Sucking mice, 2～3 day after birth, were intraperiton eally inoculated with 0.05mL F1M10 of Chen Strain Hantaan virus. At diff erent t ime point after inoculation, the brains were taken, routinely fixed and embedded in paraffin for preparing 5 μm serial sections. Traditional and confocal immun ochemical detection of viral antigens and heat shock protein 70 were performed t o exp lore the cerebral stress response after viral infection. The results showed that HSP70 immunoreactivities could be stably detected in the viral antigen positive neurons, but not in the viral antigen negative or uninfected control tissues. B y confocal microscopic examination, the HSP70 and viral antigens were colocolize d in the neuronal cytoplasm. Our result, comparable to our previous findings in human tissues and culture cells, indicated that Hantavirus infection can induce the expression of HSP70 in the infected cells, and HSP70 expression might be n ecessary but not sufficient to keep the cell survival.

Objective To detect the effect of cell density off both integrin αγβ6 expression and matrix metalloproteinase-9(MMP-9)secretion in colon cancer cells. Methods Flow cytometry was applied to analyze αγβ6 expression in human WiDr colon cancer cell lines and human HaCaT keratinocyte cells, respectively,at high-and low-cell density culture.The MMP-9 aetivity level for various coloIl cancer cell lines, WiDr and SW480 cells at high-and low-cell density culture was analyzed using Biotrak MMP-9 activity assay and Gelatin Zymography assay, respectively. Results High cell density significantly enhances integrin αγβ6 expression for WiDr cells expressing αγβ6 compared with low density,but no increase was observed for human keratinocyte HaCaT cells. Biotrak MMP-9 assay indicated that the amount of MMP-9 secreted per cell for WiDr and SW480 B6 cells at high cell density culture was(3.3±1.2)×10-7ng/cell and(27.2±3.0)× 10-7ng/cell respectively; However,at low cell density it was(1.8±0.7)× 10-7ng/cell and(10.9±2.0)×10-7 ng/cell,respectively. It was 2-3-fold higher for WiDr and SW480 β6 cells at high cell density compared with that at low cell density, but no density-dependent increase observed for SW480 wild cells lack αγβ6 expression(t=0.47,P＞0.05),MMP-9 secretion for SW480 wild cells was(3.9±1.7)× 10-7 ng/cell at hish cell density and(3.8 ±0.7)×10-7 ng/cell at low eell density(P＞0.05),respectively. Gelatin zymography assay also indicated that the level of MMP-9 in SW480 B6 cells expressing αγβ6 was evidently higher at high density than at low density, however no density-dependent increase observed for SW480 wild cells and HaCaT cells. Conclusions High cell density induces integrin αγβ6 expression and promotes MMP-9 secretion in colon cancer cells, which constitutes the basis for a self-perpetuating system of tumor infiltrating growth in colon cancer progression.

Objective To investigate the renal function changes of the children with breast milk jaundice(BMJ) and effect of early interference treatment on renal function. Methods Serum bilirubin and urine - minim protein (?2-MG,?1-MG, Alb and IgG) of the 50 patients with BMJ were measured when they were in hospital within 12 hours and the last day separately , at the same time, 20 healthy newborns had been chosen to serve as control group. Results Compared with control group, the urine minim protein of treatment group increased with the rise of serum bilirubin. When serum bilirubin was 205.2 - 256.5 ?mol/L, urine ?2- MG had mild increasing (P

Objective To investigate effects of rectal cancer to undergo total mesorectal excision (TME)or three space dissection(TSD)with pelvic autonomic nerve preservation(PANP).Methods TME or/ and TSD was applied in 247 Patients with advanced rectal cancer in which 185 cases (74.9 %) underwent PANP(Group P)including TME-PANP(Group Pro)139 cases and TSD-PANP(Group Ps)46 cases.The other 62 cases underwent none-PANP(Group P-)due to tumor invasion.Results There were no death cases for operation inall patients.Group Pm was better than Group Ps in the operation time and the difficulty of proce- dure(P0.05).Conclusion The procedure with TME to preserve pelvic autonomic nerves adapts to the majority of rectal cancer patients.TSD procedure is more complex than TME.Statistically,the survival differ- ence between Patients with TSD and with TME is no defective.The survival time is determined to the tumor's earlier diagnosis and therapy.

