Fibronectin containing extra domain A (EDA+ FN),a functional glycoprotein participating in several cellular processes,correlates with chronic liver disease.Herein,we aim to investigate the expression and secretion of EDA+ FN from hepatocytes in nonalcoholic fatty liver disease (NAFLD) and the underlying mechanisms.Circulating levels of EDA+ FN were determined by ELISA in clinical samples.Western blotting and flow cytometry were performed on L02 and HepG2 cell lines to analyze whether the levels of EDA+ FN were associated with endoplasmic reticulum (ER) stress-related cell death.Circulating levels of EDA+ FN in NAFLD patients were significantly higher than those in control subjects,and positively related with severity of ultrasonographic steatosis score.In cultured hepatocytes,palmitate up-regulated the expression of EDA+ FN in a dose-dependent manner.Conversely,when the cells were pretreated with 4-phenylbutyrate,a specific inhibitor of ER stress,up-regulation of EDA+ FN could be abrogated.Moreover,silencing CHOP by shRNA enhanced the release of EDA+ FN from hepatocytes following palmitate treatment,which was involved in ER stress-related cell damage.These findings suggest that the up-regulated level of EDA+ FN is associated with liver damage in NAFLD,and ER stress-mediated cell damage contributes to the release of EDA+ FN from hepatocytes.

Objective To develop an effective real time PCR method for genotyping mitochondrial aldehyde dehydrogenase-2(ALDH2)Glu504Lys polymorphism based on the hydrolysis probes.Methods The Mg~(2+)and probe concentration were optimized,the precision and sensitivity were also checked.The genotypings by this method were confirmed by the direct sequencing of amplified PCR products.Results The optimized Mg~(2+)and probe concentration were 2.5 mmol/L and 1.0 ?mol/L,respectively.The inter- group(n=20)and intra-group(n=20)CVs of Ct were 1.38% and 1.48 %,respectively.The method could detect human DNA in the range of 5.0?10~2-5.0 ?10~6 pg per 25 ?l reaction.The results from 150 individuals by this genotyping method are in full concordance with that by direct PCR products sequencing.Conclusion The combined merits of reliability,flexibility and simplicity should make this method suitable for routine clinical testing and cost-efficient large-scale genotyping.

Objective To compare the clinical effect of minimally invasive surgery transforaminal lumbar interbody fu-sion(MIS-TLIF)and posterior lumbar interbody fusion(PLIF)in the treatment of recurrent lumbar intervertebral disc hernia-tion(LIDH). Methods Twenty-nine patients with recurrent LIDP were selected from May 2014 to May 2016 in Weifang Peo-ple's Hospital and the clinical data were analyzed retrospectively. Thirteen patients were given MIS-TLIF(MIS-TLIF group) and sixteen patients were given PLIF(PLIF)group. The operative incision length,intraoperative bleeding volume,postoperative drainage volume,hospitalization time and complications were compared between the two groups. The lumbar function was evalu-ated with the Japanese Orthopaedic Association(JOA)score standard,and the clinical effect was compared between the two groups according to the modified Macnab standard one year after treatment. Results The operativeincision length,intraopera-tive bleeding volume,postoperative drainage volumeand hospitalization time in MIS-TLIF group were significantly less than tho-sein PLIF group (P < 0. 05). The preoperative JOA score of lumbar function in PLIF group and MIS-TLIF group was 7. 9 ± 1. 9 and 8. 0 ± 1. 6 respectively,it was 24. 0 ± 2. 7 and 24. 2 ± 2. 5 respectively at one year after treatment,there was no significant-difference in the JOA score between the two groups before and one year after operation (P > 0. 05). The JOA score atone year after operation was significantly higher than that before operation in the two groups (P < 0. 05). According to the modified Macnab standard one year after treatment,the fineness rate of the patients in PLIF group was 87. 50％(14 / 16),the fineness rate of the patients in the MIS-TLIF group was 84. 62％(11 / 13). There was no significant difference in the fineness rate be-tweenthe two groups (χ2 = 1. 380,P > 0. 05). The incidence of postoperative complications in the MIS-TLIF group and PLIF group was 7. 7％(1/ 13)and 6. 3％ (1/ 16)respectively,there was no significant difference in the incidence of postoperative complications between the two groups (χ2 = 0. 020,P > 0. 05). There were 8 cases (61. 5％)with gradeⅠfusion and 5 cases (38. 5％)with gradeⅡfusion in MIS-TLIF group,there were 9(56. 3％)with gradeⅠfusion and 7(43. 8％)with gradeⅡfu-sion,there was no significant difference in the constituent ratio with gradeⅠandⅡfusion between the two groups (χ2 = 0. 080, P >0. 05). Conclusion MIS-TLIF in treatment of recurrent LIDH has the advantages of less incision,less intraoperative bleed-ing,less postoperative drainage and shorter hospitalization time;and the clinical effect of MIS-TLIF is similar to that of PLIF.

