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This article reported the prenatal diagnosis of a fetus with ZTTK syndrome. A pregnant woman underwent preimplantation genetic diagnosis because her partner carried a balanced chromosomal translocation. Chromosomal karyotype analysis and copy number variation sequencing (CNV-seq) performed on amniocytes collected at 18 + weeks of gestation revealed no abnormalities. Ultrasonography performed at 23 +5 and 26 +3 weeks of gestation revealed severe fetal growth restriction, cerebellar dysplasia, poorly visualized sacrum and coccyx, and spina bifida. MRI of the fetal brain showed that the bilateral cerebellar hemispheres of the fetus were small and the cisterna magna was large at 23 +6 weeks of gestation. Whole exome sequencing in the pedigree identified a heterozygous variant c.2092delG (p.Glu698fs*4) in the exon 3 of the fetal SON gene, which was not inherited from the parents and proved to be a de novo mutation. Mutations in the locus are pathogenic, causing ZTTK syndrome. After genetic counseling, the pregnant woman and her family chose to terminate the pregnancy.
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This paper reported the diagnosis and treatment of two neonates with Kabuki syndrome (KS).Neither of them had typical facial features of KS during the neonatal period,but poor response,abnormal appearance and multiple organ dysplasia were observed in both.Case 1 was lost to follow up after discharge,while typical KS facial features were gradually appeared in Case 2 including eversion of lower lateral eyelids,arched eyebrows,sparse eyebrow arch,flattened nasal tip,prominent ears,during a three-month follow-up after birth.Next-generation sequencing revealed that both neonates were KS caused by lysine methyltransferase 2D (KMT2D) gene mutation,of which case 1 had a heterozygous deletion mutation ofc.13895delC (p.P4632HfsTer8) in KMT2D gene,while case 2 had a heterozygous repeat mutation of c.12809dupA (p.T4271Dfs*63) in KMT2D gene.Both cases were defined as de novo mutations and the one carried by case 2 was a newly discovered pathogenic mutation.
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Objective To investigate the effects of lentivirus-mediated RNA interference (RNAi) targeting DNA binding protein A (dbpA) on the proliferation and the biological behavior of colorectal cancer cell line SW620.Methods The experiment was divided into 3 groups:KD group (siRNA-dbpA,lentivirus interference group),CON group (non-specific sequence group) and NC group (blank control group).The lentiviral vector siRNA-dbpA was constructed and verified by PCR and DNA sequencing.SW620 cells were transfected with siRNA-dbpA plasmid,nontargeting siRNA plasmid,or empty plasmid.After 48 h the transfection,the cells were examined for dbpA expression using Western blot.After 72 hrs transfection,flow cytometry was used to detect the cell apoptosis and cell cycle changes.The cell growth inhibition rate was detected by MTT (4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide) assay,and then clone formation was detected,and the ability of SW620 cells to form tumors in vivo after dbpA was silenced was studied in nude mice.Results PCR analysis and DNA sequencing demonstrated that the RNAi sequence targeting dbpA gene was successfully inserted into the lentiviral vector.siRNA-dbpA transfection resulted in reduced expression of dbpA in SW620 cells.After transfection,the apoptosis rate of siRNA-dbpA-transfected cells increased to 26.60％ ± 0.38％,significantly higher than that in cells transfected with the nontargeting plasmid or the empty plasmid 12.54％ ± 0.25％ and 4.46％ ± 0.19％,respectively (F =28.159,P ＜0.01).The growth inhibition test indicate that the OD value of the fifth day in siRNA-dbpA group was 0.194 ±0.037,significantly lower than that in the other two groups 0.814 ±0.043 and 1.625 ±0.061,respectively(F =23.214,P ＜ 0.01).The colony formation number is 37 ± 3,64 ± 5and 175 ± 10 respectively,siRNA-dbpA is significantly higher than that in the other two groups(F =40.254,P ＜ 0.01).After the completion of nude mouse transplantation tumor model,through the detection of tumor volume,KD group (group siRNA-dbpA) tumor volume after 14 d and CON and NC group had obvious difference (F =38.256,P ＜ 0.05),and after 21d is more significant difference in tumor size (F =40.241,P ＜ 0.01),can be clearly observed after 35 d KD group (group siRNA-dbpA) growing tumors had differences with the control group (F =30.257,P ＜ 0.05).Conclusion Lentivirus-mediated RNAi targeting dbpA can effectively suppress the expression of dbpA in colorectal tumor in nude mice,it is proved that dbpA silencing has a significant inhibitory effect on the growth of living tumor cells and decrease the proliferation of the colorectal cells.
