Objective To understand the origin and variation of the hemagglutinin gene of isolates viruses from 3 novel influenza A( H1N1 ) deaths in Changsha ( A/Hunan Kaifu/SWL4142/2009 ( H1N1 ) , A/Hunan Changsha/SWL4346/2009 ( H1 N1 ) and A/Hunan Furong/SWL4224/2009( H1N1 )). Methods The nasopharyngeal swab specimens from the 3 novel influenza A( H1N1 ) deaths in Changsha were tested by RT-PCR and influenza viruses were isolated simultaneously. With the sequencing primers recommended by World Health Organization (WHO), the HA gene of sequences of 3 novel influenza A( H1N1 ) deaths were tested by CEQTM 8000 Genetic Analysis System, through dye terminator cycle sequencing. The sequencing results were submitted to GenBank, then the results were analyzed for amino acid alignment and phylogenetic tree analysis with ClustalX and Mega4.1 software. Results All the nucleotide homologies of HA gene sequences in A/Hunan Kaifu/SWL4142/2009 ( H1N1 ), A/Hunan Changsha/SWL4346/2009 ( H1N1 ) and A/Hunan Furong/SWL4224/2009( H1N1 ) are 99% as compared with the novel influenza A( H1N1 ) virus strains of A/NewYork/3502/2009 ( H1N1 ), A/Shanghai/71T/2009 ( H1N1 ) and A/Chita/01/2009 ( H1N1 )The nucleotide homology of the 3 HA gene sequences are more than 99. 5% the same compared with the novel influenza A( H1N1 ) virus strain ( A/Sichuan/1/2009( H1N1 ) ) in China. Phylogenetic tree analysis reveals that 2009 novel influenza A(H1N1 ) viruses including 3 HA gene sequences of A/Hunan Kaifu/SWL4142/2009 ( H1 N1 ), A/Hunan Changsha/SWL4346/2009 ( H1N1 ), A/Hunan Furong/SWL4224/2009( H1N1 ) had a close evolutionary relationship with the swine H1 virus isolates in North America ( A/Swine/Indiana/P12439/00), but a distant evolutionary relationship with those human seasonal A( H1 N1 ) influenza virus and avian. After comparing with genes of A/Swine/Indiana/P12439/00, we found that the HA gene sequences of the 3 viruses isolated had 28,30 and 27 amino acids with mutation respectively, but only one (R53K) amino acids mutation at 21 important antigenic sites in the 3 viruses isolated. Multiple alignment of 364 HA genes sequences of novel influenza A ( H1N1 ) viruses in the world showed they had 119 nonconserved amino acids, 5 non-conserved position at important antigenic sites. Conclusions The HA gene sequences from 3 viruses isolated in this study and other influenza A ( H1N1 ) viruses might originate from swine A( H1N1 ) in North America by variation. The 3 HA gene sequences of viruses isolated have high homology as compared with the novel influenza A ( H1N1 ) virus strains worldwide, and the 3 HA gene sequences of viruses isolated are in stable condition as the vast majority of novel influenza A( H1N1 ) virus strains in the world.

Aim To explore the lipid-lowering mecha-nisms of curcumin from the molecular levels and pro-vide scientific basis for clinical development of lipid-lowering drugs.Methods Using oil red O staining and enzymic to determinate the levels of cholesterol in HepG2 cells.Moreover,uptaking of DiI-LDL was also measured.The expressions of mRNA and protein were detected by RT-Q-PCR and Western blot.Results The red lipid droplets and the levels of TC and FC sig-nificantly increased in HepG2 cells after treated with curcumin.The orange red fluorescence was higher than that of control.Curcumin could promote the expression levels of mRNA and protein of SREBP2 and LDLR, what′s more,curcumin could reduce the expression of the mature PCSK9 level and IDOL protein.Conclu-sion Curcumin accelerates LDL-C uptake probably via downregulating the expression of PCSK9 and IDOL in HepG2 cells.

