新型CpG ODN大规模筛选及其抗病毒作用的研究
|
摘要
CpG ODN 是人工合成的含未甲基化CpG 基序的寡脱氧核苷酸,可分为三种类型:A型CpG ODN能够刺激浆细胞样树突状细胞(pDC)产生大量的IFN-α,并能强效激活NK 细胞;B 型CpG ODN 具有很强的刺激B 细胞增生能力;C型CpG ODN 兼有A 型及B 型CpG ODN 的作用。CpG ODN 能活化机体的天然免疫,诱导产生Th1 型免疫反应,在抵抗病毒感染、治疗肿瘤、抗过敏等方面有广泛的应用前景。
我们新设计了190 多条CpG ODN,经过筛选,得到A 型CpG ODN 6 条,B 型CpG ODN 8 条,C 型CpG ODN 9 条,并申请了5 项国家发明专利。
在此基础上,我们对抗病毒能力突出的CpG-302(C 型)和CpG-B4(A型)进行了较深入的研究。结果表明,CpG-302 属于一种新结构类型的C 型CpG ODN,其刺激的人PBMC 培养上清具有抗VSV、SARS-CoV 和流感病毒的作用,单独应用也能够直接保护细胞免受VSV 感染。CpG-B4 能够在一定程度上保护小鼠抵抗流感病毒的攻击,并能使乳鼠产生剂量依赖性抵抗口蹄疫病毒攻击的作用。因此,CpG-302 和CpG-B4 可能成为具有独立知识产权的新型抗病毒药物。
Athough vast progress has been made in treating the infectious diseases, virusinfection is still a major health problem. According to the data from China'sMinistry of Health, there are 120-million HBV carriers in China, nearly 10percent of its population, and 30-million of them have become active patients. Inaddition, a total of 38,000,000 cases of HCV occur in 1990’s in China and manyof which will develop into recurring hepatitis, liver cirrhosis and hepatocellularcarcinoma. Also HIV, influenza virus is associated with tens of thousands ofdeaths per year. Foot and Mouth Disease (FMD) and avian influenza infection inanimals also have substantial economic impact.
So far, it seems that recombinant interferons (IFN) is one of few effectivedrugs available in treating HBV, HCV infection and other virus-associateddiseases. However, IFN has a short half-life time and seems not effective inpatients with chronic HBV. Several side effects are also observed during thecourse of recombinant interferon treatment. So, new types of drugs are badlyneeded to be developed for controling the virus infection.
CpG ODN are artificial oligodeoxynucleotides. Based on their stimulationeffects on human PBMC, CpG ODN can be divided into 3 types: CpG-A ODNs(also known as”type D”), CpG-B ODNs (also known as “type K”) and CpG-CODNs. 2216, a published CpG ODN with the prototype sequence of CpG-AODNs, is a very potent inducer of IFN-αand IFN-γfrom pDC and NK cellsrespectively whereas stimulates weak proliferation of B cells and differentiationof plasmocytoid dendritic cells (pDC). 2006, a well-studied CpG ODN with theprototype sequence of CpG-B ODNs, is able to stimulate various functions of Bcells and induce maturation of DC, on the contrary, has little or no effects onpDC. CpG-C ODNs share the immune effects of both A-and B-Class CpG ODN.CpG ODN such as C274, M362, 2395 are all representatives of this class.
In all, CpG ODN activate the cells of innate immune system, includingdendritic cells, macrophages, and natural killer (NK) cells, and consequently
stimulate the production of Th1 cytokines and chemokines, the up-regulation ofcostimulatory molecules, which facilitate the generation of adaptive immuneresponses. So, CpG ODN manifest very promising potential in clinicalapplications and can be used as adjuvants to enhance efficacies of variouscurrent vaccines, and as mono-therapeutics to treat cancers and allergies. Accumulating clinical studies show that CpG ODN can efficiently preventvirus infections. A phase Ia study in health volunteers show that CpG ODN issafe and well tolerated over a wide dose range. A Phase Ib study showed thatCpG ODN treatment could significantly reduce early viral burden and enhanceantiviral immune response in patients who had relapsed hepatitis C after or priorIFN-αtherapy. To identify CpG ODN with potential of clinical applications, we have beencarried out a large scale screening of novel CpG ODN. About 190 CpG ODNwere designed and screened by using the inhibition of cytopathic effect (CPE)bioassay and H-Thymidine incorporation assay. In this process, we successfully 3identified a batch of novel CpG ODN, among which 6 were CpG-A ODN, 8were CpG-B ODN and 9 were CpG-C ODN. To develop novel anti-virusmedicine, we selected CpG-302 (CpG-C ODN) and CpG-B4 (CpG-A ODN) forfurther analysis.The contents of this paper are focused on the following parts:1 Screening for optimized sequences with potent effects on human PBMC We synthesized more than 190 CpG ODN. By using the inhibition ofcytopathic effect (CPE) bioassay and H-Thymidine incorporation assay, we 3tested these CpG ODN for their capacity to stimulate human PBMC toproliferate and to secrete anti-virus substances. Up to now, we have identified abatch of novel CpG ODN including 6 CpG-A ODNs, 8 CpG-B ODNs and 9CpG-C ODNs. As reported, the effects of CpG ODN is species-specific, so we tested thescreened CpG ODN for their capacities to induce the proliferation and anti-virusactivity of mouse splenocytes, rhesus PBMC and porcine PBMC. Sequencescontaining ‘GTCGTT’motif could stimulate proliferation of mouse splenocytes.CpG-667 and CpG-684, which contain ‘GTCGTT’motif, could also stimulateporcine PBMC and rhesus PBMC to proliferate. These data indicated CpG ODNcontaining ‘GTCGTT’motif had no species-specific in their function. However,as the anti-virus effects of the CpG ODN-conditioned supernatants wereconcerned, all the CpG ODN which had potent stimulatory effects on humanPBMC showed no effects on mouse splenocytes except CpG-B4 and CpG-1007,which also induced porcine and rhesus PBMC to secrete anti-virus cytokines.
CpG-302 which could simulate the proliferation of PBMC and the productionof anti-virus substances from the PBMC was also tested for its heterogeneity bythe same methods described above. CpG-302 showed no stimulatory effects onmouse splenocytes and porcine PBMC. By comparation, CpG-302 failed tostimulte rhesus PBMC to proliferate, but could stimulate rhesus PBMC toproduce anti-virus factors. These findings suggest that the CpG-302 functions ina highly species-specific way.2 Analyze the stimulatory effects of CpG-302 on human PBMC Of the 9 CpG-C ODNs, CpG-302 showed the highest activity for humanPBMC and therefore was selected for further analysis.2.1 Time-and dose-dependent anti-VSV effects induced by CpG-302 To analyze the kinetics of CpG-302 in inducing anti-virus activity, thesupernatants of cultured human PBMC stimulated by CpG-302, 2216 orControl-302 (identical to CpG-302 except the CG is reversed) were collected atdifferent time point (0.5 h～96 h) and tested for their ability to inhibit thecytopathic effect (CPE) caused by VSV on VERO E6 cells. The results showedthat anti-VSV activity induced by CpG-302 appeared at 2 h, reached peak at 12 hand maintained a plateau from 12 to 96 h. Compared to 2216, CpG-302 hadsimilar effects in terms of the speed, level and maintained time of inducedanti-VSV activities. Control-302 couldn’t induce human PBMC to produceanti-virus factors. To determine the optimal dosage of CpG ODN, human PBMC werestimulated with different doses of CpG-302, 2216 and Control-302 and thesupernatants were collected to test their abilities to protect Vero E6 cells fromVSV challenge. It was found that CpG-302 as little as 0.05μg/ml was sufficientto induce human PBMC to produce anti-virus factors, and at the dosage of 3～6μg/ml induced the highest production of the factors.2.2 Anti-influenza virus and anti-SARS-CoV activities of the supernatant of human PBMC stimulated by CpG-302 Next, we observed whether CpG-302 could induce activity against influenzavirus and SARS-CoV. The stimulated supernatants of human PBMC stimulatedby CpG-302, C274 (C-type control), 2006 (B-type control), 2216 (A-type control)and medium (negative control) were tested for its ability to inhibit the cytopathiceffect (CPE) caused by influenza virus on VERO E6 cells. The result showedthat CpG-302 induced similar anti-virus activity as 2216 but higher than C274.Anti-SARS-CoV activities of the series diluted supernatants of human PBMCstimulated with 25μg/ml, 12.5μg/ml, 6.25μg/ml and 3.13μg/ml CpG-302 werealso investigated. We found that supernatants stimulated by CpG-302 protected
Vero E6 cells from SARS-CoV infection. Together, these data suggest thatCpG-302 can stimulate human PBMC to produce anti-virus factors.2.3 Heterogeneous response of human PBMC to CpG-302 The stimulated supernatants of human PBMC from different donorsstimulated by CpG-302 were tested for its ability to inhibit the cytopathic effect(CPE) caused by VSV on VERO E6 cells, and their capacity to stimulateproliferation of human PBMC from different donors were also evaluated. Wefound that the proliferation and anti-virus effects of human PBMC stimulated byCpG-302 were heterogeneous, and the harbored mechanisms needed to befurther explored.2.4 Analysis of cytokines induced by CpG-302 Cytokines in the supernatants of human PBMC stimulated by CpG-302 werequantified by ELISA Kits. CpG-302 could induce human PBMC to produce highlevel of IFN-αeven though it was lower than that stimulated by 2216. ButCpG-302, as well as 2006 and 2216, could stimulate human PBMC to secretcertain amount of IL-6, TNF-αand MIP-1βat similar level. IFN-γproductioninduced by CpG-302 or 2216 appeared higher level than that induced by 2006. Plasmacytoid dendritic cells (pDC) are the major cells to produce largeamount of IFN-αin response to viral infection. Human pDC were isolated andpurified from buffy coat and cultured with or without CpG-302 or 2216 in themedium containing different stimulators as indicated. The supernatants fromcultured human pDC were harvested and used for detection of IFN-αand IL-12.The result showed that CpG-302 as well as 2216 could stimulate human pDC toproduce large amount of IFN-αand certain amount of IL-12 when combinedwith CD40L.2.5 Anti-VSV activities of the supernatants before or after IFN-α neutralization Type I interferon (IFN-α/β) is the major anti-virus cytokines. To test whetheranti-virus activities of the supernatants due to the type I interferon induced byCpG-302, supernatants from human PBMC stimulated by CpG-302 wereneutralized by IFN-α, IFN-βand IFN-γpolyclonal neutralizing antibodies, andthe remaining anti-virus effect were tested by bio-assay as described.Neutralization of IFN-αcould partially weaken but could’t fully abolish theanti-virus effect, whereas neutralizing IFN-βand IFN-γdid not influence theanti-virus activity, indicating that IFN-αdid contributed the anti-virus activityand except IFN-β, IFN-γother soluble factors were involved in.2.6 Activation of immune cells induced by CpG-302 Because of the unique properties of CpG-302 in anti-virus, we observed
