Correction: Hyperhomocysteinemia in ApoE-/- Mice Leads to Overexpression of Enhancer of Zeste Homolog 2 via miR-92a Regulation.

[This corrects the article DOI: 10.1371/journal.pone.0167744.].

Effect of EZH2 on Hcy-induced cholesterol accumulation of foam cells / 实用医学杂志

Objective To explore the effect of EZH2 on Hcy-induced cholesterol accumulation of foam cells. Methods THP-1 foam cells were divided into control group ,100 μmol/L Hcy group and folic acid group. Lipid droplet in foam cells was tested by Oil red O. TC and TG contents in cells were determined by enzymic meth od. H3K27me3 level and EZH2 protein expression were detected by Western-blot. EZH2 mRNA expression was assayed by q-PCR. H3K27me3 level and TC and TG contents were examined followed by overexpression or knock- down of EZH2. Results After administration of Hcy,TC and TG contents in foam cells were increased (P <0.05). H3K27me3 level and EZH2 expression were also increased(P<0.05). Overexpression of EZH2 caused the expansion of H3K27me3 level,and the TC and TC contents were also increased(P<0.05). Conclusion Regula-tion of H3K27me3 by EZH2 might be involved in Hcy-induced accumulation of cholesterol in foam cells.

Effect of EZH2 on Hcy-induced cholesterol accumulation of foam cells / 实用医学杂志

Objective To explore the effect of EZH2 on Hcy-induced cholesterol accumulation of foam cells. Methods THP-1 foam cells were divided into control group ,100 μmol/L Hcy group and folic acid group. Lipid droplet in foam cells was tested by Oil red O. TC and TG contents in cells were determined by enzymic meth od. H3K27me3 level and EZH2 protein expression were detected by Western-blot. EZH2 mRNA expression was assayed by q-PCR. H3K27me3 level and TC and TG contents were examined followed by overexpression or knock- down of EZH2. Results After administration of Hcy,TC and TG contents in foam cells were increased (P <0.05). H3K27me3 level and EZH2 expression were also increased(P<0.05). Overexpression of EZH2 caused the expansion of H3K27me3 level,and the TC and TC contents were also increased(P<0.05). Conclusion Regula-tion of H3K27me3 by EZH2 might be involved in Hcy-induced accumulation of cholesterol in foam cells.

Efficacy of montmorillonite combined with oxygen in the treatment of neonatal diaper dermatitis / 中国生化药物杂志

Objective To observe the efficacy of montmorillonite combined with oxygen in the treatment of neonatal diaper dermatitis, and to provide the basis for clinical treatment. Methods Collected 100 cases of neonatal diaper dermatitis from January 2014 to January 2017 in our hospital. The patients were divided into observation group and control group according to the order of admission. After treatment, the observation group was given montmorillonite combined with oxygen therapy, the control group was given only oxygen treatment, compared the treatment effect of the two groups of patients. Results After treatment, the total effective rate of the observation group was 100％, the total effective rate of control group was 80％, there was significant difference between the two groups in the total effective rate of clinical treatment (P<0.05).The cure time was (116.20 ± 25.79) h in the observation group and (145.72 ± 24.88) h in the control group. The difference of the cure time between the two groups was statistically significant (P<0.05). In the observation group, the erythema was found in 42 children after treatment, and the area of erosion was obviously reduced before treatment, and the surface of the lesion was dry. After treatment for 5 days, 36 cases of mild to moderate diaper dermatitis were cured. After 7 days of treatment, children are cured. In the control group, 19 cases of mild diaper dermatitis were treated with erythema dermatitis . After treatment for 5 days, the area of hips in the 22 cases was smaller than that before treatment, and the lesion was dry. After 7 days, There was no significant improvement in erosion and or exudation. Conclusion Montmorillonite powder combined with oxygen treatment of neonatal diaper dermatitis is a significant effect, is the ideal treatment for neonatal diaper dermatitis.

Effect of FABP4 DNA methylation on abnormal lipid metabolism in trophocyte treated with L-NAME / 实用医学杂志

Objective To investigate the effect of fatty acid binding protein 4(FABP4)DNA methylation on abnormal lipid metabolism in placental trophoblastic dyslipidemia. Methods Human placental trophoblast cell line(HTR-8)was treated with L-NAME of 100 μmol/L for 48 h. The lipid content in placental trophoblasts was detected by chemical enzyme-colorimetry. The FABP4 DNA methylation level in placenta trophoblasts was detected by nested-touch down methylation specific PCR (NT-MSP). the mRNA and protein expression of DNMT1 and FABP4 were detected by qRT-PCR and Western Blot,respectively,in trophoblast cells. Results The lipid content in trophoblasts significantly increased as compared with the control(P < 0.05). Expression of FABP4 mRNA and protein increased(P < 0.05),while FABP4 methylation level and expression of DNMT1 significantly decreased (P<0.05)after treatment with L-NAME. Conclusions FABP4 DNA methylation is involved in the regulation of lipid metabolism in placental trophoblastic cells of hypertensive disorder complicating pregnancy.

