Effect of polymorphonuclear leukocytes on washed platelet aggregation

AIM:To investigate the effect of non-activated or activated polymorphonuclear leukocytes(PMN) on washed platelet aggregation. METHODS:Born's method was used to determine platelet aggregation.RESULTS:non-activated PMN (5×10⁹ cells/L) significantly suppressed washed platelet aggregation induced by ADP or arachidonic acid. Aspirin enhanced this inhibition. N-formyl-methiongl-leucy-phenylalanine (fMLP)-or platelet-activating factor (PAF)-stimulated PMN strongly induced platelet aggregation, and the induction effect of PMN suspension was more active than that of PMN supernatant. Aspirin had no significant inhibitory effect on platelet aggregation induced by fMLP-or PAF-activated PMN. CONCLUSIONS:Different conditions of PMN (activated or non-activated) had the nearly opposite action on normal platelet reactivity. Briefly, non-activated-PMN inhibited platelet reactivity, whereas activated PMN stimulated it.     

In vitro anti-platelet aggregative effect of procyanidins isolated from grape seeds

AIM: To study the possible anti-platelet aggregative mechanisms of procyanidins (PC) isolated from grape seeds in vitro. METHODS: Platelet-rich plasma (PRP) and platelet-poor plasma (PPP) were prepared from the blood of healthy volunteers. PC,diphenylene iodonium(DPI,a nonspecific NADPH oxidase inhibitor) and apocynin (a specific NADPH oxidase inhibitor) were used to observe the effects on collagen-induced platelet maximum aggregation rate using platelet aggregometer. The influences of PC on platelet NADPH oxidase activity, NO content and superoxide anion (O₂^-·) level were evaluated by chemiluminescence spectrometer. The role of PC in the expression of activated platelet markers (PAC-1 and CD62P) was observed by flow cytometry. RESULTS: PC (100 μmol/L), apocynin (10 μmol/L) and DPI (100 μmol/L) significantly inhibited collagen-induced maximum platelet aggregation rate (P<0.01). In collagen-activated platelets, NO content reduced and O₂^-· level increased,both of which were recovered by PC at concentration of 100 μmol/L (P<0.05). PC also obviously inhibited NADPH oxidase activity (P<0.01), and significantly down-regulated PAC-1 and CD62P expression (P< 0.05) in platelets. CONCLUSION: Procyanidins isolated from grape seeds have the anti-platelet aggregation function through inhibiting NADPH oxidase activity, further influencing platelet NO and O₂^-· levels.     

Effects of oxidative modification of lipoproteins on blood coagulation and fibrinolysis

AIM: To study the effects of oxidative modification lipoproteins on blood coagulation and fibrino-lysis in vitro. METHODS: Normal human plasma VLDL, LDL and HDL, which were isolated by density gradient ultracentrifugation method, were oxidatively modified by Cu²⁺ and HOCl method. N-VLDL, Ox-VLDL, N-LDL, Ox-LDL, N-HDL, Ox-HDL were added to the reaction system which consisted of mixed fresh normal plasma respectively, prothrombin time (PT), activated partial thrombplastin time (APTT), plasminogen activator inhibitor 1 (PAI-1), tissue plasminogen activator (t-PA) and platelet aggregation were measured according to the direction of the kits. RESULTS: The relative electrophoretic mobility (REM), absorbance at 234nm and TBARS of oxidized VLDL, LDL and HDL mediated by HOCl or Cu²⁺ were significantly higher than that of the control group (P<0.01). N-VLDL, N-LDL and N-HDL had no effect on PT, APTT, t-PA, PAI-1 and platelet aggregation. The PT and APTT of Ox-VLDL, Ox-LDL and Ox-HDL were significantly shorter than that of the control group (P<0.05 and P<0.01). The platelet aggregation of Ox-VLDL, Ox-LDL and Ox-HDL were significantly stronger than that of the control group (P<0.01). The Ox-VLDL and Ox-LDL were higher in t-PA and lower in PAI-1 than that of the control group (P<0.05 and P<0.01), but the Ox-HDL had no influences on t-PA and PAI-1 activity. CONCLUSIONS: N-VLDL, N-LDL and N-HDL have no effects on blood coagulation and fibrinolysis in vitro. Ox-VLDL, Ox-LDL and Ox-HDL enhance blood coagulation and thrombosis. Ox-VLDL and Ox-LDL enhance t-PA activity and decreased PAI-1 activity, but Ox-HDL does not affect the fibrinolysis activity.     

