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Objective: Comparative analyses of wild-type Clostridioides difficile 630 (Cd630) strain and pathogenicity locus (PaLoc) knockout mutant (ΔPaLoc) by using RNA-seq technology. Analysis of differential expression of Cd630 wild-type strain and ΔPaLoc mutant strain and measurement of its cellular virulence changes. Lay the foundation for the construction of an toxin-attenuated vaccine strain against Clostridioides difficile. Methods: Analysis of Cd630 and ΔPaLoc mutant strains using high-throughput sequencing (RNA-seq). Clustering differentially expressed genes and screening differentially expressed genes by DESeq software. Further analysis of differential genes using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment. Finally, cytotoxicity assays of ΔPaLoc and Cd630 strains were performed in the African monkey kidney epithelial cell (Vero) and the human colonic cell (Caco-2) lines. Results: The transcriptome data showed that the ΔPaLoc mutant toxin genes tcdA and tcdB were not transcribed. Compared to the wild-type strain, CD630_36010, CD630_020910,CD630_02080 and cel genes upregulated 17.92,11.40,8.93 and 7.55 fold, respectively. Whereas the hom2 (high serine dehydrogenase), the CD630_15810 (spore-forming protein), CD630_23230 (zinc-binding dehydrogenase) and CD630_23240 (galactitol 1-phosphate 5-dehydrogenase) genes were down-regulated by 0.06, 0.075, 0.133 and 0.183 fold, respectively. The GO and KEGG enrichment analyses showed that the differentially transcribed genes in ΔPaLoc were enriched in the density-sensing system, ABC transport system, two-component system, phosphotransferase (PTS) system, and sugar metabolism pathway, as well as vancomycin resistance-related pathways. Cytotoxicity assays showed that the ΔPaLoc mutant strain lost its virulence to Vero and Caco-2 cells compared to the wild-type Cd630 strain. Conclusion: Transcriptional sequencing analysis of the Cd630 and ΔPaLoc mutant strains showed that the toxin genes were not transcribed. Those other differential genes could provide a reference for further studies on the physiological and biochemical properties of the ΔPaLoc mutant strain. Cytotoxicity assays confirmed that the ΔPaLoc mutant lost virulence to Vero and Caco-2 cells, thus laying the foundation for constructing an toxin-attenuated vaccine strain against C. difficile.
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Humanos , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/metabolismo , Células CACO-2 , Clostridioides , Clostridioides difficile/genética , Oxirredutases/metabolismo , Transcriptoma , Vacinas AtenuadasRESUMO
Objective::To study whether long-term administration of Gastrodiae Rhizoma powder can improve the learning and memory ability of APPswe/PSldE9 double transgenic (APP/PS1) Alzheimer' s disease(AD) model mice and delay the progress of AD whether these effects are related to the regulation of antioxidant stress pathway in Kelch-like epoxylopropylamine-related protein 1(Keap1)-nuclear factor E2 related factor 2 (Nrf2)/heme oxygenase(HO)-1, and further explore the neuroprotective mechanism of Gastrodiae Rhizoma powder and its role in the prevention and treatment of AD. Method::APP/PS1 double transgenic mice model, the mice consisted of five groups: normal, normal administration group, model group, Gastrodiae Rhizoma powder prevention group, Gastrodiae Rhizoma powder treatment group.The mice in the normal administration group and the Gastrodiae Rhizoma powder prevention group were given the same dose of Gastrodiae Rhizoma powder (1.5 g·kg-1) daily at the age of 8 weeks.The normal group and model group were given the same amount of normal saline at the same time, until 24 weeks old, Morris water maze was used to test the learning and memory ability of mice, and the treatment group was treated with Gastrodiae Rhizoma powder at 22 weeks old.The mice were given the same dose of Gastrodiae Rhizoma powder (1.5 g·kg-1) every day for 2 weeks.The number of crossing platform, escape latency and platform residence time of mice were detected by Morris water maze from 24 weeks old to 24 weeks old.RNA, Real-time PCR was extracted from mouse hippocampus to detect the mRNA level of Nrf2, HO-1, Keap1, and Western blot was used to detect the expression of Nrf2, HO-1, Keap1 protein in mouse hippocampus. Result::Compared with normal group, the water maze test showed that the learning and memory ability of model group was lower than that of the model group (P<0.01), and the learning and memory ability of Gastrodiae Rhizoma powder prevention group and Gastrodiae Rhizoma powder treatment group was significantly higher than that of model group (P<0.01). Compared with normal group, the levels of Nrf2, HO-1 and protein in the hippocampus in model group decreased in varying degrees (P<0.05). Compared with model group, Gastrodiae Rhizoma powder prevented Nrf2, in the hippocampus of mice in model group.The level of HO-1 in mRNA and protein increased in different degrees (P<0.05, P<0.01). Levels of Nrf2, HO-1 mRNA in Gastrodiae Rhizoma powder treatment group was significantly higher than that in Gastrodiae Rhizoma powder group (P<0.05). There was no significant difference in the expression of Nrf2, HO-1 protein.There was no significant difference in mRNA and protein levels of Keap1 among different groups. Conclusion::Morris water maze test and other results showed that Gastrodiae Rhizoma powder could improve the learning and memory ability of APP/PS1 mice, and it may enhance the expression of downstream antioxidant genes by regulating Keap1-Nrf2/HO-1 pathway.And then improve the learning and memory ability of APP/PS1 mice.