whether the effect of CpG-302 on immune cells was different with other CpGODN. Human PBMC were stimulated with CpG-302, 2216 or 2006 for 48h,stained with FITC-conjugated CD40, CD80, CD83, CD86 and HLA-A2antibodies and then analyzed on a FACSCalibur. The result showed thatCpG-302 could upregulate the expression of CD40, CD80, CD86 and HLA-A2. CD69 is an early marker of lymphoid cell activation. Human PBMCs werestimulated with different CpG ODN including CpG-302, Control-302, 2006 and2216, then stained with PE-conjugated anti-CD19, CD56, CD14, CD3 andFITC-labeled CD69 monoclonal antibodies and analyzed on a FACSCalibur.The results showed that activation of B cells (CD19+), NK cells (CD56+) andmonocytes (CD14+) stimulated by CpG-302 was much stronger than thatinduced by 2216 (A-type control). Control-302 couldn’t activate immune cells,and all the sequences tested cannot acivate T cells (CD3+). In additon, CpG-302also enhanced the lytic activity of NK cells within PBMC and could stimulatepurified human B cells to proliferate. All these results suggest CpG-302 belongto CpG-C ODNs and can activate immune cells such as B cells, pDC, NK cells,monocytes etc.2.7 Effect of poly (G) and CG motifs of CpG-302 on its stimulation of human PBMC A panel of CpG ODN (CpG-3021～CpG-3030) were synthesized based on thestructure of CpG-302 and were tested for their capacity to simulate proliferationand to induce supernatant with anti-virus effects of human PBMC. The resultsshowed that the 6 guanines (G) at the 3’end could enhance CpG-302 capacity toinduce anti-virus effects, while inhibited its capacity to simulate proliferation ofhuman PBMC. Further, the ‘TCG’motifs at 5’end and the palindromic sequencein CpG-302 were necessay for its activity.2.8 Direct inhibition of VSV on Vero E6 cells by CpG-302 Vero cells were seeded into 96-well flat-bottomed plates (3 ×104 cells/100μl/well) and cultured for 24 h to confluency. The cells were directlyincubated with CpG ODN and 10TCID50 (50% tissue culture infectious doses)of VSV for another 48h. The cytopathic effect caused by VSV was examined byusing crystal violet staining method. Anti-virus effect was expressed as ODvalues, which were correlated to the anti-virus effect of the stimulatedsupernatant. Of note, CpG-302, not 2006, 2216 or C274 could inhibit VSVinfection on Vero E6 cells. Further analysis showed that the anti-virus effect wasdependent on the phosphorothioated backbone and the sequence of CpG ODN.2.9 The function of CpG-302 related to scavenger receptor Scavenger receptor A (SR-A) can recognize a number of polyannions such as
fucoidan, dextran sulfate (DS), polyG/polyI and play important role in innateimmunity. Also, it has some effects on cells’adhesion. CpG ODN is one kind ofpolyannions and may be recognized by SR-A. Our experiments also showedCpG-302 could inhibit cells adhesion to the bottom of 96-well flat-bottomedplates. So, we analyzed whether the stimulation effects of CpG-302 could beinhibited by DS, which did not induce any detectable cytokines and could notstimulate human PBMC to proliferate. Human PBMC were incubated with 30μg/ml dextran sulfate and 3μg/mlCpG-302, 2216, 2006, C274 for 48h, the supernatants were collected andassayed for their anti-virus effects as described above. PBMC proliferation wasalso assessed by H-thymidine incorporation assay. The effect of CpG ODN on 3stimulating human PBMC to produce interferon and the other anti-virus factorscould be completely inhibited by dextran sulfate. Also dextran sulfate couldinhibit PBMC proliferation stimulated by CpG-302, but failed to inhibit theproliferation stimulated by 2006 and C274. So, there are some differencesbetween C274 and CpG-302 in spite of their similarly stimulatory effects onhuman PBMC. C274 and CpG-302 have a different structure. There are 6 guanines at the 3’end of CpG-302. In order to elucidate whether the difference between C274 andCpG-302 is owed to the 6 guanines at the 3’end of CpG-302. The sequence(CpG-3026) identical to CpG-302 except there is no guanine at the 3’end wasused to stimulate human PBMC with or without dextran sulfate. The resultdemonstrated that the difference between C274 and CpG-302 was not owed tothe existence of the 6 guanines at the 3’end of CpG-302. Further, human PBMCwas cultured with dextran sulfate alone or with partial phosphorothioated302(Part-302) and unmodified 302(PD-302), and the result showed that dextransulfate could also inhibit the stimulatory effect of CpG ODN on inducing humanPBMC to produce anti-virus factors. All these result demonstrated that theinhibitory effect of dextran sulfate was dependent on the sequence structure, noton the phosphorothioated backbone or guanine at the 3’end of CpG ODN.3 Anti-virus effects of CpG-B4 Elegant studies have shown that CpG-A ODN could stimulate lymphocytes tosecrete type I interferons (IFN-α/β), IFN-γ、IL-12、IL-18 etc, enhance the antigenpresentation ability of antigen presenting cells (APC), activate innate immuneresponse, and facilitate the aquired immune response. So, CpG-A ODN may becompetent at combating virus infection. CpG-B4 is a novel CpG-A ODNidentified in the process of large-scale screening, which could stimulate humanPBMC and mouse splenocytes to secrete large amount of anti-virus factors.
我们新设计了190 多条CpG ODN,经过筛选,得到A 型CpG ODN 6 条,B 型CpG ODN 8 条,C 型CpG ODN 9 条,并申请了5 项国家发明专利。
在此基础上,我们对抗病毒能力突出的CpG-302(C 型)和CpG-B4(A型)进行了较深入的研究。结果表明,CpG-302 属于一种新结构类型的C 型CpG ODN,其刺激的人PBMC 培养上清具有抗VSV、SARS-CoV 和流感病毒的作用,单独应用也能够直接保护细胞免受VSV 感染。CpG-B4 能够在一定程度上保护小鼠抵抗流感病毒的攻击,并能使乳鼠产生剂量依赖性抵抗口蹄疫病毒攻击的作用。因此,CpG-302 和CpG-B4 可能成为具有独立知识产权的新型抗病毒药物。
Athough vast progress has been made in treating the infectious diseases, virusinfection is still a major health problem. According to the data from China'sMinistry of Health, there are 120-million HBV carriers in China, nearly 10percent of its population, and 30-million of them have become active patients. Inaddition, a total of 38,000,000 cases of HCV occur in 1990’s in China and manyof which will develop into recurring hepatitis, liver cirrhosis and hepatocellularcarcinoma. Also HIV, influenza virus is associated with tens of thousands ofdeaths per year. Foot and Mouth Disease (FMD) and avian influenza infection inanimals also have substantial economic impact.
So far, it seems that recombinant interferons (IFN) is one of few effectivedrugs available in treating HBV, HCV infection and other virus-associateddiseases. However, IFN has a short half-life time and seems not effective inpatients with chronic HBV. Several side effects are also observed during thecourse of recombinant interferon treatment. So, new types of drugs are badlyneeded to be developed for controling the virus infection.
CpG ODN are artificial oligodeoxynucleotides. Based on their stimulationeffects on human PBMC, CpG ODN can be divided into 3 types: CpG-A ODNs(also known as”type D”), CpG-B ODNs (also known as “type K”) and CpG-CODNs. 2216, a published CpG ODN with the prototype sequence of CpG-AODNs, is a very potent inducer of IFN-αand IFN-γfrom pDC and NK cellsrespectively whereas stimulates weak proliferation of B cells and differentiationof plasmocytoid dendritic cells (pDC). 2006, a well-studied CpG ODN with theprototype sequence of CpG-B ODNs, is able to stimulate various functions of Bcells and induce maturation of DC, on the contrary, has little or no effects onpDC. CpG-C ODNs share the immune effects of both A-and B-Class CpG ODN.CpG ODN such as C274, M362, 2395 are all representatives of this class.
In all, CpG ODN activate the cells of innate immune system, includingdendritic cells, macrophages, and natural killer (NK) cells, and consequently
stimulate the production of Th1 cytokines and chemokines, the up-regulation ofcostimulatory molecules, which facilitate the generation of adaptive immuneresponses. So, CpG ODN manifest very promising potential in clinicalapplications and can be used as adjuvants to enhance efficacies of variouscurrent vaccines, and as mono-therapeutics to treat cancers and allergies. Accumulating clinical studies show that CpG ODN can efficiently preventvirus infections. A phase Ia study in health volunteers show that CpG ODN issafe and well tolerated over a wide dose range. A Phase Ib study showed thatCpG ODN treatment could significantly reduce early viral burden and enhanceantiviral immune response in patients who had relapsed hepatitis C after or priorIFN-αtherapy. To identify CpG ODN with potential of clinical applications, we have beencarried out a large scale screening of novel CpG ODN. About 190 CpG ODNwere designed and screened by using the inhibition of cytopathic effect (CPE)bioassay and H-Thymidine incorporation assay. In this process, we successfully 3identified a batch of novel CpG ODN, among which 6 were CpG-A ODN, 8were CpG-B ODN and 9 were CpG-C ODN. To develop novel anti-virusmedicine, we selected CpG-302 (CpG-C ODN) and CpG-B4 (CpG-A ODN) forfurther analysis.The contents of this paper are focused on the following parts:1 Screening for optimized sequences with potent effects on human PBMC We synthesized more than 190 CpG ODN. By using the inhibition ofcytopathic effect (CPE) bioassay and H-Thymidine incorporation assay, we 3tested these CpG ODN for their capacity to stimulate human PBMC toproliferate and to secrete anti-virus substances. Up to now, we have identified abatch of novel CpG ODN including 6 CpG-A ODNs, 8 CpG-B ODNs and 9CpG-C ODNs. As reported, the effects of CpG ODN is species-specific, so we tested thescreened CpG ODN for their capacities to induce the proliferation and anti-virusactivity of mouse splenocytes, rhesus PBMC and porcine PBMC. Sequencescontaining ‘GTCGTT’motif could stimulate proliferation of mouse splenocytes.CpG-667 and CpG-684, which contain ‘GTCGTT’motif, could also stimulateporcine PBMC and rhesus PBMC to proliferate. These data indicated CpG ODNcontaining ‘GTCGTT’motif had no species-specific in their function. However,as the anti-virus effects of the CpG ODN-conditioned supernatants wereconcerned, all the CpG ODN which had potent stimulatory effects on humanPBMC showed no effects on mouse splenocytes except CpG-B4 and CpG-1007,which also induced porcine and rhesus PBMC to secrete anti-virus cytokines.
