This study aims to establish a convenient,new and visual detection method for the field diagnosis of Pasteurella multocida(Pm).With reference to the Pm kmt1 gene conserved sequence published in GenBank,PCR amplification primers were designed,the amplified kmt1 gene was cloned into pMD19-T vector,and the recombinant plasmid standard pMD19-T-kmt 1 was estab-lished and identified by PCR and sequencing.Using pMD1 9-T-kmt 1 plasmid as template and kmt1 gene as target gene,basic primers were designed and synthesized.According to the requirements of LFD,a probe(Pm-P)was designed,and the RPA-LFD method for Pm detection was established by optimizing the reaction conditions.Specificity and sensitivity tests were carried out,and 64 clini-cal samples were tested by the method.The results showed that the established Pm RPA-LFD method could be amplified at 37 ℃ for 15 min.Escherichia coli(E.coli),Salmonella enteriditis(SE),Riemerella anatipestifer(RA),Staphylococcus,goose parvovirus(GPV),duck plague virus(DPV),Muscovy duck parvovirus(MDPV)DNA was extracted as the template,and plasmid standard pMD19-T-kmt 1 was used as the positive control.All the positive controls were negative,indicating that the method had good specificity.The plasmid standard pMD1 9-T-kmt 1 was diluted with a 10-fold ratio,and the plasmid standard with a concentration of 107-100 copies/μL was used as the template.The sensitivity was 1.50×101 copies/μ,,which was 100 times higher than that of PCR.A total of 64 clinical samples with suspected RA were subjected to testing using PCR,RPA and LAMP-LFD,with a 100％compliance rate for all three detection tests.The results show that the established RPA-LFD method has the characteristics of strong specificity,high sensitivity,fast speed and visualization,and can be applied to the field detection of Pm.

This study aims to establish a convenient,new and visual detection method for the field diagnosis of Pasteurella multocida(Pm).With reference to the Pm kmt1 gene conserved sequence published in GenBank,PCR amplification primers were designed,the amplified kmt1 gene was cloned into pMD19-T vector,and the recombinant plasmid standard pMD19-T-kmt 1 was estab-lished and identified by PCR and sequencing.Using pMD1 9-T-kmt 1 plasmid as template and kmt1 gene as target gene,basic primers were designed and synthesized.According to the requirements of LFD,a probe(Pm-P)was designed,and the RPA-LFD method for Pm detection was established by optimizing the reaction conditions.Specificity and sensitivity tests were carried out,and 64 clini-cal samples were tested by the method.The results showed that the established Pm RPA-LFD method could be amplified at 37 ℃ for 15 min.Escherichia coli(E.coli),Salmonella enteriditis(SE),Riemerella anatipestifer(RA),Staphylococcus,goose parvovirus(GPV),duck plague virus(DPV),Muscovy duck parvovirus(MDPV)DNA was extracted as the template,and plasmid standard pMD19-T-kmt 1 was used as the positive control.All the positive controls were negative,indicating that the method had good specificity.The plasmid standard pMD1 9-T-kmt 1 was diluted with a 10-fold ratio,and the plasmid standard with a concentration of 107-100 copies/μL was used as the template.The sensitivity was 1.50×101 copies/μ,,which was 100 times higher than that of PCR.A total of 64 clinical samples with suspected RA were subjected to testing using PCR,RPA and LAMP-LFD,with a 100％compliance rate for all three detection tests.The results show that the established RPA-LFD method has the characteristics of strong specificity,high sensitivity,fast speed and visualization,and can be applied to the field detection of Pm.

