Endothelial microparticle-associated protein disulfide isomerase increases platelet activation in diabetic coronary heart disease.

BACKGROUND: Endothelial microparticles (EMPs) carrying the protein disulfide isomerase (PDI) might play a key role in promoting platelet activation in diabetes. This study aimed to examine the activation of platelets, the amounts of MPs, PMPs, and EMPs, and the concentration and activity of PDI in patients with diabetic coronary heart disease (CHD) and non-diabetic CHD. METHODS: Patients with CHD (n=223) were divided as non-diabetic CHD (n=121) and diabetic CHD (n=102). Platelet activation biomarkers, circulating microparticles (MPs), the concentration of protein disulfide isomerase (PDI), and MP-PDI activity were determined. The effect of EMPs on platelet activation was investigated in vitro. Allosteric GIIb/IIIa receptors that bind to PDI were detected by a proximity ligation assay (PLA). RESULTS: Platelet activation, platelet-leukocyte aggregates, circulating MPs, EMPs, PDI, and MP-PDI activity in the diabetic CHD group were significantly higher than in the non-diabetic CHD group (P<0.05). Diabetes (P=0.006) and heart rate <60 bpm (P=0.047) were associated with elevated EMPs. EMPs from diabetes increased CD62p on the surface of the platelets compared with the controls (P<0.01), which could be inhibited by the PDI inhibitor RL90 (P<0.05). PLA detected the allosteric GIIb/IIIa receptors caused by EMP-PDI, which was also inhibited by RL90. CONCLUSIONS: In diabetic patients with CHD, platelet activation was significantly high. Diabetes and heart rate <60 bpm were associated with elevated EMPs and simultaneously increased PDI activity on EMP, activating platelets through the allosteric GPIIb/IIIa receptors.

[Comparison of Plasma Components between Frozen Plasma and Fresh Frozen Plasma].

OBJECTIVE: To compare the plasma components of frozen plasma (FP) and fresh frozen plasma (FFP). METHODS: Twenty samples of FP and 20 samples of FFP from Beijing Red Cross Blood Center were randomly selected. Immediately after plasma melting, 12 plasma components including coagulation factor, fibrinolytic system and anticoagulation protein were detected, including activated partial thromboplastin time (APTT), prothrombin time (PT), coagulation factor Ⅷ (FⅧ) activity, coagulation factor Ⅴ (FⅤ) activity, fibrinogen(FIB) level, ADAMTS-13 activity, von Willebrand factor(vWF) activity, D-dimer (D-dimer, DD), fibrin degradation products (FDP), antithrombin (AT), protein C (PC), and protein S (PS). All these coagulation components between the two types of plasma were compared and analyzed. RESULTS: Compared with FFP, APTT in FP was significantly prolonged(t=3.428, P<0.01), and PT was also significantly prolonged(z=-2.140, P<0.05), and FⅧ activity was decreased (t=-3.372, P<0.01), but all in the reference range, and PS activity was decreased(t=-2.458，P＜0.05), there was no statistical difference in the other part between two types of plasma(P>0.05). CONCLUSION: FP can substitute FFP in the treatment of some diseases, although it is lack of some coagulation factors and anticoagulation protein.

The Role of T Follicular Helper Cells and T Follicular Regulatory Cells in the Pathogenesis of Autoimmune Hemolytic Anemia.

Autoimmune hemolytic anemia (AIHA) is an acquired autoimmune disease mediated by antibodies against the patient's red blood cells. However, the underlying mechanisms for antibody production are not fully understood. Previous studies of etiology and pathogenesis of AIHA mainly focus on autoreactive B cells that have escaped tolerance mechanisms. Few studies have reported the function of TFH and TFR cells in the process of AIHA. The present study aimed to explore the potential mechanism of TFH and TFR cells in the pathogenesis of AIHA. With the model of murine AIHA, increased ratios of TFH:TFR, elevated serum IL-21 and IL-6 levels, and upregulated Bcl-6 and c-Maf expression were reported. Also, adoptive transfer of purified CD4+CXCR5+CD25- T cells from immunized mice promoted the induction of autoantibody in the AIHA mouse model. Altogether, our data demonstrate the important role of TFH cells for control and induction of AIHA. In the light of the key contributions of TFH cells to the immune response in AIHA, strategies aimed at inhibiting the TFH development or function should be emphasized.

