Objective:To elucidate the effects of calcitonin gene-re la ted peptide(CGRP) on the gene expression of osteoprotegerin(OPG) and receptor ac tivator of nuclear factor-?B ligand(RANKL) in rabbit osteoblasts. Meth ods:Osteoblasts were cultured in media containing 10 -10～10 -7 mol/L of CGRP. After 24-hour incubation,semi-quanitative RT-PCR was perfor med to detect the expression of OPG and RANKL mRNA,and with ?-actin mRNA as th e internal control. Results:CGRP increased the mRNA expressio n of OPG with the maximal effect at the concentration of 10 -8～10 -7 mol/L. CGPR downgulated the mRNA expression of RANKL dose-dependantly.C onclusion:CGRP may regulate the activities of osteoclasts by regulating gene expression of OPG/RANKL.

Human gene therapy needs to express exogenous DNA at the targeting cells,producing a practical and efficient therapeutic dosage at an approp-riate time(quantitative pharmacology)with a safe man-ner.Recombinant adeno-associated virus(rAAV)Vec-tom possess a number of properties and recent progress in rAAV production made it rapidly become the reagent of choice for therapeutic gene tmasfer.Over 60 clinical trials of gene therapy based on rAAV have been carried out.The dose response reaction between rAAV vectors and gene expression activity or clinical outcome is one of major aspects of these trials.Most studies showed that vector genomes(vg)and gene expression had a concentration-dependent relationship during a certain scope.However,gene expr~sion Can be afffected by viral serotypes,tissue tropisms,cell targeting,drug regulation,injection route,age and sex,etc.Thus,these aspects should be carefully comidered by scienti-sts,pharmacologisis and physicians during animal ex-periments or clinical trails.KEY WORDS gene therapy;viral vector;dose-re-sponse;quantitative pharmacology;clinical thempy

AIM: To investigate the effect of Shenmai Injection on the expression of bFGF and PCNA of gastric cancer in mouse. METHODS: RT-PCR was used to measure the expression of bFGF and PCNA in gastric cancer cell cultured with Shenmai Injection in three concentrations.immunohistochemistry used for protein synthesis in mouse gastric cancer model. RESULTS: 140 ?L/mL of Shenmai Injection concentration inhibited 63% of bFGF and PCNA gene expression. CONCLUSION: Shenmai Injection can inhibit the express of bFGF and PCNA.

Objective:To construct K-ras-targeted siRNAs (K-ras siRNA) and to investigate the inhibitory effects of K-ras siRNAs on the growth and migration of lung cancer A549 cells (containing mutant K-ras gene) and NCI-H446 cells (containing wild-type K-ras gene). Methods: Four K-ras siRNAs (K-ras siRNA1～K-ras siRNA3 targeting wild-type K-ras and K-ras siRNA4 targeting mutant K-ras) were designed and artificially synthesized; they were used to transfect A549 cells and NCI-H446 cells. The expressions of Ras mRNA and protein were examined by RT-PCR and Western blot-ting. The inhibitory effects of K-ras siRNAs on the proliferations of A549 and NCI-H446 cells were determined by MTT assay. The effects of K-ras siRNAs on the cell migration and apoptosis were observed by Transwell assay and Hoechst 33258 staining, respectively. Results: Mutant K-ras-targeted siRNA (K-ras siRNA4) specifically inhibited the K-ras ex-pression but had no influence on H-ras and N-ras expression in A549 cells. K-ras siRNA4 inhibited the proliferation of A549 cells but did not inhibit that of NCI-H446 cells, which contained wild type K-ras gene. K-ras siRNA4 also induced apoptosis and inhibited migration of A549 cells. Conclusion: Mutant K-ras-targeted siRNA4 can inhibit the proliferation, migration and induce apoptosis of A549 cells. It may be a potential and personalized drug for the treatment against lung cancer containing mutant K-ras gene.

