Objective:To investigate the differentially expressed genes of gastric mucosa of precancerous lesion of chronic atrophic gastritis (CAG) in rats intervented by promoting blood flow therapy of TCM,and to analyze its mechanism at genetic level. Methods:The model of precancerous lesion of CAG rats was induced by spring insertion and intragastric administration of NaCl solution and hot paste. To detect the differentially expressed genes of precancerous lesion of CAG rats by gene chip technology,and the main differentially expressed genes were indentified by real-time PCR. Results:Compared with blank group,HSP70 and ptk2 significantly reduced in natural recovery group,the HSP70 expression increased signifi cantly after being treated with supplementing qi for promoting blood flow,and ptk2 expression increased obviously after being treated with regulating qi and promoting blood flow. The results of real-time PCR in expression level of HSP70mRNA and ptk2mRNA were the same as that of the gene chip technology. Conclusion:Supplementing qi for promoting blood flow can upregulate HSP70,thus inhibit precancerous lesion of CAG in rat gastric mucosal cell apoptosis and promote mucosal repair; Regulating qi and promoting blood flow can up-regulate ptk2,sequentially regular remodeling of actin cytoskeleton and promote repairing and healing of injury gastric mucosal. These may be one of mechanism that supplementing qi for promoting blood flow and regulating qi and promoting blood flow treat precancerous lesion of CAG.

Objective To study the application of hepatamethine cyanine near-infrared fluorescence (NIRF) dye IR-783 in the mouse models of human liver cancer exenografts, and to analyze the molecular mechanisms of the NIRF dye targeting tumor cells.Methods Luciferase-tagged HepG2 cells were inoculated subcutaneously into the nude mice.We detected the correlation of NIRF intensity and bioluminescence intensity (BIL) in the tumor region.Patient-derived xenograft (PDX) model was established in mouse by subrenal capsular implantation of clinic liver cancer specimen.After injecting the IR-783 dye, the interface between mouse kidney and the xenograft tumors was confirmed by NIRF analysis, and the tumor tissue in kidney was observed by pathology using H&E staining.The expression of CEA, AFP, HIF1α and OATP3A1 in the liver cancer tissue was detected by immunohistochemical staining.The intracellular retention of NIRF dyes was observed under fluorescence microscope after adding Mito Tracker or Lyso Tracker into cultured HepG2 cells.We added IR-783 in a co-culture system of HCCs and normal liver cells to test the specifical identification ability of IR-783 of the liver cancer cells.Results There was a good correlation between NIRF intensity and BIL intensity of the subcutaneous liver cancer xenograft region in nude mice.The margin between the mouse kidney tissue and xenograft tumors was clearly identified by IR-783.Compared with normal kidney tissue, CEA, HIF1α, OATP3A1 and AFP were highly expressed in the tumor region detected by IHC staining.The NIRF dye IR-783 was mainly accumulated in the mitochondria and lysosomes of cancer cells.GFP-tagged HepG2 cells could be recognized directly, whereas red fluorescence was not detected in normal liver cells.Conclusions IR-783 is a novel near-infrared fluorescent dye with tumor targeting and imaging properties.Its targeting ability may be related to the high expression of HIF1α and OATP3A1 in the liver cancer tissue.

