Objective: To construct subtracted cDNA libraries of stomach tumors and normal stomach tissue using suppression subtractive hybridization(SSH).Methods: cDNA Library subtraction was performed using the protocol described in the Clontech PCR-Select cDNA Subtraction Kit. cDNA was synthesized from 2 μg of poly A+RNA from the tumor and normal tissues using AMV reverse transcriptase. The tester and driver cDNAs were digested with RsaⅠ, a four-base-cutting restriction enzyme that yields blunt ends. The tester cDNA was then subdivided into two portions, and each was ligated with different cDNA adaptor. Two hybridizations were performed. In the first, an excess of driver was added to each sample of tester. Hybridization kinetics led to equalization and enrichment of differentially expressed sequences. During the second hybridization, the two primary hybridization samples were mixed together without denaturing and thus the templates were generated from differentially expressed sequences for PCR amplification. Using suppression PCR, only differentially expressed sequences were amplified exponentially and after second PCR amplification the background was reduced and differentially expressed sequences were further enriched. The cDNAs were then directly inserted into a T/A cloning vector to generate a stomach tumor-specific subtracted cDNA library. Results: The amplified library contained 800 positive clones. Plasmid inserts were PCR amplified and showed 250－700 bp inserts. Conclusions: The successfully constructed subtracted cDNA library of gastrocarcinoma and normal tissue enables us to compare two populations of mRNA and obtain clones of genes that expressed in one population but not in the other.The characterization of these genes will allow them to be exploited for their diagnostic and therapeutic potentials.

Female ; Humans ; Hypothyroidism ; drug therapy ; Infant, Newborn ; Thyroxine ; poisoning ; therapeutic use

Objective To summarize the experience in laparoscopic resection of metastic liver cancer. Methods The clinical data of the patients receiving laparoscopic resection of metastic liver cancer performed in Brisbane hospital from 1999 to 2006 were retrospectively analyzed. Results A total of 28 patients received the laparoscopic resection of metastic liver cancer. Procedures included left lateral segmentectomy in 13 patients, right hemihepatectomy in 9 and segmental or subsegmental resection in 6. Survival rate and disease free survival rate in 12 patients with colorectal metastasis being followed up for 2 years were 75% and 67%. Conclusion Laparoscopic liver resection is safe and feasible in strictly selected patients with malignancy in an appropriate stage. The surgeons should have rich experience of open liver resection and advanced laparoscopic technique.

Studies have shown that women's menopause caused by permanent cessation of ovarian function is closely related to lipid metabolism disorders. Er-xian Decoction has been used in the clinical treatment for gynecological diseases and has a good effect on diseases related to reduced sex hormone function. In this study, metabolomics was performed on bilateral ovariectomized model rats within 12 weeks after modeling to mimic the physiological state of menopausal women in different menopausal stages and Er-xian Decoction dosed model rats. The results of liver oil red O staining sections showed lipid metabolic disorder of bilateral ovariectomized model rats and the regulating effects of Er-xian Decoction. 46 potential biomarkers (6 steroid hormones, 3 sphingolipids, 11 phospholipids and 26 glycerides) in plasma and 32 potential biomarkers (1 steroid hormones, 20 phospholipids and 11 glycerides) in liver were obtained based on lipidomics analysis. Then, we analyzed the differential metabolic pathways and construct the lipid metabolism network significantly regulated by Er-xian Decoction. The results provided valuable information for in-depth understanding of the gradual changes on lipid metabolism disorders under menopausal conditions and the characteristics and mechanisms of compound Er-xian Decoction's regulatory effects. The study complied with the procedures established by the Animal Experiment Ethics Committee of the Institute of Materia Medica, Chinese Academy of Medical Sciences and passed the animal experiment ethics examine (No. 00000918).

Uridine diphosphate (UDP)-GalNAc : polypeptide N-acetylgalactosaminyltransfemse (ppGalNAcT) catalyzes the initial step in mucin type O-glycosylation in the Golgi apparatus. Here generation and characterization of a polyclonal antibody to human ppGalNAcT2 were described. The subcellular location of ppGalNAeT2 in SGC7901 cell line was investigated using Western blot analysis of fractionated cell extracts and confocal microscopy with this antibody and two Golgi markers: Golgi SNARE (soluble N-ethylmalemide-sensifive factor attachment protein receptor) of 28 ku (GS28) and trans-Golgi network (TGN) 38, markers for the c/s- and trans-Golgi apparatus, respectively. Morphometric analyses indicated that ～60% of the ppGalNAcT2 signal colocalized with the GS28, while～36% of the c/s-Golgi marker colocalized with the ppGalNAeT2. Approximately 34% of the ppGalNAcT2 signal colocalized with the TGN38, whereas 38% of the trans-Golgi marker colocalized with the ppGalNAcT2. The results provide unequivocal evidence for the location ofppGalNAcT2 within the Golgi apparatus, and further highlight the importance of this organelle in the initiation of O-linked glycosylation.