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Objective Analyzing the lncRNA expression profile in peripheral blood mononuclear cells(PBMC)of primary biliary cirrhosis(PBC)patients to provide new ideas for the pathogenesis,clinical diagnosis and treatment of PBC.Methods Collected peripheral blood from 30 PBC patients and 30 healthy volunteers, then separated PBMC.Four cases from each group were selected for long-noncoding RNA (lncRNA)expression microarray detection.Reverse transcription-PCR technology in a larger sample size was used to verify the microarray results.Bioinformatic analysis such as Cis-/Trans-target genes Gene ontology(GO)and pathway analysis, co-expression networks were conducted in order to provide a theoretical basis in the pathogenesis of PBC.Transfecting small interfering RNAs(siRNAs)for linc-pbc to see changes in the expression of nuclear receptor 4A group 3(NR4A3)and forkhead boxP3(FOXP3)and cell apoptosis in transfected PBMC.Results Compared to the healthy group, 749 lncRNA and 230 coding messenger RNA(mRNA)genes were abnormally expressed.Interestingly,NR4A3 gene was down-regulated by 78%.While linc-pbc, which was about 20 000 bp downstream of NR4A3 gene, increased 2.56-fold. Then design siRNA for linc-pbc.After transfection, mRNA and protein levels of NR4A3 and FOXP3 were up-regulated.Conclusions By recruiting PRC2 complex, linc-pbc may increased the methylation level of NR4A3 gene promoter region,thus decreasing the expression of NR4A3 in PBC patients and reduced NR4A3 futher downregulated the expression of FOXP 3 and reduced the amounts of immunosuppressive Treg cells in peripheral blood and liver tissue, breaking the equilibrium state of immune tolerance, and promoted the occurrence and development of the disease.
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Objective To explore the expression and clinical significance of mismatch repair (MMR)protein and MLH1 promoter methylation testing in endometrial cancer(EC). Methods A total of 420 cases with EC diagnosed by the surgical pathology examination from the Department of Pathology of PLA General Hospital, MLH1,MSH2,MSH6 and PMS2 protein in EC were detected by immunohistochemistry and methylation-specific multiplex ligation-dependent probe amplification(MS-MLPA) testing. Results (1)Of the 420 tumor cases, the total expression loss rate of MMR protein was 34.5%(145/420), the expression loss rates of MLH1,MSH2,MSH6 and PMS2 protein were respectively 17.1%(72/420), 8.1% (34/420), 7.4%(31/420), 26.2%(110/420)and loss rates of MLH1 and PMS2,MSH2 and MSH6 were 16.7%(70/420), 6.2%(26/420). When there was a loss of MMR protein expression, any one or more protein expression deletions in MLH1, PMS2, MSH2 and MSH6, it could be Lynch syndrome related endometrial carcinoma(LS-EC). The expression loss rate of MMR protein in the poorly differentiated endometrioid adenocarcinoma was higher than that in the well differentiated endometrioid adenocarcinoma(P<0.05).(2) The expression loss rate of MMR and PMS2 protein had statistically significant between the endometrioid adenocarcinoma and non-endometrioid adenocarcinoma(P<0.01). The expression loss rate of MSH2 protein had statistically significant in the stage Ⅲ(P<0.01). Moreover, there were also significant differences in depth of myometrial invasion and lymph node metastasis between the expression loss rate of MMR protein (P<0.05).(3)The expression loss rate of MLH1 protein was 72 cases and 57 cases had MLH1 promoter methylation testing(excluding those who were not qualified for DNA testing). The positive rate was 47.4% (27/57). Therefore, these patients were sporadic endometrial cancer, not non-LS-EC. Conclusions MMR protein may be play an important role in the development of endometrial cancer and be indicated poor prognosis. Immunohistochemical staining and MLH1 promoter methylation detection may be play an important role in the screening of the LS-EC.
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Objective To explore the impacts of NPHP1 knockdown on the phenotype of Madin-Darby canine kidney (MDCK) cells. Methods The expression of NPHP1 in MDCK cells was knockdown by siRNA interference. Cells were divided into normal control group, negative control group and siRNA group. The cellular morphology and migration were observed by light microscope. The mRNA expressions and activities of matrix metalloproteinases 2 and 9 (MMP2 and MMP9) were detected by real time PCR and gelatin zymography. The mRNA and protein expressions of E-cadherin, β-catenin, zonula occluden-1 (ZO-1), ZO-1-associated nucleic acid binding protein (ZONAB) and α-smooth muscle actin (α-SMA) were analyzed by real time PCR, Western blotting and immunocytochemistry. Results Compared with those in normal control group, in siRNA group the mRNA expressions of E-cadherin, β-catenin and ZO-1 decreased, and MMP9, MMP2, α-SMA and ZONAB increased after interfering NPHP124 h (all P<0.05); the protein expressions of E-cadherin,β-catenin and ZO-1 decreased and ZONAB and α-SMA increased after 48 h (all P<0.05), and MDCK cells became elongated with enhanced migration capacity; siRNA cells had decreased expressions of E-cadherin and β-catenin on the membrane, but increased expression of ZONAB in cytoplasm and nucleoplasm after 72 h, and α-SMA was also observed in some interfered cells. Conclusions NPHP1 knockdown induces epithelial-mesenchymal transition in MDCK cells, and ZO-1/ZONAB signaling pathway was activated. These changes may associate with renal interstitial fibrosis of Nephronophthisis type I.
