Investigation of detection methods for aggregation function of platelets / 国际检验医学杂志

Objective To adopt 5 kinds of platelets aggregation function analyzer to explore and study the reliable method for detecting the platelets aggregation function and accurate parameters .Methods The platelets aggregation function in the control group and the single clopidogrel group was simultaneously detected by utilizing the ADP induced light turbidimetric platelet aggre‐gation analyzer (LTA) test ,flow cytometry for PAC‐1(receptor of Fg ,Ca2+ ,GPⅡb/Ⅲa forming complex) and CD62p(P selectin) activation percentage detection test ,Innova PL‐11 for platelets analysis test ,VerifyNow anti‐platelet therapy monitoring system and thrombelastogram(TEG) .Results (1)The 6‐parameter differences of ADP% ,PAC‐1 and CD62p receptor activation percentage ac‐tivated by ADP ,MAR% ,INHI% ,ADP induced MA value(mm) detected by TEG had statistical differences between the control group and the case group(P0 .05) .(2)ADP% was positively correlated with (100‐INHI)% ,MAR% ,ADP activated CD62p and PAC‐1 receptor activation percentage(r=0 .565 ,0 .939 ,0 .769 ,0 .583 ,P<0 .05 );while which had no correlation with ADP induced platelets aggregation value(% Agg) detected by TEG(r=0 .794 ,0 .715 ,0 .889) .Conclusion (1)Clopidogre has the anti‐platelets effect .(2)LTA is easily operating ,cheap and stable in detection results ,has high popularizing rate in hospitals and is the first option method for clinically monitoring the platelets aggregation function .

High levels of LDL-C combined with low levels of HDL-C further increase platelet activation in hypercholesterolemic patients

High levels of low-density lipoprotein cholesterol (LDL-C) enhance platelet activation, whereas high levels of high-density lipoprotein cholesterol (HDL-C) exert a cardioprotective effect. However, the effects on platelet activation of high levels of LDL-C combined with low levels of HDL-C (HLC) have not yet been reported. We aimed to evaluate the platelet activation marker of HLC patients and investigate the antiplatelet effect of atorvastatin on this population. Forty-eight patients with high levels of LDL-C were enrolled. Among these, 23 had HLC and the other 25 had high levels of LDL-C combined with normal levels of HDL-C (HNC). A total of 35 normocholesterolemic (NOMC) volunteers were included as controls. Whole blood flow cytometry and platelet aggregation measurements were performed on all participants to detect the following platelet activation markers: CD62p (P-selectin), PAC-1 (GPIIb/IIIa), and maximal platelet aggregation (MPAG). A daily dose of 20 mg atorvastatin was administered to patients with high levels of LDL-C, and the above assessments were obtained at baseline and after 1 and 2 months of treatment. The expression of platelets CD62p and PAC-1 was increased in HNC patients compared to NOMC volunteers (P<0.01 and P<0.05). Furthermore, the surface expression of platelets CD62p and PAC-1 was greater among HLC patients than among HNC patients (P<0.01 and P<0.05). Although the expression of CD62p and PAC-1 decreased significantly after atorvastatin treatment, it remained higher in the HLC group than in the HNC group (P<0.05 and P=0.116). The reduction of HDL-C further increased platelet activation in patients with high levels of LDL-C. Platelet activation remained higher among HLC patients regardless of atorvastatin treatment.

CD62pUsing Light Transmission Aggregometry and Flow Cytometry to Examine the Effect of Clopidogrel / 现代检验医学杂志

Objective To detect clopidogrel effect with light transmission aggregometry (LTA)and flow cytometry (FC). Methods ①Venous blood samples were taken from 71 inpatient with acute corotary syndrome (ACS)in PLA General Hos-pital,including unstable anqina,ST segment elevation myocardial infarction and non ST segment elevation myocardial infarc-tion (46 males,25 females)by random number table from June 2011 to March 2012,whose average age was 69(57~92).②All of them were served 160 mg aspirin and 300 mg clopidogrel after they were in hospital in the beginning,and then served with 75 mg/d clopidogrel for 6 months.On some day,firstly,they were required withdrawing drug for 10 days,and then ve-nous blood samples were separately taken from them before their served-clopidogrel again and their severd-clopidogrel 2 hours later.③The samples were assayed with LTA and FC simultaneously and the platelet aggregation rates before served-clopidogrel (ADPLTA-before serving ),platelet aggregation rates after served-clopidogrel (ADPLTA-after serving ),inhibition rates (ADPLTA-INDU ),PAC-1 activity percentage before served-clopidogrel (PAC-1 before serving ),PAC-1 activity percentage after served-clopidogrel (PAC-1 after serving ),inhibition rates (PAC-1 INHI ),CD62p activity percentage before served-clopidogrel (CD62pbefore serving ),CD62pactivity percentage after served-clopidogrel (CD62pafter serving ),inhibition rates (CD62pINHI )weregotten.All volunteers were signed informed consents and the experiment was approved by the hospital ethics committee.Re-sults ①The paired samples t-test was (t=-2.082,P =0.041)between ADPLTA-before serving (0%~97%)and ADPLTA-after serving (12%~97%),the paired samples t-test was (t = 3.663,P < 0.01)between PAC-1 before serving (15.1% ~ 78.9%)and PAC-1 after serving (14.5% ~ 78.3%);the paired samples t-test was (t = 2.082 and P = 0.041)between CD62pbefore serving (1.5% ~80.8%)and CD62pafter serving (1.4%~41.4%).②The pearson coeffcient correlation results were:ADPLTA-INDU (0%~28.2%) and PAC-1 INHI (0.6%~ 9.1%)(r = 0.297,P = 0.012);ADPLTA-INDU (0% ~ 28.2%)and CD62pINHI (0.1% ~ 48.5%)(r =0.220,P =0.065);PAC-1 INHI (0.6%~9.1%)and CD62pINHI (0.1%~48.5%)(r=0.736,P <0.001).Conclusion Because the correlation was bad between the inhibition rates of clopidogrel detected by FC and them by LTA,FC didn’t apply to clin-ical routine examination of the platelet aggregation.But it could be used to scientific researchs and auxiliary confirmation of routine examination results.

