A novel biosensing strategy on the dynamic and on-site detection of Vibrio coralliilyticus eDNA for coral health warnings.

Heat stress and coral diseases are the predominant factors causing the degradation of coral reef ecosystems. Over recent years, Vibrio coralliilyticus was identified as a temperature-dependent pathogen causing tissue lysis in Pocillopora damicornis and one of the primary pathogens causing bleaching and mortality in other corals. Yet current detection techniques for V. coralliilyticus rely primarily on qPCR and ddPCR, which cannot meet the requirements for non-invasive and real-time detection. Herein, we developed an effective electrochemical biosensor modified by an Au-MoS2/rGO (AMG) nanocomposites and a specific capture probe to dynamically detect V. coralliilyticus environment DNA (eDNA) in aquarium experiments, with a lower limit of detection (0.28 fM) for synthetic DNA and a lower limit of quantification (9.8 fg/µL, ∼0.86 copies/µL) for genomic DNA. Its reliability and accuracy were verified by comparison with the ddPCR method (P > 0.05). Notably, coral tissue started to lyse at only 29 °C when the concentration of V. coralliilyticus increased abruptly to 880 copies/µL, indicating the biosensor could reflect the types of pathogen and health risks of corals under heat stress. Overall, the novel and reliable electrochemical biosensing technology provides an efficient strategy for the on-site monitoring and early warning of coral health in the context of global warming.

Edoxaban for stroke prevention in atrial fibrillation and factors associated with dosing: patient characteristics from the prospective observational ETNA-AF-China registry.

Real-world data on effectiveness and safety of a single non-vitamin K antagonist oral anticoagulant in the Chinese population with atrial fibrillation (AF) are limited. This study reports characteristics of patients treated with edoxaban and factors associated with dosing patterns from routine care in China. ETNA-AF-China (NCT04747496) is a multicentre, prospective, observational study enrolling edoxaban-treated patients from four economic regions with a targeted 2-year follow-up. Of the 4930 patients with AF (mean age: 70.2 ± 9.5 years; male, 57.1%), the mean creatinine clearance (CrCl), CHA2DS2-VASc, and HAS-BLED scores were 71.2 mL/min, 2.9, and 1.6. Overall, 6.4% of patients were perceived as frail by investigators. Available label dose reduction criteria (N = 4232) revealed that 3278 (77.5%) patients received recommended doses and 954 (22.5%) non-recommended doses. Northeast (53.0%) and West (43.1%) regions had the highest prescriptions of 60 mg and 30 mg recommended doses, respectively. Non-recommended 30 mg doses were more frequently prescribed in patients with antiplatelet use and history of heart failure than recommended 60 mg. Multivariate analysis identified advanced age as the strongest associated factor with non-recommended doses. Frailty had the strongest association with 30 mg except for age, and history of TIA was the most relevant factor associated with 60 mg. In conclusion, patients in the ETNA-AF-China study were predominantly aged 65 years and older, had mild-to-moderate renal impairment and good label adherence. Advanced age was associated with non-recommended doses, with frailty most common for non-recommended 30 mg and a history of TIA for the non-recommended 60 mg dose.

Comprehensive transcriptomics and metabolomics analyses reveal that hyperhomocysteinemia is a high risk factor for coronary artery disease in a chinese obese population aged 40-65: a prospective cross-sectional study.