Platelet activation in diabetic mice models: the role of vascular endothelial cell-derived protein disulfide isomerase-mediated GP IIb/IIIa receptor activation.

GP IIb/IIIa receptor activation plays an important role in thrombosis. The mechanism of early activation of GP IIb/IIIa receptors in diabetic conditions remains unknown. The purpose of this study was to investigate the release of Endothelial microparticle (EMP)-associated protein disulfide isomerase (PDI) after endothelial cell injury induced in diabetes and the changes in platelet activation. We produced an animal model of type 2 diabetes mellitus using ApoE-/- mice. Normal ApoE-/- and diabetic mice were allocated to four groups (n = 15): normal diet, normal diet plus rutin, diabetic, and diabetes plus rutin. The EMP-PDI content and GP IIb/IIIa expression of mice platelets were determined. In addition, EMPs obtained from the four groups were pretreated with the PDI inhibitor rutin; then, their effects on the platelets of normal C57 mice were characterized. Compared with the normal diet group, the diabetic group had significantly increased plasma EMP-PDI content and accelerated platelet activation by increased GP IIb/IIIa expression. In conclusion, EMP-PDI promotes early platelet activation through glycoprotein (GP) IIb/IIIa receptors present on platelet surface in the diabetic state. However, this process could be partially suppressed by the administration of rutin.

PCR-ESI/MS for Molecular Diagnosis of Bloodstream Infections Directly from Blood Samples: a Systematic Review and Meta-Analysis.

BACKGROUND: PCR-ESI/MS is a commercial molecular method that can identify bloodstream infection (BSI) pathogens directly from blood. Previous studies showed its sensitivity varied greatly. Its diagnostic accuracy has not been systematically evaluated yet, thus we aimed to assess its accuracy by systematic review and meta-analysis. METHODS: Studies were searched on PubMed and Embase up to November 2017, for studies using PCR/ESI-MS to diagnose BSI directly from blood and providing sufficient data to construct two-by-two tables. Subgroup analysis and meta-regression were used to assess heterogeneity. RESULTS: A total of nine studies including 3,392 patients met the inclusion criteria. Their pooled sensitivity and specificity was 0.66 (95% CI: 0.57 - 0.74) and 0.84 (95% CI: 0.78 - 0.89), respectively. The positive and negative likelihood ratios were 4.2 (95% CI: 3.0 - 5.9) and 0.40 (95% CI: 0.31 - 0.50), respectively. The diagnostic odds ratio (DOR) was 10.61 (95% CI: 6.67 - 16.88). The area under the SROC was 0.82 (95% CI: 0.78 - 0.85). High heterogeneity was found. Subgroup analysis and meta-regression showed that regions, BSI prevalence, blood volume, and settings may cause heterogeneity. CONCLUSIONS: PCR-ESI/MS using blood specimens directly has the potential for early diagnosis of BSI, compared with blood culture. Its rule-in value is higher than rule-out value. Due to the heterogeneity of currently available studies, further high-quality studies are still needed.

Trimetazidine restores the positive adaptation to exercise training by mitigating statin-induced skeletal muscle injury.

