Labeling Imaging Research Articles

Associations of Cerebral Blood Flow Patterns With Gray and White Matter Structure in Patients With Temporal Lobe Epilepsy.

Neuroimaging studies in patients with temporal lobe epilepsy (TLE) show widespread brain network alterations beyond the mesiotemporal lobe. Despite the critical role of the cerebrovascular system in maintaining whole-brain structure and function, changes in cerebral blood flow (CBF) remain incompletely understood in the disease. Here, we studied whole-brain perfusion and vascular network alterations in TLE and assessed its associations with gray and white matter compromises and various clinical variables. We included individuals with and without pharmaco-resistant TLE who underwent multimodal 3T MRI, including arterial spin labelling, structural, and diffusion-weighted imaging. Using surface-based MRI mapping, we generated individualized cortico-subcortical profiles of perfusion, morphology, and microstructure. Linear models compared regional CBF in patients with controls and related alterations to morphological and microstructural metrics. We further probed interregional vascular networks in TLE, using graph theoretical CBF covariance analysis. The effects of disease duration were explored to better understand the progressive changes in perfusion. We assessed the utility of perfusion in separating patients with TLE from controls using supervised machine learning. Compared with control participants (n = 38; mean ± SD age 34.8 ± 9.3 years; 20 females), patients with TLE (n = 24; mean ± SD age 35.8 ± 10.6 years; 12 females) showed widespread CBF reductions predominantly in fronto-temporal regions (Cohen d -0.69, 95% CI -1.21 to -0.16), consistent in a subgroup of patients who remained seizure-free after surgical resection of the seizure focus. Parallel structural profiling and network-based models showed that cerebral hypoperfusion may be partially constrained by gray and white matter changes (8.11% reduction in Cohen d) and topologically segregated from whole-brain perfusion networks (area under the curve -0.17, p < 0.05). Negative effects of progressive disease duration further targeted regional CBF profiles in patients (r = -0.54, 95% CI -0.77 to -0.16). Perfusion-derived classifiers discriminated patients from controls with high accuracy (71% [70%-82%]). Findings were robust when controlling for several methodological confounds. Our multimodal findings provide insights into vascular contributions to TLE pathophysiology affecting and extending beyond mesiotemporal structures and highlight their clinical potential in epilepsy diagnosis. As our work was cross-sectional and based on a single site, it motivates future longitudinal studies to confirm progressive effects, ideally in a multicentric setting.

Cerebral blood flow and structural connectivity after working memory or physical training in paediatric cancer survivors – Exploratory findings

ABSTRACT Paediatric cancer survivors often suffer from cognitive long-term difficulties. Consequently, strengthening cognition is of major clinical relevance. This study investigated cerebral changes in relation to cognition in non-brain tumour paediatric cancer survivors after working memory or physical training compared to a control group. Thirty-four children (≥one-year post-treatment) either underwent eight weeks of working memory training (n = 10), physical training (n = 11), or a waiting period (n = 13). Cognition and MRI, including arterial spin labelling and diffusion tensor imaging, were assessed at three time points (baseline, post-training, and three-month follow-up). Results show lower cerebral blood flow immediately after working memory training (z = −2.073, p = .038) and higher structural connectivity at the three-month follow-up (z = −2.240, p = .025). No cerebral changes occurred after physical training. Short-term changes in cerebral blood flow correlated with short-term changes in cognitive flexibility (r = −.667, p = .049), while long-term changes in structural connectivity correlated with long-term changes in working memory (r = .786, p = .021). Despite the caution given when interpreting data from small samples, this study suggests a link between working memory training and neurophysiological changes. Further research is needed to validate these findings.

Comparison of arterial spin labeling and dynamic susceptibility contrast perfusion MR imaging in pediatric brain tumors: A systematic review and meta-analysis.

