Background Recombinant hirudin variant Ⅲ(rHV3) can effectively prevent galactose-induced human lens epithelial cells LECs injury,but little is known about the molecular mechanism of its action.Objective The present study was to investigate the effects of rHV3 on the expression of apoptosis-related genes in damaged LECs induced by galactose.Methods The rHV3 was extracted by our research group,and the biological activity of rHV3 was identified by titration of thrombase according to Markwardt's method.Human LECs (SRA01/04) were cultured using 125×10-3 mol/L D-galactose+10% FBS+D/F12 medium to establish the damaged human LECs model.rHV3 was added into the medium of the damaged human LECs model.Human LECs were cultured in D/F12 medium containing 10% FBS as normal control.The expression of apoptosis-related genes,such as aldose reductase (AR),bax,bcl2 and p53,in LECs at the mRNA level was detected using RT-PCR.The abundance ratio of target genes was presented with the absorbance (A) of gene mRNA/GAPDH mRNA.Results Compared to the normal control group,the A values of AR mRNA/GAPDH mRNA,bax mRNA/GAPDH mRNA and p53 mRNA/GAPDH mRNA were significantly elevated in model group (t=3.90E-06,t=8.44E-04,t=5.15E-08,P＜0.01).However,in the rHV3-treated group,the A values of AR mRNA/GAPDH mRNA,bax mRNA/GAPDH mRNA and p53 mRNA/GAPDH mRNA were lower than those of model group (t=5.90E-06,t=1.51E-04,t=3.42E-06,P＜0.01).The bcl2 mRNA/GAPDH mRNA was markedly downregulated in the model group when compared with the normal control group (t=1.86E-05,P＜0.01);while after rHV3 addition,bcl2 mRNA/GAPDH mRNA increased in comparison with the model group (t=8.56E-05,P＜0.01).Conclusion 125×10-3mol/L D-galactose induces the damage and apoptosis of human LECs.rHV3 likely plays a protective function on D-galactose-induced damage of human LECs by inhibiting the polyol pathway and mitochondria-mediated pathway.

Objective To quantitatively analyze total hepatitis B virus (HBV) DNA (HBV tDNA),covalently closed circular DNA (cccDNA) and HBsAg in patients with chronic hepatitis B (CHB),HBV-related liver cirrhosis (LC) and hepatocellular carcinoma (HCC),and to analyze the characteristics.Methods HBV tDNA and HBV cccDNA in the serum and liver biopsy samples were measured in 21 CHB,23 LC and 25 HCC patients by real-time polymerase chain reaction (PCR) assay. HBsAg titer was measured by chemiluminescence. Normally distributed variables among multiple groups were analyzed by ANOVA and t-test.Correlation between two variables was tested using Pearson correlation analysis.Skewed distribution was tested using Rank sum test.Results In CHB,LC and HCC patients,the serum HBV tDNA levels were (5.38±2.08),(4.96± 1.65) and (4.18 ± 0.91) lg copy/mL,respectively; the intrahepatic HBV tDNA levels in three groups were (7.18±1.91),(6.51±1.87) and (5.87± 1.47) lg copy/ug,respectively; the intrahepatic HBV cccDNA levels were (3.53±2.03),(2.63±2.13) and (0.58± 1.40) lg copy/μg,respectively; the serum HBsAg levels were (3.30±0.65),(3.12±0.52) and (2.60± 1.03) lg IU/mL,respectively.In CHB patients,the serum HBV tDNA,intrahepatic HBV tDNA,HBV cccDNA and HBsAg levels were all significantly higher than those of HCC patients (t=2.446,P=0.013; t=2.562,P=0.014;t=5.799,P＜0.01 ; t=2.709,P=0.003,respectively).However,only intrahepatic HBV cccDNA and HBsAg levels were statistically different between LC and HCC patients (t=-3.894,P＜0.01;t=-2.237,P=0.023,respectively).HBV cccDNA was all negative in the serum of 69 patients.The serum HBsAg level was positively correlated with serum HBV tDNA (r=0.290,P=0.016),intrahepatic HBV tDNA (r=0.372,P =0.002) and intrahepatic HBV cccDNA (r=0.378,P=0.001).Conclusions The levels of HBV tDNA,HBV cccDNA and HBsAg decrease gradually with the disease progression.The serum HBsAg level is positively correlated with serum HBV tDNA,intrahepatic HBV tDNA and intrahepatic HBV cccDNA.