Animals ; Brain-Derived Neurotrophic Factor ; Depression/drug therapy* ; Fluoxetine ; Mice ; Stress, Psychological ; Transcranial Magnetic Stimulation

OBJECTIVETo investigate the effect of survivin antisense oligodeoxynucleotide (ASODN) on proliferation and apoptosis of human malignant melanoma cells.

METHODShMMC A375 colonies in log growth phase were collected and divided into control group (C, without transfection), sense chain group [SC, transfected with 600 nmol/L survivin sense oligodeoxynucleotide (ODN)], mismatch chain group (MC, transfected with 600 nmol/L survivin mismatch sense ODN), liposome group (L, treated with liposome), antisense chain group (AC, transfected with survivin ASODN, and subdivided into AC 200, 400, 600 nmol/L subgroups) according to the random number table. Transfection result was observed under inverted fluorescence microscope. Inhibition rate of cell proliferation was calculated after determination of cell viability with MTT method. Cell cycle and apoptosis rate were detected with bi-variable flow cytometry. Expression of survivin protein was determined with Western blot. Activity of caspase-3 was assessed with kinase method. Data were processed with analysis of variance.

RESULTS(1) Cell transfection rates in SC, MC, AC 600 nmol/L groups were all above 80%. (2) Compared with those in SC group [(5.23 +/- 0.25)%], MC group [(5.09 +/- 0.13)%] and L group [(4.70 +/- 0.45)%], inhibition rates of cell proliferation in AC 200, 400, 600 nmol/L groups 24 hours after transfection [(10.30 +/- 0.56)%, (16.69 +/- 0.58)%, (24.67 +/- 0.67)%] were significantly increased (F = 746.91, and P values all below 0.05). As time after transfection went on, proliferation inhibition rate was increased obviously. (3) Apoptosis rate in AC 200, 400, 600 nmol/L groups 24 hours after transfection was respectively (13.5 +/- 1.9)%, (20.1 +/- 1.5)%, (32.1 +/- 2.9)%, which were significantly higher than those in C, SC, MC, and L groups [(6.5 +/- 0.6)%, (5.6 +/- 0.7)%, (6.4 +/- 1.0)%, (6.5 +/- 1.3)%, F = 139.9, P values all below 0.05]. Cells in AC group were blocked in G2/M stage. (4) Compared with those in C group, expression amount of survivin protein decreased, and caspase-3 activity obviously increased (F = 63.1, P values all below 0.05) in AC group. No significant difference in caspase-3 activity between SC, MC, L groups and C group was observed (F = 0.512, P values all above 0.05).

CONCLUSIONSSurvivin ASODN can inhibit the proliferation of hMMC A375 in a concentration-time dependent manner, and it induces G2/M stage block and promotes its apoptosis.

Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; Inhibitor of Apoptosis Proteins ; Melanoma ; metabolism ; pathology ; Microtubule-Associated Proteins ; genetics ; pharmacology ; Oligodeoxyribonucleotides, Antisense ; pharmacology ; Transfection

OBJECTIVETo investigate the expression and distribution of mast cell tryptase (MCT) in scar, and to discuss the different MCT gene expression in keloid, hypertrophic scar and normal skin.