Objective Getting the robust exogenous gene expression vector under the control of porcine insulin promoter, and to lay the foundation for pancreaticβ-cells specific transgene expressing pigs.Method Using porcine insu-lin promoter ( PIP, 1500 bp of the 5′UTR from the porcine INS gene including the first exon and the first intron) to con-struct expression vector, the HindIII restriction site which connected the sequences of PIP and EGFP was designed before ATG, named PIP-HindIII-EGFP.Considering that the different location of restriction site may affect the expression efficien-cy of the transgene, we optimized the expression vector.Firstly the HindIII restriction site was deleted to realize the seam-less connection of PIP and EGFP,the vector was named PIP-EGFP.Also we mutated the 3′intron splicing acceptor site( SA) of the first intron into HindIII restriction site, named as PIP-SA( M)-EGFP.Three different EGFP expression vectors were respectively transfected MIN-6 mouse pancreatic β-cells, pig ear fibroblasts and kidney cells.The transfected cells were cultured for 48 h and harvested for RT-PCR, flow cytometry and Western blot analysis, to analyze and compare the expres-sion efficiency of vectors.Results After transfection,green fluorescence was observed only in MIN-6 mouse pancreaticβ-cells.RT-PCR analysis and product sequencing showed that the three expression vectors did have different stability with in-tron splicing.The PIP-HindIII-EGFP construct and PIP-EGFP vector produced two kinds of mRNA with the first intron spliced and no spliced, indicating the instability of intron splicing.Mutation of the PIP splice site would cause the first in-tron not spliced, while flow cytometry and Western blot displayed that the mutation induced a most efficient expression of the downstream gene.Conclusions A robust and specific β-cells expression vector has been successfully generated by mutating the intron splicing acceptor site of the porcine insulin promoter.It provides the foundation for preparation of pigs with pancreaticβ-cells specifically expressing the transgene.

Objective To investigate the change of serum CA724, CA242 and AFP levels before and after transcatheter artery chemoembolization (TACE) treatment of primary hepatic carcinoma(PHC)patients as well as its clinic significance. Methods Patients of PHC (n=45) and healthy adults (n=40) were enrolled. Serum samples were collected from each healthy people and PHC patients 2 days before TACE,l week and 1 month after TACE. Electrochemiluminescence Immunoassay (ECLI) was used to determine serum CA724, CA242 and AFP levels, and correlations among three indexes as well as their relationships with clinical data were also analysed., Change of serum CA724 and CA242 levels before and after TACE were compared in AFP<400μg/L group and AFP≥400μg/L group. Results Serum levels of CA724, CA242 and AFP in PHC group were significantly higher than that in healthy control groups before TACE therapy(P<0.05). Positive rates of serum CA724 and AFP were higher in PHC group(P<0.01)than that in healthy control group. AFP decreased significantly at both 1 week and 1 month upon TACE treatment compared with that at before teratment(P<0.01). CA724 was significantly lower at one month after treatment than that at before treatment(P<0.05). But there was no significant difference in CA242 before and after treatment(P>0.05). CA724 and AFP expressions are associated with tumor size(P<0.05). After one month of therapy, serum CA724 level was obviously decreased in both AFP<400μg/L and AFP≥400μg/L groups. Before and after TACE therapy, there was a positive correlation between the expression of AFP and CA724(r=0.754,P<0.05). Conclusion Serum CA724 can be used as one of the tumor markers to assist the evaluation of curative effect of TACE on PHC.

Prostate cancer is one of the most common malignant tumors in men and related studies have achieved great breakthrough in recent years.But because of the lack of effective in vivo animal models, the process to translate basic research into clinical application has been severely hampered.Patient derived prostate tumor xenograft ( PDPTX) model is an ideal animal model in which freshly isolated tumor tissues from patients were inoculated into immunodeficient mice.This model can duplicate the heterogeneity of primary tumor in a better way and keep the tumor complexity at molecular, genetic and pathological levels.Particularly, the PDPTX model, in which the isolated tumor tissue is inoculated under the renal capsule, is even better, because it solves the clrawbacks of traditional subcutaneous inoculation model.In traditional mod-els, the success rate is low, it’s not easy for lower grade tumor to form xenograft, and it’s not easy to reconstruct metasta-sis, etc.PDPTX provides a more ideal in vivo model for prostate cancer studies.It has irreplaceable advantages, especially in target therapy, new drug screening and individualized tumor treatment.