BACKGROUND: Clinical observations suggest a complex relationship between obesity and coronary artery disease (CAD). This study aimed to characterize the intermediate metabolism phenotypes among obese patients with CAD and without CAD. METHODS: Sixty-two participants who consecutively underwent coronary angiography were enrolled in the discovery cohort. Transcriptional and untargeted metabolomics analyses were carried out to screen for key molecular changes between obese patients with CAD (CAD obese), without CAD (Non-CAD obese), and Non-CAD leans. A targeted GC-MS metabolomics approach was used to further identify differentially expressed metabolites in the validation cohorts. Regression and receiver operator curve analysis were performed to validate the risk model. RESULTS: We found common aberrantly expressed pathways both at the transcriptional and metabolomics levels. These pathways included cysteine and methionine metabolism and arginine and proline metabolism. Untargeted metabolomics revealed that S-adenosylhomocysteine (SAH), 3-hydroxybenzoic acid, 2-hydroxyhippuric acid, nicotinuric acid, and 2-arachidonoyl glycerol were significantly elevated in the CAD obese group compared to the other two groups. In the validation study, targeted cysteine and methionine metabolomics analyses showed that homocysteine (Hcy), SAH, and choline were significantly increased in the CAD obese group compared with the Non-CAD obese group, while betaine, 5-methylpropanedioic acid, S-adenosylmethionine, 4-PA, and vitamin B2 (VB2) showed no significant differences. Multivariate analyses showed that Hcy was an independent predictor of obesity with CAD (hazard ratio 1.7; 95%CI 1.2-2.6). The area under the curve based on the Hcy metabolomic (HCY-Mtb) index was 0.819, and up to 0.877 for the HCY-Mtb.index plus clinical variables. CONCLUSION: This is the first study to propose that obesity with hyperhomocysteinemia is a useful intermediate metabolism phenotype that could be used to identify obese patients at high risk for developing CAD.

Apoe-knockout induces strong vascular oxidative stress and significant changes in the gene expression profile related to the pathways implicated in redox, inflammation, and endothelial function.

Apolipoprotein E (APOE) was recognized as a key regulator of lipid metabolism, which prompted the Apoe-knockout (Apoe-/-) mouse to be the most widely used atherosclerotic model. However, with more and more important physiological roles of APOE being revealed, it is necessary to reacquaint its comprehensive function in the aorta. In this study, we aimed to reveal how Apoe-knockout impacts the gene pathways and phenotypes in the aorta of mice. We performed transcriptome sequencing to acquire the gene expression profile (GEP) for C57BL/6J and Apoe-/- mouse aorta, and used enrichment analysis to reveal the signal pathways enriched for differentially expressed genes (DEGs). In addition, we used immunofluorescence and ELISA to detect the phenotypic differences of vascular tissues and plasma in the two-group mice. Apoe-knockout resulted in significant changes in the expression of 538 genes, among which about 75% were up-regulated and 134 genes were altered more than twice. In addition to the lipid metabolism pathways, DEGs were also mainly enriched in the pathways implicated in endothelial cell proliferation, migration of epithelial cells, immune regulatory, and redox. GSEA shows that the up-regulated genes are mainly enriched in 'immune regulation pathways' and 'signal regulation' pathways, while the down-regulated genes are enriched in lipid metabolism pathways, 'regulation_of_nitric_oxide_synthase_activity' and the pathways involved in redox homeostasis, including 'monooxygenase regulation', 'peroxisomes' and 'oxygen binding'. A significant increase of reactive oxygen species and a remarkable reduction of GSH/GSSG ratio were respectively observed in the vascular tissues and plasma of Apoe-/- mice. In addition, endothelin-1 significantly increased in the vascular tissue and the plasma of Apoe-/- mice. Taken together, our results suggest that besides functioning in lipid metabolism, APOE may be an important signal regulator that mediates the expression of the genes related to the pathways involved in redox, inflammation, and endothelial function. Apoe-knockout-induced strong vascular oxidative stress is also the key factor contributing to atherosclerosis.

Myocyte Enhancer Factor 2A Plays a Central Role in the Regulatory Networks of Cellular Physiopathology.

Cell regulatory networks are the determinants of cellular homeostasis. Any alteration to these networks results in the disturbance of cellular homeostasis and induces cells towards different fates. Myocyte enhancer factor 2A (MEF2A) is one of four members of the MEF2 family of transcription factors (MEF2A-D). MEF2A is highly expressed in all tissues and is involved in many cell regulatory networks including growth, differentiation, survival and death. It is also necessary for heart development, myogenesis, neuronal development and differentiation. In addition, many other important functions of MEF2A have been reported. Recent studies have shown that MEF2A can regulate different, and sometimes even mutually exclusive cellular events. How MEF2A regulates opposing cellular life processes is an interesting topic and worthy of further exploration. Here, we reviewed almost all MEF2A research papers published in English and summarized them into three main sections: 1) the association of genetic variants in MEF2A with cardiovascular disease, 2) the physiopathological functions of MEF2A, and 3) the regulation of MEF2A activity and its regulatory targets. In summary, multiple regulatory patterns for MEF2A activity and a variety of co-factors cause its transcriptional activity to switch to different target genes, thereby regulating opposing cell life processes. The association of MEF2A with numerous signaling molecules establishes a central role for MEF2A in the regulatory network of cellular physiopathology.

