Objective To clone, express and purify human PIF1 protein and analyze its ATPase activity. Methods The PIF1 cDNA was amplified by PCR from HeLa cell cDNA library and inserted to pET24b with histidine tag at its terminus to form pET24b-PIF1 plasmid. The recombinant pET24b-PIF1 plasmid was transformed to RosettaTM 2 (DE3) and the expression of PIF1 protein was monitored by SDS-PAGE analysis. By using fast protein liquid chromatograph (FPLC) system, the PIF1 protein was purified by affinity chromatograph and gel filtration. The ATPase activity of PIF1 was checked by thin layer chromatograph(TLC). Results The PIF1 protein was successfully cloned and expressed in E.coli. Conclusions The purification procedure of PIF1 protein was established using FPLC. The overexpressed and the purified PIF1 helicase has DNA and Mg2+ dependent ATPase activity.

Objective To investigate the role of NF-κB in aldosterone-1％NaCl-induced renal injury in uninephrectimized SD rats and the potential mechanisms.Methods Thirty-teo male SD rats were uninephrectomized and treated for 4 weeks.Rats were divided into four groups randomly:control group (n=8),1％NaCl group (1％NaCl in chow,n=8),aldosterone group (1％NaCl in chow,0.75 μg/h aldosterone delayed relase by osmotic mini-pump,SC,n=8),PDTC group (1％NaCl in chow,0.75 μg/h aldosterone,SC,100 mg/kg PDTC,IG,n=8).Systolic blood pressure (SBP),urinary protein,renal function and renal morphologic were observed.The expression of intercellular cell adhesion molecule 1 (ICAM-1) and connective tissue growth factor (CTGF) were measured respectively by Western blotting and real-time PCR.The activity and location of NF-κB in renal cortex were detected by electrophoretic mobility shift assay (EMSA) and immunohistochemisty.Results Rats of aldosterone group exhibited higher blood pressure and more serious renal injury characterized by proteinuria,glomerular sclerosis compared with rats of the 1％ NaCl group.Protein and mRNA levels of ICAM-1 and CTGF were significantly increased inaldosterone group rats than those in 1％NaCl group (all P＜0.05).Moreover,all these changes were associated with an increase in NF-κB activity.Treatment with PDTC which is a specific inhibitor of NF-κB notably alleviated SBP,proteinuria and renal injury in aldosterone-infused rats.Furthermore,PDTC markedly reduced the expression of ICAM-1 and CTGF (all P＜0.05).Conclusion PDTC can alleviate aldosterone-1％NaCl-induced renal injury in uninephrectimized SD rats by preventing the expression of ICAM-1 and CTGF.

Wip1 is a nuclear protein and a member of serine/threonine specific protein phosphatase type 2C(PP2C)family.It was initially identified as a gene whose expression was induced in response to ? or UV radiation in a p53-dependent manner and a negative feedback regulation of p38MAPK-p53 signaling.Then,Wip1 gene was confirmed a proto-oncogene and amplified or overexpressed in several human tumor types.This review will introduce the structures and functions of Wip1 and details on the signaling process of cancer progression.

Objective To assess the protective effect of remifentanil on cultured human hepatocytes against anoxia-reoxygenation injury. Methods Cultured hepatocytes were divided into 5 groups: group C receiving normoxia as control; groups AR, R, CH, R+CH receiving 15-hour xypoxia followed by 5-hour reoxygenation (group R receiving 5 ng/ml remifentanil, group CH 10 ?mol/L chelerythrine, group R+CH 5 ng/ml remifentanil and 10 ?mol/L chelerythrine before reoxygenation). The content of MDA in the hepatocyte mitochondria were measured. The rate of apoptotic cells was measured by flow cytometry. The expression of protein kinase C mRNA was measured by RT-PCR. Results Anoxia-reoxygenation caused dramatic increase in the content of MDA, the rate of apoptotic cells and the expression of protein kinase C mRNA. The three indexes mentioned above of groups R and CH were between that of groups C and AR (P