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Objective To research the prevalence model of hepatitis B virus in the minority areas of Guizhou and to provide reference for the prevention and control virus of hepatitis B virus. Methods Using multi-stage cluster simple random sampling, four villages were selected from Leishan and Libo counties in minority areas of Guizhou. Questionnaires were investigated by trained investigators and serum hepatitis B virus five-item test results were collected from the subjects. Results A total of 1 629 participants were surveyed, the outcome showed that migrant workers’ infection rate of hepatitis B was 44.8%, and the carrying rate of HBsAg was 8.4%. The positive rates of anti-HBs and anti-HBc were 28.0% and 25.6% respectively. The infection model rate of migrant workers was 19.8%, which was lower than that of non-migrant workers (23.2%) (P>0.05).The detection rate of susceptible model in migrant workers (52.2%) was higher than that in non-migrant workers (43.4%), while the detection rate of immune mode migrant workers (28.0%) was lower than that in non-migrant workers (33.4%),which the difference was statistically significant (all P<0.05). After adjusted related factors by multivariate Logistic regression analysis model, migrant workers were still the influencing factors of vulnerability model (OR=1.568, 95% CI:1.206-2.039) compared with non-migrant workers. Conclusion There was a high susceptibility to hepatitis B virus among migrant workers in minority areas of Guizhou, In order to reduce the infection and prevalence of hepatitis B virus, we should strengthen the immunization of hepatitis B vaccine to migrant workers and to improve their specific immunity.
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Objective To investigate the expressions of receptor for advanced glycation endproducts (RAGE) and nuclear factor κB(NF-κB) in brain hippocampus of rat with chronic fluorosis,and to reveal the mechanism of brain damage resulted from chronic fluorosis.Methods Sixty clean grade SD rats were randomly divided to three groups(20 rats in each group,10 female and 10 male) fed with different contents of fluoride,control group with normal tap-water(< 0.5 mg/L fluoride),small dosage of fluoride exposure group(10 mg/L fluoride in tap-water) and large dosage of fluoride exposure group(50 mg/L fluoride) for six months.Then the rats were killed by femoral artery bleeding and hippocampus was removed.Protein and mRNA levels of RAGE and NF-κB in the hippocampus were determined by Western blotting and quantitative real time PCR,respectively.Results As compared to the control groups[(100.00 ± 2.60)%,(100.00 ± 7.80)%],the expressions of RAGE and NF-κB at protein level in the hippocampus were significantly increased in the small dosage of fluoride exposure groups [(205.00 ± 15.30)%,(156.00 ± 12.20)%] and the large dosage of fluoride exposure groups[(232.00 ± 10.90)%,(162.00 ± 9.80)%,all P < 0.05]; for the mRNA level of RAGE and NF-κB,the expressions were higher in the small dosage of fluoride exposure groups(1.27 ± 0.09,0.83 ± 0.15) and the large dosage of fluoride exposure groups (2.60 ± 0.19,1.27 ± 0.19) than those of the control groups(0.66 ± 0.18,0.32 ± 0.08,all P< 0.05).Conclusions The increased expressions of RAGE and NF-κB in the hippocampus of rat brain are caused by chronic fluorosis,and these changes may be associated with the mechanism of nerve injury.
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Objective Aim of the study is to investigate the expression of syndecan-4 and nuclear factor κB(NF-κB) in the kidney of rat with chronic fluorosis,and to reveal the mechanism of kidney damage resulted from the toxicity of excessive amount of fluoride.Methods According to body mass and sex,sixty SD rats were randomly divided to three groups according to body mass and fed with different contents of fluoride:control group with normal tap-water(< 0.5 mg/L fluoride),small dosage of fluoride exposure group (adding 10 mg/L fluoride in tap-water) and large dosage of fluoride exposure group (50 mg/L fluoride) for six months.The protein level of syndecan-4 and NF-κB in the kidney was detected by Western blotting and syndecan-4 mRNA level by quantitative real time PCR.Results As compared to the control group[(100.0 + 8.1)%],the expression of syndecan-4 at protein level in the kidney of rat was significantly increased in the small dosage of fluoride exposure group [(198.5 + 5.6)%,P < 0.05] and large dosage of fluoride exposure group [(209.2 + 13.0)%,P < 0.05]; the protein levels of NF-κB in the small dosage of fluoride exposure group[(284.4 + 11.1)%,P < 0.05] and in the large dosage of fluoride exposure group[(343.2 + 2.9)%,P < 0.05] were significantly increased than that of the control group[(100.0 ± 10.7)%].The mRNA levels of syndecan-4 in the kidney in the small dosage of fluoride exposure group and large dosage of fluoride exposure group(0.431 + 0.058 and 0.453 ± 0.065,both P < 0.05,respectively) were significantly increased than that of the control(0.128 + 0.026).Conclusions The increased expression of NF-κB in the kidney is induced by increased expression of syndecan-4,which may be involved in kidney damage of chronic fluorosis.
