Clinical Significance of Upregulation of EZH1 Expression in Hepatocellular Carcinoma Tissues.

BACKGROUND AND AIMS: The incidence and mortality of hepatocellular carcinoma (HCC) are increasing. It is urgent to develop more effective HCC biomarkers for diagnosis and treatment. This project intends to verify the expression of enhancer of zeste 1 polycomb repressive complex 2 subunit (EZH1) and its mechanism in HCC. METHODS: This study integrates global microarray and high-throughput sequencing datasets, combined with internal immunohistochemistry, to analyze the expression and prognostic value of EZH1 in HCC. Functional enrichment analysis was conducted to investigate transcriptional targets, which were achieved by intersecting HCC over-expressed genes, EZH1 co-expressed genes and putative transcriptional targets. The relationship between EZH1 and anticancer drugs was detected by drug sensitivity analysis. RESULTS: In this study, 84 datasets from 40 platforms (3,926 HCC samples and 3,428 non-cancerous liver tissues) were included to show the high expression of EZH1 in HCC. Immunohistochemistry with 159 HCC samples and 62 non-HCC samples confirmed the high expression level. HCC patients with high EZH1 expression had worse survival prognoses. Gene ontology and Reactome analysis revealed that metabolism-related pathways, including autophagy, are critical for HCC. Interestingly, as one of the EZH1 potential transcriptional targets, autophagy-related 7 (ATG7) appeared in the above pathways. ATG7 was positively correlated with EZH1, upregulated in HCC, and mediated poor prognosis. Upregulation of EZH1 was found to be in contact with HCC anti-tumor drug resistance. CONCLUSIONS: The upregulation of EZH1 expression can promote the occurrence of HCC and lead to poor clinical progression and drug resistance; these effects may be mediated by regulating ATG7.

Formononetin, J1 and J2 have different effects on endothelial cells via EWSAT1-TRAF6 and its downstream pathway.

Formononetin is a natural isoflavone compound found mainly in Chinese herbal medicines such as astragalus and red clover. It is considered to be a typical phytooestrogen. In our previous experiments, it was found that formononetin has a two-way regulatory effect on endothelial cells (ECs): low concentrations promote the proliferation of ECs and high concentrations have an inhibitory effect. To find a specific mechanism of action and provide a better clinical effect, we performed a structural transformation of formononetin and selected better medicinal properties for formononetin modifier J1 and J2 from a variety of modified constructs. The MTT assay measured the effects of drugs on human umbilical vein endothelial cell (HUVEC) activity. Scratch and transwell experiments validated the effects of the drugs on HUVEC migration and invasion. An in vivo assessment effect of the drugs on ovariectomized rats. Long-chain non-coding RNA for EWSAT1, which is abnormally highly expressed in HUVEC, was screened by gene chip, and the effect of the drug on its expression was detected by PCR after the drug was applied. The downstream factors and their pathways were analysed, and the changes in the protein levels after drug treatment were evaluated by Western blot. In conclusion, the mechanism of action of formononetin, J1 and J2 on ECs may be through EWSAT1-TRAF6 and its downstream pathways.

The Correlation of Serum Myeloid-Related Protein-8/14 and Eosinophil Cationic Protein in Patients with Coronary Artery Disease.

OBJECTIVE: To investigate the changes in serum Myeloid-Related Protein 8/14 (MRP8/14) and Eosinophil Cationic Protein (ECP) levels in patients with different types of coronary artery diseases (CAD) and assess the value of MRP8/14 and ECP detection in predicting CAD. METHODS: 178 patients were divided into CAD group including unstable angina pectoris (UAP), acute myocardial infarction (AMI), and stable angina pectoris (SAP). Thirty-six individuals with normal coronary artery served as the control group. Serum MRP8/14 and ECP were measured by ELISA. The severity of coronary artery stenosis was assessed by the numbers of involved coronary artery branches and the sum of Gensini scores. RESULTS: The MRP8/14 levels were significantly higher in AMI and UAP group than SAP and control group (P < 0.05). The levels of MRP8/14 in AMI group were also obviously higher than UAP group (P < 0.05). The ECP levels were obviously increased in AMI group, but there was no difference between SAP and UAP group (P > 0.05). The ECP was significantly increased in three impaired coronary arteries and obviously correlated with Gensini score (P < 0.01), whereas the MRP8/14 was obviously positively correlated with CRP (P < 0.01). CONCLUSIONS: Increased MRP8/14 levels suggest the instability of the atherosclerotic plaque. ECP reflects the severity of coronary arteries stenosis, predicting atherosclerosis burden. They may become the new biomarkers of CAD.