BACKGROUND: Exercise rehabilitation is demonstrated to improve the prognosis of patients with coronary heart disease (CHD). Statins, as the key medicine to lower cholesterol in CHD, result in skeletal muscle injury and impair exercise training adaptation. Energy metabolism dysfunction is identified as the potential mechanism underlying statin-induced skeletal muscle injury. In this study, we investigated the effects of the metabolic modulator trimetazidine on skeletal muscle energy metabolism and statin-associated exercise intolerance. METHODS: High-fat fed apolipoprotein E knockout (ApoE-/- ) mice were given aerobic exercise and administrated simvastatin, trimetazidine, or simvastatin plus trimetazidine by gavage. Exercise capacity was evaluated at the end of the treatment by hanging grid test, forelimb grip strength, and running tolerance test. Plasma glucose, lipid, and creatine kinase concentrations were measured at the end of the treatment. After sacrifice, gastrocnemii were stored for assessment of muscle morphology and fibre type. Energy metabolism was estimated by plasma lactic acid concentration, ragged red fibres, and glycogen stores. Activities of mitochondrial complex III, citrate synthase activity, and membrane potential were measured to assess mitochondrial function. Oxidative stress was also evaluated by superoxide in mitochondria, superoxide dismutase activity, and glutathione redox state. RESULTS: In high-fat fed ApoE-/- mice, exercise training had no effect on lipid concentrations. Lower lipid concentrations with increased creatine kinase were observed with additional simvastatin treatment. Exercise capacity increased significantly in response to exercise training alone but was blunted by the addition of simvastatin. Similarly, cross-sectional area of muscle fibres and the proportion of slow-twitch fibres increased in the exercise group but decreased in the simvastatin plus exercise group. Additionally, simvastatin increased centronucleated fibres and induced energy metabolism dysfunction by inhibiting complex III activity and thus promoted oxidative stress in gastrocnemius. We demonstrated that trimetazidine could reverse simvastatin-induced exercise intolerance and muscle damages. We also found the ability of trimetazidine in restoration of muscle fibre hypertrophy and facilitating fast-to-slow type shift. The energy metabolism dysfunction and oxidative stress in gastrocnemii were rescued by trimetazidine. CONCLUSIONS: Trimetazidine alleviated statin-related skeletal muscle injury by restoration of oxidative phenotype and increasing fibre cross-sectional areas in response to exercise training. Correspondingly, the exercise training adaptation were improved in high-fat fed ApoE-/- mice. Moreover, trimetazidine is able to exert its positive effects without affecting the beneficial lipid-lowering properties of the statins. Thus, trimetazidine could be prescribed to remedy the undesirable statins-induced exercise intolerance during cardiac rehabilitation in patients with CHD.

Inhibition of vascular smooth muscle cells premature senescence with rutin attenuates and stabilizes diabetic atherosclerosis.

Atherosclerosis is an age-associated disease; however, diabetic atherosclerosis has higher severity beyond age range for accumulative premature senescent cells in diabetes. Recent findings suggest that rutin, a flavonoid, has potential benefits for diabetic individuals. This study was designed to evaluate the effects of rutin on premature senescence and atherosclerosis. Apolipoprotein E knockout mice exhibiting insulin resistance after 6 weeks of high-fat diet were administered with a low dose of streptozotocin (STZ) to induce diabetes. After 8 weeks of STZ administration, rutin (40 mg/kg/d) was supplemented by gavage for the last 6 weeks. We evaluated the prosperity of the plaque and diabetes using serial echocardiography, histopathologic and metabolite analysis. Premature senescence induced by hydrogen peroxide in primary vascular smooth muscle cells (VSMCs) was used to analyze the underlying mechanism. Mice with diabetes showed more severe plaque burden on aortic arteries and less smooth muscle cells but larger senescent cell ratio in plaque compared with mice with control diets. Rutin significantly improves glucose and lipid metabolic disturbance in diabetes. Moreover, rutin decreased the atherosclerotic burden and senescent cell number and increased the VSMC ratio in aortic root plaque. In vitro, we demonstrated that rutin ameliorated premature senescence induced by oxidative stress, and the protective function may be mediated by inhibiting oxidative stress and protecting telomere. Rutin administration attenuates atherosclerosis burden and stabilizes plaque by improving metabolic disturbance and alleviating premature senescence of VSMCs. Inhibition of VSMCs premature senescence with rutin may be an effective therapy for diabetic atherosclerosis.