Hyperhomocysteinemia in ApoE-/- Mice Leads to Overexpression of Enhancer of Zeste Homolog 2 via miR-92a Regulation.

Hyperhomocysteinemia (HHcy) is an independent risk factor for cardiovascular diseases, such as atherosclerosis. HHcy promotes atherogenesis by modifying the histone methylation patterns and miRNA regulation. In this study, we investigated the effects of homocysteine (Hcy) on the expression of enhancer of zeste homolog 2 (EZH2), and tested our hypothesis that Hcy-induced atherosclerosis is mediated by increased EZH2 expression, which is regulated by miR-92a. The levels of EZH2 and H3K27me3 were increased in the aorta of ApoE-/- mice fed a high-methionine diet for 16 weeks, whereas miR-92a expression was decreased. Over-expression of EZH2 increased H3K27me3 level and the accumulation of total cholesterol and triglycerides in the foam cells. Furthermore, upregulation of miR-92a reduced EZH2 expression in the foam cells. These data suggested that EZH2 plays a key role in Hcy-mediated lipid metabolism disorders, and that miR-92a may be a novel therapeutic target in Hcy-related atherosclerosis.

Feasibility analysis of serum calcitonin in the treatment of pediatric respiratory tract infections / 中国生化药物杂志

Objective To study the clinical effect of serum calcitonin in the treatment of pediatric respiratory tract infections.Methods A retrospective analysis of our hospital in the patient's medical records from April 2015 to April 2016 were 80 cases of pediatric respiratory tract infection, were randomly divided into control group and observation group, 40 cases of respiratory tract infection were treated with standard antibiotic therapy in the control group, the observation group was treated with antibiotics in the treatment of serum calcitonin under the guidance of the comparative analysis of two groups of children with clinical efficacy, clinical symptoms disappear time, antibiotic use and hospital economy.Results The effective rate of control group was 70%, and the observation group was 90%.The clinical efficiency of the observation group was relatively higher than control group ( P <0.05).Compared with the control group, the clinical symptoms of the patients in the observation group were significantly lower than that of the control group(P<0.05).Patients in the observation group used antibiotics was 16 cases(40%), the antibiotic use rate was, 32 cases(80%) in the control group, the average usage time of antibiotics and the average cost of observation group was significantly lower than control group(P<0.05)..The time of hospitalization and the hospitalization expenses of,observation group were lower than control group(P<0.05).Conclusion The treatment of pediatric respiratory infection in serum calcitonin can help improve the clinical efficacy, reduce the use of antibiotics in children with high economic.

Effect of miRNA-143 on homocysteine induced vascular smooth muscle cell proliferation / 中国药理学通报

Aim To explore the effect of miRNA-143 ( miR-1 4 3 ) on homocysteine ( Hcy ) induced-vascular smooth muscle cells ( VSMCs ) proliferation and the mechanism .Methods VSMCs were cultured and in-cubated with Hcy by using primary cultured method . Then, cells were treated with different concentrations of Hcy and folate .VSMCs proliferation was determined with MTT assay , miR-143 was measured by qRT-PCR, and methylation of miR-143 was determined with meth-ylated PCR.Results After cells were treated with dif-ferent concentrations of Hcy , the proliferation of VSMCs was significantly increased , mRNA expression of miR-143 was decreased and methylation of miR-143 was increased .The proliferation of VSMCs was signifi-cantly decreased when transfected VSMCs with miR-143 precursor , and cell proliferation was increased by using miR-143 inhibitor transfection .Conclusion Hy-pomethylation of miR-143 may inhibit VSMCs prolifera-tion.

