The influenza A virus matrix protein2 gene(M2)which deleted transmembrane region was amplified by overlap extending PCR,and the multiepitope gene of hemagglutinin(HA)was PCR amplified with seven continuous synthesized segments by designing primer.The two gene segments were separately cloned into pMD18T vector to sequence analysis and prokarytic expression vector pET28a+ to construct the recombinant plasmid pET28a+M2dHA.The recombinant plasmid was transformed into E.coli BL21(DE3),and the high expression strain was obtained by screening monoclones.The recombinant protein existed as inclusion bodies,which accounted about 45% of the total cellular protein.The inclusion bodies were washed with 1% Triton X100 solution twice,and dissolved in 8 mol/L urea solution.The solution protein was purified by Ni+2 affinity chromatography,and refolded by dilution renaturation,then purified by Q Sepharose FF cation exchange column.The purity of the protein was over 90% by HPLC analysis.The result of Western blot showed it has good antigenicity and specificity.These results strongly supported for the further study of the broadspectrum influenza virus subunit vaccine.

The gene of mutated TNF-?D4 gene was amplified by overlap PCR and cloned into the prokaryotic expressive vector pBV220.TNF-?D4 contains two changes:substitutions of Pro8Arg,Ser9Lys,Asp10Arg,Ile157Phe,Leu29Ser,Arg31Val and a deletion of the N terminal four amino acids.The recombinant vector pBV220-TNF-?D4 was transformated into E.coli strain DH5?,and the high expression strain was obtained by screening monoclones.The level of expression was about 45% of total cell protein.After purification,the purity of fusion protein was above 90% by HPLC and relative ability was 8 ?107.TNF-?D4 was modificated by mPEG-ButyrALD。After purification,the purity of mPEG-TNF-?D4 was above 85% and relative ability was 8.6?107.The in vivo systemic toxicity of mPEG-TNF-?D4,which is indicated by LD50,is lower than that of rhTNF-?.These results strongly supported for the further study and exploitation of TNF-antitumor drug.

OBJECTIVETo identify the effective stress management strategies among the Chinese.

METHODSThe sample was selected from Hangzhou, Guangzhou, Chongqing and Taiyuan by using a multi-stage sampling procedure, including 3679 subjects. The data were collected using the household interviewing survey method. The Chinese perceived stress scales (CPSS) measured stress. Stress management strategies included the cognitive and behavioral ones, the former were further divided into positive, neutral and negative ones and the latter were divided into three kinds, i.e. looking for support, liberating and displacing, and relaxing and detracting. The frequency of their usage and their perceived effectiveness were assessed. Multivariable analysis was used to examine the association between various stress management strategies and stress.

RESULTSThe prevalence of health risk stress (HRS) was 44.54% (95% CI: 42.90% - 46.12%). Among the cognitive strategies, all the positive strategies and one of neutral strategies ("Suiyuan") were associated with lower HRS, and the rest of them had no effects. Among the behavioral strategies, all were associated with lower HRS except that of looking for support.

CONCLUSIONThe effective stress management strategies identified in this study might be used to develop a stress management program.

Adult ; China ; epidemiology ; Educational Status ; Female ; Humans ; Male ; Middle Aged ; Occupations ; Psychology, Social ; Sampling Studies ; Social Behavior ; Social Environment ; Stress, Psychological ; epidemiology ; psychology ; therapy ; Surveys and Questionnaires ; Urban Population

OBJECTIVETo evaluate the efficacy of intraperitoneal transplantation of microencapsulated HepG2 cells in rats with hepatolenticular degeneration (HLD).

METHODSHLD was induced by copper-overloaded diet with forage containing 1 g/kg copper sulfate and water with 0.185% copper sulfate for 12 weeks in rats. One hundred and twenty three-month-old male Wistar rats were randomly intraperitoneal injected with normal saline (NS), microencapsulated HepG2 cells or non-microencapsulated HepG2 cells 9 weeks after copper-overloaded diet. Blood or liver samples were obtained at five time points: 3, 7, 14, 21 and 28 days after transplantation (n=8). The other 8 rats receiving normal diet were used as the control group. Serum levels of ALT, AST, albumin and Cu and liver Cu contents were measured.

RESULTSSerum ALT, AST and Cu levels and liver Cu contents in the NS-treated HLD, microencapsulated HepG2 cells and non-microencapsulated HepG2 cells transplantation groups increased significantly at all time points, in contrast, serum albumin levels decreased significantly in the NS-treated HLD and non-microencapsulated HepG2 cells transplantation groups compared with those in the control group at all time points (P<0.05), but serum albumin levels in the microencapsulated HepG2 cells transplantation restored to the level of the control group 28 days after transplantation. Serum ALT, AST and Cu levels and liver Cu contents in the microencapsulated HepG2 cells and non-microencapsulated HepG2 cells transplantation groups were significantly lower, in contrast, albumin levels were higher than those in the NS-treated HLD group on almost time points (P<0.05). Serum levels of ALT, AST and Cu and liver Cu contents in the microencapsulated HepG2 cells transplantation group decreased 7 or 14 days after transplantation, while serum albumin levels increased significantly 14 days after transplantation compared with those in the non-microencapsulated HepG2 cells transplantation group (P<0.05).

