The present study observed the regulatory effect of total flavonoids of Ziziphora clinopodioides on autophagy and the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/mammalian target of rapamycin(mTOR) signaling pathways in ApoE~(-/-) mice and explored the mechanism of total flavonoids of Z. clinopodioides against atherosclerosis(AS). ApoE~(-/-) mice were fed on a high-fat diet for eight weeks to induce an AS model. The model mice were randomly divided into a model group, a positive control group, and low-, medium-and high-dose groups of total flavonoids of Z. clinopodioides, while C57BL/6J mice fed on a common diet were assigned to the blank group. The serum and aorta samples were collected after intragastric administration for 12 weeks, and the serum levels of total cholesterol(TC), triglyceride(TG), low density lipoprotein-cholesterol(LDL-C), and high density lipoprotein-cholesterol(HDL-C) were detected by an automatic biochemical analyzer. The serum expression levels of intercellular adhesion molecule-1(ICAM-1), vascular cell adhesion molecule-1(VCAM-1), matrix metalloproteinase-2(MMP-2), and matrix metalloprotei-nase-9(MMP-9) were detected by enzyme-linked immunosorbent assay(ELISA). Oil red O staining was used to observe the aortic plaque area in mice. Hematoxylin-eosin(HE) staining was used to observe the aortic plaque and pathological changes in mice. The expression of P62 and LC3 in the aorta was detected by the immunofluorescence method. The protein expression of LC3Ⅱ/Ⅰ, Beclin-1, P62, p-PI3K, p-Akt, and p-mTOR in the aorta of mice was detected by Western blot. The results showed that compared with the blank group, the serum levels of TC, TG, LDL-C, ICAM-1, VCAM-1, MMP-2 and MMP-9 in the model group were significantly increased(P<0.01 or P<0.05), the content of HDL-C was decreased(P<0.05), intra-aortic plaque area was enlarged(P<0.01), the expression of LC3 in the aorta was significantly down-regulated, P62 expression was up-regulated(P<0.01 or P<0.05), the expressions of LC3Ⅱ/Ⅰ and Beclin-1 in the aortic lysate were significantly down-regulated, and the expressions of p-PI3K, p-Akt, p-mTOR and P62 were significantly increased(P<0.01). The medium-and high-dose groups of total flavonoids of Z. clinopodioides could reduce the serum levels of TC, TG, LDL-C, ICAM-1, VCAM-1, MMP-2, and MMP-9 in AS model mice(P<0.01 or P<0.05), and increase the content of HDL-C(P<0.01 or P<0.05). The aortic plaque area of mice after middle and high doses of total flavonoids of Z. clinopodioides was significantly reduced(P<0.01), the content of foam cells decrease, and the narrowing of the lumen decreased. The total flavonoids of Z. clinopodioides significantly increased the expression of LC3 in the aorta and the expression of LC3Ⅱ/Ⅰ and Beclin-1 in the lysate, and decreased the expression of P62 in the aorta and the expression of p-PI3K, p-Akt, p-mTOR and P62 in the lysate(P<0.01 or P<0.05). The results showed that the total flavonoids of Z. clinopodioides could improve the content of blood lipids and inflammatory factors, and reduce the generation of foam cells and plaques in aortic tissue, and the mechanism may be related to the regulation of PI3K/Akt/mTOR signaling pathway.
Animals ; Mice ; Apolipoproteins E ; Atherosclerosis/genetics* ; Beclin-1 ; Cholesterol, LDL ; Intercellular Adhesion Molecule-1 ; Matrix Metalloproteinase 2/genetics* ; Matrix Metalloproteinase 9/genetics* ; Mice, Inbred C57BL ; Phosphatidylinositol 3-Kinases/metabolism* ; Plaque, Atherosclerotic ; Proto-Oncogene Proteins c-akt/metabolism* ; TOR Serine-Threonine Kinases/genetics* ; Vascular Cell Adhesion Molecule-1/genetics*

