Ji-Hong Zhao

Decreased platelet miR-223 expression is associated with high on-clopidogrel platelet reactivity

OBJECTIVES We aimed to investigate the relationship between platelet microRNA (miR-223 and miR-96) expression and clopidogrel responsiveness in patients with coronary heart disease (CHD). MATERIALS AND METHODS A total of 33 consecutive non-diabetic CHD patients scheduled for percutaneous coronary intervention were enrolled. Platelet reactivity after clopidogrel loading dose (300 mg) was determined by two methods [platelet reactivity index (PRI), measured by vasodilator-stimulated phosphoprotein (VASP) phosphorylation flow cytometry and ADP-induced platelet aggregation (PAG), measured by light transmission aggregometry]. Total platelet RNA was isolated from purified platelets (CD45 magnetic bead negative selection) to quantify miR-223 and miR-96 expression by real-time PCR. RESULTS All subjects were dichotomized according to PRI medians (normal-responders: PRI < 56.5%, n = 17 and low-responders: PRI > 56.5%, n = 16) and PAG medians (normal-responders: PAG < 43%, n = 17 and low-responders: PAG > 43%, n = 16). Compared with PRI-determined normal-responders, miR-223 expression, but not miR-96, was significantly decreased in low-responders (P = 0.037). No differential expression of miR-223 and miR-96 was observed via PAG determination between normal- and low-responders. In addition, miR-223 expression, but not miR96, was statistically correlated with PRI (Spearman r = -0.403, P = 0.020). Stepwise binary logistic regression analysis revealed that among factors that potentially influence platelet reactivity (CYP2C19*2 loss-of-function genotypes, use of calcium channel blockers/proton-pump inhibitors, age, obesity, smoking and platelet microRNAs), decreased miR-223 expression was the only independent predictor associated with the presence of PRI-determined low responders to clopidogrel (OR 0.189, 95% CI 0.043 to 0.836, P = 0.028). CONCLUSIONS The present work identifies decreased platelet miR-223 expression as a novel mechanism involved in blunted platelet response to clopidogrel in a Chinese population.

Pseudolaric acid B inhibits T-cell mediated immune response in vivo via p38MAPK signal cascades and PPARγ activation

AIMS Pseudolaric acid B (PAB) has been prescribed for its potent immunomodulatory effect. However, the detail of mechanism remains to be demonstrated. The purpose of this study is to further clarify the mechanism of PAB on T-cell mediated immune response in vivo. MAIN METHODS Investigations were carried to ascertain the pharmacological effect of PAB in a delayed-type hypersensitivity (DTH) mouse model of T-cell mediated immune response. Histological assessment was examined by hematoxylin and eosin staining. Affymetrix GeneChip® Mouse Genome 430 2.0 arrays were employed to evaluate the expression profile of PAB. Western blot was performed to detect p38MAPK signal cascades, including p38MAPK, ATF-2, MK2, and HSP27. Finally, TNF-α level was analyzed by ELISA, and Jurkat T cells were treated with PAB to determine its role on PPARγ activation using a reporter gene assay. KEY FINDINGS The results showed that PAB (5, 10, and 20mg/kg) could lead to a marked improvement for ear swelling and inflammatory infiltrate in DTH mice dose-dependently. According to the associated biological pathways from microarray analysis, PAB resulted in the restoration of abnormal immune-related gene expression linked to MAPK and PPAR signaling pathways. Moreover, PAB inhibited the activation of p38MAPK, ATF-2, MK2, and HSP27 significantly, as well as the production of TNF-α, which was reversed by GW9662, a specific antagonist for PPARγ. In addition, treatment with PAB also increased the transcriptional activity of PPARγ in a dose-dependent manner. SIGNIFICANCE These results provide us with novel insights into pharmacological action of PAB as a potential immunomodulator for the treatment of immune-related diseases.

