Objective To analyze the relationship between the expression of protection of telomeres 1 (POT1) and the pathogenesis of acute myeloid leukemia (AML). Methods 62 patients with de novo AML (case group) and 10 patients with iron deficiency anemia (control group) were enrolled in this study. The quantitative real-time polymerase chain reaction (PCR) and Western blot were used to detect the expression of POT1 in AML patients. Results There were 62 de novo AML patients, including 2 cases M1, 14 cases M2, 12 cases M3, 14 cases M4, 17 cases M5, 2 cases M6 and 1 case AML without classification. According to the risk stratification, high risk group (24 cases), medium risk group (22 cases) and low risk group (16 cases) were divided. Compared with that in the controls, POT1 expression levels in patients with AML were significantly decreased both in mRNA and protein level (P< 0.05). The relative expression levels of POT1 mRNA and protein in patients with M2, M4 and M5 were significantly lower than those in the controls (P< 0.05). The expression levels of POT1 in high risk group, medium risk group and low risk group were significantly decreased than those in the controls (P<0.05). Compared with that in the controls, The relative POT1 mRNA expression was significantly decreased in M3 patients (P< 0.05), but not in protein level. POT1 protein expression was showed both in the cytoplasm and nucleus. There was no significant difference of the expression of POT1 protein between cytoplasm and nucleus (P> 0.05). Conclusions POT1 may be involved in the pathogenesis of AML. POT1 protein expresses in both cytoplasm and nucleus, and the regulatory mechanism may be related to the telomere length.

Objective To isolate specific humanized anti-D-dimer scFv(single chain Fv) antibody from scFv phage libraries. Methods Isolate anti-D-dimer positive clones from Tomlinson I + J phage libraries by three rounds of panuing, then sequence monoclonal genes by bideoxy-mediated chain termination and express soluble scFv antibody; Pick out anti-D-dimer antibodies with high specificity and affinity by ELISA.Results After three rounds of selection from human scFv phage libraries Tomlinson I and J, 38 monclonal specific anti-D-dimer scFv fragments were selected. By polyclonal and monoclonal phage ELISA and gene sequencing, 20 different full-length monoclonal scFv phages were identified, the result of soluble scFv ELISA showed that 20 full-length monoclonal scFv were expressed smoothly. According to the result of soluble scFv ELISA, in 5 scFv antibodies with high value of A450 selected, 3 scFv antibody fragments showed high specific and affinity. Conclusion Antibody phage display was an effective, rapid method to isolate anti-D-dimer antibodies with high specificity and affinity.

In order to improve the accuracy for quantitating the bovine foamy virus(BFV)in vitro,we developed a baby hamster kidney cell(BHK)-21-derived indicator cell line containing a plasmid that encodes the firefly luciferase driven by the BFV long terminal repeat promoter(LTR,from -7 to 1012). The BFV titer could be determined by detecting the luciferase expression since the viral trans-activator BTas protein activates the promoter activity of the LTR. One clone,designated BFVL,was selected from ten neomycin-resistant clones.BFVL showed a specific and inducible dose-and time-dependent luciferase activity in response to BFV infection.Although the changes in luciferase activity of BFVL peaked at 84 h post infection,it was possible to differentiate infected and uninfected cells at 48 h post infection. A linear relationship was established between the multiplicity of infection(MOI)of BFV and the activated ratio of luciferase expression in BFVL. Moreover,the sensitivity of the BFVL-based assay for detecting infectious BFV was 10,000 times higher than the conventional CPE-based assay at 48 h post infection. These findings suggest that the BFVL-based assay is rapid,easy,sensitive,quantitative and specific for detection of BFV infection.

