We wished to study the intracellular transport of adenoviruses. We constructed a novel recombinant adenovirus in which the structural protein IX was labeled with a mini-singlet oxygen generator (miniSOG). The miniSOG gene was synthesized by overlapping extension polymerase chain reaction (PCR), cloned to the pcDNA3 vector, and expressed in 293 cells. Activation of miniSOG generated sufficient numbers of singlet oxygen molecules to catalyze polymerization of diaminobenzidine into an osmiophilic reaction product resolvable by transmission electron microscopy (TEM). To construct miniSOG-labelled recombinant adenoviruses, the miniSOG gene was subcloned downstream of the IX gene in a pShuttle plasmid. Adenoviral plasmid pAd5-IXSOG was generated by homologous recombination of the modified shuttle plasmid (pShuttle-IXSOG) with the backbone plasmid (pAdeasy-1) in the BJ5183 strain of Eschericia coli. Adenovirus HAdV-5-IXSOG was rescued by transfection of 293 cells with the linearized pAd5-IXSOG. After propagation, virions were purified using the CsC1 ultracentrifugation method. Finally, HAdV-5-IXSOG in 2.0 mL with a particle titer of 6 x 1011 vp/mL was obtained. Morphology of HAdV-5-IXSOG was verified by TEM. Fusion of IX with the miniSOG gene was confirmed by PCR. In conclusion, miniSOG-labeled recombinant adenoviruses were constructed, which could be valuable tools for virus tracking by TEM.
Adenoviruses, Human ; chemistry ; genetics ; metabolism ; Arabidopsis Proteins ; chemistry ; genetics ; metabolism ; Flavoproteins ; chemistry ; genetics ; metabolism ; Humans ; Phototropins ; chemistry ; genetics ; metabolism ; Singlet Oxygen ; chemistry ; Staining and Labeling ; Transfection

To investigate the morphogenetic process of human adenovirus type 41 (HAdV-41), 293TE cells were infected with purified wild-type HAdV-41, and ultrathin sections of infected cells were prepared and observed under a transmission electron microscope. Results showed that HAdV-41 entered host cells mainly through three ways: non-clathrin-coated pit, clathrin-coated pit, and direct penetration of plasma membrane. In addition, cell microvilli might help HAdV-41 enter cells. After entering into cells, HAdV-41 virus particles could be found in vacuoles or lysosomes or be in a free state in cytoplasm. Only free virus particles could be found near nuclear pores (NP), suggesting that the virus needed to escape from lysosomes for effective infection and viral nucleoprotein entered the nucleus through NP. Progeny viruses were as-sembled in the nucleus. Three types of inclusion bodies, which were termed as fibrillous inclusion body, condense inclusion body, and stripped condense inclusion body, were involved in HAdV-41 morphogenesis. In the late phase of viral replication, the membrane integrity of the infected cells was lost and viral particles were released extracellularly. This study reveals the partial process of HAdV-41 morphogenesis and provides more biological information on HAdV-41.
Adenovirus Infections, Human ; virology ; Adenoviruses, Human ; genetics ; growth & development ; physiology ; ultrastructure ; Cell Membrane ; virology ; Cell Nucleus ; virology ; Humans ; Virus Release ; Virus Replication

OBJECTIVETo study the surgical management of enophthalmos after severe malar maxillary complex fracture.

METHODSThe X-ray and CT examination were performed before operation to diagnose the orbital fracture and intraorbital tissue displacement. The fractured orbital rim was repositioned intraoperatively, followed by implantation of shaped titanium mesh to rebuild the orbital floor. The Medpor was inserted above the titanium mesh to correct the enophthalmos.

RESULTSFrom Sept. 2007 to Jan. 2009, 6 cases of enophthalmos after severe malar-maxillary complex fracture were treated. The enophthalmos was corrected or improved obviously in all the patients.

CONCLUSIONSThe enophthalmos after severe malar-maxillary complex fracture can be corrected or obviously improved. Shaped titanium mesh can be used to rebuild the orbital floor with the Medpor to reconstruct the intraorbital tissue volume.

Adolescent ; Adult ; Enophthalmos ; etiology ; surgery ; Female ; Follow-Up Studies ; Humans ; Male ; Maxillary Fractures ; complications ; Middle Aged ; Orbit ; surgery ; Polyethylenes ; Surgical Mesh ; Titanium ; Treatment Outcome ; Young Adult ; Zygomatic Fractures ; complications

OBJECTIVETo observe the therapeutic effect of Wenshen Huatan Recipe (WHR) on patients with early or middle stage chronic renal insufficiency (CRI) and investigate the clinical validity of applying sputum and blood stasis theory in treating CRI.