Brain tumors are a leading cause of mortality in children. Accurate tumor grading is essential to plan treatment and for prognostication. Perfusion imaging has been shown to correlate well with tumor grade in adults, however there are fewer studies in pediatric patients. Moreover, there is no consensus regarding which MR perfusion technique demonstrates the highest accuracy in the latter population. To compare the diagnostic test accuracy of dynamic-susceptibility contrast and arterial spin-labelling, in their ability to differentiate between low-and high-grade pediatric brain tumors at first presentation. Articles were retrieved from online electronic databases: MEDLINE (Ovid), Web of Science Core Collection and SCOPUS. Studies in pediatric patients with a treatment naïve diagnosed brain tumor and imaging including either ASL or DSC or both, together with a histological diagnosis were included. Studies involving adult patient or mixed age populations, studies with incomplete data and those which used dynamic contrast enhanced perfusion were excluded. The sensitivities and specificities obtained from each study were used to calculate the true-positive, true-negative, false-positive, and false-negative count. A case was defined as a histologically proven high-grade tumor. The random-effect model was used to merge statistics. Significance level was set at p < 0.05. Forest plots showing pairs of sensitivity and specificity, with their 95% confidence intervals, were constructed for each study. The bivariate model was applied in order to account for between-study variability. The SROC plots were constructed from the obtained data-sets. The AUC for the SROC of all studies was estimated to determine the overall diagnostic test accuracy of perfusion MRI, followed by a separate comparison of the SROC of ASL versus DSC studies. Small and heterogenous sample size. The diagnostic accuracy of ASL was found to be comparable and not inferior to DSC, thus its use in the diagnostic assessment of pediatric patients should continue to be supported. ASL = arterial spin labelling, DSC = dynamic susceptibility contrast, DCE = dynamic contrast-enhanced, rCBF = relative cerebral blood flow, rCBV = relative cerebral blood volume, MTT = mean transfer time, TR = repetition time, TE = echo time, SROC = summary receiver operating characteristics, HG= high-grade, LG = low-grade, AUC = area under the curve, PRISMA = Preferred Reporting Items for Systematic Reviews and Meta-Analyses.

CLINICAL APPLICATION STUDY OF 3D-ASL PERFUSION IMAGING AND MAGNETIC RESONANCE DIFFUSION IMAGING IN TRANSIENT ISCHEMIC ATTACK.

Objective: This study aimed to explore the clinical application of three-dimensional arterial spin labeling (3D-ASL) and diffusion-weighted magnetic resonance imaging (DWI) in transient ischemic attacks. Methods: Forty patients with transient cerebral ischemia in our hospital were selected and included from July 2020 to March 2022. All subjects were detected by DWI and 3D-ASL technology. The positive rate, relative cerebral blood flow (rCBF), and the receiver operating characteristic curve of the two methods in the diagnosis of transient cerebral ischemia were compared; the objective was to compare the relationship between the frequency of transient ischemic attack and hypoperfusion, and vascular stenosis. Results: The 3D-ASL examination showed two cases of hypoperfusion in the healthy control group (5.00), and the magnetic resonance imaging examination showed four cases of vascular stenosis in the healthy control group (10.00). The rCBF ratio in the cerebral ischemia group was significantly lower than that in the cerebral ischemia group, which was significantly lower than that in the healthy control group ( P < 0.05). The area under the curve (AUC) of 3D-ASL in the diagnosis of transient cerebral ischemia was 0.800, and the AUC of DWI in the diagnosis of transient cerebral ischemia was 0.725. The AUC of the combination of the two methods in transient cerebral ischemia was 0.850. There was a significant difference in the attack frequency of patients with transient cerebral ischemia with different perfusion ( P < 0.05). There was a significant difference in attack frequency between patients with transient ischemic attack and patients without vascular stenosis ( P < 0.05). Conclusion: 3D-ASL and DWI technology have higher diagnostic efficiency for transient cerebral ischemia.

Changes of cerebral structure and perfusion in subtypes of systemic sclerosis: a brain magnetic resonance imaging study.

The characteristics of brain impairment in different subtypes of systemic sclerosis (SSc) (dcSSc, diffuse cutaneous SSc; lcSSc, limited cutaneous SSc) remain unclear. This study aimed to characterize cerebral structure and perfusion changes in different subtype of SSc patients using magnetic resonance (MR) imaging. Seventy SSc patients (46.0 ± 11.7 years, 62 females) and 30 healthy volunteers (44.8 ± 13.7 years, 24 females) were recruited and underwent brain MR imaging and Montreal Cognitive Assessment (MoCA) test. Gray matter (GM) volumes were measured using voxel-based morphometry analysis on T1-weighted images. Voxel-based and regional cerebral blood flow (CBF) was calculated on arterial spin labelling images. The cerebral structural and perfusion measurements by MR imaging were compared among dcSSc, lcSSc and healthy subjects using one-way ANOVA. The correlations between clinical characteristics and MR imaging measurements were also analyzed. The dcSSc patients exhibited a significant reduction in GM volume in the para-hippocampal region (cluster p < 0.01, FWE corrected) compared with healthy volunteers. Whereas, SSc patients, particularly lcSSc patients, showed elevated CBF in cerebellum, insula, cerebral cortex, and subcortical structures (regional analyses: all p < 0.05; voxel-based analyses: cluster p < 0.01, FWE corrected). Furthermore, clinical characteristics of modified Rodnan skin score (mRSS) (r value ranged from -0.29 to -0.45), MoCA scores (r = 0.40) and antinuclear antibody (ANA) positivity (r=-0.33) were significantly associated with CBF in some regions (all p < 0.05). The manifestations of brain involvement vary among different subtypes of SSc. In addition, severe skin sclerosis may indicate higher risk of brain involvement in SSc patients.

