ObjectiveA HPLC-FLD method was developed to determine the levels of serum AAA in CRIpatients, and to studythe variationof serum AAAinCRI patientsanditsclinical significances. MethodsSerumsampleswerecollected from100healthcontrolsand 80CRI patients. According to 2002 National Kidney Foundation (NKF) staging diagnosis, CRI patients included 4 of stage 2, 12 of stage 3, 12 of stage 4, 52 of stage 5. According to pathogenesis, CRI patients were also divided into 3 groups :chronic nephritis group ( n = 32), DM group ( n = 36), hypertension group ( n = 12 ).Serums were deproteinized by equal volume of 5％ (v/v) PCA and supernate were analyzed direcdy. External standard method was used as quantitative method. The analytical column was Megres C18. 10％ acetonitrile in water was used as mobile phase. Flow rote was 1.0 ml/min. The wavelengths of fluorescence excitation and emission were changed with specific time. The levels of Tyr, Phe and Trp in CRI groups, different CKD stages and different pathogenesis were compared with healthy control groups to evaluated the sensitivity and specificity of serum AAA for CRI diagnosis. ResultsThe linear ranges of the method were 0. 550 -275.000, 3. 050 - 1220. 000 and 0. 049 -49. 000 pμmol/L for Tyr, Phe and Trp, respectively. The limit of detection (LOD) was 0.014 μmol/L for Tyr, 0.500μmol/L for Phe, and 0.005 μmol/L for Trp. The average recovery was 100. 9％, 101.3％ and 98. 5％ for Tyr, Phe and Trp, respectively. Intra-day CVwas 3. 18％ -4. 20％ ( mean was 3. 13％ )and inter-day CV was 3. 18％ -4. 20％ ( mean was 3. 58％ ). The concentration of serum AAA, Tyr and Trp and the ratio of Tyr/Phe in CRI patients were( 135.74 ±23.23 )μmol/L, (52.27 +8.25) μmol/L, (21.49 ±4.25) μmol/L and[0.87(0.68 - 1.05)]μmol/L. which were lower than that in healthy groups (t value was -14. 709, 4.452, 22. 100, U value was 266.000,respectively, P＜0. 05). The concentration of serum AAA, Tyr and Trp and the ratio of Tyr/Phe in healthy groups were ( 174. 47 ± 11.57 ) μmol/L, ( 63.53 ± 4. 68 ) μmol/L, (44. 22 ± 3. 67 ) μmol/L and[0. 97(0. 94 - 1.00)]μmoL/L. There were no statistically significant difference between the different stage of CRI. Compared with the concentration of Tyr, Phe and Trp among chronic nephritis group, DM group,hypertension group, the concentration of Tyr had no significant changes among these three kinds of diseases (P ＞ 0. 05 ). The concentration of Phe had significant changes between Chronic nephritis group and DM group, Chronic nephritis group and hypertension group ( U = 395.00, 114. 00, P ＜ 0. 05 ) ; the concentration of Trp haad significant changes between Chronic nephritis group and DM group ( U = 349.00, P ＜ 0. 05 ).The diagnostic sensitivity and specificity of serum AAA for CRI were 90％ (72/80) and 100. 0％ (100/100).ConclusionsThe method of high-performance liquid chromatography with fluorescence detection ( HPLC-FLD) is simple, rapid, sensitive and specific. Simultaneous determination of serum AAA was benefit to the diagnosis and evaluation of CRI patients.

Objective To discuss the feasibility and long-term effect of free latissimus dorsi muscle flap in repair of severe lower extremity injury in children. Methods From July 1999 to June 2004, nine child patients (at age of 6-13 years) with severe lower extremity injury involving soft tissue defects a-round the calf and the foot associated with complex open fractures, bare dislocation, and injury of the nerve, tendon and artery were repaired with free latissimus donsi flap, with flap area ranging from 30 cm ×12 cm to 10 cm × 5 cm. Results All the latissimus dorsi flaps survived, with success rate of 100%. A follow-up for 4-9 years showed that the flap had sound shape and function and normal blood supply, without significant influence on donor area. Conclusion Latissimus dorsi flap has advantages of constant anatomical site, abundant blood supply, massive area, strong anti-infection ability and less in-fluence on donor area and hence is an ideal method for repairing severe lower extremity injury in children.

OBJECTIVETo study the effects of low concentration dopamine(DA) on hydrogen peroxide-induced apoptosis in cultured rat cardiomyocytes as well as the possible molecular mechanisms.

METHODSCultured neonatal rat cardiomyocytes were randomly divided into the following groups: control group (control), hydrogen peroxide group (H2O2), pretreated with low concentration dopamine ( DA + H2O2), dopamine receptor l(DR1) antagonist group (DR1 + DA + H2O2), dopamine receptor 2(DR2) antagonist group (DR2 + DA + H2O2). The cell apoptosis was then assessed by MTT and flow cytometry. The cellular ultrastructure changes were observed by transmission electron micro- scope. The activity of lactate dehydrogenase(LDH )and superoxide dismutase (SOD) in cell medium was analyzed by colorimetry. The protein expressions of Cytochrone c, Caspase 3 and Caspase 9 were obtained by Western blot.