Risk Factors for Carbapenem-resistant Klebsiella pneumoniae Infection and Mortality of Klebsiella pneumoniae Infection.

BACKGROUND: Klebsiella pneumoniae (KP) is a pathogen commonly causing nosocomial infection. Carbapenem-resistant KP (CRKP) is more resistant to multiple antimicrobial drugs than carbapenem-susceptible KP (CSKP) isolates. The aim of the present study was to identify the risk factors for CRKP infection and the predictors of mortality among KP-infected adult patients. METHODS: Patients with CRKP and CSKP infection were categorized as the case group and control group, respectively, and we conducted a 1:1 ratio case-control study on these groups. The CRKP isolates collected were tested for antimicrobial susceptibility and presence of KP carbapenemase (KPC) gene. Clinical data were collected to identify risk factors for CRKP infection and mortality of KP infection. Risk factors were analyzed under univariable and multivariable logistic regression model. RESULTS: The independent risk factors for CRKP infection were admission to Intensive Care Unit (odds ratio [OR]: 15.486, 95% confidence interval [CI]: 3.175-75.541, P < 0.001); use of ß-lactams and ß-lactamase inhibitor combination (OR: 4.765, 95% CI: 1.508-15.055, P = 0.008); use of cephalosporins (OR: 8.033, 95% CI: 1.623-39.763, P = 0.011); fluoroquinolones (OR: 6.090, 95% CI: 1.343-27.613, P = 0.019); and indwelling of urethral catheter (OR: 6.164, 95% CI: 1.847-20.578, P = 0.003). However, older age (OR: 1.079, 95% CI: 1.005-1.158, P = 0.036), Charlson comorbidity index (OR: 4.690, 95% CI: 2.094-10.504, P = 0.000), and aminoglycoside use (OR: 670.252, 95% CI: 6.577-68,307.730, P = 0.006) were identified as independent risk factors for patient deaths with KP infection. The mortality of CRKP group was higher than that of the CSKP group. KPC gene did not play a role in the CRKP group. CRKP mortality was high. CONCLUSION: Implementation of infection control measures and protection of the immunefunction are crucial.

Overexpressing STAMP2 attenuates adipose tissue angiogenesis and insulin resistance in diabetic ApoE-/- /LDLR-/- mouse via a PPARγ/CD36 pathway.

The aim of this study was to investigate whether overexpression of STAMP2 improves insulin resistance by regulating angiogenesis in adipose tissues. The characteristics of diabetic mice were measured by serial metabolite and pathology tests. Samples were obtained from epididymal, subcutaneous and brown adipose tissues. Histological and morphological analysis demonstrated that STAMP2 gene overexpression reduced adipocyte size, angiogenesis in epididymal and brown adipose tissues. On aortic ring assay, microvessels sprouting from aortas were significantly inhibited after STAMP2 gene overexpression. The cellular effect of STAMP2 on angiogenesis was explored in human umbilical vein endothelial cells (HUVECs) model. Correlation of STAMP2 and angiogenesis was validated by Ad-STAMP2 transfection and STAMP2 siRNA inhibition. In vitro, overexpression of STAMP2 significantly inhibited endothelial cell migration, tube formation. The effects of Ad-STAMP2 transfection on HUVECs were abolished by treatment with PPARγ antagonist GW9662 (2.5 µM), and the roles of STAMP2 siRNA on HUVECs were also reversed by treatment with PPARγ agonist rosiglitazone (RSG) (0.1 mM). RT-PCR indicated that STAMP2 could regulate levels of adhesion molecules, vascular endothelial growth factor A and CD36. The expression of PPARγ and CD36 was decreased when STAMP2 was inhibited by siRNA, while PPARγ and CD36 were highly expressed after overexpression of STAMP2. Our results suggested that STAMP2 gene overexpression may improve insulin resistance via attenuating angiogenesis in epididymal and brown adipose tissues through the PPARγ/CD36 signalling pathway.