MR-conditionality failure modes: a comparison across various spinal cord stimulators.

As spinal cord stimulation (SCS) becomes a staple of chronic pain management, the SCS industry must constantly evolve to ensure safety, convenience and enhanced efficacy. Beyond waveforms and size, MR-conditionality is a key differentiator sought out by physicians and patients when choosing SCS devices. Many common SCS complications, including lead migration, can affect the MR-conditionality. The authors reviewed literature published between 2015 and 2024 using PubMed and Google Scholar databases, as well as USFDA labels for magnetic resonance imaging and technical manuals with further confirmation from local representatives. Through extensive review of the literature and direct extraction from each SCS device company, the authors aimed to investigate the specific terms of MR-conditionality under various circumstances. This article provides a collective reference of the MR-conditionality of the currently available SCS devices under normal conditions, as well as with common failure modes, such as lead migration, lead fracture and battery detachment.

An arginine-rich nuclear localization signal (ArgiNLS) strategy for streamlined image segmentation of single cells

High-throughput volumetric fluorescent microscopy pipelines can spatially integrate whole-brain structure and function at the foundational level of single cells. However, conventional fluorescent protein (FP) modifications used to discriminate single cells possess limited efficacy or are detrimental to cellular health. Here, we introduce a synthetic and nondeleterious nuclear localization signal (NLS) tag strategy, called "Arginine-rich NLS" (ArgiNLS), that optimizes genetic labeling and downstream image segmentation of single cells by restricting FP localization near-exclusively in the nucleus through a poly-arginine mechanism. A single N-terminal ArgiNLS tag provides modular nuclear restriction consistently across spectrally separate FP variants. ArgiNLS performance in vivo displays functional conservation across major cortical cell classes and in response to both local and systemic brain-wide AAV administration. Crucially, the high signal-to-noise ratio afforded by ArgiNLS enhances machine learning-automated segmentation of single cells due to rapid classifier training and enrichment of labeled cell detection within 2D brain sections or 3D volumetric whole-brain image datasets, derived from both staining-amplified and native signal. This genetic strategy provides a simple and flexible basis for precise image segmentation of genetically labeled single cells at scale and paired with behavioral procedures.

A Water-Soluble Multifunctional Probe for Colorimetric Copper Sensing, Lysosome Labelling and Live-Cell Imaging.

We report a water-soluble fluorescence and colorimetric copper probe (LysoBC1); this system can also serve for lysosome labeling and for the dynamic tracking of Cu2+ in living cells. The sensing mechanism takes advantage of the synergic action by the following three components: i) a lysosome targeting unit, ii) the spirolactam ring-opening for the selective copper chelation and iii) the metal-mediated hydrolysis of the rhodamine moiety for fluorescence enhancement. In aqueous environment the molecule acts as a fluorescent reversible pH sensor and as colorimetric probe for Cu2+ at physiological pH; the hydrolysis of the copper targeting unit resulted in a 50-fold increase of the fluorescence intensity. Most importantly, in vitro cell analyses in undifferentiated (SH SY5Y) and differentiated (d-SH SY5Y) neuroblastoma cells, LysoBC1 is able to selectively accumulate into lysosome while the copper binding ability allowed us to monitor intracellular copper accumulation into lysosome.

Nanoscale organization is changed in native, surface AMPARs by mouse brain region and tauopathy.

