Genetic Inactivation of CD33 in Hematopoietic Stem Cells to Enable CAR T Cell Immunotherapy for Acute Myeloid Leukemia.

The absence of cancer-restricted surface markers is a major impediment to antigen-specific immunotherapy using chimeric antigen receptor (CAR) T cells. For example, targeting the canonical myeloid marker CD33 in acute myeloid leukemia (AML) results in toxicity from destruction of normal myeloid cells. We hypothesized that a leukemia-specific antigen could be created by deleting CD33 from normal hematopoietic stem and progenitor cells (HSPCs), thereby generating a hematopoietic system resistant to CD33-targeted therapy and enabling specific targeting of AML with CAR T cells. We generated CD33-deficient human HSPCs and demonstrated normal engraftment and differentiation in immunodeficient mice. Autologous CD33 KO HSPC transplantation in rhesus macaques demonstrated long-term multilineage engraftment of gene-edited cells with normal myeloid function. CD33-deficient cells were impervious to CD33-targeting CAR T cells, allowing for efficient elimination of leukemia without myelotoxicity. These studies illuminate a novel approach to antigen-specific immunotherapy by genetically engineering the host to avoid on-target, off-tumor toxicity.

A RUNX1-FPDMM rhesus macaque model reproduces the human phenotype and predicts challenges to curative gene therapies.

Germ line loss-of-function heterozygous mutations in the RUNX1 gene cause familial platelet disorder with associated myeloid malignancies (FPDMM) characterized by thrombocytopenia and a life-long risk of hematological malignancies. Although gene therapies are being considered as promising therapeutic options, current preclinical models do not recapitulate the human phenotype and are unable to elucidate the relative fitness of mutation-corrected and RUNX1-heterozygous mutant hematopoietic stem and progenitor cells (HSPCs) in vivo long term. We generated a rhesus macaque with an FPDMM competitive repopulation model using CRISPR/Cas9 nonhomologous end joining editing in the RUNX1 gene and the AAVS1 safe-harbor control locus. We transplanted mixed populations of edited autologous HSPCs and tracked mutated allele frequencies in blood cells. In both animals, RUNX1-edited cells expanded over time compared with AAVS1-edited cells. Platelet counts remained below the normal range in the long term. Bone marrows developed megakaryocytic dysplasia similar to human FPDMM, and CD34+ HSPCs showed impaired in vitro megakaryocytic differentiation, with a striking defect in polyploidization. In conclusion, the lack of a competitive advantage for wildtype or control-edited HSPCs over RUNX1 heterozygous-mutated HSPCs long term in our preclinical model suggests that gene correction approaches for FPDMM will be challenging, particularly to reverse myelodysplastic syndrome/ acute myeloid leukemia predisposition and thrombopoietic defects.

A macaque clonal hematopoiesis model demonstrates expansion of TET2-disrupted clones and utility for testing interventions.

Individuals with age-related clonal hematopoiesis (CH) are at greater risk for hematologic malignancies and cardiovascular diseases. However, predictive preclinical animal models to recapitulate the spectrum of human CH are lacking. Through error-corrected sequencing of 56 human CH/myeloid malignancy genes, we identified natural CH driver mutations in aged rhesus macaques matching genes somatically mutated in human CH, with DNMT3A mutations being the most frequent. A CH model in young adult macaques was generated via autologous transplantation of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9-mediated gene-edited hematopoietic stem and progenitor cells (HSPCs), targeting the top human CH genes with loss-of-function (LOF) mutations. Long-term follow-up revealed reproducible and significant expansion of multiple HSPC clones with heterozygous TET2 LOF mutations, compared with minimal expansion of clones bearing other mutations. Although the blood counts of these CH macaques were normal, their bone marrows were hypercellular and myeloid-predominant. TET2-disrupted myeloid colony-forming units isolated from these animals showed a distinct hyperinflammatory gene expression profile compared with wild type. In addition, mature macrophages purified from the CH macaques showed elevated NLRP3 inflammasome activity and increased interleukin-1ß (IL-1ß) and IL-6 production. The model was used to test the impact of IL-6 blockage by tocilizumab, documenting a slowing of TET2-mutated expansion, suggesting that interruption of the IL-6 axis may remove the selective advantage of mutant HSPCs. These findings provide a model for examining the pathophysiology of CH and give insights into potential therapeutic interventions.

