Objectives:To explore the inhibition effect of TK/GCV and CD/5-FC system on liver metastasis of colon carcinoma.Methods:Forty nude mice were divided into 4 groups randomly,which were the control group,TK group,CD group and TK-CD group with 10 nude mice in each group.The mice in the control group were injected SW480 cells in the spleen and NS into the peritoneal cavity.The mice in other groups were all injected SW480/TK-CD cells in the spleen.And GVC,5-Fc,GVC+5F-c were injected into the peritoneal cavity of the mice in the TK group,CD group and TK-CD group respectively.Liver metastasis rate,the number of liver metastasis,the pathological changes of the tumor tissues under the electronic microscope,tumor cell's apoptotic index and life span and other indexes of nude mice in four groups were analyzed.Results:Liver metastasis rate of nude mice in every treatment group was lower than that in control group.Compared with control group,average number of liver metastasis decreased,life span of nude mice was prolonged,and the apoptosis rate of cancer cell in liver metastasis increased.The combined genes in the TK-CD group worked more markedly,and there was mutual effect between TK/GCV and CD/5-Fc.Conclusions:Synergistic effect was acquired combining TK/GCV with CD/5-FC system and it inhibited the formation of liver metastasis of colon carcinoma efficiently.

BACKGROUND:Studies have indicated that melatonin can promote the differentiation of adipose-derived stem cels into neurons, and the effect of melatonin on the osteoblasts form adipose-derived stem cels is rarely reported.
OBJECTIVE:To observe the osteogenic differentiation of adipose-derived stem cels and the effect of melatonin on the bio-viability of differentiated cels.
METHODS:(1) Adipose-derived stem cels were isolated and purified from the inguinal fat of Kunming mice by type I colagenase digestion and differential adhesion method, respectively. Immunohistochemical staining of CD44 was used as a quality control. (2) Osteogenic induction medium was added to induce osteogenic differentiation of passage 2 adipose-derived stem cels. Alkaline phosphatase staining and von Kossa method were combined to evaluate differentiation condition. (3) Melatonin at variable concentrations was added to treat mature osteocytes originated from adipose-derived stem cels and MTT was applied to determine their viability at 24 and 48 hours after culture respectively to find out optimal condition of melatonin treatment. (4) Melatonin at the optimal concentration was used to treat differentiated cels and detect alkaline phosphatase activity after 3 days and 6 days respectively.
RESULTS AND CONCLUSION: (1) After seeding for 48 hours, most cels were adherent, and after 4 days, the cels displayed multiple shapes and colonies of different sizes formed. After subculture, cel morphology homogenized as spindle shape. Cels positive for CD44 were brownish yelow, and localized mainly on the cel membrane. (2) Differentiated cels were positive for von Kossa staining and black sediments scattered in the extracelular matrix. Alkaline phosphatase expressed positively, and brown-black particles, appeared within cels. (3) Melatonin supplement improved the viability of differentiated cels; and 1, 10 and 100 μmol/L was observed as the optimal concentrations both at 24 and 48 hours. (4) The intracelular alkaline phosphatatse activity was increased with time in al the groups (P < 0.05). Compared with the blank group, the intracelular alkaline phosphatase activity in Melatonin groups (1, 10 and 100 μmol/L) had nochanges at 3 days, but significantly increased at 6 days (P < 0.05). These findings indicate that melatonin can enhance the proliferation of osteocytes differentiated from adipose-derived stem cels, and improve the activity of intracelular alkaline phosphatase.

Objective Using quantitative real-time PCR to establish a rapid specific genetic diagnostic technique for Yersinia pestis.Methods ①Four sets of specific probes and primers were designed,which targeted to chromosome genes of YPO0392,YPO1094,YPO2087 and YPO2090,respectively.②The probes and primers were tested for stability and specificity with 40 strains of Yersinia pestis and 47 strains of non-Yersinia pestis of different sources in Yunnan.③Eight positive DNA in Yulong,Yunnan,were tested with the screened probes and primers.Results ①Two sets probes and primers were selected,they were targeting YPO0392 and YPO1094,respectively.②The results were all positive of the eight positive DNA samples tested.Conclusion Two sets of primers and probes are selected for rapid specific diagnosis of Yersinia pestis.

Objective To discuss basic skills of the laparoscope doctor.Method A simulation operating system for die-spherical three-dimensional laparoscope was developed.Results The doctors trained by the simulation operating system behaved better than those untrained ones during operations.Conclusion It's a good way to enhancing basic skills of the laparoscope doctor to train them with the simulation operating system.