Association of Increased Serum Sema3E with TRIB3 Q84R Polymorphism and Carotid Atherosclerosis in Metabolic Syndrome.

OBJECTIVES: Semaphorin-3E, provoking inflammation and insulin resistance which leads to metabolic syndrome, exerts obscure effects on carotid atherosclerosis in metabolic syndrome. The Tribbles 3 Q84R polymorphism is involved in insulin resistance. This study aims to investigate whether the Tribbles 3 Q84R polymorphism has profound effects on serum semaphorin 3E and what effect semaphorin 3E exerts on carotid atherosclerosis. METHOD: A total of 518 unrelated Chinese subjects consisted of control (n=258) and metabolic syndrome (n=260) groups. Clinical and biochemical characteristics were collected. The level of serum semaphorin 3E was measured by enzyme-linked immunosorbent assay. Genotyping of the functional Tribbles 3 Q84R polymorphism was achieved by RFLP-PCR method. All subjects underwent carotid ultrasonography to determine carotid intima-media thickness (considered atherosclerosis if above 0.9 mm). RESULTS: Serum semaphorin 3E was significantly increased in metabolic syndrome (P=0.012) as compared with the control group. Among the metabolic syndrome group, those with RR84 genotype had significantly higher levels of serum semaphorin 3E than those with QQ84 or QR84 genotypes (P=0.003, P=0.004) with similarity amongst the control group. Factorial analyses showed statistically significant interactions between RR84 genotype and metabolic syndrome (P=0.019 for interaction for semaphorin 3E). Semaphorin 3E correlated positively with homeostasis model assessment insulin resistance (r=0.263, P=0.009) and carotid intima-media thickness (r=0.215, P=0.031). On partial analyses, after controlling for confounding factors, semaphorin 3E was positively correlated with carotid intima-media thickness (P=0.004). CONCLUSION: Individuals with metabolic syndrome demonstrated further increased serum semaphorin 3E, especially those with Tribbles 3 RR84 genotype. Increased serum semaphorin 3E may in part exacerbate insulin resistance and carotid atherosclerosis.

Endothelial cells microparticle-associated protein disulfide isomerase promotes platelet activation in metabolic syndrome.

BACKGROUND: Metabolic syndrome (MetS) is a common challenge in the world, and the platelet activation is enhanced in MetS patients. However, the fundamental mechanism that underlies platelet activation in MetS remains incompletely understood. Endothelial cells are damaged seriously in MetS patients, then they release more endothelial microparticles (EMPs). After all, whether the EMPs participate in platelet activation is still obscure. If they were, how did they work? RESULTS: We demonstrated that the levels of EMPs, PMPs (platelet derived microparticles) and microparticle-carried-PDI activity increased in MetS patients. IR endothelial cells released more EMPs, the EMP-PDI was more activated. EMPs can enhance the activation of CD62P, GPIIb/IIIa and platelet aggregation and this process can be partly inhibited by PDI inhibitor such as RL90 and rutin. Activated platelets stimulated by EMPs expressed more PDI on cytoplasm and released more PMPs. MATERIALS AND METHODS: We obtained plasma from 23 MetS patients and 8 normal healthy controls. First we built insulin resistance (IR) model of human umbilical vein endothelial cells (HUVECs), and then we separated EMPs from HUVECs culture medium and used these EMPs to stimulate platelets. Levels of microparticles, P-selectin(CD62P), Glycoprotein IIb/IIIa (GPIIb/IIIa) were detected by flow cytometry and levels of EMPs were detected by enzyme-linked immunosorbent assay (ELISA). The protein disulfide isomerase (PDI) activity was detected by insulin transhydrogenase assay. Platelet aggregation was assessed by turbidimetry. CONCLUSION: EMPs can promote the activation of GPIIb/IIIa in platelets and platelet aggregation by the PDI which is carried on the surface of EMPs.