Objective:To construct pEGFP-C1-Smad4 expression vector and to observe the influence of Smad4 over-expression on the proliferation of human gastric cancer SGC7901 cells and its relationship with nuclear factor kappa B (NF-?B). Methods:Recombinant expression vector pEGFP-C1-Smad4 was constructed and was used to transfect human gastric cancer SGC7901 cells. EGFP expression in transfected cells was detected by fluoroscopy. Smad4 and NF-?B expression in transfectant was examined by Western blotting. Effect of Smad4 over-expression on activation of NF-?B and proliferation of transfected SGC7901 cells were examined by electrophoretic mobility shift assay (EMSA) and MTT assay,respectively. Results:Expression of EGFP in transfected SGC7901 cells was observed under fluorescence microscope. Smad4 was over-expressed in transfected SGC7901 cells,accompanied by down-regulation of NF-?B p65 expression in the tranfectants. EMSA and MTT demonstrated that Smad4 over-expression significantly inhibited the activation of NF-?B and the proliferation of SGC7901 cells (P

Objective To explore the regulatory function of over-expression of NOTCH-1 in TNF?-induced apoptosis of gastric carcinoma carcinoma BGC-823 cells.Methods BGC-823 cells were infected with retrovirus recombined with intracellular domain of NOTCH-1.The cell clones,into which objective genes had been transfected,were obtained after screening with G-418.Before and after incubation with TNF?,MTT assay,flow cytometry,Western blot and EMSA were used to detect cell growth,apoptosis,expressions of NOTCH-1,NF-?B and caspase-3 activity.Results There was stronger expression of NOTCH-1 and its target gene HES-1 in the cells transfected with NOTCH-1 gene than that of control.After TNF? treatment,the rates of killing and apoptosis in NOTCH-1-transfected cells were all lower than those in control cells.Moreover,we showed over-expression of NOTCH-1 suppressed caspase-3 activation induced by TNF?.However,TNF?-induced activation of NF-?B was not affected by over-expression of NOTCH-1.Conclusion Over-expression of NOTCH-1 significantly protects BGC-823 cells from apoptosis and growth suppression induced by TNF?.This effect is mediated,at least in part,through inhibition of caspase-3 activation independent of NF-?B.

Objective: To elucidate the relationship between Notch3 expression and chemosensitivity of human colon carcinoma cell line SW620 to topotecan. Methods: Notch3 siRNA was transfected into SW620 cells, and the expression of Notch3 in SW620 cells was examined by Western blotting. After transfected with Notch3 siRNA for different time peri-ods, SW620 cells were further treated with topotecan, and the proliferation of SW620 cells was detected by MTT assay; the apoptosis of SW620 cells was detected by Hoechst 33342 staining and flow cytometry. Caspase-3 activation in SW620 cells was examined by caspase-3 activation kit. Results: Notch3 siRNA transfection remarkably inhibited Notch3 protein expression in SW620 cells. The IC_(50) of topotecan in Notch3 siRNA-transfected group was significantly decreased compared with that in the Ctrl siRNA group (P <0.05). Silence of Notch3 expression in SW620 cells by Notch3 siRNA remarkably promoted apoptosis (P < 0.05) and caspase-3 activation (P < 0.05) of SW620 cells induced by topotecan. Conclusion: Notch3 down-regulation by siRNA in SW620 cells can enhance the chemosensitivity cells to topotecan.

Objective To construct the recombinant retroviruses vector PCLNRX-ICN and to explore over-activation Notch1 on the growth of human colon carcinoma cells HT-29.MethodsICN(intracellular domain of Noctch1) genes were inserted into retrovirus expression vector pCLNRX.The expression vector pCLNRX-ICN and packaging vector pCL-10A1 were co-transfected into 293 package cells.The recombinant retroviruses were used to infect HT-29 cells.After being infected,proliferation of HT-29 cells was observed by MTT assay.The expression of c-Myc and ?-catenin detected by RT-PCR and Western Blot.ResultsRecombinant retrovirus vector pCLNRX-ICN was successfully constructed.Overactivation of Notch1 by over-expressing exogenous ICN significantly inhibited the growth of HT-29 cells,and down-regulated ?-catenin,a key regulator of Wnt signaling,in protein level but not in mRNA levels.However,the mRNA or protein levels of c-Myc were not affected by overactivation of Notch1.ConclusionOver-activated Notch1 signaling could inhibit the growth of HT-29 cells partly through down-regulation of Wnt signaling independent of c-Myc inhibition.

