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Objective To establish an isotope dilution liquid chromatography tandem mass spectrometry method (ID-LC-MS) for quantification of serum apolipoprotein E and phenotyping. Methods Method establishment. Samples underwent denaturing, alkylation and trypsin digestion with addition of internal standards as isotope labelling arginine. SB-C18 column was used for the liquid chromatographic separation and mass spectrometry positive ion mode and multiple reaction monitoring were employed for quantification and phenotyping. Precision, accuracy and linearity were investigated for method evaluation. 40 serum samples from Shanghai Dongfang Hospital during Oct. to Dec., 2018 were used for method comparison between ID-LC-MS and immunoassay. Deming regression and Bland-Altman were used for method comparison analysis and SPSS 24 for linearity. Results Target peptides reached their releasing maximum within 4 hours and SE did at 3 hours. 3 phenotyping of ApoE were observed, such as E3/E3, E2/E3 and E3 / E4. The imprecision of IQC was 5.2 ％ . The relative bias for low and high levels of accuracy-based samples was 7.6 ％ and 3.6 ％, respectively. Deming regression showed the intercept with 95 ％ confidence interval (CI) was 6.44-11.44 (P<0.05 and the 95％ confidence interval for the slopewas 0.77-0.89 (P<0.05). The coefficient was r=0.97. The mean difference was - 2.95 mg / L with 95 ％ CI-4.26--1.65 mg/L. The linearity covered from 16.9 to 58.5 mg/L. Conclusion ID-LC-MS can be used to quantify serum apolipoprotein E and simultaneously detect its phenotyping.
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While several lines of evidence prove that elevated concentrations of low-density lipoproteins (LDL) usually contribute to the development of atherosclerosis and its clinical consequences, high-density lipoproteins (HDL) are widely believed to exert atheroprotective effects. Hence, HDL cholesterol (HDL-C) is in general still considered as good cholesterol. Recent researches, however, suggest that this might not always be the case and that a fundamental reassessment of the clinical significance of HDL-C is warranted. The main function of HDL is to transfer the cholesterol outside the liver into the liver for catabolism.The liver′s cholesterol metabolism and other biological effects are dependent on the number of HDL particles and its proteins and lipid contents. These functions are difficult to be described simply with the HDL-C concentration. If the components of HDL particles change, they may have adverse effects on the blood vessels. Thus, high concentrations of HDL-C in plasma are not always protective factors, and some clinical trials improving HDL-C concentrations have failed to confirm a protective effect. To explore the complex relationship and pathological mechanism between HDL and atherosclerotic diseases, it is instructive for clinical application of the HDL measurement.
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Objective To investigate the clinical value of serum lipoprotein ( a ) concentration in evaluation of plaques characteristics for patients with coronary atherosclerotic heart diseases ( CAHD ) . Methods Using case-control method, Patients with suspicious CAHD, received coronary computed tomography angiography in the Shanghai East Hospital during October 2013 to June 2015 were enrolled.According to the results of coronary artery CTA, the patients were divided into two groups : the CAHD group (352 cases) and control group(438 cases) , the particle concentrations and mass concentration of lipoprotein(a), triglyceride, total cholesterol, HDL-C, LDL-C, glucose, HBA1c and hs-CRP and other tests were measured, the patients of CAHD group were divided into three subgroups by characteristics of coronary artery plaques including soft plaque (176 cases), calcified plaque (90 cases) and mixed plaque (86 cases), analysis were made with all these data.Using T test or variance analysis to compare the means between or among groups, the risk for CAHD was analyzed by logistic regression, the relationship between LP (a) -P and LP( a) -M were explored by linearly egression analysis, Conformance test were analyzed using kappa test.Results Compared with control group, the mean results of the CAHD group are significantly higher than that of control group, including LP (a) -P 18.5(8.3 -43.0))nmol/L vs.13.6 (7.6-32.4)nmol/L( t =-2.110), LP(a)-M 183(71 -361)mg/L vs.126(67 -293)mg/L(t =-2.063), age (62 ±9)years vs.(52 ±9)years(t=-7.691), hs-CRP 0.86(0.44-1.97) )mg/L vs.0.70(0.38-1.64)mg/L(t=-2.236), glucose (6.1 ±2.29 )mmol/L vs.(5.36 ±1.32 )mmol/L(t=-4.914), BA1c (6.13 ±0.98) % vs.(5.81 ±0.58) %(t=-4.842), APO(B) (1.09 ±0.33) g/L vs.