CpG-302 which could simulate the proliferation of PBMC and the productionof anti-virus substances from the PBMC was also tested for its heterogeneity bythe same methods described above. CpG-302 showed no stimulatory effects onmouse splenocytes and porcine PBMC. By comparation, CpG-302 failed tostimulte rhesus PBMC to proliferate, but could stimulate rhesus PBMC toproduce anti-virus factors. These findings suggest that the CpG-302 functions ina highly species-specific way.2 Analyze the stimulatory effects of CpG-302 on human PBMC Of the 9 CpG-C ODNs, CpG-302 showed the highest activity for humanPBMC and therefore was selected for further analysis.2.1 Time-and dose-dependent anti-VSV effects induced by CpG-302 To analyze the kinetics of CpG-302 in inducing anti-virus activity, thesupernatants of cultured human PBMC stimulated by CpG-302, 2216 orControl-302 (identical to CpG-302 except the CG is reversed) were collected atdifferent time point (0.5 h～96 h) and tested for their ability to inhibit thecytopathic effect (CPE) caused by VSV on VERO E6 cells. The results showedthat anti-VSV activity induced by CpG-302 appeared at 2 h, reached peak at 12 hand maintained a plateau from 12 to 96 h. Compared to 2216, CpG-302 hadsimilar effects in terms of the speed, level and maintained time of inducedanti-VSV activities. Control-302 couldn’t induce human PBMC to produceanti-virus factors. To determine the optimal dosage of CpG ODN, human PBMC werestimulated with different doses of CpG-302, 2216 and Control-302 and thesupernatants were collected to test their abilities to protect Vero E6 cells fromVSV challenge. It was found that CpG-302 as little as 0.05μg/ml was sufficientto induce human PBMC to produce anti-virus factors, and at the dosage of 3～6μg/ml induced the highest production of the factors.2.2 Anti-influenza virus and anti-SARS-CoV activities of the supernatant of human PBMC stimulated by CpG-302 Next, we observed whether CpG-302 could induce activity against influenzavirus and SARS-CoV. The stimulated supernatants of human PBMC stimulatedby CpG-302, C274 (C-type control), 2006 (B-type control), 2216 (A-type control)and medium (negative control) were tested for its ability to inhibit the cytopathiceffect (CPE) caused by influenza virus on VERO E6 cells. The result showedthat CpG-302 induced similar anti-virus activity as 2216 but higher than C274.Anti-SARS-CoV activities of the series diluted supernatants of human PBMCstimulated with 25μg/ml, 12.5μg/ml, 6.25μg/ml and 3.13μg/ml CpG-302 werealso investigated. We found that supernatants stimulated by CpG-302 protected
Vero E6 cells from SARS-CoV infection. Together, these data suggest thatCpG-302 can stimulate human PBMC to produce anti-virus factors.2.3 Heterogeneous response of human PBMC to CpG-302 The stimulated supernatants of human PBMC from different donorsstimulated by CpG-302 were tested for its ability to inhibit the cytopathic effect(CPE) caused by VSV on VERO E6 cells, and their capacity to stimulateproliferation of human PBMC from different donors were also evaluated. Wefound that the proliferation and anti-virus effects of human PBMC stimulated byCpG-302 were heterogeneous, and the harbored mechanisms needed to befurther explored.2.4 Analysis of cytokines induced by CpG-302 Cytokines in the supernatants of human PBMC stimulated by CpG-302 werequantified by ELISA Kits. CpG-302 could induce human PBMC to produce highlevel of IFN-αeven though it was lower than that stimulated by 2216. ButCpG-302, as well as 2006 and 2216, could stimulate human PBMC to secretcertain amount of IL-6, TNF-αand MIP-1βat similar level. IFN-γproductioninduced by CpG-302 or 2216 appeared higher level than that induced by 2006. Plasmacytoid dendritic cells (pDC) are the major cells to produce largeamount of IFN-αin response to viral infection. Human pDC were isolated andpurified from buffy coat and cultured with or without CpG-302 or 2216 in themedium containing different stimulators as indicated. The supernatants fromcultured human pDC were harvested and used for detection of IFN-αand IL-12.The result showed that CpG-302 as well as 2216 could stimulate human pDC toproduce large amount of IFN-αand certain amount of IL-12 when combinedwith CD40L.2.5 Anti-VSV activities of the supernatants before or after IFN-α neutralization Type I interferon (IFN-α/β) is the major anti-virus cytokines. To test whetheranti-virus activities of the supernatants due to the type I interferon induced byCpG-302, supernatants from human PBMC stimulated by CpG-302 wereneutralized by IFN-α, IFN-βand IFN-γpolyclonal neutralizing antibodies, andthe remaining anti-virus effect were tested by bio-assay as described.Neutralization of IFN-αcould partially weaken but could’t fully abolish theanti-virus effect, whereas neutralizing IFN-βand IFN-γdid not influence theanti-virus activity, indicating that IFN-αdid contributed the anti-virus activityand except IFN-β, IFN-γother soluble factors were involved in.2.6 Activation of immune cells induced by CpG-302 Because of the unique properties of CpG-302 in anti-virus, we observed
whether the effect of CpG-302 on immune cells was different with other CpGODN. Human PBMC were stimulated with CpG-302, 2216 or 2006 for 48h,stained with FITC-conjugated CD40, CD80, CD83, CD86 and HLA-A2antibodies and then analyzed on a FACSCalibur. The result showed thatCpG-302 could upregulate the expression of CD40, CD80, CD86 and HLA-A2. CD69 is an early marker of lymphoid cell activation. Human PBMCs werestimulated with different CpG ODN including CpG-302, Control-302, 2006 and2216, then stained with PE-conjugated anti-CD19, CD56, CD14, CD3 andFITC-labeled CD69 monoclonal antibodies and analyzed on a FACSCalibur.The results showed that activation of B cells (CD19+), NK cells (CD56+) andmonocytes (CD14+) stimulated by CpG-302 was much stronger than thatinduced by 2216 (A-type control). Control-302 couldn’t activate immune cells,and all the sequences tested cannot acivate T cells (CD3+). In additon, CpG-302also enhanced the lytic activity of NK cells within PBMC and could stimulatepurified human B cells to proliferate. All these results suggest CpG-302 belongto CpG-C ODNs and can activate immune cells such as B cells, pDC, NK cells,monocytes etc.2.7 Effect of poly (G) and CG motifs of CpG-302 on its stimulation of human PBMC A panel of CpG ODN (CpG-3021～CpG-3030) were synthesized based on thestructure of CpG-302 and were tested for their capacity to simulate proliferationand to induce supernatant with anti-virus effects of human PBMC. The resultsshowed that the 6 guanines (G) at the 3’end could enhance CpG-302 capacity toinduce anti-virus effects, while inhibited its capacity to simulate proliferation ofhuman PBMC. Further, the ‘TCG’motifs at 5’end and the palindromic sequencein CpG-302 were necessay for its activity.2.8 Direct inhibition of VSV on Vero E6 cells by CpG-302 Vero cells were seeded into 96-well flat-bottomed plates (3 ×104 cells/100μl/well) and cultured for 24 h to confluency. The cells were directlyincubated with CpG ODN and 10TCID50 (50% tissue culture infectious doses)of VSV for another 48h. The cytopathic effect caused by VSV was examined byusing crystal violet staining method. Anti-virus effect was expressed as ODvalues, which were correlated to the anti-virus effect of the stimulatedsupernatant. Of note, CpG-302, not 2006, 2216 or C274 could inhibit VSVinfection on Vero E6 cells. Further analysis showed that the anti-virus effect wasdependent on the phosphorothioated backbone and the sequence of CpG ODN.2.9 The function of CpG-302 related to scavenger receptor Scavenger receptor A (SR-A) can recognize a number of polyannions such as
fucoidan, dextran sulfate (DS), polyG/polyI and play important role in innateimmunity. Also, it has some effects on cells’adhesion. CpG ODN is one kind ofpolyannions and may be recognized by SR-A. Our experiments also showedCpG-302 could inhibit cells adhesion to the bottom of 96-well flat-bottomedplates. So, we analyzed whether the stimulation effects of CpG-302 could beinhibited by DS, which did not induce any detectable cytokines and could notstimulate human PBMC to proliferate. Human PBMC were incubated with 30μg/ml dextran sulfate and 3μg/mlCpG-302, 2216, 2006, C274 for 48h, the supernatants were collected andassayed for their anti-virus effects as described above. PBMC proliferation wasalso assessed by H-thymidine incorporation assay. The effect of CpG ODN on 3stimulating human PBMC to produce interferon and the other anti-virus factorscould be completely inhibited by dextran sulfate. Also dextran sulfate couldinhibit PBMC proliferation stimulated by CpG-302, but failed to inhibit theproliferation stimulated by 2006 and C274. So, there are some differencesbetween C274 and CpG-302 in spite of their similarly stimulatory effects onhuman PBMC. C274 and CpG-302 have a different structure. There are 6 guanines at the 3’end of CpG-302. In order to elucidate whether the difference between C274 andCpG-302 is owed to the 6 guanines at the 3’end of CpG-302. The sequence(CpG-3026) identical to CpG-302 except there is no guanine at the 3’end wasused to stimulate human PBMC with or without dextran sulfate. The resultdemonstrated that the difference between C274 and CpG-302 was not owed tothe existence of the 6 guanines at the 3’end of CpG-302. Further, human PBMCwas cultured with dextran sulfate alone or with partial phosphorothioated302(Part-302) and unmodified 302(PD-302), and the result showed that dextransulfate could also inhibit the stimulatory effect of CpG ODN on inducing humanPBMC to produce anti-virus factors. All these result demonstrated that theinhibitory effect of dextran sulfate was dependent on the sequence structure, noton the phosphorothioated backbone or guanine at the 3’end of CpG ODN.3 Anti-virus effects of CpG-B4 Elegant studies have shown that CpG-A ODN could stimulate lymphocytes tosecrete type I interferons (IFN-α/β), IFN-γ、IL-12、IL-18 etc, enhance the antigenpresentation ability of antigen presenting cells (APC), activate innate immuneresponse, and facilitate the aquired immune response. So, CpG-A ODN may becompetent at combating virus infection. CpG-B4 is a novel CpG-A ODNidentified in the process of large-scale screening, which could stimulate humanPBMC and mouse splenocytes to secrete large amount of anti-virus factors.
