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Background: Cerebral cavernous malformations, also known as cavernous angiomas, are blood vessel abnormalities comprised of clusters of grossly enlarged and hemorrhage-prone capillaries. The prevalence in the general population, including asymptomatic cases, is estimated to be 0.5%. Some patients develop severe symptoms, including seizures and focal neurological deficits, whereas others remain asymptomatic. The causes of this remarkable presentation heterogeneity within a primarily monogenic disease remain poorly understood. Methods: We established a chronic mouse model of cerebral cavernous malformations, induced by postnatal ablation of Krit1 with Pdgfb-CreERT2 , and examined lesion progression in these mice with T2-weighted 7T magnetic resonance imaging (MRI). We also established a modified protocol for dynamic contrast-enhanced MRI and produced quantitative maps of gadolinium tracer gadobenate dimeglumine. After terminal imaging, brain slices were stained with antibodies against microglia, astrocytes, and endothelial cells. Results: These mice develop cerebral cavernous malformations lesions gradually over 4 to 5 months of age throughout the brain. Precise volumetric analysis of individual lesions revealed nonmonotonous behavior, with some lesions temporarily growing smaller. However, the cumulative lesional volume invariably increased over time and after about 2 months followed a power trend. Using dynamic contrast-enhanced MRI, we produced quantitative maps of gadolinium in the lesions, indicating a high degree of heterogeneity in lesional permeability. MRI properties of the lesions were correlated with cellular markers for endothelial cells, astrocytes, and microglia. Multivariate comparisons of MRI properties of the lesions with cellular markers for endothelial and glial cells revealed that increased cell density surrounding lesions correlates with stability, whereas denser vasculature within and surrounding the lesions may correlate with high permeability. Conclusions: Our results lay a foundation for better understanding individual lesion properties and provide a comprehensive preclinical platform for testing new drug and gene therapies for controlling cerebral cavernous malformations.

Cerebral cavernous malformations (CCM), also known as cavernous angiomas, are blood vessel abnormalities comprised of clusters of grossly enlarged and hemorrhage-prone capillaries. The prevalence in the general population, including asymptomatic cases, is estimated to be 0.5%. Some patients develop severe symptoms, including seizures and focal neurologic deficits, while others have no symptoms. The causes of this remarkable presentation heterogeneity within a primarily monogenic disease remain poorly understood. To address this problem, we have established a chronic mouse model of CCM, induced by postnatal ablation ofKrit1withPdgfb-CreERT. These mice develop CCM lesions gradually over 4-6 months of age throughout of the brain. We examined lesion progression in these mice with T2-weighted 7T MRI protocols. Precise volumetric analysis of individual lesions revealed non-monotonous behavior, with some lesions temporarily growing smaller. However, the cumulative lesional volume invariably increased over time and accelerated after about 3 months. Next, we established a modified protocol for dynamic contrast enhanced (DCE) MR imaging and produced quantitative maps of gadolinium tracer MultiHance in the lesions, indicating a high degree of heterogeneity in lesional permeability. Multivariate comparisons of MRI properties of the lesions with cellular markers for endothelial cells, astrocytes, and microglia revealed that increased cell density surrounding lesions correlates with stability, while increased vasculature within and surrounding lesions may correlate with instability. Our results lay a foundation for better understanding individual lesion properties and provide a comprehensive pre-clinical platform for testing new drug and gene therapies for controlling CCM.

Significance Focused ultrasound (FUS) is a targeted and noninvasive technique that can be used to activate gas-filled microbubbles (MBs) to oscillate within the bloodstream. This technique has been used previously to open the blood–brain barrier (BBB) to facilitate the delivery of therapeutics to the surrounding brain tissue. However, disruption of the BBB may be contraindicated in certain disease contexts. Here, we utilize low-pressure FUS to oscillate the MBs just enough to transfect endothelial cells, without opening the BBB. The low-pressure FUS regimen results in enhanced gene delivery to endothelial cells, with none of the inflammatory or immune pathway up-regulation observed at higher FUS pressures., Treatment of many pathologies of the brain could be improved markedly by the development of noninvasive therapeutic approaches that elicit robust, endothelial cell-selective gene expression in specific brain regions that are targeted under MR image guidance. While focused ultrasound (FUS) in conjunction with gas-filled microbubbles (MBs) has emerged as a noninvasive modality for MR image-guided gene delivery to the brain, it has been used exclusively to transiently disrupt the blood–brain barrier (BBB), which may induce a sterile inflammation response. Here, we introduce an MR image-guided FUS method that elicits endothelial-selective transfection of the cerebral vasculature (i.e., “sonoselective” transfection), without opening the BBB. We first determined that activating circulating, cationic plasmid-bearing MBs with pulsed low-pressure (0.1 MPa) 1.1-MHz FUS facilitates sonoselective gene delivery to the endothelium without MRI-detectable disruption of the BBB. The degree of endothelial selectivity varied inversely with the FUS pressure, with higher pressures (i.e., 0.3-MPa and 0.4-MPa FUS) consistently inducing BBB opening and extravascular transfection. Bulk RNA sequencing analyses revealed that the sonoselective low-pressure regimen does not up-regulate inflammatory or immune responses. Single-cell RNA sequencing indicated that the transcriptome of sonoselectively transfected brain endothelium was unaffected by the treatment. The approach developed here permits targeted gene delivery to blood vessels and could be used to promote angiogenesis, release endothelial cell-secreted factors to stimulate nerve regrowth, or recruit neural stem cells.

The blood brain barrier (BBB) plays a critically important role in the regulation of central nervous system (CNS) homeostasis, but also represents a major limitation to treatments of brain pathologies. In recent years, focused ultrasound (FUS) in conjunction with gas-filled microbubble contrast agents has emerged as a powerful tool for transiently and non-invasively disrupting the BBB in a targeted and image-guided manner, allowing for localized delivery of drugs, genes, or other therapeutic agents. Beyond the delivery of known therapeutics, FUS-mediated BBB opening also demonstrates the potential for use in neuromodulation and the stimulation of a range of cell- and tissue-level physiological responses that may prove beneficial in disease contexts. Clinical trials investigating the safety and efficacy of FUS-mediated BBB opening are well underway, and offer promising non-surgical approaches to treatment of devastating pathologies. This article reviews a range of pre-clinical and clinical studies demonstrating the tremendous potential of FUS to fundamentally change the paradigm of treatment for CNS diseases.

Children with autism spectrum disorders often display dysregulated immune responses and related gastrointestinal symptoms. However, the underlying mechanisms leading to the development of both phenotypes have not been elucidated. Here, we showed that mouse offspring exhibiting autism-like phenotypes due to prenatal exposure to maternal inflammation were more susceptible to developing intestinal inflammation following challenges later in life. In contrast to its prenatal role in neurodevelopmental phenotypes, interleukin-17A (IL-17A) generated immune-primed phenotypes in offspring through changes in the maternal gut microbiota that led to postnatal alterations in the chromatin landscape of naïve CD4(+) T cells. The transfer of stool samples from pregnant mice with enhanced IL-17A responses into germ-free dams produced immune-primed phenotypes in offspring. Our study provides mechanistic insights into why children exposed to heightened inflammation in the womb might have an increased risk of developing inflammatory diseases in addition to neurodevelopmental disorders.