodium lauryl sulfate (SLS), carbocysteine, N-acetylcysteine etc. have shown results enhancing the drug penetration across the nail plate. Results with physical techniques such as iontophoresis, laser and Photodynamic therapy are quite promising, but the long-term suitability of these devices is in need to be determined. In this article, a brief analysis of the treatment procedures, factors affecting drug permeation across nail plate, chemical, mechanical and physical devices used to increase the drug delivery through nails for the onychomycosis management has been achieved.

Factors Affecting Depth of Penetration in Microneedling- and Laser-Assisted Drug Delivery: The Importance of Timing of Topical Application.

BACKGROUND: Microneedling- and laser-assisted drug delivery are emerging techniques used to treat various conditions. However, key parameters affecting drug penetration remain unknown. OBJECTIVE: This study aims to investigate the importance of timing of topical application, needle length, and device type for drug delivery. MATERIALS AND METHODS: Skin harvested from cosmetic surgeries was treated with black ink applied before or after treatment with a microneedling pen (MP), roller, or fractional ablative CO2 laser, and incubated for different time intervals. Ink penetration was additionally tested using different needle lengths. Sandwich estimator was used for statistical analysis. RESULTS: Ink applied before MP penetrated deeper compared to ink applied afterward at 1 and 3 hours, and roller microneedling in both the ink-before and -after scenarios at 1, 3, and 6 hours (p < .05). Microneedling demonstrated lateral extension of ink beyond microchannels with increased ink penetration over time. CO2 laser demonstrated ink localization within microthermal zones without time-dependent increases in depth after 30 minutes. Ink penetration increases by 0.06 mm per 1 mm increase in needle length. CONCLUSION: Ink applied before MP results in the deepest penetration of ink. Microneedling offers unique advantages in transdermal delivery as its channels exhibit increasing penetration over time and lateral extension of product.

Light-Driven ATP Regeneration in Diblock/Grafted Hybrid Vesicles.

Light-driven ATP regeneration systems combining ATP synthase and bacteriorhodopsin have been proposed as an energy supply in the field of synthetic biology. Energy is required to power biochemical reactions within artificially created reaction compartments like protocells, which are typically based on either lipid or polymer membranes. The insertion of membrane proteins into different hybrid membranes is delicate, and studies comparing these systems with liposomes are needed. Here we present a detailed study of membrane protein functionality in different hybrid compartments made of graft polymer PDMS-g-PEO and diblock copolymer PBd-PEO. Activity of more than 90 % in lipid/polymer-based hybrid vesicles could prove an excellent biocompatibility. A significant enhancement of long-term stability (80 % remaining activity after 42 days) could be demonstrated in polymer/polymer-based hybrids.

Ultrasonically and Iontophoretically Enhanced Drug-Delivery System Based on Dissolving Microneedle Patches.

A multifunctional system comprised of hyaluronic acid microneedles was developed as an effective transdermal delivery platform for rapid local delivery. The microneedles can regulate the filling amount on the tip, by controlling the concentration of hyaluronic acid solution. Ultrasonication induces dissolution of the HA microneedles via vibration of acoustic pressure, and AC iontophoresis improves the electrostatic force-driven diffusion of HA ions and rhodamine B. The effect of ultrasound on rhodamine release was analyzed in vitro using a gelatin hydrogel. The frequency and voltage dependence of the AC on the ion induction transfer was also evaluated experimentally. The results showed that the permeability of the material acts as a key material property. The delivery system based on ultrasonication and iontophoresis in microneedles increases permeation, thus resulting in shorter initial delivery time than that required by delivery systems based on passive or ultrasonication alone. This study highlights the significance of the combination between ultrasonic waves and iontophoresis for improving the efficiency of the microneedles, by shortening the reaction duration. We anticipate that this system can be extended to macromolecular and dependence delivery, based on drug response time.

Arabinoxylan rice bran (MGN-3/Biobran) alleviates radiation-induced intestinal barrier dysfunction of mice in a mitochondrion-dependent manner.

MGN-3 is an arabinoxylan from rice bran that has been shown to be an excellent antioxidant and radioprotector. This study examined the protective effects of MGN-3 on radiation-induced intestinal injury. Mice were treated with MGN-3 prior to irradiation, then continued to receive MGN-3 for 4 weeks thereafter. MGN-3 increased the activity of mitochondrial respiratory chain complexes Ⅰ, Ⅲ, Ⅳ and Ⅴ, the intercellular ATP content, the mitochondria-encoded gene expression and mitochondrial copy numbers in the jejunal and colonic mucosa. MGN-3 reduced the oxidative stress levels and inflammatory response indicators in the serum and jejunal and colonic mucosa. Antioxidant indicators such as superoxide dismutase, glutathione peroxidase, catalase and total antioxidant capacity were significantly increased in the serum and jejunal and colonic mucosa in the MGN-3 group. Moreover, MGN-3 decreased the gene abundances and enzymatic activities of caspase-3, 8, 9 and 10 in the jejunal and colonic mucosa. The endotoxin, diamine peroxidase, d-lactate and zonulin levels were significantly reduced in the serum and jejunal and colonic mucosa in the MGN-3 group. MGN-3 also markedly upregulated the gene abundances of ZO-1, occludin, claudin-1 and mucin 2. MGN-3 effectively attenuated radiation-induced changes in the intestinal epithelial mitochondrial function, oxidative stress, inflammatory response, apoptosis, intestinal permeability and barrier function in mice. These findings add to our understanding of the potential mechanisms by which MGN-3 alleviates radioactive intestinal injury.