Objective To investigate the clinical efficacy of endoscopic resection,laparoscopic resection and open resection in the treatment of gastric stromal tumor (GIST).Methods The retrospective cross-sectional study was conducted.The clinicopathological data of 254 GIST patients who were admitted to the First Affiliated Hospital of Army Medical University between January 2007 and June 2017 were collected.The endoscopic submucosal dissection (ESD) and laparoscopic or open wedge resection of GIST were performed according to the patients' conditions.Observation indicators:(1) surgical and postoperative recovery situations;(2) postoperative pathological examination;(3) follow-up and survival situations.Follow-up using outpatient examination and telephone interview was performed to detect postoperative adjuvant therapy and survival up to June 2017.Measurement data with normal distribution were represented as-x±s.Measurement data with skewed distribution were described as M (range).Results (1) Surgical and postoperative recovery situations:of 254 patients,112 underwent ESD,including 111 with successful operation and 1 with intraoperative conversion to open surgery due to excessive bleeding-induced blurred operating view,93 underwent successful laparoscopic wedge resection of GIST and 49 underwent successful open wedge resection of GIST.The operation time,volume of intraoperative blood loss,time for initial fluid diet intake,duration of hospital stay and hospital expenses were respectively (75±21) minutes,(6.9±0.5)mL,(2.8±0.9)days,(5.5 ± 0.2)days,(22 167±1 364) yuan in patients with ESD and (137±65) minutes,(48.1±2.6)mL,(3.9±1.4) days,(8.3 ± 2.2)days,(32 937±1 823) yuan in patients with laparoscopic operation and (168± 60) minutes,(157.2± 10.3) mL,(5.8± 1.7) days,(11.3 ± 3.5) days,(38 462± 1 961) yuan in patients with open operation.Two patients with ESD had subcutaneous emphysema and didn't receive special treatment,and then emphysema disappeared after 2 days.No complication was detected in patients with laparoscopic or open operations.(2) Postoperative pathological examination:tumor diameter in patients with ESD,laparoscopic operation and open operation was respectively (2.6± 1.6)cm,(6.1 ±2.2)cm and (6.4±2.3) cm.The cases with positive CD117,discovered on GIST-1 (DOG1),CD34 and smooth muscle actin (SMA) were 106,105,86,17 with ESD and 89,87,59,11 with laparoscopic operation and 46,47,30,8 with open operation,respectively.The extremely low risk,low risk,medium risk and high risk were respectively detected in 67,42,3,0 patients with ESD and 16,36,23,18 patients with laparoscopic operation and 7,20,14,8 in patients with open operation.(3) Follow-up and survival situations:210 of 254 patients were followed up for 6.0-120.0 months,with an average time of 36.0 months,including 86 with ESD,82 with laparoscopic operation and 42 with open operation.During the follow-up,of patients with ESD,3 patients with medium risk respectively received imatinib therapy for 7.0 months,1.5 years and 2.0 years,and postoperative gastroscopy reexaminations every 6 months,without tumor recurrence;81 with extremely low risk and low risk received postoperative gastroscopy reexaminations every 6 months and didn't receive targeted therapy,without tumor recurrence;2 died of non-stromal tumor.The postoperative average survival time,1-,3-and 5-year survival rates were respectively 56.3 months,98.8％ (81/82),91.5％ (75/82),74.4％ (61/82) in patients with laparoscopic surgery and 52.4 months,97.6％ (41/42),85.7％ (36/42),81.0％ (34/42) in patients with open surgery.Conclusions According to patients' conditions,endoscopic resection,laparoscopic resection and open resection are safe and feasible in the treatment of GIST.Endoscopic resection of GIST should be selectively applied to patients with smaller diameter,with advantages of lower hospitalization expenses and better long-term prognosis.

Objective To evaluate the clinical efficacy and analyze the prognostic factors of radiotherapy after breast-conserving surgery for stage Ⅰ-Ⅱ breast cancer patients.Methods Clinical efficacy of adjuvant radiotherapy in 1 376 patients with stage Ⅰ and Ⅱ (T1-2 N0-1 M0/T3NoM0) breast cancer after undergoing unilateral breast-conserving surgery between 1999 and 2013 was retrospectively reviewed.Among them,930 patients (67.6％) received radiotherapy combined with chemotherapy including 517 receiving radiotherapy followed by chemotherapy and 413 receiving chemotherapy followed by radiotherapy.In total,1 055 patients (76.7％) were treated with endocrine therapy.Eighty-six patients (39.6％) positive for HER-2 received targeted therapy.The overall survival (OS) and disease-free survival (DFS) rates were calculated using the Kaplan-Meier method.Univariate analysis was performed by Log-rank test and multivariate analysis was conducted by Cox regression method.Results The median follow-up time was 55 months.The quantity of patients receiving follow-up for ≥ 10 years was 90.The 5-and 10-year OS rates for all patients were 98.6％ and 91.5％,and 94.6％ and 82.8％ for 5-and 10-year DFS rates.Mutivariate analysis revealed that age (P=0.016),T staging (P =0.006),N staging (P =0.004),lymphovascular invasion (P =0.038) and time interval between radiotherapy and surgery (P=0.048) were independent prognostic factors for DFS rate.Multivariate analysis demonstrated that N staging (P=0.044) and ER (P=0.026) were independent prognostic factors for DFS in the radiotherapy alone group.Conclusions The radiotherapy-based comprehensive treatment yields favorable clinical outcomes for stage Ⅰ-Ⅱ breast cancer patients after undergoing breast conserving surgery.The prognostic factors for DFS include age,T staging,N staging,lymphovascular invasion and the time interval between radiotherapy and breast-conserving surgery.In the radiotherapy alone group,DFS rate is associated with N staging and ER level.

