Objective To observe the correlation of serum levels of cystatin C (Cys C) and plasma N-terminal pro B-type natriuretic peptide (NT-pro BNP) with classification of cardiac function in elderly patients with chronic heart failure (CHF),and to explore its clinical significance.Methods 93 patients (aged ≥ 80 years) with CHF were considered as CHF group including degree Ⅱ,Ⅲ,Ⅳ (NYHA).44 patients (aged≥ 80 years) without CHF were selected as the control group.Cystatin C,NT-pro BNP levels and left ventricular ejection fraction (LVEF) were measured respectively,then their correlations were analyzed.Results There were significant differences in levels of CysC,NT-pro BNP and LVEF between the two groups (all P＜0.01).Along with the increasing of cardiac function classification,the Cys C and NT-pro BNP levels also increased,while LVEF decreased.The increments of Cys C and NT-pro BNP levels and decrements of LVEF had statistically significant differences between the two groups (all P＜ 0.01).Correlation analysis showed that,NT-pro BNP was positively correlated with age,creatine,blood urea nitrogen,Uric acid (r=0.18,0.46,0.29 and 0.14,all P＜0.01),and negatively correlated with the LVEF (r=-0.64,P＜0.01).Cys C was positively correlated with NT-pro BNP,age,creatine,blood urea nitrogen,Uric acid (r=0.65,0.30,0.51,0.41 and 0.23,all P＜0.01),and negatively correlated with LVEF (r=-0.76,P＜0.01).After adjusting for the interference factors,above correlations still existed.Conclusions Cys C concentration is significantly increased in very old patients with chronic heart failure,and it increases along with the increasing of the heart function classification and NT-pro BNP.The serum Cys C concentration has a certain reference value for the evaluation of cardiac function in very old patients with chronic heart failure.

The levels of serum lipids, lipoproteins and apoproteins were determined in 36 age over 90 year long-living people, 43 patients with old myocardial infarction (OMI) and 56 normal subjects of the age 40 to 64 years old. The resutts showed that: the ratioes of HDL-c/TC, HDL-c/LDL-c and ApoA_1/ApoB_(100) of the long-living people were higher than that of the patients with OMI; the levels of TC, TG, LDL-c, VLDL-c, ApoB_(100) and ApoA_1 in longevity group were lower than that of OMI group; no significant difference of HDL-c level was found between longevity and OMI group. Compared with normal group, the serum concentration of ApoA_1/ApoB_(100) ratio, TG and VLDL-c in the long-living group were obviously higher, and the serum concentration of ApoB_(100), ApoA_1 and HDL-c significantly lower. No significant differences were found in the levels of TC, LDL-c, and in the ratioes of HDL-c/TC and HDL-c/LDL-c between longevity and normal group. Stepwise regression analysis showed that the serum ApoB_(100) concentration and ApoA_1/ApoB_(100) ratio was the most important factor of longevity. It was suggested that the internal balance of serum lipids and lipoprotein metabolism in long-living people was good co-ordination, and the lower ApoB_(100) level and higher ApoA_1/ApoB_(100) ratio were probably parameters of longevity.

Aim DNA vaccinating BALB/c mice with the constructed recombinant plasmid, pcDNA3, containing ROP1 gene from Toxoplasma gondii to observe the effect on the production of the cytokines, IFN- γ、 IL - 2 , and NO in the immunized mice. Methods Large-scale preparation of plasmid DNA by alkaline lysis,the DNA were injected through muscles of left leg in each mouse at the dosage of 100μg. A booster vaccine was given at the same dosage after two weeks. Control groups were injected with pcDNA3 blank plasmid and normal saline respectively. After 30,50 and 70 days of the booster injection, following tests were carried out 3 times separately :the serum IFN-γand IL-2 were detected by sandwich ELISA;the NO was detected by enzyme assay. Results The 3 times detected results of IFN-γγ、IL-2 and NO were significant higher in the vaccinated group than that of in control groups and the contents were increased with the vaccinated time prolonged. Conclusion The IFN - γγ、 IL - 2 and NO were produced by vaccinating BALB/c mice with the recombinant plasmid, pcROP1.

