Objective To evaluate the expression and its clinical significance of serum LCN-2 and MMP-9 in human breast cancer.Methods 96 patients with breast cancer,42 patients with benign breast diseases and 27 healthy controls were recrui-ted in this study.The LCN-2 and MMP-9 levels were measured by ELISA in the serum of 96 patients with breast cancer,42 patients with benign breast disease,and 27 health controls.Results The level of serum LCN-2 in the breast cancer group (89.76±28.38 ng/ml)was significantly higher than those in benign breast diseases (56.44±21.15 ng/ml)and healthy controls (44.32±16.84 ng/ml)(P<0.05).The level of serum MMP-9 in the breast cancer group (96.34±23.26 ng/ml) was significantly higher than those in benign breast diseases (28.16±11.57 ng/ml)and healthy controls (23.54±13.61 ng/ml)(P<0.05).No obvious difference in the levels of serum LCN-2 and MMP-9 was observed between benign breast disease and healthy controls (P>0.05).Correlation analysis showed that a positive correlation was found between the levels of serum LCN-2 and MMP-9 in breast cancer patients (r=0.513 5,P<0.05).The levels of serum LCN-2 and MMP-9 were significantly associated with tumor size,lymph node metastasis,the expression of ER and PR,TNM stage,but were not asso-ciated with tumor differentiation (P>0.05).The correlation analysis showed that the LCN-2 level was positive correlated with the MMP-9 level (P<0.05).Conclusion LCN-2 and MMP-9 could play a role in the tumorigenesis and development of breast cancer.Measuring of serum LCN-2 and MMP-9 may be a useful tool for diagnosis,effect monitoring and prognosis in breast cancer.

Objective To investigate the relationship of PTEN,Ki-67,type of credits of thymoma in WHO organization and Masaoka staging,and the significance of the biology's expression in thymoma.Methods The expression of PTEN and Ki-67 in 52 pieces of thymoma specimens was detected by the method of immunohistochemical SP.Results Positive expression rate of the 52 patients with thymoma was decreased gradually after WHO classification (8/10,10/14,6/12,4/11,0,0 and 10/13,12/20,6/16,0),Masaoka staging and PTEN.The difference were significant in statistics (λ2 =11.751,P =0.038 ; x2 =8.311,P =0.040).Positive expression rate of Ki-67 was gradually increased (1/10,6/14,6/12,8/11,2/3,2/2 and 3/13,9/20,10/16,3/3),the difference was also significant in statistics (x2 =11.231,P =0.047; x2 =7.904,P =0.048).Conclusions The positive expression rate of PTEN,Ki-67 are both related to clinical classification and staging closely.The detection to the positive expression rate of PTEN and Ki-67 can contribute to judging thymoma biological characteristics,thus,it will be helpful in diagnosing the thymoma,analyzing the degree of malignancy,invading and transferring ability.And then provide objective indicators for clinical diagnosis and treatment.

10.3969/j.issn.2095-4344.2013.24.024

A strain of canine parvovirus (CPV) was isolated from feces of an ill puppy in an animal hospital in Wuhan, China. It was designated as CPV/WH02/06. This isolate was identified as serotype CPV-2a by the hemagglutination test, CPV Ag detection strip, electron microscopy, and PCR. The vp2 gene was cloned and sequenced and assigned GenBank accession number EU377537. A 1242 bp segment of the 5' region of the vp2 gene was cloned and inserted into the binary vector pBI121 and used for Agrobacterium-mediated tobacco transformation. Transgenic tobacco plants were selected on MS medium supplemented with 100 μg/mL kanamycin and 100 μg/mL timentin. Integration of the vp2 gene into the tobacco genome was confirmed by PCR using T1 progeny plants, and the expression of the VP2 protein was confirmed by Western blotting.

