螺丝命名细则.doc

项 目： 螺丝命名规则编号： 制定人： 制定日期：版本：页数： 审核人：审核日期：颁发部门： 一，说明 该规则依据美制、国际标准、德制、日制、英制、国标要求制定。二，命名明细4 X 10 PW A H C （+） 螺丝直径螺丝长度螺丝头型B：球面圆柱头； C：圆柱头； F（K）：沉头； H：六角头；HW：六角头带垫圈； O：半沉头； P：平元头； R：半元头；PW：平元头带垫圈； T：大扁头； V：蘑菇头；螺丝牙型A：自攻尖尾（日标第1种）疏 AB：自攻尖尾（日标第4种）密；B：自攻平尾（日标第2种）疏；C：自攻平尾（日标第3种）密；P：双丝牙 HL：高低牙 U：菠萝牙纹；T：自攻平尾切脚 AT：自攻丝尖尾切脚 M：机械牙；BTT：B型三角牙 CCT：C型三角牙 PTT：P型三角牙 STT：S型三角牙热处理 H：有热处理 N：没热处理表面处理Zn：白锌 C：彩锌 B：蓝锌 F：黑锌 O：氧化黑 Ni：镍Cu：青铜Br：红铜 P：磷化 备注（+）：十字槽 （-）：一字槽 （T）：菊花槽 （H）：内六角 （PZ）：米字槽 （+-）：+-槽 （Y）：Y型槽 （H）：H型槽 （L）：止退花齿 （WIS）：单活动弹垫圈 （WIF）：单活动平垫圈 （WIT）：单活动外齿垫圈 （W2SF）：双活动平弹垫圈 （ W=6mm）：垫圈外径等于6mm （SUS）：不锈钢（Cu）：黄铜（Br）：红铜（8.8）：8.8级螺丝（10.9）：10.9级螺丝 （12.9）：12.9级螺丝 （R）：其它注释三，附图The authors would like to thank Johns Hopkins University for the TC-1 cells. This work was supportedby a National Health Research Institutes intramural grant (IV-103-PP-22) and grants from the NationalScience Council, which were awarded to Y.C. Song (NSC 99-2321-B-400-004-MY3) and S.J. Liu (NSC103-2321-B-400-008).Author ContributionsY.C.S. and S.J.L. designed the studies. Y.C.S. performed the research and analyzed the data. Y.C.S. andS.J.L. wrote the manuscript.Additional InformationC57BL/6 mice were immunized subcutaneously (s.c.)once with 1 g of peptide mixed with or without 10 g of CpG adjuvant. After one week, splenocytes wereharvested, and the response of IFN- -secreting cells was determined by ELISPOT after 48 h of peptidestimulation. Briefly, 2 × 105 splenocytes were incubated with 1 g/ml irrelevant peptide or RAH peptidein an anti-IFN- -coated polyvinylidene fluoride (PVDF) plate for 48 h. After incubation, the cells wereremoved, and a biotinylated anti-IFN- Ab (eBioscience, San Diego, CA, USA) was added to each well.The plates were incubated at 37 °C for 2 h. Following the addition of the avidin-HRP reagent (eBioscience,CA, USA), the assay was developed using a 3-amine-9-ethyl carbazole (AEC; Sigma-Aldrich, MO,USA) staining solution. The reaction was stopped after 46 min by placing the plate under tap water.The spots were counted using an ELISPOT reader (Cellular Technology Ltd., Shaker Heights, OH, USA).For RAH-specific T cell staining, spleens were harvested seven days after the immunizations, andRAH-specific CD8+ T cells were detected by tetramer staining using a PE-labeled RAH tetramer(Beckman Coulter, CA, USA) and a FITC-labeled anti-CD8 monoclonal antibody (mAb) (eBioscience,CA, USA). The stained RAH-specific CD8+ T cells were analyzed by flow cytometry.Supplementary information accompanies this paper at Competing financial interests: The authors declare no competing financial interests.How to cite this article: Song, Y.-C. and Liu, S.-J. A TLR9 agonist enhances the anti-tumor immunityof peptide and lipopeptide vaccines via different mechanisms. Sci. Rep. 5, 12578; doi: 10.1038/srep12578 (2015).This work is licensed under a Creative Commons Attribution 4.0 International License.The images or other third party material in this article are included in the articles CreativeCommons license, unless indicated otherwise in the credit line; if the material is not included underthe Creative Commons license, users will need to obtain permission from the license holder toreproduce the material. To view a copy of this license, visit expression of anti-apoptotic molecules such as the BCL-2 familymembers BCL-XL and CASP8 and FADD-like apoptosis regulator(CFLAR, best known as cFLIP), thereby allowing CTLs to surviveand reach neoplastic of various TLR agonists promotes the immunogenicity of DC/malignant cell fusions through the upregulation of IL-12.14,18In this setting, we used a de from Coriolus versicolor (PSK, which operates as a TLR2 agonist)and lyophilized preparations of a low-virulence strain (Su) ofStreptococcus pyogenes (OK-432, which acts as a TLR4 agonist),both of which can be produced as good manufacturing practice(GMP)-grade agents and have been previously used in the clinic asbiological response modifiers.18,19 Of note, DC/cancer cell fusionsactivated in the presence of both TLR2 and TLR4 agonists,but