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Objective To investigate the predictive and prognostic value of genetic detection of Kirsten rat sarcoma viral oncogene homolog(KRAS),Fc gamma receptors (FCGR),cytochrome P450 3A5 (CYP3A5) and CYP1A1 in patients with metastatic colorectal cancer (mCRC) receiving C225 combined with CapeOx chemotherapy.Methods Twelve KRAS wild-type (WT) mCRC patients were selected receiving C225 (cetuximab) combined with CapeOx (capecitabine/oxaliplatin) chemotherapy.KRAS,FCGR,CYP3A5,CYP1 A1 gene mutation were detected before treatment.The expressions of phosphatase and tensin homolog deleted on chromosome ten (PTEN) were detected in colorectal cancer tissues and the corresponding adjacent tissues expression with immunohistochemical staining (SP method).The relationship between FCGR,CYP3A5,CYP1A1 mutation,and PTEN expression and survival time were analyzed.Results The mutation rates of FCGR,CYP3A5,and CYP1 A1 were 16.7％ (2/12),25％ (3/12) and 16.7％ (2/12) in 12 patients with KRAS WT,respectively.PTEN was expressed mainly in the nucleus in yellowish brown.The rates of expression in the tissue adjacent to the cancer and in the tumor tissue were 100％ (12/12) and 41.7％ (5/12),respectively.PTEN expression in tumor tissue was reduced or absent.The study indicated that the objective response rate [complete response + partial response (CR + PR)] was 80％ in patients whose KRAS,FCGR,CYP3A5,and CYP1A1 were all in wild type,the CR + PR rates in FCGR,CYP3A5,and CYP1A1 gene mutation group were 0,33.6％ and 50％.The objective response rates of patients with PTEN expression or null were 50％ and 37.5％,respectively (P ＜ 0.05).The progression free survival (PFS) with KRAS/FCGR/CYP3A5/CYP1A1 wild type or mutation were 15.56 or 8.12 months (P ＜ 0.05).The overall survival (OS) with wild type or mutation of KRAS/FCGR/CYP3A5/ CYP1A1 were 25.03 or 19.21 months(P ＜0.05).The PFSs of positive or negative expression of PTEN in patients were 9.13 or 7.87 months (P ＜0.05),and the OSs of positive or negative expression of PTEN in patients were 24.25 or 18.74 months (P ＜0.05).Conclusions FCGR,CYP3A5 and CYP1A1 mutations and PTEN expression null both have been associated with resistance to cetuximab in mCRC patients.FCGR,CYP3A5,CYP1A1 and PTEN can be served as the predictive biomarkers for the response to cetuximab.
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Objective@#To investigate the localization of HBXIP protein over-expression in gastric adenocarcinoma, and its prognostic significance.@*Methods@#HBXIP localization was detected by immunofluorescence in AGS gastric cancer cell line, and by immunohistochemical staining in 97 gastric adenocarcinomas, 41 adjacent non-tumor tissues and 13 gastric adenoma tissues. Correlation between HBXIP expression and clinicopathological features of gastric cancer patients was evaluated by Chi-square and Fisher′s exact tests. Overall survival rates were calculated using Kaplan-Meier method.@*Results@#HBXIP was mainly expressed in the cytoplasm of gastric cancer. The positive and strongly positive expression rates of HBXIP protein in gastric cancers were 68.0% (66/97) and 49.5% (48/97) respectively, and were significantly higher than those in adjacent non-tumor tissues(48.8%, 20/41; 36.6%, 15/41) or gastric adenomas(2/13, 1/13; all P<0.05). HBXIP expression correlated significantly with tumor differentiation and lymph node status (P=0.007; 0.041). Kaplan-Meier survival analysis showed that the overall survival rate was significantly lower in gastric cancer patients with high HBXIP expression (P=0.015).@*Conclusions@#HBXIP expression in gastric cancer is mainly expressed in cytoplasmic, and the expression level is closely related to the prognosis. HBXIP expression status may potentially be used as an important prognostic indicator for gastric cancer.