CONCLUSIONSIntraperitoneal transplantation of microencapsulated HepG2 cells can relieve hepatic damage, reduce serum and liver Cu levels, and improve copper metabolism, therefore it is promising for the treatment of HLD.

Animals ; Copper ; Hep G2 Cells ; Hepatolenticular Degeneration ; Liver ; metabolism ; Rats ; Rats, Wistar

Acyclovir ; therapeutic use ; Anti-Inflammatory Agents ; therapeutic use ; Antiviral Agents ; therapeutic use ; Brain Stem ; pathology ; virology ; Encephalitis ; complications ; diagnosis ; drug therapy ; virology ; Herpes Zoster ; complications ; diagnosis ; drug therapy ; virology ; Herpes Zoster Oticus ; diagnosis ; drug therapy ; etiology ; virology ; Humans ; Male ; Methylprednisolone ; therapeutic use ; Middle Aged

Human parathyroid hormone peptide1-34(hPTH1-34) was highly expressed in Escherichia coli by inserting the synthesized hPTH1-34 cDNA into pThioHis, the prokaryotic expression vector. The expressed hPTH1-34 was purified by chelating sepharose immobilized metal ion affinity, reverse and filter chromatographic steps. Its purity was verified above 95% by HPLC. The quality was identified by N-terminal sequencing and MALDI-TOF-MS analysis. In vitro analysis showed the adenylate cyclase of ROS 17/2.8 cells was activated by hPTH1-34.

Purpose: The purpose of this study was to retrospectively compare the clinical characteristics and imaging features on (CEUS) of combined hepatocellular cholangiocarcinoma (CHC) with those of hepatocellular carcinoma (HCC) and cholangiocarcinoma (CC). Methods: The clinical information and CEUS features of 45 patients with CHC from 2015 to 2019 and 1-to-1-matched control subjects with HCC and CC (45 each) were compared. Results: Simultaneous elevation of α-fetoprotein (AFP) and cancer antigen (CA) 19-9 was more common in CHC than in HCC and CC. In the arterial phase, hyperenhancement (homogeneous and heterogeneous) was more common in CHC (73.3%) and HCC (100%), while peripheral rimlike enhancement was more common in CC (55.6%). In the portal phase, marked washout was significantly more frequent in CHC and CC than in HCC (42.2% and 53.3% vs. 6.7%). In the delayed phase, marked washout was more common in CHC (82.2%) and CC (93.3%) than in HCC (40.0%). The washout time (WT) was much shorter in CHC and CC than in HCC (33.8±13.1 seconds and 30.1±11.6 seconds vs. 58.4±23.5 seconds). Using the combination of simultaneous elevation of AFP and CA 19-9 with marked washout in the delayed phase and a WT <38 seconds or arterial hyperenhancement to differentiate CHC from HCC or CC, the accuracy, sensitivity, and specificity were 74.4%, 93.3%, and 55.6% and 71.1%, 80.0%, and 62.2%, respectively. Conclusion Although some CEUS imaging features of CHC, HCC, and CC overlap, the combination of tumor markers and CEUS features can be helpful in differentiating CHC from HCC and CC.

OBJECTIVETo study the inhibitory effects of insulin on nuclear factor-kappa B (NF-kappaB) nuclear translocation of vascular endothelial cells induced by burn serum and its correlative mechanism.

METHODSHuman umbilical vein endothelial cells (HUVECs) were cultured in vitro and divided into 5 groups: blank control group (BC, ordinary culture without any stimulation), normal serum control group (NS, cultured with nutrient solution containing 20% healthy human serum), burn serum stimulation group (BS, cultured with nutrient solution containing 20% burn human serum), burn serum+insulin treatment group (BI, cultured with nutrient solution containing 20% burn human serum and 1x10(-7) mol/L insulin), inhibitor pretreatment group [IP, pretreated with 50 micromol/L protein kinase B (Akt) specific inhibitor LY-294002, then cultured with the same medium as used in BI group 30 minutes later] according to the random number table. Six hours later, the injury and apoptosis of HUVECs was respectively observed by the scanning electron microscope and determined by the flow cytometry. Meanwhile, the phosphorylation of inhibitor kappa B-alpha (p-IkappaB-alpha) and Akt (p-Akt) in cytoplasm, and the content of NF-kappaB-p65 in nucleus were determined with Western blot.