Effects of bortezomib-based therapy on hemostasis system and circulating microparticles in multiple myeloma patients

AIM：To investigate the changes of hemostasis, thrombosis and total microparticles (TMPs) in multiple myeloma (MM) patients receiving bortezomib-based induction therapy (bortezomib+adriamycin+dexamethasone, PAD). METHODS：The levels of TMPs were detected by flow cytometry in 38 newly diagnosed MM patients and 30 healthy people. The changes of the platelet, coagulation, anticoagulation, fibrinolytic activation and TMPs in the MM patients before and after PAD teatment were also studied.RESULTS：Before treatment, the values of FVIII:C and vWF:Rco in MM patient were elevated ［(152.89±31.14)% and (165.69±38.43)%］, the activation of platelet aggregation was inhibited ［(63.76±21.36)%］, and the PAI level increased ［3.98(1.63)U/mL］. Compared with the healthy people, higher concentration of TMPs was observed in MM patients ［(640.65±214.22)/μL vs (134.29±63.09)/μL, P<0.01］, and the level of TMPs was positively correlated with serum β2-MG (r=0.672, P<0.01).After PAD therapy, platelet aggregation activation was restored ［(77.83±15.62)%, P=0.01］, and PAI level decreased ［0.88(1.38)U/mL, P<0.01］. The level of TMPs also decreased, after 3 cycles of PAD, to the value of (184.25±93.35)/μL. CONCLUSION：MM patients were characterized by impaired platelet, coagulation and fibrinolysis functions, and increased level of TMPs. PAD restored platelet function, decreased the levels of PAI and TMPs, which might partially explain the low incidence of thrombosis in the MM patients receiving bortezomib-based treatment.     

Effect of fraction F of naja naja atra venom on protein tyrosine phosphorylation in platelet activation

AIM: To observe the mechanism of intracellular signal transduction that fraction F of naja naja atra venom inhibits platelet aggregation. METHODS: Tests were divided into six groups: (1) blank group; (2) control group and (3)-(6) ADP plus fraction F group (doses of fraction F were 100, 30, 10, 3 mg/L, respectively). Protein tyrosine phosphorylation in platelets was assayed by Western blotting and platelet aggregation was assayed by nephelometer. RESULTS: Fraction F significantly inhibited molecular masses (MW) 76, 66 and 37.5 kD protein tyrosine phosphorylation in platelet that induced by ADP in a dose-dependent manner, in which 30 and 100 mg/L dose group showed obviously different effects when compared to control group (P<0.05). Inhibition of platelet aggregation was positive correlated with protein tyrosine phosphorylation in platelets （r=0.9367, P<0.01）. CONCLUSIONS: Fraction F of naja naja atra venom affected intracellular signal transduction pathway in platelets by inhibiting MW 76, 66 and 37.5 kD protein tyrosine phosphorylation. The result suggests that this effect may be one of the anti-thrombus mechanisms of fraction F.     

Correlation analysis between visualization of hemorrheologic changes and platelet function in patients with acute coronary syndrome after percutaneous coronary intervention