Synaptic AMPA receptors (AMPARs) on neuronal plasma membranes are correlated with learning and memory. Using a unique labeling and super-resolution imaging, we have visualized the nanoscale synaptic and extra-synaptic organization of native surface AMPARs for the first time in mouse brain slices as a function of brain region and tauopathy. We find that the fraction of surface AMPARs organized in synaptic clusters is two-times smaller in the hippocampus compared to the motor and somatosensory cortex. In 6 months old PS19 model of tauopathy, synaptic and extrasynaptic distributions are disrupted in the hippocampus but not in the cortex. Thus, this optimized super-resolution imaging tool allows us to observe synaptic deterioration at the onset of tauopathy before apparent neurodegeneration.

A multi-task generative model for simultaneous post-contrast MR image synthesis and brainstem glioma segmentation

ObjectivesThis study aims to generate post-contrast MR images reducing the exposure of gadolinium-based contrast agents (GBCAs) for brainstem glioma (BSG) detection, simultaneously delineating the BSG lesion, and providing high-resolution contrast information. MethodsA retrospective cohort of 30 patients diagnosed with brainstem glioma was included. Multi-contrast images, including pre-contrast T1 weighted (pre-T1w), T2 weighted (T2w), arterial spin labeling (ASL) and post-contrast T1w images, were collected. A multi-task generative model was developed to synthesize post-contrast T1w images and simultaneously segment BSG masks from the multi-contrast inputs. Performance evaluation was conducted using peak signal-to-noise ratio (PSNR), structural similarity index (SSIM), and mean absolute error (MAE) metrics. A perceptual study was also undertaken to assess diagnostic quality. ResultsThe proposed model achieved SSIM of 0.86 ± 0.04, PSNR of 26.33 ± 0.05 and MAE of 57.20 ± 20.50 for post-contrast T1w image synthesis. Automated delineation of the BSG lesions achieved Dice similarity coefficient (DSC) score of 0.88 ± 0.27. ConclusionsThe proposed model can synthesize high-quality post-contrast T1w images and accurately segment the BSG region, yielding satisfactory DSC scores. Clinical relevance statementThe synthesized post-contrast MR image presented in this study has the potential to reduce the usage of gadolinium-based contrast agents, which may pose risks to patients. Moreover, the automated segmentation method proposed in this paper aids radiologists in accurately identifying the brainstem glioma lesion, facilitating the diagnostic process.

GSH-AuNCs@ZIF-8 as chemiluminescence imaging label for high-throughput and ultrasensitive detection of THC and THC-COOH

Traditional quantitation methods for Δ9-tetrahydrocannabinol (THC) require multi-step sample pretreatment, specialized instruments and trained operators. There is a demand for a simple, rapid, reliable, and sensitive method for detecting THC. Herein, a novel chemiluminescence (CL) imaging protocol was developed for ultrasensitive and high-throughput determination of Δ9-tetrahydrocannabinol (THC) and its metabolite THC-COOH, as CL imaging offers advantages such as free of external light source, high sensitivity, and ease of operation. GSH-AuNCs@ZIF-8 with strong CL catalytical property was prepared by encapsulating glutathione-modified gold nanoclusters (GSH-AuNCs) into zeolitic imidazolate frameworks-8 (ZIF-8). The integration of GSH-AuNCs@ZIF-8 with a disposable immunosensor array resulted in impressively low limits of detection (down to 0.04 ng mL−1 and 0.09 ng mL−1 for THC and THC-COOH, respectively) and high throughput (90 tests h−1). When applied to rat plasma samples, it exhibited satisfactory recoveries (89.68 %-104.54 %) and yielded results consistent with those obtained using the standard method. With high sensitivity and throughput, acceptable precision, and straightforward operation, the proposed protocol demonstrated good applicability for rapid, on-site detection, and law enforcement regulation of THC. Moreover, the protocol explored the chemiluminescence property of GSH-AuNCs@ZIF-8 and utilized it as a CL imaging label, opening new directions for the application of MOF composites.

The effect of post-labeling delay on cerebral blood flow is influenced by age and sex: a study based on arterial spin-labeling magnetic resonance imaging.

