· Vascular endothelial growth factor is indispensable inducing factor in retinalangiogenesis. After the retinal neovascularization of proliferative diabetic retinopathy ( PDR ) patients, it can cause fibrovascular membrane formation, epiretinal membrane fibrosis increased, resulting in traction retinal detachment with further aggravate the condition. The recent research suggests that cytokines promote fibroblast proliferation, movement, adhesion, and secretion of extracellular matrix functions in the diabetic state of the environment changes to profibrogenic state, resulting in the accumulation and fibrosis of extracellular matrix. This paper reviewed the status quo of the correlation between vascular endothelial growth factor and fibrosis-related cytokine.

Objective To construct the recombinant plasmid of fusion protein containing respiratory syncytial virus (RSV) cytotoxicity T lymphocyte (CTL) epitope M2:81-95 and heat shock protein (HSP) 70L1, and to investigate its immunogenicity after prokaryotic expression. Methods HSP70L1 gone was cloned from SMMC7721 cells. The M2:81-95 gene fragment was amplified by polymerase chain reaction (PCR) method. Plasmid pET-HSP70L1-M2 : 81-95 (pET-HSP70L1-M2) was constructed, identified and transferred into E. coli BL21 (DE3). Expression of HSP70L1-M2 : 81-95(HSP70L1-M2) was induced by isopropy-β-D-thiogalaetosidc (IPTG). The expressed protein was purified by affinity chromatography and renatured by gradient dialysis. The BALB/c mice were immunized with this fusion protein. IgG antibodies and the subtypes were detected by enzyme-linked immunosorbent assay (ELISA), and CTL responses were determined by methyl thiazolyl tetrazolium (MTT). Results The recombinant plasmid pET-HSP70L1-M2 was successfully constructed. The fusion protein HSP70L1-M2 was expressed in E. coll. The purified protein induced strong RSV-and CTL epitope-specific CTL responses and high titer of protein specific lgG antibody 4.87±0.35. The subtypes were IgG1 (5.53±0.28) and lgG2a (4.40±0.21) and IgG1/ IgG2a ratio was balanced. The titers of lgG, IgG1 and IgG2a in PBS control group were 0.33±0.17, 0.51±0.21 and 0, respectively, which werc significantly lower than those in immunized group (t = 3.512, 3.681, 5.856; P<0.05). Conclusions The recombinant plasmid pET-HSP70L1-M2 is successfully constructed and the fusion protein is expressed and purified. HSP70L1-M2 induced strong RSV-and CTL epitope-specific CTL responses and mixed T helper cell (Th)1/Th2 response in BALB/c mice.

Objective: To investigate the regulation of respiratory syncytial virus CTL epitope fused with G protein antigen fragment G1 to the specific immunoresponses. Methods: The recombinant plasmid pET-DsbA-G1 or pET-DsbA-G1F/M2 was transferred into E.coli BL21(DE3) and the fusion protein DsbA-G1F/M2 or DsbA-G1 was expressed.The expressing product was induced and purified by affinity chromatography. The two proteins were used to immunized BALB/c mice i.p, respectively. Serum and spleen cells were collected regularly. RSV-specific CTL responses were measured by MTT, IgG and IgG1 and IgG2a antibodies by ELISA, neutralizing antibodies by plaque reduction assay. Results: The recombinant proteins were expressed successfully and the purity was over 90% after purified by affinity chromatography. The protein DsbA-G1F/M2 induced significant RSV-specific CTL responses, while DsbA-G1 without CTL epitope did not induce detctable CTL responses. Strong IgG antibody responses were elicited,indicated by both. IgG1 and IgG2a titers induced by DsbA-G1F/M2, while only IgG1 was induced by DsbA-G1.The ratio of IgG1/IgG2a was downregulated significantly. Both antigens induced high level of neutralizing antibodies, but the latter was a little lower. Conclusion: DsbA-G1F/M2, the fusion protein of a CTL epitope and G protein fragment G1 can induce both cellular immunity and humoral immunity. The activation of CTLs downregulates the ratio of IgG1/IgG2a.The more balanced immunoresponse is advantageous for improving safety of the candidate vaccine.