Objective To understand the situation of oxidative stress among diabetic nephropathy (DN) patients and observe the antioxidative effect of losartan at increasing dose in DN patients. Methods Thirty type 2 DN patients who neither smoked and nor took antioxidants were selected. The study began with an initial 4-6 weeks screening-treatment. Eligible patients then received losartan 50 mg/d daily for 8 weeks followed by losartan 100 mg/day daily for an additional 8 weeks. Blood glucose and blood pressure were closely monitored over the whole study period. All patients were followed up every other weeks, their 24-hour urine samples,fresh urine and venous blood sample were collected to measure urinary protein and creatinine excretion, urinary 8-OHdG, SOD, TAOC and MDA excretion , serum SOD, TAOC , MDA and other blood biochemistry parameters. Urinary 8-OHdG was determined by capillary electrophoresis and liquid phase chromatography. Results The total 24 hours urinary 8-OHdG excretion and the serum MDA concentration were higher than the normal values. The serum and urine SOD concentrations were lower than the normal values. There was an improvement in urinary 8-OHdG,serum and urine SOD, serum and urine MDA levels with losartan therapy. Compared with losartan 50 mg/d, the antioxidative effect of losartan 100 mg/d was more noticeable. Obvious decrease in 24-hour proteinuria on exposure to losartan was found, without severe adverse effect. Conclusions Oxidative stress damage is active in DN patients. Losartan has antioxidative effect on DN patients. Compared with losartan 50 mg/d, the antioxidative effect of losartan 100 mg/d is more marked, without increasing side effect. Losartan's antioxidative effect may be involved in its beneficial mechanisms on DN.

The aim of this study is to develop the Tripterygium glycosides nanoemulsion gels and investigate its pharmacodynamics. Oleic acid was used as oil phase, polyoxyethylene castor oil as surfaetant, and 1,2-propanediol as cosurfactant to screen the formula of Tripterygium glycoside nanoemulsion using the pseudo-temary phase diagrams. Then the nanoemulsion gels was prepared. The ICR mouse ears were sensitazated by 7% DNCB, and then were excited by 0.3% DNCB to stimulate the model of mouse chronic dermatitis and eczema. The concentrations of IFN-γ, IL-4 and IL-8 in mouse blood were determined by ELISA. The results showed that Tripterygium glycosides nanoemulsion gels could significantly inhibit the swelling of mouse ears(P < 0.01) and ameliorate the edama and erythema of model mouse ears skin. Also it could significantly decrease the expression of IFN-γ and IL-4 in model mouse blood. Tripterygium glycosides nanoemulsion gels had a good therapeutic effect on mouse model of dermatitis and eczema. It was expected to provide a new and long-acting exterernal preparation for the treatment of dermatitis and eczema.
Animals ; Chemistry, Pharmaceutical ; instrumentation ; methods ; Dermatitis ; drug therapy ; immunology ; Drug Carriers ; chemistry ; Drugs, Chinese Herbal ; chemistry ; pharmacokinetics ; Emulsions ; chemistry ; Female ; Glycosides ; chemistry ; pharmacokinetics ; Humans ; Interleukin-4 ; immunology ; Interleukin-8 ; immunology ; Mice ; Mice, Inbred ICR ; Nanoparticles ; chemistry ; Tripterygium ; chemistry

