Complement Factor H ; genetics ; Hemolytic-Uremic Syndrome ; genetics ; Humans

Objective To determine whether the susceptibility to ischemia-reperfusion injury in aged heart is higher than that in adult heart and,if so,to clarify the mechanisms underlying this change.Methods Wister rats(5-or 20-month-old)were randomly divided into 4 groups(6 animals in each group).The rats were subjected to 30 minutes of myocardial ischemia via ligating the left anterior descending coronary artery,followed by 3 hours of reperfusion(Young-MI/R group and Old-MI/R group);A silk suture around the left anterior descending coronary artery was not ligated in young and old rats(Young-sham group and Old-sham group).Myocardial apoptosis was detected by terminal deoxynueleotidyl transferase biotin-d UTP nick end labeling(TUNEL)staining and caspase-3 activity was detected by using a caspase-3 colorimetrie assay.Nitrotyrosine content,a footprint of in vivo ONOO~-formation,and total NO content were determined by ELISA and chemiluminescence method respectively.Results A significantly exacerbated cardiac reperfusion injury was found in Old-MI/R group as evidenced by increased TUNEL positive myocytes[(19.0?2.1)% vs.(14.6?1.7)%],and increased myocardial caspase-3 activity[(436?35)?mool/mg vs.(340?32)?mol/mg] compared with Young-MI/R group(P＜0.05).Aged hearts had a markedly increased basal NOx level compared with young adult hearts.Marked higher myocardial nitrotyrosine content was found in OId-MI/R group[(7.25?0.18)nmol/g]than that in Young-MI/R group[(4.68?0.15)nmol/g] (P＜0.05).Conclusions In aged hearts,high levels of NO might form highly toxie derivant, ONOO~-,and its subsequent nitrified protein.This may attribute to the increased susceptibility of the aged heart to isehemic-reperfusion injury.

Preparation of a poly (gamma-glutamic acid)-cisplatin conjugate was introduced and its in vitro antitumor effect was investigated. Poly (gamma-glutamic acids) was obtained by using fermentation methods. The hydrolyzed small molecular weight of poly (gamma-glutamic acids) was prepared by acid hydrolysis. The interaction between poly (gamma-glutamic acids) -cisplatin conjugate (PGA-CDDP) and DNA was investigated by PCR model. MTT assay was used to investigate the in vitro anticancer activity of the conjugate. Apoptosis assay of the conjugate was investigated by FCM assay and the in vivo toxicity was also proceeded. The results showed that the poly (gamma-glutamic acids) -cisplatin conjugate was obtained successfully and its yield is 10% - 12%. It has obvious antitumor effects on human liver tumor BEL7404 cells, human lung tumor H446 cells and human colon tumor RKO cells. At the same time, it also has apoptosis effects on the three kinds of tumor cell lines. The in vivo toxicity of PGA-CDDP was examined in normal mice and the results showed that the in vivo toxicity of this conjugate was significantly lower than that of free CDDP. In conclusion, the poly (gamma-glutamic acids) -cisplatin conjuate could be used as a potential clinic antitumor drug. The poly (gamma-glutamic acids) obtained by fermentation can be used as a valuable drug carrier system.
Animals ; Antineoplastic Agents ; administration & dosage ; pharmacology ; Apoptosis ; drug effects ; Cell Survival ; drug effects ; Cisplatin ; administration & dosage ; pharmacology ; Drug Carriers ; Female ; Fermentation ; Hydrogen-Ion Concentration ; Male ; Mice ; Polyglutamic Acid ; administration & dosage ; pharmacology

OBJECTIVETo investigate class I integron of Shigella flexneri, its prevalence in children, and its relation to bacterial resistance to antimicrobial agents.

METHODSTotally 51 strains of Shigella flexneri were isolated from fecal samples of children suffering from bacterial diarrhea seen between June 2004 and November 2004 at Children's Hospital of Fudan University. Polymerase chain reaction (PCR) was employed to amplify various integron markers, including intI1, gene cassette region and 3' conserved region of class I intrgron; susceptibility of Shigella flexneri strains to 7 antimicrobial agents was determined by K-B (Kriby-Bauer) method.