引文
1. Agrawal S, Temsamani J, Galbraith W, Tang J. Pharmacokinetics of antisense oligonucleotides. Clin Pharmacokinet. 1995; 28(1): 7-16
2. Ahn K, Meyer TH, Uebel S, Sempe P, Djaballah H, Yang Y, Peterson PA, Fruh K, Tampe R. Molecular mechanism and species specificity of TAP inhibition by herpes simplex virus ICP47. EMBO J. 1996; 15(13): 3247-55
3. Ahn KS, Ou W, Silver J. Inhibition of certain strains of HIV-1 by cell surface polyanions in the form of cholesterol-labeled oligonucleotides. Virology. 2004; 330(1): 50-61
4. Akira S, Takeda K. Functions of Toll-like receptors: lessons from KO mice. C. R. Biologies. 2004; 327: 581–589
5. Akira S, Takeda K. Toll-like receptor signalling. Nat Rev Immunol. 2004; 4(7): 499-511
6. Balamurugan V, Kumar RM, Suryanarayana VV. Past and present vaccine development strategies for the control of foot-and-mouth disease. Acta Virol. 2004; 48(4): 201-14
7. Ballas ZK, Krieg AM, Warren T, Rasmussen W, Davis HL, Waldschmidt M, Weiner GJ. Divergent therapeutic and immunologic effects of oligodeoxynucleotides with distinct CpG motifs. J Immunol. 2001; 167(9):4878-86
8. Ballas ZK, Rasmussen WL, Krieg AM. Induction of NK Activity in Murine and Human Cells by CpG Motifs in Oligodeoxynucleotides and Bacterial DNA. J Immunol. 1996; 157: 1840-1845
9. Becker Y. A point of view: HIV-1/AIDS is an allergy but CpG ODN treatments may inhibit virus replication and reactivate the adaptive immunity--hypothesis and implications. Virus Genes. 2005; 30(1): 127-31
10. Becker Y. CpG ODNs Treatments of HIV-1 Infected Patients May Cause the Decline of Transmission in High Risk Populations -A Review, Hypothesis and Implications. Virus Genes. 2005; 30(2): 251-66
11. Bird AP. CpG-rich islands and the function of DNA methylation. Nature. 1986; 321(6067): 209–213
12. Biron CA, Nguyen KB, Pien GC, Cousens LP, Salazar-Mather TP. Natural killer cells in antiviral defense: function and regulation by innate cytokines. Annu Rev Immunol. 1999; 17: 189-220
13. Bjersing JL, Tarkowski A, Collins LV. Anti-proliferative effects of phosphodiester oligodeoxynucleotides. Immunobiology. 2004; 209(8): 637-45
14. Blackwell SE and Krieg AM. CpG-A-Induced Monocyte IFN-γInducible Protein-10 Production Is Regulated by Plasmacytoid Dendritic Cell-Derived IFN-α. J Immunol. 2003; 170: 4061–4068
15. Bobardt MD, Armand-Ugon M, Clotet I, Zhang Z, David G, Este JA, Gallay PA. Effect of polyanion-resistance on HIV-1 infection. Virology. 2004; 325(2): 389-98
16. Brazolot Millan CL, Weeratna R, Krieg AM, Siegrist CA, Davis HL. CpG DNA can induce strong Th1 humoral and cell-mediated immune responses against hepatitis B surface antigen in young mice. Proc Natl Acad Sci U S A. 1998; 95(26): 15553-8
17. Brierley MM, Fish EN. Review: IFN-alpha/beta receptor interactions to biologic outcomes: understanding the circuitry. J Interferon Cytokine Res. 2002; 22(8): 835-45
18. Cella M, Jarrossay D, Facchetti F, Alebardi O, Nakajima H, Lanzavecchia A, Colonna M. Plasmacytoid monocytes migrate to inflamed lymph nodes and produce large amounts of type I interferon. Nat Med. 1999; 5(8): 919-23
19. Chadha KC, Dembinski WE, Dunn CB, Aradi J, Bardos TJ, Dunn JA, Ambrus JL Sr. Effect of increasing thiolation of the polycytidylic acid strand of poly I:poly C on the alpha, beta and gamma interferon-inducing properties, antiviral and antiproliferative activities. Antiviral Res. 2004; 64(3): 171-7
20. Cho JY, Miller M, Baek KJ, Castaneda D, Nayar J, Roman M, Raz E, Broide DH. Immunostimulatory DNA sequences inhibit respiratory syncytial viral load, airway inflammation, and mucus secretion. J Allergy Clin Immunol. 2001; 108(5): 697-702
21. Choi YK, Ozaki H, Webby RJ, Webster RG, Peiris JS, Poon L, Butt C, Leung YH, Guan Y. Continuing evolution of H9N2 influenza viruses in Southeastern China. J Virol. 2004; 78(16): 8609-14
22. Cocchi F, DeVico AL, Garzino-Demo A, Arya SK, Gallo RC, Lusso P. Identification of RANTES, MIP-1 alpha, and MIP-1 beta as the major HIV-suppressive factors produced by CD8+ T cells. Science. 1995; 270(5243): 1811-5
23. Coccia EM, Severa M, Giacomini E, Monneron D, Remoli ME, Julkunen I, Cella M, Lande R, Uze G. Viral infection and Toll-like receptor agonists induce a differential expression of type I and lambda interferons in human plasmacytoid and monocyte-derived dendritic cells. Eur J Immunol. 2004; 34(3): 796-805
24. Cooper CL, Davis HL, Morris ML, Efler SM, Krieg AM, Li Y, Laframboise C, Al Adhami MJ, Khaliq Y, Seguin I, Cameron DW. Safety and immunogenicity of CPG 7909 injection as an adjuvant to Fluarix influenza vaccine. Vaccine. 2004; 22(23-24): 3136-43
25. Dahl H, Linde A, Strannegard O. In vitro inhibition of SARS virus replication by human interferons. Scand J Infect Dis. 2004; 36(11-12): 829-31
26. Davis HL, Weeratna R, Waldschmidt TJ, Tygrett L, Schorr J, Krieg AM. CpG DNA is a potent enhancer of specific immunity in mice immunized with recombinant hepatitis B surface antigen. J Immunol. 1998; 160(2): 870-6
27. 戴志澄,祁国明主编.中国病毒性肝炎血清流行病学调查(1992～1995)(上 卷).北京:科学技术文献出版社,1997
28. Dong L, Mori I, Hossain MJ, Liu B, Kimura Y. An immunostimulatory oligodeoxynucleotide containing a cytidine-guanosine motif protects senescence-accelerated mice from lethal influenza virus by augmenting the T helper type 1 response. J Gen Virol. 2003; 84(Pt 6): 1623-8
29. Dong QM, He ZP, Zhuang H, Song SJ, Dai WS, Zhang SP, Chen ZH, Sun JY. Dynamics of peripheral blood B lymphocytes and natural killer cells in patients with severe acute respiratory syndrome. Zhonghua Liu Xing Bing Xue Za Zhi. 2004; 25(8): 695-7
30. Duan XZ, Wang M, Li HW, Zhuang H, Xu D, Wang FS. Decreased frequency and function of circulating plasmocytoid dendritic cells (pDC) in hepatitis B virus infected humans. J Clin Immunol. 2004; 24(6): 637-46
31. Duong FH, Filipowicz M, Tripodi M, La Monica N, Heim MH. Hepatitis C virus inhibits interferon signaling through up-regulation of protein phosphatase 2A. Gastroenterology. 2004; 126(1): 263-77
32. Ebert S, Gerber J, Bader S, Muhlhauser F, Brechtel K, Mitchell TJ, Nau R. Dose-dependent activation of microglial cells by Toll-like receptor agonists alone and in combination. J Neuroimmunol. 2005; 159(1-2): 87-96
33. Encke J, zu Putlitz J, Stremmel W, Wands JR. CpG immuno-stimulatory motifs enhance humoral immune responses against hepatitis C virus core protein after DNA-based immunization. Arch Virol. 2003; 148(3): 435-48
34. Foy E, Li K, Wang C, Sumpter R Jr, Ikeda M, Lemon SM, Gale M Jr. Regulation of interferon regulatory factor-3 by the hepatitis C virus serine protease. Science. 2003; 300(5622): 1145-8
35. French AR, Yokoyama WM. Natural killer cells and viral infections. Curr Opin Immunol. 2003; 15(1): 45-51
36. Freigang S, Probst HC, van den Broek M. DC infection promotes antiviral CTL priming: the 'Winkelried' strategy. Trends Immunol. 2005; 26(1): 13-8
37. Gilbert MJ, Riddell SR, Plachter B, Greenberg PD. Cytomegalovirus selectively blocks antigen processing and presentation of its immediate-early gene product. Nature. 1996; 383(6602): 720-2
38. Gould MP, Greene JA, Bhoj V, DeVecchio JL, Heinzel FP. Distinct modulatory effects of LPS and CpG on IL-18-dependent IFN-gamma synthesis. J Immunol. 2004; 172(3): 1754-62
39. Gretch D. Mechanism of interferon resistance in hepatitis C. Lancet. 2001; 358(9294): 1662-4
40. Groneberg DA, Poutanen SM, Low DE, Lode H, Welte T, Zabel P. Treatment and vaccines for severe acute respiratory syndrome. Lancet Infect Dis. 2005; 5(3): 147-55
41. Gruhler A, Peterson PA, Fruh K. Human cytomegalovirus immediate early glycoprotein US3 retains MHC class I molecules by transient association. Traffic. 2000; 1(4): 318-25
42. Guidotti LG, Chisari FV. Noncytolytic control of viral infections by the innate and adaptive immune response. Annu Rev Immunol. 2001; 19: 65-91
43. Guidotti LG, Rochford R, Chung J, Shapiro M, Purcell R, Chisari FV. Viral clearance without destruction of infected cells during acute HBV infection. Science. 1999; 284(5415): 825-9
44. Gurney KB, Colantonio AD, Blom B, Spits H, Uittenbogaart CH. Endogenous IFN-alpha production by plasmacytoid dendritic cells
exerts an antiviral effect on thymic HIV-1 infection. J Immunol. 2004;
173(12): 7269-76
45. Gursel I, Gursel M, Ishii KJ, Klinman DM. Sterically stabilized cationic liposomes improve the uptake and immunostimulatory activity of CpG oligonucleotides. J Immunol. 2001; 167(6): 3324-8
46. Hacker H, Mischak H, Miethke T, Liptay S, Schmid R, Sparwasser T, Heeg K, Lipford GB, Wagner H. CpG-DNA-specific activation of antigen-presenting cells requires stress kinase activity and is preceded by non-specific endocytosis and endosomal maturation. EMBO J. 1998; 17(21): 6230-40
47. Hadziyannis SJ, Sette H Jr, Morgan TR, Balan V, Diago M, Marcellin P, Ramadori G, Bodenheimer H Jr, Bernstein D, Rizzetto M, Zeuzem S, Pockros PJ, Lin A, Ackrill AM; PEGASYS International Study Group. Peginterferon-alpha2a and ribavirin combination therapy in chronic hepatitis C: a randomized study of treatment duration and ribavirin dose. Ann Intern Med. 2004; 140(5): 346-55
48. Halperin SA, Van Nest G, Smith B, Abtahi S, Whiley H, Eiden JJ. A phase I study of the safety and immunogenicity of recombinant hepatitis B surface antigen co-administered with an immunostimulatory phosphorothioate oligonucleotide adjuvant. Vaccine. 2003; 21(19-20): 2461-7
49. Harandi AM, Eriksson K, Holmgren J. A Protective Role of Locally Administered Immunostimulatory CpG Oligodeoxynucleotide in a Mouse Model of Genital Herpes Infection. Journal of virology. 2003; 77(2): 953–962.