AIM:To intuitionally observe the characteristics of blood rheology in the patients with acute coronary syndrome (ACS) after percutaneous coronary intervention (PCI) for 1 year to 3 years by micro-channel array flow analyzer (MC-FAN) combined with other platelet function indexes, and to explore the correlations between the test results of MC-FAN and platelet function. METHODS:This study brought 74 patients with ACS after PCI for 1 year to 3 years into test group, and 21 healthy subjects were enrolled as normal group. The levels of platelet aggregation test (PAgT), platelet adhesiveness test (PAdT), P-selectin, platelet-derived growth factor BB (PDGF-BB) and von Willebrand factor (vWF) were detected. MC-FAN HR300 was used to detect the transiting time (MC-FAN TT) of the blood passing through the model body capillaries. The differences of the test results between the 2 groups were compared, and the correlations between the results of MC-FAN and platelet function in the patients with ACS after PCI were also explored. RESULTS:Compared with normal group, the MC-FAN TT in test group was prolonged (P<0.01), the ability of erythrocyte deformation was weakened, and the leukocyte attaching the vascular wall and platelet adhesion and aggregation relatively increased. The levels of PAgT, PAdT, P-selectin and PDGF-BB in test group were all higher than those in normal group (P<0.01). No difference of vWF between the 2 groups was observed. The intergroup correlation analysis showed that there were correlations between MC-FAN TT and platelet function, in which 10 μL MC-FAN TT and 30 μL MC-FAN TT had the most significant correlation with P-selectin (r=0601, P<0.01; r=0334, P<0.01), 60 μL MC-FAN TT had the most significant correlation with PAgT (r=0527, P<0.01), and 100 μL MC-FAN TT had the most significant correlation with PAdT (r=0. 815, P<0.05). CONCLUSION:The visualization of hemorrheologic changes and platelet function in the patients with ACS after PCI are abnormal.There are correlations between MC-FAN TT and platelet function.The results of MC-FAN can objectively evaluate the blood rheology of the patients, and provide the reference for clinical treatment.     

Effect of atorvastatin on myocardial fibrosis induced by aldosterone

AIM: To investigate the effects and mechanisms of atorvastatin on myocardial fibrosis induced by aldosterone in SD rats. METHODS: Forty male uninephrectomized SD rats were limited to drink 1% NaCl water for 4 weeks and assigned to the follow groups: vehicle control rats (CON group); aldosterone treated rats (ALD group); spironolactone + aldosterone treated rats (SPI+ALD group); atorvastatin + aldosterone treated rats (ATO+ALD group). Blood pressure was measured by catheterization. Expression of platelet-derived growth factor (PDGF-A, PDGF-B), platelet-derived growth factor receptor (PDGFR-α, PDGFR-β) and ectodermal dysplasia-1 (ED-1) were analyzed by immunohistochemistry. Collagen volume fraction (CVF) and perivascular collagen area (PVCA) were analyzed by Sirius-Red staining. Myocardium osteopontin protein was detected by Western blotting analysis. RESULTS: Mean arterial blood pressure in ALD group, SPI+ALD group and ATO+ALD group was elevated, and significant difference was observed between the three groups and vehicle control group (P<0.01 or P<0.05). Myocardial fibrosis was observed in ALD group. Compared to other three groups, the index of CVF and PVCA was increased significantly (P<0.01 or P<0.05). No significant difference of the index of CVF and PVCA between ATO+ALD group and SPI+ALD group was observed (P>0.05). Compared to other groups, the levels of PDGF-A, PDGF-B, PDGFR-α, ED-1 and OPN in ALD group were significantly increased (P<0.01 or P<0.05). The levels of PDGF-A, PDGF-B, PDGFR-α and OPN were no significant difference between ATO+ALD group and SPI+ALD group (P<0.01 or P<0.05). However, the level of ED-1 in ATO+ALD group was significantly decreased compared to SPI+ALD group (P<0.05). No significant difference of PDGFR-β level among four groups was found (P>0.05). CONCLUSION: These results suggest that atorvastatin may attenuate myocardial fibrosis induced by aldosterone. The mechanisms concern with reduction of macrophage infiltration, expression of inflammatory cytokines OPN, partially inhibition of platelet-derived growth factor and its receptor expression.     

Application of anti-CD20 antibody in hematopoietic stem cell transplantation for sensitized recipients

AIM: To find a strategy for enhancing engraftment of hematopoietic stem cell in sensitized recipients and to study the effects of anti-CD20 antibody in hematopoietic stem cell transplantation. METHODS: BALB/c mice were sensitized by transfusions of allogeneic spleen cells on 14 d and 7 d. Anti-CD20 antibody (2 mg/mouse) was intravenously injected into sensitized recipients on 11 d. The recipients were used as experimental group, while RPMI-1640 medium (0.2 mL/mouse) was used as control. The sera and splenocytes obtained from the recipients were tested for donor reactive antibody and CD19+ B cells on 0 d. In addition, the recipients were transplanted with 1×107 C57BL/6 bone marrow cells after lethal irradiation on 0 d. The survival rates were observed and blood counts were studied post transplantation. RESULTS: The cytotoxic index in the experimental group and control group were (37.00±3.46)% and (51.80±3.49)%, respectively, and the differences were significant (P<0.01). The percentages of CD19+ B cells in experimental group and control group were (17.32±3.02)% and (34.26±2.87)%, respectively, and the differences were significant (P<0.01). All the recipients in both experimental group and control group died about 14 d post transplantation. The median time was 13 d and 11 d in experimental group and control group, respectively, and no significant difference was found between these two groups (P>0.05). Moreover, a rapid disappearance was observed in the white blood counts, hemoglobin, and platelet of dying animals, indicating the animals died from hematopoietic failure. CONCLUSION: Anti-CD20 antibody is able to deplete B cells and reduce the level of antibody in sensitized recipients, but it can’t enhance the engraftment of allogeneic hematopoietic stem cells in the sensitized recipients.     

