Objective To investigate the significance of K-ras gene status and ras protein expression in immunophenotypic classification of gastric signet ring cell carcinoma.Methods The expression of ras protein in 180 cases of gastric signet-ring cell carcinoma was detected by tissue microarray immunohistochemistry.Meanwhile,the mutation in codon 12,13 of K-ras gene was determined by using PCR-based DNA direct sequencing analysis.Results The rate of ras protein expression was 27.8％.The rate of ras protein expression in intestinal phenotype was significantly higher than those in gastric and gastrointestinal phenotypes(P<0.05).The rate of ras protein expression in cases with lymph node metastasis was significantly higher than those in cases without nodal involvement(P<0.05).The rate of ras protein expression was significantly higher in cases with deeper invasion(P<0.05).The frequency of K-ras gene mutation was 22(12.2％).All of them were found in codon 12.The types of mutation included GGT→AGT(1 case),GGT→TGT(1 case),GGT→GCT(2 cases),GGT→GTT(8 cases)and GGT→GAT(10 cases).K-ras mutation was significantly associated with intestinal phenotype(P<0.05).The rates of ras protein expression in cases with mutational type of K-ras gene was higher than those in cases with wild type(P<0.05).The ras protein expression was positively associated with K-ras gene mutation(r=0.61,P<0.05).Conclusion The ras protein expression is correlated with nodal involvement and invasion.K-ras gene mutation and expression of ras protein is related to phenotypic classification,and they might influence the phenotypic transformation in gastric signet ring cell carcinoma.

0.05).CONCLUSION:Nedaplatin combined with docetaxel is effective and safe for advanced NSCLC with high short-term efficacy and mild toxic side reaction in digestive tract and kidney.but the long-term efficacy of it require further study.

BACKGROUND: Parkinson disease(PD) is a series of clinical symptom induced by decreased dopamine (DA) in the striatum due to nigral dopaminergic neuronal degeneration. The intracerebral transplantation of secretory DA can reverse or improve the symptoms to a certain extent, but immunologic rejection is still existed.OBJECTIVE: To probe into cell transplantation with immunoisolation in treatment of in rats without application of immunosuppress and observe its mechanical intensity and the biocompatibility of microcapsule .DESIGN: Randomized controlled experiment was designed.SETTING: Biomedical Material Engineering Group, Dalian Institute of ChemicalPhysics , Chinese Academy of Sciences, and Department of Neurology, Second Hospital of Jilin University.MATERIALS: The experiment was performed in Animal Experimental Center of Second Hospital of Jilin University from August 2003 to February 2004, in which, 40 male Wistar rats were employed. PC12 cell was provided from Shanghai Institute of Cellular Biology of Chinese Academy of Sciences.METHODS: 6-hydroxydopamine solution was infused in the striatum to prepare animal model of Parkinson disease. Twenty-five rats of those had been prepared successfully and were randomized into microencapsulated cell transplantation group (12 rats), in which, 25 μL cell-loading sodium alginate-chitosan-solium alginate(ACA)microencapsul suspension (equal to 2.5×104 cells) was injected stereotaxically on two points of the right (affected side) striatum of animal model; non-microencapsulated cell transplantation group (7 rats), in which, 25 μL PC12 cell suspension (equal to 5×104cells) was injected; and empty microcapsul transplantation group (6 rats),in which, 25 μL empty microcapsules suspension was injected . On the 7th day after transplantation, in every group, apomorphine (APO) prepared with saline solution was injected (0.05 mg/kg) subcutaneously in the neck; afterwards, the revolving behavior was recorded for each rat, once per week,totally for 12 weeks. In the 12th week after operation, the rats were sacrificed with anesthesia. The brain tissue was collected for pathological observation and microcapsule were retrieved to evaluation of biocompatibility and immunoisolation.numbers before and after transplantation of each group.RESULTS:Twenty-five rats entered result analysis and the rest was sule: the retrieved ACA microcapsule was integrative in morphology,munoisolation of microcapsule: microencapsuled PC12 cells were prolifercycles before and after transplantation of each group: the records of lateral revolving of rats in every group before transplantation were not significantly different (P ＞ 0.05). In microencapsuled cell transplantation group, 2weeks later, the average number of revolving was significantly lower than that before the transplantation, or even the revolving stopped; the improved symptoms were maintained till the 12th week after transplantation. In nonmicroencapsulated cell transplantation group, the average revolving number was also significantly lower than that before the transplantation, but that on the 8th and 12th weeks was in tendency of increase, without obvious change compared with that before the transplantation (P ＞ 0.05). The revolving number before and after transplantation in non-microencapsulated transplantation group was similar[(10.5±1.4), (10.5±1.3) cyclos/min, P ＞ 0.05].microcapsule provides immune protection. The grafted encapsulated PC12cells survive for along term in the brain of rats with PD, maintain continuously the normal physiological function and improve the symptoms of PD by synthesizing and releasing DA.