Left ventricular systolic dyssynchrony in patients with isolated symptomatic myocardial bridge

Abstract Objectives. The impact of myocardial bridge (MB) on left ventricular (LV) systolic synchrony is insufficiently understood. Design. Thirty-five subjects with isolated mid-left, anterior, descending artery (LAD) MB, preserved LV ejection fraction (LVEF > 50%), and otherwise, normal coronary angiogram were identified from 3607 patients who underwent diagnostic coronary angiography and were evaluated by tissue Doppler imaging and real-time three-dimensional echocardiography (RT3DE). Control subjects consisted of 26 age and sex-matched coronary angiographically “normal” subjects. Results. MB patients were characterized by reduced, early, diastolic strain rate in LAD-supplied apical segments (lateral and anterior), with prevalence of LV systolic dyssynchrony of 25.7% (9/35). MB patients were further classified by the medians of MB stenosis and length. For MB stenosis < 52.5%, Class I: length < 17 mm (n = 7), Class II: length ≥ 17 mm (n = 10); for stenosis ≥ 52.5%, Class III: length < 17 mm (n = 10), Class IV: length ≥ 17 mm (n = 8). Binary Logistic regression model revealed that higher MB lesion classification (odds ratio: 4.944, 95%CI 1.174–20.82, P < 0.05) and hypertension (odds ratio: 15.32, 95%CI: 1.252–187.6, P < 0.05) are statistically associated with LV systolic dyssynchrony, which was independent of LV mass. Conclusions. MB in the mid LAD is associated with myocardial dyssynchrony. Hypertensive individuals and those with more severe bridging (determined by length and stenosis) tend to have an increased incidence of dyssynchrony.

The influence of different anticoagulants and time-delayed sample processing and measurements on human monocyte subset and monocyte-platelet aggregate analyses

Measuring human monocyte subsets (CD14++CD16−, CD14++CD16+, and CD14 + CD16++) and subset‐specific monocyte‐platelet aggregates (MPA) is vulnerable to analytical bias due to unavailability of a standardized methodology. We aimed to address this issue by focusing on the impacts of time‐delayed sample processing and measurement between two commonly used anticoagulants.

A novel Pseudolaric acid B derivative, Hexahydropseudolaric acid B, exterts an immunomodulatory effect in vitro/in vivo evaluation.

Identification of immunosuppressants from natural sources has a proven track record in immune mediated disorders. Pseudolaric acid B is a diterpenoid isolated from the roots of Pseudolarix amabilis, possessing potent immunomodulatory effect. However, the cytotoxicity limits its future clinical application. The purpose of this study was to investigate the immunosuppressive activity of Hexahydropseudolaric acid B, a Pseudolaric acid B derivative, on T cell-mediated immune response both in vitro and in vivo, and investigated its immunomodulatory effect to develop a more ascendant immunosuppressive agent. The results showed that Hexahydropseudolaric acid B could exert more preferable immunosuppressive activity and lower cytotoxicity than Pseudolaric acid B. Hexahydropseudolaric acid B significantly inhibited T cell proliferation activated by mitogen and alloantigen without obvious cytotoxicity in vitro. Furthermore, Hexahydropseudolaric acid B could ameliorate ear swelling in a mouse model of 2,4-dinitrofluorobenzene-induced delayed-type hypersensitivity in vivo. Mechanistic study revealed that Hexahydropseudolaric acid B could enhance regulatory T cells via promoting Foxp3 expression and TGF-β level, accompanied by attenuating Akt activation, blocking p38MAPK/MK2-HSP27 signal cascades, and up-regulating PPAR-γ expression. Taken together, these results suggest that Hexahydropseudolaric acid B exerts more preferable immunosuppressive activity than its precursor Pseudolaric acid B by affecting multiple targets, which support the need for continued efforts to characterize the efficacy of HPAB as a promising and safe candidate to treat immune-related diseases.

