To compare the activity of RGD-TRAIL in different expression systems, RGD-TRAIL in both Escherichia coli (E.coli) and Pichia pastoris was constructed and expressed. In vitro activity of RGD-TRAIL from Pichia pastoris expression system was also analyzed. Genetic engineering techniques were used to construct recombinant plasmid pET30-rgd-trail and pHBM-rgd-trail. The recombinant protein RGD-TRAIL was purified with Ni ion affinity chromatography after induction. MTT assay, ELISA, scratch wound healing, transwell migration assay and Hoechst 33342 staining were performed to detect the effects of RGD-TRAIL on proliferation, binding activity, migration and apoptosis. The expression of apoptosis-associated proteins was detected by Western blotting. Recombinant protein RGD-TRAIL was successfully expressed in a form of inclusion body in E.coli, while expressed secretorily in Pichia pastoris. It possessed more potent cytotoxicity than RGD-TRAIL in E.coli by MTT assay. The RGD-TRAIL expressed by Pichia pastoris showed powerful binding affinity with cancer cells expressing α(v), DR4, DR5 and highly potent cytotoxicity through inducing apoptosis of cancer cells. Nuclear fragmentation was examined by Hoechst 33342 staining. Cleaved PARP and caspase-3 were also detected after incubation with RGD-TRAIL. Additionally, RGD-TRAIL inhibited migration significantly in A549 and HT1080 cells. The results demonstrate that Pichia pastoris expression system is more suitable for the recombinant protein RGD-TRAIL. Its binding affinity and antitumor activity might make RGD-TRAIL a promising candidate for cancer therapy.
Antineoplastic Agents ; pharmacology ; Apoptosis ; Blotting, Western ; Cell Line, Tumor ; Chromatography, Affinity ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; Humans ; Oligopeptides ; biosynthesis ; pharmacology ; Pichia ; metabolism ; Plasmids ; Recombinant Fusion Proteins ; biosynthesis ; pharmacology ; TNF-Related Apoptosis-Inducing Ligand ; biosynthesis ; pharmacology

0.05). Compared with the healthy neonates, the concentration of blood MDA and ROS was significantly higher, and that of blood HB and SOD was significantly lower in both groups(P

Vascular endothelial growth factor(VEGF) is one of the best characterized angiogenic regulators which has many effects in promoting endotheliocyte proliferation and differentiation,increasing the microvascular permeability,inducing the angiogenesis,triggering the growth,survival and migration of tumor cells by combining its specificity receptor (VEGFR).Dysregulation of VEGF expression and signaling pathways therefore plays a pivotal role in the pathogenesis and clinical features of hematologic malignancies,direct and indirect targeting of VEGF and its receptors therefore may provide a potent novel therapeutic approach to overcome bone marrow angiogenesis and multidrug resis-tance thereby improve patient outcome.Recent years,a novel VEGF blockade system using RNA interference attracts more and more people's attention.The small interfering RNA (siRNA) targeting VEGF/VEGFR can completely inhibits the expression of VEGF and induce the silence of corresponding genes.

The sensor of naringin(NG), a non-electroactive substance, was prepared based on the molecular imprinting technique.Using cyclic voltammetry technique (scan rate is 100 mV/s), the naringin imprinting sensitive film, poly-o-aminophenol was coated on the surface of a graphite electrode in the presence of naringin which was considered as the template, and characterized by SEM and X-ray reflective spectrophotometry (XRR).Using K_3Fe(CN)_6 as an electroactive marker, the electrochemical properties of the NG sensor were investigated by CV, electrochemical impedance spectroscopy (EIS), differential pulse voltammmetric and chronoamperometric.The results showed that the imprinted electrode was significantly different from the non imprinted electrode in morphologies and electrochemical properties, and a linear relationship between the peak current and the naringin concentration was found in the range of 6.0 × 10 ~(-5)-1.4 × 10 ~(-4) mol/L with a detec tion limit of 1.6 × 10 ~(-5) mol/L.Moreover, the imprinted electrode exhibited a good selectivity and rapid response to the naringin template molecules, as well as an excellent reproducibility(RSD = 1.8 %, n=5).

Apoptosis ; Inhibitor of Apoptosis Proteins ; Myocytes, Cardiac ; cytology

OBJECTIVETo provide evidence for Chinese medical treatment of children with EB virus infection by exploring its clinical efficacy from multiple angles.

METHODSTotally 81 children patients were randomly assigned to the treatment group (46 cases) and the control group (35 cases). Patients in the treatment group took Chinese medical decoction, while those in the control received intravenous dripping of Ganciclovir and oral administration of pidotimod. The treatment period for the two groups was 2 weeks. Patients were followed-up till the 12th week. Clinical symptoms such as fever, lymphadenopathy and hepatosplenomegaly, as well as lab indices such as abnormal lymphocyte percentage, EB virus antibody, virus DNA load, T cell subsets, immunoglobulin, and so on were observed before and after treatment, at week 4 and 12 of follow-ups.