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Objective To investigate the deletion pattern of 4.8 kb mitochondrial DNA(mito-DNA) in liver,kidney,and brain of rats with chronic fluorosis and to explore the significance of mitochondria in the pathogenesis of chronic fluorosis.Methods Sixty SD rats were randomly divided into 3 groups according to body mass (20 in each group):control,low-fluoride and high-fluoride groups,and they were fed with different concentrations of fluoride in drinking water (0,10,50 mg/L,respectively) for 6 months.Mito-DNA in liver,kidney and brain was detected by real-time PCR.Results The amounts of 4.8 kb mito-DNA in liver(2.1 × 10-3,1.6 × 10-3),kidney (1.7 × 10-3,1.4 × 10-4) and brain cortex (1.5 × 10-5,1.3 × 10-5) in low-and high-fluoride groups were significantly reduced,as compared with that of control group (2.9 × 10-3,2.0 × 10-3,1.1 × 10-4,all P < 0.05).The amount of 4.8 kb mito-DNA in kidney in high-fluoride group was lower than that in low-fluoride group (P < 0.05).Conclusions Excessive fluoride intake can result in missing of 4.8 kb mito-DNA in liver,kidney and brain cortex.The abnormal of mito-DNA might be related to the dysfunction of mitochondrial respiratory chain.
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Objective To observe the expression of mitochondrial fission protein locus Fis1 and ultrastructural changes in the renal cells of rats with chronic fluorosis,and to reveal the mechanism in mitochondrial damage of the renal cells.Methods Sixty SD rats were randomly divided into 3 groups according to sex and body mass(20 in each group):control group,lower fluoride group and higher fluoride group.All the rats were fed with different doses of sodium fluoride in drinking water(0,10 and 50 mg/L,respectively).Six-month later,the expression of Fisl in renal cells was determined by real-time fluorenscence quantitative PCR and immunohistochemistry technology,the mitochondrial morphology of renal cells was observed under transmission electron microscopy (TEM).Results As compared with the control group(28.70 ± 12.41),Fis1 mRNA levels(91.48 + 34.83 and 582.09 ± 184.69) in renal cells of the lower fluoride and the higher fluoride groups were increased(all P < 0.05).As compared with the control group(10.49 ± 7.66),Fisl protein levels(16.33 ± 10.26 and 21.50 ± 5.24) in renal cells of the lower fluoride and the higher fluoride groups showed a trend of increasing,the higher fluoride group was higher than that of the control group(P < 0.05).By TEM,mitochondrial crest in renal cells of the lower fluoride and the higher fluoride groups was vague or disappeared,mitochondrial division section appeared.Conclusions Fluoride is a kind of toxicant that can cause damage to mitochondrion of renal cells,induce the expression of Fis1 in transcriptional and protein level,and lead to the obstacles of mitochondrial fusion-fission and ultrastructural abnormality of mitochondrion,which may play an important role in mechanism of mitochondrial damage in the renal cells of rats with chronic fluorosis.
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<p><b>OBJECTIVE</b>To explore the changes of protein expression of mitochondrial fission gene dynamin-related 1(Drp 1) in the cortical neurons of rats with chronic fluorosis.</p><p><b>METHODS</b>A total of 120 one-month-old SD rats (each weighing approximately 100-120 g at the beginning of the experiment) were randomly divided into three groups, and fed with the different doses of fluoride containing in drinking water (untreated control containing 0 mg/L fluoride, and low-fluoride & high-fluoride supplemented with 10 and 50 mg/L fluoride,respectively). After 3 or 6 months exposure, 20 rats from each group were killed. Then the protein expression of mitochondrial fission gene, Drp1, was detected by immunohistochemistry and western-blotting method.</p><p><b>RESULTS</b>Dental fluorosis and urinary fluorosis were obviously found in the rats exposed to fluoride. At the experiment period of 3 months, the numbers of positive cells of Drp1 detected by immunohistochemistry changed. Compared with the control group (36.3 ± 5.8), the changes in low-fluoride group (34.7 ± 4.1) showed no significant difference (t = 1.5, P > 0.05),but the increase in high-fluoride group (45.0 ± 4.7) had statistical significance (t = 8.8, P < 0.05). The western-blotting method had consistent results. Compared with the control group (0.59 ± 0.03), a significant increase of the average topical density in low- fluoride (0.62 ± 0.03) and high-fluoride (0.71 ± 0.02) groups were found (t = 0.02,0.11, P < 0.05). At the experiment period of 6 months, the numbers of positive cells of Drp1 detected by immunohistochemistry significantly changed. Compared with the control group (33.2 ± 4.4), the number in low- fluoride and high-fluoride groups were separately (36.6 ± 3.8) and (39.4 ± 4.2),both increased significantly (t = 3.5,6.3, P < 0.05). Same results could be found in western-blotting method,compared with the control group (0.65 ± 0.06), the average topical density in low- fluoride (0.80 ± 0.09) and high-fluoride (0.76 ± 0.08) groups both increased significantly (t = 0.1,0.1, P < 0.05).</p><p><b>CONCLUSIONS</b>Taking excessive amount of fluoride might result in the changes of expression of Drp1, and the neurons damage from the chronic fluorosis might be associated with the hyperfunction of mitochondrial fusion.</p>