BACKGROUND: The association of genetic variation in the IRAK-1 gene with sepsis outcome has been proved. However, few studies have addressed the impact of the IRAK-4 gene variants on sepsis risk. This study aimed to determine whether the polymorphisms in the IRAK-4 gene are associated with susceptibility to and prognosis of severe sepsis in the Chinese Han ethnic population.METHODS: In this case-control study, 192 patients with severe sepsis hospitalized in the emergency department of Zhongshan Hospital from February 2006 to December 2009 and 192 healthy volunteers were enrolled. Exclusion criteria included metastatic tumors, autoimmune diseases, AIDS or treatment with immunosuppressive drugs. This study was approved by the ethical committee of Zhongshan Hospital, Fudan University. Sepsis patients were divided into a survival group (n=124) and a non-survival group (n=68) according to the 30-day mortality. Primer 3 software was used to design PCR and sequencing primers. Genomic DNA was extracted from peripheral blood mononuclear cells. Seven tagSNPs in IRAK-4 were selected according to the data of the Chinese Han population in Beijing from the Hapmap project and genotyped by direct sequencing. The chi-square test was used to evaluate the differences in genotype and allele frequencies between the two groups.RESULTS: The distributions of all tagSNPs were consistent with Hardy-Weinberg equilibrium. The allele and genotype frequencies of rs4251545 (G/A) were significantly different between the severe sepsis and healthy control groups (P=0.015, P=0.035, respectively). Carriers of the rs4251545A had a higher risk for severe sepsis compared with carriers of the rs4251545G (OR=1.69, 95％ CI: 1.10-2.58). The allele and genotype frequencies of all SNPs were not significantly different between the survival group and non-survival group.CONCLUSION: These findings indicate that the variants in IRAK-4 are significantly associated with susceptibility to severe sepsis in the Chinese Han ethnic population.

OBJECTIVETo detect the levels of vascular endothelial growth factor (VEGF) and pigment epithelium-derived factor (PEDF) in aqueous humor of patients with active choroidal neovascularization (CNV).

METHODSAqueous humor samples were obtained from 32 patients with active CNV. The concentrations of VEGF and PEDF in aqueous humor were measured by enzyme linked immunosorbent assay (ELISA) for quantitative analysis. VEGF and PEDF in 10 samples of aqueous humor from patients with cataract were also detected by the same methods as control.

RESULTThe mean VEGF and PEDF concentrations in aqueous humor of active CNV patients were higher than those in the control group (P=0.000).

CONCLUSIONThe patients with active CNV exhibit significantly higher VEGF and PEDF levels than those in control, indicating that VEGF along with PEDF may modulate the formation of CNV.

Adult ; Aged ; Aged, 80 and over ; Aqueous Humor ; chemistry ; Choroidal Neovascularization ; metabolism ; Eye Proteins ; analysis ; Female ; Humans ; Male ; Middle Aged ; Nerve Growth Factors ; analysis ; Serpins ; analysis ; Vascular Endothelial Growth Factor A ; analysis

Apoptotic protease activating factor-1 (Apaf1) is an essential factor in intrinsic mitochondrial pathway of apoptosis activation. Apaf1 leads to the formation of apoptosome, which then proteolytically activates caspase-9. The activated caspase-9 opens the downstream signal of caspases to execute programmed cell death. Apaf-1 is important for tumor suppression and drug resistance because it plays a central role in DNA damage-induced apoptosis. Inactivation of the Apaf-1 gene is implicated in disease progression and chemoresistance of some malignancies. Further research on the Apaf-1 will contribute to develop a new type of approach to anti-cancer drugs, which might have good prospect in clinical practice. In this paper, the structure and function of Apaf-1, the mechanism involved in Apaf-1 signaling pathway, and application of Apaf-1 in tumor therapy were reviewed.
Apoptosis ; Apoptotic Protease-Activating Factor 1 ; metabolism ; Caspase 9 ; metabolism ; Humans ; Neoplasms ; therapy ; Signal Transduction

