Objective To investigate differential expression of Raf kinase inhibitor protein (RKIP)in gastric cancer and its clinical significance.Methods Pure gastric adenocarcinoma cells (GAC)and paracancerous gastric epithelial cells(NGEC)in 12 patient with gastric cancer were obtained using laser capture microdissection(LCM).After trypsin digestion all specimens were labeled with18O/16O.Quantitatively identification of differentially expressed proteins between GCC and NGEC was conducted using Nano-HPLC-MS/MS.The expression of RKIP in GAC was determined by Western blotting.Immunohistochemistry(IHC)was used to detect the expression of RKIP in gastric cancer(n=118),non-cancerous mucosa tissues(n=70)and matched positive lymph node tissues(n=35).Results A total of 78 differentially expressed proteins were identified and RKIP was downexpressed in GCC compared with NGEC(1:4.37).IHC examination revealed that positive expression of RKIP in gastric cancer had negative relation with deeper invasion,TNM stage and lymphoid node metastasis,while it had positive relation with histological differentiation(P＜0.05).Conclusion Down-regulation of RKIP in gastric cancer tissues might affect the biological behavior of gastric cancer.

OBJECTIVE:To observe clinical efficacy and safety of tegafur gimeracil oteracil potassium combined with 3D-CRT and microwave hyperthermia in the treatment of elderly patients with local advance esophageal cancer. METHODS:48 cases of local advance esophageal cancer were divided into group A,B and C according to random number table method,with 16 cases in each group according to random number table method. Group A was given 3D-CRT with total dose of 60-66 Gy totnlly for 6-7 weeks;group B was additionally given tegafur gimeracil oteracil potassium 60 mg/(m2·d),bid,on the basis of group A 3 weeks for a course of treatment,until the end of radvotherapy;group C was additionally given thermotherapy on the basis of group B 3 weeks for a course of treatment,until the end of radvotherapy. Clinical efficacy,1-year survival rate,dysphagia,weight and KPS score were compared among 3 groups,and the occurrence of toxic reactions(radioactive esophagitis,radioactive pneumonia, bone marrow suppression and gastrointestinal reaction) were observed in 3 groups. RESULTS:Total effective rate,1-year survival rate and the remission rate of dysphagia of group C were 93.75%,87.50% and 93.75%,which were significantly higher than those of group A(50.00%,50.00% and 56.25%)and B(68.75%,68.75% and 68.75%),with statistical significance(P<0.05). There was no statistical significance in the rate of weight gain and KPS score gain among 3 groups(P>0.05). The incidence of radioactive esophagitis and radioactive pneumonia in group C were significantly lower than in group A and B,with statistical significance(P<0.05);there was no statistical significance in the incidence of gastrointestinal reaction and bone marrow suppression among 3 groups (P>0.05). CONCLUSIONS:Tegafur gimeracil oteracil potassium combined with 3D-CRT and microwave hyperthermia in the treat-ment of elderly patients with local advance esophageal cancer further improves clinical efficacy and survival rate,but shows low inci-dence of toxic reaction.

Objective To construct an eukaryotic expression vector pEGFP-N1-CKB and establish a stably transfected NCI-H520 cell line.Methods Human CKB gene was amplified by PCR with human CKB cDNA library as the template and the fragment was combined with plasmid pEGFP-N1.The recombinant expression vector,pEGFP-N1-CKB,was transfected to NCI-H520 using Lipofectamin.The stably transfected cell line was established after G418 selection and the expression level of CKB gene before and after transfection was detected by Western blot.Results After identification by restriction enzyme digestion and sequencing,the eukaryotic expression vector,pEGFP-N1-CKB,was successfully constructed.The expression level of CKB in NCI-H520 transfected by pEGFP-N1-CKB was significantly higher than that in control.CKB gene had a stable transfection in NCI-H520 cells.Conclusions An eukaryotic plasmid encoding CKB (pEGFP-N1-CKB) has been constructed and a cell line expressed CKB stably has been successfully prepared.

