Nao Tan Qing ameliorates Alzheimer's disease-like pathology by regulating glycolipid metabolism and neuroinflammation: A network pharmacology analysis and biological validation.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by progressive cognitive decline and currently there are no available treatments. Alongside the conventional Aß and tau hypotheses, neuroinflammation and metabolism disruption have also been regarded as crucial hallmarks of AD. In this study, a novel Chinese formula Nao Tan Qing (NTQ) was developed and shown to improve AD. In vivo experiments showed that NTQ significantly mitigated cognitive impairment, Aß burden and neuroinflammation in a transgenic AD mouse model (5×FAD). Network pharmacology results revealed that the active components of NTQ could target inflammatory and metabolic pathways. In addition, hippocampal transcriptomics suggested that NTQ regulated signaling pathways related to inflammation and lipid metabolism. Consistently, serum metabolomics further indicated that NTQ could modulate glycolipid metabolism. In summary, a combination of systems pharmacology analysis and biological validation study demonstrates that NTQ could alleviate behavioral abnormality and pathological alterations of AD by targeting glycolipid metabolism and neuroinflammation, and is accordingly a potential therapeutic agent for AD.

Machine learning enables discovery of Gentianine targeting TLR4/NF-κB pathway to repair ischemic stroke injury.

Inflammatory response is believed to accelerate the development of stroke injury. Gentianine, an alkaloid isolated from Gentiana Scabra Bunge, shows effectiveness in anti-inflammation. In this study, the effect of Gentianine on transient middle cerebral artery occlusion (tMCAO) induced mouse model in vivo and further related mechanism in LPS-injuried microglia BV-2 cells in vitro were explored. Effect of Gentianine on tMCAO mouse demonstrated that Gentianine significantly ameliorated tMCAO induced ischemic injury by decreasing brain infarct volume and increasing the neurological score and upper limb muscle strength. Meanwhile, Gentianine significantly decreased the release of serum inflammatory cytokines. Machine learning enables that Gentianine might had anti-ischemic stroke effect through the TLR4/NF-κB signaling pathway. This was verified in vivo and in vitro. Gentianine significantly decrease the TLR4 and Iba-1 expression in vivo. These results also verified in BV-2 cells. Gentianine significantly decreased TLR4, MyD88 and NF-κB expression, as well as NO production and inflammatory cytokines release. Gentianine co-treatment with TLR4 inhibitor, further decreased TLR4, MyD88 and NF-κB expression, NO production, as well as the inflammatory cytokines. Taken together, Gentianine could be used as a potential anti-ischemic stroke agent by suppressing inflammatory responses via TLR4/NF-κB signaling pathway. This study is expected to provide an integrated traditional Chinese and western medicine solution to find potential anti-ischemic stroke compounds based on machine learning.

Metabolomic Profiling of the Synergistic Effects of Ginsenoside Rg1 in Combination with Neural Stem Cell Transplantation in Ischemic Stroke Rats.

As the greatest medical and socioeconomic problem in developing countries, stroke is the second or third leading cause of death in China and worldwide. In adult organisms suffering from stroke, transplanted stem cells have the ability to repair damaged tissues by regenerating autologous cells, while ginsenoside Rg1 could promote proliferation and differentiation of stem cells. Although obvious antistroke effects of ginsenoside Rg1 and transplanted stem cells have been verified in publications, the mechanism exploration remains challenging. Our study was carried out to investigate the synergistic effects of ginsenoside Rg1 and neural stem cell (NSC) transplantation on MCAO rats with a 1H NMR-based nontargeted metabolomics method to identify potential biomarkers and protein targets and discover the potential mechanism. NSCs transplantation after MCAO combined with ginsenoside Rg1 administration could significantly improve the cerebral infarct and neurological deficits. The treatment significantly intervened the levels of ten metabolites, and perturbed energy metabolism, amino acids metabolism, and lipids metabolism. And 11 enzymes were identified and verified as the targets of NSCs transplantation and ginsenoside Rg1 administration on MCAO rats. Our findings helped to improve the antistroke mechanism of NSCs transplantation and ginsenoside Rg1 and supply a theory basis for the combined research of stem cells and Chinese medicine in the future.

A machine learning-driven study indicates emodin improves cardiac hypertrophy by modulation of mitochondrial SIRT3 signaling.

