[Objective]To study the effect of T cell groups of CD4+,CD8+,CD4+/CD8+,IL-4,IFN-? in the hypoimmunitymice induced by hydrocortisone to discuss its function of regulating the immune function and CD4+TH cell’s inferior change of group,have offered the experiment basis for clinical practice of TAF.[Method] FACS measured the T lymphocyte phenotype;the level of cytokine IFN-r and IL-4 was detected by ELISA.[Result] TAF can raise CD4+T lymphocyte of hypoimmunitymice,suppress CD8+ T lymphocyte expression,raise CD4+/CD8+ ratio;And can make the level of serum IFN-r obviously enhanced in hypoimmunitymice,and the level of IL-4 was very low.[Conclusion] TAF can rebuild immune function of hypoimmunitymice,promote organism Th0 to split up to Th1,maintain the advantage state of TH1,and strengthen the function of cellular mediated immunoresponse.

Objective To study the expression of estrogen receptor beta (ER ?) in the hypothalamic paraventricular nucleus (PVN) of maternal mice during pregnancy and lactation. Methods Nickel ammonium sulfate intensified immunocytochemical method was employed to investigate the expression of ER ? in the postnatal developing PVN of female mice. Results ER ? immunopositive materials were predominantly localized in the magnocellular division of PVN, and sparse positive cells were found in the parvocellular division. Most of the positive materials were found in the whole cell nuclei, but no obvious cytoplasma or process immunopositive cells were detected. In the pregnant female brain, generally, the ER ? level was lower. The lowest levels of expression were found at mid pregnancy, and then peaked at peripartum (from gestational days 18 to postpartum day 1), followed by another decrease to normal adult level from postpartum day 4. Conclusion The above results suggest that during pregnancy and lactation, ER ? may be predominantly involved in the PVN regulation of parturitional process.

Objective To investigate the correlation between meteorological factors and the incidence of acute cardiovascular disease.Methods The number of calls for ambulance in case of cardiovascular disease between 2006 to 2010 was obtained from Beijing 120 emergency center and the data of meteorological factor during for the same period from Beijing Meteorological professional were collected,and then the impact of the weather factors of different seasons of weather factors on the incidence of cardiovascular disease was ere analyzed by time-stratified case-crossover design.Results The average daily incidence cases of cardiovascular disease in four seasons average daily were 5.61,4.55,5.05 and 6.03 respectively.The association between temperature Temperature and the incidence of acute cardiovascular disease was were non-linear relationship changes in spring and autumn seasons.Atmosphere Air pressure and the incidence were negatively correlated in the winter.The relative humidity of airand the incidence were related in the spring,summer,autumn.Wind and the incidence were related in the spring,summer,but negative correlation in the autumn,winter.Conclusions The incidences of acute cardiovascular disease are higher more in winter and spring than in summer and autumn.The incidences of cardiovascular disease are related with temperature,relative humidity,wind speed and other meteorological factors.,The influence of meteorological factors on the incidence of the disease delays to occur after changes in weather.

Objective To explore the correlation between quantitative value of joint bone scan by single photon emission computed tomography (SPECT) and serum bone metabolic markers in patients with active rheumatoid arthritis (RA).Method Clinical data of 60 newly diagnosed RA patients were retrospectively collected in Department of Rheumatology and Immunology from May 2013 to December 2014,including 28 cases with medical history less than 6 months and 32 from 6 months to 2 years.Serum Cterminal telopeptide of type Ⅰ collgen(CTX-Ⅰ)and N-terminal propeptide of type Ⅰ collagen(PINP) were tested by chemiluminesence.SPECT whole body bone and target joint scan before treatment was done.Thirty-nine healthy subjects as control group received ultrasound,electrocardiogram,X-ray,and whole body bone imaging examination.Quantitative values of joint imaging were calculated for the statistical analysis.Result Demographic data between RA group and control group were comparable (P ＞ 0.05),including sample,sex,age and BMI.The joint SPECT value,CTX-Ⅰ and PINP levels were all significantly higher than those of control group (P ＜ 0.01),which were 6.48 ± 1.98 versus 3.73 ± 1.t6;(0.66 ± 0.37) mg/L versus (0.58 ±0.21) mg/L;(46.35 ±28.15) mg/L versus (30.47 ± 13.75) mg/L respectively.Joint SPECT values had positive correlations with serum CTX-Ⅰ levels in all RA patients,as well as PINP in patients with disease duration 6mon-2years.And the according correlation coefficients were 0.513,0.495,0.402 (P ＜ 0.05).But SPECT value had no correlation with CTX-Ⅰ (P =0.081) in patients with disease duration less than 6 mon.The correlation coefficient was 0.336.Conclusion SPECT imaging quantitative values were positively correlated with serum bone metabolic parameters.Thus SPECT imaging alone or combined with bone markers are helpful in diagnosing active RA.