50. Hartmann G, Krieg AM. CpG DNA and LPS induce distinct patterns of activation in human monocytes. Gene Ther. 1999; 6(5): 893-903
51. Hartmann G, Krieg AM. Mechanism and function of a newly identified CpG DNA motif in human primary B cells. J Immunol. 2000; 164(2): 944-53
52. Hartmann G, Weeratna RD, Ballas ZK, Payette P, Blackwell S, Suparto I, Rasmussen WL, Waldschmidt M, Sajuthi D, Purcell RH, Davis HL, Krieg AM. Delineation of a CpG Phosphorothioate Oligodeoxynucleotide for Activating Primate Immune Responses In Vitro and In Vivo. J Immunol. 2000; 164: 1617–1624
53. Hayashi F, Means TK, Luster AD. Toll-like receptors stimulate human neutrophil function. Blood. 2003; 102(7): 2660-9
54. Hayashi M, Satou E, Ueki R, Yano M, Miyano-Kurosaki N, Fujii M, Takaku H. Resistance to influenza A virus infection by antigen-conjugated CpG oligonucleotides, a novel antigen-specific immunomodulator. Biochem Biophys Res Commun. 2005; 329(1): 230-6
55. He XS, Draghi M, Mahmood K, Holmes TH, Kemble GW, Dekker CL, Arvin AM, Parham P, Greenberg HB. T cell-dependent production of IFN-gamma by NK cells in response to influenza A virus. J Clin Invest. 2004; 114(12): 1812-9
56. Heikenwalder M, Polymenidou M, Junt T, Sigurdson C, Wagner H, Akira S, Zinkernagel R, Aguzzi A. Lymphoid follicle destruction and immunosuppression after repeated CpG oligodeoxynucleotide administration. Nat Med. 2004; 10(2): 187-92
57. Heil F, Hemmi H, Hochrein H, Ampenberger F, Kirschning C, Akira S, Lipford G, Wagner H, Bauer S. Species-specific recognition of single-stranded RNA via toll-like receptor 7 and 8. Science. 2004; 303(5663): 1526-9
58. Heller T, Saito S, Auerbach J, Williams T, Moreen TR, Jazwinski A, Cruz B, Jeurkar N, Sapp R, Luo G, Liang TJ. An in vitro model of hepatitis C virion production. Proc Natl Acad Sci U S A. 2005; 102(7): 2579-83
59. Hemmi H, Takeuchi O, Kawai T, Kaisho T, Sato S, Sanjo H, Matsumoto M, Hoshino K, Wagner H, Takeda K, Akira S. A Toll-like receptor recognizes bacterial DNA. Nature. 2000; 408(6813): 740-5
60. Hengel H, Brune W, Koszinowski UH. Immune evasion by cytomegalovirus--survival strategies of a highly adapted opportunist. Trends Microbiol. 1998; 6(5): 190-7
61. Higgins PG, Phillpotts RJ, Scott GM, Wallace J, Bernhardt LL, Tyrrell DA. Intranasal interferon as protection against experimental respiratory coronavirus infection in volunteers. Antimicrob Agents Chemother. 1983; 24(5): 713-5
62. Hofmann WP, Zeuzem S, Sarrazin C. Hepatitis C virus-related resistance mechanisms to interferon alpha-based antiviral therapy. J Clin Virol. 2005; 32(2): 86-91
63. Horner AA, Raz E. Immunostimulatory sequence oligodeoxynucleotide: A novel mucosal adjuvant. Clin Immunol. 2000; 95(1 Pt 2): S19-29
64. Hornung V, Rothenfusser S, Britsch S, Krug A, Jahrsdorfer B, Giese T, Endres S, Hartmann G. Quantitative expression of toll-like receptor 1-10 mRNA in cellular subsets of human peripheral blood mononuclear cells and sensitivity to CpG oligodeoxynucleotides. J Immunol. 2002; 168(9): 4531-7
65. Iho S, Yamamoto T, Takahashi T, Yamamoto S. Oligodeoxynucleotides containing palindrome sequences with internal 5'-CpG-3' act directly on human NK and activated T cells to induce IFN-gamma production in vitro. J Immunol. 1999; 163(7): 3642-52
66. Jacobs S, Grussendorf-Conen EI, Rosener I, Rubben A. Molecular analysis of the effect of topical imiquimod treatment of HPV 2/27/57-induced common warts. Skin Pharmacol Physiol. 2004; 17(5): 258-66
67. Jefferson T, Deeks JJ, Demicheli V, Rivetti D, Rudin M. Amantadine and rimantadine for preventing and treating influenza A in adults. Cochrane Database Syst Rev. 2004; (3): CD001169
68. Jeong SH, Qiao M, Nascimbeni M, Hu Z, Rehermann B, Murthy K, Liang TJ. Immunization with hepatitis C virus-like particles induces humoral and cellular immune responses in nonhuman primates. J Virol. 2004; 78(13): 6995-7003
69. Jiang JQ, Patrick A, Moss RB, Rosenthal KL. CD8+ T-cell-mediated cross-clade protection in the genital tract following intranasal immunization with inactivated human immunodeficiency virus antigen plus CpG oligodeoxynucleotides. J Virol. 2005 Jan;79(1):393-400
70. Jiao X, Wang RY, Qiu Q, Alter HJ, Shih JW. Enhanced hepatitis C virus NS3 specific Th1 immune responses induced by co-delivery of protein antigen and CpG with cationic liposomes. J Gen Virol. 2004; 85 (Pt 6): 1545-53
71. Jinushi M, Takehara T, Tatsumi T, Kanto T, Miyagi T, Suzuki T, Kanazawa Y, Hiramatsu N, Hayashi N. Negative regulation of NK cell activities by inhibitory receptor CD94/NKG2A leads to altered NK cell-induced modulation of dendritic cell functions in chronic hepatitis C virus infection. J Immunol. 2004; 173(10): 6072-81
72. Johnson DC, Hegde NR. Inhibition of the MHC class II antigen presentation pathway by human cytomegalovirus. Curr Top Microbiol Immunol. 2002; 269: 101-15
73. Joseph A, Louria-Hayon I, Plis-Finarov A, Zeira E, Zakay-Rones Z, Raz E, Hayashi T, Takabayashi K, Barenholz Y, Kedar E. Liposomal immunostimulatory DNA sequence (ISS-ODN): an efficient parenteral and mucosal adjuvant for influenza and hepatitis B vaccines. Vaccine. 2002; 20(27-28): 3342-54
74. Jozsef L, Khreiss T, Filep JG. CpG motifs in bacterial DNA delay apoptosis of neutrophil granulocytes. FASEB J. 2004; 18(14): 1776-8
75. Jung J, Yi AK, Zhang X, Choe J, Li L, Choi YS. Distinct response of human B cell subpopulations in recognition of an innate immune signal, CpG DNA. J Immunol. 2002; 169(5): 2368-73
76. Kadowaki N, Antonenko S, Liu YJ. Distinct CpG DNA and polyinosinic-polycytidylic acid double-stranded RNA, respectively, stimulate CD11c-type 2 dendritic cell precursors and CD11c+ dendritic cells to produce type I IFN. J Immunol. 2001; 166(4): 2291-5
77. Kadowaki N, Ho S, Antonenko S, Malefyt RW, Kastelein RA, Bazan F, Liu YJ. Subsets of human dendritic cell precursors express different toll-like receptors and respond to different microbial antigens. J Exp Med. 2001; 194(6): 863-9
78. Kaiserlian D, Dubois B. Dendritic cells and viral immunity: friends or foes? Semin Immunol. 2001; 13(5): 303-10
79. Kamstrup S, Verthelyi D, Klinman DM. Response of porcine peripheral blood mononuclear cells to CpG-containing oligodeoxynucleotides. Vet Microbiol. 2001; 78(4): 353-62
80. Kandimalla ER, Bhagat L, Cong YP, Pandey RK, Yu D, Zhao Q, Agrawal S. Secondary structures in CpG oligonucleotides affect immunostimulatory activity. Biochemical and Biophysical Research Communications. 2003; 306: 948–953
81. Kerkmann M, Rothenfusser S, Hornung V, Towarowski A, Wagner M, Sarris A, Giese T, Endres S, Hartmann G. Activation with CpG-A and CpG-B Oligonucleotides Reveals Two Distinct Regulatory Pathways of Type I IFN Synthesis in Human Plasmacytoid Dendritic Cells. J Immunol. 2003; 170(9): 4465-74
82. Kleymann G. Agents and strategies in development for improved management of herpes simplex virus infection and disease. Expert Opin Investig Drugs. 2005; 14(2): 135-61
83. Klinman DM. Immunotherapeutic uses of CpG oligodeoxynucleotides. Nat Rev Immunol. 2004; 4(4): 249-58
84. Klinman DM, Currie D. Hierarchical recognition of CpG motifs expressed by immunostimulatory oligodeoxynucleotides. Clin Exp Immunol. 2003; 133: 227–232
85. Kohl S, Thomas JW, Loo LS. Defective production of anti-herpes simplex virus antibody by neonatal mice. Reconstitution with Ia+ macrophages and T helper lymphocytes from nonimmune adult syngeneic mice. J Immunol. 1986; 136(8): 3038-44
86. Kopf M, Baumann H, Freer G, Freudenberg M, Lamers M, Kishimoto T, Zinkernagel R, Bluethmann H, Kohler G. Impaired immune and acute-phase responses in interleukin-6-deficient mice. Nature. 1994; 368(6469): 339-42
87. Kotwal GJ. Poxviral mimicry of complement and chemokine system components: what's the end game? Immunol Today. 2000; 21(5): 242-8
88. Kranzer K, Bauer M, Lipford GB, Heeg K, Wagner H, Lang R. CpG-oligodeoxynucleotides enhance T-cell receptor-triggered interferon-gamma production and up-regulation of CD69 via induction of antigen-presenting cell-derived interferon type I and interleukin-12. Immunology. 2000; 99(2): 170-8
89. Krieg AM. CpG motifs in bacterial DNA and their immune effects. Annu Rev Immunol. 2002; 20: 709-60
90. Krieg AM, Yi AK, Matson S, Waldschmidt TJ, Bishop GA, Teasdale R, Koretzky GA, Klinman DM. CpG motifs in bacterial DNA trigger direct B-cell activation. Nature. 1995; 374: 546-549
