[ ABSTRACT] AIM: To investigate the effect of miR-496 over-expression on the growth and metastasis of colon cancer cells and its molecular mechanism .METHODS:The proteins interacting with miR-496 were screened by bioinfor-matic method.The levels of miR-496, CTNNB1 mRNA and β-catenin protein in colon cancer cell lines , HT29, HCT116 and SW480, and normal colonic epithelial cell line NCM 460 were detected by real-time PCR and Western blot .HT29, HCT116 and SW480 cells were transfected with miR-496 mimics using Lipofectamine 2000 and named as HT29-miR-496 mimics, HCT116-miR-496 mimics and SW480-miR-496 mimics cells, respectively, and the cells transfected with the scramble served as negative control .The cell viability, lactate dehydrogenase (LDH) leakage, and colony formation and metastatic abilities were determined by MTT assay , LDH assay, colony formation assay and Transwell method , respective-ly.The promoter activity of miR-496 was measured using luciferase reporter gene assay .The protein levels of β-catenin, eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1), p-4E-BP1, low-density lipoprotein receptor-related protein 6(LRP6), p-LRP6, MMP-7, MMP-9, MMP-13 and TIMP-2 were monitored by Western blot.RESULTS:Endog-enous miR-406 interacted with β-catenin was found in the colon cancer cells .Low miR-496 expression in the HT29, HCT116 and SW480 cells and high miR-496 expression in NCM460 cells were detected.In contrast, high β-catenin ex-pression was found in the HT29, HCT116 and SW480 cells and low β-catenin expression was observed in the NCM460 cells.Compared with control group , the cell viability, colony formation rate and the number of metastatic cells remarkably decreased in the HT29-miR-496 mimics, HCT116-miR-496 mimics and SW480-miR-496 mimic cells (P<0.05).The promoter activity of miR-496 was significantly increased in colon cancer cells transfected with miR-496 mimics, and was 1.75, 2.04 and 1.61 times as high as control group .miR-496 over-expression inhibited β-catenin levels, and p-4E-BP1 and p-LRP6 protein levels were also reduced .siRNA-or over-expressed miR-496-mediated β-catenin down-regulation in-hibited MMP-7 and MMP-9 expression , but promoted TIMP-2 expression .CONCLUSION:The expression level of miR-496 in the colon cancer cells is low , but in the normal colonic epithelial cells is high .miR-496 over-expression inhibits the protein levels of MMP-7 and MMP-9, and promotes the protein expression of TIMP-2 via inhibiting Wnt/β-catenin path-way, thus suppressing malignant phenotype in the colon cancer cells .

Subacute thyroiditis can cause destruction of thyroid follicles and subsequent transient thyrotoxicosis.In cases of simultaneous occurrences of subacute thyroiditis and Graves' disease,the former may be missed and thus may further exacerbate thyrotoxicosis.Herein,we report in detail a case with abrupt onset of thyrotoxic heart disease when taking anti-thyroid medications,in order to call attention to the diagnosis and treatment of concomitant Graves' disease and subacute thyroiditis.

Objective To investigate the effects of paeonol on renin-angiotensin system (RAS) occurred on the development of ventricular remodeling after acute myocardial infarction (AMI) in rats. Methods The left anterior descending coronary artery was ligated to establish the model of AMI in male SD rats. Six groups were set up:sham-operation group, AMI model group, captopril control group, paeonol low dose group (6 mg/kg), paeonol middle dose group (9 mg/kg) and paeonol high dose group (12 mg/kg). Rats were given treatment for 4 weeks after the AMI model was established. HE staining was used to observe changes of myocardial tissue. Real-time PCR was used to detect the mRNA levels of angiotensinogen (AGT), angiotensin Ⅱ receptor type1(AGTR1) and endothelin (ET)-1 of six groups. Western blot assay was used to detect the protein levels of peptidyl-dipeptidase A (ACE), angiotensin Ⅱ(Ang)-Ⅱand AGTR1 in six groups. Results The transcription of AGT, AGTR1, ET-1mRNA and the expressions of ACE, Ang-Ⅱ and AGTR1 protein were significantly higher in myocardial tissue of AMI rats than those of sham-operation rats (P＜0.05). Compared with model group, the expressions of AGT, AGTR1, ET-1mRNA and ACE, Ang-Ⅱ, AGTR1 protein were significantly decreased in paeonol high dose group and captopril control group (P＜0.05). Paeonol reduced the expressions of those mRNA and protein levels in a significant dose dependent manner. Conclusion Paeonol can slow down the deterioration of the ventricular remodeling after AMI in rats, which may be related to the inhibition of over-activation of RAS.