A rapid, sensitive and simple liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the simultaneous determination of clevidipine butyrate and its primary metabolite clevidipine acid in dog blood. After one-step protein precipitation with methanol, the chromatographic separation was carried out on an Ecosil C18 column (150 mm x 4.6 mm, 5 µm) with a gradient mobile phase consisting of methanol and 5 mmol · L(-1) ammonium formate. A chromatographic total run time of 13.0 min was achieved. The quantitation analysis was performed using multiple reaction monitoring (MRM) at the specific ion transitions of m/z 454.1 [M-H]- --> m/z 234.1 for clevidipine butyrate, m/z 354.0 [M-H]- --> m/z 208.0 for clevidipine acid and m/z 256.1 [M-H]- --> m/z 227.1 for elofesalamide (internal standard, IS) in the negative ion mode with electrospray ionization (ESI) source. The linear calibration curves for clevidipine butyrate and clevidipine acid were obtained in the concentration ranges of 0.5-100 ng · mL and 1-200 ng · mL(-1), separately. The lower limit of quantification of clevidipine butyrate and clevidipine acid were 0.5 ng · mL(-1) and 1 ng · mL(-1). The intra and inter-assay precisions were all below 12.9%, the accuracies were all in standard ranges. Stability testing indicated that clevidipine butyrate and clevidipine acid in dog blood with the addition of denaturant methanol was stable under various processing and/or handling conditions. The validated method has been successfully applied to a pharmacokinetic study of clevidipine butyrate injection to 8 healthy Beagle dogs following intravenous infusion at a flow rate of 5 mg · h(-1) for 0.5 h.

[Objective] The aim of this study was to establish a quantitative SYBR Green Ⅰ real-time PCR method for detection of wide-type p53-induced phosphatase 1 (Wip1 or PPM1D) gene expression level in non-small cell lung cancer (NSCLC),and to investigate the relationship between Wip1 mRNA expression level and the clinicopathological characters.[Method] Real-time PCR was employed to determine the expression level of Wip1 mRNA in 44 specimens of NSCLC tissues and their adjacent normal tissues.[Results] In the 44 specimens,the expression of Wip1 mRNA in both cancer tissues and adjacent normal lung tissues were positive.Wip1 gene was overexpressed in 17 specimens among 44 NSCLC specimens.The rate was 38.6%.The relative level of Wip1 mRNA in NSCLC tissues was significantly higher than the adjacent normal lung tissues (Ratio ＝ 2.1644 ± 1.394,P ＜ 0.01).The expression of Wip1 mRNA was also correlated with pathological staging (F ＝ 5.08,P ＝ 0.013).[Conclusion] The established SYBR Green Ⅰ quantitative real-time PCR method can successfully detect the expression level of Wip1 mRNA.The results suggested that Wip1 may be involved in the development of NSCLC.

Objective To study the feasibility of helical tomoterapy (HT) at prescription dose escalation for nasopharyngeal carcinoma (NPC).Methods Static-IMRT (sIMRT) and HT plans were designed for 10 nasopharyngeal carcinoma patients which were treated by sIMRT and HT treatment plan.Prescription dose was escalated for each plan until any organs at risk (OARs) reached the quantitative analysis of normal tissue effects within the clinical threshold.Dosimetric factors of target and OARs were analyzed for both sIMRT and HT plans.Results Compared with sIMRT plan, prescribed dose of HT plans increased 42.6％ (t =6.373, P ＜ 0.01), and the homogeneity index was also improved (t =-2.288, P＜0.05);the conformity index decreased (P ＞ 0.05).The limits of HT prescribed dose escalation were spinal cord (2 cases), optic nerve (5 cases) and brainstem (3 cases).The limits of sIMRT prescribed dose escalation were lens (1 case), spinal cord (1 case) and parotid (8 cases).Conclusions HT could improve prescription dose of nasopharyngeal carcinoma while keeping the OARs safe.Compared with sIMRT, HT technology might be used at high dose NPC radiotherapy.

Althoughtheevidenceoftheevidence-basedmedicinehasshowedthatrecombinant tissue-type plasminogen activator can effectively open the occluded vessels, because of its short therapeutic time w indow and the risk of bleeding, the thrombolytic rate is general y low er currently. Clinical studies have show ed that telestroke can effectively shorten the treatment time of the patients, increase the thrombolytic rate, reduce the risk of bleeding, and improve the outcomes of patients. Although the application of telestroke is restricted in many w ays, such as technology, policy, and funding, w ith the grow ing maturity of the related technologies, telestroke w il play an increasingly important role in the treatment of stroke.