Four cellulolytic strains, which can be used as feed additive, were studied under the conditions of various temperature, incubation time, and anaerobic process, and examined the changes of their cell protein content, cellulase and hemi-cellulase activity. The results show: 1) The maximum cellulolytic enzyme activities were observed incubation 20h; 2) Constant medium temperature 28℃ was adequate to the growth of the 4 strains ; 3) anaerobic condition, 39℃±2℃ and fermentation 12h, 24h, 36h, the tested strains can growth well in PDA plate, however, the cellulolytic enzyme activities and growth of the tested strains were influenced adversely when fermentation 48h. The experiment provide many important basis for the strains production, storage and utilization.

Objective:To express Tumstatin_(183-230)-TRAIL fusion protein and to observe its biological functions.Methods: SOE-ing PCR was employed to amplify the recombinant sequence of Tumstatin_(183-230)and TNF-related apoptosis-inducing ligand (TRAIL_(114-281)).An expression vector pMAL-Tu-T was constructed by inserting Tu-T sequence into pMAL-c_2;the vector was used to transfect E.coli BL21(DE3)and expression of MBP-Tu-T fusion protein was induced by IPTG.Amylose Resin columns were employed to purify the fusion protein.The biological functions of MBP-Tu-T protein was examined by inhibitory test of endothelial cell proliferation,standard tumor cell cytotoxic assay,in vitro tube formation inhibition,and electron microscopic observation(apoptosis).Results:The expression rate of MBP-Tu-T fusion protein in E.coli was about 20%. Purified recombinant protein obviously inhibited endothelial cell proliferation(IC_(50)12.5?g/ml),induced apoptosis of pancreatic cancer cells,and inhibited tube formation.Conclusion:Constructed MBP-Tu-T fusion protein is bifunctional,which lays a solid foundation for further investigation of antitumor effect of Tumstatin_(183-230)-TRAIL in vivo.

Objective To investigate the effects of rapamycin on the expression of glioma patients tumor helicase RECQ1.Methods 50 glioma patients admitted to the department of neurosurgery in second hospital of hebei medical university were selected and randomly divided into 2 groups,25 patients in control group,were treated with routine admission surgical treatment;25 cases in the experimental group,firstly were given rapamycin capsule 1 mg,1 times/day orally,took 14 days in a row,and had surgical treatment after stopping drug a week.Glioma tissue samples were taken during the operation,mRNA and protein expression of tumor helicase RECQ1 were detected by reverse transcriptase polymerase chain reaction(RT-PCR)and Western blot.Results Glioma tumor helicase RECQ1 mRNA expression in the control group increased more significantly than experimental group,the optical density value in control group was(1.657 ±0.748),while the experimental group optical density value was(1.059 ±0.894),and the difference was statistically significant(P<0.05 );all organizations had the expression of tumor helicase RECQ1 protein,but gliomas tumor helicase RECQ1 protein expression in the experimental group patients(0.952 ±0.021)was significantly lower than that in the control group(1.211 ±0.024),and the difference was statistically significant(P<0.05).Conclusion Rapamycin capsule could reduce the expression of mRNA helicase RECQ1,inhibit DNA glial tumor cells of brain replication,effectively kill cancer cells,control the the progress of brain glioma,and improve prognosis,worth clinical promotion.

Objective To evaluate the application value of polymerase chain reaction (PCR)-fluorescence probe method in identifying genotypes of hepatitis C virus (HCV).Methods One hundred and sixty six serum samples from patients with chronic HCV infection were collected nationwide from March to June 2016.HCV Core-E1 gene region was amplified and sequenced by nested reverse transcription-PCR (RT nested-PCR)and genetic subtypes were analyzed by phylogenetic tree,meanwhile HCV genotypes were also determined by PCR-fluorescent probe method.Kappa test was used to compare the consistency of two methods.Results Among 166 samples detected by RT nested-PCR,the genotype of 66 samples (39.8%) was 1 b,34 (20.5%)was 2a,16 (9.6%)was 3a,27 was 3b (16.2%),23 (13.9%)was 6a.Two samples with 3b genotype detected by RT nested-PCR were identified as 1 b by PCR-fluorescent probe.The consistency rate of two methods was 98.7% (164 /166),there was no significant difference between two methods (χ2 =0.0492,P >0.05).Conclusion PCR-fluorescence probe method can accurately identify HCV genotypes and can be used in clinic.