RESULTSForty-six strains of Shigella flexneri had intI gene with a positive rate of 90.2% (46/51); 24 strains of Shigella flexneri were positive for qacEDelta1-sul1, the positive rate was 47.1% (24/51); proportion of the isolates positive for all the three regions of class I integron was 43.1% (22/51); 46 strains of intI positive Shigella flexneri were all positive for ant (3'')-I. Among 46 strains of intI positive isolates, proportions of the isolates positive and negative for qacEDelta1-sul1 were 47.8% (22/46) and 52.2% (24/46), respectively. In the class I integron positive Shigella flexneri, the resistance rates of ampicillin (chi(2) = 10.13, P < 0.01) and chloramphenicol (chi(2) = 19.97, P < 0.01) were significantly higher than those in the class I integron-negative group.

CONCLUSIONSClass I integron was detected in 90.2% of Shigella flexneri in children; carriage of class I integron is related to antimicrobial resistance of Shigella flexneri.

Anti-Bacterial Agents ; pharmacology ; Child ; DNA, Bacterial ; genetics ; Diarrhea ; microbiology ; Drug Resistance, Bacterial ; genetics ; Dysentery, Bacillary ; drug therapy ; microbiology ; Feces ; microbiology ; Humans ; Integrons ; drug effects ; genetics ; Polymerase Chain Reaction ; Retrospective Studies ; Shigella flexneri ; drug effects ; genetics ; isolation & purification

OBJECTIVETo establish a reversed-phase high-performance liquid chromatographic (RP-HPLC) method for determination of phloridzin content.

METHODSA RP-HPLC method was established for determination of phloridzin using an Inertsil ODS-3 (4.6×150 mm, 5 µm) column with the detection wavelength of 288 nm, flow rate of 1.0 ml/min, and column temperature of 25 degrees celsius;.

RESULTSThe result showed that the phloridzin had a good linear relationship when its concentration ranged between 0.5988 and 89.72 µg/ml. The regression equation was Y=46.370 X-0.6728 (r=0.9999, n=3). The average recovery of phloridzin was 99.40% with the relative standard deviations (RSD) of 0.67%.

CONCLUSIONThis method is simple, quick and accurate for determination of phloridzin content.

Chromatography, High Pressure Liquid ; methods ; Chromatography, Reverse-Phase ; methods ; Phlorhizin ; analysis

Human glutamate decarboxylase 65(hGAD65) is an enzyme that catalyzes the transformation of L-glutamic acid into ?-aminobutyric acid.It has been found that Type 1 diabetes mellitus(T1DM)is an autoimmune disease,in which pancreatic islet ?-cells are destroyed due to immune response mediated by autoantigen.hGAD65 is considered as a key autoantigen of the autoimmune response,so anti-hGAD65 antibody(hGAD65-Ab) is the most effective and specific immune marker for T1DM diagnosis,and hGAD65 can be used to detect hGAD65-Ab in serum of T1DM patients.The hGAD65 gene was cloned into pET32a(+),then the recombinant plasmid with hGAD65 was transformed into E.coli BL21(DE3) and expressed by IPTG induction.The fusion protein containing thioredox,hexahistidine and hGAD65(Trx-hGAD65) was mostly insoluble,but the band of soluble Trx-hGAD65 could also be detected by SDS-PAGE,and it was a great improvement compared with the results reported.Trx-hGAD65 was isolated from lysate and purified by immobilized metal ion affinity chromatography(IMAC).After enterokinase digestion and IMAC purification,hGAD65 with high purity was obtained.Detection of thin-layer chromatography(TLC) showed that both Trx-hGAD65 and hGAD65 had enzymatic activity,whereas Trx-hGAD65 had better stability.Furthermore,it was confirmed that Trx-hGAD65 was able to conjugate with hGAD65-Ab in the serum of T1DM patients by ELISA assay.In conclusion,Trx-hGAD65 instead of hGAD65 can be used for T1DM diagnosis,and its application in prophylaxis and therapy of T1DM is expectable.