Velocity-selective excitation: Principles and applications.

Velocity-selective (VS) excitation is a relatively new type of excitation that can be useful for generating image contrast based on spin's motion. This review aims to explain the principles of VS excitation and their utilization for clinical applications. We first review the generalized excitation k-space formalism, which reveals a Fourier relationship between sequence parameters and excitation profiles for spins with arbitrary spatial location, off-resonance, and velocity. Based on the k-space framework, we analyze practical VS excitation pulse sequences that yield sinusoidal or sinc-shaped velocity profiles. Then we demonstrate how these two types of VS excitation can be used as magnetization preparation for clinical applications, including saturation- or inversion-based arterial spin labeling and black- or bright-blood angiography. We also discuss practical considerations and issues for each application, including the determination of design parameters and the effects of MR system errors, such as magnetic field offsets and eddy currents.

Spatially and velocity-selective magnetization preparation for noncontrast-enhanced peripheral MR angiography.

The purpose of the current study was to develop spatially and velocity-selective (SVS) magnetization preparation pulses for noncontrast-enhanced peripheral MR angiography (MRA) to provide comparisons with velocity-selective (VS) MRA with comparison to velocity-selective (VS). VS preparation pulses were designed by concatenating multiple excitation steps, each of which was a combination of a hard RF pulse, VS unipolar gradient pulses, and refocusing RF pulses. SVS preparation pulses were designed by replacing the hard RF pulse with a sinc-shaped RF pulse combined with a symmetric tripolar gradient pulse (which does not perturb the velocity encoding by the VS unipolar gradient pulses). Numerical simulations were performed to verify the intended hybrid excitation selectivity of SVS pulses taking account of tissue relaxation, magnetic field errors, and eddy currents. In vivo experiments were performed in healthy subjects to verify the hybrid excitation selectivity, as well as to demonstrate the visualization of the entire peripheral arteries using six-station protocols. As demonstrated by numerical simulations, SVS preparation yielded a notch-shaped longitudinal magnetization (Mz )-velocity response within the spatial stopband (the same as VS preparation) and preserved the Mz of spins outside the stopband, regardless of its velocity. We confirmed these observations also through in vivo tests with good agreement in normalized arterial and muscle signal intensities. In six-station peripheral MRA experiments, the proposed SVS-MRA yielded significantly higher arterial signal-to-noise ratio (SNR) (51.6 ± 14.3 vs. 38.9 ± 10.9; p < 0.001) and contrast-to-noise ratio (CNR) (41.2 ± 13.0 vs. 31.3 ± 10.5; p < 0.001) compared with VS-MRA. The proposed SVS-MRA improves arterial SNR and CNR compared with VS-MRA by mitigating undesired presaturation of arterial blood upstream the imaging field of view.

Prediction and validation of hematopoietic stem and progenitor cell off-target editing in transplanted rhesus macaques.

The programmable nuclease technology CRISPR-Cas9 has revolutionized gene editing in the last decade. Due to the risk of off-target editing, accurate and sensitive methods for off-target characterization are crucial prior to applying CRISPR-Cas9 therapeutically. Here, we utilized a rhesus macaque model to compare the predictive values of CIRCLE-seq, an in vitro off-target prediction method, with in silico prediction (ISP) based solely on genomic sequence comparisons. We use AmpliSeq HD error-corrected sequencing to validate off-target sites predicted by CIRCLE-seq and ISP for a CD33 guide RNA (gRNA) with thousands of off-target sites predicted by ISP and CIRCLE-seq. We found poor correlation between the sites predicted by the two methods. When almost 500 sites predicted by each method were analyzed by error-corrected sequencing of hematopoietic cells following transplantation, 19 off-target sites revealed insertion or deletion mutations. Of these sites, 8 were predicted by both methods, 8 by CIRCLE-seq only, and 3 by ISP only. The levels of cells with these off-target edits exhibited no expansion or abnormal behavior in vivo in animals followed for up to 2 years. In addition, we utilized an unbiased method termed CAST-seq to search for translocations between the on-target site and off-target sites present in animals following transplantation, detecting one specific translocation that persisted in blood cells for at least 1 year following transplantation. In conclusion, neither CIRCLE-seq or ISP predicted all sites, and a combination of careful gRNA design, followed by screening for predicted off-target sites in target cells by multiple methods, may be required for optimizing safety of clinical development.