Background and purpose:The incidence of thyroid cancer (TC) is increasing worldwide. However, there were some differences among different regions. The purpose of this study was to investigate the incidence trends and clinicopathological characteristics of TC in Qingdao, a typical eastern coastal city, and to analyze the change in etiological spectrum of surgical thyroid diseases in recent years.Methods:A total of 2 251 patients who underwent thyroidectomy in 2014 due to thyroid nodules at the Affiliated Hospital of Qingdao University were retrospectively reviewed. The clinico-pathological characteristics were further analyzed among 1 306 patients with TC and compared with the corresponding data from the Surveillance Epidemiology and End Results (SEER) database as well as previous data from this hospital.Results:With the increasing number of thyroidectomy in Qingdao, there was also an increase in the proportion of TC in patients after thyroidectomy, from 34.8% in 2010 to 59.0% in 2014. Among those with TC, the male-to-female ratio was 1∶2.80, with a relatively high incidence among 20-54 year-old adults, who were younger than those reported in terms of distribution of age in SEER database (U=2 289,P=0.000). About 50.2% of the TC patients were overweight or obese, 78.2% TC patients had only asymptomatic nodules detected by ultrasound at initial diagnosis, while 16.6% had visible or palpable thyroid nodules. Only 5.2% presented hoarseness or other repression symptoms. Micro-carcinoma accounted for 61.7% of TC in 2014 at this hospital, which was significantly higher than the proportion in 2010 (37.7%). Lymph node involvement was significantly more frequent at this hospital than in SEER database (49.5%vs 26.0%,χ2=11.806,P=0.001). Even among patients with micro-carcinoma, 31.3% already presented lymph node metastases. The proportions of papillary, follicular, medullary and anaplastic carcinoma were 97.5%, 1.1%, 1.0% and 0.5%, respectively, among which the percentage of papillary carcinoma was higher than that in SEER database (U=4 654.5,P=0.055).Conclusion:There was an increase in the number of thyroidectomy in Qingdao, and the preoperative diagnostic accuracy of TC in this area kept rising. The in-cidence of TC was relatively high in a younger population, with more common lymph node involvements and an overweight trend. The increasing proportion of micro-carcinoma might be related to the popularization of health examination. However, the frequent lymph node metastasis in patients with micro-carcinoma is an important indicator of the invasive behavior of micro-carcinoma, which should not be overlooked.

OBJECTIVETo observe morphological changes of enteric nervous system (ENS)-interstitial cells of Cajal (ICC)-smooth muscle cell (SMC) structure injury in deep muscle nerve plexus offunctional dyspepsia (FD) rats, and the repair of Shuwei Decoction (SD) on it, and to explore its effecton FD.

METHODSTotally 72 rats were randomly divided into the control group, the model group, the lowdose SD group, the medium dose SD group, and the high dose SD group, the Mosapride group, 12 ineach group. Rats in the low dose SD group, the medium dose SD group, and the high dose SD group were intragastrically fed with SD at 0.767, 1.534, 3.068 g/mL, respectively. Rats in the Mosapride group were intragastrically fed with Mosapride (1.37 mg/kg). FD rat model with Gan depression Pi deficiency syndrome (GDPDS) was established using complex pathogenic factors. Corresponding liquors were respectively administered to rats in corresponding groups from the 3rd day after modeling. Distilled water(10 mL/kg) was administered to rats in the control group and the model group, once per day for 14 successive days. Rats were sacrificed and small intestine tissues collected for observing ENS-ICC-SMC structure injury using immunofluorescence double labeling, laser scanning confocal microscope, and transmission electron microscope at day 15. Repair of SD on it was also observed.

RESULTSENS-ICC SMC structure was incomplete, with obvious injury in mutual link of ICC, ICC, SMC, and connecting structure. ENS-ICC-SMC structure was more complete in high, medium, and low dose SD groups, with close link of ICC and SMO. Their connecting structures were in good conditions.

CONCLUSIONSD could keep the integrity of ENS-ICC-SMC structure by promoting regeneration and morphology of ICC, thereby, improving gastrointestinal movement disorder and showing therapeutic effect on FD.

Animals ; Benzamides ; pharmacology ; Drugs, Chinese Herbal ; pharmacology ; Dyspepsia ; drug therapy ; Enteric Nervous System ; drug effects ; Interstitial Cells of Cajal ; drug effects ; Morpholines ; pharmacology ; Muscle, Smooth ; drug effects ; Random Allocation ; Rats

Objective: To chine and express the recombinant human endothelial monocyte-activating polypeptide-Ⅱ(EMAP-Ⅱ)and identify its anti-tumor biological activities.Methods: EMAP-Ⅱ_(147-312)was expressed by the expression vector pMAL-p2x and E.coli BL-21 and the product was purified.The production of tissue factor(TF)in human umbili- cal vein endothelial cell ECV-304 mediated by the recombinant EMAP-Ⅱwas determined by chemiluminescence sub- strate.The promoting effect of recombinant EMAP-Ⅱon TNF?-induced ECV-304 cell.Apoptosis was determined by flow cytometry.Its inhibitory effect on human pancreaic cancer cell SW1990 proliferation was determined by MTT method. Results:DNA sequencing verified that EMAP-Ⅱwas correctly cloned.The molecular mass of the protein identified by SDS-PAGE was consistent with the theoretic value.The productivity of recombinant EMAP-Ⅱwas 500?g per 1 g bacteria (wet mass).The purified product induced expression of tissue factor(TF)in ECV-304 cells;it also enhanced the sensi- tivity of ECV-304 cells to the apoptotic effect of TNF?([16.6?2.5]% vs[25.6?2.3]%,P