Whether the effect of post-labeling delay (PLD) on cerebral blood flow (CBF) is influenced by age and sex in adults is unknown. In this study, we mainly aimed to explore the potential influence of age and sex on the effect of PLD on CBF. This prospective study enrolled 90 healthy adult volunteers (49.47±15.63 years of age; age range, 20-77 years; 47 female; 43 male). All participants underwent 3-dimensional (3D) pseudo-continuous arterial spin labeling (ASL) imaging with 3 different PLDs (1,525, 2,025, and 2,525 ms). The CBF values for each PLD, the arterial transit time (ATT), and the spatial coefficient of variation (spatial CoV) were computed for 21 regions of interest (ROIs) in every participant. Multivariate regression analysis was conducted to assess the potential influence of age and sex on the effect of PLD on CBF and the relationships among CBF, ATT, PLD, age, sex, and spatial CoV. The CBF increased for 7.32 to 9.87 mL/100 g/min as the PLD increased per 1 second in the global gray matter, bilateral frontal, temporal lobes, the vascular territories of bilateral anterior and middle carotid artery. When the age increased per 1 year, the speed of the changes for CBF decreased for 0.26 to 0.3 mL/100 g/min/s in these regions. However, the CBF decreased for 12 to 17 mL/100 g/min as the PLD increased per 1 second in the bilateral limbic lobes, insula, and deep gray matter. In these regions, the speed of the changes for CBF increased for 0.2 to 0.28 mL/100 g/min/s as the age increased per 1 year. Furthermore, compared to the female, the speed of the changes for CBF decreased for 3.58 to 4.6 mL/100 g/min/s for the male in global gray matter, bilateral frontal, limbic lobes, and the vascular territories of bilateral anterior carotid artery, and the speed increased 4.49 to 5.09 mL/100 g/min/s for the male in the limbic lobes. In addition, the CBF decreased with aging and the CBF tended to be higher in females compared to males. At the same time, we found that the ATT of all ROIs increased with age and manifested higher in males than females. Moreover, we found that CBF decreased with the increase of ATT, and the effect of ATT on CBF was less influenced by PLD. Finally, we found that the spatial CoV of ASL in certain regions increased with the increase of ATT and age, and was greater in males. The effect of PLD on CBF can be influenced by age and sex. The relationships among CBF, ATT, PLD, age, sex, and spatial CoV found in this study may have certain significance for the study of ASL imaging in the future.

Deep 3D histology powered by tissue clearing, omics and AI.

To comprehensively understand tissue and organism physiology and pathophysiology, it is essential to create complete three-dimensional (3D) cellular maps. These maps require structural data, such as the 3D configuration and positioning of tissues and cells, and molecular data on the constitution of each cell, spanning from the DNA sequence to protein expression. While single-cell transcriptomics is illuminating the cellular and molecular diversity across species and tissues, the 3D spatial context of these molecular data is often overlooked. Here, I discuss emerging 3D tissue histology techniques that add the missing third spatial dimension to biomedical research. Through innovations in tissue-clearing chemistry, labeling and volumetric imaging that enhance 3D reconstructions and their synergy with molecular techniques, these technologies will provide detailed blueprints of entire organs or organisms at the cellular level. Machine learning, especially deep learning, will be essential for extracting meaningful insights from the vast data. Further development of integrated structural, molecular and computational methods will unlock the full potential of next-generation 3D histology.

Chemical imaging unveils mitochondria as the major site of medicinal biguanide accumulation within cells

Metformin (MET), a commonly prescribed medication for managing type 2 diabetes, has demonstrated various beneficial effects beyond its primary anti-diabetic efficacy. However, the mechanism underlying MET activity and its distribution within organelles remain largely unknown. In this study, we integrate multiple technologies, including chemical labeling, immunostaining, and high-resolution microscopy imaging, to visualize the accumulation of MET in organelles of cultured cells. To achieve this objective, an alkynylated MET probe is developed that preserves biological activity similar to biguanide drugs. As determined by biorthogonal chemical labeling and imaging, the MET probe selectively localizes to substructures within cells, contrasting with its probe control. Furthermore, the MET probe can be competitively and efficiently washed out through biguanide administration, demonstrating the specific activity of this probe in monitoring the cellular dynamics of biguanide drugs. Our results indicate that the MET probe can reach near-saturated concentrations within 2 h and is rapidly eliminated within an additional 2 h once the exogenous source of the drug is removed. Furthermore, we reveal that the MET probe primarily accumulates in mitochondria, particularly within the mitochondrial matrix, and has a minor presence in other organelles, such as lysosomes and endosomes. Together, this study provides the first view of the subcellular localization of MET and lays the foundation for future investigations on its molecular targets and mechanisms of action in promoting human health.

Fluorescent Lateral Flow Immunoassay Based on Quantum Dots Nanobeads.