Objective To investigate the effects of bilateral sternocleidomastoideus contraction on neck flexion and extension. Methods The electromyograms (EMG) signals of left sternocleidomastoideus of 24 healthy persons were recorded respectively when neck relaxation and flexion in the supine position, and neck relaxation and extension in the prone position. The biomechanical measurement and analysis were done using anatomical specimens and models. Results The sternocleidomastoideus registered electrical resting potential when neck relaxation in the supine position, and when neck relaxation and extension in the prone position; the motor unit action potential of sternocleidomastoideus raised the type of mixed phase when neck flexion in the supine position. Conclusion The contractions of bilateral sternocleidomastoideus can lead to the head and neck flexion while neck joints are not fixed, and the contractions of bilateral sternocleidomastoideus can lead to the head extension under the state of fixing neck joints below atlanto-occipital.

Objective: To investigate the effect of RNA interference (RNAi) targeting AKT1 and PI3K P85 on the proliferation and invasion of breast carcinoma MCF-7 cells. Methods: The recombinant adenovirus expression vector, which contained short hairpin RNA (shRNA) targeting open reading frames of AKT1 and PI3K P85 (rAd5-siAKT1-siPI3K), was transfected into human breast carcinoma MCF-7 cells. AKT1 and PI3K P85 mRNA and protein expressions were detected by real-time PCR and Western blotting analysis. The expressions of PCNA, cyclinD1, and P53 were also detected by Western blotting analysis. The proliferation and apoptosis of MCF-7 cells were measured by MTT, flow cytometry and 2-dementinal and 3-dementional matrigel assay. Results: Recombinant adenovirus vector rAd5-siAKT1-siPI3K dramatically down-regulated AKT1 and PI3K P85 mRNA and protein expressions in MCF-7 cells; the downstream factors PCNA and cyclin D1 were also down-regulated, while P53 was up-regulated. Growth of MCF-7 cells was inhibited by over 50% in rAd5-siAKT1-siPI3K group as measured by MTT assay, and cell cycle was arrested in G_1/G_0 phase compared with untransfected and rAd5-siCtrl transfected groups. Cell growth on matrigel matrix showed normal cell shapes, while the cells in rAd5-siAKT1-siPI3K transfected group were detached from the matrix or grew in scattered clustering patterns, forming only small aggregates. Conclusion: shRNA targeting AKT1 and PI3K P85 can significantly down-regulate the expression of AKT1 and PI3K P85 in breast carcinoma MCF-7 cells, and inhibit the growth of MCF-7 cells in vitro.

Objective To investigate the expression of CXCR4 in cultured human dental pulp cells (HDPC) in vitro and the corresponding ligand SDF-1α level of HDPC supernatants stimulated by lipopolysaccharide(LPS)and tumor necrosis factor-α(TNF-α),and to explore the role of SDF-1αon the proliferation and the migration of HDPC.Methods The expression of CXCR4 in HDPC was detected by immunocytochemistry technique and indirect immunofluorescence technique.The culture supernatants of HDPC were collected after HDPC had been simulated by LPS and TNF-α of difierent concentrations for 48h and then the SDF-1α level was assayed by quantitative sandwich ELISA.Meanwhile,the effects of recombinant human SDF-1α(rhSDF-1α)on the proliferation and the migration of HDPC at different concentrations were observed by MTT and Boyden Chamber Assay.Results CXCR4 was expressed in cytomembrane of HDPC and SDF-1α was secreted into their normal cell supematants with a concentration of(4513.55±962.92)ng/L. The secretion of SDF-1α was both significantly decreased by stimulation with LPS and TNF-α(P＜0.05).In addition,rhSDF-1α stimulated the HDPC proliferation at the concentrations of 50,100,200μg/L(P＜0.01)and increased the chemotactic migration of HDPC significantly after 9h's incubation with the concentrations of 50,100μg/L(P＜0.05).Conclusions SDF-1α accelerated the proliferation and the migration of HDPC which expressed CXCR4.SDF-1-CXCR4 axis may play a role in repair of pulp injury.