Cardiac hypertrophy (CH) is an enormous risk factor in the process of heart failure development, however, there is still lack of effective treatment for CH. Mitochondrial protection is an effective way against CH. Rheum palmatum L. (rhubarb) has been used to treat chronic heart diseases such as heart failure, especially to inhibit cardiac compensatory enlargement. The aim of this study was to explore the pharmacodynamic component of rhubarb and reveal its pharmacological effects and targets in the treatment of CH. Based on network pharmacology and machine learning approach, ingredients of rhubarb and targets for CH were extracted and surflex docking was conducted for obtaining the optimal ingredient-target combination(s) and emodin-SIRT3 was identified for further functional analysis. Transverse aortic constriction or isoproterenol induced CH mice and phenylephrine injured cardiomyocytes were used to verify the mitochondria protection effect and CH improvement of emodin in vivo and in vitro by modulation of mitochondrial SIRT3 signaling. The results showed that emodin could block agonist-induced and pressure overload-mediated CH. Emodin prevented mitochondrial dysfunction and its underlying mechanism was attributed to the activation of SIRT3, but the effect was not obvious with the presence of SIRT3 inhibitors (3-TYP)/SIRT3 siRNA. Furthermore, PGC-1ɑ was involved in the process of emodin regulating SIRT3 signaling pathway as an upstream target. Our findings clarified the main material basis and mechanism of rhubarb in the treatment of CH. Emodin, as the major ingredient of rhubarb, has therapeutic potential for CH through mitochondrial protection due to the modulation of SIRT3 signaling.

The walnut transcription factor JrGRAS2 contributes to high temperature stress tolerance involving in Dof transcriptional regulation and HSP protein expression.

BACKGROUND: GRAS transcription factor (TF) family is unique and numerous in higher plants with diverse functions that involving in plant growth and development processes, such as gibberellin (GA) signal transduction, root development, root nodule formation, and mycorrhiza formation. Walnut tree is exposed to various environmental stimulus that causing concern about its resistance mechanism. In order to understand the molecular mechanism of walnut to adversity response, a GRAS TF (JrGRAS2) was cloned and characterized from Juglans regia in this study. RESULTS: A 1500 bp promoter fragment of JrGRAS2 was identified from the genome of J. regia, in which the cis-elements were screened. This JrGRAS2 promoter displayed expression activity that was enhanced significantly by high temperature (HT) stress. Yeast one-hybrid assay, transient expression and chromatin immunoprecipitation (Chip)-PCR analysis revealed that JrDof3 could specifically bind to the DOFCOREZM motif and share similar expression patterns with JrGRAS2 under HT stress. The transcription of JrGRAS2 was induced by HT stress and up-regulated to 6.73-~11.96-fold in the leaf and 2.53-~4.50-fold in the root to control, respectively. JrGRAS2 was overexpressed in Arabidopsis, three lines with much high expression level of JrGRAS2 (S3, S7, and S8) were selected for HT stress tolerance analysis. Compared to the wild type (WT) Arabidopsis, S3, S7, and S8 exhibited enhanced seed germination rate, fresh weight accumulation, and activities of catalase (CAT), peroxidase (POD), superoxide dismutase (SOD) and glutathione-S-transferase (GST) under HT stress. In contrast, the Evans blue staining, electrolyte leakage (EL) rates, hydrogen dioxide (H2O2) and malondialdehyde (MDA) content of transgenic seedlings were all lower than those of WT exposed to HT stress. Furthermore, the expression of heat shock proteins (HSPs) in S3, S7, and S8 was significant higher than those in WT plants. The similar results were obtained in JrGRAS2 transient overexpression walnut lines under normal and HT stress conditions. CONCLUSIONS: Our results suggested that JrDof3 TF contributes to improve the HT stress response of JrGRAS2, which could effectively control the expression of HSPs to enhance HT stress tolerance. JrGRAS2 is an useful candidate gene for heat response in plant molecular breeding.

Effects of Qijian mixture on type 2 diabetes assessed by metabonomics, gut microbiota and network pharmacology.