Objective To evaluate the differences, including clinical symptoms, endoscopic and histopathologic findings, status of Helicobacter pylori (Hp) infection and cytokeratin (CK) expressions, be-tween Barrett esophagus (BE) and heterotopic gastric mucosa (HGM) in upper esophagus. Methods Clinical data of 152 patients with BE and 52 patients with HGM in upper esophagus diagnosed from February 2004 to September 2005 were retrospectively studied. The parameters being compared include-ed clinical manifestations, conventional and magnifying endoscopic findings, histopathological findings, Hp infection determined by rapid urease test and Warthin-Starry staining and expression of CK phenotypes detec-ted by immunohistochemistry. Results Gastro-esophngeal reflux was observed in 64. 5% of patients with BE (98/152), higher than that in patients with HGM ( 13.5%, 7/52, χ2 = 40. 36, P < 0. 01 ). Endoscopic faveolus of BE mucosa included 46 cases of spot pattern, 65 striations and 41 villiform patterns, while those of HGM were all striation patterns. The histologic classification in BE included 56 cases of fundic type, 39 junction type and 57 specialized intestinal metaplasia, while in HGM mucesa, 31 cases of fundic type, 16 junction type and 5 antrum type were diagnosed, and no goblet cells were found. Moderate and severe infil-tration of inflammatory cells in BE mucosa was 63.2% (96/152), which was significantly higher than that in HGM mucosa (15/52, 28. 8%, P<0. 01). However, no difference was found in gastric antrum inflam-mation between the two groups (44.7%, 68/152, vs. 51.9%, 27/52, P>0.05). No difference was ob-served in prevalence of Hp infection between BE and HGM groups (P >0. 05 ), either in involved mucosa or in antrum. CK7 was not expressed in HGM or normal squamons mucosa, but was expressed in BE. CK20 and CK19 were expressed in both HGM and BE, and CK13 expression was found in some BE nmcosa including gas-tric metaplesia (55/95) and intestinal metaplasia (29/57) but not in HGM mucosa. Conclusion There are differences between HGM and BE, in regarding of reflux symptoms, magnifying endoscopic findings, histo-logical types and CKs expressions, which may be indicators to make differential diagnosis.

Objective To analyze and investigate endoscopic and pathological characteristics in the diagnosis of early esophageal carcinoma. Methods The clinical data of 62 patients with early esophageal carcinoma who had been admitted to Southwest Hospital from January 2003 to December 2007 were retrospectively analyzed. Results Of all patients, 8 had upper esophageal carcinoma, 42 had middle esophageal carcinoma and 12 had lower esophageal carcinoma. The tumor growth patterns included surface diffusion growth (n = 30), bidirectional growth (n = 11), intracavitary growth (n = 9), intra-esophageal wall growth (n = 7) and mixed growth (n = 5). The diameter of lesions ranged from 1.0 cm to 2.9 cm in majority. Surgical resection was done in 38 patients, endoscopic mucosal resection in 23 patients and endoscopic submucosal dissection in 1 patient. Postoperative pathological examination showed that 13 patients had in situ carcinoma, 22 had intramucosal carcinoma and 27 had submucosal carcinoma. The lymph node metastatic rate of intramucosal carcinoma and submucosal carcinoma were 5% (1/22) and 15% (4/27), respectively. The numbers of squamous cell carcinoma, adenocarcinoma, small cell carcinoma, sarcoma carcinoma and spindle carcinoma were 57, 2, 1, 1, 1, respectively. ConelusionsEarly esophageal carcinoma tends to appear in the middle part of esophagus, and with surface diffusion growth type. Most of the early esophageal carcinoma are squamous cell carcinoma. Endoscopy combined with lugol's iodine and methyleneblue staining is effective in detecting early esophageal carcinoma.