METHODS20 samples of keloid, 20 samples of hypertrophic scar and 20 samples of normal skin were collected. The distribution of MCT was investigated by immunofluorescence histochemistry, and the MCT mRNA expression was detected by Relative Quantification real-time fluorescent PCR.

RESULTSMCT gene was mainly located in the collagen fiber bundles of the scar, especially in the superficial layer of scar. MCT mRNA expression was significantly higher in keloid than that in hypertrophic scar and normal skin (P < 0.01). Averagely, the MCT gene expression in keloid was 2.5 times and 5.4 times of that in hypertrophic scar and normal skin.

CONCLUSIONSMCT gene may play a role in the pathogenesis of scar.

Adolescent ; Adult ; Cicatrix, Hypertrophic ; metabolism ; pathology ; Humans ; Keloid ; metabolism ; pathology ; RNA, Messenger ; genetics ; Skin ; metabolism ; pathology ; Tryptases ; genetics ; metabolism ; Young Adult

OBJECTIVETo explore the related risk factors and genetic susceptibility of coronary heart disease (CHD) in old Chinese Han patients.

METHODSIn a case-control study, we enrolled 246 patients with CHD for cases, 185 cases without CHD for control. Correlation of CHD with 15 risk factors, including sex, age, smoking, drinking, hypertension, diabetes mellitus, hyperlipidemia, homocysteine (HCY), N-terminal pro-brain natriuretic peptide (NT-pro-BNP), high-sensitivity C-reactive protein (Hs-CRP), antithrombin III (ATII), cholesterol (CHOL), triglycerides (TG), high density lipoprotein (HDL) and low density lipoprotein (LDL) were analyzed by a logistic regression analysis. Meanwhile, facilitated by USA Sequenom high-throughput single nucleotide polymorphisms (SNP) genotyping systems, 10-Plex Genes, 11 SNPs were investigated.

RESULTSAging, hypertension, lower ATIII were major risk factors for CHD (P < 0.05). Platelet glycoprotein GP1BA rs2243093 (-5T/C), angiotensin-converting enzyme ACE rs4332 (547C/T) and ATIII rs2227589 (893C/T) were associated with CHD in old Han Chinese patients. Mutant genotype CC of rs2243093 (-5T/C) compared with T + AT, P was 0.029 (OR = 3.41, CI: 1.19-9.75). Heterozygous TC of rs4332 (547C/T) was compared with CC + TT, P is 0.003 (OR = 0.56, CI: 0.38-0.82). T-allele carrier CT + CT of rs2227589 (893C/T) was compared with wild genotype CC, P = 0.003 (OR = 1.79, CI: 1.22-2.63).

CONCLUSIONOur study demonstrated that aging, hypertension, lower ATIII were the major risk factors of CHD. Three mechanisms associated with platelets, anti-coagulation system, the renin-angiotensin system were involved in coronary heart disease in the elderly.

Age Factors ; Aged ; Aged, 80 and over ; Antithrombin III ; metabolism ; Asian Continental Ancestry Group ; genetics ; Case-Control Studies ; Coronary Disease ; ethnology ; genetics ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Hypertension ; Male ; Middle Aged ; Polymorphism, Single Nucleotide ; Risk Factors

Objective To construct an aneurismal model with estrogen deficiency and investigate the mechanism of estrogen deficiency in the formation and development of intracranial aneurysm. Methods Female Wistar rats were randomly divided into experimental, sham-operative and blank control groups (n=10). Rats in the experimental group were ovariectomized and those in the sham-operative group were removed the adipose issue nearby the ovary only; while the rats in the blank control group were done nothing. Two weeks after the ovariectomized or sham operation, elastase dropped around the right external carotid artery and the crotch of the carotid artery and the carotid artery was ligated by two lines at 1.5 mm far from the crotch, and then sheared between the two lines to successfully induce the aneurysm. At 6 weeks of the successful construction of aneurysm model, the estrogen was detected and the aneurysm was harvested for pathological staining. Results The experimental group showed a lower estrogen level (105.00±12.96 pmol/L) than the sham-operative group (178.50±25.96 pmol/L) and the blank control group (180.40±18.70 pmol/L, P<0.05). Aneurismal length dilatation rates in the experimental group and the sham-operative group were (131.31±6.63)% and (109.90±3.44) %, respectively (P<0.05). Aneurismal diameter dilatation rates in the experimental group and sham-operative group were (125.10±5.49) % and (106.82±2.49) %, respectively (P<0.05). Conclusion Estrogen deficiency may promote the formation and development of the intracranial aneurysm. This experiment provides a simple model for investigating the relationship between estrogen deficiency and aneurysm development.