[Effect of Kuanxiong aerosol on coronary heart disease angina patients: a multicenter randomized controlled clinical study].

OBJECTIVE: To evaluate the anginal attack-relieving efficacy and safety of Kuanxiong Aerosol (KA) in patients with coronary heart disease (CHD). METHODS: A total of 780 patients confirmatively diagnosed as CHD angina from November 2011 to December 2012 in 13 medical centers in the mainland area were assigned to 2 groups by blocked randomization, the treatment group (376 cases) and the control group (374 cases). When the angina attacked, patients in the treatment group received sublingual spray three times, 0.6 mL each time, while those in the control group sublingually dissolved Nitroglycerin Tablet (NT), 0.5 mg each tablet. The effective rate of angina relief, efficacy of electrocardiogram (ECG), and the incidence of adverse reactions were observed. RESULTS: The 3 min and 5 min remission rates of angina attack were 53.72% (202/376) and 94.41% (355/376) in the treatment group, and 47.86% (179/374) and 90.64% (339/374) in the control group. The 95% confidence interval (CI) of the difference between the 2 groups of 3 min and 5 min remission rates of angina attacks were [(-1.84%, 12.32%) and (-1.33%, 6.85%) respectively, P > 0.05]. The total improvement rates of ST-T changes in the treatment group and the control group after treatment were 74.07% and 73.13% respectively (P > 0.05). The adverse reaction rate was 9.31 (35/376 cases) in the treatment group and 22.46% (84/374 cases) in the control group (P < 0.01). CONCLUSION: KA was not inferior to NT in relieving anginal attacks and improving ischemic ECG changes, and had obviously less adverse reaction.

Use of evidence-based pharmacotherapy for secondary prevention of coronary heart disease: a Chinese medicine hospital versus a general hospital.

OBJECTIVE: To determine differences in adherence to secondary prevention guidelines (pharmacological interventions) among coronary heart disease (CHD) patients between a Chinese medicine (CM) hospital and a general hospital in a Chinese city. METHODS: Medical records of 200 patients consecutively discharged from the CM hospital and the general hospital for CHD were reviewed to determine the proportions of eligible patients who received antiplatelet agents, ß-blockers, angiotensin-converting enzyme (ACE) inhibitors or angiotensin receptor blockers (ARBs) and statins at discharge. The effects of patient characteristics and hospital type on the use of these medicines were estimated using logistic regression models. RESULTS: Patients discharged from the CM hospitals were older; more likely females; had greater history of hyperlipidemia, cerebrovascular diseases and less smoker (P<0.01 or P<0.05). They were less likely to receive coronary angiography and percutaneous coronary intervention, and had a longer length of stay than those discharged from the general hospital (P<0.01 or P<0.05). There were no significant differences in antiplatelet agents (96% vs. 100%, P=0.121) or statins (97.9% vs. 100%, P=0.149) use between the CM hospital and the general hospital. In multivariable analyses that adjusted for patient characteristics and hospital type, there was no significant difference in use of ß-blockers between the CM hospital and the general hospital. In contrast, patients discharged from the CM hospital were less likely to receive ACE inhibitors/ARBs compared with those discharged from the general hospital (odds ratio: 0.3, 95% confidence interval: 0.105-0.854). CONCLUSION: In this study, the CM hospital provides the same quality of care in CHD for prescribing evidence-based medications at discharge compared with another general hospital except for ACE inhibitors/ARBs use.

DEP domain containing 1 is a novel diagnostic marker and prognostic predictor for hepatocellular carcinoma.

BACKGROUND: This study was conducted to determine DEPDC1 expression in hepatocelluar carcinomas (HCCs) and to reveal its potential role in diagnosis and prognosis of affected patients. MATERIALS AND METHODS: DEPDC1 expression at the mRNA level was detected by quantitative real-time PCR (qRT-PCR) in 205 cases of HCC and paired adjacent normal liver tissues, and by semi-quantitative RT-PCR in 20 cases. Survival curves were obtained by using Kaplan-Meier method and Log-rank test. Independent predictors associated with regard to disease free survival (DFS) and overall survival (OS) were identified using the Cox proportional hazard model. RESULTS: High DEPDC1 mRNA levels were detected in 144 out of 205 cases (70.24%) of HCC, significantly associated with clinicopathological parameters, including tumor size (≥4cm), alpha-fetoprotein (≥100ng/ml), B-C of BCLC stage and recurrence. Kaplan-Meier survival analysis revealed that HCC patients with high DEPDC1 expression had poor OS and DFS. Multivariate analysis demonstrated that high DEPDC1 expression was an independent predictor of OS (HR=1.651; 95% 95%CI, 1.041- 2.617; p=0.033) and DFS (HR=1.583; 95%CI, 1.01- 2.483; p=0.045). CONCLUSIONS: Our results indicate DEPDC1 might be a novel diagnostic marker and an independent prognostic predictor for HCC patients.