Objective To investigate microRNAs ( miRNAs) expression profiling of cardiomyocytes in rats with heart failure, and predict miRNAs-regulated target genes and their functions.Methods Total of 18 male SD rats weighing 200-220 g were randomly divided into 2 groups:the control group ( CON) and the heart failure group (HF).The rats in HF group were injected by adriamycin via tail vein to induce heart failure, meanwhile in CON group, rats were received an equal volume of 0.9% sodium chloride intravenously.The cardiomyocytes isolated from the rat hearts in two groups and cultured overnight.After that, total RNA was extracted and then subjected to miRNA microarray to screen differentially expressed miRNAs.The reults of microarray were further verified by quantitative real-time PCR ( qRT-PCR ) .The target genes regulated by differentially expressed miRNAs were predicted by the software of Targetscan and miRanda.Bioinformatics analysis was performed to predict the miRNAs-regulated target genes and analyze the enriched gene ontology ( GO) and signaling pathway ( KEGG Pathway) .Results The results of miRNA microarray showed that a total of 37 miRNAs were differentially expressed in HF group as compared to CON group, among which 22 miRNAs were up-regulated and 15 miRNAs were down-regulated (P<0.01, FDR<0.05).The expression of miR-133b-5p (t=14.56, P<0.01), miR-6216 (t=9.32, P<0.01) and let-7e-5p (t=13.92, P<0.01) which were detected by qRT-PCR exhibited the similar tendency of up or down regulation to those shown in microarray results.Bioinformatics analysis indicated that miRNAs-regulated target genes were significantly enriched in 31 GOs (P<0.01, FDR<0.05) and 12 signal pathways (P<0.05, FDR<0.05), among which ubiquitin-proteasome system, MAPK signaling pathway and Toll like siganling pathway exhibited a higher enrichment. Conclusion MiRNA expression profile on cardiomyocytes in rat with adriamycin-induced heart failure was significantly changed.These differentially expressed miRNAs might participate in the process of heart failing by regulating their target genes in rat cardiomyocytes.

Objective To investigate relativity between the epidemiology of HPV and cervical carcinoma in Dali region,Yunnan province,through detecting the 13 high-risk human papillomavirus infection and Thinprep cytologic test in 2153 cases.Methods Real-time PCR was used to detect the 13 high-risk HPV (16,18,31,33,35,39,45,51,52,56,58,59,68) in2153 cases and 1604 cases were checked with Thinprep cytologic test.Results In 2153 samples,260 cases were infected with HPV,with the positive rate of 12.08％.The highest positive rates were ＞60 years old (18.18％),then ＞20 and ≤30 years old (14.41％);there was no significant difference in the positive rate among the various age groups (P =0.384).There were 1465 negative for intraepithelial lesion ormalignancy (NILM) cases (91.33％),86 atypical squamous cells of undetermined significance (ASC-US) and atypical squamous cells cannot exclude HSIL (ASC-H) cases (5.36％),32 low-grade squamous intraepithelial lesion cases (LSIL) cases (2.00％),21 high-grade squamous intraepithelial lesion cases (1.31％) through Thinprep cytologic test.The correlation coefficient is 0.893.Conclusions The infection rate of HPV in Dali region,Yunnan Province,has no significant difference among the various age groups.Application of 13 high-risk HPV infection test combined with Thinprep cytologic test could be more effective in screening cervical carcinoma.

