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Objective@#Investigate the effects of inducible ppp2r1a knockout on main physiological function in adult mice and study the mechanism.@*Methods@#Ppp2r1aflox/flox mice and CAGG-CreER mice were hybridized to obtain 20 CAGG-CreER ppp2r1aflox/flox and 20 mice in homozygous group. Two groups of mice were divided into 4 groups respectively, finally we got 8 groups with 5 mice in each group. Tamoxifen was injected intraperitoneally to acquire inducible ppp2r1a knockout mice. The knockout efficiency of PP2A Aα in vital organs was measured by Western blot. At 0, 2, 4 and 6 days after injection, we measured body weight, histopathological change, peripheral blood cell counts and blood biochemical. Real-time PCR was performed to measure expression of liver glucolipid metabolism genes.@*Results@#After tamoxifen injection for 6 days, the knockout efficiency of PP2A Aα in vital organs was 35%, 12%, 15%, 60%, 69% and 72%, respectively in heart, liver, spleen, lung, kidney and brain. After tamoxifen injection for 6 days, the weight of homozygous mice was lower than that of wild type mice, with values of (17.42±1.76) g and (21.69±1.82) g, respectively (P<0.05). Moreover, the activity level, abdominal and renal fat were significantly decreased in homozygous mice. Homozygous mice survived no more than 7 days. Compared with wild type mice, the organ coefficient of spleen of homozygous mice was decreased at the 6th day, with values of (0.59±0.10)% and (0.36±0.05)% respectively (P<0.05). Obvious spleen atrophy and marked decrease of nucleated cells were showed by performing HE staining. Tunel staining revealed increased apoptosis ratio of splenic lymphocytes in homozygous mice. The levels of alanine aminotransferase (ALT) and aspartate transaminase (AST) of homozygous mice were higher than wild type mice (P<0.05). The values of ALT and AST in homozygous mice were (153.68±62.80) U/L and (193.2±44.28) U/L. The corresponding values in wild type mice were (41.02±12.91) U/L and (69.40±9.55) U/L. The above results indicated that ppp2r1a knockout caused liver damage. Blood sugar level of homozygous mice was lower than in wild type mice (P<0.05), with values of (4.20±1.99) mmol/L and (8.88±0.65) mmol/L respectively. Plasma total cholesterol (TC), high density lipoprotein (HDL) and β-hydroxybutyric acid (β-HB) level of homozygous mice were higher than those of wild type mice (P<0.05). The values of TC, HDL and β-HB in homozygous mice were (3.12±0.39), (1.53±0.38) and (2.49±0.89) mmol/L. The corresponding values in wild type mice were (1.69±0.92), (0.78±0.50) and (0.45±0.30) mmol/L respectively. The above results indicated that ppp2r1a loss interfered glucose and cholesterol metabolism. In addition, we also found that the white blood cell count (WBC) and lymphocyte count (LYM) of homozygous mice were lower than in wild type mice (P<0.05). The values of WBC and LYM in homozygous mice were (1.88±0.89)×109/L and (0.92±0.37)×109/L respectively. The corresponding values in wild type mice were (3.91±0.80)×109/L and (2.74±0.52)×109/L respectively. The mRNA levels of glucose-6-phosphatase (G6P) and phosphoenolpyruvate carboxykinase (PEPCK) of homozygous were lower than wild type mice (P<0.05). The fold change of G6P and PEPCK in homozygous mice was 0.46±0.11 and 0.72±0.07 respectively. The corresponding fold change in wild type mice was 1.02±0.07 and 1.02±0.06 respectively.@*Conclusion@#Whole body ppp2r1a is essential for the survival of adult mice, due to the important role in maintaining the metabolism of glucose and cholesterol of liver.
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Acute infection with hepatitis C virus is generally asymptomatic. Most patients can not clear the virus and are easy to develop into chronic infection, while a small number of patients even develop into cirrhosis, or liver cancer. The traditional treatment is combined pegylated interferon with ribavirin, but the antiviral effect is poor and the adverse reactions are large, which may be related to the ability of HCV to escape the host′s immune response through various mechanisms. The innate immune response is the first line of defense against pathogenic microorganisms. Retinoic acid inducible geneⅠ(RIG-Ⅰ) plays an important role in identifying HCV virus and initiating immune response as a pattern recognition receptor.