Ultrasensitive and on-site eDNA detection for the monitoring of crown-of-thorns starfish densities at the pre-outbreak stage using an electrochemical biosensor.

The coral reef crisis has significantly intensified over the last decades, mainly due to severe outbreaks of crown-of-thorns starfish (COTS). Current ecological monitoring has failed to detect COTS densities at the pre-outbreak stage, thus preventing early intervention. In this work, we developed an effective electrochemical biosensor modified by a MoO2/C nanomaterial, as well as a specific DNA probe that could detect trace COTS environmental DNA (eDNA) at a lower detection limit (LOD = 0.147 ng/µL) with excellent specificity. The reliability and accuracy of the biosensor were validated against the standard methods by an ultramicro spectrophotometer and droplet digital PCR (p > 0.05). The biosensor was then utilized for the on-site analysis of seawater samples from SYM-LD and SY sites in the South China Sea. For the SYM-LD site suffering an outbreak, the COTS eDNA concentrations were 0.33 ng/µL (1 m, depth) and 0.26 ng/µL (10 m, depth), respectively. According to the ecological survey, the COTS density was 500 ind/hm2 at the SYM-LD site, verifying the accuracy of our measurements. At the SY site, COTS eDNA was also detected at 0.19 ng/µL, but COTS was not found by the traditional survey. Hence, larvae were possibly present in this region. Therefore, this electrochemical biosensor could be used to monitor COTS populations at the pre-outbreak stages, and potentially serve as a revolutionary early warning method. We will continue to improve this method for picomolar or even femtomolar detection of COTS eDNA.

Ubiquitin-specific protease 7 regulates myocardial ischemia/reperfusion injury by stabilizing Keap1.

Myocardial ischemia/reperfusion (I/R) injury is a complex pathological process that is still not fully understood. The oxidative stress response has a critical role in the occurrence and progression of myocardial ischemia/reperfusion injury. This study investigated the specific mechanism of ubiquitin-specific protease 7 (USP7) regulation of myocardial ischemia/reperfusion injury from the perspective of proteasome degradation and its relation with the Keap1 pathway, a vital regulator of cytoprotective responses to endogenous and exogenous stress induced by reactive oxygen species (ROS) and electrophiles. Our data indicated that USP7 expression is increased during myocardial ischemia/reperfusion injury in mice, while its inhibiting suppressed the generation of oxygen free radicals and myocardial cell apoptosis, reduced myocardial tissue damage, and improved heart function. Mechanistically, USP7 stabilizes Keap1 by regulating its ubiquitination. Taken together, these findings demonstrate the potential therapeutic effect of USP7 on myocardial ischemia/reperfusion injury.

USP10 exacerbates neointima formation by stabilizing Skp2 protein in vascular smooth muscle cells.

The underlying mechanism of neointima formation remains unclear. Ubiquitin-specific peptidase 10 (USP10) is a deubiquitinase that plays a major role in cancer development and progression. However, the function of USP10 in arterial restenosis is unknown. Herein, USP10 expression was detected in mouse arteries and increased after carotid ligation. The inhibition of USP10 exhibited thinner neointima in the model of mouse carotid ligation. In vitro data showed that USP10 deficiency reduced proliferation and migration of rat thoracic aorta smooth muscle cells (A7r5) and human aortic smooth muscle cells (HASMCs). Mechanically, USP10 can bind to Skp2 and stabilize its protein level by removing polyubiquitin on Skp2 in the cytoplasm. The overexpression of Skp2 abrogated cell cycle arrest induced by USP10 inhibition. Overall, the current study demonstrated that USP10 is involved in vascular remodeling by directly promoting VSMC proliferation and migration via stabilization of Skp2 protein expression.

Ubiquitin-specific protease 2 regulates Ang â ¡-induced cardiac fibroblasts activation by up-regulating cyclin D1 and stabilizing ß-catenin in vitro.