METHODSSixty CRI patients were selected and randomly assigned to the treated group (n = 32) and the control group (n = 28), all received conventional and symptomatic treatment, but those in the treated group were given WHR additionally. Two months were taken as one treatment course and the study lasted for 3 treatment courses. Total effective rate was assessed at the end of the 2nd and 6th month, changes of clinical symptom scores of TCM, and levels of blood urea nitrogen (BUN) and serum creatinine (SCr) in the two groups were observed as well.

RESULTSThe total effective rate of the 2nd and 6th month was 68.8% and 75.0% respectively in the treated group and 42.9% and 35.7% in the control group respectively, the difference between the two groups was significant (P < 0.05). Symptoms were improving along with the increasing medication time, and the improvement between the two groups was significant (P<0.01). Levels of SCr and BUN were significantly lowered in the treated group after 2-month and 6-month treatment (P < 0.05, P < 0.01).

CONCLUSIONWHR can significantly improve the clinical effect of treatment on CRI during early-middle stage, suggesting that CRI could be treated based on sputum and blood stasis theory with definite clinical efficacy.

Adult ; Aged ; Diagnosis, Differential ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Humans ; Male ; Medicine, Chinese Traditional ; Middle Aged ; Phytotherapy ; Renal Insufficiency, Chronic ; diagnosis ; drug therapy ; Treatment Outcome

Objective To evaluate the changes of peripheral blood interleukin(IL) 2, IL 10, IL 13 in the serum of asthmatic children and their effect on the pathogenesis of asthma.Method The serum levels of IL 2,IL 10,II 13 in 16 cases of asthmatic children and 10 cases of healthy controls were measured by ELISA methods.Results The serum levels of IL 2,IL 13 in asthmatic patients were significantly higher than those in healthy controls( P

OBJECTIVETo study the clinical effectiveness of Wenshen Huatan Recipe (WHR) in treating early-middle stage chronic renal insufficiency (CRI) of Pi-Shen yang deficiency syndrome (PSYDS).

METHODSNinety early-middle stage CRI patients of PSYDS were recruited and randomly assigned to the treated group (60 cases) and the control group (30 cases) in the ratio of 2:1. All patients received conventional and symptomatic treatment, but those in the treated group received WHR additionally. The observation was lasted for six months. The total efficacy, the Chinese medicine syndrome integral, and indices of the renal functions [including serum creatinine (SCr), blood urea nitrogen (BUN), creatinine clearance (CCr), serum cystatin C (Cys-C)], blood lipids [total cholesterol (TC), triglyceride (TG), high density lipoprotein cholesterol (HDL-C), and low density lipoprotein cholesterol (LDL-C)], as well as adverse reactions were observed before and after treatment.

RESULTSAfter six months of treatment, the total effective rate in treated group was 70.0% (42/60), which was obviously higher than that in the control group [43.3% (13/30), P < 0.05]. The Chinese medicine symptoms in the treated group were obviously improved, showing statistical difference between the two groups (P < 0.01). Better effects in lowering SCr, BUN, Cys-C, and LDL-C and increasing CCr were obtained in the treated group than in the control group (P < 0.05).

CONCLUSIONWHR could postpone the progression of CRI, improve the lipid metabolism, indicating certain therapeutic efficacy could be obtained in treating early-middle stage CRI from sputum theory.

Aged ; Creatinine ; blood ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Humans ; Lipids ; blood ; Male ; Middle Aged ; Phytotherapy ; Renal Insufficiency, Chronic ; blood ; drug therapy ; physiopathology ; Yang Deficiency ; blood ; drug therapy ; physiopathology

Objective To investigate the effects of hypoxia on the features and chemoresistance of cancer stem cells in Hep-2 cells and underlying mechanism. Methods The shRNA interference recombinant plasmid targeting HIF-1α was synthesized and transfected into Hep-2 cells. The HIF-1αknockdown Hep-2 cells were established after clonal selection and the expression of HIF-1α was measured.The cellular features including proliferation,clonal formation,cell cycle,apoptosis and CD133 phenotype were measured in Hep-2 cells cultured under hypoxic condition in vitro. CD133 + cells were sorted from Hep-2 cells with flow cytometry. Clonal formation test and cisplatin treatment were carried out,and theexpressions of related genes( Oct-4,suvivin and p53 ) in CD133 + cells were measured.Results HIF-1αknockdown Hep-2 cells was successfully established,as evidenced by the reduced mRNA and proteinexpressions of HIF-lα. The Hep-2 cells cultured under hypoxic microenvironment showed higher proliferation and clonal formation activity,cell cycle arrest in G0/G1,lower apoptosis,up-regulated CD133,however the effects of hypoxia reduced in HIF-1α knockdown Hep-2 cells.CD133 + cells were successfullysorted from Hep-2 cells,and the CD133+ cells showed increased clonal formation activity and cisplatintreatment resistance in hypoxia. Also the effects of hypoxia on CD133 + cells decreased with HIF-1αknockdown,showing down-regulated Oct-4 and survivin and up-regulated p53.Conclusions Hypoixa caninduce the features of cancer stem cells in Hep-2 cells and increase proliferation,differentiation and chemoresistant ability of CD133 + cells,which might be correlated with the changes in expressions of HIF-1 αand related genes regulated by HIF-1 α.