Objective To determine whether a prophylactic tolterodine administration before surgical operation on non-urologic patients under general aneathesia can prevent the occurrence of catheter-related bladder discomfort (CRBD) ; and to assess patients’ tolerance to the symptoms as well as the impact on related consultation work of urologic surgeons.Methods One hundred and eighty cases of non-urology patients who need general aneathesia operations were divided into 2 groups:90 cases in tolterodine group and 90 in control group.The assessment of CRBD is categorized into 4 steps and statistics for adverse events ( dry mouth,dizzyness and facial flushing) was also conducted.A record of the patients’ needs for urologic surgical consultation during their reservation of catheter was also kept.SPSS 13.0 used in the statistical analysis of data in terms of X2 examination,where the divergence P ＜ 0.05 was regarded statisticly valid.Results 82 cases were followed up in the tolterodine group with a 24.4％ CRBD occurrence,which included 7.2％ shows moderate and severe symptoms,and there were also 23 cases with dry mouth ( 28.0％ ),4 cases with dizzyness (4.8％),13 cases with facial flussing ( 15.8％ ),and 1 case who needs further consultation (1.2％).In the 86 followed-up cases in control group,CRBD occurance rate was 54.7％,with 30.2％ showed moderate and severe symptoms,plus 2 cases suffered from severe consequences.Nine cases ( 10.5％) in control group requires further consultation ( X2 =19.499,P =0.000 ＜ 0.05 ).Conclusions A prophylactic tolterodine administration before surgery to the patients underwent general aneathesia can prevent the occurrence ofcatheter-related bladder discomfort (CRBD) and reduce the consultation work of urologic surgeons.Patients using tolterodine show a higher rate of adverse events,yet to which most patients can tolerate.

ObjectiveTo study the reasons and mechanism of cardiomyocyte apoptosis in chronic heart failure by using Losartan and to provide a theoretical basis for the treatment of chronic heart failure. Methods The models of chronic heart failure were produced by injecting Adriamycin and Losartan as intervention agents, the expression of apoptotic protein Bax, Bcl-2 and channel protein ERK1, JNK1 and P38MAPK were detected by immunohistochemistry and RT-PCR.Cardiomyocyte apoptosis and myocardial ultrastructure are detected by transmission electron microscopy and TUNEL staining.Results Compared with the model group of heart failure, after Losartan treatment, the ultra structure of myocardial cells were significantly improved, Apoptosis index was decreased significantly (P <0.01), The level of Bax and JNK1 decreased (Bax χ~2=6.6149, P=0.0078; JNK1 q=22.0156,P<0.01). However, the expressions of ERK1 and Bcl-2 were significantly increased (ERK1 q=15.3514,P<0.01;Bcl-2 χ~2=6.81,P=0.0074).Conclusion The effect of Losartan on chronic heart failure is related closely with the pathway of ERK1 and JNK1, and no p38 MAPK pathway.

Objective To investigate the impact of rapid eye movement (REM) sleep on the rat brain.Methods Microtubule-associated protein 2 (MAP2) and neurofilament (NF) were used as two neuronal cytoskeleton markers.The modified multiple platform method (MMPM) was used to deprived rats of sleep.The immunohistochemistry method and Western blot technique were used to detect the expression of MAP2 and NF,together with the electromicroscope study to show the neurodegeneration in rat cerebral cortex and hippocampus.Results The expression of MAP2 and NF in cerebral cortex and hippocampal neurons decreased after 5-day sleep deprivation.Some uhrastructural changes were seen in those areas as well,such as displacement of the nucleus,slightly swollen mitochondrial and endoplasmic reticulum,dissolution or condensation of sheaths of some axons.Neurons in cortex with degenerated ultrastrutrue accounted for 1.2% ,3.6% and 5.8% in tank control group,sleep deprivation 5 d group and sleep deprivation 7 d group respectively.Conclusion REM sleep deprivation can induce neurodegeneration in rat brain,but these morphological changes are tiny enough to be considered as reversible.

Aim To explore the effects of niacin on LDL-C uptake and metabolism in HepG2 cells,and to clarify the functions of niacin in lipid-lowering and slo-wing the atherosclerosis process,thus to provide a sci-entific basis for niacin as a lipid-lowering drug in clini-cal development.Methods Oil red O staining was used to observe HepG2 cells after lipid uptake.Enzy-matic method was used to determine the content of in-tracellular free cholesterol (FC)and total cholesterol (TC).The LDLR levels on the surface of cell mem-brane were detected by immunofluorescence flow cy-tometer.The mRNA and protein expressions of LDLR, SREBP2 and PCSK9 were analyzed by qPCR and Western blot.Results The results of oil red O staining showed that the rate of oil red O-positive cells and the number of red lipid droplets were significantly in-creased in niacin group than control group.Niacin sig-nificantly increased the levels of TC and FC in HepG2 cells(P <0.05 ).What’s more,niacin significantly upregulated the expression of LDLR and significantly downregulated the protein expression of PCSK9,while it had no effect on the expression of SREBP2.Conclu-sion Niacin accelerates LDL-C uptake probably via downregulating the expression of PCSK9 and reducing the degradation of LDLR protein in HepG2 cells.