CFTR suppressing hyperhomocysteinemia-induced hepatocyte damage in ApoE-/-mice / 实用医学杂志

Objective To investigate the function of CFTR in ApoE-/- mice with HHcy-induced hepato-cellular injury. Methods Thirty six 5-week old ApoE-/- mice were divided into three groups , including the ApoE-/- group, the HHcy group and the intervention group, (n = 12). Twelve normal C57BL/6J mice were fed with regular mouse diet as the normal control (SPF grade). HL-7702 human liver cells were intervened by Hcy (100 μmol/L) and 100 μmol/L Hcy + folic acid (100 μmol/L Hcy + F). The changes of Hcy, ALT and AST in the serum and the expression of CFTR mRNA and protein in liver and liver cells were detected. The concen-trations of ALT and AST in the liver cell intervened by VX-770 agonist and CFTR(inh)-172 inhibitor were mea-sured by ELISA. Results Compared with the control group , the levels of Hcy , ALT and AST were higher and the levels of CFTR mRNA and protein were lower in the Meth group (P < 0. 05 ) , while the reverse result in the Meth + F group (P < 0.05). Compared with the control group, the levels of CFTR mRNA and protein were de-creased and the levels of ALT and AST were increased in the 100 μmol/L Hcy group (P < 0.05). Compared with the 100 μmol/L Hcy group , the levels of CFTR mRNA and protein were increased and the levels of ALT and AST were decreased in the 100 μmol/L Hcy + F group (P < 0.05). Stimulated with VX-770 can reduce the concentrations of ALT and AST and the vice versa in the CFTR (inh)-17 group the concentration was increased in liver cells. Conclusion CFTR plays an important role in the regulation of hepatocellular injury by HHcy.

Role of homocysteine to promote the vascular smooth muscle cell proliferation by MiR-125 b methylation / 中国药理学通报

Aim To investigate the role of miR-125 b and its DNA methylation in homocysteine ( Hcy )-in-duced vascular smooth muscle cells( VSMCs) prolifera-tion. Methods VSMCs were stimulated with 0,50, 100, 200, 500 μmol · L-1 Hcy respectively. Then qRT-PCR was used to detect the mRNA levels of miR-125b,and nested-touchdown methylation-specific PCR ( ntMS-PCR) was used to detect the methylation levels of miR-125b. VSMCs were transfected with miR-125b precursor or the inhibitor of miR-125b ,then 3-(4,5-dimethylthiazol-2-yl)-2-5-diphenyl tetrazolium bromide ( MTT ) assay was used to reflect the proliferation of VSMCs. The distribution of CpG islands of miR-125b promoter region was analyzed by bioinformatics meth-ods. VSMCs were stimulated with 100 μmol·L-1 Hcy and transfected with or without DNA methylation inhib-itors 5-nitrogen impurity cytidine ( AZC) , then the ex-pression of miR-125b was detected by qRT-PCR. Re-sults The mRNA levels of miR-125 b were decreased in 100,200,500 μmol·L-1 Hcy group compared with 0 μmol·L-1 Hcy group. The precursor of miR-125b could inhibit the proliferation activity and the inhibitor of miR-125 b could increase the proliferation activity of VSMCs cells. Bioinformatics analysis indicated that MiR-125 b promoter region had a CpG island whose length was 792 bp ( 1881-2672 ) . The miR-125 b pro-moter region methylation levels increased after Hcy in-tervention ( P <0. 01 ) . The expression level of miR-125 b increased after AZC intervention ( P <0. 05 ) . Conclusions ① Hcy promotes vascular smooth mus-cle cell proliferation maybe by down-regulating the ex-pression of miR-125b. ② Hcy down-regulates the ex-pression of miR-125 maybe by up-regulating the methy-lation levels of miR-125b promoter region.

Role of ERO1αand its DNA methylation in homocysteine-induced inhibition of hepatocyte proliferation / 中国药理学通报

Aim To explore the role of ERO1 αand its DNA methylation in homocysteine (Hcy)-induced in-hibition of hepatocytes proliferation.Methods The hepatocytes stimulated with 0 μmol·L -1 Hcy were set as the normal group (NC group)and the hepatocytes stimulated with 1 00 μmol·L -1 Hcy as the experimen-tal group (Hcy group).Methyl thiazolyl tetrazolium (MTT)reduction assay was used to reflect the prolifer-ation of the hepatocytes;qRT-PCR and Western blot were used to detect the mRNA and protein levels of ERO1 α;the expression of green fluorescence protein was observed in hepatocytes after the recombinant plas-mid of ERO1 α was constructed,which was used to confirm if the recombinant plasmid into hepatocytes was successful,then the mRNA and protein levels of ERO1 αwere assayed and the proliferation of the hepa-tocytes was also detected;ntMSP was used to detect the change of ERO1 αDNA methylation.Results The mRNA and protein levels of ERO1 αwere decreased in Hcy group compared with NC group,and the prolifera-tion activity of hepatocytes in Hcy group was de-creased.Sequencing result showed that the recombi-nant plasmid of ERO1 αwas constructed successfully. QRT-PCR and Western blot revealed that ERO1 αwas overexpressed. The result of MTT suggested that ERO1 αoverexpression restored hepatocyte proliferation inhibited by Hcy.Hcy caused ERO1 αDNA hyperm-ethylation.Conclusions Hcy inhibits hepatocyte pro-liferation by downregulating the expression of ERO1 α, and methylation of ERO1 αpromoter may play a role in this process.

