Functional Ligand-Enabled Particle Assembly for Bio-Nano Interactions.

Functional ligands consist of a wide range of small or large molecules that exhibit a spectrum of physical, chemical, and biological properties. A suite of small molecules (e.g., peptides) or macromolecular ligands (e.g., antibodies and polymers) have been conjugated to particle surfaces for specific applications. However, postfunctionalization of ligands often presents challenges in controlling the surface density and may require the chemical modification of ligands. As an alternative option to postfunctionalization, our work has focused on using functional ligands as building blocks to assemble particles while maintaining their intrinsic (functional) properties. Through self-assembly or template-mediated assembly strategies, we have developed a range of protein-, peptide-, DNA-, polyphenol-, glycogen-, and polymer-based particles. This Account discusses the assembly of such nanoengineered particles, which includes self-assembled nanoparticles, hollow capsules, replica particles, and core-shell particles, according to three categories of functional ligands (i.e., small molecules, polymers, and biomacromolecules) that are used as building blocks for their formation. We discuss a range of covalent and noncovalent interactions among ligand molecules that have been explored to facilitate the assembly of particles. The physicochemical properties of the particles, including size, shape, surface charge, permeability, stability, thickness, stiffness, and stimuli-responsiveness, can be readily controlled by varying the ligand building block or by tuning the assembly method. By selecting specific ligands as building blocks, the bio-nano interactions (i.e., stealth, targeting, and cell trafficking) can also be modulated. For instance, particles composed mainly of low-fouling polymers (i.e., poly(ethylene glycol)) exhibit an extended blood circulation time (half-life > 12 h), while antibody-based nanoparticles demonstrate that a trade-off between stealth and targeting may be required when designing targeting nanoparticle systems. Small molecular ligands, such as polyphenols, have been used as building blocks for particle assembly as they can interact with various biomacromolecules through multiple noncovalent interactions, retain the function of biomacromolecules within the assembly, enable pH-responsive disassembly when coordinating with metal ions, and facilitate endosomal escape of nanoparticles. A perspective is provided on the current challenges associated with the clinical translation of ligand-based nanoparticles. This Account is also expected to serve as a reference to guide the fundamental research and development of functional particle systems assembled from various ligands for diverse applications.

Evaluating Middle Cerebral Artery Plaque Characteristics and Lenticulostriate Artery Morphology Associated With Subcortical Infarctions at 7T MRI.

BACKGROUND: Lenticulostriate artery (LSA) obstruction is a potential cause of subcortical infarcts. However, MRI LSA evaluation at 3T is challenging. PURPOSE: To investigate middle cerebral artery (MCA) plaque characteristics and LSA morphology associated with subcortical infarctions in LSA territories using 7-T vessel wall MRI (VW-MRI) and time-of-flight MR angiography (TOF-MRA). STUDY TYPE: Prospective. POPULATION: Sixty patients with 80 MCA atherosclerotic plaques (37 culprit and 43 non-culprit). FIELD STRENGTH/SEQUENCE: 7-T with 3D TOF-MRA and T1-weighted 3D sampling perfection with application-optimized contrast using different flip angle evolutions (SPACE) sequences. ASSESSMENT: Plaque distribution (superior, inferior, ventral, or dorsal walls), LSA origin involvement, LSA morphology (numbers of stems, branches, and length), and plaque characteristics (normalized wall index, maximal wall thickness, plaque length, remodeling index, intraplaque hemorrhage, and plaque surface morphology (regular or irregular)) were assessed. STATISTICAL TESTS: Least absolute shrinkage and selection operator regression, generalized estimating equations regression, receiver operating characteristic curve, independent t-test, Mann-Whitney U test, Chi-square test, Fisher's exact test, and intra-class coefficient. A P value <0.05 was considered statistically significant. RESULTS: Plaque irregular surface, superior wall plaque, longer plaque length, LSA origin involvement, fewer LSA stems, and shorter total and average lengths of LSAs were significantly associated with culprit plaques. Multivariable logistic analysis confirmed that LSA origin involvement (OR, 28.51; 95% CI, 6.34-181.02) and plaque irregular surface (OR, 8.32; 95% CI, 1.41-64.73) were independent predictors in differentiating culprit from non-culprit plaques. A combination of LSA origin involvement and plaque irregular surface (area under curve = 0.92; [95% CI, 0.86-0.98]) showed good performance in identifying culprit plaques, with sensitivity and specificity of 86.5% and 86.0%, respectively. DATA CONCLUSION: 7-T VW-MRI and TOF-MRA can demonstrate plaque involvement with LSA origins. MCA plaque characteristics derived from 7-T VW-MRI showed good diagnostic accuracy in determining the occurrence of subcortical infarctions. EVIDENCE LEVEL: 2 TECHNICAL EFFICACY: Stage 3.