Cardiac fibrosis, featuring abnormally elevated extracellular matrix accumulation, decreases tissue compliance, impairs cardiac function and accelerates heart failure. Mounting evidence suggests that the ubiquitin proteasome pathway is involved in cardiac fibrosis. In the present study, ubiquitin-specific protease 2 (USP2) was identified as a novel therapeutic target in cardiac fibrosis. Indeed, USP2 expression was increased in angiotensin II-induced primary cardiac fibroblasts (CFs) from neonatal rats. In addition, USP2 inhibition suppressed CFs proliferation, collagen synthesis and cell cycle progression. Furthermore, USP2 interacted with ß-catenin, thereby regulating its deubiquitination and stabilization in CFs. To sum up, these findings revealed that USP2 has a therapeutic potential for the treatment of cardiac fibrosis.

MEF2A Is the Trigger of Resveratrol Exerting Protection on Vascular Endothelial Cell.

Both resveratrol and myocyte enhancer factor 2A (MEF2A) may protect vascular endothelial cell (VEC) through activating the expression of SIRT1. However, the relationship between resveratrol and MEF2A is unclear. We aimed to investigate the deeper mechanism of resveratrol in protecting vascular endothelial cells and whether MEF2A plays a key role in the protective function of resveratrol. Human umbilical vein endothelial cell (HUVEC) was used for in vitro study, and small interfere RNA was used for silencing MEF2A. Silencing MEF2A in the vascular endothelium (VE) of ApoE-/- mice was performed by tail injection with adeno associated virus expressing si-mef2a-shRNA. The results showed that treatment of HUVEC with resveratrol significantly up-regulated MEF2A, and prevented H2O2-induced but not siRNA-induced down-regulation of MEF2A. Under various experimental conditions, the expression of SIRT1 changed with the level of MEF2A. Resveratrol could rescue from cell apoptosis, reduction of cell proliferation and viability induced by H2O2, but could not prevent against that caused by silencing MEF2A with siRNA. Silencing MEF2A in VE of apoE-/- mice decreased the expression of SIRT1, increased the plasma LDL-c, and abrogated the function of resveratrol on reducing triglyceride. Impaired integrity of VE and aggravated atherosclerotic lesion were observed in MEF2A silenced mice through immunofluorescence and oil red O staining, respectively. In conclusion, resveratrol enhances MEF2A expression, and the upregulation of MEF2A is required for the endothelial protective benefits of resveratrol in vitro via activating SIRT1. Our work has also explored the in vivo relevance of this signaling pathway in experimental models of atherosclerosis and lipid dysregulation, setting the stage for more comprehensive phenotyping in vivo and further defining the molecular mechanisms.

Allium macrostemon Saponin Inhibits Activation of Platelet via the CD40 Signaling Pathway.

Allium macrostemon saponin is a traditional Chinese medicine that exhibits anti-atherosclerosis effects. However, the mechanism of its action has not been fully clarified. Platelet activation induced by CD40L plays an important role in the process of atherosis. In the present study, we demonstrate for the first time that A. macrostemon saponin inhibits platelet activation induced by CD40L. Moreover, the effects of saponin on platelet activation were achieved by activation of the classical CD40L-associated pathway, including the PI3K/Akt, MAPK and NF-κB proteins. In addition, the present study further demonstrated that saponin exhibited an effect on the TRAF2-mediated ubiquitination degradation, which contributed to the inhibition of the CD40 pathway and its downstream members. The findings determine that A. macrostemon saponin inhibits activation of platelets via activation of downstream proteins of the CD40 pathway. This in turn affected TRAF2-associated ubiquitination degradation and caused an anti-thrombotic effect.

Myocyte enhancer factor 2A delays vascular endothelial cell senescence by activating the PI3K/p-Akt/SIRT1 pathway.