RESULTS(1) The total effective rate at week 2 was 95.6% in the treatment group, higher than that of the control group (94.3%), but there was no statistical difference between the two groups. (2) The time for defervescence, duration of pharyngeal hyperemia, duration of swollen tonsils was shorter in the treatment group than in the control group (P<0.05). The subsidence of lymphadenopathy, hepatomegaly, and abnormal lymphocytes was better in the treatment group than in the control group (P < 0.05). (3) The positive cases of peripheral blood hetero-lymphocyte was significantly reduced after treatment, at week 4 and 12 of follow-ups both in the treatment group and the control group (P < 0.01). The expression of IgA and IgM decreased after treatment in the two groups when compared with before treatment in the same group (P < 0.05, P < 0.01). IgG in the treatment group also obviously decreased after treatment, at week 4 and 12 of follow-ups (P < 0.05, P < 0.01), while it decreased only after treatment in the control group (P < 0.05). Activities of AST and ALT in the treatment group and the AST activity in the control group were markedly improved when compared with those before treatment (P < 0.05). Compared with the control group, the abnormal lymphocyte positive case number obviously decreased in the treatment group after treatment, at week 4 and 12 of follow-ups (P < 0.05). (4) After treatment, at week 4 and 12 of follow-ups, CD3+ and CD8+ significantly decreased; CD4+, CD4/CD8, and B cells significantly increased in the two groups, when compared with before treatment (P < 0.05). NK cells significantly increased more in the treatment group after treatment, at week 4 and 12 of follow-ups, higher than before treatment as well as the control group (P < 0.05). (5) EB viral DNA and EB viral CA-IgM negative conversion case numbers significantly increased in the two groups after treatment, at week 4 and 12 of follow-ups (P < 0.05). Compared with the control group, EB viral DNA and EB viral CA-IgM negative conversion case numbers significantly increased in the treatment group after treatment and at week 4 of follow-ups (P < 0.05).

CONCLUSIONSTreatment of EB virus infection by Chinese medical treatment was effective. It could promote the recovery of EB viral infection, and reduce the risk of vicious disease after EB viral infection.

Adolescent ; Child ; Child, Preschool ; Drugs, Chinese Herbal ; therapeutic use ; Epstein-Barr Virus Infections ; drug therapy ; immunology ; Female ; Herpesvirus 4, Human ; Humans ; Infant ; Male ; Phytotherapy ; T-Lymphocyte Subsets ; immunology

Objective To investigate the duration for silicon gastric tube to indwell in stomach of aged patients.Methods One hundred elderly patients needing long-term nutrition support were equally randomized into two groups: control group and observation group.The former had the gastric tubes indwelled in the stomach for 4 weeks and the latter for 3 weeks.The two groups were compared about the used tubes in terms of the color of tube at the length of the first 10cm,the indentation by reflection for 10 seconds at the tube of first 10 cm,the hardness of the tube at the length of the first 10 cm and the springback angle of tube at the first 10cm after reflection.Results The tubes in the observation group were significantly better than those in the control group in terms of color,indentation,hardness and springback angle(all P<0?01).Conclusion The duration of indwelling silicon gastric tubes in stomach can be three weeks for old patients needing long-term nutrition support via indwelling silicon tubes in their stomach.

Successive pathogenic bacteria were examined from liver,spleen, kidney and contents of intestine of diseased stone flounder (Kareius bicoloratus L.) occurring bacterial septicaemia infection caused by Aeromonas salmonicida. Identification of extensive phenotypic information to 6 pure cultures and detection of the mol% G+C ratio of the DNA to representative strains,showed that the examined bacteria belonged to a new morphovar of Acinetobacter junii, and was designated as Acinetobacter junii morphovar I. In addition, serotype, antibiotic sensitivity and pathogenicity of isolates were studied, the results showed that the 6 strains have the same K-antigen and O-antigen, there are no obvious differences in sensitivity and resistance to used 37 antimicrobial agents between strains, and have strong pathogenicity to experimental stone flounder and Bastard halibut.

Objective To examine the expression of neuron-specific enolase (NSE) in patients with remission diffuse large B-cell lymphoma (DLBCL),and to explone its prognostic value.Methods Serum NSE values were mesured by electmchemilumineseence immunoassay in 20 health donors and 40 DLBCL patients reached remission after 6-8 cycles of chemotherapy after initial diagnosis.Results Positive expressions rate of NSE in remission DLBCL patients was 50 ％,and was 10 ％ in health donors (P ＜ 0.01).In remission DLBCL patients,there was significant difference in the 2-year PFS rate between in NSE-positive group and negative group (6.6 ％ vs 20.9％,P ＜ 0.001).Serum NSE was an independent poor prognostic factor in remission DLBCL patients.Conclusion Serum NSE expressions is an independent poor prognostic factor of remission DLBCL and has significant clinical value.

Objective To evaluate the impact of epidural anesthesia with levobupivacaine combined with general anesthesia on colon surgery. Methods Sixty patients undergoing elective radical procedure for colon carcinoma were randomLy divided into four groups: saline group (group S), 0.125% levobupivacaine group (group L1), 0.25% levobupivacaine group (group L2), and 0.5% levobupivacaine group (group L3). Group S received normal saline of 10 mL epidurally and then infusion of 5 mL·h-1 until the procedure was finished; groups L1, L2, and L3 received levobupivacaine instead. Anesthetic induction was performed after epidural puncture. Mean blood pressure and heart rate were recorded at 8 time points including 5 min after entering into the operation room, 1 min after intubation, skin incision, abdominal exploration, 1 h after skin incision, completion of operation, extubation, and leaving PACU; meanwhile blood glucose and cortisol were detected, anesthesia time, time to PACU stay, bleeding, transfusion volume, adverse reaction, and doses of propofol, remifentanil, ephedrine, and fentanyl were noted. Results Time to PACU stay was longer in S group than in other 3 groups. Doses of remifentanil and fentanyl were larger in L1 group than in L2 group and L3 group. Ephedrine dose in L3 group was larger than in other 3 groups. Blood sugar in L1 group was higher than L2 group and L3 group. Cortisol in S group was higher than in other 3 group. Cortisol in L1 group was higher than in L3 group. The number of patients with hypotension was greater in L3 group than other 3 groups. Conclusions Continue epidural infusion of 0.25%levobupivacaine can reduce stress response and opioid uses, shorten PACU stay, whereas it does not increase use of ephedrine.