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Objective To investigate the role of tet methylcytosine dioxygenase 2 (TET2) in the regulation of transforming growth factor-β1 (TGF-β1) expression in human glomerular mesangial cells induced by high glucose.Methods Cultured human glomerular mesangial cells were divided into normal control group (5.5 mmol/L glucose) and high glucose group (30.0 mmol/L glucose) which was cultured for 12 h to 72 h.The gene expression of TET2 in mesangial cells were inhibited by small molecule chemical called SC1,and which were divided into high glucose group (30.0 mmol/L glucose+ DMEM),DMSO group (30.0 mmol/L glucose+0.1％DMSO) and SC1 group (30.0 mmol/L glucose+3 μmol/L SC1).The mRNA and protein expression of TGF-β1,TET1 to 3 and α-smooth muscle actin (α-SMA) was detected by quantitative real-time PCR and Western blotting.Methylation of CpG islands in the regulation region of TGF-β1 was detected by bisulfite sequencing PCR (BSP).The activity of mesangial cell proliferation was assessed by colorimetry of thiazolyl blue (MTT).Results Compared with normal control group,the mRNA and protein expression of TET2 in mesangial cells induced by high glucose was increased significantly in a time-dependent manner (all P ＜ 0.05),but the expression of TET1 and TET3 was not affected.Meanwhile methylation rate of 4 CG sites from 24 h to 72 h were decreased in the first exon of TGF-β1 (P ＜ 0.01),but not in the promoter.Compared with high glucose group,when the expression of TET2 was inhibited by SC1,the methylation rate of TGF-β1 was recovered evidently (P ＜ 0.05),the mRNA and protein expression of TGF-β1 and α-SMA was suppressed,and the proliferation of mesangial cells was decreased (all P ＜ 0.05).Conclusions Demethylation of the CpG island mediated by TET2 may play an important role in the expression of TGF-β1 and mesangial cell phenotype transformation induced by high glucose.
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ObjectiveTo investigate the expression and significance of Bax, cyclooxygenase-2 (COX2), and inhibitor of differentiation 1 (ID1) in gallbladder adenocarcinoma tissues. MethodsA total of 70 patients confirmed with gallbladder adenocarcinoma by pathological examination in the Department of Pathology, The First Hospital of Lanzhou University, from 2000 to 2013 were collected. Besides, 20 cases of high-grade intraepithelial neoplasia, 30 cases of low-grade intraepithelial neoplasia, and 20 cases of cholecystitis were selected as controls. The protein expression of Bax, COX2, and ID1 in the four groups was determined using the immunohistochemical SP method. Meanwhile, chi-square test was conducted on the data using fourfold table exact test, the correlations between Bax, COX2, and ID1 were determined by Spearman correlation analysis, and survival data were subjected to Kapan-Meier survival analysis. ResultsThe high-grade intraepithelial neoplasia group, low-grade intraepithelial neoplasia group, and cholecystitis group showed significantly different expression of Bax compared with the gallbladder adenocarcinoma group (all P＜0.05); the low-grade intraepithelial neoplasia group and cholecystitis group showed significantly different expression of COX2 and ID1 compared with the gallbladder adenocarcinoma group (all P＜0.05); the high-grade intraepithelial neoplasia group showed significantly different expression of COX2 and ID1 compared with low-grade intraepithelial neoplasia group and cholecystitis group (all P＜0.05). Survival analysis showed that patients with positive Bax had a significantly higher survival rate than those with positive COX2 and ID1 (both P＜0.05). COX2 was positively correlated with ID1 (r=0.329, P＜0.05). ConclusionBax, COX2, and ID1 may play a role in the development of gallbladder adenocarcinoma, and joint detection of Bax, COX2, and ID1 is of great significance for the prognosis of gallbladder adenocarcinoma.