Myocyte enhancer factor 2A (MEF2A) dysfunction is closely related to the occurrence of senile diseases such as cardiocerebrovascular diseases, but the underlying molecular mechanism is unclear. Here, we studied the effects of MEF2A on the senescent phenotype of vascular endothelial cells (VEC) and downstream signaling pathway, and the association between plasma MEF2A levels and coronary artery disease (CAD). Results showed that MEF2A silencing promoted cell senescence and down-regulated PI3K/p-AKT/Sirtuin 1 (SIRT1) expression. MEF2A overexpression delayed cell senescence and up-regulated PI3K/p-AKT/SIRT1. Hydrogen peroxide (H2O2) treatment induced cellular senescence and down-regulated the expression of MEF2A and PI3K/p-AKT/SIRT1. MEF2A overexpression inhibited cellular senescence and the down-regulation of PI3K/p-AKT/SIRT1 induced by H2O2. Further study revealed that MEF2A directly up-regulated the expression of PIK3CA and PIK3CG through MEF2 binding sites in the promoter region. Pearson correlation and logistic regression analysis showed that the plasma level of MEF2A was negatively correlated with CAD, and with age in the controls. These results suggested that MEF2A can directly up-regulate PI3K gene expression, and one of the molecular mechanisms of delaying effect of MEF2A on VEC cell senescence was SIRT1-expression activation through the PI3K/p-Akt pathway. Moreover, the plasma MEF2A levels may be a potential biomarker for CAD risk prediction.

MEF2A alters the proliferation, inflammation-related gene expression profiles and its silencing induces cellular senescence in human coronary endothelial cells.

BACKGROUND: Myocyte enhancer factor 2A (MEF2A) plays an important role in cell proliferation, differentiation and survival. Functional deletion or mutation in MEF2A predisposes individuals to cardiovascular disease mainly caused by vascular endothelial dysfunction. However, the effect of the inhibition of MEF2A expression on human coronary artery endothelial cells (HCAECs) is unclear. In this study, expression of MEF2A was inhibited by specific small interference RNA (siRNA), and changes in mRNA profiles in response to MEF2A knockdown were analyzed using an Agilent human mRNA array. RESULTS: Silencing of MEF2A in HCAECs accelerated cell senescence and suppressed cell proliferation. Microarray analysis identified 962 differentially expressed genes (DEGs) between the MEF2A knockdown group and the negative control group. Annotation clustering analysis showed that the DEGs were preferentially enriched in gene ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways related to proliferation, development, survival, and inflammation. Furthermore, 61 of the 578 downregulated DEGs have at least one potential MEF2A binding site in the proximal promoter and were mostly enriched in the GO terms "reproduction" and "cardiovascular." The protein-protein interaction network analyzed for the downregulated DEGs and the DEGs in the GO terms "cardiovascular" and "aging" revealed that PIK3CG, IL1B, IL8, and PRKCB were included in hot nodes, and the regulation of the longevity-associated gene PIK3CG by MEF2A has been verified at the protein level, suggesting that PIK3CG might play a key role in MEF2A knockdown induced HCAEC senescence. CONCLUSIONS: MEF2A knockdown accelerates HCAEC senescence, and the underlying molecular mechanism may be involved in down-regulation of the genes related with cell proliferation, development, inflammation and survival, in which PIK3CG may play a key role.

Resveratrol inhibits angiotensin II­induced proliferation of A7r5 cells and decreases neointimal hyperplasia by inhibiting the CaMKII­HDAC4 signaling pathway.

Resveratrol has been reported to inhibit vascular smooth muscle cell proliferation and neointimal hyperplasia following arterial injury; however, the underlying mechanisms remain unclear. The present study was designed to investigate the effects of resveratrol on angiotensin II (AngII)­induced proliferation of A7r5 cells and explore the molecular mechanisms responsible for the observed effects. Resveratrol inhibited cell proliferation and migration, and decreased the AngII­induced protein expression of α­smooth muscle actin (α­SMA), proliferating cell nuclear antigen (PCNA) and cyclin­dependent kinase 4 (CDK4). Resveratrol inhibited AngII­induced activation of intracellular Ca2+/calmodulin­dependent protein kinase II (CaMKII) and histone deacetylases 4 (HDAC4), as well as blocking AngII­induced cell cycle progression from the G0/G1 to S­phase. In vivo, 4­weeks of resveratrol treatment decreased the neointima area and the neointima/media area ratio in rats following carotid balloon injury. Resveratrol also inhibited the protein expression of total and phosphorylated CaMKII and HDAC4 in the injured arteries. In conclusion, the present study demonstrated that resveratrol attenuated AngII­induced cell proliferation and neointimal hyperplasia by inhibiting the CaMKII­HDAC4 signaling pathway. These findings suggest that resveratrol may potentially prevent arterial restenosis.

Association between HLA-DQB1 alleles and susceptibility to coronary artery disease in Southern Han Chinese.