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High mobility group box 1 protein,a kind of damage-associated molecular patterns,is a group of nonhistone DNA-binding protein in the nuclear.It not only involves in DNA replication,recombination,repair,regulation of gene transcription,but also induces activation of innate immunity and T cell generation and activates the adaptive immune response through Toll-like receptor.It is also more closely related to autoimmune diseases,inflammatory diseases,tumors and so on.Currently there are many researches on the role of TLR-mediated HMGB1 signaling pathway in diseases,which is involved in a variety of pathological processes and has a broad clinical outlook.
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Objective To detect mRNA expressions of STING and type Ⅰ interferons(IFN-α and IFN-β) in peripheral blood mononuclear ceils (PBMCs) of patients with chronic hepatitis C(CHC).Methods 113 patients with CHC and 94 healthy controls were collected from Renmin Hospital of Wuhan University during January 2015 and January 2016.Expressions of mRNA of STING,RIG-1,IFN-α and IFN-β were detected by real-time quantitative PCR,and their relative expression values were obtained by 2-△△Ct method and the results were statistically analyzed.Results The expression of mRNA of STING,RIG-1,IFN-α and IFN-β in CHC were 0.018,0.361,0.578 and 0.573 times than its in healthy controls respectively and the differences were statistically significant (t-8.28,4.26,2.18 and 2.07,all P ＜ 0.05).Pearson correlation analysis showed that mRNA expressions of STING in healthy controls were positively correlated with that of IFN-α mRNA and IFN-β mRNA (r=0.487 and 0.207,all P＜0.05),which had no correlation in CHC (r=0.174 and 0.091,all P＞0.05).The expressions of STING mRNA was positively correlated with that of RIG-1 mRNA in healthycontrols (r=0.222,P＜0.05),but there was no correlation of expression in CHC between STING mRNA and RIG-1 mRNA (r=-0.029,P＞0.05).Conclusion Compared with healthy controls,the expression of STING,RIG-1,IFN-α and IFN-β mRNA were all reduced.There was positive correlation between STING and RIG-1,IFN-α and IFN-β in healthy controls,but no correlation in CHC.
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Objective@#To investigate the effect of polycyclic aromatic hydrocarbons (PAHs) exposure on the level of histone H3Ser10 phosphorylation (p-H3S10) and DNA damage degree in peripheral blood lymphocyte (PBLCs).@*Method@#75 coke oven workers from Benxi steel plant in Liaoning Province of China (PAHs-exposed group) and local 50 hot rolling workers (control group) were recruited in this study with age, working years, labor intensity and high temperature for matching factors using cluster sampling method in 2014. HPLC-fluorescence was performed to determine the level of urinary 1-hydroxypyrene (1-OHP), DNA damage and specific histone modification were measured in PBLCs of the subjects through comet assay and ELISA assay, respectively. Linear regression model analysis was used to analyze the differences among PAHs exposure, DNA damage and p-H3S10 level in two groups. The Mediation analysis was used to analyze the regulated relationships between urinary 1-OHP, DNA damage and histone modification through the bootstrap method.@*Results@#Age of the control and the exposed group were (45.32±8.32) and (43.87±5.67) years old (P=0.284). The concentration of urinary 1-OHP, OTM value, Tail DNA% and p-H3S10 level in exposure group were higher than that in control group, while the M (P5-P95) of p-H3S10 levels in control and exposed group were 2.21 (0.68-4.71), 4.54 (1.85-23.91) (P<0.001). The degree p-H3S10 level was increased after the subgroups which were (2.59±1.19)%, (3.24±2.81)%, (5.55±3.25)%, (8.77±7.84)%, respectively, divided by quantitated 1-OHP concentration as P0-P25, P26-P50, P51-P75 and P76-P100 (P<0.001). We also found the correlations between urinary 1-OHP and p-H3S10 level or OTM value or Tail DNA%, β (95%CI) were 0.264 (0.167-0.360), 0.500 (0.299-0.702), and 0.510 (0.384-0.671), respectively (P<0.001). Similar result was also observed between p-H3S10 level and OTM value or Tail DNA%, β (95%CI) were 0.149 (0.073-0.226) and 0.220 (0.132-0.308) (P<0.001). Moreover, the mediation effect value of DNA damage on PAHs induced p-H3S10 alteration was 0.054(P=0.040).@*Conclusion@#The results suggested that PAHs exposure could induce DNA damage and an increase in histone H3Ser10 phosphorylation in PBLCs. Particularly, the alteration of H3S10 phosphorylation may play an important role in regulating cell DNA damage repair.