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Animais , Masculino , Ratos , Água Potável , Química , Dinaminas , Genética , Metabolismo , Intoxicação por Flúor , Metabolismo , Fluoretos , Urina , Fluorose Dentária , Metabolismo , Dinâmica Mitocondrial , Neurônios , Metabolismo , Patologia , Ratos Sprague-DawleyRESUMO
<p><b>OBJECTIVE</b>To investigate the effects of α3 neuronal nicotinic acetylcholine receptor (nAChR) on apoptosis and p38 signal transduction pathway in SH-SY5Y cells and to assess the roles of α3 nAChR in the pathogenesis of Alzheimer's disease (AD).</p><p><b>METHODS</b>The levels of α3 nAChR mRNA and protein were measured by real-time PCR and Western blot, respectively, in SH-SY5Y cells transfected with α3 nAChR siRNA. The mRNA level of bcl-2 and bax was measured by the real-time PCR. The siRNA transfected SH-SY5Y cells and control were then treated with 10 µmol/L Aβ25-35 for another 48 h, and the change in apoptotic rate and the levels of p-p38 and p38 were measured by flow cytometry and Western blot. Subsequently these SH-SY5Y cells were exposed to a blocker of p38 protein, and the apoptotic rate was measured again.</p><p><b>RESULTS</b>Compared to the controls, the expression of α3 nAChR at mRNA and protein levels in the SH-SY5Y cells transfected with α3 nAChR siRNA decreased by 95% and 86%, respectively; the mRNA levels of bax increased 2.11 times and that for bcl-2 decreased 0.53 times. The apoptotic rate was unaffected (3.40% ± 0.20%); but it increased after Aβ25-35 treatment (24.52% ± 1.59%); the level of p-p38 protein also increased by 178% in the α3 nAChR inhibited cells treated with Aβ25-35. Compared to controls, the Aβ25-35-treated SH-SY5Y cells and the Aβ25-35-treated and siRNA-transfected cells both showed a reduction in apoptosis after treatment with p38 blocker, especially in the former.</p><p><b>CONCLUSION</b>The siRNA silencing of α3 nAChR mRNA may enhance the effect of Aβ25-35 on the cell apoptosis by increasing the levels of p38 protein and bax mRNA and decreasing the level of bcl-2 mRNA, which may play a role in the pathogenesis of AD.</p>
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Humanos , Doença de Alzheimer , Peptídeos beta-Amiloides , Metabolismo , Apoptose , Linhagem Celular Tumoral , Inativação Gênica , Neuroblastoma , Metabolismo , Patologia , Fragmentos de Peptídeos , Metabolismo , Proteínas Proto-Oncogênicas c-bcl-2 , Genética , Metabolismo , RNA Mensageiro , Metabolismo , RNA Interferente Pequeno , Genética , Receptores Nicotínicos , Genética , Metabolismo , Transdução de Sinais , Transfecção , Proteína X Associada a bcl-2 , Genética , Metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno , MetabolismoRESUMO
<p><b>OBJECTIVE</b>To analyze the population genetics characteristics of mitochondrial DNA (mtDNA) in Gelao, Mulao, Maonan ethnic groups from Guizhou.</p><p><b>METHODS</b>Minisequenceing and restriction fragment length polymorphism (RFLP) were used to analyze 12 single nucleotide polymorphism (SNPs) of mitochondrial DNA in the 3 ethnic groups.</p><p><b>RESULTS</b>A total of 30 haplotypes were detected in 156 samples. The distribution of H1, H23 had differed between Mulao, Maonan and Gelao, respectively, and so did M7 among the three groups. The difference was statistically significant (P < 0.05). Mulao, Maonan had respectively differed from Gelao and the difference was also statistically significant (P < 0.05).</p><p><b>CONCLUSION</b>There was a great similarity in the distribution of haplotypes of the mtDNA among the three ethnic groups, except for some difference in the distribution of certain haplotypes.</p>
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Humanos , Masculino , Povo Asiático , Etnologia , Genética , China , Etnologia , DNA Mitocondrial , Genética , Linhagem , Polimorfismo GenéticoRESUMO