Quantum dots, also known as semiconductor nanocrystals, are novel fluorescent labels for biological imaging and sensing. However, quantum dot-antibody conjugates with small dimensions (~10 nm), prepared through laborious purification procedures, exhibit limited sensitivity in detecting certain trace disease markers using lateral flow immunoassay strips. Herein, we present a method for the preparation of quantum dot nanobeads (QDNB) using a one-step emulsion evaporation method. Using the as-prepared QDNB, a fluorescent lateral flow immunoassay was fabricated to detect disease biomarkers using C-reactive protein (CRP) as an example. Unlike single quantum dot nanoparticles, quantum dot nanobead-antibody conjugates are more sensitive as immunoassay labels due to signal amplification by encapsulating hundreds of quantum dots in one polymer composite nanobead. Moreover, the larger size of QDNBs facilitates easier centrifugation separation when conjugating QDNBs with antibodies. The fluorescent lateral flow immunoassay based on QDNBs was fabricated, and the CRP concentration in the sample was measured in 15 min. The test results can be qualitatively assessed under UV light illumination and quantitatively measured using a fluorescent reader within 15 min.

Insomnia in patients with MRI-negative epilepsy: The associated factors and 3D-pCASL cerebral blood flow perfusion changes

ObjectiveThis study aimed to identify the factors associated with insomnia in MRI-negative epilepsy and uncover the underlying pathological mechanism driving insomnia within the context of epilepsy. MethodsWe conducted a retrospective study of patients with MRI-negative epilepsy recruited consecutively from December 2021 to December 2022. All subjects completed the Insomnia Severity Index (ISI), Epworth Sleepiness Scale (ESS), Self-rating Anxiety Scale (SAS), and Self-rating Depression Scale (SDS). Additionally, some subjects underwent the three-dimensional pseudo continuous arterial spin labeling(3D-pCASL) imaging examination. Bilateral frontal lobe, temporal lobe, hippocampus, thalamus, amygdala, caudate nucleus and lenticular nucleus were selected as regions of interest(ROI) and cerebral blood flow(CBF) values were measured in these regions. Subjects were classified into insomnia (ISI ≥ 10) or non-insomnia (ISI < 10) groups, and univariate and stepwise logistic regression analyses were employed to identify the factors associated with insomnia. Furthermore, CBF values in each ROI were compared between the two groups to identify the brain regions potentially related to the underlying pathological mechanism of insomnia in epilepsy. ResultsA total of 73 patients with MRI-negative epilepsy were recruited in this study(men, 49.3 %). Among them, 14 patients(19.2 %) had insomnia. Univariate regression revealed that nocturnal seizures, number of anti-seizure medication(ASM), anxiety, use of valproic acid(VPA), depression, and excessive daytime sleepiness(EDS) may be associated with insomnia in MRI-negative epilepsy (all p＜0.05). Stepwise regression demonstrated that nocturnal seizures, anxiety, and EDS were independently associated with insomnia in MRI-negative epilepsy (OR[95 %CI]P: 14.64[2.02–106.27]0.008,49.35[3.06–796.61]0.006, 13.28[1.25–140.66]0.032, respectively). Furthermore, CBF values in the left amygdala were significantly lower in patients with MRI- negative epilepsy who had insomnia. ConclusionThe prevalence of insomnia in MRI-negative epilepsy is 19.2%. Nocturnal seizures, anxiety, and EDS were independently associated with insomnia in MRI-negative epilepsy. The noteworthy decrease in CBF values in the left amygdala might be connected to the underlying pathological mechanism of insomnia in epilepsy.

In vivo Labeling and Intravital Imaging of Bacterial Infection using a Near-infrared Fluorescent D-Amino Acid Probe.

Bacterial infections still pose a severe threat to public health, necessitating novel tools for real-time analysis of microbial behaviors in living organisms. While genetically engineered strains with fluorescent or luminescent reporters are commonly used in tracking bacteria, their in vivo uses are often limited. Here, we report a near-infrared fluorescent D-amino acid (FDAA) probe, Cy7ADA, for in situ labeling and intravital imaging of bacterial infections in mice. Cy7ADA probe effectively labels various bacteria in vitro and pathogenic Staphylococcus aureus in mice after intraperitoneal injection. Because of Cy7's high tissue penetration and the quick excretion of free probes via urine, real-time visualization of the pathogens in a liver abscess model via intravital confocal microscopy is achieved. The biodistributions, including their intracellular localization within Kupffer cells, are revealed. Monitoring bacterial responses to antibiotics also demonstrates Cy7ADA's capability to reflect the bacterial load dynamics within the host. Furthermore, Cy7ADA facilitates three-dimensional pathogen imaging in tissue-cleared liver samples, showcasing its potential for studying the biogeography of microbes in different organs. Integrating near-infrared FDAA probes with intravital microscopy holds promise for wide applications in studying bacterial infections in vivo.