Objective To summarize the clinical features of the allergic bronchopulmonary aspergillosis (ABPA) in children,in order to improve the understanding for ABPA and make early diagnosis and treatment of the disease.Methods A retrospective study was performed on ABPA patients diagnosed in Department of Respiratory Medicine,Beijing Children's Hospital Affiliated to Capital Medical University from March 2010 to December 2013.The clinical features,laboratory results,image characteristics and the prognosis information were reviewed.Results Eight ABPA cases met the diagnostic criterion.All patients were school-age children (7 years and 2 months to 13 years and 8 months old).Cough (8 cases),productive sputum (8 cases),wheezing (5 cases),fever (4 cases) and hemoptysis (3 cases) were the main clinical features.Six of the 8 patients showed eosinophilia.IgE level was elevated in 7 patients (1.000-5.000 IU/L).All 8 patients were allergic to aspergillus fumigatus,while only 2 cases were positive in sputum culture for aspergillus fumigatus.CT scans showed pulmonary opacities in all 8 cases,while 7 patients had typical central bronchiectasis.Seven patients were treated with the regimen,which included glucocorticosteroid,antifungal agents (voriconazole or itraconazole) and regular bronchoscope.The symptoms of all treated patients relieved,the total serum IgE level and eosinophil cell count decreased spontaneously after the therapy.Conclusions ABPA is rare in children and the clinical features are non-specific.If the patient has elevated total IgE level in serum and eosinophilia,especially in patients with underling diseases,ABPA should be suspected.The positive result of specific antibodies to aspergillus fumigatus and central bronchiectasis on the radiology may give the suggestive diagnosis.ABPA patients generally have good response to the therapy of glucocorticosteroid and antifungal agents.

Objective To analyze and evaluate the precision ,accuracy and linearity of the chemiluminescence method for detec‐ting plasma BNP .Methods According to the experimental schemes of CLSI EP15‐A2 ,EP6‐A files and other relevant documents , the precision ,accuracy and linearity of the Siemens Centaur XP chemiluminescence instrument for detecting BNP ,and the detection results were compared with the performance declared by manufacturer or the quality target formulated by laboratory .Results The imprecision of BNP detected by the chemiluminescence method was less than 1/3 TEa regulated by the Clinical Laboratory Center of Ministry of Health (allowable total error);the variation coefficient index (CVR) of internal quality control data was less than ± 2;the relative bias of the results of external quality control blind samples with the target values were less than Tea regulated by the Ministry of Health ;the linear evaluation results showed that BNP was once linearity in the range of 5 .3‐4696 .7 pg/mL .Conclusion The precision ,accuracy and linearity of the Siemens Centaur XP chemiluminescence instrument for detecting BNP can accord withthe quality objectives requirements and meet the clinical needs .

Objective To summarize the experiences in the diagnosis and treatment of the hepatic hereditary hemorrhagic telangiectasia (HHHT).Methods The clinical data of 15 HHHT patients who were admitted to the Qilu Hospital,People's Hospital of Mengyin,People's Hospital of Liaocheng,Henan Provincial People's Hospital,the Second Hospital of Hebei Medical University,First Affiliated Hospital of Zhejiang University were retrospectively analyzed.The clinical manifestation,features of imaging and laboratory examination were summarized,and the diagnosis,treatment and prognosis of the disease were investigated.Results HHHT patients had nonspecific symptoms in the early stage,and some patients presented with right upper quadrant discomfort,shortness of breath,anemia and liver bruit.The condition of HHHT patients could be worsened by liver cirrhosis or portal hypertension rapidly.The results of color doppler ultrasound and computed tomography showed intrahepatic telangiectasia,arteriovenous fistula and hepatic artery aneurysm in the 15 patients.Digital subtraction angiography was not clear enough for 2 HHHT patients with more than 1 enlarged hepatic arteries,but computed tomographic angiography was feasible.According to the degree and stages of the HHHT,all the 15 patients were divided into asymptomatic HHHT,simple HHHT and complex HHHT.Among the 6 patients who underwent surgical treatment,5 received ligation or banding of the enlarged hepatic arteries with subsequent disappearance of symptoms.Three patients received interventional treatment,and the treatment for 1 patient with complex HHHT was failed,and the patient died 30 months after medical treatment.Six patients were treated by conservative treatment,2 patients of them had no symptoms at the beginning,then they suffered from hepatic dysfunction and ascites at 21 and 35 months,respectively,and 1 of them died 6 months later.Four patients received medical treatment,and the results of color doppler ultrasound and computed tomography showed the pathological changes were aggravated gradually.Conclusions Telangiectasia,intrahepatic arteriovenous fistula and hepatic artery aneurysm are the main imaging characteristics of HHHT,and imaging diagnosis has significant value in the diagnosis of HHHT.HHHT is a progressive disease,early,active and individualized treatment is beneficial to the patients.The outcome of ligation or banding of the hepatic arteries is satisfactory.

