In recent years, cytokinesis-block method was used to analyze cytomics indicators including micronucleus, nuclear bridge, nuclear bud, nuclear division index, cell apoptosis and cell necrosis. In public health, it has become the common method to explore the impacts of different population structure, environment and occupational exposure for genomic instability, chromosome breakage, chromosome loss and cell proliferation. This article reviews and discusses the application of using cytokinesis block method to analyze cytomics indicators in public health field.

Objective:To investigate the role of CD46 and Nectin-4 on Measles virus (MV) infecting human pulmonary alveolar epithelial cells (HPAEpiC),and the interaction between CD46 and Nectin-4.Methods: Measles virus was divided into pre-infection group and 2 h-infection group,HPAEpiCs treatment with anti-CD46 antibody and/or anti-Nectin-4 antibody as experimental groups,and untreated HPAEpiCs as a control.The variation of viral replication level was detected.A Co-immunoprecipitation assay (Co-IP) was used to explore whether CD46 and Nectin-4 had interactive relationship in MV infection.Results: Compared with the control group,MV titers were reduced in HPAEpiCs of the pre-infection group treated with anti-CD46 and anti-Nectin-4 respectively (48.03% and 49.53%).Furthermore,virus titers showed a more reduction in which treated with anti-CD46 and anti-Nectin-4 antibodies (27.15%,P<0.01).Western blot and Real-time PCR showed that anti-CD46 antibody and anti-Nectin-4 antibodies decreased the rate of MV infection.In the 2 h-infection group,however,the treatment with anti-CD46 and anti-Nectin-4 could significantly reduce the MV titer and NP protein in HPAEpiCs.The Co-IP assay showed that there were interaction between CD46 and Nectin-4.Conclusion: CD46 and Nectin-4 mediated MV infecting HPAEpiCs.Moreover,CD46 and Nectin-4 may play a synergetic role in MV infection,which could enhance the infection effect.

Objective:To explore the feasibility of the optimized cytokinesis-block (CB) assay on radiation-induced nucleoplasmic bridge (NPB), and to provide a scientific basis for the application of NPB in biological dose estimation.Methods:Human peripheral blood in vitro was irradiated with 2 Gy 60Co γ-rays at a dose rate of 1 Gy/min (0 Gy control group). According to the culture time after irradiation, blood samples were divided into group 48, 56, 68 and 72 h. Cytochalasin-B (Cyt-B) with a concentration of 6 μg/ml was added into the samples at 28 h and harvested at 48, 56, 68 and 72 h after irradiation, respectively. On the other hand, the blood samples were treated with different concentration of Cyt-B i. e., 0.6, 1, 2, 6 and 10 μg/ml at the beginning of culture (0 h) and harvested at 68 h after irradiation. The proportion of mononucleated, binucleated and multinucleated cells, radiation-induced NPB and micronucleus (MN) frequencies were analyzed. Results:The nuclear division index (NDI) and proportion of binucleated cells at 2 Gy and 0 Gy had tendency of increasing with cell culture time. NPB frequencies (0.023 0-0.033 0/cell) and MN frequencies had no significantly difference ( P> 0.05). With the increase of Cyt-B concentration, NDI and the proportion of binucleated cells in group 2 Gy and 0 Gy also increased, but NPB frequencies (0.023 0-0.047 0/cell) had no significant difference ( P> 0.05). MN frequencies of group 10 μg/ml were significantly lower than that of group 6 μg/ml ( U=2.74, P< 0.01). Conclusions:Cell culture time and Cyt-B concentration had no significant influence on radiation-induced NPB frequencies, suggesting that NPB could be obtained by appropriately reducing cell culture time and Cyt-B could be added into blood samples at the beginning of culture. But this protocol reduced the number of cells for further analysis, and thus its feasibility for dose estimation still need to be studied.

Objective To explore the influences of the final concentration and adding time of Cytochalasin-B (Cyt-B) on radiation-induced nucleoplasmic bridges (NPB) in cytokinesis-block assay.Methods Hunan peripheral blood samples were divided into 5 final concentration groups (group 2,4,6,8,10 μg/ml) according to different final concentrations of Cyt-B.Moreover,blood samples were divided into 4 adding time groups (group 0,28,40,44 h) according to different adding times of Cyt-B.Blood samples were irradiated with 0 (sham irradiation) and 2 Gy 60Co-rays in vitro,at a dose rate of 1 Gy/min.A cytokinesis-block assay was carried out to prepare NPB samples.The percentages of mononucleated,binucleated and multinucleated cells,as well as the frequencies of NPB and micronucleus (MN) in binucleated cells were analyzed using an optical microscope.Results Nuclear division index (NDI) and the percentages of binucleated cells increased with increased concentration of Cyt-B,and decreased with delayed adding time of Cyt-B (except group 0 h) in both final concentration groups and adding time groups.After exposed to 2 Gy,NPB frequencies were no significant difference (except group 0 h).MN frequencies had the trend of decreased with the increased concentration of Cyt-B,but no significant difference with adding time of Cyt-B.Conclusions In cytokinesis-block assay,different final concentration and adding time of Cyt-B may induce to the variation of NPB frequencies,but there was no significant difference.Appropriate increased final concentration or ahead adding time of Cyt-B can increase the percentage of binucleated cells that help to improve the efficiency of analysis.