RESULTSCompared with hydrogen peroxide group, low concentration dopamine(10 µmol/L) decreased the apoptosis rate and the expression of protein of apoptosis related protein, enhanced SOD activity, decreased LDH activity. DR1 antagonist SCH-23390 treatment inhibited dopamine induced cardiac protective effect. DR2 antagonist haloperido treatment had no changes compared with dopamine group.

CONCLUSIONAbove findings indicate that low concentration dopanine inhibits apoptosis induced by hydrogen peroxide in neonatal rat cardiomyocytes, which is partly associated with the activation of DR1.

Animals ; Apoptosis ; Benzazepines ; Caspase 3 ; metabolism ; Caspase 9 ; metabolism ; Cells, Cultured ; Dopamine ; pharmacology ; Hydrogen Peroxide ; L-Lactate Dehydrogenase ; metabolism ; Myocytes, Cardiac ; drug effects ; Rats ; Rats, Wistar ; Receptors, Dopamine D1 ; metabolism ; Superoxide Dismutase ; metabolism

Objective To investigate the numbers of corpus luteum and ovarian follicles and compare the levels of serum prolactin (PRL), luteinizing hormone (LH), follicle-stimulating hormone (FSH) and estradiol (E2 ) in different phases of estrus cycle in female gerbils .Methods Consecutively taking vaginal smears of the gerbils and directly examined under light microscope to distinguish the four phases of the estrus cycle .Hematoxylin and eosin staining was used to histological examination of the gerbil ovaries , and to detect the levels of serum PRL , LH, FSH and E2 by ELISA assay during estrus cycle .Results The proportion of cornified vaginal exfolliated cells could be the basis to distinguish four phases respectively:proestrus, oestrus, metoestrus, and dioestrus.Moreover, there were no significant differences between the numbers of ovarian follicles in different phases of estrus cycle .The numbers of corpus luteum in preoestrus were significantly lower than that in the other phases of estrus cycle ( P <0.05 ) .The levels of serum PRL and LH were increasing constantly from preoestrus to dioestrus , and both reached a peak at dioestrus ( P<0.05 ) .The levels of serum FSH and E2 both peaked at preoestrus , and were significantly higher than those at oestrus , metoestrus and dioestrus ( P<0.05).Conclusions There are no significant differences between the numbers of ovarian follicles in different phases of estrus cycle .Gonadotropin , prolactin and estradiol paly important roles in the regulation of estrous cycle .The phases during which surges of FSH and E 2 occur in Mongolian gerbils are similar to those of rats and mice , while the PRL and LH are different .Our findings provide further reference to the study of reproductive physiology of Mongolian gerbils .

This study determined the whole genome sequence and phylogenetic characteristics of a rat coronavirus.Nucleic acids were extracted from rat intestinal tissues collected in Inner Mongolia,and high-throughput sequencing was performed.A novel alphacoronavirus was present in the samples.The complete genome was amplified with PCR and RACE.Multiple se-quence alignment and a phylogenetic tree were constructed in MEGA.The whole genome of the rat coronavirus,denoted NMR-13,was 27 674 bp and included two non-coding regions and eight open reading frames,successively 5'UTR-ORF1ab-S-ORF3-E-M-ORF6-N-ORF8-3'UTR.Sequence identity analysis indicated that NMR-13 was most closely related to alphacoronavirus,which shared 91.3％nt identity with strain FiCoV/UMN2020.NMR-13h shared the next highest-sequence identitywiththe strains Lucheng/Lijiang-170,Lucheng/Ruian-83 and Lucheng/Lijiang-71 found in Zhejiang Province,China(79.49％,80.6％and 81.0％,respectively).Phylogeneticanalysis indicated that NMR-13 clustered with FiCoV/UMN2020.Recombination analy-sis indicated no recombination phenomenon.A rat coronavirus was isolated in this study,thus enriching the diversity of known alphacoronaviruses,and providing a reference for understanding the molecular genetic characteristics and molecular evolution of mouse coronaviruses in China.

BACKGROUND:Influenza virus reverse genetics has reached a level of sophistication where one can confidently generate virus entirely form cloned cDNA. This system makes it possible to generate attenuated live virus vaccine candidate. We tried to generate human influenza A viruses encoding altered viral NS1 proteins in various available cell lines. MATERIALS AND METHODS:Eight (HA, NA, NP, M, NS, PBI, PB2, PA) viral and four (PB1, PB2, PA, NP) expression plasmids were generated from A/T exas/ 36/91 influenza virus by RT-PCR and cloning with POL-I and pGEM-T vector. Two NS1 mutant cDNA (NS1-126delta, NS1-99delta) were also generated. We transfected these plasmids into the 293T/MDCK, 293, CEF and Vero cells and incubated with culture media for 2-3 days. And then, we inoculated cell soups into the embryonated eggs. After 3-4 days of incubation, we harvested allantoic fluid and checked viral titer by HA assay. Finally we did RT-PCR and sequencing to confirm the virus. RESULTS:Finally we got the NS1 mutant A/Texas influenza viruses from 293T/MDCK cells, but not from FDA approved cells. However, whereas 293 cells are capable of being transfected and of growing the NS1 mutant viruses with low titer, CEF cells are only capable of growing this mutant viruses. CONCLUSION:293 and CEF cells could not be used alone for acquiring NS1 mutant A/Texas influenza viruses. However, 293/CEF co-culture seems to be a resonable next step for NS1 mutant virus rescue for human using.
Cell Line ; Clone Cells ; Cloning, Organism ; Coculture Techniques ; Culture Media ; DNA, Complementary ; Eggs ; Humans* ; Influenza, Human* ; Orthomyxoviridae ; Ovum ; Plasmids ; Reverse Genetics ; Vero Cells

