Objective To explore the role of ripply1 in zebrafish dorsal-ventral development .Methods Using ze-brafish whole-mount in situ hybridization to examine the ripply1 expression pattern in early embryo development .To analyse the expression pattern changes of dorsal-ventral marker genes at shield stage and the morphological changes at 24 hpf (hours post-fertilization) after overexpression of ripply1 by injecting synthetic mRNA at 1-cell stage.Using Tol2 transposon technology to obtain a ripply1 promoter driven GFP transgenic fish and to identify promoter region that recapitulates endoge -nous ripply1 expression pattern .Results The in situ hybridization results revealed that ripply1 specifically expresses in the future dorsal region at shield stage .Overexpression of ripply1 caused an enhanced expression of dorsal marker genes and a reduction of ventral marker genes .Embryos overexpressing ripply1 also showed severely dorsalized phenotype , with enlarged head, reduced ventral yolk extension , and shortened posterior trunk and tail regions , and the formation of a secondary trunk axis.Transgenic fish revealed the maternal expression of ripply1 and suggested that a 1.2 kb promoter-driven GFP is able to recapitulate the endogenous gene expression pattern .Conclusion ripply1 may participate in the early development of dor-sal-ventral axis in zebrafish embryo .

Objective To develop a method for the determination of DBP and DEHP in cosmetics by GC-MS. Methods The samples was extracted by methanol or acetic ether, DBP and DEHP were separated by GC and determined by MS. Results The linear range, regression equation of calibration curve and correlation coefficient were 0.5-100.0 ?g/ml, y=2?106 x-2?106, 0.999 6 for DBP respectively and 5.5-110.0 ?g/ml, y=988 223 x-7?106, 0.995 9 for DEHP respectively. Based on threefold ratio of signal and noise, as the sample was 1.0 and 0.2 g respectively, the volume was 10.0 ml, the detection limits were 1.0 and 5.0 mg/kg respectively for both DBP and DEHP. The recovery rates were 90.8%-119.0% for DBP and 90.4%-115.3% for DEHP. Relative standard deviations were 4.8%-9.8% for DBP and 6.0%-8.6% for DEHP. Conclusion This method is sensitive, accurate and high reproducible, and was applicable to the determination of DBP and DEHP in the cosmetics.

Objective To construct a human endostatin adenovirus vector and investigate its inhibitory effect on pancreatic carcinoma in nude mice.Methods Animal model of pancreatic carcinoma bearing nude mice was established by subcutaneous injection of SW1990 cells.All mice were randomized into Ad-hEnd group,Ad-LacZ group and control group with 8 mice in each group.The endostatin gene recombinant adenovirus were intratumorally injected every two days for 4 times.The rate of tumor growth was observed.lmmunohistochemical staining was employed to investigate the expression of vascular endothelial growth factor (VEGF) and micro-vessel density (MVD).TUNEL in situ was used to examine tumor cell apoptosis.Results The tumor formation rate was 100%.4 weeks later,the volumes of the tumors were (921.9±279.7 )mm3,(2804.4±553.5 )mm3 and ( 3040.6±487.6 ) mm3 in Ad-hEnd group,Ad-LacZ group and control group,respectively;the weights of the tumors were (1.19±0.18 ) g,( 2.38±0.42 ) g and ( 2.41±0.47 ) g,respectively;the VEGF positive rates were (36.3±7.1 )%,(81.2±6.6)% and (79.4±6.2)%,respectively;the levels of MVD were 12±4,27±5 and 25±6,respectively;the apoptotic rates were (31.2 ±5.4) %,( 9.4±4.9 ) % and ( 8.5±3.7 ) %,respectively.Compared with Ad-LacZ group and control group,the parameters in Ad-hEnd group were statistically different (P ＜0.01 ).The difference betweon Ad-LacZ group and control group was not statistically different.Conclusions Human endostatin gene mediated by recombinant adenovirus could inhibit tumor growth,angiogenesis and promote cell apoptosis of pancreatic carcinoma and could be used as geue therapy for pancreatic carcinoma.