Accumulated evidence indicates that polymorphisms in human leukocyte antigens (HLA) are associated with susceptibility to coronary artery disease (CAD). However, whether HLA-DQB1 alleles are correlated with susceptibility to CAD is unclear. In this study, significantly lower frequencies of the allele groups (DQB1*03:01:01G and DQB1*05:03:01G) and the genotypes (DQB1*03:01:01G/DQB1*03:01:01G and DQB1*03:01:01G/DQB1*05:03:01G) were observed in the CAD group compared with that in the controls. However, notably higher frequencies of DQB1*04:01:01G and genotype DQB1*05:01:01G/DQB1*03:01:01G were observed in the CAD patients than in the controls. Further analysis in subgroups showed that DQB1*03:01:01G was present at a significantly lower frequency in both female and male CAD patients compared with the corresponding controls; however, DQB1*04:01:01G was overtly high only in male CAD patients. CAD patients with diabetes showed a negative association with DQB1*03:01:01G and DQB1*05:03:01G and a positive association with DQB1*04:01:01G, DQB1*03:02:01G and DQB1*03:03:02G. Results of logistic regression analysis indicated that DQB1*03:01:01G and DQB1*05:03:01G were significantly associated with reduced susceptibility to CAD, but DQB1*04:01:01G, DQB1*03:02:01G and DQB1*03:03:02G had no correlation with CAD. Together, these findings indicate that CAD in Southern Han Chinese is negatively associated with HLA-DQB1*03:01:01G and DQB1*05:03:01G, and males with HLA-DQB1*04:01:01G are likely to have high risk for CAD.

Genotype Frequencies of CYP2C19, P2Y12 and GPIIIa Polymorphisms in Coronary Heart Disease Patients of Han Ethnicity, and Their Impact on Clopidogrel Responsiveness.

To investigate the genotype frequencies of cytochrome P450, family2, subfamily C, polypeptide19 (CYP2C19); P2Y12 receptor; and glycoprotein IIIa polymorphisms in patients with coronary heart disease and their impact on clopidogrel responsiveness and major adverse cardiac events (MACEs).A total of 146 coronary heart disease patients of Han ethnicity, on a clopidogrel regimen, were enrolled. Polymerase chain reaction and DNA sequencing were used to detect the genotype and allelic frequencies of CYP2C19 ((*)2,(*)3,(*)17), P2Y12 (C34T, G52T, T744C) and GPIIIa (T1565C) polymorphisms. Clinical and laboratory data were compared between the high on-treatment platelet reactivity (HTPR) versus normal groups.HTPR was identified in 35 (24%) patients. CYP2C19(*)2 (G681A) polymorphism was found to be significantly associated with HTPR (P < 0.05). A allele frequencies were significantly higher in the HTPR group versus the normal group (P < 0.05). On logistic regression analysis, CYP2C19(*)2 (G681A) polymorphism was found to be an independent risk factor associated with HTPR. No link could be established between genetic polymorphisms and recurrence of MACEs, or between HTPR and recurrence of MACEs.The genetic polymorphisms in CYP2C19(*)2 were closely associated with HTPR. The frequency of the A allele of CYP2C19(*)2 was significantly associated with HTPR, with A allele carriers being more likely to develop HTPR.

Inhibition of platelet activation and aggregation by furostanol saponins isolated from the bulbs of Allium macrostemon Bunge.

Three new furostanol saponins (FSs) were recently isolated from the dried bulbs of Allium macrostemon and were shown to have antiplatelet effects. This study investigated the inhibitory capabilities of these compounds on adenosine diphosphate (ADP)-induced human platelet activation. FS-1, when compared with the other 2, had a potent inhibitory effect on ADP-induced platelet aggregation and on the expression of P-selectin and integrin ß-3. FS-1 also inhibited Ca mobilization and significantly decreased phosphorylated AKT expression in ADP-activated platelets. The suppression by FS-1 of ADP-induced platelet activation and aggregation shown in this study indicate its potential for therapeutic applications.

New steroidal glycosides isolated as CDL inhibitors of activated platelets.