Oxidized low-density lipoprotein induces autophagy in macrophages via CD36-mediated oxidative stress

AIM: To investigate the effect of oxidized low-density lipoprotein (ox-LDL) on autophagy in macrophages and the underlying molecular mechanisms. METHODS: RAW264.7 macrophages were pretreated with 2 mg/L anti-CD36 monoclonal antibody (anti-CD36 mAb), 5 μmol/L diphenyleneiodonium (DPI), 3 mmol/L 3-methyladenine (3-MA) or 1 μmol/L rapamycin for 1 h and then treated with ox-LDL (100 mg/L) for 12 h. The viability of the cells was measured by MTT assay. The activities of lactic dehydrogenase (LDH) in the medium and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, superoxide dismutase (SOD) in the cells as well as the levels of intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) were determined to characterize the membrane integrity and the oxidative stress, respectively. The protein levels of beclin-1 and microtubule-associated protein 1 light chain 3-II (LC3-II), 2 important molecular markers of autophagy, were examined by Western blotting. RESULTS: ox-LDL induced autophagy in RAW264.7 macrophages as assessed by upregulation of beclin-1 and LC3-II. Similar to 3-MA, an autophagy inhibitor, anti-CD36 mAb significantly inhibited the ox-LDL-induced upregulation of beclin-1 and LC3-II. Anti-CD36 mAb suppressed the ox-LDL-induced oxidative stress as revealed by decreased NADPH oxidase activation, ROS and MDA generation as well as increased SOD activity. Similar results were observed in the cells pretreated with DPI, a NADPH oxidase inhibitor. Moreover, DPI significantly inhibited the ox-LDL-induced upregulation of beclin-1 and LC3-II. Inaddition, the decrease in the cell viability and increase in LDH release induced by ox-LDL were promoted by 3-MA and blocked by rapamycin (an autophagy inducer). CONCLUSION: ox-LDL induces autophagy in RAW264.7 macrophages, which may be involved in CD36-mediated ox-LDL uptake and subsequent activation of oxidative stress, and moderate activation of autophagy may protect macrophages from ox-LDL-induced injury.     

The biological characteristics of cytokine-induced killer cells

AIM:To investigate the biological characteristics of cytokine-induced killer (CIK) cellsin vitro.METHODS:The non-adhere peripheral blood monoclear cells from healthy donors were induced into CIK cells in the presence of IFN-γ, IL-1β, IL-2 and anti-CD3 antibody. LAK (lymphokine activated killer) cells were prepared as a control. The cellular phenotype were detected by FCM and immunocytochemistry and the cytotoxicity was measured by LDH release assay.RESULTS:After 2 weeks of induction, the proliferation rate of CIK cells reached a peak and the proportion of CD3⁺ population was above 95%, and then the cells growth entered to plateau phase at week 3. The proportion of CD3⁺CD56⁺ NKT subset cells was 16.5% on day 15 and it had no obvious variety between 2 and 4 weeks. Correspondingly, LAK cells grew slowly and had lower proliferation rate compared with the CIK cells (P<0.01). CIK cells showed higher specific lysis rates to BeL-7402 hepatoma cells than those of LAK cells at different effector to target ratio (P<0.01). Immunocytochemical staining showed the CIK cells highly co-expressed HLA-DR and CD54 antigens. The NKT cells were slightly bigger than CD3⁺CD56^- cells and a large quantity of pseudopodia were observed on their surface.CONCLUSION:The CIK cells have higher proliferation potency and stronger cytotoxicity to lyse tumor cells than LAK cellsin vitro.Within the span of time from 14 to 21 days, the proliferation rate and the proportion of CD3⁺CD56⁺ subset of CIK cells all reach peaks. Therefore, CIK cells in this period are suitable for clinical application.     