PURPOSE: The present study aimed to investigate the value of apolipoproteins, including ApoA-1, ApoC-III, and ApoE, in patients with small cell lung cancer (SCLC) as potential biomarkers for diagnosis, prognosis, and cancer progression. MATERIALS AND METHODS: Lung samples were collected from 89 patients with SCLC. Nineteen lung samples from non-small cell lung cancer (NSCLC) patients and 12 normal lung tissues were used as controls. Expression profiles of ApoA-1, ApoC-III, and ApoE in different samples were examined using immunohistochemical methods, and the expression levels were correlated with cancer types, treatment, and outcomes using chi-square and Mann-Whitney tests. RESULTS: Expression of ApoA-1 and ApoC-III in SCLC was significantly different, compared with that in NSCLC and normal lung tissues, and was correlated with recurrence of SCLC. Patients undergoing neoadjuvant chemotherapy before surgery showed significantly reduced expression of ApoA-1 and increased expression of ApoC-III and ApoE. Nevertheless, the expression levels of ApoA-1, ApoC-III, and ApoE were not correlated with SCLC staging. CONCLUSION: ApoA-1 and ApoC-III may be used as differentiating and predictive markers for SCLC. ApoA-1, ApoC-III, and ApoE may be used to monitor the efficacy of chemotherapy.
Adult ; Aged ; Apolipoprotein A-I/*genetics ; Apolipoprotein C-III/*genetics ; Apolipoproteins E/*genetics ; Biomarkers/analysis ; Case-Control Studies ; Female ; Gene Expression Regulation ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Neoplasm Recurrence, Local ; Predictive Value of Tests ; Prognosis ; RNA, Messenger/*genetics ; Small Cell Lung Carcinoma/*diagnosis/genetics

OBJECTIVEWe aimed to investigate the cumulative effect of high CRP level and apolipoprotein B-to-apolipoprotein A-1 (ApoB/ApoA-1) ratio on the incidence of ischemic stroke (IS) or coronary heart disease (CHD) in a Mongolian population in China.

METHODSFrom June 2003 to July 2012, 2589 Mongolian participants were followed up for IS and CHD events based on baseline investigation. All the participants were divided into four subgroups according to C-reactive protein (CRP) level and ApoB/ApoA-1 ratio. Cox proportional hazard models were used to estimate the hazard ratios (HRs) and 95% confidence intervals (CIs) for the IS and CHD events in all the subgroups.

RESULTSThe HRs (95% CI) for IS and CHD were 1.33 (0.84-2.12), 1.14 (0.69-1.88), and 1.91 (1.17-3.11) in the 'low CRP level with high ApoB/ApoA-1', 'high CRP level with low ApoB/ApoA-1', and 'high CRP level with high ApoB/ApoA-1' subgroups, respectively, in comparison with the 'low CRP level with low ApoB/ApoA-1' subgroup. The risks of IS and CHD events was highest in the 'high CRP level with high ApoB/ApoA-1' subgroup, with statistical significance.

CONCLUSIONHigh CRP level with high ApoB/ApoA-1 ratio was associated with the highest risks of IS and CHD in the Mongolian population. This study suggests that the combination of high CRP and ApoB/ApoA-1 ratio may improve the assessment of future risk of developing IS and CHD in the general population.

Adult ; Apolipoproteins A ; classification ; genetics ; metabolism ; Apolipoproteins B ; genetics ; metabolism ; C-Reactive Protein ; genetics ; metabolism ; Cohort Studies ; Coronary Disease ; epidemiology ; etiology ; Gene Expression Regulation ; Humans ; Mongolia ; epidemiology ; Prospective Studies ; Risk Factors ; Stroke ; epidemiology ; etiology ; Young Adult

We evaluated the association of the APOE polymorphism with serum C-reactive protein levels and white blood cell count in two large population-based studies in Korean. The datasets included the Dong-gu study (n = 8,893) and the Namwon Study (n = 10,032). APOE genotypes were identified by polymerase chain reaction-restriction fragment length polymorphism. Multivariable linear regression analysis was performed to evaluate the relationship of APOE genotypes with C-reactive protein levels and white blood cell count with adjustments for age, sex, body mass index, smoking, diabetes, hypertension, and serum lipids. In the multivariate model, carriers of E3E4 or E4E4 genotype had significantly lower C-reactive protein levels compared with carriers of E3E3 genotype group (0.50 mg/L vs. 0.67 mg/L; 0.37 mg/L vs. 0.67 mg/L, respectively, for the Dong-gu Study and 0.47 mg/L vs. 0.66 mg/L; 0.45 mg/L vs. 0.66 mg/L, respectively, for the Namwon Study). However, there was no difference in white blood cell count among APOE genotypes. We found that the APOE E4 allele is associated with lower C-reactive protein levels, but not white blood cell count. Our results suggest that APOE genotype may influence C-reactive protein levels through non-inflammatory pathway.
Aged ; Apolipoproteins E/*genetics ; C-Reactive Protein/*metabolism ; Female ; Genetic Association Studies ; Genotype ; Humans ; Inflammation/*blood/immunology ; Leukocyte Count ; Male ; Middle Aged ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Polymorphism, Single Nucleotide/genetics ; Prospective Studies ; Republic of Korea

OBJECTIVETo study the effects of ApoE gene polymorphism on anti-inflammatory action of Xuezhikang Capsule.