(1.03 ±0.29) g/L( t=-2.407), all of the P values <0.05;The relative risk(RR)of age, glucose, LP( a)-P and LP ( a)-M are 1.067, 2.377, 1.384 and 1.342 respectively; Among the three types of plaques groups,the mean differences of age, TC, HDL-C, LDL-C and LP ( a)-P are statistically significant ( F=6.276,3.060,3.127,4.723,2.878;all of the P<0.05);The median of LP ( a)-P in the soft plaque group 20.3(8.3-48.2)nmol/L is higher than that of the mixed plaque group 15.7(7.3-26.0)nmol/L(P<0.05 ) and calcified plaque group 15.6 ( 8.1 -23.1 ) nmol/L ( P <0.05 ).The linearly regression equation of LP ( a) -M and LP( a)-P is Y=6.646X, r=0.939; Consistency test indicate the two methods are not consistent when used for grouping ( Kappa value is 0.557 ).Conclusions Serum concentration of lipoprotein(a) is an independent risk factor of CAHD, and the particle concentration of LP(a) is closely related to the characteristics of the plaques, especially to the soft plaque.
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Objective To investigate the clinical application value of serum lipoprotein associated phospholipase A2(Lp‐PLA2) in coronary atherosclerotic heart diseases(CAD) .Methods Using the case‐control study ,790 patients with coronary computed tomography angiography (CTA) in our hospital from October 2013 to June 2015 were selected and divided into the CAD group (352 cases) and control group (438 cases) according to the results of coronary artery CTA .According to the number of coronary artery lesion vessels the CAD group was re‐divided into three subgroups :single branch coronary artery lesion (118 cases) ,double branch coronary arterial lesions(n=107) and multiple branch coronary arterial lesions(132 cases) .The levels of Lp‐PLA2 ,hs‐CRP , TG ,TC ,HDL‐C ,LDL‐C ,glucose ,HbA1c and other indexes were measured and comprehensively analyzed .The t test or variance a‐nalysis was used to compare the means between or among groups .The correlation of different indicators was analyzed with the Pearson linear correlation analysis .Results Compared with the control group ,the CAD group was significantly higher than the con‐trols in the levels of Lp‐PLA2 ,hs‐CRP ,age ,GLU ,HbA1c and ApoB ,the differences were statistically significant(P0 .05) .Conclusion Serum Lp‐PLA2 level increase is a risk factor of CAD and could be used to assess coronary arterial atherosclerosis and number of coronary arterial lesions .
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Objective To explore the serum level of Glucagon like peptide-1 in late-onset Alzheimer′s patients and its clinical significance.Methods Case control study.Collecting cerebral vascular disease fifty-five cases, diagnosed with late-onset Alzheimer′s disease sixty-one cases, type 2 diabetes mellitus fifty-one cases , type 2 diabetic patients combined with late-onset Alzheimer′s disease thirty-seven patients from the Shanghai East Hospital and partly Pudong area elderdly hospital during October 2013 to March 2014, and forty healthy persons as normal control from physical examination center of Shanghai East Hospital during September 2013 to February 2014.Measuring the concentrations of GLP-1,β-amyloid, Tau protein and other routinely used clinical tests in the serum of patients from the normal controls , cerebrovascular disease , late-onset Alzheimer′s disease, type 2 diabetes and type 2 diabetes mellitus combined with late-onset Alzheimer′s disease by ELISA method developed in our laboratory.The blood samples were also collected at three fixed time including fasting time ,1 hour after taking glucose , 2 hour after taking glucose, the concentrations of GLP-1 were determined in the LOAD group , T2DM group and the T2DM combined with LOAD group and normal control group.The concentrations of serum GLP-1 among groups were compared with single factor analysis of variance , and the concentrations of serum GLP-1 between the two groups were compared using LSD-t test.Analysing the correlation between GLP-1 and other indicators with Pearson analysis.Results The fasting GLP-1 levels of LOAD group were ( 123.4 ±20.8 ) nmol/L, and they were highest between the normal control group (78.6 ±6.0) nmol/L and the cerebral blood vessel disease group(89.0 ±8.7)nmol/L (F values were 3.46 and 1.98, P0.05).Deficient secretion of GLP-1 after taking glucose 1 hour in most of the patients of T2DM combined with LOAD group (99.1 ±14.2) nmol/L, LOAD group(73.9 ±6.6 ) nmol/L and T2DM group (96.3 ±7.0 ) nmol/L could be concluded .The GLP-1 levels of T2DM combined with LOAD group after taking sugar 2 hour were (115.4 ±18.6)nmol/L ,and were higher than that of normal levels (63.3 ±6.2) nmol/L after taking sugar 2 hour(t=4.49,P0.05).Pearson correlation analysis showed that the relationship of the levels of GLP-1 with Aβ( 1-42 ) and the levels of GLP-1 after taking glucose 1 h and 2 h were positively relative, and its coefficients of correlation were 0.401,0.436,0.722.Conclusions LOAD and T2MD are similar, and they have GLP-1 secretion shortage phenomenon after taking glucose , so monitoring dynamic change of GLP-1 after taking glucose may contribute to the auxiliary diagnosis of LOAD.