Ensuring both velocity and spatial responses robust to B0/B1+ field inhomogeneities for velocity-selective arterial spin labeling through dynamic phase-cycling.

PURPOSE: To evaluate both velocity and spatial responses of velocity-selective arterial spin labeling (VS-ASL), using velocity-insensitive and velocity-compensated waveforms for control modules, as well as a novel dynamic phase-cycling approach, at different B0 / B1+ field inhomogeneities. METHODS: In the presence of imperfect refocusing, the mechanism of phase-cycling the refocusing pulses through four dynamics was first theoretically analyzed with the conventional velocity-selective saturation (VSS) pulse train. Numerical simulations were then deployed to compare the performance of the Fourier-transform based velocity-selective inversion (FT-VSI) with these three different schemes in terms of both velocity and spatial responses under various B0 / B1+ conditions. Phantom and human brain scans were performed to evaluate the three methods at B1+ scales of 0.8, 1.0, and 1.2. RESULTS: The simulations of FT-VSI showed that, under nonuniform B0 / B1+ conditions, the scheme with velocity-insensitive control was susceptible to DC bias of the static spins as systematic error, while the scheme with velocity-compensated control had deteriorated velocity-selective labeling profiles and, thus, reduced labeling efficiency. Through numerical simulation, phantom scans, and brain perfusion measurements, the dynamic phase-cycling method demonstrated considerable improvements over these issues. CONCLUSION: The proposed dynamic phase-cycling approach was demonstrated for the velocity-selective label and control modules with both velocity and spatial responses robust to a wide range of B0 and B1+ field inhomogeneities.

Brain MRI radiomics analysis may predict poor psychomotor outcome in preterm neonates.

OBJECTIVES: This study aimed to apply a radiomics approach to predict poor psychomotor development in preterm neonates using brain MRI. METHODS: Prospectively enrolled preterm neonates underwent brain MRI near or at term-equivalent age and neurodevelopment was assessed at a corrected age of 12 months. Two radiologists visually assessed the degree of white matter injury. The radiomics analysis on white matter was performed using T1-weighted images (T1WI) and T2-weighted images (T2WI). A total of 1906 features were extracted from the images and the minimum redundancy maximum relevance algorithm was used to select features. A prediction model for the binary classification of the psychomotor developmental index was developed and eightfold cross-validation was performed. The diagnostic performance of the model was evaluated using the AUC with and without including significant clinical and DTI parameters. RESULTS: A total of 46 preterm neonates (median gestational age, 29 weeks; 26 males) underwent brain MRI (median corrected gestational age, 37 weeks). Thirteen of 46 (28.3%) neonates showed poor psychomotor outcomes. There was one neonate among 46 with moderate to severe white matter injury on visual assessment. For the radiomics analysis, twenty features were selected for each analysis. The AUCs of prediction models based on T1WI, T2WI, and both T1WI and T2WI were 0.925, 0.834, and 0.902. Including gestational age or DTI parameters did not improve the prediction performance of T1WI. CONCLUSIONS: A radiomics analysis of white matter using early T1WI or T2WI could predict poor psychomotor outcomes in preterm neonates. KEY POINTS: • Radiomics analysis on T1-weighted images of preterm neonates showed the highest diagnostic performance (AUC, 0.925) for predicting poor psychomotor outcomes. • In spite of 45 of 46 neonates having no significant white matter injury on visual assessment, the radiomics analysis of early brain MRI showed good diagnostic performance (sensitivity, 84.6%; specificity, 78.8%) for predicting poor psychomotor outcomes. • Radiomics analysis on early brain MRI can help to predict poor neurodevelopmental outcomes in preterm neonates.

Non-contrast-enhanced abdominal MRA at 3 T using velocity-selective pulse trains.

