Objective To use HSV 1 tk (herpes simplex virus type Ⅰthymidine kinase)/GCV (ganciclovir) in androgen independent prostate cancer cells(C4 2,PC3) in vitro in order to provide useful basis for clinical use. Methods HSV 1 tk gene was ligated to a pN 2A retroviral vector. Recombinant DNA molecules being introduced into a packaging cell line PA317,the high titer virus producer cells (VPC) were screened.The integration and expression of HSV 1 tk gene in VPC was observed by PCR and RT PCR. VPC was co cultured with these cancer cells in the light of 1∶1,1∶2,1∶4,1∶8.Cell viability (cytotoxicity) was assessed by SRB (sulforhodamine B protein dye binding)after the first day,the third day,the fifth and the seventh day. Results The highest titer VPC producing HSV 1 tk gene was isolated. Retrovirus mediated HSV 1 tk gene therapy was effective and active against such prostate cancer cells.The best one was co culture of VPC and cancer cells at 1∶1 and the fifth day followed by GCV. Compared with C4 2, PC3 decreased remarkably.The activation of apoptosis and other ways failed to be found. Conclusions Retrovirus mediated HSV 1 tk gene therapy in Vitro directly killed the tumor cells by cytolytic activity.

In order to optimize acupuncture treatment protocol for functional defecation disorders, literature during past 20 years is reviewed, and factors which influences acupuncture therapeutic effect are analyzed from aspects of acupoint, acupuncture technique, etc. As for the selection of acupoint, more attention should be paid on the use of Baliao, especially Zhongliao (BL 33) and Xialiao (BL 34); when Baliao is deeply needled, it is essential to acquire scientific technique. The relationship between acupuncture parameter (including electroacupuncture waveform and frequency), acupuncture techniques (including reinforcing and reducing technique, quantity of stimulation, etc. ) and acupuncture efficacy is complicated, and the scientific values of present research conclusion are in need, of further improvement. The diagnosis and treatment awareness on types of functional defecation should be strengthened, and the mental health of the constipation patients should be concerned. The combination of acupuncture and biofeedback training can have a synergistic effect, which is benefit to achieve a better long-term effect. Based on this, acupuncture treatment protocol for functional defecation disorders can be optimized to further improve the efficacy.
Acupuncture Points ; Acupuncture Therapy ; methods ; Constipation ; physiopathology ; therapy ; Defecation ; Female ; Humans ; Male

Objective To investigate the distribution characteristics and risk factors of infectious pathogens after open limb fracture surgery.Methods From January 2016 to December 2018,180 patients with open limb fracture admitted to Hangzhou Hospital of Zhejiang Medical and Health Group were selected to observe the infection after operation.Pathogens were isolated and identified by automatic biological analyzer and bacterial identification system,and drug resistance test was carried out by K-B method.Multivariate logistic regression analysis was used to analyze the risk factors of infection after open limb fracture surgery.Results Among 180 cases of open limb fracture,29 cases had postoperative infection,the infection rate was 16.11％.Among 29 cases of post-operative infection,34 strains of pathogenic bacteria were isolated,including 19 strains of Gram-positive bacteria,13 strains of Gram-negative bacteria and 2 strains of fungi.The resistance rate of Staphylococcus aureus to penicillin G was high,and the resistance rate was 80.00％.Univariate analysis showed that there were no statistically significant differences in gender,BMI,injury site,smoking history,hypertension and intraoperative bleeding between the two groups (x2 =0.252,0.416,0.734,0.856,0.572,all P ＞ 0.05).There were statistically significant differences in age,diabetes mellitus,operation time,hospitalization time,wound contamination and drainage tube placement time between the two groups (x2 =21.537,9.664,17.244,15.459,24.327,19.804,all P ＜ 0.05).The single factor analysis showed that age ＞ 60 years old,diabetes mellitus,operation time ＞ 3h,hospitalization time ＞ 14d,wound contamination and drainage tube placement time ＞ 4d were independent risk factors for infection after open limb fracture surgery.Conclusion Postoperative infection rate of open limb fracture is high.Gram-positive bacteria are the main pathogenic bacteria.Postoperative infection is affected by many factors.In order to prevent postoperative infection,specific measures should be taken and antibiotics should be used reasonably in strict accordance with drug sensitivity test.