The levels of NO decrease induced apoptosis in human placental trophoblast cells through oxidative stress / 中国药理学通报

Aim To investigate the possible mecha-nisms of the levels of NO decrease induced apoptosis in human placental trophoblast cells. Methods Human placental trophoblast cells ( HTR-8 ) were cultured in 5 ml DMEM-F12 culture medium with 37℃ 5% CO2 . Then, the old culture medium was discarded and re-placed with 10,100,500,1 000 μmol·L-1 L-NAME, and the group without L-NAME was set as the control group, cultured for 48h. The effects of L-NAME on the survival of cells were detected by methylthiazolyldiphe-nyl tetrazolium bromide ( MTT); the content of NO in cells was tested by nitrate reductive enzymatic;trans-mission electron microscopy, flow cytometry analysis and Annexin-V FITC dyeing were used to test the effects of L-NAME on apoptosis in HTR-8 cells;restore Fe3+ colorimetric assay was applied for detection of to-tal antioxidant capacity ( T-AOC ) , xanthine oxidase for detection of superoxide dismutase ( SOD) activity, and thiobarbituric acid colorimetry for determination of content of MDA. Results Compared with the control group, the survival rate of HTR-8 cells and the levels of NO in 100,500,1 000 μmol·L-1 L-NAME group were significantly reduced(P<0.05,P<0.01). Flow analysis and Annexin-V FITC staining showed that L-NAME could induce cell apoptosis in a dose-dependent manner. The number of cell apoptosis was negatively correlated with the content of NO ( r = -0.5210 ) in HTR-8 cells. Transmission electron microscopy results showed that compared with the control group, the ex-perimental group's cell nucleus shape was irregular, nuclear pyknosis in irregular shape, the chromatin ag-glutination or side the collection, mitochondrial swell-ing or enrichment, crest fracture or dissolved, even vanished, forming the vacuole, especially in 100 μmol ·L-1 L-NAME group, the apoptotic bodies obviously appeared. At the same time, T-AOC, SOD levels in HTR-8 cells decreased ( P <0.05 ) , and the MDA content increased ( P<0.05 ) . The number of cell ap-optosis was negatively correlated with the level of T-AOC ( r= -0.3212 ) , SOD ( r= -0.2779 ) in HTR-8 cells , while positively correlated with the content of MDA(r=0.2807). Conclusion Oxidative stress may play an important role in the levels of NO decrease in-duced apoptosis in human placental trophoblast cells.

Homocysteine-mediated PPARalpha,gamma DNA methylation and its potential pathogenic mechanism in monocytes.

Homocysteine (Hcy) is an independent risk factor for cardiovascular disease, but the molecular mechanisms causing atherosclerosis in monocytes remain poorly characterized. The objective of the present study was to investigate the effects of Hcy on DNA methylation of PPARalpha,gamma and the underlying mechanism of PPARalpha,gamma expression that was induced by Hcy in monocytes. About 50, 100, 200, and 500 microM Hcy were added to the monocytes cultured for 48 h. PPARalpha,gamma that acted as lipid sensors and bind with mM affinities to ligands of antiatherosclerosis were determined by real-time reverse transcription-polymerase chain reaction and Western blotting in monocytes. Here, we used a high-throughput quantitative methylation assay that utilizes fluorescence-based real-time polymerase chain reaction to determine the levels of the PPARalpha,gamma DNA methylation. S-adenosylmethionine (SAM) level and S-adenosylhomocysteine (SAH) level were detected by high performance liquid chromatography. Results indicated that the levels of PPARalpha,gamma promoter methylation in monocytes cultured with Hcy were increased in comparison with the control group, and the peak was in the 100 muM Hcy group, however, a dose-dependent increase with increasing Hcy was not seen. Hcy also decreased mRNA and protein levels of PPARalpha,gamma in monocytes. Further, with the addition of Hcy, the levels of SAH were elevated, the levels of SAM and the ratio of SAM/SAH were lower, and the activity of C-5MT-ase was increased. In conclusion, these results suggest that PPARalpha,gamma DNA methylation induced by Hcy may represent an important mechanism to explain atherosclerosis, which may become a therapeutic target for preventing atherosclerosis induced by Hcy.

