Objective To explore the expressions and significances of apoptotic modulating genes Fas and Bcl-2 mRNA in peripheral blood monocytes(PBMC)of patients with myasthenia gravis (MG). Methods The levels of Fas and Bcl-2 mRNA in PBMC were detected by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) in 44 patients with MG as well as 30 healthy donors served as a normal control (NC).Results (1) The level of Fas mRNA in MG group was no significantly difference compared with NC group. The level of Bcl-2 mRNA in MG group was higher than that in NC group(P

High-risk human papillomavirus infection is the major cause of tumors such as cervical cancer and head and neck cancer.MicroRNAs (miRNAs) are 20-25nt small non-coding RNAs,which are involved in the proliferation,apoptosis,invasion and migration of tumors.HR-HPVs,which integrate into the host genome,encode three oncogenes:E6,E7 and E5 and enhance carcinogenesis by regulating downstream miRNAs and genes e.g.p53 and pRb.

Aim To study the effect of (S) -1, 8-(2-methyl phosphate ethoxy )-6-fluorine-7-( 4-methyl- pi-perazine-1-base )-3-[ S-benzyls-based-4-( for nitroben-zene methylene group amino )-1 , 2 , 4-all triazole-3 base]-quinoline ( 1-H )-4-ketone ( M18 ) on apoptosis of hepatocarcinoma SMMC-7721 cells in vitro. Meth-ods With different concentrations of M18 at different time used to treat SMMC-7721 cells, human breast cancer MB-231cells, human colon cancer HCT-116 cells, human hepatocarcinoma HEPG-2 cells, mouse bone marrow mesenchymal stem cells ( BMSCs ) in vitro,the inhibition effects of M18 on cell proliferation were examined by MTT assay. Cell apoptosis was de-termined using Hoechst 33258 fluorescence staining and TUNEL method. Mitochondrial membrane poten-tial (△ψm ) was measured using a high content screening image system. Protein expression of caspase-3 , p53 and cytochrome C was detected with Western blot analysis. Results Treatment with M18 ( 4 ~32μmol·L-1 ) potently inhibited the proliferation of the cancer cells in time-and dose-dependent manners ( the IC50 value at 24 h in SMMC-7721 cells, MB-231cells,
HCT-116 cells and HEPG-2 cells was 8. 65 μmol · L-1 , 9. 37 μmol · L-1 , 12. 74 μmol · L-1 and 9. 40μmol · L-1 , respectively ) . In contrast, M18 had weak cytotoxicity against BMSCs with IC50 value of 38. 96 μmol·L-1 . Levofloxacin had weak cytotoxicity against SMMC-7721 cells with IC50 value of 735. 10μmol·L-1 . Treatment of SMMC-7721cells with differ-ent concentrations of M18 for 24 h increased the per-centage of the apoptosis cells ( P <0. 05 ) and de-creased the mitochondrial membrane potential. In ad-dition, M18 increased protein expression of p53, caspase-3 and the cleaved activated forms of caspase-3 in SMMC-7721 cells. Treatment of SMMC-7721 cells with M18 significantly increased cytochrome C in the cytosol, and decreased cytochrome C in the mitochon-drial compartment. Conclusion The mitochondrial-dependent pathways are involved in M18 induction of apoptosis of SMMC-7721 cells.

Objective To study features of the MRI and clinic in a family with pure hereditary spastic paraplegia (PHSPG) type 6.Methods Target loci (SPG3, 4, 6, 8 10 and 12) linkage analysis was performed in a SPG pedigree having 6 affected individuals using microsatellite markers and NIPA1 gene was screened for mutation by PCR-amplification and sequencing. MRI of brain and cervical and thoracic spinal cord were examined in these 6 patients and 6 normal controls matched for age and sex by two independent radiologists blinded to the clinical diagnosis. Cross-sectional areas and anteroposterior and transverse diameters of the spinal cord at the levels of C2～3, C7, T1～4, T9 were measured and data was statistically analyzed using the student's t test. Results A missense mutation of 316g→c in NIPA1 was identified in the affected subjects, presumably resulting in substitution of glutamic acid for arginine in residue 106. Evaluation of the brain MRI images revealed non-specific brain abnormalities. All patients presented thinning of cervical and upper thoracic spine with atrophy in both gray and white matter and enlarged subarachnoid cavity. In severe atrophic segments, a distinct boundary between grey and white matter was observed and the lesions in grey matter presented literal high intensity spots or patches with clear boundary on transaxial T2-weighted images (T2WI) and high signal intensity longitudinal strip on the sagittal T2WI. Cross-sectional areas and anteroposterior and transverse diameters of the spinal cord at C2～3, C7, T1～4 were significantly smaller in patients than in controls, while at the T9 level only transverse diameter showed significant difference (7.22±0.08 vs 8.17±0.41, t=2.870, P=0.046). Conclusions These findings indicate that the disease process in patients with SPG6 might be confined to the cervical and thoracic spinal cord, with atrophy in both white and grey matter having a distinct boundary.