OBJECTIVETo investigate the value of preoperative barium contrast examination for the diagnosis and operative planning in gastric cancer.

METHODSClinical data of 229 gastric cancer patients were analyzed retrospectively. Lesions were divided into three parts: the cardiac, the body, and the antrum. The diagnostic accuracy of localization and the extent of tumor between gastroscopy alone and gastroscopy plus barium contrast were compared with the results of surgical findings.

RESULTSThe diagnostic accuracy of localization and the extent of tumor for gastroscopy in the cardiac, the body and the antrum cancers were 100% and 78.4%, 94.6% and 86.5%, 98.1% and 84.6%, respectively, while for gastroscopy plus barium contrast were 100% and 84.8%, 100% and 91.9%, 99.0% and 90.4%, respectively. The diagnostic accuracy of both the localization and the extent of tumor were not significantly different between gastroscopy alone and gastroscopy plus barium contrast (P>0.05). Diagnostic accuracy of the length of esophagus infiltrated by cardiac cancer in gastroscopy was 60.6%, while in gastroscopy plus barium contrast was 90.9%, which was significantly different (P<0.05). Gastroscopy plus barium contrast was more accurate in predicting the possibility of thoracotomy in cardiac cancer infiltrating the lower esophagus.

CONCLUSIONSIt is necessary to perform preoperative barium contrast examination in cardiac cancer patients, so as to identify whether the lower esophagus is infiltrated and to measure the length of lesion, which can provide evidences for making a decision of thoracotomy. For gastric body and antrum cancer, there is no indication for barium contrast examination if gastroscopy findings are satisfied.

Adenocarcinoma ; diagnostic imaging ; pathology ; surgery ; Barium ; Contrast Media ; Female ; Humans ; Male ; Middle Aged ; Radiography ; Retrospective Studies ; Stomach Neoplasms ; diagnostic imaging ; pathology ; surgery

OBJECTIVETo investigate the effects of silencing ADP-ribosylation factor 6 (Arf6) on the proliferation, migration, and invasion of prostate cancer cell line PC-3 and the possible molecular mechanisms.

METHODSThree Arf6-specific small interfering RNA (siRNA) were transfected into cultured prostate cancer cell line PC-3. Arf6 expression was examined by real-time PCR and Western blotting. MTT assay, wound healing assay, and Transwell migration and invasion assay were used to observe the effect of Arf6 silencing on the proliferation, migration, and invasion ability of PC-3 cells. The levels of phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2), ERK1/2, p-AKT, AKT and Rac1 were detected by Western blotting.

RESULTSTransfection of siRNA-3 resulted in significantly decreased Arf6 mRNA and protein expression with inhibition rates of (91.88±3.13)% and (86.37±0.57)%, respectively. Arf6 silencing by siRNA-3 markedly suppressed the proliferation, migration and invasion of PC-3 cells and reduced the expression levels of p-ERK1/2 and Rac1.

CONCLUSIONSilencing of Arf6 efficiently inhibits the proliferation, migration, and invasion of PC-3 cells in vitro, and the underlying mechanisms may involve the down-regulation of p-ERK1/2 and Rac1.

ADP-Ribosylation Factors ; genetics ; metabolism ; Cell Line, Tumor ; Cell Movement ; Down-Regulation ; Humans ; Male ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Neoplasm Invasiveness ; Prostatic Neoplasms ; pathology ; RNA Interference ; RNA, Messenger ; genetics ; metabolism ; RNA, Small Interfering ; genetics ; Real-Time Polymerase Chain Reaction ; Transfection ; Wound Healing ; rac1 GTP-Binding Protein ; metabolism