Homocysteine-mediated expression of SAHH, DNMTs, MBD2, and DNA hypomethylation potential pathogenic mechanism in VSMCs.

Homocysteine (Hcy) is a well-established risk factor for atherosclerosis and may cause dysregulation of gene expression, but the characteristics and the key links involved in its pathogenic mechanisms are still poorly understood. The aim of this study was to explore (i) the effects of Hcy on DNA methylation in vascular smooth muscle cells (VSMCs) and (ii) the underlying mechanism of Hcy-induced changes in DNA methylation patterns in relation to atherosclerosis. We examined the levels of gDNA methylation, namely, the Alu and line-1 element sequences, which can serve as a surrogate marker for gDNA methylation, and also investigated the effects of Hcy on the intracellular S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH) concentrations as well as the expressions of SAH hydrolase (SAHH), DNA methyltransferase3a (DNMT3a), DNMT3b, and methyl-CpG-binding domain 2 (MBD2). We found that clinically relevant levels of Hcy (0-500 microM) induced elevation of SAH, declination of SAM and SAM/SAH ratio, and reduction in expression of SAHH and MBD2, but increased the activity of DNMT3a and DNMT3b compared to the control group (p < 0.05). We found also that the genome-wide hypomethylation is a common feature of gDNA in the VSMCs cultured with Hcy. In conclusion, these results suggest that Hcy-induced DNA methylation may be an important potential pathogenic mechanism in the development of atherosclerosis, and may become a therapeutic target for preventing Hcy-induced atherosclerosis.

Homocysteine-mediated Aberrant DNA Methylation in Vascular Smooth Muscle Cells and Its Potential Pathogenic Mechanism / 生物化学与生物物理进展

Hyperhomocysteinemia, which is an independent risk factor for atherosclerosis, may cause aberrant methylation and dysregulation of gene expression, but the characteristics of the aberrant methylation and its key links involved in its pathogenic mechanisms are still poorly understood. The effect of hyperhomocysteine on DNA methylation in vascular smooth muscle cells, its characteristics and the underlying mechanism of Hcy-induced changing in DNA methylation patterns were investigated. Clinical relevant concentrations of homocysteine was added into the cultured vascular smooth muscle cells of the Homo sapien umbilical vein for 24 h. The level of SAM and SAH was detected by HPLC. The activity of SAH Hydrolase was detected by real-time quantitative reverse transcription-PCR and Western blotting analysis. The level and patterns of DNA methylation were measured by endogenous C-5 DNA methyltransferase(C-5 MT-ase) activity and capacity of genomic DNA to accept methyl groups and methylation-dependent restriction analysis. The results indicated that an increased Hcy concentration induced elevated SAH, declined SAM and the ratio of SAM/SAH, reduced expression of SAH Hydrolase, but increased activity of C-5MT-ase. The methylation status of gDNA analyzed by methyl-accepting capacity of gDNA uncovered a demethylation process in gDNA, or homocysteine-caused hypomethylation in gDNA.With different methylation-dependent restriction endonucleases, the aberrant demethylation was found to prefer C↓CGG sequences to CpG islands. The impacts of different dosage of Hcy showed that the varied detrimental effects of Hcy could be attributed to different concentrations via different mechanisms. In mild and moderate hyperhomocysteinemia, the Hcy may primarily influence the epigenetic regulation of gene expression through the interference of transferring methyl-group metabolism, while in more higher Hcy concentration, the notorious impacts may be more directly caused via oxidative stress, apoptosis, inflammation etc.

Effects of matrine on ATP binding cassette transporter A1 and cholesterol in monocyte cells / 基础医学与临床

Objective To study the effect of matrine on ATP binding cassette transporter A1 (ABCA1) expression and cholesterol in monocyte derived foam cells. Methods The lipid peroxide in cells was measured by TBARS method; The foam cells were examined by oil red O stain. The accumulation of cholesterol in monocyte was measured by fluorescence spectrophotometric method;ABCA1 mRNA and its protein level were determined by RT-PCR and Western blot, respectively. Results In the homonocytes incubated with PMA and low density lipoprotein (LDL), the intracellular accumulated total cholesterol (TC), free cholesterol (FC), cholesteryl ester (CE) and lipid peroxide increased obviously, and a huge amount of foam cells were found by oil red O stain. This was accompanied by a reduction in the membrane content of ABCA1. matrine alter ABCA1 mRNA abundance, indicating that increased ABCA1 transcription, enhanced mRNA level.Conclusion The mechanism of anti-artherosclerosis by matrine may be related to reduce cholesterol and to increase the level of ABCA1 expression.