Polysaccharide extracted from laminaria japonica aresch inhibits platelet activation and protects endothelial cells

AIM: To study the relationship between the effects of polysaccharide L01 extracted from laminaria japonica aresch on platelet activation and endothelial cells. METHODS: A rat model of endothelial injury was established via injecting adrenaline. The percentage of platelet adhesion was evaluated by filtration method, the activation of platelet aggregation was observed on a glass plate with collodion membrane, the content of vWF in rat plasma was measured by ELISA, the damaged degree of aortic vascular endothelial was evaluated by immunity histochemistry. RESULTS: The percentage of platelet adhesion and aggregation in model group were higher than those in NS group from the 3th and 4th day during the model made (P<0.05, P<0.01). The percentage in both L01 high-dose group (50 mg/kg) and low-dose group (10 mg/kg) at the 4th and 5th day was lower than that in model group (P<0.05, P<0.01). The content of vWF in rat plasma in model group was higher than that in NS group and in L01 high-dose group at 4th day (P<0.05). The same results were presented by the comparison among model group and NS group, both L01 high-dose group and low-dose group at the 5th (P<0.05). The measure of intact endodermis lengths (μm) stained by immunohistochemistry demonstrated that the length in model group was shorter than that in NS group (P<0.05), whereas the length in L01 high-dose group and low-dose group was obviously longer than that in model group (P<0.05) at the 4th and 5th day. CONCLUSION: The inhibitory effect of L01 on platelet activation may be related with its protective effect on vascular endothelial cells.     

Effect of nitric oxide on adiponectin-induced inhibition of platelet aggregation in hyperlipidemia rats

AIM: To investigate the mechanism that adiponectin inhibits platelet aggregation via nitric oxide (NO) signaling pathway. METHODS: Adult rats were fed with normal or high-fat diet for 14 weeks. Their platelets were immediately isolated and treated with or without recombinant full-length adiponectin (rAPN). The platelet aggregation, NO and superoxide production, endothelial nitric oxide synthase (eNOS)/inducible NOS (iNOS) expression, and antioxidant capacity were determined. RESULTS: Treatment with rAPN inhibited platelet aggregation induced by hyperlipidemia (P<0.05). Interestingly, total NO, a crucial molecule depressing platelet aggregate and thrombus formation, was significantly reduced, rather than increased in rAPN-treated platelets. Treatment with rAPN significantly decreased superoxide production by 62% (P<0.05) and increased antioxidant capacity by 38% (P<0.05) in hyperlipidemic platelets. Importantly, hyperlipidemia-induced reduction of eNOS phosphorylation and increase in iNOS expression were markedly reversed by rAPN treatment (P<0.05 and P<0.01, respectively). CONCLUSION: Adiponectin is an adipokine that inhibits platelet aggregation by enhancing eNOS activation and attenuating oxidative/nitrative stress including blockage of iNOS expression and superoxide production.     

Protective effect of aFGF delivery by ultrasound-targeted microbubble destruction on left ventricular function in diabetic cardiomyopathy rats

AIM：To observe the protective effect of delivery of acidic fibroblast growth factor (aFGF) to myocardium by ultrasound-targeted microbubble destruction (UTMD) on left ventricular function in diabetic cardiomyopathy (DCM) rats and to investigate the possible mechanisms. METHODS：Twenty-four rats were intraperitoneally injected with streptozocin to induce DCM and were randomly divided into DCM group and aFGF treatment group. Twelve healthy rats served as normal controls. The rats in aFGF treatment group were infused with SonoVue-aFGF mixed fluid through tail vein and UTMD was simultaneously performed. Four weeks after intervention, all rats underwent cardiac catheterization to mea-sure left ventricular end-systolic pressure (LVESP), left ventricular end-diastolic pressure (LVEDP) and the maximal increase/decrease rate of left ventricular pressure (LV±dp/dtmax). The microvessel density (MVD) of rat myocardial tissues was measured by immunohistochemical staining for CD31. The myocardial collagen volume fraction (CVF) was determined by improved Masson staining. The apoptotic index (AI) was detected by TUNEL method. RESULTS：Four weeks after intervention, the LVESP and LV±dp/dtmax in aFGF treatment group were significantly increased compared with DCM group (P<0.01), while the LVEDP in aFGF treatment group was significantly lower than that in DCM group (P<0.01). The MVD in aFGF treatment group was significantly increased compared with DCM group (P<0.01), but the CVF and AI in aFGF treatment group were significantly lower than those in DCM group (P<0.01). CONCLUSION: Delivery of aFGF to diabetic myocardium by UTMD could improve the left ventricular function of DCM rats and may be a new feasible therapeutic method for DCM.     