METHODSOne hundred and two patients with hyperlipidemia (as the treated group) and one hundred healthy volunteers (as the control group) were enrolled in the case-control study. Total DNA of the peripheral blood was extracted and ApoE genotypes were determined by PCR sequence analysis. The serum levels of tumor necrosis factor alpha (TNF-alpha), interleukin 6 (IL-6), and high sensitivity C reactive protein (hs-CRP)were measured in all subjects. The changes of TNF-alpha, IL-6, and hs-CRP were detected before and after 6-week Xuezhikang Capsule treatment, thus analyzing the correlation between ApoE gene polymorphism and changes of each inflammatory factor.

RESULTSThe frequency of E3/3 genotype was 86% (86/100 cases)in the control group, significantly higher than that of the treated group (62.7%, 64/102 cases). The frequency of E3/4 genotype was 6% (6/100 cases) in the control group, significantly lower than that of the treated group (21.6%, 22/102 cases; both P < 0.05). Compared with the control group, the serum levels of TNF-alpha, IL-6, and hs-CPR were higher in the treated group before treatment (P < 0.05). In hyperlipidemia patients with E3/4 + E4/4 genotype, the serum level of TNF-alpha was higher than that of E3/3 genotype (P < 0.05); the serum level of IL-6 was higher than that of E2/E2 + E2/E3 genotype (P < 0.05); the serum level of hs-CRP was higher than that of E2/E2 + E2/E3 and E3/E3 genotype (P < 0.05). But there was no statistical difference in the serum levels of TNF-alpha, IL-6, or hs-CPR between E3/3 and E2/E2 + E2/E3 genotype. After 6-week intervention of Xuezhikang Capsule, the serum levels of TNF-alpha, IL-6, and hs-CRP were lower in the treated group (P < 0.05), but the serum levels of TNF-alpha and IL-6 were still higher than those of the control group (P < 0.05). But there was no statistical difference in the decrement of TNF-alpha, IL-6, or hsCRP among E2/E2 + E2/E3, E3/E3, or E3/4 + E4/4 genotypes (P > 0.05).

CONCLUSIONSThe distribution of ApoE gene polymorphism is different between the hyperlipidemia patients and the healthy people. Chronic inflammatory reactions exist in hyperlipidemia patients, especially in those with e4 allele. Xuezhikang Capsule showed anti-inflammatory effects, but ApoE gene polymorphism did not affect its effects.

Adult ; Aged ; Anti-Inflammatory Agents ; therapeutic use ; Apolipoproteins E ; genetics ; C-Reactive Protein ; metabolism ; Case-Control Studies ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Genotype ; Humans ; Hyperlipidemias ; drug therapy ; genetics ; Interleukin-6 ; blood ; Male ; Middle Aged ; Phytotherapy ; Polymorphism, Genetic ; Tumor Necrosis Factor-alpha ; blood

OBJECTIVETo develop a tight tetracycline-controlled HCV-C double transgenic mouse model.

METHODSBy crossbreeding of ApoE-rtTA-tTS transgenic mice with TRE-HCV-C transgenic mice, the double transgenic mice were produced in the F1 generation. The presence of HCV-C and tTS gene in the F1 generation was confirmed by PCR, followed by further identification and quantification of the transgene using Southern blot hybridization. The expression of HCV-C in the liver of the mouse model was detected immunohistochemically.

RESULTS AND CONCLUSIONTwo transgenic mice were obtained, which contained ApoE-rtTA-tTS and TRE-HCV-C genes in the genome. Five founders contained HCV-C gene as confirmed by PCR and Southern blot hybridization. The tight tetracycline-controlled system may facilitate further study of HCV-C gene expression and gene therapy of hepatic cellular carcinoma.

Animals ; Apolipoproteins E ; genetics ; Blotting, Southern ; Breeding ; Crosses, Genetic ; Female ; Gene Expression Regulation, Viral ; drug effects ; Hepacivirus ; genetics ; immunology ; Hepatitis C Antigens ; genetics ; immunology ; Male ; Mice ; Mice, Transgenic ; Polymerase Chain Reaction ; Tetracycline ; pharmacology ; Trans-Activators ; genetics ; Viral Core Proteins ; genetics

OBJECTIVETo observe the regulatory effect of assorted use of Chinese drugs for detoxifying and activating blood circulation on serum high sensitive C-reactive protein (hs-CRP) in apolipoprotein E gene knock-out [ApoE (-/-)] mice.