By discussing different ways on prediction, prevention and treatment of femur head necrosis (FHN), to provide a theory reference for future clinical application. By searching, reading and summarizing related-literatures through CNKI, VIP, CBM and foreign-related literature, to sum up the relevance methods and techniques of currently used. It was found that imaging study (especially X-ray and MRI) was of great importance in prediction. There were several ways (both non-surgical or surgical) for prevention and treatment. FHN collapse was affected by many factors, but most of the researches were all focus on one aspect of the mechanism and based on small samples. It is necessary to have a research with a large sample and to compare the effect on different treatment. Early and effective imaging inspection is needed for high risk group of FHN; possibility of collapse should be predicted for existing FHN; core decompression or vascularized bone grafting are required for collapse of high risk group. Early prediction and treatment are essential for FHN patients.
Bone Transplantation ; Decompression, Surgical ; Femur Head Necrosis ; complications ; diagnostic imaging ; surgery ; Humans ; Magnetic Resonance Imaging ; Tomography, X-Ray Computed

Objective To explore the effect of associated deficiency of selenium,protein and vitamin E(VE)on the thyroid iniury and thyroid hormone metabolism of the rats in a long-term.Methods The Wistar rats were randomly divided into four groups:Group A with selenium deficiency and low protein and VE;Group B with selenium deficiency,low protein but adequate VE;Group C,adequate selenium and protein but low VE;Group D,adequate selenium.protein and VE.The rats were killed at the end of 26th week.Glutathione peroxidase(GSH-Px)activity in the rat blood and type I 5'-deiodinase activity of the rat liver were determined.The content of triiodothyronine(T3),tetraiodothyronine(T4),thyrotropic-stimulating hormone(TSH),activated oxygen(ROS)and malonaldehyde(MDA)were detected in serum. The changes of thyroid histopathology were observed under light microscope.Results ①The interactive effect of selenium+protein and VE was not significant on GSH-Px and ID I activity(F=0.003,0.871,P＞0.05),but it was significant on MDA and ROS content(F=13.057,6.706,P＜0.05 or＜0.01). ②Selenium+protein and VE could influence T3 and T4 content(F=431.977,28.271,6.570,41.419,P＜0.05).The interactive effect of selenium±protein and VE was not significant on T3 and T4 content(F=0.871,0.136,P＞0.05).Whether in the condition of low selenium and protein or supplementary,T4 contents of supplementary VE group[(79.095±12.199),(64.392±6.261)μg/L]were respectively higher than the low VE group[(61.068±6.648),(44.176±7.090)μg/L],the difference being statistically significant(t = 3.670,6.045, P ＜ 0.01). In the condition of low VE, T3[(0.718 ± 0.079)μg/L] and T4[(44.176 ±7.090)μg/L] content of supplementary selenium and protein group was lower than that in the low selenium and protein group[ (0.966 ± 0.156), (61.068 ± 6.648)μg/L], the difference being statistically significant (t = 4.568,4.916, P ＜0.01 ). With supplementary VE, T4 content of supplementary selenium and protein group[ (64.392 ± 6.261 )μg/L] was lower than that in the low selenium and protein group [(79.095 ± 12.199)μg/L], the difference being statistically significant (t = 3.033, P ＜ 0.01 ). ③Degeneration and necrosis of follicular epithelial cell were induced by diet of low selenium, protein and VE, which could be relieved by supplymentary VE. The sparseness of intracavitary glue was observed occationally in the supplementary selenium and protein but low VE group. Conclusions Long-term deficiency of selenium, protein and VE results in the decrease of the selenoenzymes of rats, which causes accumulation of the oxidative products, as well as thyroid pathological injury and thyroid hormone metabolism disorder, but supplement of adequate VE can reduce the oxidative damage in rats having low selenium and protein diet.

