Objective To detect the expression of cyclooxygenase(COX) in human gastric cancer cell lines and its subcellular location of the isoforms. Methods Immunohistochemistry、RTPCR combined with laser scanning cofocal microscopy (LSCM) were used to investigate expression and distribution of COX. Results Positive staining of COX2 and COX1 protein was seen in human gastric cancer cell line MKN45、SGC7901 and AGS. However, the COX2 staining was absent and COX1 staining was weak in MGC803; though their mRNA in all the four cell lines. When compared to COX1,COX2 showed a stronger signal at both protein and mRNA levels of the gastric cancer cell lines, which was confirmed by double labeling and LSCM. The quantitative analysis of fluorescein intensity indicated the pixel intensity peak of COX2 reached 50～70, while COX1 only 10. LSCM also showed that COX2 was both cytoplasmic and nuclear envelope staining but COX1 only in cytoplasm. Conclusions In human gastric cancer, there is stronger expression of COX2 than COX1, and different distribution of the two isoforms implies their distinct roles in cell function.

Objective To investigate the abnormal expression of microRNA (miRNA)-25 in the serum of gastric cancer patients and its clinical significance. Methods In Xijing Hospital of Digestive Diseases,Fourth Military Medical University,86 gastric cancer patients with operation and completed follow-up data,70 gastric adenoma patients and 80 healthy controls were selected as study objects.Total RNA was isolated from the serum. After the stable and sensitive miRNA-25 absolute quantity detection method established,the serum levels of miRNA-25 in gastric carcinoma patients,gastric adenoma patients and healthy controls were tested according to this method. The expression differences of miRNA-25 in the serum of patients with gastric cancer and gastric adenoma and healthy controls were analyzed with statistic analysis,and the correlation between miRNA-25 expression level and clinic pathological features of gastric cancer was also analyzed. Results The expression level of miRNA-25 in the serum of gastric cancer patients (135. 6 fmol/μg total RNA) was significantly higher than that of gastric adenoma patients (67. 7 fmol/μg total RNA) and healthy controls (62. 2 fmol/μg total RNA)(P＜0. 01). The receiver operating characterisstic curve of miRNA-25 indicated that serum miRNA-25 with good specificity and sensitivity in gastric cancer diagnosis (AUC=0. 827). The serum level of miRNA-25 in gastric cancer patients with lymph node metastasis [(148. 3±10. 2) fmol/μg total RNA] or clinicopathological stage Ⅲ /Ⅳ patients [(146. 7±9.5) fmol/μg total RNA] was significantly higher than that of gastric cancer patients without lymph node metastasis [(120. 3±10. 1)fmol/μg total RNA] or clinicopathological stage Ⅰ/Ⅱpatients [(119. 4±12. 2) fmol/μg total RNA] (P＜0.05). The correlation statistical analysis result indicated that there was no significant difference in survival period between serum miRNA-25 highly expressed and lowly expressed gastric cancer patients (P＞0. 05).Conclusion Serum miRNA-25 testing maybe helpful in diagnosis and prognosis of gastric cancer.

Objective To establish a stable and reliable mouse model as an alternative to the traditional model of impaired glucose tolerance induced by calorie restriction and its effect on glucose homeostasis.Methods Forty 16-week-old SPF C57BL/6J mice (half male and half female) were randomly divided into four groups by sex and the way of feeding.The mice in the ad libitum (AL) group had free access to basic diet, while the mice in the intermittent fasting (IF) group had normal diet and fasting on alternate days, with free access to water on the fasting days.The changes of body weight and blood glucose concentration in each group were monitored, and intraperitoneal glucose tolerance test and insulin tolerance test in mice were performed before and after the 12-week IF treatment.Results At 12 weeks after IF treatment, the body weight and blood glucose concentration of mice did not show significant difference.After i.p.injection of glucose, the blood glucose concentration of IF mice was less increased than the AL group, and after the insulin injection, the blood glucose concentration was more decreased.Compared to the AL group, the areas under the curve of tolerance test in the IF group were significantly decreased (P < 0.05).Conclusions After IF treatment, the mice show an enhanced sensitivity to insulin and improved glucose tolerance.This establishment method of mouse model of intermittent fasting is easy and simple, therefore, can be used as an effective alternative to traditional calorie restriction model of impaired glucose tolerance.