Objective To investigate the role of interleukin-17 (IL-17) in spinal dorsal horns in neuropathic pain (NP) in rats and its effect on activation of astrocytes.Methods In vivo experiment Sixty-four male SPF Sprague-Dawley rats,aged 6-8 weeks,weighing 180-200 g,were randomly divided into 3 groups using a random number table:control group (group C,n =16),sham operation group (group S,n =24) and group NP (n =24).The animals were anesthetized with intraperitoneal pentobarbital sodium,the L5,6 spinal nerves of the left side of the rat were gently separated and exposed,tightly ligated with 5-0 silk suture and transected.In group S,the L5,6 spinal nerves of the left side of the rat were only exposed.In group C,no operation was performed.Mechanical pain threshold was measured at day 1 before operation and days 1,3,5,7,10 and 14 after operation.The expression of IL-17,IL-6,IL-1β and tumor necrosis factor-alpha (TNF-α) mRNA in the spinal dorsal horn was determined using quantitative real-time PCR at day 7 and day 14 after operation.At day 7 after operation,the activation of astrocytes in the spinal dorsal horn was detected.In vitro experiment Primarily cultured astrocytes of neonatal rats were randomly divided into 4 groups using a random number table:control group (group C,n=22),10 ng/ml IL-17 group (I10 group,n=18),50 ng/ml IL-17 group (I50 group,n-18) and 100 ng/ml IL-17 group (I100 group,n=22).In I10,I50 and I100 groups,the astrocytes were incubated with the culture medium containing 10,50 and 100 ng/ml IL-17,respectively.The proliferation of astrocytes was detected by MTT at 24,48 and 72 h of incutation or culture.The expression of IL-6,IL-1β and TNF-α mRNA was determined using quantitative real-time PCR.Results In vivo experiment Compared with group C,the mechanical pain threshold was significantly decreased at 3-14 days after operation,the expression of IL-17,IL-6 and IL-1β mRNA in the spinal dorsal horn was up-regualted at 7 days after operation,and the activation of astrocytes was increased in group NP,and no significant change was detected in the mechanical pain threshold at each time point after operation in group S.In vitro experiment Compared with group C,the proliferation of astrocytes was significantly increased at 48 h of incubation in I10 and I50 groups,the proliferation of astrocytes was significantly increased at 48 and 72 h of incubation,and the expression of IL-6 and IL-1β mRNA was up-regulated in I100 group,and no significant change was found in the proliferation of astrocytes in group S.Conclusion Up-regulated expression of IL-17 in spinal dorsal horns may be involved in the maintenance of NP,and the mechanism is related to promoted activation of astrocytes and induced inflammatory responses in rats.

OBJECTIVE：To construc t the evaluation system of the research quality of medical insurance budget impact analysis （BIA），and to provide feasible evaluation tool for related departments as medical insurance department. METHODS ：Based on BIA guidance documents and relevant empirical literatures of ISPOR ，Canada，Poland，the United States and other countries ， combined with expert interview ，the relevant elements of medical insurance negotiation BIA material were confirmed （including key elements and adjuctive elements ）. The scale and system was established to calculate total score of BIA research quality evaluation. RESULTS ：Key elements included three data blocks as target population ，market situation and treatment cost ，involving 14 key elements such as total population ，new drug scenario market share ，treatment cost ，etc.. According to the degree of compliance，0-3 points were assigned and the lowest score after normalization was taken as the basic score of BIA research quality. The adjunctive elements included five data blocks as title & abstract ，research background ，analysis framework ，result presentation and other ，including 23 adjunctive elements such as title ，abstract，research angle ，research time limit ，etc.. According to whether there is quality grade difference ，the elements were divided into type A and type B ；the grade score （0-4 points）and 0/1 score（1 point for yes and 0 point for no ）were used respectively ，and the additional score of BIA research quality was obtained after calculation and addition. According to the addition of different weights （0.67 and 0.33）of basic score and additional score ，the total score system of BIA research quality evaluation could be calculated. CONCLUSIONS ：This study successfully constructed a new BIA quality evaluation system ，which can be used for the quality evaluation of BIA research submitted by medical insurance drug negotiation.