BACKGROUNDIt is widely accepted that tumor necrosis factor-α (TNF-α) plays an important role in the pathogenesis of emphysema. This study aimed at investigating the protective effects of anti-TNF-α antibody, infliximab, in the development of emphysema induced by passive smoking in rats.

METHODSThirty-nine rats were randomly divided into a normal control group (group 1), an emphysema group (group 2), and an infliximab-intervention group (group 3). Rat models of emphysema were established by exposure to cigarette smoking daily for 74 days. After 1 month, the infliximab intervention group was treated with infliximab via subcutaneous injection. The levels of TNF-α, IL-8 and vascular endothelial growth factor (VEGF) in bronchoalveolar lavage fluid (BALF) were measured with enzyme linked immunosorbent assay (ELISA). The number and classification of cells in the BALF were measured. Lung tissue sections stained by hematoxylin and eosin (HE) were observed, and mean linear intercept (MLI) and mean alveolar numbers (MAN) were measured. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) methods were used to examine the percentage of positive cells and distribution of apoptotic cells.

RESULTSThe levels of TNF-α and IL-8 in BALF were higher in group 2 than in group 1 and group 3. The MLI was greater in group 2 than that in group 1 and group 3 while MAN was decreased. The concentration of VEGF in BALF of group 2 was significantly decreased as compared with group 1. The total cells and neutrophils number was significantly increased in group 2 as compared with group 1 and group 3, so was the percentage of neutrophils. The number of TUNEL positive cells in the alveolar septa was significantly increased in group 2 as compared with group 1 and group 3.

CONCLUSIONInfliximab protects against cigarette smoking-induced emphysema by reducing airway inflammation, attenuating alveolar septa cell apoptosis and improving pathological changes.

Animals ; Antibodies, Monoclonal ; therapeutic use ; Bronchoalveolar Lavage Fluid ; chemistry ; cytology ; Infliximab ; Interleukin-8 ; metabolism ; Male ; Pulmonary Alveoli ; cytology ; drug effects ; Pulmonary Emphysema ; chemically induced ; metabolism ; prevention & control ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Tobacco Smoke Pollution ; adverse effects ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVEMutations in NPHS2 mapped to 1q25-q31 and encoding podocin, which is exclusively expressed in glomerular podocytes, are responsible for autosomal recessive familial steroid-resistant nephrotic syndrome (SRNS) with minor glomerular abnormalities or focal segmental glomerulosclerosis. Different groups from European and North American countries have screened NPHS2 mutations in familial SRNS with recessive inheritance, documenting a mutation detection rate of 45% - 55% in families. This study aimed to examine mutations in the NPHS2 gene in Southern Chinese Han ethnic group patients with familial SRNS.

METHODSGenomic DNA from 3 probands from Southern Chinese Han families with autosomal recessive SRNS, and their siblings and parents was isolated and analyzed for all eight exons, exon-intron boundaries and promoter of NPHS2 using the polymerase chain reaction and direct sequencing.

RESULTSNo mutation of NPHS2 in all eight exons and exon-intron boundaries was identified in the 3 probands. However, a polymorphism of 954T > C in exon 8 was detected in all the 3 probands and some controls, and 5 variants of NPHS2 promoter, -1709G > A, -1000A > T, -670C > T, -116C > T and -51G > T, were identified in some patients and controls, indicating that these variants are polymorphisms. One heterozygous variant of NPHS2 promoter, -1715A > G, was also identified in one proband and her mother whose urinalyses were normal, whereas it was not found in any of the 50 controls. There was no significant difference in the allelic frequencies of -1709G > A, -1000A > T, -670C > T, -116C > T and -51G > T polymorphisms between the patients and controls.