Objective To investigate the variation of biomarker of coagulation, anti-coagulation, fibrinolysis in elderly critical patients and find out whether they are related to the disease severity. Methods Sixty-seven patients were no less than 60 years old. Eligible criteria: coincidence with the diagnostic criteria of systemic inflammatory response syndrome (SIRS) and APACHE Ⅱ score was no less than 10 scores. Blood sample was drawn from the venous for the test of biomarker (APTT, PT, TT, D-D, Fib, AT-Ⅲ , PC, PAI-1). According to the existent status,all the patients were divided into two groups:survival group (43 cases) and death group(24 cases) ,meanwhile,according to the diagnostic criteria of MODSE,all the patients were divided into MODSE group (30 cases) and non-MODSE group (37 cases). Results There were significant differences in APACHE Ⅱ score between MODSE group and non-MODSE group, survival group and death group [(25.83 ± 1.19) scores vs(18.1±20.73) scores and(18.81±0.72) scores vs(26.50 ± 1.42) scores](P <0.01). The PT and D-D in MODSE group anti death group were higher than those in non-MODSE group and survival group, the differences were significant (P <0.05),while the activity of AT-Ⅲand PC in MODSE group and death group were lower than those in non-MODSE group and survival group, the differences were significant (P <0.05). The PT,D-D and PAI-1 were positively correlated to APACHE Ⅱ score (related coefficients were 0.328, 0.308, 0.335,P <0.05). The AT-Ⅲ and PC were negatively correlated to APACHE Ⅱ score (related coefficients were -0.469, -0.559,P <0.01). Conclusions The abnormality of eoagnlation-fibfinolysis system exists in elderly critical patients. The extended PT, elevated D-D and PAI-1 ,descended PC and AT-Ⅲ are the hints of disease severity and poor prognosis.

To compare the difference in tumor immunity and autoimmunity elicited by adenovirus (Ad) encoding human or murine tyrosinase-related protein 2 (AdhTRP2 or AdmTRP2), and to find the most effective way to induce immunity by AdhTRP2 or AdmTRP2, C57BL/6 mice were immunized with AdhTRP2 or AdmTRP2 intramuscularly at different doses of 10(5), 10(6), 10(7) and 10(8) separately (10 mice for each dose). Two weeks after the immunization, in vivo CTL assay and intracellular staining (ICS) of IFN-gamma were carried out to analyze the dose-effect relationship. Tumor growth and vitiligo (as an sign of autoimmunity) were observed until 3 months after challenge with 10(5) B16F10 tumor cells. The results showed that Ad encoding AdmTrp2 induced weak tumor immune response. Similar immunization with AdhTrp-2 elicited stronger protective immunity. CTL activity and IFN-gamma-produced CD8+T cells were directly proportional to dose of AdhTrp2 or AdmTrp2. Moreover, AdhTrp2 group showed tumor rejection in 100% of challenged mice till the end of 3rd month while 60% of mice immunized with AdmTrp2 were protected against tumor. In the whole process of this experiment, no vitiligo was observed in mice immunized either with AdhTrp2 or AdmTrp2. It is concluded that anti-melanoma responses induced by genetic vaccination expressing xenoantigens breaks immune tolerance effectively and is able to elicit strong antigen-specific cytotoxic T cell response without vitiligo.
Adenoviridae/metabolism ; Antineoplastic Agents/*pharmacology ; Cell Line, Tumor ; Cytokines/metabolism ; Immune System ; Immune Tolerance ; Interferon-gamma/metabolism ; Intramolecular Oxidoreductases/*biosynthesis ; Intramolecular Oxidoreductases/*genetics ; Mice, Inbred C57BL ; T-Lymphocytes, Cytotoxic/*metabolism ; Vitiligo/*metabolism