The gene coding for beta-glycosidase (EC3.2.1.21) from Thermus nonproteolyticus HG102 has been cloned and expressed in E. coli. The gene open reading frame was 1311 bp and it codes for 436 amino acids. The deduced amino acid sequence of the enzyme showed identity with members of glycosyl hydrolase family I. The enzyme had high content of hydrophobic amino acid (Ala 12.8%, Leu 10.9%), Arg(9.6%), Glu(9.4%) and Pro(8.0%), but low content Cys(0.45%) and Met (0.9%). From the alignment of enzyme amino acid sequence with other glycosyl hydrolase family I members, Glu164 and Glu338 were predicated as the proton donor and nucleophile group. The DNASTAR program was used to predict the secondary structure. According to the Chou-Fasman model, the enzyme has 41.4% of alpha-helics, 16.2%, beta-strands, 14.4%, beta-turns. 14 of the 35 Pro were located at the second sites of beta-turns. Hydrophobic interaction, ion bond, alpha-helics and Pro had important contribution to Tn-gly thermostability.
Amino Acid Sequence ; Cloning, Molecular ; Escherichia coli ; genetics ; Glycoside Hydrolases ; biosynthesis ; classification ; genetics ; Hot Temperature ; Molecular Sequence Data ; Open Reading Frames ; genetics ; Phylogeny ; Protein Structure, Secondary ; physiology ; Recombinant Proteins ; biosynthesis ; genetics ; Sequence Analysis, DNA ; methods ; Sequence Homology ; Thermus ; enzymology ; beta-Glucosidase

Objective To investigate the basic distribution of endemic areas in the type of drinking water arsenism and in Xiantao City,Hubei Province,and to offer a scientific basis for control and prevention.Methods According to"the Chinese Scheme of Implementing Surveillance of Distribution of Endemic Arsenism",considering with the special geography feature of Xiantao,both sampling and overall survey were used in 7 towns chosen.The water arsenic content was determined by half quantitative fast reagent-box method.We began to search for clues and patients according to the endemic areas and the families with high arsenic wells.Results High endemic arsenic water sources were distributed in 7 the towns(districts or farms).In 81 villages of Xiantao City,35 villages had drinking water arsenic content exceeding 0.05 mg/L,accounting for 43.21%(35/81).In 4020 screened wells,269 had arsenic content higher than the national standard,the detective rate of high arsenic wells(more than 0.05 mg/L)was 6.69%(269/4020),with the highest rate in Shahu Seed Plant being 13.56%(115/848).The population exposed to high arsenic was 1091,in a rate of 5.75%(1091/18 975),in which 281 children were exposed in a rate of 5.82%(281/4826).In Shahu Seed Plant,467 people including 129 children were exposed to high arsenic, accounting for 13.26%(467/3522)and 12.91%(129/999),respectively.Conclusions High arsenic sources widely exist in Xiantao City,especially in Shahu Seed Plant,where arsenic content,the exposed rate of population and children are high.Therefore,prevention and control should be carried out in the southeast as soon as possible,as well as in other places where situation is less serious.

OBJECTIVETo analyze the in-hospital mortality and factors affecting in-hospital mortality for patients with transposition of the great arteries (TGA) undergoing arterial switch operation (ASO).

METHODSBetween January 2004 and December 2007, ASO was performed in 169 patients [129 male, 40 female; mean age (11.71 ± 26.3) months] with TGA. The patients were divided in intact ventricular septum group (n = 56): TGA with intact ventricular septum and ventricular septal defect group (n = 113): TGA with ventricular septal defect. Multiple logistic regression analysis was performed to identify the risk factors of in-hospital mortality.

RESULTSThe overall in-hospital mortality was 11.24% (19/169). The yearly in-hospital mortality was similar between intact ventricular septum group and ventricular septal defect group. With the improvement of perioperative treatment, the in-hospital mortality decreased from 16.67% in 2004 to 3.92% in 2007. The multivariate analysis revealed that body weight ≤ 3 kg (OR: 4.571, P = 0.0409), complicating ventricular septal defect (OR: 4.444, P = 0.0406), complex TGA (OR: 4.321, P = 0.0140), coronary anomalies (OR: 4.867, P = 0.0104) and non-type A coronary arteries (OR: 3.045, P = 0.0243) were independent predictors for poor early postoperative survival.

CONCLUSIONBody weight ≤ 3 kg, complicating ventricular septal defect, complex TGA, coronary anomalies are independent predictors for increased in-hospital mortality in patients with transposition of TGA and undergoing arterial switch operation.

