To better understand the cleavage efficiency of muhiribozyme system on its RNA substrate in the presence and absence of divalent magnesium and monovalent sodium ions.we constructed pGEM-Coat'A,pGEM-Coat'A196Rz plasmids and pGEM-MDRl target plasmid.They were applied to transcribe RNAs with SP6/T7 transcription kit.Cleavage reactions were carried out in cell-free system and reaction products were analyzed by electrophoresis on 6% denaturing polyacrylamide gels in TBS buffer.The gels were dried and exposed to X-ray films for autoradiography.The Image J software was employed to analyze the dried gels.The results indicated that the cleavage efficiency of the muhiribozyme was dependent on the concentration of divalent Mg2+.The cleavage products increased with the concentrations of divalent Mg2+ and were Mg2+ concentration and time dependent.No cleavage product was obtained in the presence of lower than 200 mmol/L Na+ alone.On the contrary,monovalent Na+ inhibited the Mg2+ -induced cleavage reaction in Na+ and Mg2+ coexistance.The cleavage rate was significantly lower than that observed with divalent Mg2+ alone.These results suggested that divalent Mg2+ was required for muhiribozyme on substrate cleavage reaction in the physical condition,whereas monovalent Na+ was not.

Four crystalline forms of clopidogrel bisulfate were characterized by analytical techniques. Aiming to research the absorption characteristics of clopidogrel bisulfate polymorphs after taken orally by rat, and to estimate the influence of crystal form to pharmacodynamic action, four crystalline forms of clopidogrel bisulfate were administered intragastrically to rats, and high performance liquid chromatography (HPLC) was used to measure the contents of clopidogrel bisulfate and its metabolite in rat plasma. The metabolite of clopidogrel bisulfate was detected in rat plasma. There were significant deviations among four crystalline forms in the areas under curve of the metabolite of clopidogrel bisulfate. We concluded that the different crystal forms of clopidogrel bisulfate showed different pharmacokinetic characteristics, which might affect pharmacodynamic action.

Three pSIREN-siRNA plasmids were constructed using a pSIREN-RetroQ vector to silence the expression of muhidrug resistance-associated protein (MRP1) gene, and subsequently characterized by restriction endonuclease digestion and DNA sequencing. A truncated MRP1 and a full-length MRP1 were cloned into pEGFP-N2 and PeDNA3.1 respectively as pEGFP-MRP1T and pcDNA-MRP1. The plasmid pEGFP-MRP1T was co-transferred with each of the three pSIREN-siRNAs into HEK293 cells for MRP1T-GFP targeted silencing, and pSIREN-siRNA1 was used as the negative control, pSIREN-siRNA2 and pSIREN-siRNA3 appeared to be more effective to silence MRP1T-GFP compared to pSIREN-siRNA1 as shown by fluorescence microscopy. For the silencing of full-length MRP1 expression, HEK293 ceils were co-transferred with pcDNA-MRP1 and either of the three pSIREN-siRNAs, then subjected for Western blot analysis and MTT assays, pSIREN-siRNA2 and pSIREN-siRNA3 were able to inhibit the expression of 190 kD MRP1, but not pSIREN-siRNA1. The MDR of MRP1-transfected HEK293 ceils was abolished with pSIREN-siRNA2 or pSIREN-siRNA3 transfections. RNA secondary structure predictions demonstrated that the mRNA local free energy (△G) of the siRNA1 targeted sequence was lower, as the GC content and Tm value of siRNA1 were higher than those of siRNA2 and siRNA3. These data suggest that the local structure siRNAs and target mRNA may influence the silencing efficiency of MRP1 expression.

OBJECTIVETo explore the current application and features of Aconite prescriptions with incompatible herbs in grade A class three hospitals in east China and central China through a clinical study and comparative analysis.

METHODClinical prescriptions containing Aconite with incompatible herbs were collected. Association rules were utilized to analyze the compatible features of these herbs.