Objective To analyze the clinical manifestation and genetic testing in a patient with Adams-Oliver syndrome (AOS) and summarize clinical and genetic characteristics of the dedicator of cytokinesis (DOCK) 6 gene related AOS through reviewing related references.Methods Information of the proband who was hospitalized in Affiliated Children Hospital of Capital Institute of Pediatrics in October 2016 and her family members as well as their DNA samples were collected.The gene sequencing was performed using next generation sequencing technology.Using "Adams-Oliver syndrome"and "DOCK6" as key words,the relevant articles were searched from the Pubmed,China National Knowledge Internet and Wanfang databases and reports of 19 cases were reviewed.Results The proband is an eight months old girl.She presented with severe developmental delay,terminal transverse limb defects and visual loss after birth,and then suffered from tonic seizures and myoclonic seizures at two months old.By physical examination she was found to have esotropia and visual loss.The distal phalanx and nail of the right second-fourth fingers were absent,while the phalangette of the left second-fourth fingers and bilateral distal phalanges of toes were short with small nails attachment.Thyroid function test showed hypothyroidism.The ocular fundus examination showed the residual vitreous artery in the left eye and the retinal pigment degeneration in the right eye.CT scan showed multiple bilateral periventricular calcification and cranial magnetic resonance imaging showed bilateral periventricular lesion.Two heterozygous mutations were identified in DOCK6 gene:one was a known pathogenic mutation (p.L1064Vfs*60),and the other was a novel splice site mutation (c.873+ 1G＞A).By analyzing this case and reported 19 cases,the common performances of DOCK6 gene related AOS included terminal transverse limb defects (20/20),aplasia cutis congenita (18/20),ocular abnormalities (13/20),seizures (12/20),mental retardation (12/20),microcephaly (10/20),cardiovascular malformations (10/20),intrauterine growth retardation (6/20).The mutation of the DOCK6 gene was found to be dominated by frameshift mutation and splice site mutation.Conclusions If either terminal transverse limb defects or aplasia cutis congenita was detected in a patient,AOS should be under consideration.In addition,autosomal recessive inheritance,nervous system and eyes involvement will further indicate DOCK6 gene related AOS.

[Abstract] Objective: To investigate the effect of lncRNA MAFG-AS1/ miR-11181-3p/GLG1 axis on cell migration, invasion and aerobic glycolysis of gastric cancer (GC) cells and its possible mechanism. Methods: AGS, a GC cell line with relatively high expression of MAFG-AS1, was selected as the study object. qPCR was used to detect RNA expression levels of MAFG-AS1, miR-11181-3p and GLG1. Transwell and glycolysis analysis were used to investigate cell migration, invasion and aerobic glycolysis. Bioinformatics analysis and Dual luciferase reporter gene assay were used to analyze the interaction among MAFG-AS1, miR-11181-3p and GLG1. Results: Knockdown of MAFG-AS1 significantly up-regulated miR-11181-3p and down-regulated GLG1 expression (both P<0.01), and significantly inhibited migration, invasion and aerobic glycolysis of GC cells (all P<0.01). Luciferase reporter gene assay confirmed that MAFG-AS1 competitively sponged miR-11181-3p (P<0.01). Inhibition of miR-11181-3p or overexpression of GLG1 partially reversed the inhibitory effect of MAFG-AS1 knockdown on GC cell migration, invasion, and aerobic glycolysis (all P<0.05 or P<0.01). Conclusion: MAFG-AS1 promotes cell migration, invasion and aerobic glycolysis of GC cells via miR-11181-3p/GLG1 axis, and may be a potential molecular target for GC diagnosis and therapy.