Objective To investigate the endoscopic and ultrasonic endoscopic features of colorectal carcinoid and the indications of endoscopic treatment. Methods The clinical data of 22 patients with colorectal carcinoid who had been admitted to our hospital from 2002 to 2007 were collected. The endoscopic and ultrasonic endoscopic features and the relationship between the features and invasion depth of colorectal carcinoid were analyzed. Results Under the endoscope, early carcinoid presented submucosa tumor with 1.5cm in diameter, and yellow or white smooth surface; advanced carcinoid presented submucosa tumor with 0.8-3.0cm in diameter, and yellow or white little nodus or ulcerative surface. The ultrasonic endoscopic feature of the colorectal carcinoid was orbicular-ovate low level echo tumor with punctiform slightly high-level echo and an unsharpness edge. Sixteen mucosal layer-cancers and submucosal layer-cancers were removed by endoscopic mucosal resection, and 10 of them were additional treated by argon plasma coagulation. After a follow-up period of 4-36 months, no recurrence was observed. Conclusions Endoscopy and endoscopic ultrasonography are effective methods to diagnose colorectal carcinoid and its invasion depth. Endoscopic treatment is a simple, safe and effective means to treat the early colorectal carcinoid tumors.

Objective To assess the potential diagnostic value of the potential of flexible image col-or enhancement system(FICE)with magnifying endoscopy for superficial esophageal lesions by observing the intrapapillary capillary loops(IPCL)in esophageal mucosa. Methods IPCL in patients with esophageal diseases were studied with Fujinon EG-590ZW FICE endoscopy. The relationship between changes of IPCL in 31 cases of superficial esophageal lesions and pathological findings was studied. Results The vascular patterns of IPCL were analyzed and classified as follows: Type Ⅰ,often seen in normal esophagus,were e- venly distributed with regular forms. TypeⅡ,protracted,often occured in esophagitis. Type Ⅲ showed two or three of such four changes as,dilatation,wave-like inflection,irregular caliber and deformity,which was mainly seen in dysplasia. While Type Ⅳshowed all the four changes mentioned above and frequently ap- peared in esophageal cancer. Of the 17 cases of esophagitis,fifteen cases were TypeⅡIPCL and the others Type Ⅲ. Of the 10 cases of dysplasia,eight were Type Ⅲ IPCL and two high-grade dysplasia showed Type Ⅳ IPCL. One sml early esophageal cancer showed Type Ⅲ IPCL and all three advanced esophageal cancer showed TypeⅣ IPCL. Conclusion The esophageal microvessels can be clearly seen by FICE with magnif- ying endoscopy,and the differentiation of superficial esophageal lesions can be fairly performed with the ob- servation of IPCL,it gives an important practical siginificance in diagnosing tumorous lesions.

Objective To clarify the changes of ultrastructure of interstitial cells of Cajal in the small intestine of diabetic rats. Methods Rats were randomly divided into diabetic group and control group. The rate of the small intestinal transit was measured and the tissues of the small intestine were observed by transmission electron microscopy at 3 months after the establishment of rat model of diabetics. Results In the diabetic rats, the rate of small intestinal transit was delayed as compared with that in the control group. The number of the gap junction of interstitial cells of Cajal in the small intestine of diabetic rats decreased significantly, and the rest structures were damaged. Damaged organelles and formation of vacuoles were also found. Conclusion The changes of ultrastructure of interstitial cells of Cajal in the small intestine may be one of the mechanisms resulting in slow rate of the small intestinal transit in diabetic rats.

Objective To study the effect of carbachol(CCH) on membrane potential of smooth muscle cells of gastric antrum of diabetic rats to verify the mechanism of gastric motility disturbance.Methods Fifty healthy Wistar rats of 2-month old,weighing 100-160 g,were divided into the control group(n=20) and diabetes mellitus group(n=30).Three months after the establishment of the rat model of diabetes by intraperitoneal injection of 60 mg/kg streptozotocin,gastric emptying time and gastric electrical activity were measured,and the resting membrane potentials of smooth muscle cells in antrum were measured by confocal laser scanning microscopy,then treated with carbachol of 10~(-9) mmol/L,10~(-8) mmol/L,10~(-7) mmol/L,the membrane potentials were measured.Results As compared with the normal rats,gastric emptying time in diabetic rats was significantly longer and abnormal gastric electric rhythm was significantly increased,the abnormal rhythm index(ARI) and the coefficient of variation(CV) of slow wave frequency in diabetic rats were significantly higher,but the resting membrane potentials remained unchanged and the sensitivity of CCH-induced membrane depolarization was increased.Conclusion The increase of sensitivity of CCH-induced membrane depolarization may be involved in the diabetes-induced gastric motor disorders.