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Objective To explore the lung developmental disorder of rats with gestational diabetes mellitus (GDM) via investigating the GDM rat fetal lung structures and expression of pulmonary surfactant proteins (SP)-B,SP-C,thyroid transcription factor (TTF)-1 and pleiomorphic adenoma gene like (PLAGL)-2.Methods Sprague-Dawley rats were used to construct the GDM model.Twenty GDM rats were used as GDM group and 20 normal pregnant rats as control group.Cesarean section was performed on day 21 of gestation and random blood sugar was detected,and fetal rats were counted and weighed.Ultrastructure of the fetal lungs was studied by transmission electron microscopy.Sixty fetal rats were selected randomly in each group,and 360 paraffin sections were made from fetal lungs.One hundred discontinuous paraffin sections were picked up in each group to observe morphological and structural changes under optical microscope.The other one hundred discontinuous paraffin sections were picked up in each group to detect the location and expression of SP-B,SP-C,TTF-1 and PLAGL-2 protein by immunohistochemistry.Nine fetal rats were selected randomly to detect the expression level of SP-B,SP-C,TTF-1 and PLAGL-2 proteins in fetal lung tissues by Western blotting.Twenty seven fetal rats were selected randomly to detect the mRNAs level of SP-B,SP-C,TTF-1 and PLAGL-2 by real-time quantitative polymerase chain reaction.Independent sample t-test was used for statistical analysis.Results The average random blood glucose level in GDM group was significantly higher than that in control group [(26.8± 2.8) vs (4.9± 0.5) mmol/L,t=-34.05,P=0.00].The average weight of fetal rats in GDM group was higher than that in control group [(5.6±0.6) vs (5.2±0.5) g,t=-1.97,P=0.03].Alveolar number (10.1 ± 1.6 vs 12.1 ± 1.3) and alveolar area [(986.9 ± 5.5) vs (1 257.3± 5.0) μ m2] in GDM group was less than that in control group (t=9.84 and 27.53,both P ＜ 0.05).Alveolar septum [(11.5±6.2) vs (9.9±4.3) μm] in GDM group was higher than that in control group (t=-2.17,P ＜ 0.05).Microvillus in type] cells were short and the number of lamellar bodies was significantly decreased in GDM group.SP-B,SP-C,TTF-1 and PLAGL-2 proteins were distributed in the cytoplasm in granular form.The average value of absorbance of SP-B,SP-C,TTF-1 and PLAGL-2 proteins in GDM group was 1.15±0.12,1.23±0.06,0.87±0.21 and 1.21 ±0.18 respectively;and that in control group was 1.22±0.05,1.31 ±0.14,1.12±0.09 and 1.33 ±0.07 respectively.The value in GDM group was lower than that in control group (t=2.40,2.35,4.89,and 2.77 respectively,all P ＜ 0.01).The expression level of SP-B,SP-C,TTF-1 and PLAGL-2 proteins in GDM group was 0.57± 0.09,0.45±0.03,1.50±0.04 and 1.11 ±0.04 respectively;and that in control group was 0.81 ±0.03,0.66±0.04,1.69±0.05 and 1.46±0.07 respectively.The value in GDM group was lower than that in control group (t=1 1.77,11.09,8.80 and 13.37,respectively,all P ＜ 0.01).The mRNA level of SP-B,SP-C,TTF-1 and PLAGL-2 in GDM group was 0.60±0.04,0.79±0.04,0.81 ±0.03 and 0.79±0.05 respectively;and that in control group was 1.06±0.19,1.03±0.24,1.03±0.18 and 1.02±0.19 respectively.The value in GDM group was lower than that in control group (t=6.80,2.98,3.54 and 3.54 respectively,all P ＜ 0.01).Conclusions The protein expression level of SP-B and SP-C in fetal lungs of GDM rats decreases obviously,possibly because of the down-regulation of the gene expression of TTF-1 and/or PLAGL-2.The pathological changes in fetal lungs of GDM rats might be associated with the descending level of SP-B and SP-C protein.
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Objective To investigate the effects of zinc fingers and homeoboxes 3 (ZHX3) silence on expressions of smad3, smad4 and RUNX2 in bone marrow mesenchymal stem cells (BMSCs). Methods ZHX3 low expression vector (ZHX3 silent group) was constructed and was transfected to rat BMSCs. Empty vector was transfected into BMSCs and was used as vehicle control group, and wild type BMSCs was used as the control group. The cell transfection rate was measured under a fluorescence microscope, and then the successful transfection was identified. The immunocytochemistry and immu?noblotting methods were used to detect the expression levels of smad3, smad4 and RUNX2. Results (1) Cells with BMSCs phenotype can be obtained by recovery culturing. (2) After transfection, the green fluorescent protein was found in ZHX3 si?lence group and vehicle control group. Blank control group showed no significant fluorescence. The expression level of ZHX 3 was significantly lower in ZHX3 silence group than that of vehicle control group. (3) Results of immunofluorescence asssay showed that the positive expressions of smad3 and smad4 were located in nucleus and cytoplasm, the positive expression of RUNX2 was mainly located in nucleus. Positive cells were observed in three groups. There was no significant difference in fluorescence intensity between the control group and the vehicle control group, but the fluorescence intensity was significant?ly lower in ZHX3 gene silence group than that of two control groups. (4) There were no significant differences in expressions of smad3, smad4 and RUNX2 betweem control group and the vehicle control group, but they were significantly higher than those of ZHX3 silence group(P < 0.05). Conclusion ZHX3 gene silence can delay vitro osteogenesis of BMSCs, which may play a role by the down-regulated expression levels of smad3, smad4 and RUNX2.
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Runx2 is a transcription factor belongs to RUNXX family, and it is one of the important factors involved in expression and participation in regulation of mammary specific genes in the mammary gland. The overexpression is also as-sociated with certain characteristics of breast cancer. This article aims to summarize the recent progress of the role that Runx2 plays in breast cancer.