Detection of the Platelet Activation Markers CD62p and PAC-1 in Patients with Hypertension and Coronary Heart Disease / 现代检验医学杂志

Objective To investigate the clinical significance of platelet activation markers CD62p and PAC-1 in patients with hypertension and coronary heart diease.Methods To select patients who were in the Second Affiliated Hospital of Dalian Medical University between January 2012 and September 2012,42 patients with hypertension,46 patients with coronary heart disease.Flow cytometry was used to test the positive percentage of the platelet surface activation markers CD62p and PAC-1 .To observe the difference among hypertension group,coronary heart disease group and health control group;hyper-tension and coronary heart disease group.Results The positive percentage of platelet activation markers CD62p and PAC-1 in hypertension (36.36±9.62,7.18±8.20)%,coronary heart disease group (42.74±14.60,8.81±12.53)% showed sta-tistically significant differences (t=4.150~5.853,P<0.01)compared with healthy control group (26.82±9.13 ,1.09± 1.05)%.The positive percentage of CD62p in coronary heart disease group (42.74±14.60)% was also higher than that of hypertension (36.36±9.62)%,and the difference between them was of statistical significance (t=2.444,P<0.05).Con-clusion The hypertensive patients and coronary heart disease were in pre-thrombotic state,the activation of platelet increase significantly.Molecular markers should be measured in hypertensive patients and coronary heart disease for prevention and treatment of thrombotic disease complication.

Expression and clinical significance of platelet activating factor (PAC-1), CD629P and thrombus precursor protein (TpP) in severe sepsis / 中华急诊医学杂志

Objective To investigate the expression and clinical significance of platelet activating factor [PAC]-1, CD62P and TPP hi severe sepsis. Method Patients with severe sepsis who were admitted into the EICU of Subei People's Hospital from April 2007 to March 2008 were included. Patients with severe sepsis (Group Ⅲ)were treated according to the treatment guidelines for severe sepsis, and were divided, according to their clinical records, into those who survived and those who died within 28 days of admission. Patients admitted during the same period with symptoms of infection but without severe sepsis were included as the General Infected Group (Group Ⅱ). A Control Group (Group Ⅰ) comprised patients who visited the hospital over the same period for physical examination or the healthy volunteers. The group members were all included randomly, and the gender and sex of patients in all three groups were similar. Patients with acute brain infarction, acute coronary syndrome,serious diabetes, hyperlipidemia, malignant tumor, leukemia, primary liver, renal and hematopoietic system dis-eases,long-term bedridden patients, pregnant women, and patients taking hormone treatment or hranunosuppres-sants were excluded from the study. Morning venous blood was collected and ELISA and Flow Cytometry performed on the fwst day of admission for Groups Ⅰ- and Ⅱ, and on the first, third and fifth day after admission for Group Ⅲ, to determine the TpP,PAC-1 and CD62P respectively; and the Marshall score was determined. Data were ana-lyzed by SPSS 12.0 software. For continuous variables, comparisons among groups were analyzed by ANOVA.Levene's and LSD test were applied to assess homogeneity. Bivariate test is applied to Correlation Analysis. P<0.05 was regarded as a statistically significant difference. Results There were a total of 20 patients each in GroupⅠ-and GroupⅡ, and 30 in Group Ⅲ; of these, 19 were classed as survivors and 11 died during the 28-day peri-od. On the first day of admission, there were no significant differences in PAC-1, CD62P or TpP expression between Groups Ⅰ- and Ⅱ(P>0.05); however, Group Ⅲ was significantly different compared with both Group Ⅰ and Group Ⅱ (both:P<0.05). The expression of PAC-1, CD62P and TpP tended to decline in the survivor group,and became normal with the treatment process, while the expression of PAC-1 ,CD62P and TpP in the patients who died remained high, and even increased significantly over time. On the first day, the expression of CD62P and TpP in the patients who survived and in those who died was not significantly different (P>0.05); on the third day,however, a significant difference appeared with values of (2.89±1.48) % vs. (5.04±2.57) % (P<0.01) for CD62P, and (5.24±2.22) mg/L vs. (9.20±1.93) mg/L (P<0.01) for TpP. The expression of PAC-1 was significantly different between the two subgroups on the first day, with values of (3.15±0.42)% vs. (5.30±.48)% (P<0.01). The Marshall score of the two groups showed similar changes. Correlation analysis showed that PAC-1, CD62P and TpP were significantly correlated with the Marshall score. Conclusions Platelet activation and microthrombosis existing in the early stage of severe sepsis work together in the early hypercoagulable state.They both play important roles in disease development and progression. The dynamic detection of CD62P and TpP is beneficial to the diagnosis and prognosis of severe sepsis.PAC-1 appears to hold a risk stratification effect, as pa-tients with high expression of PAC-1 in the early stage show poor prognosis. Therefore, PAC-1 could be used as a marker of severe sepsis and poor prognsis.