Qijian mixture, a new traditional Chinese medicine (TCM) formula comprising of Astragalus membranaceus, Ramulus euonymi, Coptis chinensis and Pueraria lobata, was designed to ameliorate the type 2 diabetes (T2D), and its safety and efficacy were evaluated in the research by metabonomics, gut microbiota and system pharmacology. To study the hypoglycemic effect of Qijian mixture, male KKay mice (28-30 g, 8-9 week) and C57/BL6 mice (18-19 g, 8-9 week) were used. Thirty KKay diabetic mice were randomly distributed into 5 groups, abbreviated as Model group (Model), Low Qijian Mixture group (QJM(L)), High Qijian Mixture group (QJM(H)), Chinese Medicine (Gegen Qinlian Decoction) Positive group (GGQL), and Western Medicine (Metformin hydrochloride) Positive group (Metformin). C57/BL6 was considered as the healthy control group (Control). Moreover, a system pharmacology approach was utilized to assess the physiological targets involved in the action of Qijian mixture. There was no adverse drug reaction of Qijian mixture in the acute toxicity study and HE result, and, compared with Model group, Qijian mixture could modulate blood glycemic level safely and effectively. Qijian Mixture was lesser effective than metformin hydrochloride; however, both showed similar hypoglycemic trend. Based on 1H NMR based metabonomics study, the profoundly altered metabolites in Qijian mixture treatment group were identified. Qijian mixture-related 55 proteins and 4 signaling pathways, including galactose metabolism, valine, leucine and isoleucine degradation metabolism, aminoacyl-tRNA biosynthesis metabolism and alanine, aspartate and glutamate metabolism pathways, were explored. The PCoA analysis of gut microbiota revealed that Qijian mixture treatment profoundly enriched bacteroidetes. In addition, the system pharmacology paradigm revealed that Qijian mixture acted through TP53, AKT1 and PPARA proteins. It was concluded that Qijian mixture effectively alleviated T2D, and this effect was linked with the altered features of the metabolite profiles and the gut microbiota.

Sonic hedgehog is a regulator of extracellular glutamate levels and epilepsy.

Sonic hedgehog (Shh), both as a mitogen and as a morphogen, plays an important role in cell proliferation and differentiation during early development. Here, we show that Shh inhibits glutamate transporter activities in neurons, rapidly enhances extracellular glutamate levels, and affects the development of epilepsy. Shh is quickly released in response to epileptic, but not physiological, stimuli. Inhibition of neuronal glutamate transporters by Shh depends on heterotrimeric G protein subunit Gαi and enhances extracellular glutamate levels. Inhibiting Shh signaling greatly reduces epileptiform activities in both cell cultures and hippocampal slices. Moreover, pharmacological or genetic inhibition of Shh signaling markedly suppresses epileptic phenotypes in kindling or pilocarpine models. Our results suggest that Shh contributes to the development of epilepsy and suppression of its signaling prevents the development of the disease. Thus, Shh can act as a modulator of neuronal activity, rapidly regulating glutamate levels and promoting epilepsy.

Significance of Resveratrol in Clinical Management of Chronic Diseases.

Resveratrol could be beneficial to health and provides protection against a wide array of pathologies and age-associated problems, as evident from preclinical studies. However, a comparison of animal and human studies reveals that this dietary polyphenol cannot protect against metabolic diseases and their associated complications. The clinical outcomes are affected by many factors such as sample size. This article not only presents a comprehensive review of the current advances concerning the dose, the extent of absorption, interaction and toxicity of resveratrol in human studies, but also describes its therapeutic effects against several chronic diseases such as diabetes mellitus, obesity, cardiovascular diseases, cancer and aging and the related diseases.

In planta characterization of a tau class glutathione S-transferase gene from Juglans regia (JrGSTTau1) involved in chilling tolerance.

KEY MESSAGE: JrGSTTau1 is an important candidate gene for plant chilling tolerance regulation. A tau subfamily glutathione S-transferase (GST) gene from Juglans regia (JrGSTTau1, GeneBank No.: KT351091) was cloned and functionally characterized. JrGSTTau1 was induced by 16, 12, 10, 8, and 6 °C stresses. The transiently transformed J. regia showed much greater GST, glutathione peroxidase (GPX), superoxide dismutase (SOD), and peroxidase (POD) activities and lower H2O2, malondialdehyde (MDA), reactive oxygen species (ROS), and electrolyte leakage (EL) rate than prokII (empty vector control) and RNAi::JrGSTTau1 under cold stress, indicating that JrGSTTau1 may be involved in chilling tolerance. To further confirm the role of JrGSTTau1, JrGSTTau1 was heterologously expressed in tobacco, transgenic Line5, Line9, and Line12 were chosen for analysis. The germinations of WT, Line5, Line9, and Line12 were similar, but the fresh weight, primary root length, and total chlorophyll content (tcc) of the transgenic lines were significantly higher than those of WT under cold stress. When cultivated in soil, the GST and SOD activities of transgenic tobacco were significantly higher than those of WT; however, the MDA and H2O2 contents of WT were on average 1.47- and 1.96-fold higher than those of Line5, Line9, and Line12 under 16 °C. The DAB, Evans blue, and PI staining further confirmed these results. Furthermore, the abundances of NtGST, MnSOD, NtMAPK9, and CDPK15 were elevated in 35S::JrGSTTau1 tobacco compared with WT. These results suggested that JrGSTTau1 improves the plant chilling tolerance involved in protecting enzymes, ROS scavenging, and stress-related genes, indicating that JrGSTTau1 is a candidate gene for the potential application in molecular breeding to enhance plant abiotic stress tolerance.