Association of single nucleotide polymorphisms in IL-12 and IL-27 genes with colorectal cancer risk.

OBJECTIVES: Interleukin-12 (IL-12) plays an important role in antitumor immunity. Interleukin-27 (IL-27) is a novel IL-12 family member. The present studies demonstrate that IL-27 mediates potent antitumor activity. However, No studies have examined the association of these polymorphism with colorectal cancer (CRC). Therefore, we investigated the relationship of IL-12 and IL-27 gene polymorphisms and CRC. DESIGN AND METHODS: We analyzed polymorphisms of IL-12 gene 16974 A/C and IL-27 gene -964 A/G, 2905 T/G, 4730 T/C in 410 patients with CRC and 450 controls, using PCR-RFLP method. RESULTS: There were no significant differences in the genotype and allele frequencies of IL-12 and IL-27 gene polymorphisms between the group of patients with CRC and the controls. Furthermore, no association was found between IL-12 family gene polymorphisms and different clinical stages in patients with CRC. CONCLUSION: These findings suggest that IL-12 and IL-27 gene polymorphisms may not be involved in susceptibility to CRC.

Curculigoside attenuates human umbilical vein endothelial cell injury induced by H2O2.

AIM OF THE STUDY: Vessel endothelium injury caused by reactive oxygen species (ROS) including H(2)O(2) plays a critical role in the pathogenesis of cardiovascular disorders. Therefore, agents or antioxidants that can inhibit production of ROS has highly clinical values in cardiovascular therapy. Curculigoside is the major bioactive compounds present in Curculigo orchioides, and possess potent antioxidant properties against oxidative stress insults through undefined mechanism(s). The present study was designed to test the hypothesis that curculigoside can inhibit H(2)O(2)-induced injury in human umbilical vein endothelial cells. MATERIALS AND METHODS: Human umbilical vein endothelial cells (HUVECs) were treated with curculigoside in the presence/absence of hydrogen peroxide (H(2)O(2)). The protective effects of curculigoside OP-D against H(2)O(2) were evaluated. RESULTS: HUVECs incubated with 400 µM H(2)O(2) had significantly decreased the viability of endothelial cells, which was accompanied with apparent cells apoptosis, the activation of caspase-3 and the upregulation of p53 mRNA expression. In addition, H(2)O(2) treatment induced a marked increase of MDA, LDH content and in intracellular ROS, decreased the content of nitric oxide (NO) and GSH-Px activities in endothelial cells. However, pretreatment with 0.5.5,10 µM curculigoside resulted in a significant recovery from H(2)O(2)-induced cell apoptosis. Also, it decreased other H(2)O(2)-induced damages in a concentration-dependent manner. Furthermore, pretreatment with curculigoside decreased the activity of caspase-3 and p53 mRNA expression, which was known to play a key role in H(2)O(2)-induced cell apoptosis. CONCLUSION: The present study shows that curculigoside can protect endothelial cells against oxidative injury induced by H(2)O(2), suggesting that this compound may constitute a promising intervention against cardiovascular disorders.

Evaluation of recombinant endostatin in the treatment of atherosclerotic plaques and neovascularization in rabbits.

OBJECTIVE: Atherosclerotic plaques and neovascularization play an important role in the course of coronary atherosclerosis. This study evaluated the effect of recombinant endostatin on experimental atherosclerotic plaques and neovascularization in rabbits. METHODS: Eighteen healthy male rabbits were divided into three groups: control group, atherosclerotic model group, and recombinant endostatin treated group. The atherosclerotic model was established via a high-cholesterol diet after balloon catheter injury. The subject weights, serum total cholesterol, creatine kinase-myocardial band fraction (CKMB), and matrix metalloproteinase-2 (MMP-2) were measured. Six weeks after treatment, the aortic roots were taken for pathological assay. The thickness ratio of the intima to media was measured by hematoxylin and eosin (HE) staining, and the number of neovessels was measured by immunohistochemistry via monoclonal antibody CD31 staining. RESULTS: The weight, plasma total cholesterol, and CKMB were not significantly different between the atherosclerotic model group and the recombinant endostatin treated group, but much higher than those of the control group (P<0.05). The thickness ratio of the intima to media in the recombinant endostatin treated group was distinctly less than that in the atherosclerotic model group (P<0.05). The number of neovessels decreased dramatically (P<0.05) and the content of MMP-2 decreased slightly without statistical difference (P>0.05) in the recombinant endostatin treated group, compared to the atherosclerotic model group. CONCLUSIONS: Recombinant endostatin is able to inhibit the growth of neovascularization in the atherosclerotic plaque and the development of plaque.