The structure of cognitive strategies for wayfinding decisions.

Literature proposes five distinct cognitive strategies for wayfinding decisions at intersections. Our study investigates whether those strategies rely on a generalized decision-making process, on two frame-specific processes-one in an egocentric and the other in an allocentric spatial reference frame, and/or on five strategy-specific processes. Participants took six trips along a prescribed route through five virtual mazes, each designed for decision-making by a particular strategy. We found that wayfinding accuracy on trips through a given maze correlated significantly with the accuracy on trips through another maze that was designed for a different reference frame (rbetween-frames = 0.20). Correlations were not significantly higher if the other maze was designed for the same reference frame (rwithin-frames = 0.19). However, correlations between trips through the same maze were significantly higher than those between trips through different mazes that were designed for the same reference frame (rwithin-maze = 0.52). We conclude that wayfinding decisions were based on a generalized cognitive process, as well as on strategy-specific processes, while the role of frame-specific processes-if any-was relatively smaller. Thus, the well-established dichotomy of egocentric versus allocentric spatial representations did not translate into a similar, observable dichotomy of decision-making.

The impact of mindfulness therapy combined with mentalization-based family therapy on suicidal ideation in adolescents with depressive disorder: randomized intervention study.

BACKGROUND: Adolescents with depression who engage in non-suicidal self harming behaviors are more likely to adopt negative coping strategies when faced with negative events. Therefore, these patients should be introduced to positive coping strategies. Evidences have showed that mindfulness-based interventions can positively impact the psychology of patients with mental disorders. This study was to explore the impact of a combination of mindfulness therapy and mentalization-based family therapy (MBFT) on suicidal ideation in adolescents with depressive disorder. METHODS: Eighty adolescent patients with depression and suicidal ideation admitted to our hospital from September 2021 to February 2022 were selected as subjects. They were divided into a control group and a study group using the random number table method, with each group comprising 40 subjects. The control group received MBFT, whereas the study group received both mindfulness therapy and MBFT. The psychological status and suicidal ideations of the two groups were compared before and after the intervention. RESULTS: The psychological health scores of both groups of patients were lower after the intervention, with the scores of the study group being lower than those of the control group (P < 0.05). The scores on the suicidal ideation scales for both groups were lower after intervention, and the study group scored lower than the control group (P < 0.05). The absolute values of the differences in psychological health scale scores and suicidal ideation scale scores before and after the intervention were higher in the study group than in the control group (P < 0.05). CONCLUSION: The combination of mindfulness therapy and MBFT can improve the psychological condition of adolescents with depression, reduce their suicidal ideations, and help them develop a healthy and positive outlook toward life, making this method worthy of clinical recommendation.

Particle Engineering via Supramolecular Assembly of Macroscopic Hydrophobic Building Blocks.

Tailoring the hydrophobicity of supramolecular assembly building blocks enables the fabrication of well-defined functional materials. However, the selection of building blocks used in the assembly of metal-phenolic networks (MPNs), an emerging supramolecular assembly platform for particle engineering, has been essentially limited to hydrophilic molecules. Herein, we synthesized and applied biscatechol-functionalized hydrophobic polymers (poly(methyl acrylate) (PMA) and poly(butyl acrylate) (PBA)) as building blocks to engineer MPN particle systems (particles and capsules). Our method allowed control over the shell thickness (e.g., between 10 and 21 nm), stiffness (e.g., from 10 to 126 mN m-1 ), and permeability (e.g., 28-72 % capsules were permeable to 500 kDa fluorescein isothiocyanate-dextran) of the MPN capsules by selection of the hydrophobic polymer building blocks (PMA or PBA) and by controlling the polymer concentration in the MPN assembly solution (0.25-2.0 mM) without additional/engineered assembly processes. Molecular dynamics simulations provided insights into the structural states of the hydrophobic building blocks during assembly and mechanism of film formation. Furthermore, the hydrophobic MPNs facilitated the preparation of fluorescent-labeled and bioactive capsules through postfunctionalization and also particle-cell association engineering by controlling the hydrophobicity of the building blocks. Engineering MPN particle systems via building block hydrophobicity is expected to expand their use.