OBJECTIVETo investigate the protect effects of sodium ferulate (SF) on the daunormbicin(DNR-induced cardiotoxicity in juvenile rats.

METHODSForty male juvenile SD rats were randomly divided into control group (Control), daunorubicin group (DNR), sodium ferudate treatment group (DNR + SF), sodium ferudate group (SF) (n = 10) . Juvenile rats were intraperitoneally treated with DNR (2.5 mg/kg every week for a cumulative dose of 10 mg/kg) preparation immature myocardial injury model in presence with SF (60 mg/kg) oral treat- ment for 25 days. The left ventricular pressure and its response to isoproterenol were measured using left ventricular catheter. Rat myocardium myocardial pathology specimens and ultrastructure changes were also observed. The expression of cardiac Troponin I (cTNI) was detected by Western blot and RT-PCR. Results: SF treatment could inhibit the decreasing of heart rates induced by DNR damage (P < 0.05); it could increase the left ventrivular end diastolic pressure(LVEDP), heart rate, the maximal left ventrivular systolic speed(LVP + dp/dtmax) and the maximal left ventrivular diastolic speed (LVP-dp/dtmax) responding to isoproterenol stimulation(P < 0.01); SF also could improve the myocardial ultrastructure injuries and inhibit the decreasing of cTNI expression caused by DNR damages (P < 0.05).

CONCLUSIONSF treatment could alleviate the decreasing of cardiac reservation induced by DNR damages in juvenile rats, which might be related to its reversing the effects on the cardiac systolic and diastolic function injuries and its inhibiting effects on the decreasing of cTNI expression caused by DNR. The mechanism of SF preventing daunorubicin-induced cardiotoxicity in juvenile rats is relevant to inhabited cardiac Troponin I expression.

Animals ; Blood Pressure ; Cardiotoxicity ; drug therapy ; Coumaric Acids ; pharmacology ; Daunorubicin ; toxicity ; Heart ; physiopathology ; Heart Rate ; Isoproterenol ; Male ; Myocardium ; pathology ; Protective Agents ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Troponin I ; metabolism

The aim of this study was to synthesize 1-(5-(hydroxymethyl)-4-methyl-3-oxo-3,4-dihydro-pyrazin-2-yl) guanidine (mucunaguanide),a pyrazinone alkaloid compound extracted from the seed of Stizotobium cochinchinensis.Starting from benzyloxy acetaldehyde,following four steps in reaction including condensation,cyclization,substitution and hydrogenation reaction,mucunaguanide was synthesized with total yield of 27.9％,and purity of more than 97.5％.Structures of all the intermediates and target compound were confirmed by IR,1H NMR and MS.This synthetic process was characterized with raw materials available,simple in operation with much milder reaction conditions,and was an ideal method of synthesis of this compound.

Objective To develop a clinically useful assay for detecting the mutations of HBV pre-C/BCP based on the pyrosequencing and accuracy, reproducibility and reliability of this assay was evaluated. Methods The pyrosequencing primers for HBV pre-C/BCP mutation were designed through the cluster analysis among one hundred HBV gene sequences. After the amplification of the fragment of pre-C/BCP with the template of pre-C/BCP mutation plasmids, the pyrosequencing method for pre-C/BCP detection was initially set up with this standard sample. The accuracy, reliability and reproducibility of the pre-C/BCP pyrosequencing were confirmed through the pre-C/BCP plasmids as a standard sample when compared with Sanger/genechip sequencing method pre-C/BCP pyrosequencing assay was applied for detecting pre-C/BCP mutation types of 60 chnical serum samples in HBV patients. Results The pre-C/BCP mutation detection assay based on pyrosequencing has been established in our study. The coincidence rate between pyrosequencing and Sanger squencing was 100%. The coincidence rate between the result of pyrosequencing and of genechip method was 91.7%. The reproducibility of this assay was 97. 8%. It indicates the pre-C/BCP pyrosequencing is a high-accurate method with, good-reproducibility and high-reliability. And multi-site detection can be achieved by pyrosequencing one time. A rare mutation T1758C was also detected. Conclusion Pyrosequencing for pre-C/BCP mutations assay is high-throughout method for simultaneous detection of multi-site mutation.

To study the effect of hemoporphyrin derivative(HPD) combined with laser irradiation on human breast tumor cell line MCF-7 and normal human umbilical cord blood-derived hematopoietic cells by using MTT assay. The results showed that HPD plus laser irradiation was more efficient for killing MCF-7 cells than normal human umbilical cord blood-derived hematopoietic cells. The photochemical effect with laser irradiation were higher than with light irradiation and it's effect on MCF-7 cells was higher gradually with the increase of HPD concentration.

Adolescent ; Bacteria ; isolation & purification ; Child ; Child, Preschool ; Humans ; Infant ; Infant, Newborn ; Respiratory Tract Infections ; microbiology