The polymorphism of XRCC3 codon 241 and AFB1-related hepatocellular carcinoma in Guangxi population, China.

PURPOSE: The relationship between aflatoxin B1 (AFB1) exposure and hepatocellular carcinoma (HCC) has been previously demonstrated and supported with strong epidemiological evidence. However, the role of genetic polymorphism of X-ray cross-complementing group 3 (XRCC3) codon 241 (namely: Thr241Met), which may be involved in the repair of DNA double-strand breaks caused by carcinogens such as AFB1, been less well elaborated. METHODS: We conducted a case-control study including 491 cases and 862 controls to evaluate the associations between this polymorphism and HCC risk for Guangxi population by means of polymerase chain reaction-restriction fragment length polymorphism analysis. RESULTS: We found that individuals with the XRCC3 genotypes with codon 241 Met (namely XRCC3-TM or XRCC3-MM) had an increased risk of HCC than those with the homozygote of XRCC3 codon 241 Thr alleles (namely XRCC3-TT, adjusted odds ratios 2.22 and 7.19; 95% confidence intervals 1.72-2.88 and 4.52-11.42, respectively). The risk of HCC, moreover, did appear to differ more significantly among individuals featuring high-level AFB1-DNA adducts, whose adjusted odds ratios (95% confidence intervals) were 11.59 (5.73-23.47) and 37.54 (16.32-86.32), respectively. CONCLUSIONS: These findings support the hypothesis that the XRCC3 Thr241Met polymorphism may be associated with the risk of AFB1-related HCC among the Guangxi population.

Icariin enhances endothelial nitric-oxide synthase expression on human endothelial cells in vitro.

Icariin, a flavonoid isolated from Epimedii herba, is considered to be the major therapeutical constituent of E. herba. The aim of this study was to investigate the possible protective effects and to clarify the mechanism of icariin on endothelial cells in vitro. Incubation of human umbilical vein endothelial cells (HUVEC) derived EA. hy926 cells with icariin(0.1, 1, 10 micromol l(-1)) from 6 h to 72 h, then the production of NO was measured to evaluate the protective effects of icariin. RT-PCR was employed to confirm the mRNA expression of endothelial nitric oxide synthase (eNOS). Western blotting was used to evaluate the protein expression of eNOS. NO production was enhanced in a time- and concentration-dependent manner (P<0.05), which was well matched with the expression of eNOS mRNA (up to 2.4-fold) and protein (up to 2.5-fold) after long-term incubation with icariin in endothelial cells (P<0.05). Moreover, activated NF-kappaB was increased in EA. hy926 cells incubated with icariin for 24 h, in association with an increase in the expression of eNOS gene. In addition to its long-term effects on eNOS expression, icariin also enhanced the production of bioactive NO in the short-term (after a 5 min incubation, P<0.05). In concert with other effects, the protective effects of icariin on endothelial cells may contribute to the cardiovascular protective effects.

Vasorelaxant effects of icariin on isolated canine coronary artery.

Owing to its cardiovascular therapeutical effects, icariin, a flavonoid isolated from Epimedii herba, is considered to be the major active constituent of Epimedii herba. The aim of this study is to investigate the effect of icariin on precontracted coronary artery isolated from canine. Coronary artery segments were isolated from normal anesthetized Beagle dogs and cut into 5-mm rings. The rings were mounted in an organ chamber and contracted by either 40 mM KCl or 10 microM PGF2alpha, and vasorelaxant tone to icariin was measured. Treatment of icariin could significantly produce a relaxation of precontracted coronary arterial rings with intact endothelium in a concentration-dependent manner. Comparatively, the vasorelaxation disappeared in denuded-endothelium rings. Furthermore, the vasorelaxant effect of icariin was blocked by Nomega-Nitro- L-arginine Methyl Ester (L-NAME), 1H-[1, 2, 4]-oxadiazolo [4, 3-a] quinoxalin-1-one (ODQ) but not by indomethacin and glibenclamide, respectively. Tetraethylammonium (TEA) could partly antagonize the vasorelaxant effect triggered by icariin. There was no significant gene expression difference of the endothelial nitric oxide synthase (eNOS) gene in coronary arterial rings among the different concentrations of icariin by RT-PCR, but the activity of eNOS was increased in a concentration-dependent manner after icariin exposure. These results suggest that icariin produces NO-dependent relaxation in the isolated canine coronary artery, and the possible mechanism is involved in the activation of eNOS protein and NO-cGMP pathway.