Objective:To screen radiosensitive lipid metabolites in rat small intestine and analyze their metabolic pathways, in order to provide scientific basis for radiation enteropathy biomarkers.Methods:The total body irradiation of 60Co γ rays was performed to rats with different doses of 0, 1, 2, 3, 5 and 8 Gy. The changes of lipids in small intestine were studied by targeted lipidomics method based on liquid chromatography coupled mass spectrometry (LC-MS). Results:Fifteen lipids in small intestine were screened as radiosensitive metabolites at 3 d after irradiation, including 4 up-regulated lipids and 11 down-regulated lipids( t=-6.395, 5.998, 5.836, -5.503, -5.449, -5.422, 4.841, 4.802, 4.621, 4.457, 4.426, 4.373, 4.110, 3.945, 3.902, P< 0.05 and FDR < 0.05). The metabolic pathways of sphingolipid, glycerophosphoplipid were significantly enriched. Four phosphatidyl serines (PS)increased while 1 phosphatidic acid(PA), 2 sphingomyelins(SM) and 4 fatty acids(FA)decreased in a good dose-response manner( R2> 0.80, P< 0.05), which were more potential radiation enteropathy biomarkers. Conclusions:Lipid metabolites in rat small intestine were significantly changed after the rat was total body irradiated with 60Co γ-rays.Eleven lipids with good dose-response relationship were more potential to be radiation enteropathy biomarkers.

OBJECTIVETo investigate the association of the polymorphisms of cytochrome P450 2C9 (CYP2C9) exon 4 608T/G, 561A/C, 537A/C and 527A/C, and -65G/C with warfarin sensitivity.

METHODSA total of 102 patients under warfarin anticoagulant therapy were selected. During follow-up, warfarin dosage and associated Prothrombin Time-International Normalized Ratio (P-INR) values were recorded. Simultaneous monitoring of incidence of bleeding and thrombosis adverse effect was recommended. Genetic polymorphisms of the above mentioned loci were identified by polymerase chain reaction and DNA sequencing.

RESULTSThe average age of the 102 patients was (62.1+/-10.5) years. The body mass index (BMI) was (24.7+/-3.8) kg/m2. Mean daily warfarin requirement was from 1.250 to 5.077 mg/day when therapeutic PT-INR (1.5-2.5) was maintained. DNA sequencing showed no polymorphisms of 608T/G, 561A/C, 537A/C, 527A/C in CYP2C9 exon 4. Warfarin daily dosage in CYP2C9 exon 4 -65C carriers was 3.106+/-0.619 mg/d, while it was (2.555+/-0.708) mg/d in individuals with wild-type -65G (P=0.020). Receiver operating characteristic (ROC) analysis showed that warfarin daily dosage of more than 2.5 mg/d can be used to predict the CYP2C9 exon 4 -65GC genotype (AUC: 0.770, P=0.005, 95%CI:0.626-0.915). Logistic regression indicated that BMI was an independent factor of bleeding during anti-coagulation therapy (OR=0.794, 95%CI: 0.651-0.970, P=0.024).

CONCLUSIONThe Chinese population are, generally, warfarin-sensitive. Exon 4 of the CYP2C9 gene is highly conserved in this population. The warfarin maintenance dosage in CYP2C9 exon 4 -65CG carriers was significantly higher than those with wild-type -65GG. The clinical significance needs further investigation with more large-scale, multi-center trials.

Adult ; Alleles ; Anticoagulants ; pharmacology ; therapeutic use ; Aryl Hydrocarbon Hydroxylases ; genetics ; Cytochrome P-450 CYP2C9 ; Exons ; genetics ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Male ; Middle Aged ; Point Mutation ; Polymorphism, Genetic ; Thrombosis ; drug therapy ; Warfarin ; pharmacology ; therapeutic use

Objective:To summarize the relationship between the clinicopathological features and prognosis of immunoglobulin A nephropathy (IgAN) after renal transplantation.Methods:A total of 34 patients with IgAN after renal transplantation confirmed by renal biopsy were enrolled. And another 34 patients with primary IgAN confirmed by initial renal biopsy were adopted as controls. Clinical and pathological features of two groups were compared to explore the relationship between clinicopathological features and prognosis of allograft IgAN.Results:As compared with primary IgAN group, renal function in allograft IgAN group included serum creatinine [(158.5±75.9) vs (84.8±26.8) umol/L], urea nitrogen [(9.7±6.1) vs (5.2±1.4) mmol/L], uric acid [(406.7±87.8) vs (359.0±92.6) umol/L], estimated glomerular filtration rate {(57.4±25.4) vs (91.2±28.6) [ml/(min·1.73m 2)]}. All were statistically significantly higher ( P<0.05) while other parameters showed no differences. Pathologically, the proportion of T1 type (50.0% vs 17.6%) of renal tubular atrophy/interstitial fibrosis was significantly higher in allograft IgAN group than control group ( P<0.05). Furthermore, univariate and multivariate Logistic regression analyses were performed between various pathological parameters and prognosis in allograft IgAN patients. It indicated that the degree of mesangial hyperplasia of patients with transplanted IgAN had a significantly negative impact on the prognosis. Conclusions:The clinicopathological features of patients with allograft IgAN show no difference from those of patients with primary IgAN. And among patients with allograft IgAN, those with severe mesangial hyperplasia often have a worse prognosis.