Objective: To study the protective effects of Panax quinquefolium 20s-protopanaxdiolsaponins extracted from leaves of P.quinquefolium（PQDS）on acute myocardial infarction（AMI）in dogs. Method：The parameters of myocardial infart size , the serum CK and LDH activity, myocardial metabolism,free radicals and coronary circulation etc were determined by using the model of ligation of LAD in the anaesthetized open-chest dogs. Result：In dogs treated with PQDS（in a dosage of 10 and 20 mg*kg-1 iv infusion），the myocardial infarct size, the activity of serum CK ,LDH and the contents of serum FFA and LPO were decreased，whereas the activity of serum SOD and GSH-Px increased markedly.At the same time, myocardial blood flow was increased and coronary vascular resistance decreasedsignificantly. Conclusion：PQDS has protective effect on myocardial ischemia by modifying metabolic dysfunction of FFA, inhibiting oxygen free radicalmediated peroxidation of membrane lipids ， enhancing endogenous antioxidase activity and increasing myocardial blood supply.

Meridians and Acupoints Science is a basic course and important component of acupuncture and Tuina science. This course contains considerable theoretical and basic knowledge to be memorized, which causes some difficulties for students to study and inevitably influences the learning effect. In view of that, the authors implemented reforms in interest teaching practice in teaching of Meridians and Acupoints Science. Reforms including the use of interesting memory method, string comparison of acupoints, interspersed discussion of cases, presentation of famous doctors' experience in acupoint application and development of the second class activities have achieved some success. That has aroused the students' enthusiasm for learning and improved the quality of teaching.

The systematic treatment based on gemcitabine plus cisplatin is recommended as the current standard chemotherapy for unresectable or metastatic biliary tract cancers.However,the exact benefits from the recognized regime are still dismal.We thus elicit this study in an attempt to analyze whether targeted therapy coupled with various chemotherapy could produce improvement of survival benefits.The clinical trials were searched electronically from databases till July 2016 published in English and Chinese.Nine hundred and sixty-four patients from 7 trials were identified in our analysis.The overall analysis achieved a significantly higher overall response rate (ORR) among the patients treated with targeted drugs plus chemotherapy than chemotherapy alone (OR=1.87;95％ CI:1.37-2.57;P=0.000),but failed in the overall progression-free survival (PFS) [mean difference (MD)=0.63;95％ CI:-0.45-1.72;P=0.26] and overall survival (OS) (MD=-0.67;95％ CI:-2.54-1.20;P=0.49).In the sub analysis,better ORR was obtained with the addition of EGFR (OR=1.75;95％ CI:1.20-2.56;P=0.004) and VEGFR (OR=2.5;95％ CI:1.28-4.87;P=0.007) targeted therapy.Furthermore,the sub analysis of EGFR target showed an significant improvement on PFS (MD=l.36;95％ CI:0.29-2.43;P=0.01).No significant differences were observed in the incidences ofneutropenia (OR=1.37;95％ CI:0.89-2.12),thrombocytopenia (OR=l.40;95％ CI:0.83-2.39),anemia (OR=l.21;95％ CI:0.62-2.38),peripheral neuropathy (OR=1.52;95％ CI:0.81-2.88),increased AST/ALT (OR=l.40;95％ CI:0.82-2.39) as well as fatigue (OR=1.65;95％ CI:0.96-2.84) in either of the treatment groups.In conclusion,better ORR associated with chemotherapy combined with targeted therapy (both targeting EGFR and VEGF) is found in the present mcta-analysis without the cost of increased unacceptable toxicities,but regretfully not for the OS.The sub-analysis of targeting EGFR instead of VEGF obtains a superior PFS.Otherwise,there is no statistically significant difference in the overall PFS between the combination regime and chemotherapy alone.Given the paucity of favorable data,we need further studies to characterize optimal targeted agents to confirm the potential value to biliary tract cancer.