[Elevated serum secretory type II phospholipase A2 in patients with coronary heart disease].

OBJECTIVE: To measure the serum level of secretory type II phospholipase A2 (sPLA2) in patients with coronary heart disease and investigate the possible relationship with IL-8 and LPA. METHODS: A total of 110 patients with acute coronary syndrome (ACS), 63 patients with stable coronary heart disease (SCHD) group and 89 non-CHD control patients were studied. Serum levels of sPLA2, IL-8, LPA and hs-CRP were measured and the correlation among these parameters was observed. RESULTS: The levels of serum sPLA2 [(68 +/- 17) U/ml], IL-8 [(182 +/- 80) pg/ml] and LPA [(2.85 +/- 0.36) micromol/L] were significantly higher in CHD patients than those in controls [sPLA2: (55 +/- 12) U/ml; IL-8: (119 +/- 33) pg/ml; LPA: (2.34 +/- 0.36) micromol/L, all P < 0.01], and sPLA2 and IL-8 were also significantly higher in ACS patients [sPLA2: (71 +/- 18) U/ml; IL-8: (195 +/- 78) pg/ml] than those in SCHD patients [sPLA2: (63 +/- 12) U/ml; IL-8: (159 +/- 79) pg/ml, both P < 0.01]. Serum sPLA2 level was positively correlated with hs-CRP, IL-8 and LPA (r = 0.203, P = 0.007; r = 0.658, P < 0.01; r = 0.231, P = 0.005, respectively). The relative risk of having CHD is 6.248 (P < 0.01) with the sPLA2 level above 63.75 U/ml. CONCLUSION: Elevated serum sPLA2 level is a risk factor for CHD.

Protective effects of silybin on human umbilical vein endothelial cell injury induced by H2O2 in vitro.

Oxidative injury induces cellular and nuclear damages that lead to cell injury. Agents or antioxidants that can inhibit production of reactive oxygen species can prevent injury. We tested the hypothesis that silybin can inhibit H2O2-induced injury in human umbilical vein endothelial cells. Eighteen hours of treatment with 750 micromol l(-1) H2O2 significantly stimulated expression of caspase-3 and cell apoptosis. In addition, it is observed that H2O2 increased the amounts of malondialdenhyde (MDA), the dehydrogenase (LDH) leakage, and decreased the activity of GSH-Px and NO contents in ECV-304 cells. In the H2O2 apoptosis model, the addition of 6.25-25 mg/L of silybin, which has in vitro radical scavenging activity, partially restored cell viability with a reduction in H2O2-induced apoptotic DNA damage, and decreased the expression of caspase-3. Moreover, it decreased other H2O2-induced damage in a concentration-dependent manner. The endothelial cell apoptosis was detected by AO/EB dual staining as well as flow cytometry, and the activity of pro-apoptotic factor caspase-3 was detected by immunocytochemical method. Our results suggest that the antioxidant, silybin, protects ECV-304 cells against H2O2-induced injury probably through its antioxidant activity, increasing the NO content, the activity GSH-Px and inhibiting signaling pathways mediated by caspase-3.

Protective effects of icariin on human umbilical vein endothelial cell injury induced by H2O2 in vitro.

Icariin is a flavonoid isolated from Epimedium and is considered to be the major pharmacological active component of Epimedii Herba. In the present investigation, we studied and confirmed the protective activity of icariin on H2O2-induced injury in human umbilical vein endothelial cell line: ECV-304. Eighteen-hour treatment with 750 micromol l(-1) H2O2 significantly decreased the viability of ECV-304 cells, which was accompanied with apparent apoptotic features, including distinct cell morphological alteration and the increase of caspase-3 expression. In addition, it is observed that H2O2 increased the amounts of malondialdenhyde (MDA) and the dehydrogenase (LDH), and decreased the content of nitric oxide (NO) in ECV-304 cells. However, pretreatment with 0.1-50 micromol l(-1) icariin resulted in a significant recovery from H2O2-induced cell apoptosis. Also, it decreased other H2O2-induced damage in a concentration-dependent manner. Furthermore, pretreatment with icariin decreased the expression of caspase-3, which was known to be involved as a key role executor in H2O2-induced cell apoptosis. The endothelial cells apoptosis were detected by acridine orange/ethidium bromide (AO/EB) dual staining as well as flow cytometry, and the expression of pro-apoptotic factor caspase-3 were detected by immunocytochemical method. Taken together, these data suggest that protective effects of icariin against oxidative injuries of ECV-304 cells may be achieved via decreasing of caspase expression.