This paper focuses mainly on the study of optimal fermentation conditions of S-adenosyl-L-methionine.Effects of carbon sources,nitrogen sources,inorganic constituents,growth factors and adding time of L-methionine on the yield,the content and biomass of S-adenosyl-L-methionine are studied.And ingredients of the culture medium are also optimized by the method of uniform design.The final optimum culture medium contains: glucose 30 g,Yeast powder 11 g,(NH_4)_2SO_4 12 g,K_2HPO_4?3H_2O 5 g,KH_2PO_4 10 g,MnSO_4?H_2O 0.09 g,ZnSO_4?7H_2O 0.14 g,MgCl_20.5 g,CaCl_2 0.3 g,CuSO_4 0.005 g per liter. Using that optimum culture medium,the yield of S-adenosyl-L-methionine can reach 0.9 g/L in Erlenmeyer flask which is 30 % higher than before.Experiment on 5 L fermenter reveals that the accumulation of S-adenosyl-L-methionine can reach 2.66 g/L.Biomass is 23.4 g/L.

Objective To study the effects of glycogen synthase kinase-3?(GSK-3?)and free radical on neuron apoptosis of hypoxic-ischemic brain damage(HIBD)in newborn rats.Methods Eighty 7-day-old neonatal rats were randomly divided into 2 groups:normal group and hypoxic-ischemic(HI)group.Rats in HI group were subjected to left common carotid artery ligation,exposed to 80 mL/L oxygen and 920 mL/L nitrogen gas in 37 ℃ closed container for 2.5 h.Rats in 2 groups were killed at 6 hours,24 hours,48 hours,72 hours,5 days after hypoxia respectively.The neuron apoptosis was detected by flow cytometry.The activity of GOD-PX and the contents of SOD and MDA were detected by spectrophotometry,the level of GSK-3? was mensurated by Enzyme-linked immumosorbent assay(ELISA).Results The rates of neuronal apoptosis in HI group were significantly higher than those in normal group at 6,24,48 and 72 hours after hypoxia,respectively(Pa

SCHWABE Company in German is the first and largest manufacturer of Ginkgo biloba preparation. The company not only has leading technology in this field, but also protects its own market effectively through the high quality of patent drafting and exactly patent layout. Based on multi-angle analysis for patent portfolio of G. biloba preparation at application time, legal status, globally layout, Chinese layout, the article provides technical reference of research and development of G. biloba, also provides valuable experience of traditonal Chinese medicine patent portfolio layout for Chinese enterprises.
Drug Industry ; economics ; legislation & jurisprudence ; trends ; Ginkgo biloba ; chemistry ; Humans ; Patents as Topic ; legislation & jurisprudence ; Phytotherapy ; economics ; trends ; Plant Preparations ; isolation & purification ; Technology, Pharmaceutical ; economics ; trends