Objective To observe the effects of treadmill training on the recovery of neurological function and the expression of HSP70 and C-MYC in the brains of rats with focal cerebral ischemia. Methods Forty-two male adult Sprague-Dawley rats were randomly divided into a sham group ( n =6), a model group (n =18) and a treadmill exercise group (n=18). Focal cerebral ischemia was induced by right middle cerebral artery occlusion (MCAO) in the model group and exercise group using a modified version of Longa's method. The rats in the treadmill exercise group were given treadmill training 6 d per week for 2 weeks after 24 h of MCAO. By contrast, the rats in the sham group and the model group were reared in standard cages. Before the rats were sacrificed at the 3rd, 7th and 14th d after MCAO, their neurological functions were tested using modified neurological severity scores ( mNSS) , and the mRNA and protein levels of HSP70 and C-MYC were detected using a reverse transcriptase-polymerase chain reaction (RT-PCR) , immunohistochemistry and Western blotting. Results Neurological function in the exercise group at the 7th and 14th days after MCAO had improved significantly compared with the control and model groups.The mRNA and protein levels of HSP70 and C-MYC were significantly upregulated at the 7th and 14th days. Conclusions Treadmill training can improve neurological function by upregulating the expression of HSP70 and C-MYC in the ischemic brain after MCAO.

Objective To study the effect of hypoxia on the expression of interleukin 6 (IL 6) in cocultured pulmonary arterial smooth muscle cells (PASMC) Methods Rat PASMC were cocultured with rat lung microvascular endothelial cells, and randomly divided into 5 groups: normal group (N), 2 h hypoxia (H2), 6 h hypoxia (H6), 12 h hypoxia (H12), and 24 h hypoxia (H24) The expression level of IL 6 mRNA in PASMC was detected with RT PCR, and the activity of IL 6 in the supernatant with radioimmunoassay Results The expression level of IL 6 mRNA increased in PASMC in H2, reached the highest in H6, and decreased in H12, but still higher than that of N The changes of the IL 6 activity in the supernatant as well as IL 6 mRNA expression in the cells were in a time dependent manner Conclusion Hypoxia can enhance the expression of IL 6 in cocultured PASMC And it may activate others genes which regulate the hypoxic response of PASMC and signal transduction

Objective To express the anti-IL-1βscfv and soluble TNF receptor 1 (sTNFR1),and analyze their bio-activities.Methods sTNFR1 was obtained by RT-PCR from the total RNA of HeLa cells,and fused with IL-1βscfv by the hinge fragment of IgG molecule.The fusion gene IL-1scfv:TNFR1 was cloned into the expression vector pET27b(+).The fusion protein was expressed and purified from inclusion bodies.Results The ELISA analysis showed that the fusion protein could bind hIL-1β and hTNF-α respectively in a dose-dependent manner,indicating that scfv and sTNFR in the fusion protein can form the correct spatial configuration.The dolt-blot analysis showed that the fusion protein could concurrently bind with hIL-1β and hTNF-α,indicating that the combination of the two parts of the fusion protein does not influence each other for binding to their target molecules.The bioactivity assay showed that the fusion protein could inhibit both the cytotoxicity of hTNF-α on L929 cells and hIL-1β-induced proliferation of L929 cells,indicating that the fusion protein has the ability to neutralize hTNF-α and hIL-1β.Conclusion A bispecific fusion protein IL-1scfv:TNFR1 was successfully constructed.The fusion protein has the ability to inhibit the biological activity of hTNF-α and hIL-1β,and provides a drug candidate for the treatment of rheumatoid arthritis.

The aim of the study is to establish a platform to deliver therapeutic proteins into target cells through a polyarginine-based cell penetrating peptide. To facilitate the expression of therapeutic proteins, a pSUMO (Small Ubiquitin-like Modifier)-R9-EGFP (enhanced green fluorescence protein) prokaryotic expression vector was constructed. After induction, the fusion protein SUMO-R9-EGFP was efficiently expressed. To validate the cell penetrating ability of the fusion protein, HepG2 cells were incubated with the purified R9-EGFP or EGFP protein as control, internalization of the fluorescent proteins was examined by either flow cytometry or confocal microscopy. The result obtained by flow cytometry showed that the R9-EGFP fusion protein could efficiently penetrate into the HepG2 cells in a dose and time-dependent manner. In contrast, the fluorescence was barely detected in the HepG2 cells incubated with EGFP control. The fluorescence intensity of the R9-EGFP treated cells reached plateau phase after 1.5 h. The result obtained by confocal microscopy shows that R9-EGFP efficiently entered into the HepG2 cells and was exclusively located in the cytoplasm, whereas, no fluorescence was detected in the cells incubated with the EGFP control. The heparin inhibition experiment showed that heparin could inhibit penetrating effect of the R9-EGFP protein by about 50%, suggesting that the penetrating ability of the fusion protein is heparin-dependent. In summary, the study has established a platform to deliver therapeutic proteins into target cells through a polyarginine-based penetrating peptide.
Cell-Penetrating Peptides ; biosynthesis ; genetics ; pharmacology ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; biosynthesis ; genetics ; Hep G2 Cells ; Humans ; Peptides ; genetics ; metabolism ; Protein Transport ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; pharmacology

The aim of this project is to establish a fibroblast growth factor-21 (FGF-21) signaling pathway targeted cell model, for screening a class of FGF-21 receptor agonists as anti-diabetic candidates. FGF-21 requires beta klotho transmembrane proteins as co-receptor for the activation of tyrosine kinase FGF receptor (FGFR) signaling, thereby activating a series of intracellular signaling pathways and regulating gene transcription for glucose metabolism. Firstly a recombinant plasmid expressing co-receptor beta klotho and EGFP reporter genes was constructed. After introducing the recombinant plasmid into package cells, the cell culture supernatant was used to infect 3T3-L1 cells, which were then screened for stably expressing beta klotho gene. Administration of FGF-21 increased the expression of GLUT1 and stimulated GLUT1-mediated glucose uptake. This novel cell model can be conveniently used in high-throughput drug screening of FGF-21 or FGF-21 analogues.