OBJECTIVETo generate phage-displayed anti-idiotypic antibody single chain variable fragments (anti-Id ScFv) to MG7 monoclonal antibody (McAb) directed against gastric carcinoma so as to lay a foundation for developing anti-Id ScFv vaccine of the cancer.

METHODSBalb/c mice were immunized i.p. with MG7 McAb conjugated with keyhole limpet hemocyanin (KLH), and mRNA was isolated from the spleens of the immunized mice. Heavy and light chain (VH and VL) genes of antibody were amplified separately and assembled into ScFv genes with a linker DNA by PCR. The ScFv genes were ligated into the phagemid vector pCANTAB5E and the ligated sample was transformed into competent E. coli TG1. The transformants were infected with M13K07 helper phage to yield recombinant phages displaying ScFv on the tips of M13 phage. After 4 rounds of panning with MG7, the MG7-positive clones were selected by ELISA from the enriched phages. The types of the anti-Id ScFv displayed on the selected phage clones were preliminarily identified by competition ELISA.

RESULTSThe VH, VL and ScFv DNAs were about 340 bp, 320 bp and 750 bp respectively. Twenty-four MG7-positive clones were selected from 60 enriched phage clones, among which 5 displayed beta or gamma type anti-Id ScFv.

CONCLUSIONThe anti-Id ScFv to MG7 McAb can be successfully selected by recombinant phage antibody technique, which paves a way for the study of prevention and cure of gastric carcinoma by using anti-Id ScFv.

Animals ; Antibodies, Anti-Idiotypic ; biosynthesis ; genetics ; Antibodies, Monoclonal ; genetics ; immunology ; Bacteriophages ; genetics ; Cloning, Molecular ; Immunoglobulin Fragments ; biosynthesis ; genetics ; Immunoglobulin Heavy Chains ; biosynthesis ; genetics ; Immunoglobulin Light Chains ; biosynthesis ; genetics ; Immunoglobulin Variable Region ; biosynthesis ; genetics ; Mice ; Mice, Inbred BALB C ; RNA, Messenger ; genetics ; Stomach Neoplasms ; immunology ; Vaccines, DNA ; genetics ; immunology

OBJECTIVE:To study the effect of peiminine on increasing the chemosensitivity of 5 kinds of cancer cells. METH-ODS:Using human esophageal cancer Eca-109 cell,human breast cancer MCF-7 cell,human small cell lung cancer A549 cell,hu-man hepatoma HepG2 cell and human cervical cancer HeLa cell as objects,MTT colorimetric method was used to detect the growth inhibition rate of above-mentioned 5 kinds of cancer cells after treated by peiminine with maximal non-toxic mass concentra-tion(20 μg/mL)and adriamycin with different gradient mass concentrations(0.026-2.1,0.026-2.1,0.125-2.0,0.125-2.0,0.0625-0.10μg/mL)for 72 h. The half inhibitory concentration(IC50)was calculated. Crystal violet staining method was adopted to observe the proliferation of above-mentioned cancer cells after treated by peiminine with maximal non-toxic mass concentration and adriamycin with low mass concentrations(0.02,0.005,0.04,0.02,0.01 μg/mL)for 7 d. Solvent control,single use of peiminine and adriam-ycin control were conducted. RESULTS:Compared with single use of adriamycin,the combination use of peiminine and adriamy-cin can improve the growth inhibition rate of 5 kinds of cancer cells to certain degree,most of the differences were statistically sig-nificant (P<0.05 or P<0.01);and IC50 was obviously decreased,with statistical significances (P<0.05 or P<0.01). Compared with solvent control,single use of peiminine or adriamycin had no obvious effects on the proliferation of above-mentioned cancer cells in 7 d,and the combination use of peiminine and adriamycin can obviously inhibit the proliferation of above-mentioned cancer cells in 7 d. CONCLUSIONS:Peiminine can enhance the sensitivity of above-mentioned-mentioned 5 kinds of cancer cells to cer-tain degree,showing certain chemosensitivity increasing effect.