Development of antiviral carbon quantum dots that target the Japanese encephalitis virus envelope protein.

Japanese encephalitis is a mosquito-borne disease caused by the Japanese encephalitis virus (JEV) that is prevalent in Asia and the Western Pacific. Currently, there is no effective treatment for Japanese encephalitis. Curcumin (Cur) is a compound extracted from the roots of Curcuma longa, and many studies have reported its antiviral and anti-inflammatory activities. However, the high cytotoxicity and very low solubility of Cur limit its biomedical applications. In this study, Cur carbon quantum dots (Cur-CQDs) were synthesized by mild pyrolysis-induced polymerization and carbonization, leading to higher water solubility and lower cytotoxicity, as well as superior antiviral activity against JEV infection. We found that Cur-CQDs effectively bound to the E protein of JEV, preventing viral entry into the host cells. In addition, after continued treatment of JEV with Cur-CQDs, a mutant strain of JEV was evolved that did not support binding of Cur-CQDs to the JEV envelope. Using transmission electron microscopy, biolayer interferometry, and molecular docking analysis, we revealed that the S123R and K312R mutations in the E protein play a key role in binding Cur-CQDs. The S123 and K312 residues are located in structural domains II and III of the E protein, respectively, and are responsible for binding to receptors on and fusing with the cell membrane. Taken together, our results suggest that the E protein of flaviviruses represents a potential target for the development of CQD-based inhibitors to prevent or treat viral infections.

Subarachnoid extension and unfavorable outcomes in patients with supratentorial intracerebral hemorrhage.

OBJECTIVE: Our study aimed to investigate the association between the subarachnoid extension of intracranial hemorrhage (SAHE) and clinical outcomes in patients with supratentorial intracerebral hemorrhage (ICH). METHODS: We analyzed the data from a prospective, multi-center, and registry-based database. Two experienced investigators independently assessed ICH imaging data. We compared baseline characteristics and follow-up outcomes. Multivariable logistic regression analysis was used to evaluate the association between SAHE and poor clinical outcomes. We also performed Kaplan-Meier curves and Cox proportional hazards regression analyses to analyze whether SAHE was relevant to a higher mortality rate. RESULTS: A total of 931 patients were included in this study (SAHE vs. no SAHE, 121 [13.0%] vs. 810 [87.0%]). Patients with SAHE had more severe neurological deficits, higher scores of the mRS, and more remarkable mortality rates at follow-up (all p values < 0.05). In multivariable-adjusted models, SAHE was independently associated with a higher risk of poor outcomes (adjusted OR [95%CI]: 2.030 [1.142-3.608] at 3 months; 2.348 [1.337-4.123] at 1 year). In addition, SAHE remained an independent association with an increased death rate at 1 year (adjusted HR [95%CI], 1.314[1.057-1.635]). In the subgroup analysis, the correlation between SAHE and prognosis exists in patients with lobar or deep ICH. CONCLUSIONS: SAHE is independently associated with poor outcomes in patients with supratentorial ICH. It may provide a promising target for developing new predictive tools targeting ICH.

Single-cell RNA sequencing to reveal non-parenchymal cell heterogeneity and immune network of acetaminophen-induced liver injury in mice.