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Objective To summarize the clinical and radiological features of DYT6 dystonia with mutations based on the data of our patient cohort as well as the report by others.Methods Clinical data of the 11 patients with DYT6 dystonia in Peking Union Medical College Hospital from June 2009 to May 2012 were retrospectively reviewed and analyzed.Clinical data included gender,onset age,initiative symptom of onset,the sites of involvemet,family history,etc.All patients were examined for brain MRI scan,6 patients were examined for DTI.Results Of the eleven gene-confirmed DYT6 dystonia patients,7 were male and 4 were female,with an onset-age ranged from 5 years to 36 years,the mean age of onset was 19.4years.Eight patients had a family history.There were 10 patients with early onset dystonia and only 1 patient with late onset dystonia.The most common site of onset was the neck (7/11),and the next was the right arm,1-5 body areas were affected at the time of neurological assessment,the average amount was 2.8,and the most frequently affected anatomical site was the neck (10/11),next came lower face,jaw and tongue.Among all the patients,6 patients presented with segmental dystonia,4 patients presented with focal dystonia,only 1 patient presented with generalized dystonia.All the patients with thanatos-associated protein domain-containing apoptosis-associated protein (THAP) domain affected had a family history,but the patients with the same mutant gene varied with clinical manifestation.Only 1 patients with non-THAP domain affected had a family history,but in most families,there were adult asymptomatic mutant gene carriers.Mutations within the THAP domain were associated with an earlier age of onset than non-THAP domain (17.3 and 21.8 years old).Routine MRI of all patients were normal and DTI of 6 patients showed that fractional anisotropy values in the bilateral sensorimotor area in DYT6 dystonia were reduced.A detailed description of a patient with TOR1A and THAP1 gene mutations was given.Conclusions Early onset dystonia is the main manifestation in patients with DYT6 dystonia in China.The most common site of onset is the neck,and the next is the right arm.The most frequently affected anatomical site is the neck,next come lower face,jaw and tongue.Laryngeal dystonia is absent.The patients with same mutant gene show high heterogeneity in the clinical manifestations,mutations within the THAP domain of THAP1 tend to manifest at an earlier age and higher penetration than mutations localized to non-THAP domain.Reduction of fractional anisotropy values indicates that the axonal integrity and coherence in the region of sensorimotor area is damaged in DYT6 dystonia.
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Objective To explore influence of JAK/STAT3 signaling pathway on the apoptosis of HCC cells.Methods DNA-vector-based RNAi approach silence was used to down-regulate STAT3 expression in Bel-7402 cells.According to the STAT3 cDNA sequence in the GeneBank database,the plasmid pGCsi.U6/neoRFP-STAT3 that was designed for expression of STAT3 siRNA was constructed and synthesized,and then transfected into the Bel-7402 cells with iipofectamine 2000.The apoptotic rate was measured with flow cytometry (FCM) and annexinV/PI apoptosis detection kit staining.The mitochondrial membrane potential (BΨm) was visualized by the JC-1 fluorescence staining and the inverted fluorescence microscope.Moreover,the expression of caspase-3 protein was analyzed by Western blotting.Results The apoptotic ratio of STAT3-siRNA group was (38.82 ± 0.88) ％,which was significantly higher than that in other group [control group (9.22 ± 0.38) ％,scramble-siRNA group (16.47 ± 1.04) ％,P ＜ 0.05].The mitochondrial membrane potential of STAT3-siRNA group observed by the JC-1 fluorescence staining was decreased significantly[(91.33 ± 1.78) ％] and [(89.90 ± 1.92) ％ vs (59.06 ± 1.89) ％,P ＜ 0.05].The Western blot results showed that the protein expression of active caspase-3 in STAT3-siRNA group was significantly higher than other groups (0.48 ± 0.05 vs 0.22 ± 0.04 and 0.26 ± 0.06,P ＜ 0.05).Conclusions STAT3 gene silencing significantly improves the apoptotic effect in the Bel-7402 ceils.