DIPG-73. FOCUSED ULTRASOUND FOR TREATMENT OF CHILDREN DIAGNOSED WITH DIFFUSE MIDLINE GLIOMAS

Abstract BACKGROUND Diffuse midline glioma (DMG) is a highly aggressive and fatal brain tumor that predominantly impacts children and young adults. The infiltrative pattern and rapid growth characteristics of DMG causes a significant challenge in finding an effective treatment. Additionally, the blood-brain barrier (BBB) poses a significant challenge by restricting the passage of therapeutic drugs to tumor bed. Currently the established standard of care is radiotherapy (RT), which is transitory. Thus there is a need for improved treatment options. OBJECTIVE Focused ultrasound (FUS) is an advancing non-invasive technology that has been used to open BBB and enhance drug delivery. We hypothesize that a combination therapy of FUS with pharmacological agents will be an effective and non-invasive treatment option for children diagnosed with DMG. METHOD Syngeneic DMG murine models were used to study the combination therapy of FUS with ONC201, FUS with anti-PD1 and FUS with RT. FUS mediated BBB opening was monitored with contrast-enhanced T1-weighted MRI. FUS-mediated drug delivery was validated with western blot, assessment of ROS, and optical label imaging. Kaplan Meier analysis was used to assess survival benefits. Single cell RNA-sequencing (scRNA-Seq) analysis is being done to assess the treatment effects of FUS and RT combination therapy. RESULTS/CONCLUSION We found that FUS and ONC201 combination therapy resulted in a decrease in NDUFA12 protein (a biomarker of ONC201 response) and an increase in ROS production compared to ONC201 monotherapy. For FUS and anti-PD1 combination therapy, an increased anti-PD1 delivery to the brainstem was detected. The overall survival of mice treated with FUS and anti-PD1 combination therapy increased when compared to anti-PD1 alone. Our preliminary scRNA-Seq analysis showed that combination therapy of FUS and RT induces innate immune response compared to the untreated group. Combination therapy with focused ultrasound shows a promising outcome by increasing the efficacy of therapeutic drugs and immunotherapy.

Multiplexed MicroRNA biomarker detection by bridging lifetime filtering imaging and dynamic chemical labeling

MicroRNAs (miRs) have emerged as promising biomarkers for early disease diagnosis and personalized treatment monitoring. However, their clinical utility has been hampered by technical limitations. Dynamic chemical labeling (DCL) based on capturing abasic PNA probes and reactive nucleobases, known as SMART bases, is a PCR-free approach that has proven very useful for the direct interrogation of circulating miRs. In this work, we expand the palette of tools available for the detection of DCL miR by synthesizing a new SMART nucleobase called SMART-C-Eu. This nucleobase contains a stable lanthanide cryptate. Using this SMART-C-Eu base and time-gated (TG) luminescence imaging, we successfully detect and quantify miR-122–5p in human serum samples. miR-122–5p is a well-known biomarker for drug-induced liver injury. Through a bead-counting analysis approach, statistical robustness is improved and miR-122–5p concentrations are detected in the nanomolar range. Furthermore, we extend this approach to multiplexed detection of three different miRs (miR-371a-3p, miR-451a-5p, and miR-122–5p) using spectral and temporal filtering. Importantly, we designed a user-independent multiplexed analysis using machine learning algorithms for automatic bead classification. Although the sensitivity of this technique must be further improved to detect miRs at lower concentrations, the method represents a significant advancement in miR analysis by combining ML segmentation using lifetime and intensity images. In addition, the technique offers multiplexing capabilities and the potential for automation, paving the way for more accurate and robust clinical applications in the future.

1569-P: Quantification of Postprandial Hepatic Glucose Disposal in Roux-en-Y Gastric Bypass and Non-operated Controls Using Stable Isotope Labelling and Liver Metabolic Imaging

1569-P: Quantification of Postprandial Hepatic Glucose Disposal in Roux-en-Y Gastric Bypass and Non-operated Controls Using Stable Isotope Labelling and Liver Metabolic Imaging