PURPOSE: Most existing non-contrast-enhanced methods for abdominal MR arteriography rely on a spatially selective inversion (SSI) pulse with a delay to null both static tissue and venous blood, and are limited to small spatial coverage due to the sensitivity to slow arterial inflow. Velocity-selective inversion (VSI) based approach has been shown to preserve the arterial blood inside the imaging volume at 1.5 T. Recently, velocity-selective saturation (VSS) pulse trains were applied to suppress the static tissue and have been combined with SSI pulses for cerebral MR arteriography at 3 T. The aim of this study is to construct an abdominal MRA protocol with large spatial coverage at 3 T using advanced velocity-selective pulse trains. METHODS: Multiple velocity-selective MRA protocols with different sequence modules and 3D acquisition methods were evaluated. Sequences using VSS only as well as SSI+VSS and VSI+VSS preparations were then compared among a group of healthy young and middle-aged volunteers. Using MRA without any preparations as reference, relative signal ratios and relative contrast ratios of different vascular segments were quantitatively analyzed. RESULTS: Both SSI+VSS and VSI+VSS arteriograms achieved high artery-to-tissue and artery-to-vein relative contrast ratios above aortic bifurcation. The SSI+VSS sequence yielded lower signal at the bilateral iliac arteries than VSI+VSS, reflecting the benefit of the VSI preparation for imaging the distal branches. CONCLUSION: The feasibility of noncontrast 3D MR abdominal arteriography was demonstrated on healthy volunteers using a combination of VSS pulse trains and SSI or VSI pulse.

The impact of aging on primate hematopoiesis as interrogated by clonal tracking.

Age-associated changes in hematopoietic stem and progenitor cells (HSPCs) have been carefully documented in mouse models but poorly characterized in primates and humans. To investigate clinically relevant aspects of hematopoietic aging, we compared the clonal output of thousands of genetically barcoded HSPCs in aged vs young macaques after autologous transplantation. Aged macaques showed delayed emergence of output from multipotent (MP) clones, with persistence of lineage-biased clones for many months after engraftment. In contrast to murine aging models reporting persistence of myeloid-biased HSPCs, aged macaques demonstrated persistent output from both B-cell and myeloid-biased clones. Clonal expansions of MP, myeloid-biased, and B-biased clones occurred in aged macaques, providing a potential model for human clonal hematopoiesis of indeterminate prognosis. These results suggest that long-term MP HSPC output is impaired in aged macaques, resulting in differences in the kinetics and lineage reconstitution patterns between young and aged primates in an autologous transplantation setting.

Cerebral blood volume mapping using Fourier-transform-based velocity-selective saturation pulse trains.

PURPOSE: Velocity-selective saturation (VSS) pulse trains provide a viable alternative to the spatially selective methods for measuring cerebral blood volume (CBV) by reducing the sensitivity to arterial transit time. This study is to compare the Fourier-transform-based velocity-selective saturation (FT-VSS) pulse trains with the conventional flow-dephasing VSS techniques for CBV quantification. METHODS: The proposed FT-VSS label and control modules were compared with VSS pulse trains utilizing double refocused hyperbolic tangent (DRHT) and 8-segment B1-insensitive rotation (BIR-8). This was done using both numerical simulations and phantom studies to evaluate their sensitivities to gradient imperfections such as eddy currents. DRHT, BIR-8, and FT-VSS prepared CBV mapping was further compared for velocity-encoding gradients along 3 orthogonal directions in healthy subjects at 3T. RESULTS: The phantom studies exhibited more consistent immunity to gradient imperfections for the utilized FT-VSS pulse trains. Compared to DRHT and BIR-8, FT-VSS delivered more robust CBV results across the 3 VS encoding directions with significantly reduced artifacts along the superior-inferior direction and improved temporal signal-to-noise ratio (SNR) values. Average CBV values obtained from FT-VSS based sequences were 5.3 mL/100 g for gray matter and 2.3 mL/100 g for white matter, comparable to literature expectations. CONCLUSION: Absolute CBV quantification utilizing advanced FT-VSS pulse trains had several advantages over the existing approaches using flow-dephasing VSS modules. A greater immunity to gradient imperfections and the concurrent tissue background suppression of FT-VSS pulse trains enabled more robust CBV measurements and higher SNR than the conventional VSS pulse trains.