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Objective To explore the effects of Helicobacter pylori (H.pylori) on the expression of catalytic subunit of the DNA-dependent protein kinase (DNA-PKcs) and Ku70/Ku80 heterodimer in gastric mucosa epithelial cells in vivo and in vitro.Methods After treated with H.pylori for one,three,six,12 and 24 hours,the expressions of DNA-PKcs and Ku70/Ku80 heterodimer in gastric epithelial cells (GES) 1 and gastric adenocarinoma cells (AGS) were detected by Western blot.Mongolian gerbils were gavaged with H.pylori,and were sacrificed after infected for six and 12 months.The gastric mucosa tissues were taken for immunohistochemistry to detect the expressions of DNA-PKcs and Ku70/Ku80 heterodimer at protein level.The data were analyzed by t test and chi-square test.Results After H.pylori infection for one hour,the relative quantity of the expression of DNA-PKcs in GES-1 was 1.16±0.09,which was higher than that of non infected group (1.04±0.31) and the difference was statistically significant (t=4.67,P＜0.05).After infected by H.pylori for one,three,six,12 and 24 hours,the relative quantities of the expressions of Ku70/Ku80 heterodimer in GES-1 were 1.58±0.32,1.84±0.40,1.97±0.35,3.72±1.42 and 3.74±1.56,respectively,all were higher than that of non infected group (1.24±0.31) and the differences were statistically significant (t=3.57,4.20,5.03,8.11 and 8.14,all P＜0.05).The relative quantities of the expressions of Ku70/Ku80 heterodimer in AGS were 4.69 ± 0.87,3.67 ± 0.67,2.41±0.24,1.35±0.35 and 1.32±0.10 after H.pylori infected for one,three,six,12 and 24 hours,respectively,all were lower than that of no H.pylori infected group (4.84 ± 0.76) and the differences were statistically significant (t=34.13,27.68,19.81,4.47 and 5.69,all P＜0.05).In Mongolian gerbil models,DNA-PKcs did not express in H.pylori negative group (0/25),the total positive rate of H.pylori infected group was 98.1％ (53/54),the difference between the two groups was statistically significant (x2 =74.55,P＜0.01).The total positive rate of Ku70/Ku80 heterodimer in H.pylori negative group was 92.0％ (23/25) and in H.pylori infected group was 68.5％ (37/54),the difference between the two groups was statistically significant (x2=5.16,P＜0.05).Conclusion H.pylori infection affected cellular DNA damage repair through changing the expression of DNA-PKcs and Ku70/Ku80 heterodimer in gastric mucosa in vivo and in vitro,which may cause gastric mucosal lesions.
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Objective To explore the relationship among single nucleotide polymorphism (SNP) of excision repair cross-complementing 1 (ERCC1) gene,chemotherapy sensitivity and clinical outcomes of epithelial ovarian cancer (EOC) patients treated with platinum.Methods Six tag single nucleotide polymorphisms (tagSNP;rs11615,rs3212986,rs735482,rs3212955,rs12610134 and rs3212958) were chose from ERCC1 gene.The genotypes of 6 tagSNP were determined by Snapshot method in 220 EOC patients.Primary clinical outcomes parameter contained EOC patients'responses to platinum-based chemotherapy,progression-free survival (PFS) and overall survival (OS) were analysed.Results The rs11615 C/T SNP of ERCC1,CC,CT and TT genotype frequencies were 53.1％,45.6％,1.4％ in responders to platinum-based chemotherapy,while 52.0％,35.6％,12.3％ in non-responders,respectively,in which there was significant difference between the two groups(P =0.002).Compared with the patients with CC genotype,the patients carrying TT genotype had a significantly poor response to platinum-based chemotherapy (OR =6.22,95％ CI:1.12-34.42).Similarly,the genotypes frequencies distribution of rs11615 C/T SNP of ERCC1 was different between the recurrence and non-recurrence group,death and survival group (all P ＜ 0.05).Kaplan-Meier survival analysis showed that the genotypes frequencies distribution of rs11615 C/T SNP of ERCC1 was associated with PFS and OS(P ＜ 0.01) of EOC patients.Cox's multivariate analysis suggested that patients with TT genotype had a shorter PFS (HR =2.19,95 ％ CI:1.14-4.22,P =0.009) and OS (HR =2.22,95 ％ CI:1.06-4.64,P =0.021) compared with those carrying CC genotype [adjusting for age,International Federation of Gynecology and Obstetrics (FIGO) stage,pathological type,grade and tumor residual size].The genotypes frequencies distribution of rs3212986,rs735482,rs3212955,rs12610134 and rs3212958 SNP of ERCC1 did not show the significant difference between the responders to platinum-based chemotherapy and non-responders.The other 5 tagSNP may not be associated with the PFS and OS of EOC patients (all P ＞ 0.05).Conclusion The rs 11615 SNP of ERCC1 may become a valuable prognostic biomarker for EOC patients treated with platinum-based chemotherapy.
