Objective To investigate the effects of preconditioning with emulsified isoflurane(eISO)on inflammatory response to myocardial ischemia-reperfusion(I/R)injury in rats.Methods Forty SD rats of both sexes weighing 250-280 g were randomly allocated into 4 groups(n =10 each):groupⅠ sham operation(S);group Ⅱ myocardial I/R + normal saline(NS); group Ⅲ I/R + eISO and group Ⅳ I/R + 30％ intralipid(Ⅱ.)(vehicle for eISO).Myocardial ischemia was induced by occlusion of anterior descending branch of left coronary artery for 30 min followed by 180 min reperfusion.NS,30％ intralipid and elSO 2 ml/kg were infused iv over 30 min at 30 min before myocardial ischemia in groups Ⅱ,Ⅲ and Ⅳ respectively.The animals were killed at the end of 180 min repeffusion.Their hearts were removed for determination of infarct size and myocardial NF-κB p65 and ICAM-1 expression(by immuno-histochemistry)and plasma concentration of TNF-t(by radioimmunoassay).Results Myocardial I/R induced myocardial infarct and significantly increased plasina TNF-a concentration and myocardial ICAM-1 and NF-κB p65 expression in gro up Ⅱ,Ⅲ and Ⅳ as compared with'sham operation group (Ⅰ).Plasma TNF-a concentration and myocardial ICAM-1 and NF-kB p65 expression were significantly lower in group Ⅲ(eISO)than in group Ⅱ and Ⅳ.Conclusion Down-regulation of myocardial NF-kB and ICAM-1 expression and inhihition of inflammatoy response are involved in the mechanism by which preconditioning with iv elso protects against myocardial I/R injury.

BACKGROUND: The effect of Chinese traditional medicine 3'-meisoindigo as well as indimbin derivatives on normal immunocytes is less reported while it is used for antitumor.OBJECTIVE: To investigate the effect of 3'-meisoindigo on the proliferation of the splenocytes and thymocytes of C56BL/6 mouse.DESIGN, TIME AND SETTING: The randomized cytopathology observation was performed between August 2007 and January 2008 at Zhujiang Hospital of Southern Medical University, Guangzhou, Guangdong Province, China.MATERIALS: C57BL/6 mice, clean, male, 6-8 weeks old, weighing (20±2) g.METHODS: The thymus gland and the spleen of C57BL/6 mice were ground to get the single-cell suspension and cells were treated by 5, 10, 15, 20 and 25 μ mol/L 3'-meisoindigo,respectively. Cells without any drug treatment were used as blank control and cells treated by concanavalin A were used as positive control.MAIN OUTCOME MEASURES: Proliferation of splenocytes and thymocytes detected using MTT method; IL-12 activity in culture supematant detected using ELISA method; the cell cycle, apoptosis rate, cell death rate and intracellular reactive oxygen species level detected using flow cytometry; the mRNA level of bcl-2 and cdk2 detected using reverse transcription-polymerase chain reaction; the expression rate of Bcl-2, CDK2 and Bax detected using fluorescence microscope.RESULTS: After treating for 24 hours, 15, 20 and 25 μ mol/L 3'-meisoindigo can significantly inhibit the proliferation of thymocytes and splenocytes (P < 0.05) and the inhibition was dose-dependent and time-dependent. Cells resumed proliferation after removing the 3'-meisoindigo, although they had been treated by 25 μ mol/L 3'-meisoindigo for 72 hours. The secretion of IL-12 was markedly reduced in all 3'-meisoindigo groups versus control groups at each time point (P < 0.05). 15 μ mol/L 3'-meisoindigo could decrease the mRNA expression level of apoptosis-related protein bcl-2 and cyclin cdk2 gene, decrease the expression level of BCL-2 protein and CDK protein, increase Bax expression level, decrease Bcl-2/Bax ratio markedly and started apoptosis.15 μ mol/L 3'-meisoindigo arrested cells in the G2/M stage of the thymocytes and splenocytes, and intracellular reactive oxygen species level elevated dose-dependently and time-dependently.CONCLUSION: In certain concentration range, 3'-meisoindigo can reversibly inhibit the proliferation of thymocytes and splenocytes of C57BL/6 mouse and can inhibit IL-12 secretion in parallel, and start apoptosis.