Arteries ; surgery ; Body Weight ; Cardiac Surgical Procedures ; mortality ; Child, Preschool ; Female ; Heart Septal Defects, Ventricular ; complications ; mortality ; surgery ; Hospital Mortality ; Humans ; Infant ; Logistic Models ; Male ; Risk Factors ; Transposition of Great Vessels ; mortality ; surgery

Beta-glycosidase (Tngly) from the thermophilic eubacterium Thermus nonproteolyticus HG102, which is a thermostable monomeric protein and adopts the (beta/alpha)8 barrel fold, is an excellent model system to be investigated for the thermostable mechanism, activity and substrate specificity. Here, based on the analysis of structural basis for thermostability of Tngly (Wang et al, 2003) and comparison of other proteins structure of homofamily, Glu164 and Glu338 may act as proton donor and nucleophile in the hydrolysis reaction respectively; proline located at N1 of alpha-helix and arginine which can form ion link may contribute to the thermostability. We aim to further identify the critical sites and the amino acid residue(s) responsible for the activity, the thermal stability and the substrate specificity. Mutations had been constructed by site-directed mutagenesis. They are Glu164Gln, Glu338Ala, Pro316Gly, Arg325Leu, Pro344Phe, Pro356Ala and Pro316Gly/Pro356Ala. All mutant proteins were purified to SDS-PAGE purity. Changes in the conformations were examined by means of CD. The Glu338Ala mutant showed no detectable hydrolysis activity, but can synthesize oligosaccharides, as expected for the residue acting as the nucleophile of the reaction. The Glu164 acts as the general acid/base catalyst in the hydrolysis reaction. Changes in stabilities of mutants compared with wild-type were determined by means of heat inactivity experiment. These results indicate that the amino acid residue of proline that is located at N1 positions of alpha-helix, and Arg325 that form salt bridge between alpha-helices 5 and alpha-helices 6, are the critical sites to protein thermostabilization.
Bacterial Proteins ; genetics ; metabolism ; Enzyme Stability ; Hot Temperature ; Hydrolysis ; Mutagenesis, Site-Directed ; Mutation ; Structure-Activity Relationship ; Thermus ; enzymology ; genetics ; beta-Glucosidase ; genetics ; metabolism

Fruit ripening is associated with a number of physiological and biochemical changes. They include degradation of chlorophyll, synthesis of flavor compounds, carotenoid biosynthesis, conversion of starch to sugars, cell wall solublisation and fruit softening. These changes are brought about by the expression of specific genes. People are interested in the molecular mechanism involved in the regulation of gene transcription during fruit ripening. Many fruit-specific promoters such as PG, E4, E8, and 2A11 have been characterized and shown to direct ripening-specific expression of reporter genes. AGPase plays the key role in catalyzing the biosynthesis of starch in plants. It is a heterotetrameric enzyme with two small subunits and two large subunits, which are encoded by different genes. In higher plants, small subunits are highly conserved among plant species and expressed in all tissues. And the large subunits are present at multiple isoforms and expressed in a tissue-specific pattern. In fruits, the expression pattern of the large subunits varies with plant species. That made it important to study the transcriptional regulation of the large subunits of AGPase in different plant species. Northern-blot analysis indicates in watermelon, an isoform of the large subunits Wml1 expressed specifically in fruits, not in leaves. The 5' flanking region of Wml1, which covers 1573bp, has been isolated through the method of uneven PCR. And transient expression assay has shown that the 1573bp (named WSP) can direct fruit-specific expression of GUS gene. Our goal in this study was to scan the promoter region for main regulatory regions involved in fruit-specific expression. A chimaeric gene was constructed containing the WSP promoter, the beta-glucuronidase (GUS) structural sequence as a reporter gene and the nopaline synthase polyadenylation site (NOS-ter). The plasmid pSPA was digested with Hind III + Hinc II and promoter fragment of 1573bp (from 180bp to 1752bp) was cut out and cloned into Sma I sites of pBluescript SK(-), to produce pBSPA-16. The same insert was then cut out with Hind III + BamH I, and ligated with transient expression vector pBI426 digested by HindIII + Bgl II to produce pISPA-16. Three 5'-end deletions of the promoter were obtained and fused to GUS gene in plant transient expression vector pBI426: the 1201bp fragment (from 551bp to 1752bp) was generated by digestion of pBSPA-16 with BamH I + SnaB I, the 898bp fragment (from 854bp to 1752bp) by BamH I + EcoRV. Both fragments were ligated with pBluescript SK(-) digested by BamH I + Sma I, to produce pBSPA-12 and pBS-PA-9. The inserts were cut out with HindmIII + BamH I and ligated with pBI426 digested by Hind III + Bgl II, to produce pISPA-12 and pISPA-9. The 795bp fragment (from 957bp to 1752bp) was generated by digestion of pSPA with Hinc II + EcoR I, promoter fragment was cut out and cloned into Sma I sites of pBluescript SK(-), to produce pBSPA-8. The same insert were cut out with Hind III + BamH I, and ligated with transient expression vector pBI426 digested by Hind III + Bgl II. The 1573bp fragment and three 5'-end deletions were delivered into watermelon leaf, stem, flower and fruit of different development stages (5, 10, 20 days after pollination) via particle bombardment using a biolistic PDS-1000/He particle gun. Bombardment parameters were as follows: a helium pressure of 1200 psi, vacuum of 91432.23Pa, 7 cm between the stopping screen and the plate. Histochemical assay were done on all the tissues bombarded after incubation for 2 days. The 1573bp fragment had the strongest promoter activity, and can induce GUS expression in fruits of 5 and 20 days after anthesis and flowers, but not in fruits of 10 days after anthesis, leaves and stems. Fragments of 1201bp and 898bp can induce GUS expression only in fruits of 20 days after anthesis, and with lower expression levels than 1573bp. Fragment of 795bp was not able to direct GUS expression in any of the tissues bombarded (data not shown). It can be concluded that of the 1573bp, 1201 bp, 898bp Wml1 5'flanking regions include the necessary information directing fruit-specific expression. Deletion from 180bp to 551bp doesn't affect the fruit-specificity of the promoter, but lowered the expression level. There may be some cis-acting elements located in this region, which can enhance external gene expression in later stages of fruit development. Deletion from 854bp and 958bp led to loss of GUS expression. This region includes the necessary information needed for gene expression as well as the regulatory elements for fruit-specific transcription.
Citrullus ; genetics ; Fruit ; genetics ; Gene Expression Regulation, Plant ; genetics ; Promoter Regions, Genetic ; genetics ; Regulatory Sequences, Nucleic Acid ; genetics ; physiology