Objective To explore the curative effect of fibular flap with limbs composite soft-tissue.Methods From February,2013 to February,2016,13 cases with body severe trauma patients were treated,which including 5 cases of upper limbs and 8 cases of lower limbs,and all existed bone defect,soft tissue defect and trunk vessel defect.Three cases with limbs distal non blood supply were emergency treated with debridment and flow-through fibular flap transplantation renovation,peroneal artery repairing defective blood vesscls to rcstorc limbs distal blood supply,fibular flap repairing bone defect,skin flap repairing soft tissue defect.The limb blood supply for other 10 cases were in good condition,but one case with main artery defect did the second phase of fibular flap transplantation and repaired defective blood vessels,bone and skin soft tissue synchronously according to wound condition.According to the postoperative observation for flap survival and appearance,X-ray films to observe fracture healing after 6 weeks,three months and 6 months of operation as well as evaluating limb function recovery,then analyzed the results.Results Flaps survived successfully for 11 cases,and flaps for the other 2 cases were partial necrosis.One Case was edge flap necrosis,heal scabby after dressing,and the other case was necrosis for 1/3 of the area,but the deep fascia survival,and the skin graft healing after dressing.One case with forearm rolling was in vascular crisis after operation,but tbe crisis was relieved after detection,and fingers blood supply was recovered.All the patients were followed up for 6 to 36 months(mean,14 months).All flaps were survived,fractures healed well and limbs distal blood supply was good.Bone healing time was 8 to 24 weeks,and patients with lower limbs injury could bear load after 3 to 8 months.Lower limbs restored walking function.Upper limbs and hands restored rotation function.Transplant flapshad good elasticity and satisfactory appearance.Conclusion Using fibular flap to repair defective blood vessels,bone and soft tissue synchronously,not only can rescue the limbs on the verge of amputation,but also can repair defective composite tissue and get a good prognosis.It is an effective method for open injuries severely treatment in clinic.

OBJECTIVETo observe the protective effect of metformin on the endothelial function and the mechanisms in rats with low-density lipoprotein (LDL) injection.

METHODSA single dose (4 mg/kg) of natural LDL was injected through the sublingual vein of rats to induce vascular endothelial dysfunction. Blood samples were then collected from the rats to detect the concentrations of malondialdehyde (MDA) and nitric oxide (NO), activity of superoxide dismutase (SOD) and serum lipid levels. The thoracic aorta of rats was obtained to assay acetylcholine (ACh)-induced endothelium-dependent relaxation (EDR) and sodium nitroprusside (SNP)-induced endothelium-independent relaxation. The effects of metformin pretreatment on the endothelial functions in the rats were investigated.

RESULTSA single-dose LDL significantly inhibited ACh-induced EDR without affecting SNP-induced endothelial-independent relaxation. The injection decreased serum NO and elevated serum MDA level, but had no effect on serum lipid level. Metformin markedly attenuated LDL-induced inhibition of EDR, serum MDA elevation, and serum NO reduction without affecting the serum lipid levels.

CONCLUSIONMetformin provides protection against vascular endothelial dysfunction induced by LDL in rats, the mechanism of which is probably associated with protection of endothelium-dependent relaxation factor and inhibition of the oxidative stress.

Animals ; Endothelium, Vascular ; drug effects ; physiopathology ; Endothelium-Dependent Relaxing Factors ; metabolism ; Lipoproteins, LDL ; administration & dosage ; Male ; Malondialdehyde ; blood ; Metformin ; pharmacology ; Nitric Oxide ; blood ; Oxidative Stress ; drug effects ; Rats ; Rats, Sprague-Dawley ; Superoxide Dismutase ; metabolism ; Vasodilation ; drug effects ; physiology

BACKGROUNDVisfatin, a visceral fat-derived adipocytokine, plays a significant physiological function in lipid metabolism. However, the precise function of visfatin and its regulation by thyroid hormones are still unknown. This study observed the plasma visfatin concentrations in subjects with hyperthyroidism and hypothyroidism in vivo, and investigated the possible regulation mechanism between visfatin and tri-iodothyronine (T3) in vitro as a further interpretation.