METHODSApoE (-/-) mice of 13-week old were devided into two groups, 12 in Group A fed with common forage and 98 in Group B with high lipid forage. Besides, 12 C57 BL/6J mice, 13-week old, were set as Group C fed with common forage. After being fed for 19 weeks, and the formation of vulnerable plaque had been confirmed in 2 mice of Group B, the other 96 mice were sub-divided into 8 groups, 12 in each group. Except that 0.4 mL normal saline was infused to the Group B1 as well as Group A and C, the other 7 sub-groups of Group B were respectively medicated with various drugs as follows: B2, Polydatin (PD, an extract from giant knotweed rhizome for detoxicating) 26. 6 mg/kg; B3, Xiongshao Capsule (XS, a Chinese herbal preparation for activating blood circulation), 110 mg/kg; B4, PD 53.2 mg/kg + XS 220 mg/kg; B5, PD 26. 6 mg/kg + XS 110 mg/kg; B6, PD 13.3 mg/kg + XS 55 mg/kg; B7, Lovastatin 3.3 mg/kg; and B8, Xuezhikang (XZK, a Chinese patent drug) 0.2 g/kg, all were administered by dissolving in 0.4 mL of distilled water for gastrogavage. The serum contents of hs-CRP were detected, with blood sample drawing from inferior vena cava, using enzyme-linked immunosorbent assay 17 weeks after medication.

RESULTSThe hs-CRP level was significantly higher in Group B1 than in Groups A and C (P <0.05, P <0.01). Comparisons between various sub-groups of Group B in hs-CRP content showed that hs-CRP in Group B2, B4 and B7 was lower than that in B1 (P <0.01), hs-CRP in Group B4 was the lowest, and hs-CRP was lower in Group B2 than in Group B3.

CONCLUSIONAssorted use of Chinese drugs for detoxifying and activating blood circulation could reduce serum hs-CRP level in ApoE (-/-) mice.

Animals ; Apolipoproteins E ; genetics ; metabolism ; Atherosclerosis ; drug therapy ; genetics ; metabolism ; physiopathology ; Blood Circulation ; drug effects ; C-Reactive Protein ; metabolism ; Disease Models, Animal ; Drugs, Chinese Herbal ; administration & dosage ; Female ; Humans ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; Random Allocation

OBJECTIVETo investigate the effects of the -384A>C polymorphism in the promoter region of endothelial lipase (EL) gene on serum lipid and apolipoprotein levels in healthy normolipidemic (HTG) and endogenous hypertriglyceridemic (HTG) subjects.

METHODSTwo hundred and fourteen healthy normolipidemic and 103 endogenous hypertriglyceridemic subjects from a population of Chinese Han nationality in Chengdu area were studied using restriction fragment length polymorphism (RFLPs). Serum lipids were measured by enzymatic kits and apolipoproteins AI, AII, B100, CII, CIII and E were measured by the radial immunadiffussion kits.

RESULTSThe frequency of the C allele at the -384A>C site in EL gene in the population (0.178) was higher than that of Japanese population (0.119) and Japanese Americans (0.115) (P < 0.01 and P < 0.01), respectively. No significant difference between normolipidemic and HTG groups was found in both allele and genotype frequencies. In normal group, subjects of the C allele carriers (A/C and C/C genotype carriers) had a higher serum mean concentration of TC, LDL-C and nHDL-C when compared with those of genotype AA (5.23 +/- 0.74 mmol/L vs 4.93 +/- 0.74 mmol/L, P=0.025; 3.27 +/- 0.74 mmol/L vs 2.98 +/- 0.80 mmol/L, P=0.038; 3.81 +/- 0.73 mmol/L vs 3.49 +/- 0.85 mmol/L, P=0.031, respectively). Similar result was only observed in female subgroup when male and female subgroups were further separated. No significant changes of lipid and lipoprotein levels were observed in the polymorphism in HTG group.

CONCLUSIONThese results suggest that the -384A>C polymorphism in the promoter region of the endothelial lipase gene is associated with serum TC, LDL-C, and nHDL-C levels in healthy Chinese subjects in Chengdu area, but not associated with the lipid levels in the endogenous hypertriglyceridmic group.