Objective To assess the effective length of jejunal graft when the 3~(rd) intestinal artery is u- tilized as vascular pedicle and afford a reliable theoretic base for clinical esophageal reconstruction.Methods In 32 formalin preserved and 21 fresh cadaver specimens,the diameter of 1st to 5th intestinal arteries and diameter of arterial arches are measured with linear calibre.Measure the length of jejunum that can be harves- ted as graft when the arches are extended.In the 21 fresh specimens,the 1st,2nd,4th and 5th intestinal ar- teries are ligated,acetic ester stained with red dye were injected into the lumen of 3rd intestinal artery via catheter.Extent of distribution of the arteries to the jejunum was observed.And then red ABS solution was in- jected into the 3rd intestinal artery to make into cast specimen.The blood supply distribution of jejunum through 3rd intestinal artery-arterial arch and communicating system were observed again.Results The di- ameter of the 3rd intestinal artery was the largest among the 1st to 5th intestinal arteries.The length of jejunum vascularized by 3rd intestinal artery can be as long as (142.2?62.3) (69.0～206.60cm) in acetic ester in- filtrated specimens.While in ABS east specimen,the average available extent of donor jejunum was(30.8?7.3) (23.0～37.3cm).Conclusion As observed by this applied anatomy study,the jejunum graft vascu- larized by 3rd intestinal artery alone has sufficient length to meet the need of esophageal reeonstrution.

Objective To observe the clinical efficacy of balance acupuncture predominantly by puncturing Jiaji points (EX-B2) from C4 to T1and from T12 to L1in treating hemiplegic balance disturbance after cerebral stroke. Method A total of 180 hemiplegia patients were randomized into 3 groups, 60 cases in balance acupuncture group, 60 cases in ordinary acupuncture group, and 60 cases in basic control group. After 1-month treatment and 3 months after the treatment, the motor function (Fugl-Meyer Assessment, FMA), balance function (Berg Balance Scale, BBS; Timed Up and Go Test, TUGT), and activities of daily living (ADL) (Barthel Index, BI) were observed.Result After 1-month treatment and 3 months after the treatment, limb function, balance ability and ADL were significantly different from those before the treatment in balance acupuncture group (P<0.01); after 1-month treatment, limb function in balance acupuncture group was significantly different from that in basic control group (P<0.05), and the differences were more statistically significant in comparing the rest indexes between the two groups (P<0.01); there were significant differences between ordinary acupuncture group and basic control group (P<0.05). Three months after the treatment, there was a significant difference in comparing balance function between balance acupuncture group and basic control group (P<0.05), and the differences were more statistically significant in comparing the rest indexes between the two groups (P<0.01), there was no significant difference in comparing balance function between ordinary acupuncture group and basic control group (P>0.05).Conclusion In combination with basic treatment, balance acupuncture works better than ordinary acupuncture and basic control in improving limb function, ADL and balance function of hemiplegia patients.

OBJECTIVETo construct the recombinant virus-like particles containing HCV envelope glycoprotein E1E2 based on sindbis virus vector.

METHODSThe gene encoding HCV envelope glycoprotein E1E2 was cloned into sindbis virus vector to construct recombinant plasmids pBR-XJE1E2 and pVA-XJE1E2, and transfect them into BHK-21 cells to obtain recombinant virus-like particles. The expression of E1 and E2 protein were verified by Western Blot and indirect immunofluorescent assay (IFA).

RESULTSThe results of restriction enzyme digestion, PCR and sequencing analysis showed that the recombinant plasmids were constructed successfully. And the results of RT-PCR, Western blotting and IFA detection showed that the transfect cells could package HCV-like particles of expressing structural proteins E1E2.

CONCLUSIONThe recombinant expression plasmids pBR-XJE1E2 and pVA-XJE1E2 based on sindbis virus vector could package HCV-like particles in eukaryotic cell, which provides a foundation for further study of its in vivo animal immune response.

Animals ; Cells, Cultured ; Cricetinae ; Genetic Vectors ; Hepacivirus ; genetics ; Plasmids ; Recombination, Genetic ; Sindbis Virus ; genetics ; Viral Envelope Proteins ; genetics