Objective To evaluate the effect of gender matching on the outcomes of livingdonor renal transplantation.Methods A total of 419 cases of living-donor renal transplantation in our center were divided into male-donor-male-recipient (MDMR) group,male-donor-female-recipient (MDFR) group,female-donor-male-recipient (FDMR) group,female-donor-female-recipient (FDFR)group.The outcomes including graft and patient survival,acute rejection and renal function were analyzed retrospectively.Results Compared to MDMR group,MDFR group and FDFR group had lower Scr [(80.7±17.9),(87.4±21.9) μmol/L vs (120.3±72.5) μmol/L,all P ＜ 0.05] and uric acid (UA) [(318.1 ± 86.4),(303.5 ± 66.9) μmol/L vs (358.4 ± 77.8) μmol/L,P ＜ 0.05] 6 months after operation.Compared to MDFR group,FDMR group had higher Scr[(117.7±27.4) μmol/L vs (80.7±17.9) μmol/L,P ＜ 0.01],UA [(371.0±92.4) μmol/L vs (318.1±86.4) μmol/L,P ＜ 0.05] and lower glomerular filtration rate (GFR) [(70.4± 17.8) ml/min vs (79.6± 18.9) ml/min,P ＜ 0.05].Compared to FDMR group,FDFR group had lower Scr [(87.4±21.9) μmol/L vs (117.7±27.4) μmol/L,P ＜ 0.01] and UA [(303.5±66.9)μmol/L vs (371.092.4) μmol/L,P＜ 0.01].Compared to MDFR group,FDFR group showed lower GFR [(72.4±25.3) ml/min vs (82.7± 18.7) ml/min,P ＜ 0.05] 1 year after operation.Compared to MDMR group,FDFR group showed lower UA [(322.9±69.7) μmol/L vs (376.0±66.2) μmol/L,P ＜ 0.05] 2 years after operation.Compared to FDMR group,FDFR group showed lower Scr [(88.7 ±27.0) μmol/L vs (112.7±27.8) μmol/L,P ＜ 0.05] and UA [(318.3 ±61.2) μmol/L vs (396.2± 100.3) μmol/L,P ＜ 0.05] 3 years after operation.5 years after operation,there were no significant differences in above indexes,the incidence of slow graft function,acute rejection and survival of graft and patient among groups.Conclusions Male recipients of female donors have the worst renal function while female recipients have better outcomes after operation.

Objective To investigate the age-related change of secretory leukocyte protease inhibitor (SLPI) content in stimulated whole saliva. Methods Ninety-four healthy adults aged 20-94 yrs were enrolled. Samples of whole stimulated saliva were collected from all subjects, the SLPI content was analyzed by enzyme-linked immunosorbent assay (ELISA). Results The flow rate of the SLPI in older groups was lower than in the young group (P0. 05) , but the contents of SLPI secretion per time-unit of the three older groups[(2. 12?1. 70), (2. 17?2. 65) , (1. 91?2. 51) ng/min, all P

Severe Acute Respiratory Syndrome(SARS)is a deadly infectious disease caused by SARS Coronavirus(SARS-CoV).Inactivated SARS-CoV has been explored as a vaccine against SARS-CoV.However,safe and potent adjuvants,especially with more efficient and economical needle-free vaccination are always needed more urgently in a pandemic.The development of a safe and effective mucosal adjuvant and vaccine for prevention of emergent infectious diseases such as SARS will be an important advancement.PIKA,a stabilized derivative of Poly(I:C),was previously reported to be safe and potent as adjuvant in mouse models.In the present study,we demonstrated that the intraperitoneal and intranasal co-administration of inactivated SARS-CoV vaccine together with this improved Poly(I:C)derivative induced strong anti-SARS-CoV mucosal and systemic humoral immune responses with neutralizing activity against pseudotyped virus.Although intraperitoneal immunization of inactivated SARS-CoV vaccine alone could induce a certain level of neutralizing activity in serum as well as in mucosal sites,co-administration of inactivated SARS-CoV vaccine with PIKA as adjuvant could induce a much higher neutralizing activity.When intranasal immunization was used,PIKA was obligatorily for inducing neutralizing activity in serum as well as in mucosal sites and was correlated with both mucosal IgA and mucosal IgG response.Overall,PIKA could be a good mucosal adjuvant candidate for inactivated SARS-CoV vaccine for use in possible future pandemic.