91. Krug A, Rothenfusser S, Hornung V, Jahrsdorfer B, Blackwell S, Ballas ZK, Endres S, Krieg AM, Hartmann G. Identification of CpG oligonucleotide sequences with high induction of IFN-α/βin plasmacytoid dendritic cells. Eur. J. Immunol. 2001; 31: 2154-2163
92. Krug A, Rothenfusser S, Selinger S, Bock C, Kerkmann M, Battiany J, Sarris A, Giese T, Speiser D, Endres S, Hartmann G. CpG-A oligonucleotides induce a monocyte-derived dendritic cell-like phenotype that preferentially activates CD8 T cells. J Immunol. 2003; 170(7): 3468-77
93. Krug A, Towarowski A, Britsch S, Rothenfusser S, Hornung V, Bals R, Giese T, Engelmann H, Endres S, Krieg AM, Hartmann G. Toll-like receptor expression reveals CpG DNA as a unique microbial stimulus for plasmacytoid dendritic cells which synergizes with CD40 ligand to induce high amounts of IL-12. Eur J Immunol. 2001; 31(10): 3026-37
94. Latz E, Schoenemeyer A, Visintin A, Fitzgerald KA, Monks BG, Knetter CF, Lien E, Nilsen NJ, Espevik T, Golenbock DT. TLR9 signals after translocating from the ER to CpG DNA in the lysosome. Nat Immunol. 2004; 5(2): 190-8
95. Le Bon A, Tough DF. Links between innate and adaptive immunity via type I interferon. Curr Opin Immunol. 2002; 14(4): 432-6
96. Lee SW, Song MK, Baek KH, Park Y, Kim JK, Lee CH, Cheong HK, Cheong C, Sung YC. Effects of a hexameric deoxyriboguanosine run conjugation into CpG oligodeoxynucleotides on their immunostimulatory potentials. J Immunol. 2000; 165(7): 3631-9
97. Leifer CA, Verthelyi D, Klinman DM. Heterogeneity in the human response to immunostimulatory CpG oligodeoxynucleotides. J Immunother. 2003; 26(4): 313-9
98. Leydet A, Moullet C, Roque JP, Witvrouw M, Pannecouque C, Andrei G, Snoeck R, Neyts J, Schols D, De Clercq E. Polyanion inhibitors of HIV
and other viruses. 7. Polyanionic compounds and polyzwitterionic
compounds derived from cyclodextrins as inhibitors of HIV
transmission. J Med Chem. 1998; 41(25): 4927-32
99. Li S, Labrecque S, Gauzzi MC, Cuddihy AR, Wong AH, Pellegrini S, Matlashewski GJ, Koromilas AE. The human papilloma virus (HPV)-18 E6 oncoprotein physically associates with Tyk2 and impairs Jak-STAT activation by interferon-alpha. Oncogene. 1999; 18(42): 5727-37
100. Liang H, Reich CF, Pisetsky DS, Lipsky PE. The Role of Surface Ig Binding in the Activation of Human B Cells by Phosphorothioate Oligodeoxynucleotides. Scand J Immunol. 2001; 54: 551-563
101. Liu B, Mori I, Hossain MJ, Dong L, Takeda K, Kimura Y. Interleukin-18 improves the early defence system against influenza virus infection by augmenting natural killer cell-mediated cytotoxicity. J Gen Virol. 2004; 85(Pt 2): 423-8
102. Malmgaard L. Induction and regulation of IFNs during viral infections. J Interferon Cytokine Res. 2004; 24(8): 439-54
103. Manzel L, Macfarlane DE. Lack of immune stimulation by immobilized CpG-oligodeoxynucleotide. Antisense Nucleic Acid Drug Dev. 1999; 9(5): 459-64
104. Mao QG, Luo KX, Zhang MX, Liu D, Hou JL. Generation of neutralizing anti-interferon antibodies and factors influencing their production in chronic hepatitis B patients receiving recombinant interferon-alpha treatment Di Yi Jun Yi Da Xue Xue Bao. 2003; 23(12): 1319-22
105. Marshall JD, Fearon K, Abbate C, Subramanian S, Yee P, Gregorio J, Coffman RL, Van Nest G. Identification of a novel CpG DNA class and motif that optimally stimulate B cell and plasmacytoid dendritic cell functions. J Leukoc Biol. 2003; 73(6): 781-92
106. McMichael AJ. Environmental and social influences on emerging infectious diseases: past, present and future. Philos Trans R Soc Lond B Biol Sci. 2004; 359(1447): 1049-58
107. Messina JP, Gilkeson GS, Pisetsky DS. Stimulation of in vitro murine lymphocyte proliferation by bacterial DNA. J Immunol. 1991; 147(6): 1759-64.
108. Miller DM, Cebulla CM, Sedmak DD. Human cytomegalovirus inhibition of major histocompatibility complex transcription and interferon signal transduction. Curr Top Microbiol Immunol. 2002; 269: 153-70
109. Moldoveanu Z, Love-Homan L, Huang WQ, Krieg AM. CpG DNA, a novel immune enhancer for systemic and mucosal immunization with influenza virus. Vaccine. 1998; 16(11-12): 1216-24
110. Mutwiri G, Pontarollo R, Babiuk S, Griebel P, van Drunen Littel-van den Hurk S, Mena A, Tsang C, Alcon V, Nichani A, Ioannou X, Gomis S, Townsend H, Hecker R, Potter A, Babiuk LA. Biological activity of immunostimulatory CpG DNA motifs in domestic animals. Vet Immunol Immunopathol. 2003; 91(2): 89-103
111. Muzio M, Bosisio D, Polentarutti N, D'amico G, Stoppacciaro A, Mancinelli R, van't Veer C, Penton-Rol G, Ruco LP, Allavena P, Mantovani A. Differential expression and regulation of toll-like receptors (TLR) in human leukocytes: selective expression of TLR3 in dendritic cells. J
Immunol. 2000; 164(11): 5998-6004
112. Neyts J, Reymen D, Letourneur D, Jozefonvicz J, Schols D, Este J, Andrei G, McKenna P, Witvrouw M, Ikeda S, et al. Differential antiviral activity of derivatized dextrans. Biochem Pharmacol. 1995; 50(6): 743-51
113. Nichani AK, Kaushik RS, Mena A, Popowych Y, Dent D, Townsend HG, Mutwiri G, Hecker R, Babiuk LA, Griebel PJ. CpG oligodeoxynucleotide induction of antiviral effector molecules in sheep. Cell Immunol. 2004; 227(1): 24-37
114. Nicholson KG, Wood JM, Zambon M. Influenza. Lancet. 2003; 362(9397): 1733-45
115. Nomura H, Tanimoto H, Kajiwara E, Shimono J, Maruyama T, Yamashita N, Nagano M, Higashi M, Mukai T, Matsui Y, Hayashi J, Kashiwagi S, Ishibashi H. Factors contributing to ribavirin-induced anemia. J Gastroenterol Hepatol. 2004; 19(11): 1312-7
116. Oaks JL, McGuire TC, Ulibarri C, Crawford TB. Equine infectious anemia virus is found in tissue macrophages during subclinical infection. J Virol. 1998; 72(9): 7263-9
117. Olbrich AR, Schimmer S, Dittmer U. Preinfection treatment of resistant mice with CpG oligodeoxynucleotides renders them susceptible to friend retrovirus-induced leukemia. J Virol. 2003; 77(19): 10658-62
118. Olbrich AR, Schimmer S, Heeg K, Schepers K, Schumacher TN, Dittmer U. Effective postexposure treatment of retrovirus-induced disease with immunostimulatory DNA containing CpG motifs. J Virol. 2002; 76(22): 11397-404
119. Ozato K, Tsujimura H, Tamura T. Toll-like receptor signaling and regulation of cytokine gene expression in the immune system. Biotechniques 2002; 66-68,70,72
120. Oxford JS. Influenza A pandemics of the 20th century with special reference to 1918: virology, pathology and epidemiology. Rev Med Virol. 2000; 10(2): 119-33
121. Park W, Keeffe EB. Diagnosis and treatment of chronic hepatitis B. Minerva Gastroenterol Dietol. 2004; 50(4): 289-303
122. Park Y, Lee SW, Sung YC. Cutting Edge: CpG DNA inhibits dendritic cell apoptosis by up-regulating cellular inhibitor of apoptosis proteins through the phosphatidylinositide-3'-OH kinase pathway. J Immunol. 2002; 168(1): 5-8
123. Pestka S, Krause CD, Walter MR. Interferons, interferon-like cytokines, and their receptors. Immunol Rev. 2004; 202: 8-32
124. Pfeffer K, Matsuyama T, Kundig TM, Wakeham A, Kishihara K, Shahinian A, Wiegmann K, Ohashi PS, Kronke M, Mak TW. Mice deficient for the 55 kd tumor necrosis factor receptor are resistant to endotoxic shock, yet succumb to L. monocytogenes infection. Cell. 1993; 73(3): 457-67
125. Pyles RB, Higgins D, Chalk C, Zalar A, Eiden J, Brown C, Van Nest G, Stanberry LR. Use of immunostimulatory sequence-containing oligonucleotides as topical therapy for genital herpes simplex virus type 2 infection. J Virol. 2002; 76(22): 11387-96
126. Qiao M, Murata K, Davis AR, Jeong SH, Liang TJ. Hepatitis C virus-like particles combined with novel adjuvant systems enhance virus-specific immune responses. Hepatology. 2003; 37(1): 52-9
127. Rehermann B, Nascimbeni M. Immunology of hepatitis B virus and hepatitis C virus infection. Nat Rev Immunol. 2005; 5(3): 215-29
128. Ronco LV, Karpova AY, Vidal M, Howley PM. Human papillomavirus 16 E6 oncoprotein binds to interferon regulatory factor-3 and inhibits its transcriptional activity. Genes Dev. 1998; 12(13): 2061-72
129. Rossol S, Marinos G, Carucci P, Singer MV, Williams R, Naoumov NV. Interleukin-12 induction of Th1 cytokines is important for viral clearance in chronic hepatitis B. J Clin Invest. 1997; 99(12): 3025-33
130. Rothenfusser S, Hornung V, Krug A, Towarowski A, Krieg AM, Endres S, Hartmann G. Distinct CpG oligonucleotide sequences activate human gamma delta T cells via interferon-alpha/-beta. Eur J Immunol. 2001; 31(12): 3525-34.