The Synthesis and Evaluation of Arctigenin Amino Acid Ester Derivatives.

The use of arctigenin (ARG), a traditional medicine with many pharmacological activities, has been restricted due to its poor solubility in water. Five amino acid derivatives of ARG have been synthesized using glycine, o-alanine, valine, leucine, and isoleucine, which have t-butyloxy carbonyl (BOC) as a protective group. In this study, we examined the effects of removing these protective groups. The results showed that the amino acid derivatives have better solubility and nitrite-clearing ability than ARG. Among the compounds tested, the amino acid derivatives without protective group were the best. Based on these results, ARG and its two amino acid derivatives without protective group (ARG8, ARG10) were selected to evaluate their anti-tumor activity in vivo at a dosage of 40 mg/kg. The results indicated that ARG8 and ARG10 both exhibit more anti-tumor activity than ARG in H22 tumor-bearing mice. The tumor inhibition rates of ARG8 and ARG10 were 69.27 and 43.58%, which was much higher than ARG. Furthermore, the mice treated with these compounds exhibited less damage to the liver, kidney and immune organs compared with the positive group. Furthermore, ARG8 and ARG10 improved the serum cytokine levels significantly compared to ARG. In brief, this study provides a method to improve the water solubility of drugs, and we also provide a reference basis for new drug development.

Intraplaque neovascularization combined with plaque elasticity for predicting ipsilateral stroke in patients with asymptomatic mild carotid stenosis.

Background: Intraplaque neovascularization (IPN) is a biomarker for vulnerable atherosclerotic plaques and can be effectively visualized via contrast-enhanced ultrasound (CEUS). Plaque elasticity is influenced by elements such as lipid core and fibrosis and can be quantitatively assessed on shear wave elastography (SWE). Studies combining the use of CEUS and SWE for the assessment of stroke risk are currently lacking. Our study thus aimed to determine the predictive value of IPN combined with plaque elasticity among patients with asymptomatic carotid plaque. Methods: Consecutive patients with mild carotid stenosis who underwent CEUS and SWE were retrospectively analyzed. IPN was graded according to the presence and location of microbubbles within the plaque, while plaque elasticity was measured in terms of mean shear wave velocity (SWV). All patients were followed up for 6 months to monitor the development of ischemic stroke. The predictive values of IPN and SWV, individually and in combination, were assessed. Results: A total of 121 patients were included, of whom 95 (78.5%) were male. The mean age was 63.1±10.7 years. Both grade 2 IPN [hazard ratio (HR) =2.37, 95% confidence interval (CI): 1.58-9.65; P=0.039] and SWV (HR =0.43, 95% CI: 0.20-0.95; P=0.038) were independently associated with future ischemic stroke events. The combined model demonstrated a significantly better predictive performance (HR =3.243, 95% CI: 1.87-6.17; P=0.027). Conclusions: The combination of IPN and SWV demonstrated significantly better predictive value for the risk of stroke. Our combined model thereby has the potential to guide the clinical stratification and management of patients with asymptomatic mild carotid stenosis.

Quantitative evaluation using carotid ultrasonography-based high-frame-rate vector flow imaging in patients with low carotid stenosis.