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Objective To investigate the expressions of signal transducer and activator of transcription factor 3 (STAT3),Bcl-2,and matrix metalloproteinase 2 (MMP2) in colorectal adenomas and adenocarcinomas,and the relationship of those factors with the colorectal clinicopathological features and prognosis of intestinal adenocarcinomas,and explore their roles in invasion,metastasis,and prognosis of a colorectal cancer.Methods The samples were selected from Dongyang City People's Hospital from January 2011 to January 2012,including 50 cases of paraffin-coded colorectal mucosas with chronic inflammation,50 cases of paraffincoded colorectal adenomas,and 100 cases of paraffin-coded colorectal adenocarcinomas (35 cases with metastasis and 65 cases without distant metastasis).Envison two method was used to detect the expressions of STAT3,Bcl-2,and MMP2 in each sample.Results The expressions of STAT3,Bcl-2,and MMP2 in colorectal adenomas and adenocarcinomas were significantly higher than those in colorectal mucosas with chronic inflammation (P ＜ 0.05).The expressions of STAT3 and MMP2 in colorectal adenocarcinomas with distant metastasis were significantly higher than that those without distant metastasis (P ＜ 0.05).The expression of BCL-2 had no significant difference between colorectal adenocarcinomas with and without distant metastasis (P ＞ 0.05).Conclusions STAT3,Bcl-2 and MMP2 were associated with occurrence and development of colorectal carcinoma.STAT3 was associated with distant metastasis of colorectal carcinoma and might indicate a poor prognosis.STAT3 might be used as a candidate clinical sign for recurrence,metastasis,and poor survival prognosis of colorectal adenocarcinoma.
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Objective To study the relationship between Nogo-B and transforming growth factor-β1 (TGF-β1)/Smad2 signaling pathway in mice models of hepatic fibrosis.Methods Twenty four healthy male ICR mice were divided into two groups,with 6 in the control group and 18 in the model group.Mice in the model group were further divided into three subgroups according to different time points:subgroups of 4,8 and 12 weeks,with 6 mice in each subgroup.Hepatic fibrosis of mice was induced by subcutaneous injection of carbon tetrachloride (CCl4).The histopathologic changes of the liver were observed by optical microscope using hematoxylin-eosin and Masson trichrome stainings of the liver tissues.Expressions of Nogo-B,Smad2 and TGF-β1 mRNA and proteins in liver were detected by reverse transcription-polymerase chain reaction (RT-PCR),Western blot and immunohistochemistry assays,respectively.Means among groups were compared by univariate analysis of variance.Results The hepatic fibrosis models were successfully induced by CCl4 injection.The expressions of two subtypes of Nogo-B,Nogo-B1 and Nogo-B2 mRNA in normal livers were 0.140±0.050 and 0.104±0.023,but both significantly increased in the livers of mice in the 12 week model subgroup (1.054±0.040 and 0.500±0.057,F=431.41 and 135.46,respectively; both P＜0.01).The Nogo-B protein was mainly expressed in nonparenchymal cells of the liver,and was hardly expressed in hepatocytes.Linear correlation analysis showed that the expressions of Nogo-B mRNA and proteins were positively correlated with Smad2 and TGF-β1 mRNA and proteins (all P＜0.01),which were considered to participate in the signaling pathway of hepatic fibrosis.Conclusion Nogo-B might play a role in the development and progression of hepatic fibrosis by participating in TGF-β1/Smad2 signaling pathway.
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Objective To detect the excision repair cross-complementing gene 1 (ERCC1) and thymidylate of the acid synthase (TS) in non-small cell lung cancer (NSCLC) and its adjacent tissue,and investigate the relationship of the expression of ERCC1 and TS with the clinical characteristics of NSCLC and prognosis for NSCLC individual therapy to provide experimental basis.Methods The protein expression levels of ERCC1 and TS in 50 cases of postoperative NSCLC cancer and adjacent tissue were detected by immunohistochemical method and the relationship among the expression of ERCC1,TS,and overall survival of patients with NSCLC Phase (OS),disease progression time (TTP),the median OS,and median TTP was analyzed.Results (1)There was an obvious difference between the expression of ERCC1,TS in cancer and paraneoplastic tissue of NSCLC,which had statistically significance (64.00％ vs 20.00％,x2 =19.87,P ＜ 0.01 ;48.00％ vs 24.00％,x2 =6.25,P ＜ 0.05) ; (2)The continued investigation in the patients who received postoperative cisplatin or carboplatin chemotherapy showed that the 0S of negative expression of ERCC1 was significantly longer than the positive one (19.10 vs 10.00 months;x2 =8.133,P =0.002),so was median TTP (15.30 vs 9.00 months; x2 =7.410,P =0.003).The median 0S of the negative expression of TS,was significantly longer than the positive one (17.80 vs 11.00 months,x2 =7.001,P =0.008),so was median TTP (11.40 vs 6.80 months; x2 =5.884,P =0.026).Conclusions ERCC1 and TS protein may become sensitive predictors of platinum chemosensitivity for NSCLC patients ; the detection of combined with ERCC1 and TS would contribute to the selection of individualized treatment programs for NSCLC.