Objective To investigate the role of p38 mitogen-activated protein kinase (p38MAPK) signaling pathway in reduction of myocardial ischemia-reperfusion (I/R) injury by morphine preconditioning in the rats with chronic heart failure in vitro.Methods Adult male Sprague-Dawley rats,weighing 200-230 g,aged 6-7 weeks,in which doxorubicin 2 mg/kg was injected via the tail vein once a week for 6 consecutive weeks to induce chronic heart failure,were studied.At the end of 8th week,30 rats with chronic heart failure were randomly divided into 5 groups (n =6 each) using a random number table:sham operation group (group Sham),I/R group,morphine preconditioning group (group MPC),SB203580 (p38MAPK inhibitor) + morphine preconditioning group (group SBM),and SB203580 group (group SB).The hearts were quickly excised and passively perfused in a Langendorff apparatus and subjected to 30 min of occlusion of the anterior descending branch of the left coronary artery followed by 120 min of reperfusion to establish the model of myocardial I/R injury.After equilibration,the hearts were subjected to 3 cycles of 5 min perfusion with K-H solution containing morphine 1 μmol/L at 5-min intervals before ischemia in group MPC.In group SBM,the hearts were perfused with K-H solution containing SB203580 (5 μmol/L) for 45 min starting from l0 min before morphine preconditioning until 5 min of ischemia.In group SB,morphine preconditioning was not performed,and the hearts were only perfused with K-H solution containing SB203580 (5 μmol/L) starting from 40 min before ischemia until 5 min of ischemia.At 15 min of equilibration (baseline),5 and 10 min of reperfusion,the coronary effluent was collected to detect the activity of lactate dehydrogenase (LDH) using the chemical colorimetry.At 10 min of reperfusion,the expression of phosphor-p38MAPK (p-p38MAPK) in the myocardium was determined by Western blot in Sham,I/R and MPC groups.At 120 min of reperfusion,the area at risk (AAR),total areas of right and left ventricles (LV+RV),and infarct size (IS) were measured,and the IS/AAR ratio was calculated.Results Compared with group Sham,the LDH activity in coronary effluent during reperfusion and IS/AAR ratio were significantly increased in the other groups,and the expression of p-p38MAPK was significantly up-regulated in I/R and MPC groups (P＜0.05).Compared with group I/R,the LDH activity in coronary effluent during reperfusion was significantly decreased,the expression of p-p38MAPK was significantly up-regulated,and the IS and IS/AAR ratio were significantly decreased in group MPC (P＜0.05),and no significant change was found in the LDH activity in coronary effluent,IS and IS/AAR ratio in SBM and SB groups (P＞0.05).Compared with group MPC,the LDH activity in coronary effluent during reperfusion was significantly increased,and the IS and IS/AAR ratio were significantly increased in group SBM (P＜0.05).Conclusion The mechanism by which morphine preconditioning reduces myocardial I/R injury is related to activation of p38MAPK signaling pathway in the rats with chronic heart failure in vitro.

Objective To summarize the experience of surgical management of postoperative intrahepatic duct stones and strictures with choledochoscope.Methods The clinical data of 1241 patients with intrahepatic duct stones and strictures who were admitted to the Shenzhen People's Hospital from January 2000 to December 2011 were retrospectively analyzed.All patients were divided into postoperative choledochoscopy group (652 patients were treated by choledochoscope through T-tube fistula) and conservative treatment group (589 patients were treated by cholangic and antibiotic drugs) according to treatment plan.The clinical data including stricture alleviation,incidence of complications and treatment efficacies of the 2 groups were compared using the chi-square test.Results In the postoperative choledochoscopy group,the intrahepatic duct stones of 630 patients were completely removed,with the removal rate of 96.63％ (630/652),which was significandy higher than 76.23％ (449/589) of the conservative treatment group (x2=113.407,P ＜ 0.05).A total of 1237 intrahepatic duct strictures were detected in the postoperative choledochoscopy group,including 698 cases of membranous stricture (the length of strictured bile duct ＜ 2 mm),529 cases of spool-shaped stricture (2 mm ≤ the length of strictured bile duct ≤5 mm),8 cases of long tube-shaped stricture (the length of strictured bile duct ＞ 5 mm),2 cases of crackshaped stricture.The alleviation rate of the stricture was 99.35％ (1229/1237).The conditions of 6 cases of long tube-shaped stricture and 2 cases of crack-shaped stricture were not alleviated.There were 986 cases of intrahepatic duct strictures in the conservative treatment group,and the stricture alleviation rate was 80.43％ (793/986).There was a significant difference in the stricture alleviation rate between the 2 groups (x2=238.994,P ＜ 0.05).Nine patients in the postoperative choledochoscopy group had severe complications.Five patients had intrahepatic duct hemorrhage with a volume of blood loss more than 1500 ml,and they were treated by balloon dilatation hemostasis.One patient with repeated intrahepatic duct bleeding received hepatectomy.Two patients with severe intrahepatic cholangitis received choledochoscopy.Two patients had incomplete formation of sinus and they received implantation of biliary stents under duodenoscope.Conclusion Standard surgical management with choledochoscope can effectively alleviate postoperative intrahepatic duct stones and strictures.