Adult ; Aged ; Aged, 80 and over ; Alleles ; Apolipoproteins C ; genetics ; Asian Continental Ancestry Group ; genetics ; Female ; Gene Frequency ; Humans ; Hypertriglyceridemia ; genetics ; Lipase ; genetics ; Male ; Middle Aged ; Polymorphism, Genetic ; Polymorphism, Restriction Fragment Length ; Population Groups ; genetics ; Promoter Regions, Genetic ; genetics ; Triglycerides ; blood ; Young Adult

BACKGROUND: Ischemic stroke is a complex condition influenced by many factors. Previous studies have demonstrated that inflammatory markers might play a role in such vascular diseases. Therefore the purpose of this study was to compare the expression of inflammatory markers in Korean ischemic stroke patients and to investigate their relationship to APOE polymorphism. METHODS: The patient group consisted of 275 patients with large artery atherosclerosis (LAA, n=169) and small artery occlusion (SAO, n=106). One hundred and nineteen age matched healthy subjects were recruited as the control group. Serum levels of three inflammatory markers (matrix metalloproteinase, MMP-9; tissue inhibitor of metalloproteinase-1, TIMP-1; and high-sensitivity C-reactive protein, hs-CRP) were measured in each patient by using commercially available kits. Comparison of clinical risk factors, inflammatory marker levels, and APOE genotypes between the stroke patient group and control group and between the two patient subgroups was assessed. RESULTS: Comparison of the stroke group to control group showed significantly elevated levels of circulating MMP-9 (P<0.01) and hs-CRP (P=0.01). Comparison between the individual subgroups revealed a significantly higher level of only TIMP-1 in the LAA subgroup compared to the SAO subgroup (P<0.01). There was no significant difference in inflammatory marker levels among each allele carrier. CONCLUSIONS: The present study revealed the obvious tendency of increased circulating inflammatory markers in the patients with acute ischemic attack, especially MMP-9 and hs-CRP. Our observations suggest that measurement of serum MMP-9, TIMP-1, and hs-CRP levels may be useful in the diagnosis of ischemic stroke patients.
Aged ; Apolipoproteins E/*genetics ; Biological Markers/blood ; Brain Ischemia/complications/*diagnosis ; C-Reactive Protein/analysis ; Carotid Artery Diseases/complications ; Female ; Genotype ; Humans ; Inflammation Mediators/*blood ; Korea ; Male ; Matrix Metalloproteinase 9/blood ; Middle Aged ; Polymorphism, Genetic ; Stroke/*diagnosis/etiology/immunology ; Tissue Inhibitor of Metalloproteinase-1/blood

OBJECTIVETo investigate the changes of several protein markers in a metastatic colorectal carcinoma model by serum proteomic analysis.

METHODSThe pEGFP-N1 plasmid with enhanced expression of green fluorescence protein (EGFP) was transfected into human colon carcinoma cell line SW480 to obtain a stable SW480-EGFP cell line, the SW480-EGFP cells were then injected subcutaneously into nude mice. The harvested tumor cells were implanted orthotopically into the colon of the nude mice. Real-time tumor growth and metastasis formation were visualized by whole-body fluorescent imaging system. Serum samples at different metastatic stages were collected and differential proteomic profiles were investigated by two-dimensional gel electrophoresis (2DE) and matrix-assisted laser absorption/ionization time of flight mass spectrometry (MALDI-TOF MS).

RESULTSThe SW480- EGFP cells enabled to express EGFP stably. The rates of subcutaneous and orthotropic tumor formation were 100%. The metastasis rates to local lymph nodes, liver and lung were 100%, 40% and 30%, respectively. Furthermore, 5 differentially expressed proteins were analyzed by serum proteome technologies, including haptoglobin alpha chain, apolipoprotein E, apolipoprotein A-IV, Ig kappa chain V region chain L and transferrin.

CONCLUSIONSVisualized metastatic model of colorectal carcinoma was successfully established. Several differentially expressed serum proteins collected at different stages after the occurrence of metastasis were identified. These differentially expressed proteins may be candidate serum biomarkers for diagnosis and therapeutic evaluation of colorectal carcinoma metastasis.

Animals ; Apolipoproteins A ; blood ; Apolipoproteins E ; blood ; Biomarkers, Tumor ; blood ; Blood Proteins ; analysis ; Cell Line, Tumor ; Colorectal Neoplasms ; blood ; genetics ; pathology ; Electrophoresis, Gel, Two-Dimensional ; Green Fluorescent Proteins ; genetics ; metabolism ; Haptoglobins ; analysis ; Humans ; Immunoglobulin kappa-Chains ; blood ; Liver Neoplasms ; secondary ; Lung Neoplasms ; secondary ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Transplantation ; Neoplasms, Experimental ; blood ; genetics ; pathology ; Proteomics ; methods ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Transfection ; Transferrin ; analysis ; Transplantation, Heterologous