A novel coronavirus, severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV), has been identified as the causal agent of SARS. Spike (S) protein is a major structural glycoprotein of the SARS virus ...A novel coronavirus, severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV), has been identified as the causal agent of SARS. Spike (S) protein is a major structural glycoprotein of the SARS virus and a potential target for SARS-specific cell-mediated immune responses. A panel of S protein-derived peptides was tested for their binding affinity to HLA-A*0201 molecules. Peptides with high affinity for HLA-A*0201 were then assessed for their capacity to elicit specific immune responses mediated by cytotoxic T lymphocytes (CTLs) both in vivo, in HLA-A2.1/K b transgenic mice, and in vitro, from peripheral blood lymphocytes (PBLs) harvested from healthy HLA-A2.1(+) donors. SARS-CoV protein-derived peptide-1 (SSp-1 RLNEVAKNL), induced peptide-specific CTLs both in vivo (transgenic mice) and in vitro (human PBLs), which specifically released interferon-gamma (IFN-γ) upon stimulation with SSp-1-pulsed autologous dendritic cells (DCs) or T2 cells. SSp-1-specific CTLs also lysed major histocompatibility complex (MHC)-matched tumor cell lines engineered to express S proteins. HLA-A*0201-SSp-1 tetramer staining revealed the presence of significant populations of SSp-1-specific CTLs in SSp-1-induced CD8+ T cells. We propose that the newly identified epitope SSp-1 will help in the characterization of virus control mechanisms and immunopathology in SARS-CoV infection, and may be relevant to the development of immunotherapeutic approaches for SARS.展开更多
目的预测并筛选免疫反应性较强的四跨膜蛋白超家族成员1(transmembrane 4 L6 family member 1,TM4SF1)人类白细胞抗原A2(human leukocyte antigen A2,HLA-A2)限制性T淋巴细胞(cytotoxic lymphocyte,CTL)表位肽。方法联合4种软件(BIMAS、...目的预测并筛选免疫反应性较强的四跨膜蛋白超家族成员1(transmembrane 4 L6 family member 1,TM4SF1)人类白细胞抗原A2(human leukocyte antigen A2,HLA-A2)限制性T淋巴细胞(cytotoxic lymphocyte,CTL)表位肽。方法联合4种软件(BIMAS、SYFPEITHI、IEDB、PROPRED I)对TM4SF1的HLA-A2限制性CTL表位进行预测,并采用预培养法酶联免疫斑点法(enzyme-linked immunospot assy,ELISOPT)、直接法ELISOPT对所测得表位肽的免疫反应性进行鉴定。结果 4种不同软件共预测出10条表位多肽,对其中4条(P1、P2、P8、P10)进行免疫反应性验证。多肽活化CTL能力结果显示,预培养法ELISPOT产生斑点形成细胞(spvt forming cell,SFC)的净值T明显高于直接法ELISPOT:[(阳性对照肽:322±8 vs 169±22,P<0.05)、(多肽P1:114±10 vs 39±7,P<0.05)、(多肽P10:156±31 vs 52±8,P<0.05)]。单个活化CTL细胞分泌INF-γ因子的水平检测结果显示,预培养法ELISPOT产生斑点的平均大小明显大于直接法ELISPOT[(阳性对照肽:21.91±2.45 vs 13.80±1.76,P<0.05)、(多肽P1:12.90±0.88 vs 8.31±1.40,P<0.05)、(多肽P10:17.50±3.85 vs 11.96±0.61,P<0.05)]。表位多肽P1、P10均具有免疫原性,P10的免疫活性更高。结论预培养法ELISPOT检测多肽的免疫反应性较直接法ELISPOT灵敏性更高,多种软件联合ELISPOT预培养法可有效筛选出免疫反应性更强的卵巢癌相关抗原TM4SF1 HLA-A2限制性CTL优势表位,其中P10的免疫活性最高。展开更多
Human immunodeficiency virus type-1 （HIV-1）-specific dendritic cell （DC） vaccines have been used in clinical trials. However, they have been found to only induce some degree of immune responses in these studies. W...Human immunodeficiency virus type-1 （HIV-1）-specific dendritic cell （DC） vaccines have been used in clinical trials. However, they have been found to only induce some degree of immune responses in these studies. We previously demonstrated that the HIV-1 Gag-specific Gag-Texo vaccine stimulated Gag-specific effector CD8＋ cytotoxic T lymphocyte （CTL） responses, leading to completely protective, but very limited, therapeutic immunity. In this study, we constructed a recombinant adenoviral vector, adenovirus （AdV）4-1BBL, which expressed mouse 4-1BB ligand （4-1BBL）, and generated transgenic 4-1BBL-engineered OVA-Texo/4.1BSL and Gag-Texo/4.1BSL vaccines by transfecting ovalbumin （OVA）-Texo and Gag-＇rexo cells with AdV4.1BBL, respectively. We demonstrate that the OVA-specific OVA-Texo/4.ZSSL vaccine stimulates more efficient OVA-specific CTL responses （3.26%） compared to OVA-Texo-activated responses （1.98%） in wild-type C57BIJ6 mice and the control OVA-TeXO/Nu, vaccine without transgenic 4-1BBL expression, leading to enhanced therapeutic immunity against 6-day established OVA-expressing B16 melanoma BL6-1OovA cells. OVA-Texo/4.1BBL-stimulated CTLs, which have a CD44＋CD62Lhigh IL-7R＋ phenotype, are likely memory CTL precursors, demonstrating prolonged survival and enhanced differentiation into memory CTLs with functional recall responses and long-term immunity against BL6-1OovA melanoma. In addition, we demonstrate that OVA-Texo/4_ZBBL-Stimulated CTLs up- and downregulate the expression of anti-apoptosis （Bcl2110, Naipl, No13, Pak7 and Tnfrsfllb） and pro-apoptosis （Casp12, Trp63 and Trp73） genes, respectively, by RT2 Profiler PCR array analysis. Importantly, the Gag-specific Gag-Texo/4.1BBL vaccine also stimulates more efficient Gag-specific therapeutic and long-term immunity against HLA-A2/Gag-expressing B 16 melanoma BL6-1OGag/A2 cells than the control Gag-TeXO/NuH vaccine in transgenic HLA-A2 mice. Taken together, our novel Gag-Texo/4-ZBBL vaccine, which is capable of stimulating potent Gag-specific therapeutic and long-term immunity, may represent a new immunotherapeutic vaccine for controlling HIV-1 infection.展开更多
文摘A novel coronavirus, severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV), has been identified as the causal agent of SARS. Spike (S) protein is a major structural glycoprotein of the SARS virus and a potential target for SARS-specific cell-mediated immune responses. A panel of S protein-derived peptides was tested for their binding affinity to HLA-A*0201 molecules. Peptides with high affinity for HLA-A*0201 were then assessed for their capacity to elicit specific immune responses mediated by cytotoxic T lymphocytes (CTLs) both in vivo, in HLA-A2.1/K b transgenic mice, and in vitro, from peripheral blood lymphocytes (PBLs) harvested from healthy HLA-A2.1(+) donors. SARS-CoV protein-derived peptide-1 (SSp-1 RLNEVAKNL), induced peptide-specific CTLs both in vivo (transgenic mice) and in vitro (human PBLs), which specifically released interferon-gamma (IFN-γ) upon stimulation with SSp-1-pulsed autologous dendritic cells (DCs) or T2 cells. SSp-1-specific CTLs also lysed major histocompatibility complex (MHC)-matched tumor cell lines engineered to express S proteins. HLA-A*0201-SSp-1 tetramer staining revealed the presence of significant populations of SSp-1-specific CTLs in SSp-1-induced CD8+ T cells. We propose that the newly identified epitope SSp-1 will help in the characterization of virus control mechanisms and immunopathology in SARS-CoV infection, and may be relevant to the development of immunotherapeutic approaches for SARS.