Unenhanced Velocity-Selective MR Angiography (VS-MRA): Initial Clinical Evaluation in Patients With Peripheral Artery Disease.

BACKGROUND: Safe and accurate imaging of the peripheral arterial system is important for diagnosis and treatment planning of patients with peripheral artery disease (PAD). PURPOSE: To evaluate image quality and diagnostic performance of unenhanced magnetic resonance angiography (MRA) based on velocity-selective (VS) magnetization preparation (termed VS-MRA). STUDY TYPE: Prospective. POPULATION: Thirty-one symptomatic PAD patients underwent VS-MRA. Twenty-four of them underwent clinical digital subtraction angiography (DSA) examination, 18.8 ± 5.2 days after the MR scans. FIELD STRENGTH/SEQUENCE: 1.5T MRI that included VS-MRA (homemade research sequence) and phase-contrast flow imaging (clinical sequence). ASSESSMENT: Image quality (0: nondiagnostic, 3: excellent) and stenosis severity (0: normal, 3: occlusion) of VS-MRA images were assessed independently by three reviewers. Arterial signal-to-noise-ratio (SNR) and artery-to-muscle contrast-to-noise ratio (CNR) were calculated. STATISTICAL TESTS: The sensitivity and specificity of VS-MRA were calculated for the detection of significant stenosis (>50%) with DSA as the reference standard. Interobserver agreement among the three reviewers was evaluated by using Cohen κ-statistics. RESULTS: The image quality score of VS-MRA was 2.7 ± 0.5 for Reader 1, 2.8 ± 0.5 for Reader 2, and 2.8 ± 0.4 for Reader 3; SNR and CNR were 37.8 ± 12.5 and 30.5 ± 11.8, respectively. Segment-based analysis revealed that VS-MRA had sensitivities of 85.3%, 74.5%, and 78.4%, respectively, for the three reviewers, and specificities of 93.5%, 96.8%, and 95.2%. The interobserver agreement for the stenosis grading was good, as demonstrated by Cohen κ values of 0.76 (Reader 1 vs. Reader 2), 0.82 (Reader 1 vs. Reader 3), and 0.79 (Reader 2 vs. Reader 3). DATA CONCLUSION: Unenhanced VS-MRA allows clear depiction of the peripheral arteries and accurate stenosis grading, as evidenced by high image quality scores and strong agreement with DSA. LEVEL OF EVIDENCE: 2 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2019;49:744-751.

Current Strategies to Enhance Adipose Stem Cell Function: An Update.

Mesenchymal stem cells (MSCs) emerged as a promising therapeutic tool targeting a variety of inflammatory disorders due to their multiple remarkable properties, such as superior immunomodulatory function and tissue-regenerative capacity. Although bone marrow (BM) is a dominant source for adult MSCs, increasing evidence suggests that adipose tissue-derived stem cells (ASCs), which can be easily obtained at a relatively high yield, have potent therapeutic advantages comparable with BM-MSCs. Despite its outstanding benefits in pre-clinical settings, the practical efficacy of ASCs remains controversial since clinical trials with ASC application often resulted in unsatisfactory outcomes. To overcome this challenge, scientists established several strategies to generate highly functional ASCs beyond the naïve cells, including (1) pre-conditioning of ASCs with various stimulants such as inflammatory agents, (2) genetic manipulation of ASCs and (3) modification of culture conditions with three-dimensional (3D) aggregate formation and hypoxic culture. Also, exosomes and other extracellular vesicles secreted from ASCs can be applied directly to recapitulate the beneficial performance of ASCs. This review summarizes the current strategies to improve the therapeutic features of ASCs for successful clinical implementation.

Characterization and suppression of stripe artifact in velocity-selective magnetization-prepared unenhanced MR angiography.