Objective To investigate the surgical effort of transurethral bipolar plasma kinetic resection (PKR-BT) on who cannot tolerate radical cystectomy in elderly patients with high-risk infiltration bladder urothelial cancer.Methods From 2010 January to 2013 September,data from 27 cases of elderly had been reviewed.The risk of invasive bladder urothelial cancer were treated with PKR-BT treatment.All patients met WHO elderly standards in this group of patients with severe diabetes was 7 cases,and other comlications.Results All 27 patients had been successful in operation and postoperative recovery.Patients were followed up for 4-45 months,8 cases occured recurrence,6 cases of PKR-BT again,2 cases missed,complete follow up 25 patients still survived,the median follow-up time was 23.6 months,the longest survival was 45 months.Conclusion PKR-BT in the treatment of invasive bladder urothelial cancer patients is safe and effective,it can be used as one of the alternate treatment for such patients.

Objective To explore the related factors and nursing interventional methods of depression in patients with motor aphasia after stroke. Methods The Stroke Aphasic Depression Questionnaire-Hospital version (SADQ-H) was used to diagnose depression of 271 patients with motor aphasia after stroke. The association of post-stroke depression with social factors, vascular risk factors and stroke features were analyzed. The relationship between depression severity and aphasia severity were analyzed. A total of 166 patients with depression were divided into the control group and the observation group to carry out different nursing interventional methods. Results The incidence of post-stroke depression in patients with motor aphasia was 61.3%(166/271). The occurrence of post-stroke depression was correlated with female (χ2=5.580, P=0.02) , age less than 60 years old (χ2=4.390, P=0.04) , living alone (χ2=5.400, P=0.02) , recent negative events (χ2=4.420, P=0.04). The multi-factor Logistic regression analysis showed that the occurrence of post-stroke depression was independently associated with female (OR=2.58, 95%CI 2.12-3.86, P<0.01), age less than 60 years old(OR=2.62, 95%CI 2.16-3.94, P<0.01), living alone(OR=2.59, 95%CI 2.13-3.92, P<0.01). After 4 weeks of nursing intervention, there were significant differences in SADQ-H scores (t=2.94, P=0.002) and distribution of Boston Diagnostic Aphasia Examination (BDAE) (χ2=14.045,P=0.000) between the control group and the observation group. Conclusions Female, ages less than 60 years old, living alone are independent risk factors of post-stroke depression in motor aphasia patients. The degree of depression is associated with aphasia severity. The early nursing intervention can improve the depression status and promote recovery of the language function of patients with motor aphasia after stroke.

The mechanism of how stroma plays an important role in tumor carcinogenesis is now a hotspot. To delineate the features of stromal protein between nasopharyngeal carcinoma (NPC) and normal nasopharyngeal mucosa(NNM), laser capture microdissection (LCM) was performed to purify stromal cells from the NPC and NNM, respectively. The protein expressed profiles of the stroma of NPC and NNM were compared using fluorescent two-dimensional difference gel electrophoresis (2D-DIGE) and 34 differential protein spots between tumor stroma and normal stroma were chosen to be identified by mass spectrometry (MS). A total of 20 differential proteins were identified, and three differential proteins (CapG, L-plastin and S100A9) were selectively further validated by Western blotting and immunohistochemical analysis to confirm the results of 2D-DIGE. 2D-DIGE patterns of the stroma of NPC and NNM were established for the first time, the results suggested that differentially expressed proteins in the stroma of NPC and NNM may be useful for understanding the relationship between NPC cells and their surrounding microenvironment. Further studying of these proteins will be helpful to elucidate the mechanisms of NPC carcinogenesis and provide new thoughts on therapy of NPC through stroma.

Objective To search for lymph node metastasis-associated proteins in human lung squamous carcinoma (hLSC).Methods Laser capture microdissection (LCM) was used to purify the target cells from lung primary tumor and matched lymph node metastatic tumor in hLSC. Two-dimensional gel electrophoresis (2-DE) was performed to separate the total proteins of microdissected tumor cells from lung primary tumor and matched lymph node metastatic tumor. PDQuest software was applied to analyze 2-DE images. Differential protein spots between the two types of tissues were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS). The expression of Rho-GDIα, one of the differential proteins, in the microdissected lung primary tumor cells (LPTC) and matched lymph node metastatic tumor cells (LNMTC) was detected by Western blot. Results In the present study, 2-DE patterns of microdissected LPTC and LNMTC were established, and 22 differential proteins in the above two tissues were identified, of which 14 were down-regulated in LNMTC and 8 were up-regulated in LNMTC.Conclusion The 22 differential proteins may play some roles in the process of lymph node metastasis in hLSC, and the data provide new clues for metastasis-associated biomarker screen and mechanism of hLSC.