Pseudolaric acid B attenuates atherosclerosis progression and inflammation by suppressing PPARγ-mediated NF-κB activation

Aims/objective: Atherosclerosis is a progressive disease of large arteries characterized with chronic inflammation and aberrant immune response. Pseudolaric acid B (PB) has been found to exert multiple effects by inhibiting inflammatory response. However, there is no comprehensive assessment of the effects of PB on atherosclerosis using relevant in vivo and in vitro models. Material and methods: Male ApoE−/− mice were treated with PB orally with a high fat diet (HFD) to clarify its anti‐atherosclerotic activities. RAW264.7 macrophage line, a well‐accepted cell model of atherosclerosis, was used to investigate anti‐inflammatory effects and molecular mechanisms of PB. Results: PB significantly attenuated atherosclerotic lesions by modulating plasma lipid profiles as well as inhibiting inflammatory responses in macrophages of atherosclerotic mice. Meanwhile, PB markedly suppressed the expression of pro‐inflammatory cytokines, and regulated cholesterol efflux related genes in oxidative low density lipoprotein (ox‐LDL)‐loaded macrophages. The cellular uptake of Dil‐labeled ox‐LDL was significantly inhibited by PB either. Moreover, the ability of PB to suppress nuclear factor kappa B (NF‐&kgr;B) and activate peroxisome proliferator‐activated receptor gamma (PPAR&ggr;) was confirmed using luciferase reporter assays. Conversely, the selective PPAR&ggr; antagonist GW9662 reversed the influence of PB in macrophages. Conclusion: Together, these findings indicate that PB exerts its protective effects on atherosclerosis by inhibiting macrophage‐mediated inflammatory response and cellular ox‐LDL uptake, and promoting cholesterol efflux by suppressing NF‐&kgr;B activation PPAR&ggr;‐dependently. Therefore, PB may be a promising agent for inflammatory and atherosclerotic diseases. HIGHLIGHTSWe have identified a novel action of Pseudolaric acid B (PB) on atherosclerosis.PB enhances cholesterol efflux related genes in lipid‐loaded macrophages, thereby reducing the cellular uptake of ox‐LDL.PB alleviates the progression of atherosclerosis through inhibiting inflammatory response both in vitro and in vivo.The mechanisms of PB might be related to the inhibition of NF‐&kgr;B activation via a PPAR‐&ggr;‐dependent manner.

Discordance Between VASP Phosphorylation and Platelet Aggregation in Defining High On-Clopidogrel Platelet Reactivity After ST-Segment Elevation Myocardial Infarction

To investigate potential clinical characteristics associated with discordance between platelet vasodilator-stimulated phosphoprotein phosphorylation (VASP-P) flow cytometry (FCM) assay and light transmission aggregometry (LTA) in defining high on-clopidogrel platelet reactivity (HPR) after ST-segment elevation myocardial infarction (STEMI). In this study, platelet responsiveness was measured by the above 2 methods simultaneously on day 1 and on day 6 of STEMI onset in 90 consecutive patients who underwent primary percutaneous coronary intervention. The FCM-derived platelet reactivity index and LTA-derived platelet aggregation rate were both significantly reduced after dual antiplatelet therapy on day 6. Multiple variable-adjusted logistic regression analysis revealed that smoking (odds ratio [OR]: 4.507, 95% confidence interval [CI]: 1.123-18.09, P = .034) and onset-to-admission time (per 1 hour increase, OR: 1.196, 95% CI: 1.023-1.398, P = .025) both were independent predictors for the discordance between the 2 methods. Additionally, improved correlation and concordance was observed in nonsmokers compared with smokers. Our data show that smoking and prolonged onset-to-admission time are associated with discordance between platelet VASP-P and LTA in defining HPR after STEMI, which should be considered when planning personalized antiplatelet therapy.

GW25-e3231 Increased Admission CD14++CD16– Monocyte to Lymphocyte Ratio Predicts Early Cardiovascular Events in Patients with ST-Elevation Myocardial Infarction

To investigate the relationships between admission monocyte subsets and early cardiovascular outcomes in patients with ST-elevation myocardial infarction (STEMI) undergoing primary coronary interventions (PCI). Flow cytometry (FCM) analysis of monocyte subsets was carried out upon admission in 100