OBJECTIVE: To explore the role of quantitative evaluation using carotid ultrasonography (US)-based high-frame-rate vector flow (V Flow) imaging in patients with low carotid stenosis. METHODS: This single-centre cross-sectional study consecutively recruited volunteers without carotid plaque and patients with low carotid stenosis from August 2022 to May 2023. Patients were divided into symptomatic and asymptomatic groups according to their head CT or MRI results within 8 weeks. All V Flow imaging examinations were performed using a Mindray Resona R9 US system. The wall shear stress (WSS) values, oscillatory shear index (OSI) values, and turbulence (Tur) indexes in the normal common carotid artery (CCA), normal carotid bifurcation (CB), and on the upstream and downstream surface of carotid plaque were measured. Pearson Chi-square test and Fisher exact test were used for counting data according to their type. For measurement data, independent sample t test and non-parametric rank sum test were used. RESULTS: The results proved that patients have higher WSS values and Tur indexes of CB than volunteers, and higher WSS values were detected on the surface of the plaques in symptomatic patients. What's more, the downstream side of the plaque was more vulnerable to plaque rupture than the upstream side due to more dynamic blood flow. CONCLUSION: Therefore, carotid US-based high-frame-rate V Flow imaging provides reliable mechanical biomarkers for assessing the haemodynamic change in patients with low stenosis. Our study may provide a new imaging tool for monitoring the progression of atherosclerosis and aiding the management of early atherosclerotic patients. ADVANCES IN KNOWLEDGE: Our study firstly investigated the difference of V Flow parameters on the surface of carotid plaques between symptomatic and asymptomatic patients with low carotid stenosis, which is expected to provide haemodynamic information and the mechanical basis for plaque rupture.

Associations between carotid plaque shape, biomechanical parameters, and ischemic stroke in mild carotid stenosis with a single plaque.

PURPOSE: This cross-sectional cohort-comparison observational study investigated the value of high-frame-rate vector flow (V Flow) imaging for evaluating differences in carotid plaque shape and biomechanical parameters in patients with mild stenosis according to a recent history of ipsilateral ischemic stroke. METHODS: The present study included 352 patients from February 2023 to October 2023, who were categorized as symptomatic or asymptomatic based on a history of recent ischemic stroke and ipsilateral ischemic lesions detected on head computed tomography or magnetic resonance imaging. A Mindray Resona R9 system was used for B-mode ultrasonography and V Flow imaging. The upstream and downstream surfaces of the plaques were examined at the carotid bifurcation for wall shear stress (WSS), oscillatory shear index (OSI), and turbulence index, which performed peri-plaque biomechanical condition. Multivariable logistic regression models were used to determine associations between plaque shape, V Flow parameters, and ischemic stroke. RESULTS: Symptomatic patients exhibited higher WSS values for the upstream and downstream surfaces of carotid plaque, as well as higher OSI and turbulence index values for the downstream surface. Type Ⅲ plaques and higher WSS and OSI values for the downstream surface of the plaque were significantly associated with ischemic stroke. Type Ⅲ plaques were more prevalent in symptomatic patients and demonstrated much higher WSS and OSI values for the downstream plaque surface in both groups. CONCLUSION: High-frame-rate V Flow imaging could assess peri-plaque biomechanical forces and may provide effective imaging biomarkers for early prediction of ischemic stroke in patients with mild stenosis.

TRPC5 channel is the mediator of neurotrophin-3 in regulating dendritic growth via CaMKIIα in rat hippocampal neurons.

Neurotrophin-3 (NT-3) plays numerous important roles in the CNS and the elevation of intracellular Ca(2+) ([Ca(2+)](i)) is critical for these functions of NT-3. However, the mechanism by which NT-3 induces [Ca(2+)](i) elevation remains largely unknown. Here, we found that transient receptor potential canonical (TRPC) 5 protein and TrkC, the NT-3 receptor, exhibited a similar temporal expression in rat hippocampus and cellular colocalization in hippocampal neurons. Stimulation of the neurons by NT-3 induced a nonselective cation conductance and PLCγ-dependent [Ca(2+)](i) elevation, which were both blocked when TRPC5, but not TRPC6 channels, were inhibited. Moreover, the Ca(2+) influx through TRPC5 induced by NT-3 inhibited the neuronal dendritic growth through activation of calmodulin-dependent kinase (CaMK) IIα. In contrast, the Ca(2+) influx through TRPC6 induced by NT-4 promoted the dendritic growth. Thus, TRPC5 acts as a novel and specific mediator for NT-3 to regulate dendrite development through CaMKIIα.

Machine learning and experimental screening of chromatin regulator signatures and potential drugs in hepatitis B related hepatocellular carcinoma.