Uridine diphosphate (UDP)-GalNAc : polypeptide N-acetylgalactosaminyltransfemse (ppGalNAcT) catalyzes the initial step in mucin type O-glycosylation in the Golgi apparatus. Here generation and characterization of a polyclonal antibody to human ppGalNAcT2 were described. The subcellular location of ppGalNAeT2 in SGC7901 cell line was investigated using Western blot analysis of fractionated cell extracts and confocal microscopy with this antibody and two Golgi markers: Golgi SNARE (soluble N-ethylmalemide-sensifive factor attachment protein receptor) of 28 ku (GS28) and trans-Golgi network (TGN) 38, markers for the c/s- and trans-Golgi apparatus, respectively. Morphometric analyses indicated that ～60% of the ppGalNAcT2 signal colocalized with the GS28, while～36% of the c/s-Golgi marker colocalized with the ppGalNAeT2. Approximately 34% of the ppGalNAcT2 signal colocalized with the TGN38, whereas 38% of the trans-Golgi marker colocalized with the ppGalNAcT2. The results provide unequivocal evidence for the location ofppGalNAcT2 within the Golgi apparatus, and further highlight the importance of this organelle in the initiation of O-linked glycosylation.

High mobility group A2 protein (HMGA2), an architectural factor, is highly expressed in various cancer types including lung cancers. It is a candidate target for cancer therapy. RNAi is an effective gene silencing method with low cost and less time-consuming. It is possible to exploit this technology in therapy. Here, 5 siRNAs targeting Hmga2 gene (HMGA2 siRNA1-5) were designed and synthesized. MTT assay, colony formation assay, transwell assay and flow cytometry were used to evaluate the effects of these siRNAs on lung cancer cell lines (NCI-H446 and A549). Results from cell proliferation, clone formation, migration and apoptosis showed that HMGA2 siRNA1, 3, 5 could affect these aspects for both lung cancer cell lines. Among the five siRNAs, HMGA2 siRNA5 showed the greatest inhibition effects. The inhibition effects of HMGA2 siRNA5 are sequence specific and are not due to the induction of interferon response. Taken together, siRNAs targeting Hmga2 gene are potential candidates for lung cancer gene therapy.

Random amplified microsatellite polymorphism (RAMP) markers were used to access the genetic diversity among 112 samples of nine populations of Dendrobium officinale Kimura et Migo. Using 16 informative primers, 123 bands were amplified and 86 (69.92%) were polymorphic. The polymorphic bands from three to eight could be detected for each RAMP primer, with a mean of 5, indicating abundant genetic diversity among populations. Genetic similarity coefficients ranged from 0.250 to 0.813. UPGMA dendrogram illustrated 9 populations clustered into 3 groups, and the cluster pattern showed correlation with the locations of the D. officinale populations. These results were supported by the previous conclusions that were achieved by other molecular markers, and RAMP is proved to be effective for evaluating the genetic diversity of wild populations of Dendrobium officinale.