METHODSThe experiment in vivo included clinical subjects (57 patients with thyroid dysfunction and 29 euthyroid healthy volunteers) and an animal model (24 Wistar rats). All subjects were divided into hyperthyroidism, hypothyroidism and euthyroidism groups, with plasma thyroid hormones, thyrotropin, visfatin and triglyceride concentrations determined. Visfatin mRNA expression in visceral fat and liver of rats was detected by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR). The experiment in vitro studied 3T3-L1 cells and visfatin mRNA expression under nine different T3 concentrations (0, 0.1, 0.25, 0.5, 1, 5, 10, 20, 100 nmol/L) using quantitative real-time RT-PCR.

RESULTSClinical subjects and animal models showed elevated plasma visfatin concentrations in the hyperthyroidism group (20.466 ng/ml (15.263, 26.795 ng/ml) and (1209.164±165.292) ng/L) and hypothyroidism group (12.457 ng/ml (11.115, 15.454 ng/ml) and (1205.425±109.200) ng/L) compared to euthyroidism group (6.891 ng/ml (5.888, 8.803 ng/ml) and (926.650±54.002) ng/L, P<0.001). For animal models, visfatin mRNA expression in visceral fat in the hyperthyroidism and hypothyroidism groups increased about 3.33-fold and 1.98-fold compared to the euthyroidism group (P<0.001), which was positively correlated with plasma visfatin concentrations (r=0.713, P<0.001). However, no significant group difference (P>0.05) and correlation (r=0.121, P=0.572) was found in the liver. T3 induced a remarkable increase of visfatin mRNA expression in 3T3-L1 cells at low concentrations (0-0.5 nmol/L T3) followed by a sharp decrease at higher concentrations (0.5-100 nmol/L T3), with an inflection point at 0.5 nmol/L T3.

CONCLUSIONElevated circulating visfatin levels in subjects with hyperthyroidism and hypothyroidism are possibly due to an increase of visfatin mRNA expression in visceral fat, and a nonlinear regulation mechanism on visfatin mRNA expression under various T3 concentrations might be involved.

3T3-L1 Cells ; Adult ; Animals ; Female ; Humans ; Hyperthyroidism ; blood ; genetics ; metabolism ; Hypothyroidism ; blood ; genetics ; metabolism ; Male ; Mice ; Middle Aged ; Nicotinamide Phosphoribosyltransferase ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Triiodothyronine ; blood

Background and purpose:Adenosine triphosphate-binding cassette superfamily G member 2 (ABCG2), which has been found over-expressed in a variety of cancer cells, takes part in the drug resistance of cancer through effux of anticancer drugs. The purpose of this study was to investigate the mechanisms of human glioblastoma cells sensitivity to pyropheophorbide-a methyl ester (MPPa)-mediated photodynamic therapy (PDT) eradicating tumour cells and its relationship to ABCG2.Methods:U87 and A172 glioma cell lines in the logarithmic growth phase were selected and exposed to the treatment of MPPa-PDT and MPPa-PDT+fumitremorgin C (FTC) respectively. The cell viability was measured with the use of CCK-8 assay. The expression of ABCG2 was detected by Western blot. The intracellular contents of MPPa in each group without illumination were tested by lfow cytometry. Flow cytometry with AnnexinⅤ-FITC/PI double staining was used to detect the cell apoptotic rate. DCFH-DA staining was used to assess the generation of intracellular reactive oxygen species (ROS).Results:The MPPa-mediated PDT could eradicate A172 and U87 cancer cells in an energy-dependent manner. The light energy density in A172 was 8 times of that in U87 when the cell viability reached median lethal dose after MPPa-mediated PDT. The high expression of ABCG2 in A172 cells affected the accumulation of intracellular MPPa. Inhibition of ABCG2, not only could enhance the eradicating effect of MPPa-PDT on A172 cells, but also could increase the yield of ROS triggered by MPPa-PDT and the accumulation of intracellular MPPa.Conclusion:The human glioblastoma cell line A172 is insensitive to MPPa-mediated PDT. The mechanism may relate to ABCG2, which decreases the MPPa content in cancer cells through effux of MPPa, resulting in decline of cytotoxicity.