OBJECTIVETo assess the efficacy of sacral neuromodulation (SNM) in patients with intractable constipation.

METHODSA total of 7 patients with intractable constipation were treated with pereutaneous test stimulation of the S3 nerve root and were assessed by sacral never stimulation system in our department from January 2013 to January 2014. Four of these 7 patients received operation for constipation before. The efficacy was assessed by bowel habit diary, clinic constipation scores, subjective questionnaire and clinical signs.

RESULTSThe constipation symptoms were improved significantly in all the 7 patients. The frequency and volume of defecation per week were increased obviously, and the average urine was increased. Six patients underwent permanent implantation of the SNS system. After a median 4 months follow-up, the defecation frequency increased from 0.6 ± 0.5 to 8.0 ± 2.5 per week (P<0.01), and the defecation time decreased from (22.9 ± 11.5) to (3.7 ± 0.8) min (P<0.01). The Cleveland clinic constipation score decreased from 24.6 ± 4.2 to 9.0 ± 0.9 (P<0.01), and the visual analogue scale(VAS) score increased from 8.1 ± 0.9 to 82.5 ± 5.2 (P<0.01).

CONCLUSIONSNM is a clinically efficacious, minimally invasive and safe new technique, which offers an alternative treatment for the patients with intractable constipation resistant to conservative treatment, especially for the patients refractory to traditional operations.

Constipation ; therapy ; Defecation ; Electric Stimulation Therapy ; Humans ; Sacrum ; Treatment Outcome

Objective To investigate the clinicopathological and prognostic significance of lysosome-associated membrane protein-2a (LAMP2a) expression in the development of colorectal cancer.Methods Immunohistochemistry was performed to detect the LAMP2a expression in colorectal cancer,normal mucosa,and adenoma tissues.Results LAMP2a expression in colorectal cancer and adenoma was higher than that in normal mucosa (P ＜ 0.05).Further,LAMP2a expression in colorectal cancer was negatively correlated with TNM staging and lymph node metastasis (P ＜ 0.05);however,the invasion and liver metastasis,depth of invasion and differentiation had no correlation with the lymph node (P ＞ 0.05).Kaplan-Meier analysis showed that LAMP2a expression in colorectal cancer is not significantly correlated with good prognosis (P ＞ 0.05).Univariate survival analysis showed that lymphatic invasion,vascular invasion,lymph node metastasis,TNM stage,liver metastasis,distant metastasis,and differentiation were correlated with poor prognosis of colorectal cancer (P ＜ 0.05).The Cox proportional hazards regression model showed that the depth of invasion and distant metastasis were important factors affecting the survival period of patients with colorectal cancer (P ＜ 0.05).Conclusion LAMP2a may play a role in the carcinogenesis and progression of colorectal cancer,and may be used as a molecular marker for the biological behavior of colorectal cancer.

Ubiquitin specific protease 33 (USP33) is a multifunctional protein regulating diverse cellular processes. The expression and role of USP33 in lung cancer remain unexplored. In this study, we show that USP33 is down-regulated in multiple cohorts of lung cancer patients and that low expression of USP33 is associated with poor prognosis. USP33 mediates Slit-Robo signaling in lung cancer cell migration. Downregulation of USP33 reduces the protein stability of Robo1 in lung cancer cells, providing a previously unknown mechanism for USP33 function in mediating Slit activity in lung cancer cells. Taken together, USP33 is a new player in lung cancer that regulates Slit-Robo signaling. Our data suggest that USP33 may be a candidate tumor suppressor for lung cancer with potential as a prognostic marker.
Blotting, Western ; Cell Line, Tumor ; Cell Movement ; genetics ; physiology ; Cohort Studies ; Down-Regulation ; Female ; Gene Expression Regulation, Neoplastic ; HEK293 Cells ; Humans ; Immunohistochemistry ; Intercellular Signaling Peptides and Proteins ; metabolism ; Kaplan-Meier Estimate ; Lung Neoplasms ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; Nerve Tissue Proteins ; metabolism ; Prognosis ; RNA Interference ; Receptors, Immunologic ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; genetics ; physiology ; Ubiquitin Thiolesterase ; genetics ; metabolism