CONCLUSIONNPHS2 mutations are not a major cause of familial steroid-resistant nephrotic syndrome in Southern Chinese Han ethnic group included in the study.

Adolescent ; Asian Continental Ancestry Group ; genetics ; Child ; Child, Preschool ; Female ; Gene Frequency ; Humans ; Infant ; Intracellular Signaling Peptides and Proteins ; genetics ; Male ; Membrane Proteins ; genetics ; Mutation ; Nephrotic Syndrome ; ethnology ; genetics ; Pedigree

OBJECTIVEA stable primary breast cancer model in liver-specific insulin-like growth factor 1 (IGF-1) deficient (LID) mice and control mice was established. To screen apoptosis related genes expression in different serum IGF-1 levels by gene chip and flow cytometry.

METHODSThe LID mice and control mice were used. Induction of breast cancer was achieved by using the 7,12-dimethylbenz(a) anthracene. Ginsenoside Rg3 was used to interfering therapy treatment. The incidence of breast cancer in every group was compared, and expression of apoptosis associated genes was detected by gene chip and flow cytometry.

RESULTSThe incidence of tumor in none ginsenoside Rg3 injected control mice was 66.7%. The incidence of tumor in ginsenoside Rg3 injected LID mice was 12.0% which was significantly lower than any other group (P < 0.05). The apoptosis percentage in none ginsenoside Rg3 injected control mice was (2.7 +/- 0.7)%. The apoptosis percentage in ginsenoside Rg3 injected LID mice was (14.0 +/- 1.7)%. The results of gene chip indicated that in contrast to LID mice, LTA, LTB, TNF-alpha, TRAIL, TRANCE, BLK, BOK, CASP8, TRAF5, and APAF1 genes were down-regulated, and LTBR, TRAF4 genes were up-regulated in the breast cancer tissues of control mice. Application of ginsenoside Rg3 therapy could change the expression of these genes.

CONCLUSIONSCirculating IGF-1 levels play a role in the onset and development of breast cancer. Degrade serum IGF-1 level is able to promote apoptosis by affecting the expression of a series of apoptosis related genes consequently inhibit the growth of breast cancer. There was a synergistic effect with the application of ginsenoside Rg3.

Animals ; Apoptosis ; Breast Neoplasms ; metabolism ; pathology ; Cell Proliferation ; Disease Models, Animal ; Female ; Insulin-Like Growth Factor I ; genetics ; metabolism ; Mice ; Mice, Knockout ; Oligonucleotide Array Sequence Analysis

OBJECTIVETo compare the protective activity of liver injury induced by D-galactosamine (GalN) between Huangqin-Tang and their metabolites by human intestinal bacteria(HIB).

METHODThe liver injuries in conventional and pseudo-germfree mice were induced by GalN. After oral administration of Huangqin-Tang and their metabolites mixtures by HIB, the serum transaminase (ALT and AST) activities were detected.

RESULTIn conventional mice, large and medium doses (20 and 10 g.kg-1) of Huangqin-Tang decoction significantly reduced the increase of serum ALT activity after 18 h GalN treatment. In pseudo-germfree mice, metabolites significantly reduced the ALT levels. However, Huangqing-Tang didn't affect the ALT levels in this kind of mice. To all of the animals, AST levels remained the same after oral Huangqin-tang or their metabolites.

CONCLUSIONThe metabolism by intestinal bacteria plays a role in pharmacological effects of constituents of Chinese herbal medicine. The metabolites of the constituents by intestinal bacteria were the real active components in vivo.

Administration, Oral ; Alanine Transaminase ; blood ; Animals ; Aspartate Aminotransferases ; blood ; Bacteria ; metabolism ; Chemical and Drug Induced Liver Injury ; Drugs, Chinese Herbal ; isolation & purification ; metabolism ; pharmacology ; Galactosamine ; Intestines ; microbiology ; Liver Diseases ; metabolism ; Male ; Mice ; Plants, Medicinal ; chemistry ; Protective Agents ; metabolism ; pharmacology