<p><b>OBJECTIVE</b>To observe the mitochondrial fragmentation and the expression of mito-fusion 1 gene in the cortical neurons of rats with chronic fluorosis, and to reveal their roles in mitochondria damage to neurons due to chronic fluorosis.</p><p><b>METHODS</b>SD rats were divided randomly into three groups of 20 each (a half females and a half males housed individually in stainless-steel cages), and fed with the different doses of fluoride containing in drinking water (untreated control containing 0 mg/L fluoride, and low-fluoride and high supplemented with 10 and 50 mg/L fluoride, respectively). After 3 or 6 months exposure, the mitochondrial morphology of the neurons in rat brains were observed by transmission electron microscopy (TEM), then the expression of mitochondrial fusion gene, Mfn1, were detected by immunohistochemistry and western-blotting, respectively.</p><p><b>RESULTS</b>Dental fluorosis was obvious in the rats exposed to excessive fluoride in their drinking water, that is, (16 rats out of 20) numbers of I° detal fluorosis in the low-fluoride group, and (11 rats out of 20) numbers of I° and (9 rats out of 20) numbers of II° detal fluorosis in the high-fluoride group were observed after 3 months exposure. Moreover, (14 rats out of 20) numbers of I° and (6 rats out of 20) numbers of II° detal fluorosis in the low-fluoride group and (6 rats out of 20) numbers of Io, (13 rats out of 20) numbers of II°, and (1 rats out of 20) numbers of III° detal fluorosis in the high-fluoride group were observed after 6 months exposure. And both of untreated controls without detal fluorosis were also observed. The urinary level of fluoride in the low-fluoride group (3.30 ± 1.18) mg/L and in the high-fluoride group (5.10 ± 0.35) were observed after 3 months exposure (F = 3.18, P < 0.05). Moreover, the urinary level of fluoride in the low-fluoride group (4.16 ± 1.39) mg/L and in the high-fluoride group (5.70 ± 1.70) mg/L were also observed after 6 months exposure (F = 3.17, P < 0.05). The normal mitochondrial morphology of neurons in rats without fluorosis was observed after 3 and 6 months, while the abnormal mitochondrial morphology of neurons with fluorosis was shown, presenting mitochondrial fragmentation with swollen cristae and even the fragmented, shortened or stacked punctuate membranes (section observation of three bullous mitochondrial-mitochondrial fission process) by TEM. As compared with controls (53.0 ± 4.54 and 1.21 ± 0.18) at the experiment period of 3 months, Mif1 protein analysis with immunocytochemical (the numbers of positive cells: 51.09 ± 6.25) and western-blotting (1.22 ± 0.26) were no significant difference for low fluoride group (t = 1.7, 1.1, P > 0.05); Mif1 protein analysis with immunocytochemical (the numbers of positive cells: 59.71 ± 5.64) and western-blotting (1.66 ± 0.20) were significantly increasing for high fluoride group (t = 2.1, 2.1, P < 0.05). As compared with controls (36.43 ± 4.04 and 1.00 ± 0.13) at the experiment period of 6 months, Mif1 protein analysis with immunocytochemical (the numbers of positive cells 20.05 ± 4.55 and 17.10 ± 3.86) and western-blotting (0.64 ± 0.08 and 0.39 ± 0.06) were significantly decreasing for the two fluoride group (t = 2.1, 2.2; 2.2, 2.2 respectively, all P value were < 0.05).</p><p><b>CONCLUSIONS</b>Taking excessive amount of fluoride might result in the mitochondrial fragmentation for the changed expression of Mfn1, and the neurons damage from the chronic fluorosis might be associated with the dysfunction of mitochondrial fusion.</p>
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Animais , Feminino , Masculino , Ratos , Água Potável , Química , Intoxicação por Flúor , Metabolismo , Patologia , Fluorose Dentária , Metabolismo , Proteínas de Membrana , Metabolismo , Mitocôndrias , Patologia , Proteínas Mitocondriais , Metabolismo , Neurônios , Metabolismo , Patologia , Ratos Sprague-DawleyRESUMO
<p><b>OBJECTIVE</b>To study the frequency of a 9 bp deletion polymorphism of mitochondrial DNA (mtDNA) in ethnic Miao, Buyi and Dong populations from Guizhou province.</p><p><b>METHODS</b>Polymerase chain reaction-polyacrylamide gel electrophoresis (PCR-PAGE) was used to detect the 9 bp deletion. The result was verified with DNA sequencing.</p><p><b>RESULTS</b>Two polymorphisms, including a standard pattern and a short pattern (the 9 bp deletion), were found among the three ethnic groups. The frequency of short pattern in 304 males was 23.0%. Respectively, those of Miao, Buyi and Dong ethnics were 28.6%, 26.8% and 13.7%. A statistically significant difference was detected among the three groups (P<0.05).</p><p><b>CONCLUSION</b>The frequencies of the 9 bp polymorphism were relatively high among ethnic Miao, Buyi and Dong populations from Guizhou, and there was a significant difference between the three.</p>
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Humanos , Masculino , Sequência de Bases , China , Etnologia , DNA Mitocondrial , Genética , Deleção de Genes , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Polimorfismo Genético , Análise de Sequência de DNARESUMO
<p><b>OBJECTIVE</b>To investigate allelic frequencies of interluekin-10 (IL-10) gene promoter in Miao, Dong and Buyi ethnics of Guizhou.</p><p><b>METHODS</b>TaqMan MGB-based real-time PCR was used to determine the genotypes of IL-10 -819 and IL-10 -592 in 589 Miao, Dong and Buyi ethnics of Guizhou.</p><p><b>RESULTS</b>The allelic frequency of IL-10 -819 in Miao ethnics was significantly different from those in Dong or Buyi ethnics. Allelic frequencies of IL-10 -592 in Miao ethnics was significantly different from those in Dong or Buyi ethnics. In Miao, Dong and Buyi ethnics, the distributions of genotype frequencies of IL-10 -819 and IL-10 -592 were statistically different from Han ethnics from Guizhou and Taiwan of China as well as South Koreans.</p><p><b>CONCLUSION</b>There is a heterogeneity in the frequencies of polymorphisms of IL-10 promoter among different ethnic groups.</p>