Objective To investigate the clinical features and treatment of solid pseudopapillary neoplasm (SPN) of the pancreas.Methods The retrospective cross-sectional study was conducted.The clinicopathological data of 69 patients with SPN of the pancreas who were admitted to the Qilu Hospital of Shandong University from January 2012 to July 2017 were collected.Serum tumor markers detection,enhanced computed tomography (CT) and magnetic resonance imaging (MRI) of abdomen were carried out preoperatively for all the patients,and a part of the patients received endoscopic ultrasonography (EUS).Surgery plans were formulated after completion of examinations.Observation indicators:(1) clinical features;(2) treatment situation;(3) results of pathological examination;(4)follow-up.All the patients were followed up via outpatient examination and telephone interview to detect the survival and tumor recurrence and metastasis till July 2017.Measurement data with normal distribution were presented by (x)±s and were compared by Student's t test.Count data were compared by chi-square test.Results (1) Clinical features:① epidemiologic features:the ratio of male to female was 1∶5.9;patients were aged between 9 and 65 years,including 40 under 30 years and 29 above or equal to 30 years.The onset age was (34± 15)years for male patients and (28 ± 11)years for female patients,respectively,with no statistically significant difference (t=1.364,P＞0.05).Of 69 patients,SPN was located at pancreatic uncinate process in 25 patients,at neck of pancreas in 12 patients,body and tail of pancreas in 32 patients.② Medical history:history of acute or chronic pancreatitis and abdominal trauma were denied by all the 69 patients.③ Clinical manifestation:26 patients had no obvious symptoms and were detected by physical examination;31 patients had discomfort in upper abdomen,nausea and vomiting;other patients were admitted to the hospital because of upper abdominal mass (10 patients),jaundice (1 patient) or nausea,constipation (1 patient).④ Laboratory examination:the levels of preoperative carcinoembryonic antigen (CEA) and CA19-9 were normal.⑤ Imaging examination:plane scan of the CT examination showed round or round-like low-density shadows in the 69 patients,including 51 of cystic solid lesion,13 of solid lesion and 5 of cystic lesion.Complete capsules were observed in 64 patients,blurred boundary between pancreas and adjacent viscera in 5 patients,calcified foci in the pancreatic parenchyma and capsules in 14 patients.Ten patients received MRI examination,and the T1-weighted images showed equal or slightly lower signal,T2-weighted images showed slightly higher signal in the plane scan,and T1-weighted and T2-weighted images of the tumor capsule showed continuous or non-continuous ring-like signal.The results of enhanced scan showed slightly heterogeneous enhancement of the capsule and the parenchyma of the pancreas in the arterial phase,and progressive enhancement in the venous and delayed phase,while the enhancement degree was lower than that of the normal pancreas parenchyma.The parenchyma was cloud-,papillaor mural nodule-like enhanced.Obvious enhancement was observed in capsule while not in the cystic components.The boundaries of the tumors in 5 patients were clear under EUS.Hypo-,iso-and hyperechoic regions were found in the masses,and the masses were confirmed as cyst-solidary type.Obvious calcified foci were found in 1 patient.(2) Treatment outcome:twenty-seven patients received laparoscopic surgery,including tumor expiration in 13 patients,distal pancreatectomy with preservation of spleen in 8 patients,distal pancreatectomy combined with splenectomy in 2 patients,middle pancreatectomy in 2 patients,pancreaticoduodenectomy with preservation of pylorus in 1 patient,pancreatic head resection with preservation of duodenum under the assistance of laparoscopy in 1 patient.Forty-two patients received open surgery,including tumor expiration in 12 patients,distal pancreateetomy with preservation of spleen in 10 patients,distal pancreatectomy combined with spleneetomy in 6 patients,middle pancreatectomy in 5 patients,pancreaticoduodenectomy in 7 patients (with preservation of pylorus in 2 patients) and pancreatic head resection combined with preservation of duodenum in 1 patient.One patient with SPN + hepatic metastasis received distal pancreatectomy+ metastatic foci resection in the lesser omental bursa,and then followed by radiofrequency ablation in the hepatic metastatic foci.Postoperative complications:21 of 69patients had postoperative complications,including 17 intestinal fistulas,2 abdominal bleedings,1 incomplete obstruction,1 pleural effusion + atelectasis,and all of them were cured by symptomatic treatment.