In recent years,cancer immunotherapy has attracted considerable attention in the field of biotherapy for the development of chimeric antigen receptor T (CAR-T) cell.CAR-T cells are capable of recognizing tumor cell surface antigens leading to kill tumor cells.Anti-CD19 CAR-T has achieved remarkable success in the treatment of hematopoietic malignancies.Whether it can benefit solid tumor patients to the same extent still faces great challenges,and it has been a focus of attention in immunotherapy for tumors.Compared with hematological malignancies,the microenvironment of solid tumor is more complex,and the screening of tumor specific antigen is more difficult.The infiltration of CAR-T cells from the blood system to the tumor site needs to overcome the tumor stromal barrier.Although part of CAR-T cells can infiltrate into tumor site,its function may be quickly inhibited by multiple factors in microenvironment.In this paper,authors discussed the challenges that CAR-T is facing for solid tumor treatment,including the specificity of the tumor antigen,the heterogeneity of the solid tumor antigen and the inhibitory effect of the tumor microenvironment,and proposed potential strategies to possibly overcome these hurdles.Authors believe that with the development of biotechnology,it will provide more abundant technical means to optimize the structure and function of CAR-T,and CAR-T will make breakthrough progress in the treatment of solid tumor.

Objective:To compare the knowledge of endoscopic screening of gastrointestinal cancers between digestive healthcare workers and new chatbots (chatGPT and new Bing).Methods:A test with twenty-three questions of endoscopic screening of gastrointestinal cancers was conducted, focusing on the appropriate age of screening, high-risk factors, the follow-up time, and the advantages and risks of digestive endoscopy. Digestive healthcare workers were invited to complete the test through electronic questionnaires. New Bing and chatGPT were used to answer each question for 10 rounds. The primary endpoint was the correct rate of all answers. The answer accuracy between digestive healthcare workers and new chatbots were compared using variance analysis, and the factors that affected the accuracy of the answers in digestive healthcare workers were explored using univariate and multivariable liner regression analysis.Results:The results of the test completed by 76 digestive healthcare workers (21 residents, 28 digestive nurses, and 27 digestive doctors) were analyzed. The accuracies were 36.4%±10.9%, 34.5%±10.2%, 52.2%± 12.6%, 46.3%±9.8% and 67.1%±9.3% in residents, digestive nurses, digestive doctors, chatGPT, and new Bing, respectively, with significant difference ( F=22.6, P<0.001). The accuracy was highest in new Bing ( P<0.001). The accuracy was comparable between chatGPT and digestive doctors (LSD- t=-1.398, P=0.166), and both higher than that of digestive nurses (LSD- t=2.956, P=0.004; LSD- t=5.955, P<0.001) and residents (LSD- t=2.402, P=0.018; LSD- t=4.951, P<0.001). Furthermore, the accuracy was comparable between digestive nurses and residents (LSD- t=-0.574, P=0.567). Compared with new Bing, digestive doctors had lower accuracy in answering questions related to adverse events of screening, follow-up recommendation of intestinal metaplasia, high risk factors and screening methods for colon cancer ( P<0.05), but higher accuracy in answering questions related to endoscopic adverse events and screening methods for esophageal cancer ( P<0.05). Multivariable liner regression analysis showed that being digestive doctors ( β=11.7, t=3.054, P=0.003) and questionnaire response time (≥7.6 min) ( β=7.8, t=2.894, P=0.005) were independent factors for the answer accuracy of digestive healthcare workers. Conclusion:Compared with digestive healthcare workers, New chatbots—new Bing has higher accuracy in answering gastrointestinal cancer screening-related questions, but performs poorly in answering questions such as adverse events of endoscopy and screening methods for esophageal cancer.