Effects of cimetidine on platelet function and thrombosis

AIM:To observe the effects of cimetidine(Cim) on platelet function and thrombosis. METHODS:After incubated with Cimin vitro, rat platelets were activated with ADP or thrombin. The platelet aggregation, platelet malondialdehyde(MDA) formation, platelet intracellular free calcium( [Ca²⁺]_i), and thromboxane B₂ (TXB₂) were measured. The effects of Cim on electric-induced thrombosis in rat carotid artery were examined. RESULTS:Cim potentiated ADP induced platelet aggregation, increased the thrombin induced [Ca²⁺]_i and MDA formation, decreased TXB₂. Also, Cim shortened the duration of electric-stimulated occlution time in rat carotid artery. CONCLUSION:Cim increased platelet function and accelerated thrombosis.     

Prolactin induces interleukin-6 expression in PBMCs of rheumatoid arthritis patients

ATM: To investigate the correlation between serum prolactin (PRL) levels and disease activity in rheumatoid arthritis (RA) patients, and the regulatory role of PRL in interleukin-6 (IL-6) release from peripheral blood mononuclear cells (PBMCs), and to explore the MAPK-related mechanism of IL-6 release in PBMCs. METHODS: The clinicopathologic and hematologic parameters of 40 new-onset RA patients in the Department of Rheumatology of our hospital between March and September 2015 were collected. Chemilumineseent immunoassay (CLIA) was used to detect the serum PRL levels in the 40 RA patients and 20 healthy controls. The levels of IL-6 secretion by the PBMCs were evaluated using ELISA. Quantitative real-time PCR was employed to examine the mRNA expression of prolactin receptor (PRLR). MAPK pathway protein p-p38 levels were evaluated by Western blot. RESULTS: Serum PRL level in the RA patients was significantly higher than that in the healthy controls (P<0.01). Serum PRL level in active RA patients was significantly higher than that in inactive RA patients (P<0.01). Serum PRL level was positively correlated with DAS28, ESR and CRP (P<0.01). The expression of PRLR in the PBMCs was markedly increased in the RA patients than that in the healthy samples (P<0.01). Exposure of the PBMCs to PRL in the culture increased the release of IL-6, which was abolished by PRLR gene silencing or blocking the MAPK pathway.CONCLUSION: Serum PRL level is related to DAS28, ESR and CRP of RA patients and could be used as a predictor of disease activity. PRL/PRLR-p38 MAPK-IL-6 pathway may play a central role in the pathogenesis of RA.     

Role of GRP78 in alteration of myocardium induced by intestinal endotoxemia in cirrhotic rats