The role of non-parenchymal cells (NPCs) in the early phase of acetaminophen (APAP)-induced liver injury (AILI) remains unclear. Therefore, single-cell sequencing (scRNA-seq) was performed to explore the heterogeneity and immune network of NPCs in the livers of mice with AILI. Mice were challenged with saline, 300 mg/kg APAP, or 750 mg/kg APAP (n = 3 for each group). After 3 h, the liver samples were collected, digested, and subjected to scRNA-seq. Immunohistochemistry and immunofluorescence were performed to confirm the expression of Makorin ring finger protein 1 (Mkrn1). We identified 14 distinct cell subtypes among the 120,599 cells. A variety of NPCs were involved, even in the early stages of AILI, indicating highly heterogeneous transcriptome dynamics. Cholangiocyte cluster 3, which had high deleted in malignant brain tumors 1 (Dmbt1) expression, was found to perform drug metabolism and detoxification functions. Liver sinusoidal endothelial cells exhibited fenestrae loss and angiogenesis. Macrophage cluster 1 displayed a M1 polarization phenotype, whereas cluster 3 tended to exhibit M2 polarization. Kupffer cells (KCs) exhibited pro-inflammatory effects due to the high expression of Cxcl2. qRT-PCR and western blotting verified that the LIFR-OSM axis might promote the activation of MAPK signaling pathway in RAW264.7 macrophages. Mkrn1 was highly expressed in the liver macrophages of AILI mice and AILI patients. Interaction patterns between macrophages/KCs and other NPCs were complex and diverse. NPCs were highly heterogeneous and were involved in the immune network during the early phase of AILI. In addition, we propose that Mkrn1 may serve as a potential biomarker of AILI.

Choice between decision-making strategies in human route-following.

To follow a prescribed route, we must decide which way to turn at intersections. To do so, we can memorize either the serial order of directions or the associations between spatial cues and directions ("at the drug store, turn left"). Here, we investigate which of these two strategies is used if both are available. In Task S, all intersections looked exactly alike, and participants therefore had to use the serial order strategy to decide which way their route continued. In Task SA, each intersection displayed a unique spatial cue, and participants therefore could use either strategy. In Task A, each intersection displayed a unique cue, but the serial order of cues varied between trips, and participants therefore had to use the associative cue strategy. We found that route-following accuracy increased from trip to trip, was higher on routes with 12 rather than 18 intersections, and was higher on Task SA than on the other two tasks, both with 12 and with 18 intersections. Furthermore, participants on Task SA acquired substantial knowledge about the serial order of directions as well as about cue-direction associations, both with 12 and with 18 intersections. From this we conclude that, when both strategies were available, participants did not pick the better one but rather used both. This represents dual encoding, a phenomenon previously described for more elementary memory tasks. We further conclude that dual encoding may be implemented even if the memory load is not very high (i.e., even with only 12 intersections).

Nanostructured particles assembled from natural building blocks for advanced therapies.

Advanced treatments based on immune system manipulation, gene transcription and regulation, specific organ and cell targeting, and/or photon energy conversion have emerged as promising therapeutic strategies against a range of challenging diseases. Naturally derived macromolecules (e.g., proteins, lipids, polysaccharides, and polyphenols) have increasingly found use as fundamental building blocks for nanostructured particles as their advantageous properties, including biocompatibility, biodegradability, inherent bioactivity, and diverse chemical properties make them suitable for advanced therapeutic applications. This review provides a timely and comprehensive summary of the use of a broad range of natural building blocks in the rapidly developing field of advanced therapeutics with insights specific to nanostructured particles. We focus on an up-to-date overview of the assembly of nanostructured particles using natural building blocks and summarize their key scientific and preclinical milestones for advanced therapies, including adoptive cell therapy, immunotherapy, gene therapy, active targeted drug delivery, photoacoustic therapy and imaging, photothermal therapy, and combinational therapy. A cross-comparison of the advantages and disadvantages of different natural building blocks are highlighted to elucidate the key design principles for such bio-derived nanoparticles toward improving their performance and adoption. Current challenges and future research directions are also discussed, which will accelerate our understanding of designing, engineering, and applying nanostructured particles for advanced therapies.

Supramolecular Polyphenol-DNA Microparticles for In Vivo Adjuvant and Antigen Co-Delivery and Immune Stimulation.