Objective In this study,we investigated the relationship between oxidative stress,selenoproteins level and onset of Keshan disease (KD) through detecting the expression of 8-hydroxy-2-deoxy guanosine (8-OH-dG),thioredoxin reductase 1 (TrxR1) and glutathione peroxidase 1 (GPx1) in myocardial tissue.Methods Myocardium samples of autopsy patients including 8 cases of KD (KD group included 4 acute KD and 4 chronic KD) and 9 cases of non-KD (control group) were immunohistochemically stained for 8-OH-dG,TrxR1 and GPx1.The staining intensities subsequently quantified by using Olympus Image-Pro Plus 6.0 software.Results The positive rate of 8-OH-dG expression in myocardial nuclei was higher in the case group[(68.6 ± 20.4)％] than that of the control group[(2.4 ± 1.5)％,t =8.515,P ＜ 0.05].In addition,the positive rate of 8-OH-dG expression in acute KD[(91.7 ± 3.7)％] was significantly higher than that of chronic KD[(53.2 ± 7.9)％,t =6.409,P＜0.05].The distribution of TrxR1 and GPx1 was not associated with the distribution of myocardial damage.The expression of these two selenoproteins in KD group (401340 ± 59865,497590 ± 197082) were both lower than that of control group(2790300 ± 379298,1348400 ±615840; t =-28.493,-6.016,respectively,all P＜0.01).Conclusions Oxidative damage is detected in myocardium tissue of KD,and 8-OH-dG expression is associated with the degree of myocardial damage in KD.Selenoproteins,TrxR1 and GPx1,may be closely related to the pathogenesis of KD.

OBJECTIVETo study the correlation between polyoma virus load and hemorrhagic cystitis after allogeneic stem cells transplantation for prevention of hemorrhagic cystitis.

METHODSBlood and urine specimens were collected from 40 healthy persons, 40 patient with stem cells transplantation and 20 cases complicated with hemorrhagic cystitis for determination of VP1 gene of polyomaviruses BK virus (BKV)/Jamestown Canyon virus (JCV) and simian virus 40 (SV40) by polymerase chain reaction (PCR) and EvaGreen stain fluorescence quantitative assay.

RESULTSIn the peripheral blood, all genes of BKV/JCV and SV40 were negative, while BKV gene in urine and blood from healthy persons and patient with stem cells transplantation was 15% (6/40) and 100% (40/40), respectively. The gene of JCV was positive in 10% (4/40) and 12% (5/40), the gene of SV40 was negative.

CONCLUSIONGenes of BKV and JCV was detectable in urine specimens of healthy persons and there was a correlation between the load of polyomavirus and incidence of hemorrhagic cystitis.

Capsid Proteins ; genetics ; Cystitis ; diagnosis ; etiology ; virology ; DNA, Viral ; blood ; genetics ; urine ; Hematopoietic Stem Cell Transplantation ; adverse effects ; Hemorrhage ; diagnosis ; etiology ; virology ; Humans ; Polymerase Chain Reaction ; methods ; Polyomavirus ; genetics ; growth & development ; Polyomavirus Infections ; complications ; virology ; Viral Load

OBJECTIVETo explore the correlation between the polyomavirus DNA load and the dose of immunosuppressant in patients with allogene bone marrow transplantation (allo-BMT) for preventing the development of post-transplantational hemorrhagic cystitis.

METHODSSerial blood and urine samples from 122 cases of allo-BMT recipients were obtained and DNA was extracted from urine samples. Polyomavirus DNA-specific probe was synthesized and Fluorescence quantitative polymerase chain reaction was used for detecting the polyomavirus DNA loads and Fluorescence polarization immunoassay (FPIA) was performed for determining the dose of immunosuppressant Cyclosporin A (CsA) in blood.

RESULTSThe altered polyomavirus DNA load in urine was followed by concentration of CsA in blood. When the concentration of CsA in blood was higher than 86-105 ng/ml, the positive rate of polyomavirus DNA load was significantly increased and both presented the linable correlation.

CONCLUSIONIn immunosuppression condition, polyomavirus DNA load correlated to the dose of immunosuppressant, which increased the risk of post-transplantational hemorrhagic cystitis.

Bone Marrow Transplantation ; adverse effects ; Cyclosporine ; adverse effects ; blood ; Cystitis ; prevention & control ; virology ; DNA, Viral ; genetics ; urine ; Dose-Response Relationship, Drug ; Humans ; Immunosuppressive Agents ; adverse effects ; blood ; Polyomavirus ; genetics ; isolation & purification ; Polyomavirus Infections ; prevention & control ; virology ; Transplantation, Homologous ; adverse effects ; Viral Load