In order to screen EGFR-regulated secreted proteins in human nasopharyngeal carcinoma(NPC), and to reveal the role and mechanism of epidermal growth factor receptor(EGFR) in the pathogenesis of NPC. NPC cell line CNE2 cells were cultured in serum-free medium and stimulated by transforming growth factor-? (TGF-?) for 24 h in experimental group. Control CNE2 cells were cultured at the same condition but without TGF-? stimulation. The culture medium of control and experimental cells was desalted and concentrated through ultrafiltration to prepared the total secreted proteins. Two-dimensional gel electrophoresis (2-DE) was used to separate the secreted proteins of control and experimental cells, PDQuest software was applied to analyze 2-DE images, and the differential protein spots between the control and experimental cells were identified by desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The 2-DE patterns of the secreted proteins of TGF-? stimulated and un-stimulated CNE2 cells were established, 22 differential proteins spots between the two groups of cells were found, and 8 non-redundant proteins were identified with MALDI-TOF-MS, the functions of which were involved in invasion, metastasis, apoptosis and proliferation of cancer cells. The data will be valuable for further to study the role and mechanism of EGFR in the pathogenesis of NPC.

Objective To screen for new methylation association genes in HL-60 to reveal the pathogenesis of leukemia, and provide important theoretical and scientific basis for the prevention and cure of leukemia. Methods Two-dimensional fluorescence difference gel electrophoresis (F-2D-DIGE) was performed to separate the total proteins from acute myelogenous leukemia (AML) cell line HL-60 cells with or without 5-aza-2-deoxycytidine (5-aza-2-dC) treatment. Imaging software Decyder 6.5 and PDQuest were used to detect the differential expression protein spots, and matrix-assisted laser desorption/ionizaion time-of-flight mas spectrometer (MALDI-TOF MS) was adopted to identify the differential expression proteins. Results F-2D-DIGE maps of 5-aza-2-dC-untreated HL-60 and-treated HL-60 cells were established. A total of 53 differential protein spots were detected, and 35 differential proteins were successfully identified. Of the identified proteins, 32 proteins were up-regulated, and 3 proteins were down-regulated in HL-60 cells after 5-aza-2-dC treatment.Conclusion Thirty-five differential proteins may be associated with methylation in HL-60 cell line, which provides the important clues for epigenetic study of leukemia.

Objective To identify proteins eliciting a humoral immune response in patients with nasopharyngeal carcinoma (NPC) by a serological proteome analysis, and provide candidate biomarkers for diagnosis and treatment of NPC.Methods Two-dimensional (2-DE) electropboresis was used to separate total cellular proteins from 19 NPC tissues.Separated proteins were transferred onto PVDF membranes and sera from 19 NPC patients and 19 healthy subjects were individually screened by western blotting for antibodies that react against separated proteins.Each tissue samples was subjected to three 2-DE gels and coomassie staining was performed in one of them.The protein spots which selectively reacted with the patient sera were excised from the preparative gels and subjected to further analysis of MOLDI-TOF MS and ESI-QTOF MS/MS.The proteins were identified based on peptide mass fingerprints and peptide sequence tags followed by searching database.Results In this study, 13 NPC associated antigens (HSP 70, HAS,HSP 60, CK 15, LAP 3, α-enolase, EBP 1, CK 19, ribosomal protein P 0, pyrovate dehydrogenase E1,guanine nucleotide-binding protein, prohibitin, Rho-GDI 2) that elicited an antibody response in most of NTC patients were identified.The positivities of these proteins were more than 21% all in NPC patients, but were lower or even absent in normal subjects.Conclusion These 13 NPC associated antigens and their autoantibodies may be useful for NPC diagnosis and treatment.