PURPOSE: To characterize and suppress stripe artifact associated with velocity-selective (VS) magnetization for unenhanced MRA. METHODS: Extended phase graph formalism was used to show that the stripe artifact contains multiples of the fundamental frequency that is determined by the area of unipolar VS gradient. Four VS preparation pulses whose excitation profiles are spatially shifted by quarter the fundamental period of the stripes, were applied alternately. For further suppression of the artifact, k-space data at kz = 0 were averaged over the 4 VS preparations. The proposed schemes were tested in a chicken breast phantom and healthy human subjects. RESULTS: When the standard VS preparation scheme was used, stripe artifact was shown in all the reconstructed images and appeared as artifactual peaks in k-space that corresponded to the first and second order harmonics of the fundamental frequency. Alternate application of the 4 phase-shifted VS preparation pulses suppressed the stripes, but not completely, as evidenced by residual erroneous peaks in k-space. After the k-space averaging, the stripe artifact was nearly eliminated. CONCLUSION: Stripe artifact in VS-MRA consists of multiples of the fundamental frequency and can be effectively suppressed through alternate application of phase-shifted VS preparations along with k-space averaging.

Whole-brain arteriography and venography: Using improved velocity-selective saturation pulse trains.

PURPOSE: To develop velocity-selective (VS) MR angiography (MRA) protocols for arteriography and venography with whole-brain coverage. METHODS: Tissue suppression using velocity-selective saturation (VSS) pulse trains is sensitive to radiofrequency field (B1 +) inhomogeneity. To reduce its sensitivity, we replaced the low-flip-angle hard pulses in the VSS pulse train with optimal composite (OCP) pulses. Additionally, new pulse sequences for arteriography and venography were developed by placing spatially selective inversion pulses with a delay to null signals from either venous or arterial blood. The VS MRA techniques were compared to the time-of-flight (TOF) MRA in six healthy subjects and two patients at 3T. RESULTS: More uniform suppression of stationary tissue was observed when the hard pulses were replaced by OCP pulses in the VSS pulse trains, which improved contrast ratios between blood vessels and tissue background for both arteries (0.87 vs. 0.77) and veins (0.80 vs. 0.59). Both arteriograms and venograms depicted all major cervical and intracranial arteries and veins, respectively. Compared to TOF MRA, VS MRA not only offers larger spatial coverage but also depicts more small vessels. Initial clinical feasibility was shown in two patients with comparisons to TOF protocols. CONCLUSION: Noncontrast-enhanced whole-brain arteriography and venography can be obtained without losing sensitivity to small vessel detection. Magn Reson Med 79:2014-2023, 2018. © 2017 International Society for Magnetic Resonance in Medicine.

Free breathing three-dimensional late gadolinium enhancement cardiovascular magnetic resonance using outer volume suppressed projection navigators.

PURPOSE: To develop a three-dimensional, free-breathing, late gadolinium enhancement (3D FB-LGE) cardiovascular magnetic resonance (CMR) technique, and to compare it with clinically used two-dimensional breath-hold LGE (2D BH-LGE). METHODS: The proposed 3D FB-LGE method consisted of inversion preparation, inversion delay, fat saturation, outer volume suppression, one-dimensional projection navigators, and a segmented stack of spirals acquisition. The 3D FB-LGE and 2D BH-LGE scans were performed on 29 cardiac patients. Qualitative analysis and quantitative analysis (in patients with scar) were performed. RESULTS: No significant differences were noted between the 3D FB-LGE and 2D BH-LGE data sets in terms of overall image quality score (2D: 4.69 ± 0.60 versus 3D: 4.55 ± 0.51, P = 0.46) and image artifact score (2D: 1.10 ± 0.31 versus 3D: 1.17 ± 0.38; P = 0.63). The average difference in fractional scar volume between the 3D and 2D methods was 1.9% (n = 5). Acquisition time was significantly shorter for the 3D FB-LGE over 2D BH-LGE by a factor of 2.83 ± 0.77 (P < 0.0001). CONCLUSIONS: The 3D FB-LGE is a viable option for patients, particularly in acute settings or in patients who are unable to comply with breath-hold instructions. Magn Reson Med 77:1533-1543, 2017. © 2016 International Society for Magnetic Resonance in Medicine.

Mesenchymal Stem Cell Therapy for Inflammatory Skin Diseases: Clinical Potential and Mode of Action.