Objective To investigate the effect of Glycyrrhizin combined with puerarin on serum Leptin and insulin resistance in non-alcoholic fatty liver disease(NAFLD) patients.Methods One hundred and twenty patients with NAFLD were randomized into 4 groups,which were control group,compound Glycyrrhizin group,puerarin group,combined group,and each group was 30 cases.Patients in control group were received the regular liver protecting therapy including vitamins,amino acids,glucurolactone,in compound Glycyrrhizin group were given 60 ml compound glycyrrhizin solution (iv),in puerarin group were given puerarin at dose of 400 mg by intravenous infusion,and in combined group were given both compound glycyrrhizin and puerarin combination.All treatment period was 4 weeks.The levels of serum serum alanine aminotransferase (ALT),aspartate aminotransferase (AST),total cholesterol (TC),triglyceride (TG),leptin (LP),fasting blood glucose (FBG) and insulin(INS) were measured,and the insulin resistance index(IRI) was calculated.The liver CT image of patients were performed by Germany Siemens dual source CT instrument.Results The levels of serum ALT,AST,TC,TG,LP and IRI in control group at before and after treatment were ((83.08 ± 115.68) U/L vs.(43.32 ±11.72) U/L,(52.12±15.62) U/Lvs.(36.08 ±7.28) U/L,(6.20±1.30) mmol/Lvs.(5.60 ±0.70) mmol/L,(2.70 ±0.50) mmol/L vs.(2.10 ±0.40) mmol/L,(14.63 ±3.26) μg/L vs.(7.61 ± 2.46) μg/L,(7.9 ± 1.8) vs.(7.0 ± 1.2)),and the difference were statistically significant (t =12.828,4.244,16.648,21.442,3.341,16.152 respectively,P ＜ 0.01).The levels of serum ALT,AST,TC,TG,LP and IRI in compound glycyrrhizin group after treatment were ((43.28 ± 11.06) U/L,(37.28 ± 7.22) U/L,(5.70± 0.80) mmol/L,(2.20 ± 0.50) mmol/L,(7.89 ± 2.26) μg/L,(7.1 ± 1.6) respectively,significant different from before treatment ((83.06 ± 14.38) U/L,(51.68 ± 15.48) U/L,(6.30 ± 1.50) mmol/L,(2.60 ± 0.40) mmol/L,(15.13 ± 3.87) μg/L,(7.8 ± 2.2) respectively,t =8.893,4.225,16.520,24.708,6.353,21.137 respectively,P ＜ 0.01).The levels of serum ALT,AST,TC,TG,LP and ISI in puerarin group after treatment were (44.26 ± 9.68) U/L,(36.86 ± 6.88) U/L,(5.60 ± 0.70) mmol/L,(2.26 ± 0.48) mmol/L,(6.89 ± 2.18) μg/L,(7.0 ± 1.8) respectively,significant different from that before treatment ((82.68±14.36) U/L,(50.06±15.23) U/L,(6.20±1.60) mmol/L,(2.70±0.52) mmol/L,(15.68 ±3.26)μg/L,(7.7 ±2.8) respectively,t =7.087,8.138,18.159,7.244,7.470,32.283 respectively,P ＜ 0.01).The levels of serum ALT,AST,TC,TG,LP and ISI in combined treatment group after treatment were (22.28 ± 9.38)U/L,(28.48 ± 9.06) U/L,(5.00 ± 0.60) mmol/L,(1.70 ± 0.40) mmol/L,(4.63 ± 2.36) μg/L,(6.20± 1.6) respectively,significantly different from that before treatment ((84.62 ± 14.88) U/L,(49.12 ± 16.56)U/L,(5.70 ± 1.60) mmol/L,(2.78 ± 0.50) mmol/L,(14.78 ± 3.68) μg/L,(7.6 ± 2.1),t =14.255,11.272,8.371,9.941,8.102,37.626,P ＜ 0.01).The levels of serum ALT,AST,TC,TG,LP and ISI of patient were no significant difference before treatment,but after treatment,these indexes in combined therapy group were the lowest among 4 groups (P ＜ 0.05).And there were no significant difference among control group,compound glycyrrhizin group,puerarin group (P ＞ 0.05).Conclusion Compound glycyrrhizin combined with puerarin is proved to be an effect treatment method for NAFLD through decreasing serum leptin,improving insulin resistance.

Objective To investigate the relationship between expression of active form of caspase-3 and cell cycle in Fas-mediated apoptosis of B lymphocytoma cell line MML-1. Methods MML-1 cells were incubated with agonistic anti-Fas antibody for different time,and cell apoptosis was induced.Cell apoptotic rates were analysed by flow cytometry,and sensitivity of MML-1 cells to apoptosis was determined.The expression of active form of caspase-3 was analysed by double staining with PI-Triton X and FITC-active caspase-3.Cyclin A,B_1 and E were selected as cell cycle markers for S,G_2/M and G_1 phase of MML-1 cells,and the expression of active form of caspase-3 was detected by flow cytometry. Results The cell apoptotic rate reached 56% after induction by Fas for 6 h.After induction by Fas for 4 h,the active form of caspase-3 was mainly expressed in cells of G_1 phase,while rarely in cells of S and G_2/M phase.Cells with negative cyclin A and B_1 and positive cyclin E expressed active form of caspase-3. Conclusion The expression of active form of caspase-3 in MML-1 cells mediated by Fas might be cell cycle dependent.Cells entering into late G_1 and early S phase first express active form of caspase-3,and their sensitivity to Fas-mediated apoptosis is the highest.

Aim To investigate the pharmacodynamic experiment and molecular mechanisms of a diterpenoid from cortex pseudolaricis, pseudolaric acid B ( PB ) , on immunoregulation. Methods The mouse models of contact hypersensitivity ( CHS) were induced by 2,4-dinitrofluorobenzene ( DNFB ) . Then , the ear swelling and spleen index were measured after administered o-rally with PB. The pathological changes such as in-flammatory cell infiltration in ear skin were observed by hematoxylin and eosin ( HE ) staining. Besides, the expression of peroxisome proliferater-activated receptorγ ( PPARγ) and the phosphorylation of Akt were ana-lyzed by Western blot. The activity of PPARγ was fur-ther detected by luciferase reporter gene assay. Results The results showed that PB could both alleviate the ear thickness, inhibit the spleen index, and reduce the inflammatory degree of their ear skin, which might be involved in inducing PPARγexpression and activation, associated with suppressing Akt signaling pathway. Conclusion It is suggested that PB might regulate PPARγ-related Akt pathway, which indicates the pos-sibility of developing PB as a novel immunoregulation agent for treating inflammatory-immune disease.