OBJECTIVE Dynamics of serum and urine metabolites in hepatic injury rats induced by ethanolic extract from Rhizoma Dioscoreae Bulbiferae(RDB)was investigated by 1H-NMR-based metabo?nomic methods in order to discover early biomarkers of liver toxicity induced by RDB. METHODS Rats were ig adminisetred with RDB at a dose of 5 g·kg-1 for 28 d. Rats were sacrificed 3,7,14 and 28 d af?ter RDB administration,as well as after a recovery period,respectively. Blood was taken for routine bio?chemical analysis by an automatic biochemical analyzer. Liver/body mass indexes were calculated ,and liver pathological changes were observed with hematoxylin-eosin staining. Urine samples were collected before and 3,7,14 and 28 d after RDB administration,respectively,as well as after withdrawal. Metabo?nomic analysis was carried out for serum and urine samples. Principal component analysis and orthogonal partial least squares-discriminant analysis were used for screening and identifiying early biomarkers. RESULTS Compared with the control group,total bilirubin (TB) and total cholesterol (TC) values were increased in 3-28 d in RDB group(P<0.05,P<0.01). Total bile acid(TBA)was elevated in 7-28 d (P<0.05,P<0.01). TB,TC and TBA became normal after discontinuation with RDB. There was no significant difference between RBD-treated group and control group in the activity of glutamic-pyruvic transaminase and glutamic-oxaloacetic transaminase,and the content of glucose also was not different between the two groups. The ratio of liver/body mass was elevated at 3-28 d(P<0.01)but returned to normal after withdraval of RDB. The enlargement and necrosis of hepatocytes were observed 7 d after RDB administration,and lesion degree was aggravated with the extension of RDB delivery time. Meta?bonomic analysis showed that the serum lipids (low density lipoprotein/very low density lipoprotein (LDL/VLDL),glutamic acid,choline phosphate and glycerolphosphatecholine were increased in the early stage. Pyruvate and N-acetylglutamate were decreased in urine. These metabolites became normal 7 d after discontinuation with RDB. CONCLUSION The serum lipids (LDL/VLDL),glutamic acid,glycerol phosphate choline,as well as urine pyruvic acid salt and N-acetyl glutamate may be used as the early biomarkers for liver toxicity induced by RDB.

Mitochondria play a central role in the regulation of energy metabolism and signal transduction in eukaryotic cells. Although many fluorescent labeling strategies have been developed for mitochondrial studies, the methods that enable labeling efficiency assessment at the single-mitochondrion level are still lacking. By employing the unique advantages of high sensitivity flow cytometry ( HSFCM ) in the sensitive, rapid, and quantitative multiparameter analysis of individual mitochondria, here we examined the performance of several different mitochondrial labeling strategies from the perspectives of brightness, labeling ratio, and stability. Mitochondria isolated from HeLa cells transfected with pAcGFP1-Mito plasmid upon transient or stable transfections, and mitochondria directly labeled with MitoTracker Green or SYTO 62 were analyzed by a laboratory-built three-channel HSFCM. Upon the quantitative measurement of fluorescence brightness, it was found that the fluorescence intensity of green fluorescent protein ( GFP ) in mitochondria isolated from cells with stable transfection was about 17. 7-fold higher than the transient transfection ones, and was approximately two orders of magnitude brighter than mitochondria labeled with MitoTracker Green. On the other hand, the fluorescence signal of SYTO 62 labeling decreased upon washing, indicating its rapid dissociation rate. The strong fluorescence intensity and good labeling stability make stable transfection an efficient method to label mitochondria. The experimental results demonstrates that HSFCM provides a powerful analytical tool to assess the performance of mitochondrial fluorescence labeling via high throughput single mitochondria analysis.