Three new compounds were isolated from the dried bulbs of Allium macrostemon Bunge. Their structures were elucidated from their spectral data as (25R)-26-O-beta-D-glucopyranosyl-5alpha-furostane-3beta,12beta,22,26-tetraol-3-O-beta-D-glucopyranos-yl (1-->2) [beta-D-glucopyranosyl (1-->3)]-beta-D-glucopyranosyl (1-->4)-beta-D-galactopyranoside (1), (25R)-26-O-beta-D-glucopyranosyl-5alpha-furostane-3beta,12alpha,22,26-tetraol-3-O-beta-D-glucopyranosyl (1-->2) [beta-D-glucopyranosyl (1-->3)]-beta-D-glucopyranosyl (1-->4)-beta-D-galacto- pyranoside (2) and (25R)-26-O-beta-D-glucopyranosyl-5beta-furostane-3beta,12alpha,22,26-tetraol-3-O-beta-D-glucopyranosyl (1-->2)-beta-D-galactopyranoside (3), respectively. The inhibition effect of all compounds on CD40 ligand (CD40L) expression on the membrane of activated platelets stimulated by ADP was tested. Compounds 1 and 2 exhibited significant inhibitory activities in a dose dependent manner (P < 0.05), suggesting their potential application as CD40L inhibitors.

[Role of sphingosine 1-phosphate receptor signaling in hematopoietic stem/progenitor cell transmigration].

OBJECTIVE: To determine the role of sphingosine 1-phosphate receptor (S1PRs ) signaling in CD34+ hematopoietic stem/progenitor cell transmigration. METHODS: CD34(+) cells were separated by Ficoll density gradient centrifugation and incubated in DMEM medium with 10% fetal calf serum. The cells were pretreated by FTY720, with or without pertussis toxin (PTX) and antiCXCR4 mAb in the medium, followed by addition of 100 ng/ml SDF-1 into the lower chamber of a Costar 24-well transwell. The migrated cells were counted using FACS and the migrating rates were determined. The expressions of sphingosine 1-phosphate receptors were analyzed in CD34(+) cells before and after the transmigration by reverse transcriptase- polymerase chain reaction (RT-PCR). Cord blood CD34(+) cells were treated with or without FTY720 (10(+) mol/L), and the expressions of CD49d (VLA-4), CD11a (LFA-1), and CD62L (L-selectin) were analyzed at 1, 8, and 16 h after the treatment. RESULTS: While FTY720 did not affect spontaneous migration, a substantial increase of SDF-1-induced transmigration was observed in the presence of FTY720 (15.26 2.14 to 28.64 2.37). The FTY720-enhanced transmigration was completely blocked by addition of PTX or antiCXCR4 mAb. S1p1-5 was expressed in fresh isolated cord blood CD34(+) cells. The migrating cells stimulated by FTY720 and SDF-1 only expressed S1P1, S1P3, and S1P4. The expressions of CD49d, CD11a and CD62L on CD34(+) cells treated with FTY720 remained unchanged at the selected time points as compared with the control. CONCLUSIONS: S1PRs are involved the transmigration of CD34(+) cells. The activation of S1PRs results in increased chemotactic response of CD34(+) to SDF-1. These effects are mediated through CXCR4 and PTX-sensitive Gi proteins. Only the CD34(+) cells expressing the specific receptors can rapidly transmigrate. The activation of the S1PRs does not affect the expressions of the adhesion molecules on cord blood CD34(+) cells.

[Targeted adhesion of platelet receptor-specific microbubble agent to thrombus in vitro].

OBJECTIVE: To examine the thrombus-targeting effect of platelet receptor-specific lipid microbubbles. METHODS: The targeted microbubbles were prepared by coupling Arg-Gly-Asp-Ser (RGDS) with the lipid microbubbles, which were added to the microthrombus generated by platelet aggregation. The effects of the targeted microbubbles on the ultrasonic signal was observed in an artificial thrombus model. RESULTS: The targeted microbubbles were adhesive to the microthrombus, while the non-targeted microbubbles did not possess this property. The ultrasonic signal of the thrombus border was enhanced significantly after the addition of the targeted microbubbles. CONCLUSIONS: Platelet receptor-specific microbubbles possess significant adhesive property to the thrombus and can improve signal-to-noise ratio of the thrombus, suggesting the potential value of the targeted microbubbles for clinical application in the diagnosis and treatment of thrombus.