AIM: To study the chemical constituents of Illicium micranthum. METHODS: Various column chromatographic techniques were applied to isolation and purification and the structures were elucidated by NMR and MS. RESULTS: Seven compounds were isolated from the part of ethyl acetate and identified as Neomajucin(1),2-oxo-3,4-dehyhroxyneomajucin(2),Transcinnamic acid(3),3,3′,4′,5,7-pentahydroxyflavan(4),Shikimic acid(5),Kaempferol(6),Kaempferol-3-O-?-L-rhamnopyranosyl-(1→6)-?-D-glucopyranoside(7). CONCLUSION: The above compounds are obtained from this plant for the first time.

To improve the expression of heterologous genes using baculovirus expression system, we constructed a novel shuttle vector based on the Bm-Bacmid. In the Bm-Bacmid, partial sequences of Chitinase and Cystein Protease were replaced with a tandem cassette of Cm and egfp through homologous recombination. Bombyx mori bidensovirus (BmBDV) ns1 under the control of polyhedrin promoter was inserted into the modified Bm-bacmid by transposition. For comparison, BmBDV ns1 under the control of polyhedrin promoter was also cloned in the wild type Bm-bacmid. The resulting Bm-bacmids were transfected into the cultured BmN cells to prepare recombinant virus to infect silkworms for expression of BmBDV ns1. Total proteins of hemocyte from infected silkworms were subjected to Western blotting and ELISA analysis. The yield of BmBDV NS1 1 with the modified vector was three times as much as that with the unmodified vector. The method to improve the yield of BmBDV NS1 in silkworms will facilitate the function and three-dimensional structure study of BmBDV NS1.
Animals ; Baculoviridae ; Bombyx ; virology ; Cell Line ; Chitinases ; Cysteine Proteases ; Genetic Vectors ; Insect Viruses ; Promoter Regions, Genetic ; Transfection

Baculovirus gene expression is the most popular method to make target protein in cultured insect cells. To fast determine the generation of recombinant virus in cultured cells, donor plasmid of pFastBacI was modified by introducing egfp cassette. In the modified vector, egfp cassette was under the control of ie1 promoter, and target gene cassette was under the control of polyhedron promoter. To evaluate the convenience of the genetically modified donor plasmid used in eukaryotic expression, ns1 gene from Bombyx mori bidensovirus was ligated into the donor plasmid to generate recombinant plasmid pFastBacI-[P(ie1)-egfp-sv40]-[P(polh)-ns1-sv40]. Then the plasmid was transformed into DH10B competent cells containing Bm-Bacmid vector to produce the final recombinant Bm-Bacmid with the help of transposase. The resulting recombinant Bm-Bacmid was transfected into BmN cells to generate recombinant virus, which was easily and rapidly judged by green fluorescent signal observed in BmN cells. After infection for 96 h, the BmN cells were harvested and the total protein extracted from the infected BmN cells was subjected to Western blotting analysis. The result showed that a specific protein band about 36 kDa was detected, indicating that NS1 protein was successfully expressed in the BmN cells. In conclusion, the expression of NS1 protein with the modified expression system is useful for further research on the function of NS1 protein.
Animals ; Baculoviridae ; genetics ; Bombyx ; virology ; Cell Line ; Cells, Cultured ; Gene Expression ; Genetic Vectors ; Plasmids ; Promoter Regions, Genetic ; Transfection ; Viral Nonstructural Proteins ; biosynthesis ; genetics

OBJECTIVE:To establish the quality standard for Mian medicine Indigofera stachyoides. METHODS:Medicinal properties and microscopic characteristics were identified;TLC was used for the qualitative identification;HPLC was conducted for content determination of epicatechin and 2α,3α-epoxy-5,7,3′,4′-tetra-hydroxy-flavan-(4β→8)-epicatechin(reference A):the col-umn was Comatex TM C18 with mobile phase of acetonitrile- water at a flow rate of 1.0 ml/min,and detection wavelength was 210 nm,column temperature was 30 ℃,injection volume was 5 μl. RESULTS:It showed clear microscopic identification map. I. stachyoides TLC had clear spots and well separated. The linear range was 0.274 5-4.392 0 μg (r=0.999 6) for epicatechin,and 0.103 0-1.648 3 μg for reference A(r=0.999 4),RSDs of precision,stability and reproducibility tests were lower than 3%,recov-eries were 99.76%-104.82%(RSD=2.42%,n=6) and 97.98%-104.99%(RSD=2.75%,n=6) respectively. CONCLUSIONS:The established standard can be used for the quality control of I. stachyoides.