AIM:To investigate the relationship between the thrombin-activatable fibrinolysis inhibitor (TAFI) level in plasma and atherosclerosis, the TAFI level in plasma and blood fat and blood clotting index.METHODS: Totally 40 healthy male Wistar rats were randomly divided into 4 groups (n=10), control group, high lipid group, high lipid +vitamin D3 overload group, and high lipid +vitamin D3+endothelium injure group by treating the animals with normal diet, high lipid, high lipid+ vitamin D overload, and high lipid+vitamin D overload+endothelium injury, respectively, for inducing three stages of AS in rats. Then, the total cholesterol (TC), total triglyeride (TG), high density lipoprotein (HDL), low density lipoprotein (LDL), and prothrombin time (PT), activated partial thromboplasin time (APTT), fibrinogen (Fib), the activity of TAFI were measured.RESULTS: TC, TG, LDL-C, Fib and the activity level of TAFI in plasma in three model groups increased gradually compared with the control (P

Objective To construct and identify the screening vector pUC18-EGFP,using EGFP as an indicator. Methods The EGFP gene was prepared by PCR and cloned into pUC18 resulting the vector pUC18-EGFP. Then DNA fragment was inserted into the MCS of pUC18-EGFP to test its practicability based on green fluorescence. Results The pUC18-EGFP was confirmed correctly by restriction enzyme analyses. The pUC18-EGFP was used to select recombinants. The green strains on the plate were confirmed by restriction enzyme and DNA analyses,which were E.coli harboring recombinants. Conclusion The screening vector PUCl8-EGFP was constructed successfully. Thus,we can select the positive clones on plates based on the green fluorescence of EGFP.

Objective　To construct the eukaryotic expression recombinant plasmid, pcIFN-γ, as a genetic adjuvant and observe the immune responses elicited by pcDNA3-rhoptry protein 1 （pc-ROP1） combined with pcIFN-γ against Toxoplasma gondii (T. gondii) infection in mice. Methods　A fragment of the IFN-γ gene was directly inserted into the pcDNA3 plasmid and identified by two restriction endonucleases digestion. pcIFN and pcROP1 DNA was injected into the left leg muscle of mice at a dosage of 100*!μg, and a booster vaccination was given at the same dosage after two weeks. Control groups were injected with pcDNA3 blank plasmid or normal saline. At 30, 50 and 70 days after booster injection, kinetic tests were carried out: MTT assay for the proliferation response of T lymphocyte cells and the activity of NK cells, sandwich ABC-ELISA for the determination of IFN-γ, IL-2 and IL-10; a serum enzymetic aassay for nitric oxide (NO) in sera and ELISA for the titer of IgG antibody in sera. Results　The recombinant plasmid, pcIFN-γ was constructed. The proliferation response of spleen T lymph cells, NK cell killing activity, and serum levels of IFN-γ, IL-2 and NO in mice injected with pcROP1 and pcIFN-γ were higher than in those injected with pcROP1 alone. There was no difference in IgG antibody levels between the two groups. Conclusion　The genetic adjuvant, pcIFN-γ, could enhance the cellular immune response induced by DNA vaccine of pcROP1 in mice against Toxoplasma gondii infection.

Objective　To observe the immune responses elicited in BALB/c mice by a DNA vaccine. A gene encoding rhoptry protein 1 (ROP1) from Toxoplasma gondii (T. gondii) was cloned into vector pcDNA3. Methods　Amplifyied gene fragments coding for ROP1 from the genomic DNA of T.gondii ZS2 were inserted into cloning vector, pUC18, and sub-cloned into pcDNA3. Mice were injected at a dosage of 100?μg recombinant plasmid DNA by intramuscular injection and boosted after 2 weeks. pcDNA3 and normal saline were used as control. 30, 50 and 70 days after the second immunization, NK cell activity, T lymphocyte proliferation and sub-clusters and serum IgG antibody were assayed. Results　The specific gene fragment coding for ROP1 was amplified and a pcROP1 recombinant was constructed. At 30 days after immunization, the spleens of the mice were obviously enlarged evidently. NKC activity and the proliferation of spleen T lymphocytes seen on MTT assay were higher in pcROP1 group than in the controls. The number of CD4+ T cells exhibited no obvious increase compared with that of the control, but CD8+ T cells were obviously increased (P<0.05). At 90 days after vaccination, the titer of IgG antibody in the serum of vaccinated mice was positive (1∶100). Conclusion　pcROP1 was constructed and it could elicit both cellular and humoral immune responses in immunized mice.

OBJECTIVETo characterize the immune response induced by SAG1 encoding plasmid combined with IL-2 gene adjuvant in mice and to assess the protective effect of this vaccination against toxoplasmosis.

METHODSMice were co-injected intramuscularly with plasmid encoding Toxoplasma gondii SAG1 plus murine IL-2 expression vector at a dose of 100 microg. Booster immunizations were employed 2 more times at 3-week interval. As controls, mice were inoculated with PBS or empty plasmid pcDNA3. Humoral and cellular responses were assayed using ELISA for the determination of Ab, Ab isotype and IFN-gamma, as well as IL-4. To detect the integration and dissemination of DNA in the injected mice, PCR and in situ hybridization were performed. All mice were then infected with highly virulent RH tachyzoites of Toxoplasma gondii intraperitoneally.