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Objective To detect the expression of retinoic acid-inducible gene Ⅰ ( RIG-Ⅰ) in peripheral blood mononuclear cells (PBMCs) of patients with chronic hepatitis C (CHC), and to explore its correlations with type Ⅰinterferon gene expression and serum level of hepatitis C virus .Methods A total of 101 na?ve patients with CHC were enrolled from Renmin Hospital of Wuhan University during July 2014 and July 2015, and 69 healthy subjects served as controls .The expressions of RIG-Ⅰ, IFNαand IFNβmRNA in PBMCs and serum level of HCV RNA were detected by real-time quantitative polymerase chain reaction.Pearson correlation analysis was performed to investigate the correlations of RIG-Ⅰ mRNA expression with IFNα/βmRNA expression and serum level of HCV RNA .Results The relative expressions of RIG-Ⅰ, IFNα, IFNβmRNA to healthy controls in CHC group were 0.082, 0.022 and 0.156, respectively (t=7.83, 3.65 and 2.13, all P0.05).Conclusion Expression of RIG-ⅠmRNA decreases in patients with CHC , and is positively correlated with that of IFNαand IFNβmRNA.
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Hepatitis C virus (HCV)was easily developed into hepatitis,cirrhosis and liver cancer after infecting human.Cur-rently,the traditional method of treatment of HCV was pegylated interferon and ribavirin program,which was ineffective and had poor side effects,while the new direct antiviral drugs were expensive.Therefore,looking for cheap and non-toxic side effects of treatment was the focus of current research.More and more evidence indicate that micro-Ribose Nucleic Acid (miRNAs)play an important role in the development of liver disease,regeneration and functional regulation.This review studies the relationship between miR-122 and Hepatitis C virus and hepatocellular carcinoma.
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Objective To investigate mRNA expressions of Toll-like receptors (TLR3 and TLR7)and type Ⅰ interferons (IFN-α and IFN-β) in peripheral blood mononuclear cells (PBMCs) of patients with chronic hepatitis C (CHC).Methods Thirty patients with CHC and 30 healthy controls were collected from Renmin Hospital of Wuhan University during September 2013 and May 2014.Liver function was detected using enzyme-linked immunosorbent assay (ELISA).mRNA expressions of TLR3,TLR7,IFN-α and IFN-β were detected by real-time quantitative PCR,and their relative expression values were obtained by 2-△△Ctmethod.The t test was performed for the comparison of mean differences between CHC patients and healthy controls,and Pearson analysis was used to analyze the correlations between TLR mRNA and IFN mRNA,liver function,HCV RNA load.Results Taking mRNA expressions in healthy control group as 1,the relative mRNA expressions of TLR3,TLR7,IFN-α and IFN-β in CHC group were 0.086,0.633,0.145 and 0.423 respectively (t =4.25,2.39,2.37 and 2.80,all P ＜ 0.01).Pearson correlation analysis showed that mRNA expressions of TLR3 and TLR7 were positively correlated with that of IFN-β (r =0.381 and 0.487,all P ＜ 0.05),but not correlated with IFN-α mRNA expression,HCV RNA load,ALT and AST (all P ＞ 0.05).Conclusion The mRNA expressions of TLR3,TLR7 and IFN-α,IFN-β decrease in CHC patients,and the mRNA expressions of TLR3 and TLR7 are positively correlated with that of IFN-β,indicating that increasing the expression of TLRs might be a new strategy in CHC treatment.