RESULTThis analysis found that the frequently used incompatible herba; pairs are Aconiti Lateralis Radix Praeparata-Pinelliae Rhizoma, with the support rate of 44.45%, occupying nearly half of the surveyed prescriptions; Pinelliae Rhizoma is the most frequently used herb in the two areas, with support rate up to 76.24%. Among the top 10 herbal pairs in the support rate, except for Aconiti Lateralis Radix Praeparata and Pinelliae Rhizoma, the top 10 herbs in Central China were mostly for warming the middle jiao and tonifying qi, such as Zingiberis Rhizoma, Atractylodis Macrocephalae Rhizoma and Codonopsis Radix; Whereas those in east China were mostly for activating and nourishing blood, such as Angelicae Sinensis Radix, Chuanxiong Rhizoma, and Salviae Miltiorrhizae Radix et Rhizoma. Among the top 10 herbal pairs in the support rate, except for Aconiti Lateralis Radix Praeparata-Pinelliae Rhizoma, the core herbal pairs applied in central China were mainly for resolving phlegm and warming the middle jiao, such as Pinelliae Rhizoma-Glycyrrhizae Radix et Rhizoma, Pinelliae Rhizoma-Zingiberis Rhizoma; Whereas those in east China were principally for activating blood and tonifying qi, like Atractylodis Macrocephalae Rhizoma and Pinelliae Rhizoma, Angelicae Sinensis Radix and Pinelliae Rhizoma. Among the core herbal groups in the two areas, the most frequently used herbal groups in the two areas are Aconiti Lateralis Radix Praeparata, Glycyrrhizae Radix et Rhizoma and Pinelliae Rhizoma with the support rate of 59.73%, accounting for the highest proportion among all of herbal groups.

CONCLUSIONThere are the combined clinical application of Aconite with incompatible herbs, mostly with Aconiti Lateralis Radix Praeparata-Pinelliae Rhizoma, but with differences in the combined application in east China and central China.

Aconitum ; chemistry ; Drug Incompatibility ; Drug Prescriptions ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Humans ; Pinellia ; chemistry ; Sophora ; chemistry

One 51 years old man was admitted to the rheumatology department with a history of prominent eyes, headache and blurred vision for half year. The main manifestations included retrobulbar inflammatory pseudotumor and retroperitoneal fibrosis. He was initially diagnosed as granulomatosis with polyangiitis. Prednisone and cyclophosphamide were administrated and effective. New mass of dura mater and osteosclerosis presented during follow up. Finally Erdheim Chester disease(ECD) was diagnosed by biopsy and pathological examination. Vemurafenib, a v-raf murine sarcoma viral oncogenes homolog B1 (BRAF) inhibitor, 480 mg was given twice a day. The patient′s condition is stable and still in follow-up. Although ECD is a rare histiocytosis, clinicians should pay attention to its manifestations and differential diagnoses.

Objective To investigate the protective effects of purified effective component group in extract from Xiaoshuan Tongluo(CGXT)formula on chronic brain ischemia in rats.Methods CGXT 75,150,and 300 mg/kg or vehicle were ig administered daily for four weeks to rats with bilateral common carotid arteries ligation(BCCAL).From the day 24 to 28 after BCCAL,Morris water maze was performed to assess the learning and memory impairment of rats.Four weeks after BCCAL,brain gray and white matter damage were assessed.Results In Morris test,the mean escape latency of rats in the CGXT(150 and 300 mg/kg)groups was significantly shorter than that in the vehicle group.CGXT also attenuated the neuronal damage in hippocampus and cortex and reduced the pathological damage in the optic tract and corpus callosum.Conclusion CGXT could improve learning and memory impairment resulted from BCCAL in rats.These results provide the experimental basis for the clinical use of CGXT in stroke treatment and may help in investigation of multimodal therapy strategies in ischemic cerebrovascular diseases including stroke.

Objective To develop and validate a simple,rapid,sensitive,and reproducible HPLC method for simultaneous determination of baicalein and its metabolite baicalin in dog plasma and for the subsequent pharmacokinetic study after iv administration to dogs.Methods An accurate and reproducible HPLC-UV method was developed and validated for simultaneous determination of baicalein and baicalin in dog plasma,using luteolin as internal standard.The analytes were separated by an Agilent Zorbax SB-C<,18> column(250 mm×4.6 mm,5μm)and the column temperature was maintained at 40 ℃.The mobile phase was a binary mixture of acetonitrile and water(27:73),containing 0.05% phosphoric acid in water,with a flow rate of 1.0 mL/min.The UV detector was set al 276 nm.Results Linear relationships were validated over the range of 0.05-25 μg/mL for baicalein and 0.05-20 μg/mL for baicalin.The intra-and inter-day precision values for all samples were within 8.0%,using relative standard deviation.This method was successfully applied to the pharmacokinetic studies in dogs after iv administration of baicalein.Baicalein was converted to baicalin quickly.Cmax values were 21.13 μg/mL at 0.05 h for baicalein and 1.57 μg/mL at 0.5 h for baicalin,areas under the plasma concentration-time curve were 4.97 h.μg/mL for baicalein and 0.63 h.μtg/mL for baicalin,and the elimination half-life is 0.50 h for baicalein and 0.75 h for baicalin,respectively.Conclusion The method is able and sufficient to be used in drug metabolism and pharmacoklnetic studies of baicalein.