(3)Pathological examination:the resection margins of 69 patients were negative.The mean diameter of the tumor was (7±4) cm (21 patients with tumor diameter ＜ 5 cm,and 48 with tumor diameter ≥5 cm).The tumor diameters of 4 in 10 male patients were above or equal to 5 cm,and the number was 44 in 59 female patients,with statistically significant difference (x2 =4.828,P＜0.05).The tumor diameters of 32 in 40 patients who aged under 30 years were above or equal to 5 cm,and the number was 16 in 29 patients who were aged above or equal to 30 years,with statistically significant difference (x2=4.895,P＜0.05).Solid,pseudo-papillary and cystic regions in the SPN tissues were seen under the light microscope.Tumor cells were surrounded the blood vessels and were arranged in the nest or sheet shape in the solid region;blood vessels were surrounded by one or multiple layers of tumor cells in the axis or pseudopapillary shape in the pseudopapillary region;large amount of mucus and clusters of blood cells were seen in the cystic regions.The result of immunohistochemistry showed that the positive rates of α1-antitrypsin,vimentin,β-catenin,progesterone receptor,CD10,synaptophysin and chromogranin A were 100.0％ (39/39),96.6％ (28/29),95.7％ (45/47),94.4％ (51/54),92.5％ (49/53),72.9％ (35/48) and 5.6％ (3/54),respectively.(4) Follow-up:63 of 69 patients were followed up for 1-68 months,with median time of 29 months.No SPN recurrence or metastasis was detected.One patient died of lung cancer at postoperative month 35 and other patients survived well.Conclusions SPN of pancreas is mostly detected in young female patients,and it could be solid or cystic.Abdominal enhanced CT or MRI examination could clarify the diagnosis.EUS-fine needle aspiration examination could provide pathological evidence for definitive diagnosis.Typical cellular morphology and pseudopapillary regions may provide hints for the diagnosis of SPN,and the diagnosis could be clarified when combined with the detection of vimentin,α 1-antitrypsin or other indexes.Complete resection of SPN and ensure negative resection margin are fundamental principles of treatment.

Objective To identify Mobiluncus species with matrix-assisted laser desorption/ioniza-tion time-of-flight mass spectrometry (MALDI-TOF MS) and 16S rRNA gene sequencing and to analyze anti-biotic resistance genes and phylogenetic relationships among Mobiluncus species with whole genome sequen-cing. Methods Twenty-five Mobiluncus strains were isolated after anaerobic culture of 65 vaginal secretion samples of patients with bacterial vaginosis (BV) and identified to species level by MALDI-TOF MS and 16S rRNA gene sequencing. The whole genome DNA of each strain was extracted for next-generation sequencing. SPAdes was used to assembly genomes. Resistance genes were searched in ARGD database. The core ge-nome of all isolates was analyzed by Harvest to construct phylogenetic tree. Results MALDI-TOF MS could only identify Mobiluncus curtisii, while 16S rRNA gene sequencing could identify both Mobiluncus mulieris and Mobiluncus curtisii. Results of the whole genome sequencing showed that tetracycline resistance gene tet ( o) and macrolides resistance gene erm(x) were the two predominant acquired resistance genes of Mobilun-cus with a positive rate of 84. 7% and 61. 5% respectively. Intra-species relationships of the two Mobiluncus species were close, but a distant phylogenetic relationship was found between the two species. Conclusion This study shows that MALDI-TOF MS can't be used to identify Mobiluncus mulieris at present. Mobiluncus strains have potential resistance to tetracycline and macrolides. Intra-species evolution of Mobiluncus is slow, which indicates that there is no growing trend towards new species for the time being.