RESULTS(1) Compared with those in BC group, HUVECs in BS group shrank obviously with irregular nuclear structure, and intercellular links jagged or vanished. Slight change was observed in HUVECs structure in NS and BI groups, with the cell ductility and nuclear structure much better than those in BS group. (2) The apoptosis rates of HUVECs in BS group [(28.5+/-2.3)%], BI group [(22.3+/-1.8)%], and IP group [(29.7+/-2.4)%] were all obviously higher than that in BC group [(15.7+/-2.2)%, F=14.288, P<0.05 or P<0.01]. There was no significant statistical difference between NS group [(17.0+/-2.5)%] and BC group in apoptosis rate (F=14.288, P>0.05). The apoptosis rate of HUVECs in BI group was obviously lower than that in BS group (F=14.288, P<0.05). (3) Compared with those in BC group, the protein expressions of p-IkappaB-alpha in cytoplasm and NF-kappaB-p65 in nucleus were up-regulated, and the protein expression of p-Akt in cytoplasm was down-regulated in BS and IP groups. The expression levels of the three proteins in NS and BI groups were close to those in BC group.

CONCLUSIONSInsulin could inhibit the IkappaB phosphorylation, and then restrict NF-kappaB nuclear translocation and improve the vascular endothelial cells function accordingly through regulating phosphatidylinositol 3 kinase/Akt pathway.

Apoptosis ; Burns ; blood ; Cells, Cultured ; Endothelial Cells ; metabolism ; Endothelium, Vascular ; cytology ; metabolism ; Humans ; I-kappa B Proteins ; metabolism ; Insulin ; pharmacology ; NF-kappa B ; metabolism ; Phosphorylation ; Serum ; metabolism ; Umbilical Veins ; cytology

OBJECTIVETo establish of acute respiratory distress syndrome (ARDS) model in canine after inhalation of perfluoroisobutylene (PFIB), and to observe the progressing of lung injury, and to study the mechanisms of injury.

METHODSA device of inhalation of PFIB for canine was made. The concentration of PFIB was 0.30 - 0.32 mg/L. Serum IL-6 and IL-8 were dynamically measured. Clinical manifestations, pathology of organs in canine were observed.

RESULTS(1) During inhalation, the concentration of PFIB remained stable; (2) After inhalation, blood arterial oxygen partial pressure fell gradually, and eventually met the criteria for diagnosing ARDS; (3) The level of IL-8 in serum rises significantly after inhalation (P < 0.05), whereas that of IL-6 was not obviously altered (P > 0.05); (4) Within 6 hours after inhalation, no abnormality in canine was observed, but afterwards symptoms gradually appeared, and typical breath of ARDS, such as high frequency and lower level could be seen in later phase; (5) Pathological examination showed severe congestion, edema and atelectasis in most part of both lungs, and signs of anoxia in other organs.

CONCLUSIONS(1) The device designed is capable of ensuring control of inhalation of PFIB; (2) Exposure to PFIB for 30 mins, canines all met the criteria for diagnosing ARDS 22 hours after inhalation, therefore the modeling is successful; (3) PFIB specifically damages the lung by causing excessive inflammation.

Administration, Inhalation ; Animals ; Disease Models, Animal ; Dogs ; Female ; Fluorocarbons ; toxicity ; Interleukin-6 ; blood ; Interleukin-8 ; blood ; Lung ; drug effects ; pathology ; Male ; Random Allocation ; Respiratory Distress Syndrome, Adult ; blood ; chemically induced

OBJECTIVETo evaluate the efficacy and clinical significance of 64-multislice spiral computed tomography angiography(MSCTA) with image fusion for the anatomy of perigastric arteries.

METHODSA total of 53 patients underwent abdominal 64-MSCTA, among whom 26 patients with gastric cancer underwent gastrectomy. Using volume rendering techniques, computed tomography angiography(CTA) of perigastric arteries and the stomach were reconstructed respectively, and then the images were fused together. The branching pattern of the celiac trunk and the origins and courses along the stomach of the 10 perigastric arteries were assessed. The accuracy, sensitivity, and specificity of 64-MSCTA were determined based on intraoperative findings.

RESULTSCTA clearly showed the celiac trunk. The most common branching pattern of the celiac trunk was Michels type I( in 46 patients(86.8%). The anatomy of perigastric arteries and stomach could be clearly demonstrated from any angle according to image fusion. The left gastric artery and the right gastroepiploic artery were shown in 100%, the left gastroepiploic artery 94.3%(50/53), the right gastric artery 83.0%(44/53), short gastric artery 58.5%(31/53), posterior gastric artery 49.1%(26/53), the replaced left hepatic artery 15.1%(8/53). The accessory left hepatic artery, accessory left gastric artery and replaced right hepatic artery were all identified in 7.5%(4/53) patients. The accuracy of preoperative CTA in term of correctly identifying perigastric arteries ranged from 84.6% to 100%, the sensitivity 82.6% to 100%, and the specificity was 100% for all the perigastric arteries.

CONCLUSIONS64-MSCTA can clearly reveal individual perigastric arteries. The anatomy of the stomach and perigastric arteries can be shown in vivo by fused image, and can provide guidance for gastrectomy.

Adult ; Aged ; Angiography ; methods ; Arteries ; Female ; Humans ; Image Processing, Computer-Assisted ; Male ; Middle Aged ; Preoperative Care ; Sensitivity and Specificity ; Stomach ; blood supply ; Stomach Neoplasms ; diagnostic imaging ; surgery ; Tomography, Spiral Computed ; Young Adult