DNA-based materials have attracted interest due to the tunable structure and encoded biological functionality of nucleic acids. A simple and general approach to synthesize DNA-based materials with fine control over morphology and bioactivity is important to expand their applications. Here, we report the synthesis of DNA-based particles via the supramolecular assembly of tannic acid (TA) and DNA. Uniform particles with different morphologies are obtained using a variety of DNA building blocks. The particles enable the co-delivery of cytosine-guanine adjuvant sequences and the antigen ovalbumin in model cells. Intramuscular injection of the particles in mice induces antigen-specific antibody production and T cell responses with no apparent toxicity. Protein expression in cells is shown using capsules assembled from TA and plasmid DNA. This work highlights the potential of TA as a universal material for directing the supramolecular assembly of DNA into gene and vaccine delivery platforms.

Macromolecular Engineering of Thermoresponsive Metal-Phenolic Networks.

Dynamic nanostructured materials that can react to physical and chemical stimuli have attracted interest in the biomedical and materials science fields. Metal-phenolic networks (MPNs) represent a modular class of such materials: these networks form via coordination of phenolic molecules with metal ions and can be used for surface and particle engineering. To broaden the range of accessible MPN properties, we report the fabrication of thermoresponsive MPN capsules using FeIII ions and the thermoresponsive phenolic building block biscatechol-functionalized poly(N-isopropylacrylamide) (biscatechol-PNIPAM). The MPN capsules exhibited reversible changes in capsule size and shell thickness in response to temperature changes. The temperature-induced capsule size changes were influenced by the chain length of biscatechol-PNIPAM and catechol-to-FeIII ion molar ratio. The metal ion type also influenced the capsule size changes, allowing tuning of the MPN capsule mechanical properties. AlIII-based capsules, having a lower stiffness value (10.7 mN m-1), showed a larger temperature-induced size contraction (∼63%) than TbIII-based capsules, which exhibit a higher stiffness value (52.6 mN m-1) and minimal size reduction (<1%). The permeability of the MPN capsules was controlled by changing the temperature (25-50 °C)─a reduced permeability was obtained as the temperature was increased above the lower critical solution temperature of biscatechol-PNIPAM. This temperature-dependent permeability behavior was exploited to encapsulate and release model cargo (500 kDa fluorescein isothiocyanate-tagged dextran) from the capsules; approximately 70% was released over 90 min at 25 °C. This approach provides a synthetic strategy for developing dynamic and thermoresponsive-tunable MPN systems for potential applications in biological science and biotechnology.

Engineering Poly(ethylene glycol) Nanoparticles for Accelerated Blood Clearance Inhibition and Targeted Drug Delivery.

Surface modification with poly(ethylene glycol) (PEGylation) is an effective strategy to improve the colloidal stability of nanoparticles (NPs) and is often used to minimize cellular uptake and clearance of NPs by the immune system. However, PEGylation can also trigger the accelerated blood clearance (ABC) phenomenon, which is known to reduce the circulation time of PEGylated NPs. Herein, we report the engineering of stealth PEG NPs that can avoid the ABC phenomenon and, when modified with hyaluronic acid (HA), show specific cancer cell targeting and drug delivery. PEG NPs cross-linked with disulfide bonds are prepared by using zeolitic imidazolate framework-8 NPs as templates. The reported templating strategy enables the simultaneous removal of the template and formation of PEG NPs under mild conditions (pH 5.5 buffer). Compared to PEGylated liposomes, PEG NPs avoid the secretion of anti-PEG antibodies and the presence of anti-PEG IgM and IgG did not significantly accelerate the blood clearance of PEG NPs, indicating the inhibition of the ABC effect for the PEG NPs. Functionalization of the PEG NPs with HA affords PEG NPs that retain their stealth properties against macrophages, target CD44-expressed cancer cells and, when loaded with the anticancer drug doxorubicin, effectively inhibit tumor growth. The innovation of this study lies in the engineering of PEG NPs that can circumvent the ABC phenomenon and that can be functionalized for the improved and targeted delivery of drugs.