Ionizing radiation increases the endothelial permeability and the transendothelial migration of tumor cells through ADAM10-activation and subsequent degradation of VE-cadherin.

BACKGROUND: We analyzed the changes in permeability of endothelial cell layers after photon irradiation, with a focus on the metalloproteases ADAM10 and ADAM17, and on VE-cadherin, components crucial for the integrity of endothelial intercellular junctions, and their roles in the transmigration of cancer cells through endothelial cell monolayers. METHODS: Primary HUVEC were irradiated with 2 or 4 Gy photons at a dose rate of 5 Gy/min. The permeability of an irradiated endothelial monolayer for macromolecules and tumor cells was analyzed in the presence or absence of the ADAM10/17 inhibitors GI254023X and GW280264X. Expression of ADAM10, ADAM17 and VE-Cadherin in endothelial cells was quantified by immunoblotting and qRT. VE-Cadherin was additionally analyzed by immunofluorescence microscopy and ELISA. RESULTS: Ionizing radiation increased the permeability of endothelial monolayers and the transendothelial migration of tumor cells. This was effectively blocked by a selective inhibition (GI254023X) of ADAM10. Irradiation increased both, the expression and activity of ADAM10, which led to increased degradation of VE-cadherin, but also led to higher rates of VE-cadherin internalization. Increased degradation of VE-cadherin was also observed when endothelial monolayers were exposed to tumor-cell conditioned medium, similar to when exposed to recombinant VEGF. CONCLUSIONS: Our results suggest a mechanism of irradiation-induced increased permeability and transendothelial migration of tumor cells based on the activation of ADAM10 and the subsequent change of endothelial permeability through the degradation and internalization of VE-cadherin.

Colloids, nanoparticles, and materials for imaging, delivery, ablation, and theranostics by focused ultrasound (FUS).

This review focuses on different materials and contrast agents that sensitize imaging and therapy with Focused Ultrasound (FUS). At high intensities, FUS is capable of selectively ablating tissue with focus on the millimeter scale, presenting an alternative to surgical intervention or management of malignant growth. At low intensities, FUS can be also used for other medical applications such as local delivery of drugs and blood brain barrier opening (BBBO). Contrast agents offer an opportunity to increase selective acoustic absorption or facilitate destructive cavitation processes by converting incident acoustic energy into thermal and mechanical energy. First, we review the history of FUS and its effects on living tissue. Next, we present different colloidal or nanoparticulate approaches to sensitizing FUS, for example using microbubbles, phase-shift emulsions, hollow-shelled nanoparticles, or hydrophobic silica surfaces. Exploring the science behind these interactions, we also discuss ways to make stimulus-responsive, or "turn-on" contrast agents for improved selectivity. Finally, we discuss acoustically-active hydrogels and membranes. This review will be of interest to those working in materials who wish to explore new applications in acoustics and those in acoustics who are seeking new agents to improve the efficacy of their approaches.

Enhanced Antitumor Efficacy Achieved Through Combination of nsPEFs and Low-Dosage Paclitaxel.

Looking for a safe and effective cancer therapy for patients is becoming an important and promising research direction. Nanosecond pulsed electric field (nsPEF) has been found to be a potential non-thermal therapeutic technique with few side effects in pre-clinical studies. On the other hand, paclitaxel (PTX), as a common chemotherapeutic agent, shows full anti-tumor activities and is used to treat a wide variety of cancers. However, the delivery of PTX is challenging due to its poor aqueous solubility. Hence, high dosages of PTX have been used to achieve effective treatment, which creates some side effects. In this study, nsPEF was combined with low-level PTX, in order to validate if this combined treatment could bring about enhanced efficacy and allow reduced doses of PTX in clinical application. Cell proliferation, apoptosis, and cell cycle distribution were examined using MTT and flow cytometry assay, respectively. Results showed that combination treatments of nsPEF and PTX exhibited significant synergistic effects in vitro. The underlying mechanism might be that these two agents acted at different targets and coordinately enhanced MDA-MB-231 cell death.

MRI-Guided Focused Ultrasound for Targeted Delivery of rAAV to the Brain.

Recombinant adeno-associated viral (rAAV) vectors are a promising tool for therapeutic gene delivery to the brain. However, the delivery of rAAVs across the blood-brain barrier (BBB) and entry into the brain remains a major challenge for rAAV-based gene therapy. To circumvent this limitation, transcranial MRI-guided focused ultrasound (MRIgFUS) combined with intravenously injected microbubbles has been used to transiently and reversibly increase BBB permeability in targeted brain regions. Systemic administration of rAAVs at the time of sonication with focused ultrasound (FUS) facilitates the passage of rAAVs through the BBB and into the brain parenchyma. We and others have demonstrated that FUS-mediated rAAV delivery to the brain results in efficient transduction and transgene expression in vivo. Using this approach, the dose of intravenously injected rAAV variants that can cross the BBB can be reduced by 100 times, achieving significant transgene expression in the brain parenchyma with reduced peripheral transduction. Moreover, this strategy can be used to deliver rAAV variants that do not cross the BBB from the blood to selected brain regions. Here, we provide a detailed protocol for FUS-induced BBB permeability for targeted rAAV delivery to the brain of adult mice and rats.