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Humanos , Alelos , Povo Asiático , Etnologia , Genética , China , Etnologia , Frequência do Gene , Genética Populacional , Genótipo , Interleucina-10 , Genética , Polimorfismo de Nucleotídeo Único , Grupos Populacionais , Genética , Regiões Promotoras GenéticasRESUMO
Objective The aim of the study was to investigate the expression of P-glycoprotein (P-gp)and nuclear factor κB (NF-κB) in the cerebral cortex of rat with chronic fluorosis,and to reveal the mechanisms of damaged nervous system resulted from the toxicity of fluoride.Methods Sixty SD rats were randomly divided into three groups.The rats in each group were given drinking water containing different levels of fluoride:control group less than 0.5 mg/L,small amount of fluoride exposure group 10.0 mg/L and large amount of fluoride exposure group 50.0 mg/L.The animals were examined at the sixth month after initiating the experiment.Protein levels of P-gp and NF-κB in brain tissues were detected by immunocytochemistry and Western blotting,and the P-gp protein and mRNA level by quantitative real time PCR method.Results As compared to the control group(28.21 ±6.13),the numbers of positive staining cells by P-gp antibody in the cortex of rat brains were significantly increased in both the small and the large amount of fluoride exposure groups[(48.46 ± 8.00),(53.72 ± 9.15),respectively,all P < 0.05] ; the protein levels in the control group(100.00 ± 3.86)% detected by Western blotting were significantly increased in the cortex of rat brains treated with fluoride in both the small and the large amount of fluoride exposure groups[(189.47 ± 3.14)%,(191.36 ± 11.09)%,respectively,all P < 0.05].The significantly increased expression of NF-κB at the protein level was observed in the cortex of rat brains of the small and the large amount of fluoride exposure groups[(365.97 ± 6.04)% and (417.15 ± 10.89)%,respectively] as compared with the control group[(100.00 ± 10.07)%,all P < 0.05].The mRMA level of P-gp in the cortex of rat brains of the small and the large amount of fluoride exposure groups(2396 ± 427,3479 ± 371,respectively) were higher than that of the control group(260 ± 106,all P < 0.05).Conclusion The increased expressions of P-gp and NF-κB in the cortex of rat brains are induced by chronic fluorosis,which might be connected with the mechanism of brain damages.
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ObjectiveTo investigate the transcriptional changes of nitochondria fission and fusion gene loci and reactive oxygen species (ROS) level in cortical neurons of rats with chronic fluorosis,and to reveal their roles in mitochondria damage due to chronic fluorosis.MethodsSD rats were fed with different doses of fluoride through drinking water[< 0.5(control),10,50 mg/L,respectively] for 3 and 6 months.The level of ROS and mRNA contents of mitochondria fission gene loci Drp1/Fis1 and fusion gene locus Mfn1 in the cortical neurons of rat brains were detected with ROS fluorescent probe and real-time PCR,respectively.ResultsAs compared with control group [10.43 ± 5.98,(3.4 ± 0.6) × 103,(8.8 ± 1.4) × 10,(1.2 ± 0.2) × 102] at the experiment period of 3 months,the level of ROS and mRNA contents of mitochondria fusion gene locus Mfn1 and fission gene loci Drp1/Fis1 in the cortical neurons were obviously increased in the rats fed with 50 mg/L fluoride through drinking water[25.48 ± 6.09,(1.0 ± 0.2) × 1011,(3.0 ± 1.6) × 103,(8.9 ± 3.6) × 102,all P < 0.05],whereas no significant changes were found in the rats fed with 10 mg/L fluoride[11.67 ± 3.49,(3.1 ± 0.3) × 104,(6.7 ± 2.7) × 10,(5.0 ± 0.9) × 10,all P >0.05].Furthermore,at 6 months of the experiment the increases in ROS level(63.02 ± 8.15,65.60 ± 7.40) and mRNA contents of mitochondria fission gene loci Drp1/Fis1 [(2.0 ± 0.8) × 106,(4.0 ± 0.6) × 105,(3.8 ± 1.3) × 103,(1.3 ± 0.2) × 103] and the decrease in mitochondrial fusion gene locus Mfn1[(3.0 ± 0.4) × 106、(4.0 ± 0.9) × 104]were observed in the cortical neurons of the rats fed with 10 mg/L and 50 mg/L fluoride as compared with the control group[25.26 ± 6.41,(3.0 ± 0.8) × 109,(5.1 ± 0.8) × 103,(2.8 ± 0.7) × 102,all P < 0.05].Conclusions Excessive intake of fluorine leads to elevated ROS levels,and decreased transcription of mitochondrial fusion gene loci Mfn1,which is positively correlated with the time and dose-exposed to fluoride.The changes of mitochondrial fission and fusion gene loci in the cortical neurons may be related to high level of oxidative stress induced by chronic fluorosis.