Objective To investigate the dose-response pattern on the inducible expression of PIG3 mRNA in normal human lymphoblastoid AHHI cells by 60Co γ-rays,and its possibility for developing novel radiation biodosimeter.Methods Laser confocal fluorescent microscopy was used to detect the γ-H2AX foci,a biomarker of DNA double-strand break.After irradiation with 0,1,2,4,6,8 and 10 Gy of 60Coγ- rays,AHH-1 cells were harvested at 4,10 and 24 h post-irradiation,and subjected to total RNA extraction and detection of PIG3 mRNA expression by quantitative real-time PCR.Results PIG3 protein foci were formed in the nuclei at 30 min after irradiation,and a part of these PIG3 foci were colocalized with γH2AX foci.After irradiation,PIG3 mRNA level was enhanced with the increased time of postirradiation,and remained an increased level at least till 24 h.The radiation-inducible expression of PIG3 mRNA was demonstrated in a dose-dependent manner.The dose-dependent range at 4 h post-irradiation was 0 - 6 Gy,and at 10 h and 24 h was 0 - 10 Gy.Conclusions PIG3 involves in the cellular response to DNA double-strand break.The dose-dependent inducible response of PIG3 mRNA expression might provide a valuable candidate for developing a novel radiation biodosimeter.

Objective To investigate the effect of Tip60 on the cellular radiosensitivity,and to explore the related mechanism.Methods siRNA and anacardic acid (AA,an inhibitor of Tip60 acetyltransferase) were used to inhibit Tip60 expression and its acetyltransferase activity,respectively.Radiosensitivity was analyzed by colony-forming ability assay.γ-H2AX foci were detected to analyze the DNA double-strand break (DSB).Immunoprecipitation was used to determine the interaction of proteins.Results siRNA-mediated silencing of Tip60 led to enhanced sensitivity of U2OS cells at 1,2 Gy after γ-ray irradiation,but had no significant effect at 4 Gy post-irradiation ( t =3.364,3.979,P ＜ 0.05 ).γ-H2AX foci detection indicated that Tip60 silencing resulted in a decreased capability of DNA doublestrand break repair at 1,4 and 8 h after irradiation( t =3.875,3.183 and 3.175,respectively,P ＜ 0.05 ).The interaction of Tip60 and DNA-PKcs was prompted by ionizing radiation.Anacardic acid largely abrogated the phosphorylation of DNA-PKcs at T2609 site induced by irradiation.Conclusions Tip60plays a role in the cellular response to ionizing radiation-induced DNA damage through,at least in part,interacting with DNA-PKcs and regulating its phosphorylation.

Objective To evaluate the radioprotective effect of vanillin derivative VND3207 on DNA damage induced by different LET ionizing radiation.Methods The plasmid DNA in liquid was irradiated by 60Co γ-rays, proton or 7Li heavy ion with or without VND3207.The conformation changes of plasmid DNA were assessed by agarose gel electrophoresis and the quantification was done using gel imaging system.Results The DNA damage induced by proton and 7Li heavy ion was much more serious as compared with that by 60Co γ-rays, and the vanillin derivative VND3207 could efficiently decrease the DNA damage induced by all three types of irradiation sources, which was expressed as a significantly reduced ratio of open circular form (OC) of plasmid DNA.The radioprotective effect of VND3207 increased with the increasing of drug concentration.The protective efficiencies of 200 μmol/L VND3207 were 85.3% (t =3.70,P =0.033), 73.3% (t = 10.58, P =0.017)and 80.4% (t =8.57,P =0.008)on DNA damage induction by 50 Gy of γ-rays, proton and 7Li heavy ion, respectively.It seemed that the radioprotection of VND3207 was more effective on DNA damage induced by high LET heavy ion than that by proton.Conclusions VND3207 has a protective effect against the genotoxicity of different LET ionizing radiation, especially for γ-rays and 7 Li heavy ion.

OBJECTIVETo explore the mechanism of paraquat (PQ) poisoning and to observe the changes in inflammatory cytokines in PQ-exposed rats treated in different ways.

METHODSFifty 8-week-old clean male Wistar rats were randomly divided into high-dose curcumin plus conventional treatment group, low-dose curcumin plus conventional treatment group, high-dose curcumin group, PQ poisoning group, and blank control group. On days 1, 3, 5, 7, 14, and 21 after PQ exposure, serum levels of transforming growth factor-β₁(TGF-β₁) , tumor necrosis factor-α (TNF-α) , and interleukin-6 (IL-6) were measured. The pathological changes in lung tissue were evaluated by HE staining.