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Objective To compare the difference of kidney function evaluated by using 3 different estimated glomerular filtration rate(GFR) equations in populations .Methods Retrospectively analyzed 65 856 patients who measured serum creatinine and Cysta‐tin C at the same time ,and come from the outpatients or inpatients of the hospital .The estimated GFR (eGFR) were calculated through 3 equations ,then compared the eGFR in the population and among different groups according to different kidney functions , and then grouped the people enrolled in the study again according to the eGFR calculated by using the 3 different equations and compared the differences among groups .Results Compared with the eGFR calculated by using Creatinine equation ,the correlation coefficients of the eGFRs calculated by using the other two equations were 0 .81 and 0 .90 ,respectively ,both P<0 .05 ;The differ‐ence between the means of eGFR were 6 .19 and 1 .79 mL/(min × 1 .73 m2 ) respectively with obvious significance (P<0 .01) ,in consistency analysis .There were obvious overestimation of kidney function when using Creatinine equation to calculate eGFR .Con‐clusion There is consistence and obvious difference by using the 3 CKD‐EPI′s eGFR equations .Physicians should choose suitable equations to evaluate kidney function in different populations .
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To prove the influence of protein isoaspartyl-methyltransferase ( PIMT ) on the cell apoptosis of fibroblast-like synoviocytes of RA.Methods: The expression vector of PIMT was constructed and transfected in to the RA-FLS, the impact of PIMT on the cell apoptosis of RA-FLS was observed by overexpressing the vector of PIMT.Results:The mRNA and protein level of PIMT in RA-FLS was increased after transfected the vector of PIMT into RA-FLS;compared with the normal cultured RA-FLS and the RA-FLS transfected with the empty vectors ,the cell apoptosis level was also increased.Conclusion:The decreased expression level of PIMT in RA-FLS is an important reason for reduce apoptosis of RA-FLS,and PIMT can affect the imbalance of proliferation/ap-optosis in the RA-FLS.
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Lipoprotein (a) [Lp (a)] is an LDL-like molecule consisting of an apolipoprotein (B) [Apo (B)] particle attached by a disulphide bridge to apolipoprotein (a) [Apo (a)].Because of the LPA gene polymorphisms,there exists Lp (a) molecular heterogeneity in the human population.Recently,several observations have pointed out that elevated serum Lp (a) levels may be an independent risk factor of cardiovascular disease and an predictive indicator of cardiovascular disease.Recent findings suggest that Lp (a)-lowering therapy might be beneficial in patients with high Lp(a) levels.Currently,there are some problems in Lp(a) assay,including clinically diverse methods,the Lp (a) molecular heterogeneity between reference materials and test samples.Standardization of the Lp (a) reference reagents and measurement methods may be helpful for the wide application of Lp(a) in clinical medicine and be helpful for assuring the role of Lp(a) as an independent risk factor and predictive indicator of cardiovascular diseases.