Histone deacetylase (HDAC) is a kind of protease,which plays an important role in the structural modification of chromosomes and the regulation of gene expression.Its excessive expression in cancer cells causes acetylation imbalance,which is closely related to the occurrence of tumor.The high efficiency and low toxicity of histone deacetylase inhibitor (HDACi) has been widely recognized as anti-tumor drug with the deepening of the study in epigenetics.It is expected to bring more breakthroughs in the treatment of tumor.

BACKGROUND:Influenza virus reverse genetics has reached a level of sophistication where one can confidently generate virus entirely form cloned cDNA. This system makes it possible to generate attenuated live virus vaccine candidate. We tried to generate human influenza A viruses encoding altered viral NS1 proteins in various available cell lines. MATERIALS AND METHODS:Eight (HA, NA, NP, M, NS, PBI, PB2, PA) viral and four (PB1, PB2, PA, NP) expression plasmids were generated from A/T exas/ 36/91 influenza virus by RT-PCR and cloning with POL-I and pGEM-T vector. Two NS1 mutant cDNA (NS1-126delta, NS1-99delta) were also generated. We transfected these plasmids into the 293T/MDCK, 293, CEF and Vero cells and incubated with culture media for 2-3 days. And then, we inoculated cell soups into the embryonated eggs. After 3-4 days of incubation, we harvested allantoic fluid and checked viral titer by HA assay. Finally we did RT-PCR and sequencing to confirm the virus. RESULTS:Finally we got the NS1 mutant A/Texas influenza viruses from 293T/MDCK cells, but not from FDA approved cells. However, whereas 293 cells are capable of being transfected and of growing the NS1 mutant viruses with low titer, CEF cells are only capable of growing this mutant viruses. CONCLUSION:293 and CEF cells could not be used alone for acquiring NS1 mutant A/Texas influenza viruses. However, 293/CEF co-culture seems to be a resonable next step for NS1 mutant virus rescue for human using.
Cell Line ; Clone Cells ; Cloning, Organism ; Coculture Techniques ; Culture Media ; DNA, Complementary ; Eggs ; Humans* ; Influenza, Human* ; Orthomyxoviridae ; Ovum ; Plasmids ; Reverse Genetics ; Vero Cells

Four new triterpenoids, together with six known analogues, were isolated from an aqueous extract of the Ziziphus jujuba var. spinosa seeds, by multiple column chromatographic separation methods using stationary phases of macroporous adsorption resin, MCI resin, normal phase silica gel, Sephadex LH-20, and Toyopearl HW-40C as well as preparative thin-layer chromatography and reversed-phase HPLC. Their structures were determined by spectroscopic data analysis, the new structures were trivially named jujubaceanothoside A (1), 23-epijujuboside A (2), and jujubosides J and K (3 and 4), while the known analogues were identified as jujubosides A-C (5-7) and II (8), alphitolic acid (9), and betulinic acid (10). The structure of 1 was confirmed by single crystal X-ray diffraction.

Objective To explore the risk factors of recurrent low hemoglobin(RLH) level among students from 6 to 13 years old, and to formulate strategies and policies in this regard. Methods Surveillance on hemoglobin concentration was conducted among 71 742 students aged from 6 to 13y between 2013-2014 based on the annual physical examination for primary and middle school in Minhang District.Of those students, 670 were diagnosed with low hemoglobin level according to WHO criteria.A 1 ︰ 1 matched case-control study was conducted based on gender, age and school type.Questionnaire surveys for data collection were analyzed using Cox′s proportional hazards regression. Results Factors as pregnancy anemia(OR=2.32, 95%CI:1.49-3.63), non-pregnancy anemia(OR=4.65, 95%CI:1.22-17.69), maternal anemia(OR=2.51, 95%CI:1.50-4.21), drinking strong tea(OR=2.56, 95%CI:1.27-5.13) and dietary bias(OR=1.94, 95%CI:1.45-2.59) were risk factors of recurrent low hemoglobin level.Chewing pencils(OR=1.80, 95%CI:1.25-2.59) or toys(OR=1.79, 95%CI:1.10-2.86) and wasting(OR=3.37, 95%CI:1.11-10.21)might be the risk factors. Conclusion The risk factors of recurrent low hemoglobin level among students aged from 6 to 13 years are related to maternal anemia status and their dietary habits.We should strengthen education for women of child-bearing age, and help to develop healthy eating practices for their babies.Emaciated students should be focused on in this regard.