Aim To observe the influences of dexmen-detomidine on the spontaneous contraction of duodenal smooth muscle of rabbits in vitro and explore the mech-anisms.Methods The rabbits ( male or female ) were stunned and the duodenums were isolated .The sam-ples of duodenal segments were connected with tension transducer , which were then put into oxygen saturation Krebs-Henseleit ( K-H) solution .The influences of dex-mendetomidine on amplitude ( AM ) and frequency ( FR ) of duodenal smooth muscle were recorded by BL-420 F biological signal processing system .The cu-mulative dosing method was used to observe the differ-ent concentrations of dexmedetomidine on duodenal smooth muscle spontaneous contraction .Glibenclamide ( Gli) was added to K-H solution before dexmendeto-midine.In the calcium-free K-H solution, calcium chloride and rynodine were added before dexmendeto-midine.The mechanisms of dexmendetomidine were studied .Results ① Dexmendetomidine reduced the amplitude of spontaneous contraction of duodenal smooth muscle in rabbits in a dose-dependent manner ( P<0.05 or P<0.01 ) , while the frequency was not obviously influenced ( P >0.05 ) .② Gli ( P <0.05 ) partly abolished the inhibitory effects of dexmendetomi-dine on duodenal smooth muscle .③ Dexmendetomi-dine inhibited the contraction of duodenum smooth muscle induced by calcium chloride ( P <0.05 ) and rynodine ( P<0.05 ) application into calcium-free K-H solution.Conclusion Dexmendetomidine inhibits the spontaneous contraction of duodenal smooth muscle of rabbits in vitro.The mechanisms may be related to ac-tivating ATP sensitive potassium channels , inhibition of the extracellular calcium influx via cell membrane and intracellular calcium release via sarcoplasmic reticulum in duodenal smooth muscle .

Objective To assess the role of health education in outcomes of diabetes mellitus among high-risk populations.Methods The community physicians who participated this investigation received standardized training,and 307 community residents at high risk of developing diabetes obtained three-month intense health education and nine-month follow-up study.Paired t-test,and Analysis of Variance were used for data analysis.Results After systematic health education,professional level of community physicians was improved.Cognitive level of health knowledge was also significantly improved (5.5 vs 12.6,t=-28.511,P＜0.05).In addition,health knowledge of variant age (F=4.036,P＜0.05),education level (F=15.27,P＜0.05) and occupation (F=9.80,P＜0.05) subgroups was significantly increased.In comparison with baselines,the scores of each age subgroups (F=0.204,P＞0.05) showed no significant differences,although scores of different education level (F=4.71,P＜0.05) and occupation (F=4.87,P＜0.05) subgroups were significantly different.The risk factors of diabetes were effectively controlled.Conclusions Health education should be the key to health management of diabetes,which plays important roles in improving cognitive level of health knowledge among populations at high risk of developing diabetes and reducing the incidence of this condition.

Objective To investigate the mechanism and effect of the spinal cord outlet of the skull base on Chiari Ⅰ malformation with syringomyelia.Methods The cervical spinal cord stem angle (Anbc),slope angle of cervical vertebra (Ansc) of Chiari Ⅰ malformation were measured.In foramen magnum (Llf) and anterior vertebral canal level (Laf),spinal canal(Ac),spinal cord (As) and inferior hernia area (Ah) were measured.Angle,area and ratio were compared in Chiari Ⅰ malformation with syringomyelia,Chiari Ⅰ malformation without syringomyelia and normal control group.Results Ansc,Anbc-Ansc had significant differences among control group and Chiari Ⅰ malformation patients (all P＜0.001).In Llf,Laf,As had significant differences among three groups (all P＜0.05),further comparison of the two showed there were significant differences between Chiari Ⅰ malformation with syringomyelia patients and control group,Chiari Ⅰ malformation without syringomyelia patients and control group in Llf(all P＜0.05).In Llf,Laf,Ac in Chiari Ⅰ malformation with syringomyelia was smaller than control group (P＜0.05).Ah in Llf,Lafand Lh in Llf had no statistical significant difference between Chiari Ⅰ malformation with and without syringomyelia patients (all P＞0.05).In Llf,Laf,As/Ac had statistical significant difference among Chiari Ⅰ malformation with and without syringomyelia patients,control group (all P＜0.001),further comparison of the two showed As/Ac in Llf had statistical significance difference between Chiari Ⅰ malformation with syringomyelia patients and control group (P＜0.05),As/Ac in La had statistical significance difference between Chiari Ⅰ malformation with syringomyelia patients and control group,between Chiari Ⅰ malformation without syringomyelia patients and control group (all P＜0.05),Conclusion The cervical spinal cord,Ansc reducing,narrow vertebral proportion increase are important factors to promote Chiari Ⅰ malformation syringomyelia.