The relationship between the activity of platelet and the chronic cor pulmonale / 中国综合临床

Objective To investigate the clinical significance of piatelet activity in patients with chronic cor Dulmonale.Methods PAC-1 and CD62p was measured with flow cytometry in whole blood samples from 40 patients and 30 normal controls.The pulmonary arterial pressure was detected through Doppler echocardiography.The arterial partial pressure of oxygen and the carbon dioxide were also analyzed.Results PAC-1 and CD62p increased significantiy (P<0.01).Conclusion Platelet activity is positively related to pulmonary artrial systolic pressure,CO2 partial pressure,and negatively related to O2 partial pressure.

Regulation of Prolactin Gene Expression by Hypothalamic Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) in the Female Rat Pituitary Gland / 대한내분비학회지

BACKGROUND: Pituitary adenylate cyclase-activating polypeptide (PACAP) plays the role of a hypophysiotropic factor, which regulates the synthesis and secretion of pituitary hormones through the hypothalamo-hypophysial portal system. No clear evidence has yet been reported regarding the regulation of prolactin (PRL) by PACAP. In the present study, we tested a hypothesis that PACAP regulates the synthetic machinery of PRL during the estrus cycle and pubertal process using intracerebroventricular (i.c.v.) injection of an antisense oligodeoxynucleotide (ODN) against type I PACAP receptor (PAC1). METHODS: An RNase protection assay (RPA) was used to determine the pattern of hypothalamic PACAP and PAC1 mRNA expressions during the estrus cycle. Antisense PAC1 ODN was administered via i.c.v. injection to the female rats in normal estrus cycle of pubertal process. Northern blot analysis was used to determine the mRNA ievel of PRL in the pituitary gland. RESULTS: 1) PACAP mRNA in the medial basal hypothalamus was significantly increased at the diestrus I, while PAC1 mRNA showed no significant change. 2) PRL mRNA level of pituitary was increased by an injection of antisense PAC1 ODN at the proestrus and estrus stages. 3) PRL mRNA level of pituitary was significantly decreased by antisense PAC1 ODN injection at stage of prepuberty and initiate puberty, while its level was increased at stage of puberty. CONCLUSION: These data suggest that PACAP suppresses PRL mRNA synthesis through the PAC1 signaling pathway in the certain estrus cycle environments. It may be also involved in the regulation of pituitary PRL gene expression during the pubertal process

Effects of Cytokine on Platelet Activation / 대한수혈학회지

BACKGROUND: It has been reported that the cytokine released from the white blood cells (WBC) in the stored platelet concentrates may induce febrile nonhemolytic transfusion reactions. Few studies, however, have examined the effects of cytokine on platelet activation. METHODS: The platelets from healthy donors were incubated with interleukin (IL) at the final concentration of IL-8 0.5 ng/mL, 3 ng/mL, 5 ng/mL, 10 ng/mL, 1000 ng/mL, and IL-6 0.1 ng/mL, 0.5 ng/mL, 1 ng/mL, 5 ng/mL, respectively. The activated platelets were stained with monoclonal antibodies of PAC-1 and CD62, and were analyzed by flow cytometry. RESULTS: The ranges of mean percent of PAC-1 and CD62 expressions on platelets were 61.1-71.4% and 9.0-12.5% by incubating with IL-8, and 69.0-73.4% and 13.0-13.9% with IL-6, respectively. There were no significant differences of PAC-1 and CD62 expressions on platelets among the various concentrations of IL-8 nor IL-6. In 0.5 ng/mL concentration, the mean percentage of CD62 expressions on platelets by incubating with IL-6 (13.8%) were greater than that of IL-8 (9.0%). CONCLUSION: During the period of the storage of platelet concentrates, the IL-6 and IL-8 secreted by WBC can activate platelets and thus the suppression of the cytokine secretion may be important for increasing post-transfusional survival of platelets.