OBJECTIVETo summarize the resectable rate of advanced gastric cancer characterized by abdominal lymph node metastases with the arteriovenous neoadjuvant FLEOX chemotherapy plus appropriate nutritional support.

METHODSArteriovenous neoadjuvant FLEOX chemotherapy was administered to 50 patients of gastric cancer with advanced abdominal lymph node metastases. Of the 50 patients, 42 had never received any previous therapy and preoperative CT scanning revealed unresectable tumors because of advanced lymph node (station No.3, 7, 9, 12) or distant lymph node (No.14,16) metastases. The other 8 were characterized with relapse of severe lymph node metastases or with unresectable lymph node metastases demonstrated by exploratory laparotomy. Arteriovenous neoadjuvant FLEOX chemotherapy was conducted as follows: from day 1 to day 5, 5-FU 370 mg/m(2) and leukovorin 30 mg/kg intravenously, at day 6 and day 20, CDDP 70 mg/m(2) and epotoside 70 mg/m(2) intraarterially. This FLEOX regimen was repeated every five weeks for two or three courses. Out of the 50 patients,12 malnutritional cases received parenteral and/or enteral nutritional support according to the nutritional condition. Arteriovenous neoadjuvant FLEOX chemotherapy was then administered after the improvement of nutritional state. Their nutritional support methods were adapted to their chemotherapy as well.

RESULTSAll the patients' general conditions and symptoms were improved significantly. For the 50 cases, the imageological and histological response rate (CR+PR) was 84.0%, and curative resection rate was 78.0%. Thus, 39 patients underwent subtotal or total gastrectomy, even combined organ resection, with D(2)+alpha or D(3) lymphadenectomy. Despite neoadjuvant chemotherapy, all malnutritional cases had significant weight gain after nutritional support, and other nutritional indexes,such as serum albumin, also resumed to normal.

CONCLUSIONArteriovenous neoadjuvant FLEOX chemotherapy proves favorable therapeutic effect for gastric cancer with advanced abdominal lymph node metastases, and downstages inoperable metastatic lymph nodes for radical operation. This combined modality regimen and nutritional support may play an important role in the treatment of advanced gastric cancer.

Adult ; Aged ; Chemotherapy, Adjuvant ; Female ; Gastrectomy ; Humans ; Male ; Middle Aged ; Neoadjuvant Therapy ; methods ; Nutritional Support ; Stomach Neoplasms ; surgery ; therapy