ObjectiveTo investigate the mechanisms and the effects of magnesium Valproate on the expressions of the kinin B1 and B2 receptors in the hippocampus of the juvenile rats submitted to pilocarpine model of epilepsy.Methods 35 healthy Wistar juvenile rats were randomly divided into six groups,that is the model groups:Ⅰ group,Ⅱ group,Ⅲ group,and intraperitoneal injection of saline water control groups:Ⅰ a group,Ⅱ a group,Ⅲ a group,after succession of 15 rats to kindle to establish the model of epilepsy by pilocarpine.To collect hippocampus tissue after the rats were to put to death,and to compared the expression levels of kinin B1 and B2 receptor mRNA by RT-PCR and western blot in the hippocampus of rats.ResultsBy treated with magnesium valproate,kinin B1 receptor mRNA (0.38 ± 0.051 ) and protein expressions(0.58 ± 0.057 ) decreased and kinin B2 receptor mRNA (0.48 ±0.056 ) and protein expressions(0.48 ± 0.044 ) increased in Ⅰ group,compared with that (0.76 ±0.068,0.89 ± 0.034;0.28 ± 0.034,0.32 ± 0.039 ) of Ⅰ a group(P ＜ 0.05 ).Compared with control group,there were more significant upregulation of kinin B1 receptor mRNA and protein expressions (P＜0.05) in the Ⅰ and the Ⅱ groups and there were no alteration in Ⅲ group.The expressional levels of B2 receptor mRNA and protein were upregulated in the Ⅰ,Ⅱ and Ⅲ groups.ConclusionThe kinin B1 and B2 receptor may play a role in the onset and maintenance of epilepsy.The magnesium valproate increased the expressional levels of kinin B2 receptor,and decreased the expressional levels of kinin B1 receptor.

Objective To investigate the usefulness of Utstein template to guide the assessment and study of cardiopulmonary resuscitation (CPR) in our medical practice because it has been popularized in many countries since 1991.Methods A prospective observational study using Utstein CPR registry form to evaluate the epidemiological features and outcomes of 511 patients resuscitated in the emergency department.Results Of 511 CPR patients registered,higher cardiac arrest rates were observed in the group of patients aged 40- 70 years. In 511 CPR patients registered,preexisting chronic diseases were common including cardiovascular diseases ( 190,37.2％ ) cerebrovascular diseases (48,9.4％ ) and respiratory diseases (39,7.6％ ).Of them,173 cardiac arrest patients (33.9％) had underlying cardiac causes,such as acute myocardial infarction (AMI) found in 109 (21.3％) patients,and ventricular fibrillation witnessed during first cardiac monitoring in eighty ( 15.7％ ) patients.The restoration of spontaneous circulation (ROSC) rate and survival rate at discharge of in - hospital cardiac arrest (IHCA) patients were 47.0％ and 13.5％respectively,but 16.7％ and 4.7％ respectively in the out - hospital cardiac arrest (OHCA) patients (P ＜0.01,both ). Conclusions This study indicated that the cardiovascular diseases, cerebrovascular diseases,and respiratory diseases were the most common preexisting chronic diseases found in cardiac arrest patients.Myocardial infarct,stroke and trauma were the most common precipitation causes of cardiac arrest in the recruited patients.The rates of ROSC and survival at discharge were significantly higher in statistics in patients with IHCA than those in ones with OHCA.

Objective This study was aimed to characterize the swine leukocyte antigen( SLA) class I genes of GGTA1 -/ - Wuzhishan minipigs and compare their similarity to human leukocyte antigen( HLA) . It has important implica?tions for understanding the cellular rejection in xenotransplantation. Methods Specimens of ear tissue from six founding GGTA1 -/ - Wuzhishan minipigs were collected, and the SLA class I genes (SLA?1, SLA?3, SLA?2) were amplified by RT?PCR. Purified products were cloned into pEASY?T1 vectors and sequenced, followed by BLAST alignment and using bioin? formatc analysis to characterize the SLA class I genes and compare with the similarity to HLA. Results A total of six al?leles were detected, among them alleles were previously reported (SLA?1?0703,SLA?2?1102, SLA?3?0401, SLA?3?0403), and the other were novel (SLA?1?0401wz01, SLA?2?11wz01). The homology between alleles of SLA class I genes in Wuzhishan minipigs and HLA was from 70?5% to 72?1%. The homology analysis of critical amino acid residues on HLA binding with human CD8 + molecules showed that SLA?1?0401wz01, SLA?1?0703, SLA?2?11wz01, SLA?2?1102 and SLA?3?0401 occurred mutant at amino acid positions 225 and 228 ( T→S,T→M) , whereas the other loci were highly conserved. There was a high homology at amino acid level between SLA?2?11wz01, SLA?2?1102 and HLA class I genes which are NK cell KIRs binding sites. Conclusions The amino acid sequences of SLA class I genes of GGTA1 -/ -Wuzhishan minipigs have a high homology to HLA. From the point of view of cell?mediated xenograft rejection, the amino acid sequences of SLA class I genes of GGTA1 -/ - Wuzhishan minipigs have a high homology to HLA, therefore, Wzhishan minipigs may become a good potential donor for pig?human xenotransplantation.