Nonviral mcDNA-mediated bispecific CAR T cells kill tumor cells in an experimental mouse model of hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide and the adoptive immunotherapy of which is worth studying. CD133, a kind of cancer stem cell (CSC) antigen, together with glypican-3 (GPC3) has been proved to be highly expressed in HCC cells and both of them are used as targets to generate chimeric antigen receptor (CAR) T cells. But there are limitations like "off-target" toxicity, low transfection efficacy and weak antitumor ability in CAR T cells treatment. METHODS: The peripheral blood was acquired from healthy donors and T cells were separated by density-gradient centrifugation. We used an electroporation system to deliver anti-CD133 and anti-GPC3 single chain Fragment variable (scFv) structures as target genes into the T cells. The cell membrane was opened by the momentary electric current effect, and the target gene was delivered into the cell by non-viral minicircle DNA (mcDNA) vector. The flow cytometry and western blot assays were used to detect whether the two scFv were simultaneously transfected and the transfection efficacy of this bispecific CAR T cell generation method. We respectively detected the in vitro and in vivo tumor-suppression efficacy of CAR T cells through the CCK-8 assays and the HCC xenograft mice models. The CoG133-CAR T cells containing both CD133 and GPC3 antigen recognition sites were the effector cells. CD133-CAR T cells and GPC3-CAR T cells were defined as single-targeted control groups, normal T and mock T cells were defined as blank control groups. RESULTS: The mcDNA vector accommodated two target gene structures successfully transfected to generate bispecific CAR T cells. The detection methods on gene level and protein level confirmed that CoG133-CAR T cells had considerable transfection efficiency and exhibited both antigen-binding capacity of CD133 and GPC3. Compared to single-targeted CAR T cells or control T cells, CoG133-CAR T cells performed enhanced eliminated efficacy against CD133 and GPC3 double-positive HCC cell line in vitro and HCC xenograft mice in vivo. Hematoxylin and eosin (H&E) staining indicated no fatal "off-target" combination existed on CoG133-CAR T cells and major organs. CONCLUSION: Our study suggests that it is with higher efficiency and more safety to prepare bispecific CAR T cells through non-viral mcDNA vectors. CoG133-CAR T cells have enhanced tumor-suppression capacity through dual antigen recognition and internal activation. It provides an innovative strategy for CAR T therapy of HCC, even solid tumors.

Early neurological deterioration in acute ischemic stroke patients after intravenous thrombolysis with alteplase predicts poor 3-month functional prognosis - data from the Thrombolysis Implementation and Monitor of Acute Ischemic Stroke in China (TIMS-China).

BACKGROUND: We aimed to investigate the risk factors of early neurological deterioration (END) after intravenous thrombolysis with recombinant tissue-type plasminogen activator (rt-PA) and the relationship between END and poor 3-month functional outcomes. METHODS: Patients who accepted intravenous recombinant rt-PA were enrolled continuously. END was defined as an increase in National Institute of Health Stroke (NIHSS) score ≥ 4 points or death within 24 hours after intravenous thrombolysis. The modified Rankin Scale (mRS) score was recorded to evaluate the functional outcome of stroke, and the poor 3-month prognosis was defined as an mRS score ≥ of 3. Univariate and multivariate analyses were used to analyze the risk factors of END. The relation between END and 3-month functional outcome was analyzed by multivariate logistic regression analysis. RESULTS: A total of 1107 patients (mean age, 63.42 ± 11.33 years; 673 males) were included in the final analysis, and 81(7.32%) patients had END. In multivariate analysis, the serum glucose level was significantly associated with END; the odds ratio was 1.10 (95% CI 1.03 to 1.18, p = 0.004). The multivariate logistic analysis showed END has a notable association with the poor 3-month functional recovery even after adjusting for confounding factors; the adjusted OR was 8.25 (95% CI 3.77 to 18.03, p < 0.0001). CONCLUSIONS: The initial serum glucose level might be an independent risk factor of END, and END might predict a poor 3-month prognosis.

Extracranial carotid plaque hemorrhage predicts ipsilateral stroke recurrence in patients with carotid atherosclerosis - a study based on high-resolution vessel wall imaging MRI.