Aim To study the progestogen effect and activity of triphasic contraceptive tablets.Methods The progestogen activity of triphasic contraceptive tablets (kalirui and trinordiol) was evaluated through uterus weight increase test of female young mice and endometrium translation examination of female young rabbits. Radialized receptor analysis was used to determine the effect of the triphasic contraceptive tablets on progestogen receptor by the uterus histiocyte from the adult female rabbits whose ovaries had been taken away a week ago. Results Compared with the negative control, triphasic contraceptive tablets kalirui andtrinordiol in three phase groups not only enhanced uterus weight of female young mice, middle and high dose groups obviously accelerated the uterus weight increase ( P

Objective To study vascular anatomy between the pancreatic head and duodenum,providing anatomy basis for performing surgery of pancreatic head,duodenum and distal common bile duct in surgical practice. Methods Anatomy study was performed in 30 formaldehyde fixed and 10 fresh bodies in reference to blood supply to duodenum,the distal common bile and Vater ampulla. Results The anterior and posterior pancreaticoduodenal arterial arcade gives off branches to descending and horizontal portion of the duodenum. The posterior superior pancreaticoduodenal artery goes to distal common bile duct. The papilla artery arising from the posterior superior pancreaticoduodenal artery goes to Vater ampulla. Conclusions The pancreaticoduodenal anterior and posterior arterial arcades are main arteries that give off branches to the descending and horizontal portion of the duodenum,distal common bile duct and the Vater ampulla,hence should be carefully protected in duodenum-preserving resection of the pancreatic head.

To investigate the safety, feasibility and effectiveness of transrectal high-intensity focused ultrasound (HIFU) in the ablation of canine prostate, 20 dogs were divided randomly into 5 groups. Sixteen canine prostates were treated with the third-generation transrectal HIFU device (Sonablate-500). Transrectal ultrasound images of the prostate and prostatic urethra were observed preoperatively and postoperatively. Serial study was performed 30 min, 30 days, 60 days and 180 days after the therapy. The rectum, periprostatic tissues, and prostate were excised en bloc and the tissues were fixed for gross and histological analysis. Our results showed that the average maximal diameter of prostatic urethra was 0.59+/-0.11 cm before the operation and 2.57+/-0.98 cm 60 days after the operation. The volume of prostate was 6.5+/-3.12 cm(3) before the treatment while the volume was 4.13+/-0.23 cm(3) 60 days after the treatment and the differences were statistically significant (P<0.05). Histologically, there was a clear demarcation between the necrotic area of the treated tissues and the unaffected surrounding tissues. All the necrotic tissues in the targeted zone broke off and the prostatic urethra became cavitary 60 days later. The more frequent complications were urinary retention and frequency and hematuria. No rectal injury occurred during the treatment. It is concluded that the third-generation transrectal HIFU is capable of destroying prostatic tissue, substantially increasing the width of the prostatic urethra without causing injury to the adjacent tissues. The risk of postoperative complications associated with HIFU was low. HIFU may become a safe, effective and minimally invasive alternative for the treatment of prostatic diseases.

BACKGROUND: It is easy to established human solid tumor nude mouse model, but for leukemia which is difficult. We inhibited immune system further by radioactive ray or CTX, to decrease cost and increase the stability.OBJECTIVE: To establish a human acute myeloblastic leukemia M2 Kasumi-1 models containing AML/ETO positive genes in BALB/c nude mouse. METHODS: Nude mice were randomly divided into three groups: CTX group was injected CTX 2 mg/day in abdominal cavity for two days, and injected 8×10~5/mouse Kasumi-1 cells in caudal vein next day; irradiation group was exposed to total body irradiation, and injected 8×10~5/mouse Kasumi-1 cells in caudal vein that day; untreated group was inoculated with 8×10~5/mouse Kasumi-1 cells by caudal vein injection. Three additional mice were considered as the normal control group. The blood smearing and bone morrow slides were detected, immunity type of BMC was detected using flow cytometry, loading of leukemic cellular tumor was detected using RT-PCR, and positive ratio of AML/ETO fusion gene was detected using FISH method. RESULTS AND CONCLUSION: After inoculated into untreated nude mice by caudal vein injection for 14 days, the ratio of leukemia cell in blood smearing was 3.5%, and over 40% in bone marrow slides, which was equal to the results of FISH and FCM. The increasing of tumor loading was time-dependent. For irradiation group and CTX treated group, the tumor loading was higher that untreated group, and the cells also survived more than 60 days. AML/ETO band was observed by RT-PCR in all experimental groups, for normal mice it was negative. The results indicated that the systemic disseminated leukemia model was established successfully by caudal vein injection 8×10~5/mouse Kasumi-1 cells in the three experimental groups.