Many evidences have confirmed that chromatin regulator factors (CRs) are involved in the progression of cancer, but its potential mechanism of affecting hepatitis B related hepatocellular carcinoma still needs to be studied. Our study detected the CRs that affect hepatitis B related hepatocellular carcinoma (HBV-HCC) through machine learning analysis, conducted the analysis of immune cells, constructed the relevant risk model and immune function infiltration, and predicted the potential therapeutic drugs. We found that these CRs were significantly related to the immune cells of Macrophages, B cells, CD8+T cells, etc., and PBK, AURKA, TOP2A and AURKB were the potential risk CRs of HBV-HCC. The expression levels of these four CRs increased in HepG2.2.15 cells and the liver of HBV-HCC patients, consistent with the predicted risk model. Subsequently, ten potential drugs closely related to the risk CRs were finally obtained, experimental research on resveratrol has shown that it can inhibit the proliferation of HepG2.2.15 cells and potentially inhibit the occurrence and development of HBV-HCC. Our study provides novel insights into the function of CRs in HBV-HCC and certain ideas for more accurate targeted therapy.Communicated by Ramaswamy H. Sarma.

Manganese-enriched photonic/catalytic nanomedicine augments synergistic anti-TNBC photothermal/nanocatalytic/immuno-therapy via activating cGAS-STING pathway.

As the clinical efficacy of immunotherapy for triple-negative breast cancer (TNBC) remains limited, exploring new immunotherapy approaches is still indispensable. Mn2+ has been proven as a cGAS-STING agonist to remarkably enhance antitumor immunity. Here, we report a combined tumor-therapeutic strategy based on Prussian blue (PB)-mediated photothermal therapy with Mn2+-augmented immunotherapy by synergistically activating the cGAS-STING pathway. Mn-enriched photonic nanomedicine (MnPB-MnOx) were constructed by integrating MnOx onto the surface of Mn-doped PB nanoparticles. All components of MnPB-MnOx are biocompatible and biodegradable, wherein sufficient Mn are endowed through rational nanostructure design, conferring easier cGAS-STING activation. Additionally, tumor hyperthermia strengthened by MnPB under near-infrared light radiation, synergistic with the generation of reactive oxygen species catalyzed by MnOx, double hits cancer cells to release abundant tumor-associated antigens for further promoting immune response stimulation. The local anti-TNBC efficacy of photothermal/immuno-therapy has been proven effective in subcutaneous 4T1-bearing mice. Especially, it has been systematically demonstrated in bilateral orthotopic 4T1-bearing mice that the as-proposed treatment could successfully activate innate and adaptive immunity, and local therapy could engender systemic responses to suppress the distant tumors. Collectively, this work represents a proof-of-concept for a non-invasive Mn-based tumor-immunotherapeutic modality, providing a paradigm for the immunotherapy of metastatic-prone tumors.

Associations of plaque morphology and location with Intraplaque neovascularization in the carotid artery by contrast-enhanced ultrasound imaging.

Objective: Intraplaque neovascularization (IPN) is a known indicator of plaque vulnerability, and is thus considered a predictor of stroke. The morphology and location of the carotid plaque may be correlated with plaque vulnerability. Therefore, our study aimed to examine the associations of carotid plaque morphology and location with IPN. Methods: A total of 141 patients with carotid atherosclerosis (mean age, 64.99 ± 10.96 years) who underwent carotid contrast-enhanced ultrasound (CEUS) between November 2021 and March 2022 were retrospectively analyzed. IPN was graded according to the presence and location of microbubbles within the plaque. The association of IPN grade with carotid plaque morphology and location was evaluated using ordered logistic regression. Results: Of the 171 plaques, 89 (52%) were IPN Grade 0, 21 (12.2%) were Grade 1, and 61 (35.6%) were Grade 2. IPN grade significantly associated with both plaque morphology and location, with higher grades observed among Type III morphology and common carotid artery plaques. Significant negative association was further shown between IPN grade and serum high-density lipoprotein cholesterol (HDL-C) level. Plaque morphology and location, and HDL-C remained significantly associated with IPN grade after adjusting for confounding factors. Conclusion: The location and morphology of carotid plaques were significantly associated with the IPN grade on CEUS, and therefore show potential as biomarkers for plaque vulnerability. Serum HDL-C was also identified as a protective factor against IPN, and may play a role in the management of carotid atherosclerosis. Our study provided a potential strategy for identification of vulnerable carotid plaques and elucidated the important imaging predictors of stroke.