Inflammatory skin disorders that cause serious deterioration of the quality of life have become one of the major public concerns. Despite their significance, there is no fundamental cure to date. Mesenchymal stem cells (MSCs) possess unique immunomodulatory properties which make them a promising tool for the treatment of various inflammatory diseases. Our recent preclinical and clinical studies have shown that MSCs can be successfully used for the treatment of atopic dermatitis (AD), one of the major inflammatory skin diseases. This observation along with similar reports from other groups revealed the efficacy and underlying mechanisms of MSCs in inflammatory dermatosis. In addition, it has been proposed that cell priming or gene transduction can be novel strategies for the development of next-generation high-efficacy MSCs for treating inflammatory skin diseases. We discuss here existing evidence that demonstrates the regulatory properties of MSCs on immune responses under inflammatory conditions.

Cathepsin S contributes to microglia-mediated olfactory dysfunction through the regulation of Cx3cl1-Cx3cr1 axis in a Niemann-Pick disease type C1 model.

Microglia can aggravate olfactory dysfunction by mediating neuronal death in the olfactory bulb (OB) of a murine model of Niemann-Pick disease type C1 (NPC1), a fatal neurodegenerative disorder accompanied by lipid trafficking defects. In this study, we focused on the crosstalk between neurons and microglia to elucidate the mechanisms underlying extensive microgliosis in the NPC1-affected brain. Microglia in the OB of NPC1 mice strongly expressed CX3C chemokine receptor 1 (Cx3cr1), a specific receptor for the neural chemokine C-X3-C motif ligand 1 (Cx3cl1). In addition, a high level of Cx3cl1 was detected in NPC1 mouse-derived CSF due to enhanced catalytic activity of Cathepsin S (Ctss), which is responsible for Cx3cl1 secretion. Notably, nasal delivery of Cx3cl1 neutralizing antibody or Ctss inhibitor could inhibit the Cx3cl1-Cx3cr1 interaction and support neuronal survival through the suppression of microglial activation, leading to an improvement in the olfactory function in NPC1 mice. Relevant in vitro experiments revealed that intracellular cholesterol accumulation could act as a strong inducer of abnormal Ctss activation and, in turn, stimulated the Cx3cl1-Cx3cr1 axis in microglia via p38 mitogen-activated protein kinase signaling. Our data address the significance of Cx3cl1-Cx3cr1 interaction in the development of microglial neurotoxicity and suggest that Ctss is a key upstream regulator. Therefore, this study contributes to a better understanding of the crosstalk between neurons and microglia in the development of the neurodegeneration and provides a new perspective for the management of olfactory deficits and other microglia-dependent neuropathies. GLIA 2016;64:2291-2305.

Noncontrast-enhanced peripheral venography using velocity-selective magnetization preparation and transient balanced SSFP.

PURPOSE: To develop a three-dimensional (3D) noncontrast-enhanced (NCE) peripheral magnetic resonance venography (MRV) method and demonstrate its feasibility in vivo. METHODS: The proposed MRV pulse sequence consisted of a velocity-selective (VS) inversion preparation module, inversion delay time (TI), fat inversion pulse, and 3D balanced steady-state free precession (bSSFP) dummy excitations and readout. The VS preparation module inverted arterial blood, which recovered close to zero magnetization during TI. The TI and the number of dummy excitations (Nnum ) were numerically optimized for maximizing vein-to-background contrast and tested in a healthy subject. The proposed MRV of the entire peripheral system, using four-station acquisition, was performed in six healthy subjects and three peripheral artery patients. RESULTS: The numerical optimization yielded TI = 350 ms and Ndum = 40, which was supported by the largest vein contrast among the parameters chosen around the optima on in vivo venograms. Four-station peripheral MRV using the optimized parameters well visualized all major deep veins with high vein-to-background contrast. The relative vein contrast ratios were 0.80 ± 0.08, 0.75 ± 0.07, and 0.84 ± 0.06 against the arteries, muscle, and fat, respectively. CONCLUSION: The proposed NCE MRV using VS preparation and transient bSSFP can generate high-contrast peripheral venograms directly with a single acquisition.