not DC/malignant cell fusions that were left unstimulatedor were exposed to either TLR agonist alone, overcame theimmunosuppressive activity of tumor-derived molecules suchas transforming growth factor 1 (TGF1). In particular,TLR2/4-activated DCs (or the corresponding fusions): (1) exhibitincreased expression levels of MHC class II molecules and CD86on the cell surface; (2) manifest an improved fusion efficacy;(3) produce elevated levels of IL-12; (4) simultaneously activateCD4+ and CD8+ T cells, which secrete high levels of interferon (IFN); (5) potently induce antigen-specific CTL activity;and (6) manifest a superior efficacy in inhibiting the generationof CD4+CD25+FOXP3+ Tregs.20 Nonetheless, when DC/cancercell fusions are generated with neoplastic cells producing extremelyhigh levels of TGF1, they inhibit the activity of CTLs in vitro.Therefore, incorporating the simultaneous activation of multipleTLRs and the blockade of immunosuppressive that are intrinsicallyproduced by DC/neoplastic cell fusions may significantly enhancethe therapeutic potential of this approach.Improving the Immunogenicity of Malignant CellsMost, if not all, malignant cells secrete multipleimmunosuppressive mediators such as TGFmolecules normally inhibit the initiation of efficient CTLresponses,21 the microenvironment of malignant cells used forthe generation of DC/cancer cell fusions immunostimulatory. Several strategies to inhibit the production ofimmunosuppressive factors by cancer cells have been developed,including the administration of neutralizing antibodies22 and smallchemical inhibitors,23 as well as the transfection of specific smallinterferingRNAs (siRNAs)24 or constructs coding for a solublevariant of the TGF receptor.25 Also heat-shock proteins (HSPs),which have recently been implicated in the immunogenicity ofapoptotic and necrotic cells, might constitute effective adjuvantfor boosting the efficacy of DC/neoplastic cell fusions.26,27 HSPsgenerally operate as chaperons for a wide panel of peptides,including antigenic peptides, and HSP/peptide complexes notonly can be efficiently taken up by DCs through specific receptors,but also can be presented in molecules the DC surface.28 We have previously reported thatTLR2-stimulated DCs fused with heat-treated cancer cells areimmunogenic, as demonstrated by: (1) the upregulation of multipleHSPs, MHC class I and II molecules, TAAs, CD80, CD86, CD83,and IL-12; (2) their ability producing high levels of IFN; and (3) the capacity to efficientlyelicited antigen-specific CTL activity.26 More recently, we havedemonstrated that the secretion of TGF1, IL-10 and VEGRfrom whole cancer cells is significantly limited upon exposure topharmaceutical grade ethanol, a maneuver that does not reduce theevels of MHC class I molecules and TAAs on the cell surface.27Moreover, ethanol, employed at concentrations that affect tumorgrowth, promoted the upregulation of HSPs. HSPs exposed bycancer cells can be recognized by DCs via TLR4, facilitating theiractivation and promoting antigen processing and presentation.29Of note, malignant cells that undergo immunogenic apoptosisectopically expose the Ca2+-binding chaperone calreticulin (CRT)on the cell surface, allowing TAAs to efficiently traffic to theDC antigen-presenting compartment.30 Moreover, high-mobilitygroup box 1 (HMGB1) passively released from dying neoplasticcells can stimulate antigen processing and presentation in DCs viaa TLR4-dependent signaling pathway.31,32 Therefore, the exposureof CRT and the release of HMGB1 by ethanol-treated malignantcells enhance the immunogenicity of DC/cancer cell fusions.27Importantly, fusions involving DCs and ethanol-treated cancercells activate T cells to produce high levels of IFN, boosting theelicitation of antigen-specific CTL response in vitro.27 In addition,HSP70-peptide complexes derived from DC/cancer cell fusionsappear to possess superior immunogenic properties as comparedwith similar complexes obtain from neoplastic cells.33Synergistic Effects of Fusions Generated