RESULTSCompared with the blank control group, the high-dose curcumin plus conventional treatment group, low-dose curcumin plus conventional treatment group, high-dose curcumin group, and PQ poisoning group had significantly increased serum levels of TGF-β₁, TNF-α, and IL-6 (P<0.05) , and the three cytokines in each group reached peak levels on day 14 after exposure. Compared with the PQ poisoning group, the high-dose curcumin group had significantly reduced serum levels of TGF-β₁, TNF-α, and IL-6 (P<0.05). On day 21 after exposure, there were no significant differences in serum levels of TGF-β₁, TNF-α, and IL-6 between the high-dose curcumin plus conventional treatment group and the low-dose curcumin plus conventional treatment group (P>0.05). The HE staining revealed alveolar inflammatory changes on days 1~7 and massive pulmonary fibrosis on days 14~21 in the high-dose curcumin plus conventional treatment group, low-dose curcumin plus conventional treatment group, high-dose curcumin group, and PQ poisoning group, but the above changes were milder in the high-dose curcumin group than in the PQ poisoning group.

CONCLUSIONFor rats with PQ poisoning, curcumin can significantly reduce inflammatory response and pathological changes in lung tissue and inhibit and delay the development and progression of body injury.

Animals ; Curcumin ; pharmacology ; Cytokines ; blood ; Interleukin-6 ; blood ; Lung ; pathology ; Male ; Paraquat ; poisoning ; Pulmonary Fibrosis ; pathology ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar ; Transforming Growth Factor beta1 ; blood ; Tumor Necrosis Factor-alpha ; blood

Objective To summarize the clinical features of global developmental delay (GDD) children,and to explore the relationship between severity of GDD and social-culture factors.Methods Sign the informed consent before enrollment.Collect clinical data in detail about 100 GDD children (GDD group) and 95 children with normal development (healthy control group),and analyze their regular clinical data,physical examination,intellectual disability test,electroencephalography (EEG) and cranial imaging test.Spearman rank correlation was used to analyze the differences of social and cultural factors between GDD group and healthy control group,such as maternal reproductive age,parental education level and family economic status.At the same time,we compared the lag degree of total developmental quotient and the degree of developmental retardation of five energy areas with the above factors.Results Significant associations were found between GDD and maternal/paternal education,economic level of family,but no sgnificant association was found between maternal age and GDD.And analysis in the relationship between severity of delay in all domains of the child's developmental status about language and social-culture factors,we only obtained the severity of delay in abilities about language is related with maternal education.Spearman rank correlation statistics explains that if there are the lower level of education with mothers,the delay of language domain will more severe (rs =-0.505,P ＜ 0.05).Conclusions Significant associations were found between GDD and maternal/paternal education,economic level of family.The higher maternal education was an important protective factor against risk of GDD.Improving the cognition of parents in child health care,early be diagnosed,early be intervened,is the most important for children with language development.

Objective:To construct and identify the monoclonal antibody of ATP synthase beta subunit (ATP5B) with high purity, and to lay foundation for further study.Methods:The ATP5Bgene was amplified by PCR and cloned into the pET28avector and transformed into E.coli BL21 (DE3) .The protein expression was induced by IPTG and then the fusion protein was purified by nickel affinity chromatography column.The protein purity was detected by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) .Three female Balb/C mice were immunized with purified fusion protein and the tail vein blood was taken to detect the titer of ATP5Bantibody by indirect enzyme-linked immunosorbent assay (ELISA) .The spleen cells from the immunized mice with the highest serum titer were mixed with the SP2/0cells to establish the hybridoma cells and the fused cells were screened by indirect ELISA and monoclonally cultured.Karyotype analysis were performed in the positive cells.The hybridoma cells were intraperitoneally injected into 12weeks old BALB/C mice to estabilish the ascites models.The titer of ascites was detected by indirect ELISA.The purity of the antibody was detected by SDS-PAGE.The antibody subtype was detected by ELISA.Results:After PCR amplification, a specific band of 1 455bp was obtained, and the pET28aempty vector was ligated to obtain a recombinant pET28a/ATP5Bvector.The target protein was expressed in the IPTG-induced bacteria solution;the SDS-PAGE results showed that the protein band was found at51 000.The indirect ELISA results showed that the serum titer of the venous blood of immunized mice was up to1:64 000.In karyotype analysis, the total number of chromosomes in hybridoma cells was about the sum of myeloma cells and normal mouse spleen cells.The mouse ascites was prepared with the hybridoma cell line, and the highest titer of the antibody was 1:240 000.The subtype of the monoclonal antibody produced by the hybridoma cells was IgG1.Conclusion:The monoclonal antibody against ATP5Bprotein is successfully prepared by cloning, expressing and purifying the recombinant protein.