ObjectiveTo investigate the effect of Bufeitang on intestinal flora of rats with lung Qi-deficiency syndrome of chronic obstructive pulmonary disease（COPD）， and to explore the mechanism of traditional Chinese medicine in regulating intestinal flora and thus restoring the balance of lung-gut axis. MethodA total of 84 rats were randomly divided into 7 groups， including blank group， model group， fecal bacterial transplantation（FMT） group， dexamethasone group and low， medium and high dose groups of Bufeitang， 12 rats in each group. Except for the blank group， cigarette and sawdust fumigation combined with intratracheal instillation of lipopolysaccharide（LPS） were used to establish the COPD rat model with lung Qi-deficiency syndrome in all other groups. The low， medium and high dose groups of Bufeitang were intragastric administrated with Bufeitang（3.645， 7.29， 14.58 g·kg^-1）， the FMT group was given fecal bacteria liquid enema（10 mL·kg^-1）， dexamethasone group was given dexamethasone acetate tablet suspension by gavage（0.135 mg·kg^-1）， the blank group and model group were given equal amount of distilled water. Fresh feces were collected after 28 d of continuous intervention for 16S rRNA gene sequencing. Lung and colon tissues were stained with hematoxylin-eosin（HE） for pathomorphological observation， and enzyme-linked immunosorbent assay （ELISA） was performed to detect the contents of tumor necrosis factor-α（TNF-α） and interleukin-8（IL-8） in lung tissues. ResultCompared with the blank group， the model group showed severe abnormal lung tissue structure with alveolar atrophy and collapse accompanied by severe inflammatory cell infiltration. Compared with the model group， the extent of injury was significantly improved， and inflammatory cell infiltration was reduced with basically normal alveolar structure in the high dose group of Bufeitang. Compared with the blank group， the model group had severely abnormal colonic tissue structure， the epithelial cells in the mucosal layer were eroded and shed， the number of inflammatory cells increased， the submucosal layer was edematous and the gap was enlarged. Compared with the model group， the extent of damage was significantly improved in the medium and high dose groups of Bufeitang， the epithelial cells in the mucosal layer were neatly and closely arranged， with only a small amount of inflammatory cell infiltration and no significant degeneration. Compared with the blank group， the TNF-α and IL-8 levels of lung tissue in the model group were significantly increased（P<0.01）. Compared with the model group， the TNF-α and IL-8 levels of lung tissues in the low， medium and high dose groups of Bufeitang were significantly decreased（P<0.01）. Bufeitang significantly modulated the number of bacteria species as well as alpha and beta diversity of model rats， corrected the return of intestinal flora to normal abundance and diversity， and positively regulated 4 differential phyla（such as Firmicutes， Proteobacteria） and 13 differential genera（such as Turicibacter， Lactobacillus， Anaerobiospirillum， Intestinimonas） in COPD model rats with lung Qi-deficiency syndrome， and down-regulated 2 carbohydrate metabolic pathway functions， including the pentose phosphate pathway（non-oxidative branch） Ⅰ and the Calvin-Benson-Bassham cycle. ConclusionBufeitang can modulate the abundance and diversity of intestinal flora species， affect the function of metabolic pathways， repair the structure of lung and colon tissues， regulate the level of inflammatory factors， and thus improve COPD with lung Qi-deficiency syndrome. The mechanism may be related to its regulation of inflammation-related intestinal flora to restore the balance of lung-gut axis in COPD with lung Qi-deficiency syndrome.