ISG15, the first ubiquitin-like molecule identified two decades ago, is encoded by interferon stimulated gene 15 ( ISG 15), where its robust expression can be induced by viral infections or interferon treatments. ISG 15 conjugate to other proteins as the ubiquitin and was found to be involved in innate immune response. However, the functions of ISG15 modification remained unclear. We cloned bovine ISG15(bisg15) into a prokaryotic expression vector pET28a( + ) with a His-tag to generate a soluble form of bISG15 fusion protein, and purified with Ni-NTA Sepharose chromatography. The purified protein was concentrated and used to immune Balb/c mice to raise the antiserum, which could specifically recognize bISG15 expressed in eukaryotic cells by Western blot analysis. The concentrated bISG15 protein and its antiserum were then used to establish an in vitro bISG15 modification system. Our studies have demonstrated that cellular proteins could be conjugated to bISG15 with this system.

Virus infection or interferon can stimulate robust expression of the protein ISG15 that is encoded by interferon stimulated gene 15, which was the first unbiquitin-like molecule identified two decades ago. While ubiquitin and its many important functions have been well established, the functions of ISG15 and its post-translational conjugation are still largely unknown . Recently, some specific enzymes have been identified to be involved in the ISG15 modification system, suggests that ISG15 and its modification system play important roles in the innate immune response and regulation of interferon signaling. The history of ISG15 discovery and its biochemical characterization were briefly introduced. Then such topics as the ISG15 gene expression and the ISG15 modification will be focued on, and finally summarize new findings which have implications for ISG15 and its modification system in immunology and interferon signal transduction were summarized.

Objective To develop prototype foamy virus (PFV) detection method by loop-mediated isothermal amplification. Methods Three pairs of primers targeting core region of PFV integrase were designed in this study and Bst DNA polymerase was used to amplify target sequence at 63℃. The system was established with all the conditions optimized. Results The method was established with the plasmid containing target sequence as the template. This method could specifically detect PFV infectious clone, no crossreaction was observed with human immunodeficieney virus infectious clone, bovine immunodefieiency virus infectious clone and bovine foamy virus infectious clone as templates. The detection capability of this system was 50 copy, one order more sensitive than PCR. The amplification could be finished in 15 min and human genomic DNA did not adversely affect the amplification efficiency. Conclusion The PFV detection method by loop-mediated isothermal amplification was established and it had potential usefulness in PFV detection.

The ubiquitin-like modifier bISG15 is an antiviral protein found in fetal bovine lung (FBL) cells.Bovine Herpesvirus 1(BHV-1),which is a viral pathogen of cattle,can infect FBL cells and induce cytopathic effects.Real-time PCR assays showed that BHV- 1 's infection could repress the basal or inducible transcription of bISG15 in FBL cells.It demonstrates that this repression effect depends on BHV-1 viral infection and new protein synthesis.Our previous work showed that bIRF-3 was the key factor in the stimulation of bISG 15 in FBL cells,so the effect of BHV-1 viral protein on bIRF-3 activating the promoter of bISG15 was confirmed.The luciferase assay showed the BHV-1 viral protein bICP0 inhibited the activation of bISG15 promoter stimulated by bIRF-3.Taken together,our work suggested that BHV-I had some molecular mechanism to resist the cellular bISG15'santiviral functions.

We described the first results of a quantitative ultra performance liquid chromatographytandem mass spectrometry method for a novel antimicrobial peptide (phylloseptin,PSN-1).Chromatographic separation was accomplished on a Waters bridged ethyl hybrid (BEH) C18 (50 mm × 2.1 mm,1.7 μm) column with acetonitrile-water (25∶75,v/v) as isocratic mobile phase.Mass spectrometry detection was performed in the positive electrospray ionization mode and by monitoring of the transitions at m/z 679.6/120,509.6/120 (PSN-1) and m/z 340.7/165 (Thymopentin,IS).Protein precipitation was investigated and the recovery was satisfactory (above 82％).The method was shown to be reproducible and reliable with intra-day precision below 5.3％,inter-day precision below 14.2％,and linear range from 0.02 to 2 μg/mL with r＞0.994.The method was successfully applied to a pharmacokinetic study of PSN-1 in rats after intravenous administration.

0 05) CONCLUSION:UH is effective and safe in treatment of congestive heart failure and its ARDs are rare It is worth popularizing in clinical use

We described the first results of a quantitative ultra performance liquid chromatographytandem mass spectrometry method for a novel antimicrobial peptide （phylloseptin, PSN-1）. Chromatographic separation was accomplished on a Waters bridged ethyl hybrid （BEH） C18 （50mm× 2.1 mm, 1.7 μm） column with acetonitrile-water （25：75, v/v） as isocratic mobile phase. Mass spectrometry detection was performed in the positive electrospray ionization mode and by monitoring of the transitions at m/z 679.6/120, 509.6/120 （PSN-1） and m/z 340.7/165 （Thymopentin, IS）. Protein precipitation was investigated and the recovery was satisfactory （above 82%）. The method was shown to be reproducible and reliable with intra-day precision below 5.3%, inter-day precision below 14.2%, and linear range from 0.02 to 2 lag/mL with r〉0.994. The method was successfully applied to a pharmacokinetic study of PSN-1 in rats after intravenous administration.

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.

Besides its function as a pathogenicity determinant, the Tombusvirus P19 also serves as a suppressor of RNA interference (RNAi) by sequestering intracellular small RNAs such as the small interfering RNAs (siRNAs) and microRNAs (miRNAs). However, the effect of P19 on mammalian cells has not been evaluated before. A human embryonic kidney 293 cell line that stably expressed p19 (HEK293-p19) was generated. Flow cytometric analysis revealed that over-expression of P19 caused a significant accumulation of G2/M phase cells. Cell proliferation assays demonstrated a reduced DNA replication and cell growth in HEK293-p19 cells. Moreover, p19 altered the expression profiles of a number of cell cycle regulators in HEK293 cells, such as upregulafion of cyclin A1, CDK2, CDK4, CDK6, p18, cyclin D2, p19INK4d and E2F1, and downregulation of p15, cyclin A2, cyclin B1 and cyclin E1. Thus, the data strongly indicate that p19 might influence multiple G2/M regulators to cause G2/M arrest.

Objective:To explore the etiology and treatment of one case of bilateral mandibular second molar impaction with paradental cyst, and to provide a reference for its diagnosis and treatment. Methods:Root canal treatment of the left mandibular first molar of the patient was performed before operation.The left mandibular second molar of the patient was removed;the residual dental follicle, the granulation tissue and the cyst wall were stroken off under local anesthesia.The diamond ball was used to polish the wound cavity and sharp bone edge, and to mill the distal apical part of left mandibular first molar.The tissue removed during the procedure was used for the pathological examination.Results:The X-ray image showed that the bilateral mandibular second molar was impacted with the left mandibular first molar root's absorption, and there was a clear round-like density reduction zone around the second molar crown.The pathologic result was paradental cyst.Conclusion:Dental impaction complicated with paradental cyst could occur in other tooth position except for the third molar.Its diagnosis should be combined with the clinical manifestations, the pathologic manifestations and the medical imaging.Multidisciplinary consultation is in favor of its diagnosis and treatment.