文摘目的预测并筛选免疫反应性较强的四跨膜蛋白超家族成员1(transmembrane 4 L6 family member 1,TM4SF1)人类白细胞抗原A2(human leukocyte antigen A2,HLA-A2)限制性T淋巴细胞(cytotoxic lymphocyte,CTL)表位肽。方法联合4种软件(BIMAS、SYFPEITHI、IEDB、PROPRED I)对TM4SF1的HLA-A2限制性CTL表位进行预测,并采用预培养法酶联免疫斑点法(enzyme-linked immunospot assy,ELISOPT)、直接法ELISOPT对所测得表位肽的免疫反应性进行鉴定。结果 4种不同软件共预测出10条表位多肽,对其中4条(P1、P2、P8、P10)进行免疫反应性验证。多肽活化CTL能力结果显示,预培养法ELISPOT产生斑点形成细胞(spvt forming cell,SFC)的净值T明显高于直接法ELISPOT:[(阳性对照肽:322±8 vs 169±22,P<0.05)、(多肽P1:114±10 vs 39±7,P<0.05)、(多肽P10:156±31 vs 52±8,P<0.05)]。单个活化CTL细胞分泌INF-γ因子的水平检测结果显示,预培养法ELISPOT产生斑点的平均大小明显大于直接法ELISPOT[(阳性对照肽:21.91±2.45 vs 13.80±1.76,P<0.05)、(多肽P1:12.90±0.88 vs 8.31±1.40,P<0.05)、(多肽P10:17.50±3.85 vs 11.96±0.61,P<0.05)]。表位多肽P1、P10均具有免疫原性,P10的免疫活性更高。结论预培养法ELISPOT检测多肽的免疫反应性较直接法ELISPOT灵敏性更高,多种软件联合ELISPOT预培养法可有效筛选出免疫反应性更强的卵巢癌相关抗原TM4SF1 HLA-A2限制性CTL优势表位,其中P10的免疫活性最高。
文摘Human immunodeficiency virus type-1 （HIV-1）-specific dendritic cell （DC） vaccines have been used in clinical trials. However, they have been found to only induce some degree of immune responses in these studies. We previously demonstrated that the HIV-1 Gag-specific Gag-Texo vaccine stimulated Gag-specific effector CD8＋ cytotoxic T lymphocyte （CTL） responses, leading to completely protective, but very limited, therapeutic immunity. In this study, we constructed a recombinant adenoviral vector, adenovirus （AdV）4-1BBL, which expressed mouse 4-1BB ligand （4-1BBL）, and generated transgenic 4-1BBL-engineered OVA-Texo/4.1BSL and Gag-Texo/4.1BSL vaccines by transfecting ovalbumin （OVA）-Texo and Gag-＇rexo cells with AdV4.1BBL, respectively. We demonstrate that the OVA-specific OVA-Texo/4.ZSSL vaccine stimulates more efficient OVA-specific CTL responses （3.26%） compared to OVA-Texo-activated responses （1.98%） in wild-type C57BIJ6 mice and the control OVA-TeXO/Nu, vaccine without transgenic 4-1BBL expression, leading to enhanced therapeutic immunity against 6-day established OVA-expressing B16 melanoma BL6-1OovA cells. OVA-Texo/4.1BBL-stimulated CTLs, which have a CD44＋CD62Lhigh IL-7R＋ phenotype, are likely memory CTL precursors, demonstrating prolonged survival and enhanced differentiation into memory CTLs with functional recall responses and long-term immunity against BL6-1OovA melanoma. In addition, we demonstrate that OVA-Texo/4_ZBBL-Stimulated CTLs up- and downregulate the expression of anti-apoptosis （Bcl2110, Naipl, No13, Pak7 and Tnfrsfllb） and pro-apoptosis （Casp12, Trp63 and Trp73） genes, respectively, by RT2 Profiler PCR array analysis. Importantly, the Gag-specific Gag-Texo/4.1BBL vaccine also stimulates more efficient Gag-specific therapeutic and long-term immunity against HLA-A2/Gag-expressing B 16 melanoma BL6-1OGag/A2 cells than the control Gag-TeXO/NuH vaccine in transgenic HLA-A2 mice. Taken together, our novel Gag-Texo/4-ZBBL vaccine, which is capable of stimulating potent Gag-specific therapeutic and long-term immunity, may represent a new immunotherapeutic vaccine for controlling HIV-1 infection.