AIM: To explore the role of glucose-regulated protein 78 (GRP78) in the alteration of myocardium induced by intestinal endotoxemia in cirrhotic rats. METHODS: Fifty-one male Wistar rats were randomly divided into liver cirrhosis groups of 4-week, 6-week and 8-week, and normal control groups at corresponding time points. The cardiac functions of the 8-week rats were measured. Tumor necrosis factor α(TNF-α) and malondialdehyde(MDA) in myocardial tissues were detected. The number of myocardial cells and the collagen volume fraction (CVF) were determined with toluidine blue and van Giesan staining, respectively. The expression of GRP78 and hypoxia-inducible facotr 1α(HIF-1α) was analyzed by the method of immnunohistochemistry. RESULTS: Compared with normal control group at corresponding time point, left ventricular end-diastolic pressure(LVEDP) and ±LV dp/dt_max in 8-week group were significantly decreased (P<0.05). The levels of TNF-α, MDA and CVF, the protein expression of GRP78 and HIF-1α in the myocardial tissues were significantly increased in every model group (P<0.05), and the number of myocardial cells was gradually decreased (P<0.05). Elevated levels of endotoxin in plasma were positively correlated with the levels of alanine aminotransferase (ALT),homocysteine (Hcy) and TNF-α in plasma, the levels of TNF-α, MDA and CVF, and protein levels of GRP78 and HIF-1α in the myocardial tissues (P<0.05). Elevated protein expression of GRP78 in the myocardial tissues was positively correlated with the levels of ALT, Hcy in plasma and MDA, CVF, HIF-1α protein in the myocardial tissues (P<0.05). CONCLUSION: Intestinal endotoxemia induced by liver cirrhosis may directly or indirectly lead to endoplasmic reticulum stress and overexpression of GRP78. GRP78 may be a key molecule in the pathogenesis of myocardial remodeling and functional alteration induced by liver cirrhosis.     

Effects of different group B streptococci strains on platelet activation

AIM: To explore the ability of different group B streptococci (GBS) strains on inducing platelet activation. METHODS: Six strains of GBS, separated from the septic patients with thrombocytopenia, were used as the inducers. Light transmission aggregometry was used to measure platelet aggregation. Scanning electron microscopy (SEM) was performed to investigate the interaction of platelets with bacteria. The expression of platelet CD62P, Toll-like receptor 2 (TLR2) and TLR4 was determined by flow cytometry and Western blotting. Furthermore, the activity of platelet TLR2 (or TLR4) was blocked by anti-TLR2 (or anti-TLR4) monoclonal antibody, and the platelet aggregation induced by GBS was detected. RESULTS: Only 3 of 6 GBS strains isolated from the septic patients induced platelet aggregation and up-regulated the expression of CD62P and TLR2 in the platelets (P < 0.05), but not TLR4. Incubation with anti-TLR2 antibody, but not anti-TLR4 antibody, significantly blocked platelet aggregation induced by GBS.CONCLUSION: Some GBS strains from the patients are able to trigger platelet activation in vitro, and platelet TLR2 may play an important role in the interaction between GBS and platelets.     

Effect of ERK1/2 phosphorylation on the aggregation of the rat platelets in vitro

AIM: To study the influence of PD098059 on the rat platelet aggregation rate and the phosphorylation of ERK1/2 induced by the different agonists, and to observe the effects of phosphorylation of ERK1/2 on the platelet aggregation. METHODS: The maximal aggregation rate (MAR) was measured by nephelometry. The inhibitory rate of PD098059 and the appearing time of MAR were also observed. ERK1/2 phosphorylation was detected by Western blot. RESULTS: The phosphorylation of ERK1/2 was detected during aggregation induced by thrombin and ADP. PD098059 inhibited the MAR and phosphorylation of ERK1/2. Effects of PD098059 were different on the aggregation induced by thrombin and ADP. CONCLUSIONS: The phosphorylation of ERK1/2 is one of the cellular signal transduction mechanisms of platelets aggregation. Phosphorylation of ERK1/2 plays different roles during the platelet aggregation induced by thrombin and ADP.     

Role of juglone in regulating oxidative stress and apoptosis in ovarian cancer SKOV3 cells

AIM: To study the cytotoxicity of juglone on ovarian cancer SKOV3 cells. METHODS: The activity of SKOV3 cells was detected by MTT assay. The cells apoptosis was examined by flow cytometry. The level of reactive oxygen species (ROS) was measured by DCF-DA staining. The protein levels of cytochrome C (Cyt C) and activated caspase-3 were evaluated by Western blot. RESULTS: MTT assay showed that juglone significantly inhibited the growth of SKOV3 cells in dose- and time-dependent manners (P<0.05). The early apoptotic rate and late apoptotic rate of SKOV3 cells in 50 μmol/L juglone group at 24 and 48 h were higher than those in control group (P<0.01).Moreover, juglone induced ROS accumulation, and increased the protein levels of Cyt C and activated caspase-3.CONCLUSION: Juglone inhibits the cell growth and induces the apoptosis of SKOV3 cells by ROS accumulation.