RESULTSSignificant increases in specific IgG levels were observed in mice after immunization three times with SAG1 expression plasmid. With respect to the IgG isotype, co-inoculation of IL-2 expression plasmid enhanced the level of IgG2a and the production of IFN-gamma. Challenging mice by vaccinating with combined plasmids with RH tachyzoites resulted in prolonged survival.

CONCLUSIONHumoral and cytokine responses elicited by SAG1 DNA immunization can be modulated by co-inoculation with IL-2 expression plasmid. The use of DNA vaccine in combination with an appropriate cytokine gene to prevent T. gondii infection warrants further investigation.

Animals ; Antibodies, Protozoan ; blood ; Antigens, Protozoan ; Cytokines ; biosynthesis ; Female ; Immunization ; Immunoglobulin G ; blood ; classification ; Interleukin-2 ; genetics ; Mice ; Protozoan Proteins ; genetics ; Protozoan Vaccines ; immunology ; Toxoplasma ; immunology ; Toxoplasmosis, Animal ; prevention & control ; Vaccines, DNA ; immunology

Objective To evaluate retrograde transpopliteal access for femoral-popliteal artery total occlusion with blind puncture.Methods Clinical data of 22 cases admitted from Sep 2014 to June 2016 undergoing endovascular treatment of the femoral-politeal artery occlusion with transpopliteal artery retrograde access by blind puncture were analyzed.Results A total of 22 patients underwent retrograde popliteal access with blind puncture after antegrade attempts failed.Puncture above the knee was performed in 18 cases and below the knee in 4 cases.The average increase of ABI was 0.57.Hematoma of puncture site was observed in 2 patients,other complications included pneumonia in 1 case and renal insufficiency in 2 cases.The mean follow-up time was (13 ± 5)months.Restenosis occurred in 8 patiens(36.4％)during the follow-up time.The primary patency was (86.4 ± 0.07) ％ at 6 months and (70.7-± 0.12) ％ at 12 months.There was no major amputation rate and mortality during the follow-up.Conclusions Retrograde transpopliteal access for femoral-popliteal artery occlusion with blind puncture is safe and effective.

OBJECTIVETo evaluate the immune responses induced by experimental DNA construct encoding Toxoplasma gondii (T. gondii) surface antigen1 (SAG1) and rhoptry protein 1 (ROP1) in mice as a hybrid gene.

METHODSTruncated SAG1 and ROP1 DNA fragments were amplified using polymerase chain reaction (PCR) and inserted into pEGFP-N3 vector to construct recombinant plasmid pSAG1-ROP1. NIH3T3 mammalian cells were transiently transfected with the DNA construct. Female BALB/c mice were given three intramuscular injections of 10 micro g plasmid DNA entrapped in liposome. Four weeks after the final booster injection, blood samples were collected and subjected to enzyme-linked immuno sorbent assay (ELISA) to investigate humoral and cell-mediated immune responses. Reversal transcript-polymerase chain reaction (RT-PCR) was used to evaluate the transcription of inoculated DNA-liposome complex in the injected site. Dot-blot hybridization was employed in order to detect whether or not the injected DNA was incorporated into the genomic DNA of the immunized mice.

RESULTSGreen fluorescence was observed in pSAG1-ROP1-transfected cells. Western blot analysis showed antibody recognition of the expressed SAG1-ROP1 was between 58 kDa and 75 kDa. No expression was observed in blank control plasmid-transfected cells. The sera of immunized mice exhibited antibodies to T. gondii tachyzoites and primarily interferon-gamma and interlukin-2. RT-PCR showed that the duration of transcribed inoculated liposome entrapped DNA in the injected muscular tissue was at least ten days post the first injection. Dot-blot hybridization revealed that the presence of foreign DNA in the splenocytes and peripheral blood leukocytes was transient and that no foreign DNA had inserted into the genomic DNA of mice immunized with pSAG1-ROP1.

CONCLUSIONSImmunization with a liposome-encapsulated DNA construct encoding the T. gondii SAG1 and ROP1 can induce humoral and cell-mediated immune responses.

Animals ; Antibody Formation ; Antigens, Protozoan ; genetics ; DNA ; Enzyme-Linked Immunosorbent Assay ; Female ; Genetic Vectors ; Immunity, Cellular ; Immunization ; Liposomes ; Membrane Proteins ; genetics ; Mice ; Mice, Inbred BALB C ; Protozoan Proteins ; genetics ; Toxoplasma ; genetics ; immunology ; Transfection