Carotid plaque score and ischemic stroke risk stratification through a combination of B-mode and contrast-enhanced ultrasound in patients with low and intermediate carotid stenosis.

Objective: The occurrence of ischemic stroke (IS) is closely related to the characteristics of carotid plaque (CP). Due to the effect of stroke risk stratification based on B-mode ultrasound (US) and contrast-enhanced ultrasound (CEUS) that has not been studied in patients with low and intermediate carotid stenosis, we construct and validate a CP score and ischemic stroke risk stratification (ISRS) using a combination of B-mode and CEUS, in order to provide new convenient strategies to stratify these patients to prevent stroke. Materials and methods: This retrospective study evaluated 705 patients with low and intermediate carotid stenosis who underwent B-mode and CEUS from November 2021 to April 2023. Qualitative B-mode and CEUS features of carotid plaques were analyzed using a univariable and multivariable logistic regression to construct the CP score. Then, we combined the CP score with Essen stroke risk score (ESRS) to develop ISRS. Results: This study included a total of 705 patients with low and intermediate carotid stenosis, of which 394 were symptomatic patients (with a mean age of 71.03 ± 10.48 years) and 311 were asymptomatic patients (with a mean age of 65.13 ± 10.31 years). Plaque echogenicity, plaque morphology, carotid intima-media thickness in B-mode US and intraplaque neovascularization grading and perfusion pattern in CEUS were significantly associated with IS. The ISRS incorporating these five predictors and ESRS showed good discrimination and calibration in both primary cohort [area under the curve (AUC), 0.91; Hosmer-Lemeshow test, p = 0.903] and validation cohort (AUC, 0.84; Hosmer-Lemeshow test, p = 0.886). Conclusion: We developed an effective and practical tool to identify and stratify patients with low and intermediate carotid stenosis, based on the CP score and ISRS estimation. Our study may provide new insights into managing patients with no indication of surgery.

Membrane depolarization increases membrane PtdIns(4,5)P2 levels through mechanisms involving PKC ßII and PI4 kinase.

In a previous study, we showed that membrane depolarization induced elevation of membrane phosphatidylinositol 4,5-bisphosphates (PtdIns(4,5)P(2), also known as PIP(2)) and subsequently increased the KCNQ2/Q3 currents expressed in Xenopus oocytes through increased PI4 kinase activity. In this study, the underlying mechanism for this depolarization-induced enhancement of PIP(2) synthesis was further investigated. Our results indicate that activation of protein kinase C (PKC) isozyme ßII was responsible for the enhanced PIP(2) synthesis. We found that phorbol-12-myristate, 13-acetate (PMA), an activator of PKC, mimicked the effects of the membrane depolarization by increasing KCNQ2/Q3 activity, elevating membrane PIP(2) levels and increasing activity of PI4 kinase ß. Furthermore, membrane depolarization enhanced PKC activity. The effects of both depolarization and PMA were blocked by a PKC inhibitor or PI4 kinase ß RNA interference. Further results demonstrate that the depolarization selectively activated the PKC ßII isoform and enhanced its interaction with PI4 kinase ß. These results reveal that the depolarization-induced elevation of membrane PIP(2) is through activation of PKC and the subsequent increased activity of PI4 kinase ß.

TRPC6 inhibited NMDA receptor activities and protected neurons from ischemic excitotoxicity.

Excitotoxicity induced by NMDA receptor-mediated intracellular Ca(2+) ([Ca(2+) ](i)) overload is a major cause of delayed neuronal death in cerebral ischemia. Transient receptor potential canonical (TRPC) 6 protects neurons from ischemic brain damage. However, the mechanisms by which TRPC6 protects neurons are largely unknown. Here, we reported that TRPC6 suppressed the [Ca(2+)](i) elevation induced by NMDA and protected neurons from excitotoxicity. Over-expressing or down-regulating TRPC6 suppressed or aggravated Ca(2+) overload under excitotoxicity, respectively. TRPC6 protected cultured neurons from damage caused by NMDA toxicity or oxygen glucose deprivation (OGD). Moreover, the infarct volume in TRPC6 transgenic (Tg) mice was smaller than that in wild-type (WT) littermates. The TRPC6 Tg mice had better behavior performance and lower mortality than their WT littermates. Thus, TRPC6 inhibited NMDA receptor-triggered neurotoxicity and protected neurons from ischemic brain damage. Increase in TRPC6 activity could be a potential strategy for stroke prevention and therapy.