Objective To establish a patient-derived gastric cancer xenograft( PDX) model in nude mice and to in-vestigate the application of near infrared fluorescent ( NIRF) dye IR-783 in in vivo imaging of gastric cancer xenograft mod-els.Methods Fresh human gastric cancer tissue was taken and transplanted into the subrenal capsule of nude mice to es-tablish the xenograft model.When the transplanted tumors grew,took part of the tumor tissue to do HE staining and compare the structural characteristics with the primary tumor.Another portion of the tumor was xenografted into nude mice subcutane-ously.Twenty days later,the tumor-bearing mice were injected intraperitoneally with IR-783 dye (10μM) in a dose of 100 mg/20 g.The intensity of the tumor image was monitored by optical NIRF imaging.The correction between tumor volume and fluorescence intensity was analyzed.Finally,the expression of OATP1B3 and HIF1αin the xenografted tumor tissue was detected by immunohistochemistry.Results We successfully established three patient-derived xenograft ( PDH) models of human gastric cancer.The transplanted tumor tissues maintained the histological characteristics of the primary tumor well.NIRF signal can be detec ted in subrenal capsule of the xenografted nude mice.The correlation between tumor size and fluorescence intensity in the PDX models reached higher than 98%.Strong positive expressions of HIF1αand OATP1B3 in the tumor tissues were detected.Conclusions NIRF dye IR-783 can be specifically accumulated at the tumor site,therefore, can be used to detect PDX in vivo early.The tumor targeting property may be related to the expression of OATP1B3 and HIF1α.

Objective To study the tumor targeting ability and application of farnesylthiosalicylic Acid (FTS) and heptamethine carbocyanine fluorescent dye-mediated near-infrared imagine in living animals, and confirm the inhibitory effect of this compound on growth of tumor cells.Methods Human breast cancer cell line MCF-7, glioma cell line U251 and prostate cancer cell line PC3 were cultured to logarithmic growth phase, and different concentrations of FTS and FTS-IR783 were added, respectively.We observed the inhibitory effect of those two compounds on the growth of tumor cells.Under fluorescence microscopy, specific accumulation of FTS-IR783 in these tumor cells was observed.The tumor cells (1×106) were transplanted subcutaneously into nude mice.These mice were subjected to intraperitoneal injection of FTS-IR783 (10 nmol/mouse) two weeks later.In the in vivo imaging, near infrared fluorescence signal and tumor volume were measured and their correlation was analyzed.Results Compared with FTS, FTS-IR783 significantly inhibited the growth of MCF-7, U251 and PC3 cells in vitro.FTS-IR783 was specifically uptaken by these three kinds of tumor cells, showing strong near infrared fluorescence in cell agglomerates.After subcutaneous injection of FTS-IR783, the correlation between fluorescence intensity and tumor volume was 0.987, 0.998 and 0.971, respectively.Conclusions The compound of FTS conjugated with near infrared fluorescent dye IR-783 can specifically recognize tumor cells, in both in vitro and in vivo imaging.At the same time, the compound can significantly inhibit the growth of tumor cells, and may be expected to become a new potential targeted drug.