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Objective To investigate the clinical value of pregnancy-associated plasma protein A (PAPP-A) in patients with acute coronary syndrome (ACS).Methods The study subjects comprised of 249 patients with ACS [ 50 patients with ST elevation acute myocardial infarction( STEMI),43 patients with non-ST elevation acute myocardial infarction (NSTEMI), 156 patients with unstable angina pectoris (UAP) ] from July 2008 to December 2009 at Shanghai East Hospital affiliated to Tongji University.The patients were divided into single-vessel lesions group,double-vcssel lesions group and three-vessel lesions group according to the coronary artery stenosis.A group of 205 healthy subjects admitted for health physical examination were used as conuols.Blood samples were collected from ACS patients and control subjects.Serum PAPP-A,creatine kinaseisoenzyme MB (CK-MB),high-sensitivity troponin-T (hs-TnT) were measured by clectrochemiluminesence and high-sensitivity C-reactive protein (hs-CRP) was measured by immune scatter turbidity method.Analysis of variance ANOVA and Kruskal-Wallis H test were used for statistical analysis.Results Serum PAPP-A in the STEMI,NSTEMI,UAP and normal control group were 10.45(5.54 - 16.08),6.56(4.68 - 9.55),5.70(4.12 - 8.50),5.23 (4.00 - 5.88) mIU/L,respectively,and the difference was statistically significant (H =43.94,P ＜ 0.01 ).Pairwise comparison showed that PAPP-A in STEMI,NSTEMI,UAP were significantly higher than the healthy control group and differences were statistically significant ( t =6.80,2.46,1.72,P ＜ 0.05 ).The PAPP-A had a sensitivity of 52.2％ and specificity of 80％ in diagnosis of ACS.The positive rate of PAPP-A was 44.5％ (69/155) in patients with negative hs-TnT.The serum levels of PAPP-A in the three-vessel lesions group [7.72(5.03 -12.46) mIU/L] was higher than that in the single,double groups [ 5.35 ( 4.14 - 8.59 ),6.05 (4.42 -9.58 ) mIU/L ],and the difference had statistical significance( t =2.00,1.57,P ＜ 0.05 ).There was obvious correlation between the level of serum PAPP-A and the level of hs-CRP ( r =0.524,P ＜ 0.05 ),and there was weak correlation between the PAPPA and CK-MB or hs-TnT (r=0.326,0.343,P＜0.05).Conclusions The results shows that PAPP-A is elevated in patients with ACS.It may be used as a serum biomarker for vulnerable plaques in patients with ACS and has a clinical value for ACS diagnosis especially in patients with negative hs-TnT.
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Objective To study the effects of citrullinated vimentin (cVim) on the maturation and immunologic function of dendritic cells (DCs) from rheumatoid arthritis (RA) peripheral blood.Methods In the present study,mononuclear cells were isolated from the peripheral blood of patients with RA and cultivated in media containing GM-CSF and IL-4 to generate immature DCs (imDCs).The imDCs generated were stimulated with citrullinated vimentin and vimentin.LPS was used as the positive control and PBS was used as the negative control.The expression of surface molecules on the DCs,such as CD14,CD80,CD83,CD86,MHC Ⅰ and MHC Ⅱ were analyzed with FACS.The capability of the stimulatory activity of the DCs on allogeneic T cells in mixed reaction was tested by MTS.t-test was used for statistical analysis.Results Compared to untreated DCs,DCs treated with LPS increased the expression levels of MHC Ⅱ,CD80,CD83 and CD86 (1.07±0.14,1.25±0.13,1.90±1.08,2.44±0.65,P＜0.05),while cVim increased the expression levels of MHC Ⅱ ( 1.18±0.09,P＜0.05) and CD83 ( 1.97±0.99,P＜0.01 ),and Vim decreased the expression levels of CD80 (0.82±0.18,P＜0.01 ).It was demonstrated that the expression levels of MHC Ⅱ on DCs pulsed with cVim were significantly higher than that of the DCs with LPS,but the expression levels of CD80 and CD86 were not significantly different.The expression levels of MHC Ⅱ and CD83 on DCs pulsed with cVim were significantly higher than that of the DCs with Vim.The mixed lymphocyte reaction showed that the DCs induced by LPS and cVim trigerred the proli-feration of allogenic T cells obviously.Conclusion This result suggests that cVim could promote the phenotypic maturation of DCs and increase the expression of costimulatory molecules.