Objective:To explore the influence of dose-rate on radiation-induced gene expression in human peripheral blood.Methods:Human peripheral blood ex vivo was irradiated with 0, 1, 2, 4 and 6 Gy of 60Co γ-rays with different dose-rates of 0.2, 1 and 2 Gy/min. Human blood cells were harvested at 24 h post-irradiation. Following RNA isolation, the mRNA expression levels ofCDKN1A, MDM2, PCNA, FDXR, GADD45A, PHPT1, ASTN2, TNFSF4, POLH, GDF-15 and PPM1D were measured by quantitative real-time PCR. The stepwise regression method was used to establish the gene combination models. Results:The relative mRNA expression levels of 11 genes significantly increased in a dose-dependent manner within the dose range of 0-6 Gy with three dose-rates of irradiation ( R2=0.744-0.998, P< 0.05). Following the exposure to 2 Gy(0.2 Gy/min) 60Co γ-rays, the expression levels of CDKN1A, FDXR, PHPT1 and TNFSF4 genes were significantly higher than that of the 1 and 2 Gy/min groups ( t=3.73, 5.73, 2.44, 2.77, 3.53, 2.68, 2.43, 2.05, P< 0.05). With 6 Gy irradiation, the changes of radiation-induced PPM1D expression level under a dose rate of 2 Gy/min were significantly higher than other two dose-rates( t=3.82, 2.54, P< 0.05). The combined expression model at different dose rates was composed of 2-3 genes, and the R2values of regression equations were 0.976, 0.964 and 0.951, respectively. Conclusions:In a certain range, the dose-rate may affect the changes of radiation-induced gene expression in human peripheral blood.

Objective:To explore the changes of oxidative stress(OS),DNA damage and the occurrence of cellular premature aging of human immortalized keratinocytes(HaCaT)after that was radiated by X-ray with different doses.Methods:HaCaT cells were radiated by X-ray,and they were divided into 0 Gy group,5 Gy group and 10 Gy group according to the irradiation dose.The levels of intracellular reactive oxygen species(ROS)were detected by 2,7-Dichlorofluorescein diacetate(DCFH-DA)fluorescent probe,and the intracellular content of malondialdehyde(MDA)of lipid peroxidation products and the activity of superoxide dismutase(SOD)were measured by colorimetry.Immunofluorescence staining was used to detect the phosphorylated histone 2A variant(γ-H2AX)in HaCaT cells that were radiated by X-ray with different doses.Cell count kit-8(CCK-8)was used to detect the effect of X-ray with different doses on the proliferation of HaCaT cells after X-ray with different doses radiated them.β-Galactosidase staining was used to detect the proportion of premature aging cells.The changes of p21 and p53 protein expressions after X-ray irradiation were detected by Western blot.Results:After HaCaT cells were radiated by X-ray for 24h,the fluorescence intensity of 2',7'-Dichlorofluorescein(DCF)in 5 Gy and 10 Gy groups were significantly higher than that in the 0 Gy group,and the MDA contents of them were significantly higher than that in the control group,and the SOD activities of them were significantly lower than that in the control group(F=38.35,92.22,5.22,P<0.05),respectively.The change of γ-H2AX focus showed a dose-dependent significant increase at 1 h after irradiation,and the difference between them and control group was statistically significant(F=129.3,P<0.05).At 6h,24h and 48h after X-ray radiated HaCaT cells,the cell proliferation abilities of 5 Gy group and 10 Gy group were significantly decreased than that of 0 Gy group(F=116.41,62.20,34.29,P<0.01),and the β-Galactosidase activity of the two groups were significantly increased than that of 0 Gy group,and the difference was significant(F=1629.22,P<0.01).At 72h after X-ray with different doses radiated HaCaT cells,the expression levels of p21 and p53 proteins of 5 Gy group and 10 Gy group increased,and the differences of them among three groups were significant(F=104.4,66.69,P<0.01),respectively.Conclusion:Ionizing radiation can induce the occurrences of oxidative stress and DNA damage in HaCaT cells,and cause the occurrence of cellular premature aging.

Based on an overreview of Chinese radiological health standards, the Chinese radiological health standards currently in effect for biodosimetry were analyzed with respect to their current status, application and existing problems. Furthermore the improvement measures and development trends were put forward.