Objective To explore the impact of four different anesthetic drugs commonly used in animal experi-ments on cardiovascular system in rats.Methods Electrocardiogram ( ECG) and blood pressure were dynamically recor-ded by a BioPac MP150 system after anesthesia.In addition, the blood glucose at different time points and hepatic func-tion, kidney function, cardiac enzymes and electrolytes at the end of the test were collected.Rusults Chloral hydrate caused severe ventricular arrhythmia.Isoflurane had inhibitory effect on the heart rate.Pentobarbital sodium induced a in-crease of ECG P wave.Urethane caused J point elevation of ECG.Blood pressure in the urethane-and pentobarbital sodi-um-treated groups were increased.Chloral hydrate caused CK to be raised, while isoflurane showed the opposite effect on CK and CKMB.Alanine aminotransferase and aspartate aminotransferase in the pentobarbitol sodium and isoflurane groups were decreased.Creatinine in the chloral hydrate, pentobarbital sodium and isoflurane groups were lower, and the serum sodium and potassium were decreased in the four groups.Conclusions Chloral hydrate has obvious effect on the cardio-vascular system, and is not suitable for animal studies on cardiovascular diseases.Pentobarbital sodium, urethane, isoflu-rane can be chosen for animal studies on cardiovascular diseases.

Objective To evaluate the effects of Ulinastatin (UTI) on the expressions of TNF-α,IL-6 and neurons apoptosis in cerebral cortex of rats after cardiopulmonary resuscitation (CPR).Methods Thirty-six healthy male adult Wistar rats were induced ventricular fibrillation untreated for 7 min and then received CPR.The animals were infused UTI 100 000 U/kg or phosphate-buffered solution (PBS) at once after ROSC.At 2,4 and 8 h after ROSC,cerebral cortex were removed to determine the mRNA expressions and levels of TNF-α protein and IL-6 protein,the translocation ratio of NF-κB p65 from cytoplasm to nucleus and the apoptotic neurons.Results The plasma levels of TNF-α (ng/mL) in animals of UTI group were (17.7 ± 1.4),(21.9 ± 2.1) and (17.1 ± 0.6) at 2,4 and 8 h after ROSC respectively,and significantly lower than those in PBS group at the given intervals.Mean while,the levels of IL-6 (ng/mL) were (208.9 ± 14.1),(281.5 ±25.9) and (251.8 ± 15.3) at 2,4 and 8 h after ROSC respectivèly in animals of UTI group,and lower than those in PBS group.The expressions of TNF-α mRNA and IL-6 mRNA and protein levels of TNF-α and IL-6 in UTI group were both lower than those in PBS group at given intervals,respectively.The translocation ratio of NF-κB p65 from plasma to nucleus in PBS group at each given interval after ROSC was significantly higher than that in UTI group.The number of viable neurons in cerebral cortex in UTI group was higher than that in PBS group,while the number apoptosis neurons was fewer in UTI group.Conclusions UTI attenuated the general inflammatory response after ROSC in rat,decreased the activation of NF-κB pathway,and subsequently attenuated the expression of TNF-α and IL-6,and finally decreased the neurons apoptosis.

Objective To establish a rapid fluorescent inhibition test (RFFIT) for testing rabies virus neutralizing antibody and the titer of rabies virus neutralizing antibody.CVS-11 was used as the standard challenge virus,and three generations prepared for the establishment of the virus library.Methods International standard for rabies immunoglobulin was used as the reference serum.RFFIT test was established under consulting the protocol of Institute of Pasteur,and its specificity,stability and reproducibility were validated.Results We established the RFFIT which showed both good specificity ( 100% ) and reproducibility (P＞0.5).Conclusion The establishment of RFFIT test perfected the rabies laboratory techniques and would enhance the overall ability in detecting rabies in China.