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ObjectiveTo observe the distribution of vitamin D receptor(VDR) gene polymorphisms in coal-burning borne fluorosis in Guizhou province and investigate the relationship between VDR gene polymorphisms and the susceptibility to coal-burning borne fluorosis.MethodsOne hundred and fifty villagers from non-improving cooking stove villages were selected as a non-intervention group in Bijie area,Guizhou province where coal-burning borne fluorosis was prevailing; 150 villagers were chosen from cooking stove improved villages as a intervention group; 150 villagers were selected from non-endemic area Changshun county as a control group.DNA was extracted from peripheral blood samples of these people.Genotype of VDR gene Bsm Ⅰ and Fok Ⅰ loci were detected using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP).ResultsDistribution of Bsm Ⅰ polymorphism site of VDR gene of control group [AA:19.3% (29/150),AG:39.3% (59/150),GG:41.3%(62/150)],was compared with that[AA:4.7%(7/150),AG:14.0%(21/150),GG:81.3%(122/150)] of the non-intervention group and that[AA:7.3%(11/150),AG:23.3%(35/150),GG:69.3%(104/150)] of intervention group,and the difference was statistically significant(X2 =56.6,P < 0.05).The frequency of VDR-Fok Ⅰ loci in non-intervention group [TT:29.3%(44/150),TC:55.3%(83/150),CC:15.3%(23/150)] and intervention group [TT:32.7%(49/150),TC:55.3%(83/150),CC:12.0%(18/150)] was compared with that [TT:45.3%(68/150),TC:48.7%(73/150),CC:6.0%(9/150)] of control group,and the difference was statistically significant(X2 =11.9,P < 0.05).Univariate analysis showed that individuals carrying the GG genotype had increased risk of suffering fluorosis than individuals carrying the AA and AG genotypes(OR values were 6.2,3.2,all P < 0.05),while carrying the TC and CC genotype had increased risk of suffering fluorosis than individuals carrying the TT genotype (OR values were 1.3,2.8,1.3,2.1,all P < 0.05).ConclusionVDR gene polymorphisms may be one of the predisposing factors of coal-burning borne fluorosis.
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Objective To investigate the association between both regulated upon activation normal T cell expressed and secreted (RANTES)-403G/A,-28C/G gene polymorphism and the susceptibilities to hepatitis B virus (HBV) infection,among people with Dong and Han ethnicities,in Guizhou.Methods A total of 229 individuals with HBV persistence infection,161 HBV clearanced patients and another 200 controls were recruited to conduct a case-control study among residents with Dong or Han ethnicities.Allelic frequencies of both RANTES-403G/A and-28C/G were identified by TaqMan-MGB probe.Results Both RANTES-403G/A and-28C/G polymorphism in the HBV-persistent group,when compared to the HBV-clearances group,no significant difference was found (P>0.05).Results from the univariate analysis showed that subjects carrying-403AG and-28GG genotype had higher risk on the susceptibility to HBV persistence infection.The distributions of RANTES-28C/G gene polymorphism between Dong minority and Han ethnicities regarding HBV persistence showed statistically significant difference (P<0.05).There was no difference on the distributions of RANTES-403G/A gene polymorphism between Dong minority and Han ethnicities.Conclnsion Patients that carrying both RANTES-403AG and-28GG genotype had higher risk on the persistence to HBV,while RANTES-403A had contributed to the clearance of HBV infection.