Objective To investigate the therapeutic effects and mechanism of biliopancreatic diversion (BPD) surgery on type 2 diabetes mellitus(T2DM) in GK rats.Methods 16 GK rats were randomly divided into 2 groups:BPD surgery group included 10 rats undergoing BPD surgery,sham-BPD group included 6 rats undergoing a sham operation.Fasting plasma glucose,insulin,glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide(GIP)were detected one week before BPD surgery and the 1st week,4th week,10th week,26th week after BPD surgery.Oral glucose tolerance test(OGTT) and insulin tolerance test(ITT) were done in the 10th week after BPD surgery.Results There was no statistical difference in fasting plasma glucose,insulin,plasma GLP-1 or GIP between the 2 groups before surgery.Plasma glucose had significant reduction in BPD group compared to that in the sham group(P ＜0.05) and insulin level had no significant difference between the 2 groups.Rats in BPD group had significant improvement in glucose tolerance and insulin sensitivity compared to those in the sham group.Serum level of GLP-1 was significantly elevated in BPD group compared to that before surgery (P =0.0337 at the 1st week after surgery; P =0.0002 at the 4th week after surgery,P ＜ 0.0001 at the 10th week after surgery,P ＜0.0001 at the 26th week after surgery) and that in sham-BPD group(P =0.0354 at the 1st week after surgery,P =0.0032 at the 4th week after surgery,P =0.0001 at the 10th week after surgery,P ＜0.0001 at the 26th week).Serum level of GIP was significantly lowered in BPD group compared to that before surgery(P =0.0189 at the 1st week after surgery; P =0.0007 at the 4th week after surgery,P =0.0003 at the 10th week after surgery,P ＜0.0001 at the 26th week after surgery) and that in sham-BPD group(P =0.0089 at the 1st week after surgery,P =0.0002 at the 4th week after surgery,P =0.0006 at the 10th week after surgery,P ＜0.0001 at the 26th week after surgery).The difference had statistical significance (P ＜0.05).Conclusion BPD surgery can significantly reduce fasting plasma glucose,improve glucose tolerance and insulin sensitivity.The change of serum levels of GLP-1 and GIP may play the major role in BPD treatment of diabetes mellitus.

Objective To detect the expression of BP1 gene in thyroid cancer and its relationship with clinicopathological features of thyroid cancer.Methods BP1 gene expression in 60 cases of thyroid cancer tissues and 20 cases of normal thyroid tissues were detected by in situ hybridization and immunohistochemistry.Results The positive expression rate of BP1 mRNA was 78.3 ％ (47/60) in the 60 cases of thyroid cancer tissues while it was 20％ (4/20) in the 20 cases of normal thyroid tissues detected by in situ hybridization.The difference had statistical significance (P ＜ 0.05).Of the 3 pathological types of thyroid cancer,the positive expression rate of papillary carcinoma was 81.6％ (40/49),follicular carcinoma 85.7％ (6/7),and medullary carcinoma 25.0％ (1/4).The expression of BP1 mRNA had statistical difference between medullary carcinoma and other pathological types like papillary carcinoma and follicular carcinoma (P ＜ 0.05).The positive expression rate of BP1 protein was 93.3％ (56/60)in the 60 cases of thyroid cancer tissues while it was 10.0％ (2/20) in the 20 cases of normal thyroid tissues detected by immunohistochemistry.The difference had statistical significance(P ＜0.05).Conclusion BP1 gene expression is up-regulated in human thyroid cancer and it is related to tumor stage and pathological type but not related to patients' age,sex or lymph node metastasis.