131. Schattenberg D, Schott M, Reindl G, Krueger T, Tschoepe D, Feldkamp J, Scherbaum WA, Seissler J. Response of human monocyte-derived dendritic cells to immunostimulatory DNA. Eur J Immunol. 2000; 30(10): 2824-31
132. Sha Q, Truong-Tran AQ, Plitt JR, Beck LA, Schleimer RP. Activation of airway epithelial cells by toll-like receptor agonists. Am J Respir Cell Mol Biol. 2004; 31(3): 358-64
133. Siegal FP, Kadowaki N, Shodell M, Fitzgerald-Bocarsly PA, Shah K, Ho S, Antonenko S, Liu YJ. The nature of the principal type 1 interferon-producing cells in human blood. Science. 1999; 284(5421): 1835-7
134. Sikorski M, Zrubek H. Long-term follow-up of patients treated with recombinant human interferon gamma for cervical intraepithelial neoplasia. Int J Gynaecol Obstet. 2003; 82(2): 179-85
135. Silvera P, Savary JR, Livingston V, White J, Manson KH, Wyand MH, Salk PL, Moss RB, Lewis MG. Vaccination with gp120-depleted HIV-1 plus immunostimulatory CpG oligodeoxynucleotides in incomplete Freund's adjuvant stimulates cellular and humoral immunity in rhesus macaques. Vaccine. 2004; 23(6): 827-39
136. Skowronski DM, Astell C, Brunham RC, Low DE, Petric M, Roper RL, Talbot PJ, Tam T, Babiuk L. Severe acute respiratory syndrome (SARS): a year in review. Annu Rev Med. 2005; 56: 357-81
137. Sparwasser T, Koch ES, Vabulas RM, Heeg K, Lipford GB, Ellwart JW, Wagner H. Bacterial DNA and immunostimulatory CpG oligonucleotides trigger maturation and activation of murine dendritic cells. Eur J Immunol. 1998; 28(6): 2045-54
138. Sun J, Wang Z, Ma S, Zeng G, Zhou Z, Luo K, Hou J. Clinical and virological characteristics of lamivudine resistance in chronic hepatitis B patients: a single center experience. J Med Virol. 2005; 75(3): 391-8
139. Sun S, Zhang X, Tough D, Sprent J. Multiple effects of immunostimulatory DNA on T cells and the role of type I interferons. Springer Semin Immunopathol. 2000; 22(1-2): 77-84
140. Svelander L, Erlandsson Harris H, Lorentzen JC, Trollmo C, Klareskog L, Bucht A. Oligodeoxynucleotides containing CpG motifs can induce T cell-dependent arthritis in rats. Arthritis Rheum. 2004 Jan;50(1):297-304
141. Takeshita A, Imai K, Hanazawa S. CpG motifs in Porphyromonas gingivalis DNA stimulate interleukin-6 expression in human gingival
fibroblasts. Infect Immun. 1999; 67(9): 4340-5
142. Takeshita F, Gursel I, Ishii KJ, Suzuki K, Gursel M, Klinman DM. Signal transduction pathways mediated by the interaction of CpG DNA with Toll-like receptor 9. Semin Immunol. 2004; 16(1): 17-22
143. Talon J, Horvath CM, Polley R, Basler CF, Muster T, Palese P, Garcia-Sastre A. Activation of interferon regulatory factor 3 is inhibited by the influenza A virus NS1 protein. J Virol. 2000; 74(17): 7989-96
144. Thimme R, Bukh J, Spangenberg HC, Wieland S, Pemberton J, Steiger C, Govindarajan S, Purcell RH, Chisari FV. Viral and immunological determinants of hepatitis C virus clearance, persistence, and disease. Proc Natl Acad Sci U S A. 2002; 99(24): 15661-8
145. Tokunaga T, Yamamoto H, Shimada S, Abe H., Fukuda T, Fujisawa Y, Furutani Y, Yano O, Kataoka T, Sudo T, Makiguchi N and Suganuma T. Antitumor activity of deoxyribonucleic acid fraction from Mycobacterium bovis BCG. L. Isolation, physicochemical characterization, and antitimor activity. J. Natl. Cancer Inst., 1984; 72: 955-62
146. Tortorella D, Gewurz BE, Furman MH, Schust DJ, Ploegh HL. Viral subversion of the immune system. Annu Rev Immunol. 2000; 18: 861-926
147. Trevani AS, Chorny A, Salamone G, Vermeulen M, Gamberale R, Schettini J, Raiden S, Geffner J. Bacterial DNA activates human neutrophils by a CpG-independent pathway. Eur J Immunol. 2003; 33(11): 3164-74
148. Tsai SL, Sheen IS, Chien RN, Chu CM, Huang HC, Chuang YL, Lee TH, Liao SK, Lin CL, Kuo GC, Liaw YF. Activation of Th1 immunity is a common immune mechanism for the successful treatment of hepatitis B and C: tetramer assay and therapeutic implications. J Biomed Sci. 2003; 10(1): 120-35
149. Tseng CT, Klimpel GR. Binding of the hepatitis C virus envelope protein E2 to CD81 inhibits natural killer cell functions. J Exp Med. 2002; 195(1): 43-9
150. Turner RB, Felton A, Kosak K, Kelsey DK, Meschievitz CK. Prevention of experimental coronavirus colds with intranasal alpha-2b interferon. J Infect Dis. 1986; 154(3): 443-7
151. Verani A, Gras G, Pancino G. Macrophages and HIV-1: dangerous liaisons. Mol Immunol. 2005; 42(2): 195-212
152. Verthelyi D, Ishii KJ, Gursel M, Takeshita F, Klinman DM. Human peripheral blood cells differentially recognize and respond to two distinct CpG motifs. J Immunol. 2001; 166(4): 2372-7
153. Verthelyi D, Klinman DM. Immunoregulatory activity of CpG oligonucleotides in humans and nonhuman primates. Clin Immunol. 2003; 109(1): 64-71
154. Vollmer J, Weeratna R, Payette P, Jurk M, Schetter C, Laucht M, Wader T, Tluk S, Liu M, Davis HL, Krieg AM. Characterization of three CpG oligodeoxynucleotide classes with distinct immunostimulatory activities. Eur. J. Immunol. 2004; 34: 251–262
155. Wagner M, Poeck H, Jahrsdoerfer B, Rothenfusser S, Prell D, Bohle B, Tuma E, Giese T, Ellwart JW, Endres S, Hartmann G. IL-12p70-dependent Th1 induction by human B cells requires combined activation with CD40 ligand and CpG DNA. J Immunol. 2004; 172(2): 954-63
156. Weeratna RD, McCluskie MJ, Xu Y, Davis HL. CpG DNA induces stronger immune responses with less toxicity than other adjuvants. Vaccine. 2000; 18(17): 1755-62
157. Weighardt H, Feterowski C, Veit M, Rump M, Wagner H, Holzmann B. Increased resistance against acute polymicrobial sepsis in mice challenged with immunostimulatory CpG oligodeoxynucleotides is related to an enhanced innate effector cell response. J Immunol. 2000; 165(8): 4537-43
158. Weiner GJ. The immunobiology and clinical potential of immunostimulatory CpG oligodeoxynucleotides. J Leukoc Biol. 2000; 68(4): 455-63
159. Wenzel J, Uerlich M, Haller O, Bieber T, Tueting T. Enhanced type I interferon signaling and recruitment of chemokine receptor CXCR3-expressing lymphocytes into the skin following treatment with the TLR7-agonist imiquimod. J Cutan Pathol. 2005; 32(4): 257-62
160. Wieland S, Thimme R, Purcell RH, Chisari FV. Genomic analysis of the host response to hepatitis B virus infection. Proc Natl Acad Sci U S A. 2004; 101(17): 6669-74
161. 许正锯,王崇国,李旭红,杨红,张启华。干扰素抗体与干扰素α-2a 治疗慢性乙型肝炎疗效的关系。中华传染病杂志。1998;16(3):26-27
162. Yamamoto S, Kuramoto E, Shimada S, Tokunaga T. In vitro augmentation of natural killer cell activity and production of interferon-alpha/beta and -gamma with deoxyribonucleic acid fraction from Mycobacterium bovis BCG. Jpn J Cancer Res. 1988; 79(7): 866-73
163. Yamamoto S, Yamamoto T, Kataoka T, Kuramoto E, Yano O, Tokunaga T. Unique palindromic sequences in synthetic oligonucleotides are required to induce IFN [correction of INF] and augment IFN-mediated [correction of INF] natural killer activity. J Immunol. 1992; 148(12):4072-6
164. Yamamoto T, Yamamoto S, Kataoka T, Tokunaga T. Lipofection of synthetic oligodeoxyribonucleotide having a palindromic sequence of AACGTT to murine splenocytes enhances interferon production and natural killer activity. Microbiol. Immunol. 1994; 38(10): 831–836
165. Yi AK, Klinman DM, Martin TL, Matson S, Krieg AM. Rapid immune activation by CpG motifs in bacterial DNA. Systemic induction of IL-6 transcription through an antioxidant-sensitive pathway. J Immunol. 1996; 157(12): 5394-402
166. Yi AK, Yoon JG, Krieg AM. Convergence of CpG DNA-and BCR-mediated signals at the c-Jun N-terminal kinase and NF-kappaB activation pathways: regulation by mitogen-activated protein kinases. Int Immunol. 2003; 15(5): 577-91
167. Yu D, Zhu FG, Bhagat L, Wang H, Kandimalla ER, Zhang R, Agrawal S. Potent CpG oligonucleotides containing phosphodiester linkages: in vitro and in vivo immunostimulatory properties. Biochem Biophys Res Commun. 2002; 297(1): 83-90
168. Yu H, Babiuk LA, van Drunen Littel-van den Hurk S. Priming with CpG-enriched plasmid and boosting with protein formulated with CpG
2. Ahn K, Meyer TH, Uebel S, Sempe P, Djaballah H, Yang Y, Peterson PA, Fruh K, Tampe R. Molecular mechanism and species specificity of TAP inhibition by herpes simplex virus ICP47. EMBO J. 1996; 15(13): 3247-55