ObjectiveTo investigate the effects of ziprasidone on the behavior and the expression of pERK1/2 in posttraumatic stress disorder(PTSD) model rats.Methods 24 adult male SD rats weighing (200 ±20) g were randomly divided into four groups (n =6):control group,single prolonged stress and foot shock (SPS&S) group,ziprasidone group and ziprasidone + U0126 group.The fear response to environment,high alertness,and anxiety & depression behavior of rats were tested by the open field,elevated plus-maze,and the expression of pERK1/2 was measured by Western blot.ResultsIn open field test(OFT),the SPS&S group( (76.23 ± 54.76) cm for horizontal motion distance,(4.60 ± 1.14) for the number of entering central region) showed significant difference compared with control group ( (343.77 ± 74.22 ) cm,( 12.40 ± 3.36 ) ) or ziprasidone group ( ( 274.98± 83.56) cm,( 12.00 ± 2.92) ) (P ＜ 0.01 ),but showed no significant difference with ziprasidone + U0126 group ( ( 138.14 ± 41.98) cm,(5.00 ± 1.58) ) (P ＞ 0.05 ).The results of elevated plus maze (EPM) were in accordance with the results of OFT.The expression of pERK1/2 in SPS&S group and ziprasidone + U0126 group showed significant decrease when compared with control group or ziprasidone group (P ＜ 0.01 ).ConclusionZiprasidone can obviously improve fear response to environment,high alterness and anxiety & depression behavior of rats,and these effects of ziprasidone may be carried out by up-regulation the expression of pERK1/2.

The Borf1 protein is encoded by an immediate-early gene of the bovine foamy virus (BFV) and plays a key role in the viral life cycle. Borf1 is a DNA binding protein which can transactivate both the long terminal repeat (LTR) and the internal promoter (IP) of BFV by specifically binding to the transactivation responsive element (TRE). To analyze the subcellular localization of Borf1 during the BFV life cycle, this gene was cloned into a prokaryotic expression vector and expressed in a soluble form. After the purification and immunization, we raised the mouse anti-Borf1 serum with a high titer based on ELISA results. Western blot analysis showed that the antiserum could specifically recognize the Borf1 protein that was expressed in 293T cells. With this specific serum, we revealed the nuclear and cytoplasmic localization of Borf1 in HeLa cells that was transfected with Borf1. Moreover, the immuno-fluorescence assay also showed that the localization of Borf1 during the infection and transfection of BFV was identical.

Objective To study the anticoagulation therapy of lower molecular weight heparin in the treatment of patients with acute pancreatitis.Methods Seventy-three patients with acute pancreatitis were divided into anticoagulation group(n=38)and control group(n=35).The serological indexes and prognosis of patients were detected.Results Anticoagulation treatment with lower molecular weight heparin significantly decreased the white blood cell count,increased the oxygen partial pressure in arterial blood,shoaened the duration of hospitalization,and reduced the aggravation rate,secondary operation rate and mortality of patients with acute pancreatitis.Conclusions Anticoagulation treatment with lower molecular weight heparin is safe,effective and can improve the prognosis of patients with acute pancreatitis.

Viruses (e.g. Human immunodeficiency virus, Human simplex virus and Prototype foamy virus) are obligate intracellular parasites and therefore depend on the cellular machinery for cellular trafficking. Bovine foamy virus (BFV) is a member of the Spumaretrovirinae subfamily of Retroviruses, however, details of its cellular trafficking remain unknown. In this study, we cloned the BFV gag gene into prokaryotic expression vector pET28a and purified the denaturalized Gag protein. The protein was used to immunize BALB/c mouse to produce antiserum, which could specifically recognize the BFV Gag protein in BFV-infected cells through western blot assay. Additionally, these results demonstrated that both the optimal and suboptimal cleavage of Gag protein occur in BFV-infected cells. Subsequently, the Gag antiserum was used to investigate subcellular localization of BFV. In immunofluorescence microscopy assays, colocalization microtubules (MTs) and assembling viral particles were clearly observed, which implied that BFV may transport along cellular MTs in host cells. Furthermore, MTs-depolymerizing assay indicated MTs were required for the efficient replication of BFV. In conclusion, our study suggests that BFV has evolved the mechanism to hijack the cellular cytoskeleton for its replication.

Objective The aim of the experiment is to make an intracranial aneurysm model in canine.Methods A digital tube was made based on raw magnetic resonance images of the human intracraaial carotid artery.Then 6 tubes were made in the 3 D rapid prototyping machine and coated with silicone.Finally we isolated the common carotid arteries of 6 canines and made them go through the tubes and anastomosed them end-to-side to get the aneurysm model.Six stents were implanted after one week.Results Six aneurysm models were successfully made in canines.The parent artery had similar geometrv of the human carotid siphon.All the aneurysms and parent arteries were patent in one week's follow-up.One canine died of excessive anesthesia after stentingr Two vaseular models kept patent in one month without stenosis.The other 3 had some stenosis on the bends of the vessel.Conclusions The aneurysm model in tIle experiment has high flexibility and reliability.The model provides an effective tool for research and testing neurovascular devices.It's also a useful device to train the neuroradiologists and interventional physicians.