BACKGROUND: Intraplaque hemorrhage (IPH) is a hallmark of carotid plaque vulnerability. We aim to investigate the association between IPH and recurrent ipsilateral ischemic stroke. METHODS: Patients with a recent stroke or transient ischemic attack (TIA) were prospectively recruited and underwent an ultrasonographic examination and carotid HR VWMRI on the side consistent with symptoms. Carotid plaque was defined as carotid intima-media-thickness (IMT) by ultrasound≥1.5 mm. IPH was determined that the ratio of the plaque signal intensity relative to that of adjacent muscle was > 1.5. All enrolled patients were clinically followed until an ipsilateral ischemic stroke, TIA, carotid endarterectomy (CEA)/carotid artery stenting (CAS), or death within 12 months. Univariate analysis was used to analyze the correlation between clinical characteristics and IPH. Kaplan-Meier survival analysis and a log-rank test were used to compare recurrence-free survival time between the IPH and non-IPH groups. Cox regression models evaluated IPH as the predictor of ipsilateral stroke recurrence. RESULTS: A total of 171 patients (mean age, 60.13 ± 10.04 years; 118 males) were included in the final analysis. Thirty-two patients (18.7%) showed carotid IPH. During the follow-up, patients with carotid IPH suffered 60.9% (14 of 23) of recurrent ipsilateral strokes and 60.0% (3 of 5) TIA. Multivariate Cox regression analysis proved IPH as a strong predictor of ipsilateral stroke; the adjusted hazard ratio (HR) was 6.64 (95% confidence interval [CI], 2.84-15.54, P < 0.001). Meanwhile, Cox regression analysis also proved that IPH could predict recurrent ischemic events; the adjusted HR was 8.08 (95% CI, 3.65-17.91, P < 0.001). CONCLUSIONS: Carotid intraplaque hemorrhage is strongly associated with recurrent ischemic events and could predict recurrent ipsilateral stroke.

Protecting mitochondria via inhibiting VDAC1 oligomerization alleviates ferroptosis in acetaminophen-induced acute liver injury.

Acetaminophen (APAP) overdose is a common cause of drug-induced liver injury (DILI). Ferroptosis has been recently implicated in APAP-induced liver injury (AILI). However, the functional role and underlying mechanisms of mitochondria in APAP-induced ferroptosis are unclear. In this study, the voltage-dependent anion channel (VDAC) oligomerization inhibitor VBIT-12 and ferroptosis inhibitors were injected via tail vein in APAP-injured mice. Targeted metabolomics and untargeted lipidomic analyses were utilized to explore underlying mechanisms of APAP-induced mitochondrial dysfunction and subsequent ferroptosis. As a result, APAP overdose led to characteristic changes generally observed in ferroptosis. The use of ferroptosis inhibitor ferrostatin-1 (or UAMC3203) and iron chelator deferoxamine further confirmed that ferroptosis was responsible for AILI. Mitochondrial dysfunction, which is associated with the tricarboxylic acid cycle and fatty acid ß-oxidation suppression, may drive APAP-induced ferroptosis in hepatocytes. APAP overdose induced VDAC1 oligomerization in hepatocytes, and protecting mitochondria via VBIT-12 alleviated APAP-induced ferroptosis. Ceramide and cardiolipin levels were increased via UAMC3203 or VBIT-12 in APAP-induced ferroptosis in hepatocytes. Knockdown of Smpd1 and Taz expression responsible for ceramide and cardiolipin synthesis, respectively, aggravated APAP-induced mitochondrial dysfunction and ferroptosis in hepatocytes, whereas Taz overexpression protected against these processes. By immunohistochemical staining, we found that levels of 4-hydroxynonenal (4-HNE) protein adducts were increased in the liver biopsy samples of patients with DILI compared to that in those of patients with autoimmune liver disease, chronic viral hepatitis B, and non-alcoholic fatty liver disease (NAFLD). In summary, protecting mitochondria via inhibiting VDAC1 oligomerization attenuated hepatocyte ferroptosis by restoring ceramide and cardiolipin content in AILI.

Commutability assessment of reference materials for homocysteine.

OBJECTIVES: Commutability of reference materials is essential for ensuring the traceability of patient measurement results and the technical basis for the use of reference materials. Commutability is only relevant for matrixed reference material; it is a prerequisite for the accuracy and authenticity of calibration methods. In this study, we evaluated the commutability of reference materials for homocysteine. METHODS: Five conventional measurement methods were applied to simultaneously measure 30 serum samples and seven homocysteine reference materials from the National Institute of Standards and Technology and the National Institute of Metrology. Liquid chromatography tandem-mass spectrometry was used as a reference method. Two methods were used to evaluate the commutability of the seven reference materials according to the Clinical and Laboratory Standards Institute EP30-A and the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) commutability assessment document. RESULTS: Among 35 combinations of the five conventional methods and seven reference materials, after evaluation in accordance with the EP30-A, the seven reference materials passed the commutability assessment, and 34 combinations were commutable. According to the IFCC, the commutability evaluation of 28 combinations was conclusive (commutable or non-commutable), while results for the remaining seven combinations could not be determined. CONCLUSIONS: The homocysteine reference materials showed good commutability. The sensitivity of the measurement procedure, measurement deviation and uncertainty, and differences in the "measurand" selected by different methods may affect the evaluation results. Additionally, different judgment standards for different methods may explain the observed variations in evaluation results.