OBJECTIVE: To select specific DNA aptamer for determining ketamine by FluMag-SELEX. METHODS: Based on magnetic beads with tosyl surface modification as solid carrier and ketamine as target, a random ssDNA library with total length of 78 bp in vitro was compounded. After 13 rounds screening, DNA cloning and sequencing were done. Primary and secondary, structures were analyzed. The affinity, specificity and Kd values of selected aptamer were measured by monitoring the fluorescence intensity. RESULTS: Two ssDNA aptamers (Apt#4 and Apt#8) were successfully selected with high and specific abilities to bind ketamine as target with Kd value of 0.59 and 0.66 μmol/L. The prediction of secondary structure was main stem-loop and G-tetramer. The stem was the basis of stability of aptamer's structure. And loop and G-tetramer was the key of specific binding of ketamine. CONCLUSION FluMag-SELEX can greatly improve the selection efficiency of the aptamer, obtain the ketamine-binding DNA aptamer, and develop a new method for rapid detection of ketamine.
Aptamers, Nucleotide/metabolism* ; DNA ; DNA, Single-Stranded/genetics* ; In Vitro Techniques ; Ketamine/metabolism* ; Oligonucleotides ; SELEX Aptamer Technique/methods*

Objective The Rex shunt has been used to treat children with extrahepatic portal hypertension by creating a venous bypass between the superior mesenteric vein and the left portal vein to circumvent the extrahepatic portal venous obstruction.This retrospective study aimed to analyse our results in the use of this novel approach.Methods 52 patients with cavernous transformation and obstructed main portal veins were treated by Rex shunts.Results The age of children was 1.4 ～ 12 year,the mean age was 3.7 years.The patients were followed up from 1.5 to 5 years.In 48 patients,there was no recurrence of gastrointestinal bleeding after surgery making an efficacy rate of 92.3％.In the recurrence group,the postoperative venous pressure in the superior mesenteric vein was (27.6 ± 3.2) cmH2O (1 cmH2O =0.098kPa) which was significantly higher than the non-recurrence group (23.5 ± 3.1)cmH2O.The difference between the pre-and post-Rex shunting was significantly lower in the recurrence group (5.7 ± 1.8)cmH2O than the non-recurrence group (11.7 ± 3.3) cmH2 O,P ＜ 0.05.Thus,a low reduction in postoperative pressure was an early manifestation of poor prognosis.Conclusions The Rex shunt was safe and efficacious.The degree of reduction in postoperative venous pressure in the superior mesenteric vein could be used to predict recurrence of gastrointestinal bleeding.

Objective To study the state of erythrocyte immune function in neonates with hyperbilirubinemia,and analyze the influence of various clinical status on erythrocyte immune function.Methods Fifty-two neonates with hyperbilirubinemia were enrolled and 104 healthy neonates as the control group.The adherence rate of complement 3b-receptor on the surface of red blood cell(RBC-C3bRR) and the immune complex adherence rate of red blood cell(RBC-ICR) were detected with erythrocyte saccha-romycete rosettet test.Results 1.The level of RBC-C3bRR in neonates with hyperbilirubinemia was lower than that in control group,and the level of RBC-ICR in neonates with hyperbilirubinemia was higher than that of control group(Pa0.05).3.Comparing the neonates with unconjugated bilirubin of different concentrations,there were significant difference in RBC-ICR(Pa0.05).4.There were positive correlation between RBC-ICR and bilirubin,unconjugated bilirubin in the neonates(Pa0.05).Conclusion Erythrocyte immune function in neonates with hyperbilirubinemia is obviously lower than that of control group and it is influenced by the concentratron of bilirubin and the time of phototherapy.