Objective To characterize the effect of the ~1H-MRS scan parametem, including the type of coil, TE,NSA and VOI, on shimming, water suppression, spectral signal to noise ratio(SNR)and the stability of the baseline of liver in vivo. Methods ~1H-MRS of liver in vivo was performed prospectively on GE Signa Excite HD 3.0 T system in 46 volunteers. Point-resolred spectroscopy(PRESS)sequence with built-in body coil and eight-channel torso phased-array coils was applied. After the localized scan,the first PRESS sequence with a TR of 1500 ms,TE of 30 ms. VOI of 2 cm×2 cm×2 cm and NSA of 64 times was acquired using eight-channel torso phased-array coils.(The first PRESS sequence parametem was deemed as A).Then,the sequence was repeated with alteration of the three parameters including the type of coil,TE and size of VOI.(Changed parameters deem as B).The data were analyzed with the Wilcoxon matched pairs signed test.0 mark:A is similar to B,1 mark:A better than B,-1 mark:A worse than B.Results SNR(-1 mark 0 pair,0 mark 1 pair,1 mark 10 pair,Z=-3.162,P=0.002)was better in data(n=11)with eight-channel torso phased-array coils(A)than that with the built-in body coil(B),but the autoshimming line width with eight-channel torso phased-array coils were inferior to those with built-in body coil (-1 mark 8 pair,0 mark 2 pair,1 mark 1 pair,Z=-2.511,P=0.012).SNR was better in data(n=13)with TE of 30 ms(A)than that at the sequence with TE of 90 ms(B)(-1 mark 2 pair,0 mark 0 pair,1 mark 11 pair,Z=-2.496,P=0.013).whereas baseline stability was,poorer in the former(-1 mark 10 pair,0 mark 2 pair,1 mark 1 pair,Z=-2.333,P=0.020).SNR at the sequence(n=10)with VOI of 2 cm×2 cm×3 cm(B)was better(-1 mark 6 pair,0 mark 4 pair,1 mark 0 pair,Z=-2.449,P=0.014)than that at the sequence(n=29)with VOI of 2 cm ×2 cm × 2 cm(A),but poorer(-1 mark 0 pair,0 mark 5 pair,1 mark 5 pair,Z=-2.041,P=0.041)auto-shimming line width was shown. By comparison the sequences with NSA of 128 times(B)and NSA of 64 times(A),the former could provide better spectrum SNR(-1 mark 21 pair,0 mark 7 pair,1 mark 1 pair,Z=-4.264,P=0.000).Conclusion It is more easy to achieve a homogeneous bo magnetic field using a small size of VOI and builtin body coli.The sequence with VOI of 2 cm ×2 cm ×3 cm.NSA of 128 times is recommended for clinical use. Increase VOI and NSA are helpful to improve SNR. Longer TE is helpful to improve baseline stability.

Bel1, a transactivator of prototype foamy virus (PFV), plays pivotal roles in the replication of PFV. Previous studies have shown that Bel1 bears a nuclear localization signal (NLS), but its amino acid sequence remains unclear and the corresponding importins have not been identified. In this report, we inserted various fragments of Bel1 into an EGFP-GST fusion protein and investigated their subcellular localization by fluorescence microscopy. We found that the 215PRQKRPR221 fragment could direct nuclear localization, which accords with the consensus sequence K(K/R)X(K/R) of monopartite NLS. Point mutation experiments revealed that K218, R219, and R221 are essential for the nuclear localization of Bel1. The results of the GST-pulldown showed that the Bel1 fragment with residues 215-223, which bears the NLS, interacts with KPNA1, KPNA6, and KPNA7. This result suggests that KPNA1, KPNA6, and KPNA7 maybe involved in Bel1 nuclear translocation.
Cell Line ; Cell Nucleus ; genetics ; metabolism ; virology ; Humans ; Nuclear Localization Signals ; genetics ; metabolism ; Protein Binding ; Protein Transport ; Retroviridae Infections ; genetics ; metabolism ; virology ; Retroviridae Proteins ; chemistry ; genetics ; metabolism ; Spumavirus ; chemistry ; genetics ; physiology ; Trans-Activators ; chemistry ; genetics ; metabolism ; alpha Karyopherins ; genetics ; metabolism