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<p><b>OBJECTIVE</b>To investigate the influence of inhibited α7 neuronal nicotinic acetylcholine receptor (nAChR) by small interference RNA (siRNA) in SH-SY5Y cells and to explore the connection of these changes with the β-amyloid precursor protein (APP) metabolism and the pathogenesis of Alzheimer's disease (AD).</p><p><b>METHODS</b>The siRNA of α7 nAChR was transfected into SH-SY5Y cells, and the expression of α7 nAChR and two subtypes of β-secretases (BACE1 and BACE2) at mRNA and protein levels was studied by real-time PCR and Western blot, respectively. The variation of Aβ(1-42) content was detected by ELISA.</p><p><b>RESULTS</b>As compared with controls, the expression of α7 nAChR at mRNA and protein levels in the SH-SY5Y cells transfected with the α7 nAChR siRNA were decreased by 84% and 79% (P < 0.01), respectively. The expressions of BACE1 mRNA and protein levels was increased by 527% and 71% (P < 0.01), respectively, while the expression of BACE2 decreased by 58% and 75% (P < 0.01), respectively. The Aβ(1-42) content increased by 208% (P < 0.01).</p><p><b>CONCLUSIONS</b>An inhibited α7 nAChR mRNA induced by siRNA may markedly stimulate the production of Aβ through the mechanism of increased expression of BACE1 and inhibited expression of BACE2, which may be related to the pathogenesis of AD.</p>
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Humanos , Secretases da Proteína Precursora do Amiloide , Genética , Metabolismo , Peptídeos beta-Amiloides , Metabolismo , Ácido Aspártico Endopeptidases , Genética , Metabolismo , Linhagem Celular Tumoral , Neuroblastoma , Metabolismo , Patologia , Fragmentos de Peptídeos , Metabolismo , Interferência de RNA , RNA Mensageiro , Metabolismo , RNA Interferente Pequeno , Genética , Receptores Nicotínicos , Genética , Metabolismo , Transfecção , Receptor Nicotínico de Acetilcolina alfa7RESUMO
Objective To investigate the association between interleukin-10 (IL-10) gene promoter microsatellite polymorphisms and the susceptibility to hepatitis B virus infection in Han,Yi and Yao ethnicities in GuiZhou province.Methods 500 volunteers were selected from Guizhou province.Ailelic frequency of IL-10.G and IL-10.R loci was identified by short tandom repeat polymerase chain reaction.The relativity between allelic frequency and HBV infection was analyzed.Results Genotype data from H-W analysis on all the IL-10 polymorphisms indicated that it was a random distribution.Very high HBV infection rates were found in the native ethnic minorities of Guizhou province.The overall HBV infection rate among the total population was 67.00%,with the HBV infection rates of Yi nationality in Weining,Yi nationality in Qianxi,Yao nationality in Libo and Han nationality in Libo as 51.85%,42.86%,79.52% and 84.30%,respe~vely.The polymorphisms distribution of IL- 10.G and IL- 10.R were statistically different among the ethnic groups (P< 0.05 ).The polymorphisms distribution of IL-10.R had no significant difference between HBV infection group and non-infection group,as well as among HBV natural removal group and non-infected group in all the ethnic groups.The frequency of IL-10.G 459 bp (19CA) was significantly higher in non-infection group than in the infected group (P< 0.05 ).The frequency of IL-10.G 471 bp (25CA) was significantly higher in the non-infection group than in the HBV natural removal group(P<0.05).The polymorphisms distribution of IL-10.G did not show significant difference between the HBV infection group and the HBV natural removal group in all the ethnic groups.We did not find any differences in allelic and genotypic frequencies of IL-10.G between infection group and non-infection group in Yi nationality in Weining,and Yao nationality in Libo (P>0.05),as well as HBV natural removal group and non-infected group (P>0.05).Conclusion The polymorphisms distribution of IL-10.R and IL-10.G did not show significant difference in Yi,Yao and Han ethnics population living in Guizhou province.IL-10.G seemed to influence the susceptibility of HBV infection in Han,Yao and Yi ethnics population of Guizhou province.
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<p><b>OBJECTIVE</b>To investigate the expressions of mRNA and protein of p38, Osx, PI3K, Akt1 in the rats bone with chronic fluorosis.</p><p><b>METHODS</b>Dental fluorosis were observed and the fluoride contents in the urine and bone were detected by fluorin-ion selective electrode. The morphologic changes and ultrastructure of rats' bone were observed by light and electronic microscopy. The expressions of protein and mRNA of p38, Osx, PI3K and Akt1 were detected by immunohistochemistry and real-time PCR, respectively. The contents of BALP and BGP in serum were detected by ELISA.</p><p><b>RESULTS</b>The rates of dental fluorosis in the fluorosis rats were increased, and the fluoride contents in bone and urine of the fluorosis rats were increased compared to the control group, the difference was statistically significant (P < 0.05). The bone trabeculae thickness and density and the thickness of bone cortex in fluorosis rats were remarkably increased, the space of bone trabeculae was reduced, and in accordance with the matching morphometrical indices, the difference was statistically significant (P < 0.05) as compared with the control rats. The contents of BALP [(54.61 ± 2.27) U/L] and BGP [(2.38 ± 0.16) µg/L]in the fluoride groups were higher than those in the control group, the difference was statistically significant (P < 0.05). Ultrastructurally, the broadening of the osseouslacuna was observed. The reduced protuberances of the osteocytes, the unclear organelle structure, pyknosis, karyotheca increasation and edged chromatin were also observed. Compared to the control group, the expressions of protein and its mRNA of p38, Osx, PI3K and Akt1 were higher in the fluorosis rats than those in the control rats, and the difference was statistically significant (P < 0.05). There is no any expression of p38, Osx, PI3K and Akt1 in the osteocytes in fluorosis rats.</p><p><b>CONCLUSIONS</b>The over-expression of p38, Osx, PI3K and Akt1 in bone tissue of fluorosis rats may relate to the accumulation of fluorine in the body. The bone injury mainly occur in the stage of the differentiation and proliferation. The upregulation of P38MARK signal path and PI3K/Akt1 signal path may be involved in the pathogenesis of bone injury caused by fluoride.</p>