Combined hepatocellular-cholangiocarcinoma (CHC) is a mixed tumor containing elements of both hepatocellular carcinoma (HCC) and cholangiocarcinoma (CC).Its remarkable histological heterogeneity has been linked to putative hepatic progenitor cell (HPC) origin.However,detailed histological or phenotypic description is rarely documented.In the present study,we reassessed 68 cases previously diagnosed as hepatitis B-related CHCs by immunohistochemistry and double-fluorescence immunostaining,focusing on HPC associated phenotypic observation of intermediate area of the tumor.It was found that tumor cells showed remarkable heterogeneity in intermediate area.Tumor cells with intermediate morphology between hepatocytes and cholangiocytes were oval-shaped and small with scant cytoplasm and hyperchromatic nuclei,arranging in solid nests mostly.By Keratin 7 (K7) staining,it appeared that the nests of tumor cells represented a maturation process from the undifferentiated small cells to mature hepatocytes through the "transitional" cells.Then,these small cells were further confirmed with intermediate phenotype as HPC by exploring immature hepatocellular marker and HPC/biliary markers co-localization.In conclusion,the HPC associated trait in CHC can be interpreted by HPC origin or gain of"stemness" by dedifferentiation.It is still too soon to give a final word that it is innate or acquired signature of HPC associated trait in CHC.

Objective To study the effects of mild hypothermia (MH) on blood coagulation and cerebral microcirculation in rabbits after cardiopulmonary resuscitation (CPR). Method A total of 24 New Zealand rabbits were randomly (random number) divided equally into normothermic group (NT) and MH group. CPR model was established by ventricular fibrillation induced by using alternating current. The rabbits of NT group were observed for 12 h in room temperature after restoration of spontaneous circulation (ROSC). The mild hypothermia was induced in the rabbits of group MH by surface cooling after ROSC, and maintained for 12 h after the aimed low temperature reached. The PT (prothrombin time), APTT (activated partial thromboplastin time), INR (international normalized ratio of prothrombin), D-dimmer (DD) , blood platelet count (BPC) , anti-thrombin Ⅲ activity (AT-Ⅲ) and protein C activity (PC) were measured before CPR and 4 h, 8 h and 12 h after ROSC, and at the same time the cerebral microcirculation was measured by using PERIMED Multichannel Laser Doppler system. One-way ANOVA or Mann-Whitney rank was used to determine the statistical significance between two groups. LSD-t test was used for multiple comparisons,t test for comparisons of means between two independent samples, and Pearson correlation test for correlation analysis. Results The PT, APTT and INR showed a trend of gradually shortening during the course. The APTT in 12 h after ROSC was significantly shorter than that before CPR (23.32 ±5.19 vs. 29.53 ±5.10,P = 0.025), and the activity of AT- Ⅲ and PC were decreased significantly. Compared with the group NT,the PT, APTT and INR in group MH were increased significantly, while there were no differences in the activity of AT- Ⅲ, PC and D-D between two groups. The rates of cerebral microcirculation in group NT before CPR and 4 h, 8 h and 12 h after ROSC were 401.60 ± 11.76 mL/min, 258.86 ± 34. 58 mL/min,317.59 ± 23.36 mL/min and 371.98 ± 5.79 mL/min, respectively, and those in group MT were 398.18 ±12.91 mL/min, 336.19 ± 19.27 mL/min, 347.76 ± 13.80 mL/min and 383.78 ± 3.29 mL/min, respectively. There were significant differences between two groups at each interval after ROSC (4 h: t = - 6.025,df=16, P=0.000;8 h: t= -2.942, df=12, P=0.012;12 h: t= -3.959, df=8, P=0.004). The Pearson correlation test showed that the rate of cerebral microcirculation was positive correlated with APTT after ROSC (4 h:R =0.503,P=0.033;8 h:R=0. 565,P=0. 035;12 h:R=0. 774,P=0. 009), and was not correlated with the other blood coagulants. Conclusions The mild hypothermia led to the inhibition of blood coagulation and improved the cerebral microcirculation concomitantly, which may be one of the mechanism of cerebral protection.