The Ratio of Urine N-Terminal Pro B-Type Natriuretic Peptide to Cyclic Guanosine Monophosphate Predicts Emergency Department Visits for Heart Failure.

INTRODUCTION: Emergency department (ED) visits for decompensated heart failure (HF) are frequent and associated with poor long-term outcomes. Plasma N-terminal pro B-type natriuretic peptide (NT-proBNP) and cyclic guanosine monophosphate (cGMP) are used in diagnosis and prognosis of HF patients, while clinical values of urine NT-proBNP/cGMP ratio have been rarely explored. This study aims to compare the predictive values of urine NT-proBNP/cGMP ratio versus plasma NT-proBNP for ED visits for decompensated HF. METHODS: This prospective study included 126 HF patients with reduced left ventricular ejection fraction (<50%) and without chronic kidney disease. Baseline data included demographics, co-morbidities, and co-medications. Medical records were used to determine the incidence of ED visits for decompensated HF during the 3 months following the last visit. RESULTS: Patients with subsequent ED visits had significantly higher levels of plasma and urine NT-proBNP and urine cGMP in than those without. Multivariate Cox regression analysis disclosed that Lg10urine NT-proBNP/cGMP was an independent risk factor for subsequent ED visits (OR = 3.267; 95% CI: 1.105-9.663; p = 0.032). ROC analysis revealed an Lg10urine NT-proBNP/cGMP ratio optimal cut-off value of 0.1706 (AUC, 0.700; 95% CI: 0.543-0.857; p = 0.036) for predicting subsequent HF-related ED visits. CONCLUSION: A single measurement of urinary NT-proBNP/cGMP ratio is predictive of subsequent ED visits for decompensated HF. This noninvasive and easy measurement may be a clinically useful tool for identifying a subset of patients at higher risk of ED visits.

HAT2CH2 score performance predicting neurologic events after cardiac implantable electronic device.

Objectives: The HAT2CH2 score has been evaluated for predicting new-onset atrial fibrillation in several clinical conditions, but never for adverse neurologic events. We aimed to evaluate the effectiveness of HAT2CH2 score in predicting neurologic events in patients with cardiac implantable electronic device (CIED), comparing with atrial high-rate episodes (AHRE). Methods: This case-control study enrolled 314 consecutive patients aged 18 years or older with CIED implantation between January 2015 and April 2021. Patient data were analyzed retrospectively. The primary endpoint was subsequent neurologic events (NE) after implantation. AHRE was defined as > 175 bpm (Medtronic®) lasting ≥ 30 seconds. Variables associated with independent risk of NE were identified using multivariate Cox regression analysis with time-dependent covariates. Results: Patients' median age was 73 years and 61.8% of them were male. During follow-up (median 32 months), 18 NE occurred (incidence rate 2.15/100 patient-years, 95% CI 1.32-4.30). Multiple Cox regression analysis showed that the HAT2CH2 score (HR 2.424, 95% CI 1.683 - 3.492, p < 0.001) was an independent predictor for NE. Optimal HAT2CH2 score cutoff value was 3 with highest Youden index (AUC, 0.923; 95% CI, 0.881-0.966; p < 0.001). Both AHRE ≥ 1 minute and HAT2CH2 score ≥ 3 had the highest AUC of the receiver-operating characteristic (0.898, 95% CI, 0.831-0.965, p < 0.001). Significant increase was observed in NE occurrence rates using the HAT2CH2 score (p < 0.001). Conclusion: The HAT2CH2 score and episodes of AHRE lasting ≥ 1 minute are independent risk factors for NE in patients with CIED.