3. Ahn KS, Ou W, Silver J. Inhibition of certain strains of HIV-1 by cell surface polyanions in the form of cholesterol-labeled oligonucleotides. Virology. 2004; 330(1): 50-61
4. Akira S, Takeda K. Functions of Toll-like receptors: lessons from KO mice. C. R. Biologies. 2004; 327: 581–589
5. Akira S, Takeda K. Toll-like receptor signalling. Nat Rev Immunol. 2004; 4(7): 499-511
6. Balamurugan V, Kumar RM, Suryanarayana VV. Past and present vaccine development strategies for the control of foot-and-mouth disease. Acta Virol. 2004; 48(4): 201-14
7. Ballas ZK, Krieg AM, Warren T, Rasmussen W, Davis HL, Waldschmidt M, Weiner GJ. Divergent therapeutic and immunologic effects of oligodeoxynucleotides with distinct CpG motifs. J Immunol. 2001; 167(9):4878-86
8. Ballas ZK, Rasmussen WL, Krieg AM. Induction of NK Activity in Murine and Human Cells by CpG Motifs in Oligodeoxynucleotides and Bacterial DNA. J Immunol. 1996; 157: 1840-1845
9. Becker Y. A point of view: HIV-1/AIDS is an allergy but CpG ODN treatments may inhibit virus replication and reactivate the adaptive immunity--hypothesis and implications. Virus Genes. 2005; 30(1): 127-31
10. Becker Y. CpG ODNs Treatments of HIV-1 Infected Patients May Cause the Decline of Transmission in High Risk Populations -A Review, Hypothesis and Implications. Virus Genes. 2005; 30(2): 251-66
11. Bird AP. CpG-rich islands and the function of DNA methylation. Nature. 1986; 321(6067): 209–213
12. Biron CA, Nguyen KB, Pien GC, Cousens LP, Salazar-Mather TP. Natural killer cells in antiviral defense: function and regulation by innate cytokines. Annu Rev Immunol. 1999; 17: 189-220
13. Bjersing JL, Tarkowski A, Collins LV. Anti-proliferative effects of phosphodiester oligodeoxynucleotides. Immunobiology. 2004; 209(8): 637-45
14. Blackwell SE and Krieg AM. CpG-A-Induced Monocyte IFN-γInducible Protein-10 Production Is Regulated by Plasmacytoid Dendritic Cell-Derived IFN-α. J Immunol. 2003; 170: 4061–4068
15. Bobardt MD, Armand-Ugon M, Clotet I, Zhang Z, David G, Este JA, Gallay PA. Effect of polyanion-resistance on HIV-1 infection. Virology. 2004; 325(2): 389-98
16. Brazolot Millan CL, Weeratna R, Krieg AM, Siegrist CA, Davis HL. CpG DNA can induce strong Th1 humoral and cell-mediated immune responses against hepatitis B surface antigen in young mice. Proc Natl Acad Sci U S A. 1998; 95(26): 15553-8
17. Brierley MM, Fish EN. Review: IFN-alpha/beta receptor interactions to biologic outcomes: understanding the circuitry. J Interferon Cytokine Res. 2002; 22(8): 835-45
18. Cella M, Jarrossay D, Facchetti F, Alebardi O, Nakajima H, Lanzavecchia A, Colonna M. Plasmacytoid monocytes migrate to inflamed lymph nodes and produce large amounts of type I interferon. Nat Med. 1999; 5(8): 919-23
19. Chadha KC, Dembinski WE, Dunn CB, Aradi J, Bardos TJ, Dunn JA, Ambrus JL Sr. Effect of increasing thiolation of the polycytidylic acid strand of poly I:poly C on the alpha, beta and gamma interferon-inducing properties, antiviral and antiproliferative activities. Antiviral Res. 2004; 64(3): 171-7
20. Cho JY, Miller M, Baek KJ, Castaneda D, Nayar J, Roman M, Raz E, Broide DH. Immunostimulatory DNA sequences inhibit respiratory syncytial viral load, airway inflammation, and mucus secretion. J Allergy Clin Immunol. 2001; 108(5): 697-702
21. Choi YK, Ozaki H, Webby RJ, Webster RG, Peiris JS, Poon L, Butt C, Leung YH, Guan Y. Continuing evolution of H9N2 influenza viruses in Southeastern China. J Virol. 2004; 78(16): 8609-14
22. Cocchi F, DeVico AL, Garzino-Demo A, Arya SK, Gallo RC, Lusso P. Identification of RANTES, MIP-1 alpha, and MIP-1 beta as the major HIV-suppressive factors produced by CD8+ T cells. Science. 1995; 270(5243): 1811-5
23. Coccia EM, Severa M, Giacomini E, Monneron D, Remoli ME, Julkunen I, Cella M, Lande R, Uze G. Viral infection and Toll-like receptor agonists induce a differential expression of type I and lambda interferons in human plasmacytoid and monocyte-derived dendritic cells. Eur J Immunol. 2004; 34(3): 796-805
24. Cooper CL, Davis HL, Morris ML, Efler SM, Krieg AM, Li Y, Laframboise C, Al Adhami MJ, Khaliq Y, Seguin I, Cameron DW. Safety and immunogenicity of CPG 7909 injection as an adjuvant to Fluarix influenza vaccine. Vaccine. 2004; 22(23-24): 3136-43
25. Dahl H, Linde A, Strannegard O. In vitro inhibition of SARS virus replication by human interferons. Scand J Infect Dis. 2004; 36(11-12): 829-31
26. Davis HL, Weeratna R, Waldschmidt TJ, Tygrett L, Schorr J, Krieg AM. CpG DNA is a potent enhancer of specific immunity in mice immunized with recombinant hepatitis B surface antigen. J Immunol. 1998; 160(2): 870-6
27. 戴志澄,祁国明主编.中国病毒性肝炎血清流行病学调查(1992～1995)(上 卷).北京:科学技术文献出版社,1997
28. Dong L, Mori I, Hossain MJ, Liu B, Kimura Y. An immunostimulatory oligodeoxynucleotide containing a cytidine-guanosine motif protects senescence-accelerated mice from lethal influenza virus by augmenting the T helper type 1 response. J Gen Virol. 2003; 84(Pt 6): 1623-8
29. Dong QM, He ZP, Zhuang H, Song SJ, Dai WS, Zhang SP, Chen ZH, Sun JY. Dynamics of peripheral blood B lymphocytes and natural killer cells in patients with severe acute respiratory syndrome. Zhonghua Liu Xing Bing Xue Za Zhi. 2004; 25(8): 695-7
30. Duan XZ, Wang M, Li HW, Zhuang H, Xu D, Wang FS. Decreased frequency and function of circulating plasmocytoid dendritic cells (pDC) in hepatitis B virus infected humans. J Clin Immunol. 2004; 24(6): 637-46
31. Duong FH, Filipowicz M, Tripodi M, La Monica N, Heim MH. Hepatitis C virus inhibits interferon signaling through up-regulation of protein phosphatase 2A. Gastroenterology. 2004; 126(1): 263-77
32. Ebert S, Gerber J, Bader S, Muhlhauser F, Brechtel K, Mitchell TJ, Nau R. Dose-dependent activation of microglial cells by Toll-like receptor agonists alone and in combination. J Neuroimmunol. 2005; 159(1-2): 87-96
33. Encke J, zu Putlitz J, Stremmel W, Wands JR. CpG immuno-stimulatory motifs enhance humoral immune responses against hepatitis C virus core protein after DNA-based immunization. Arch Virol. 2003; 148(3): 435-48
34. Foy E, Li K, Wang C, Sumpter R Jr, Ikeda M, Lemon SM, Gale M Jr. Regulation of interferon regulatory factor-3 by the hepatitis C virus serine protease. Science. 2003; 300(5622): 1145-8
35. French AR, Yokoyama WM. Natural killer cells and viral infections. Curr Opin Immunol. 2003; 15(1): 45-51
36. Freigang S, Probst HC, van den Broek M. DC infection promotes antiviral CTL priming: the 'Winkelried' strategy. Trends Immunol. 2005; 26(1): 13-8
37. Gilbert MJ, Riddell SR, Plachter B, Greenberg PD. Cytomegalovirus selectively blocks antigen processing and presentation of its immediate-early gene product. Nature. 1996; 383(6602): 720-2
38. Gould MP, Greene JA, Bhoj V, DeVecchio JL, Heinzel FP. Distinct modulatory effects of LPS and CpG on IL-18-dependent IFN-gamma synthesis. J Immunol. 2004; 172(3): 1754-62
39. Gretch D. Mechanism of interferon resistance in hepatitis C. Lancet. 2001; 358(9294): 1662-4
40. Groneberg DA, Poutanen SM, Low DE, Lode H, Welte T, Zabel P. Treatment and vaccines for severe acute respiratory syndrome. Lancet Infect Dis. 2005; 5(3): 147-55
41. Gruhler A, Peterson PA, Fruh K. Human cytomegalovirus immediate early glycoprotein US3 retains MHC class I molecules by transient association. Traffic. 2000; 1(4): 318-25
42. Guidotti LG, Chisari FV. Noncytolytic control of viral infections by the innate and adaptive immune response. Annu Rev Immunol. 2001; 19: 65-91
43. Guidotti LG, Rochford R, Chung J, Shapiro M, Purcell R, Chisari FV. Viral clearance without destruction of infected cells during acute HBV infection. Science. 1999; 284(5415): 825-9
44. Gurney KB, Colantonio AD, Blom B, Spits H, Uittenbogaart CH. Endogenous IFN-alpha production by plasmacytoid dendritic cells
exerts an antiviral effect on thymic HIV-1 infection. J Immunol. 2004;
173(12): 7269-76
45. Gursel I, Gursel M, Ishii KJ, Klinman DM. Sterically stabilized cationic liposomes improve the uptake and immunostimulatory activity of CpG oligonucleotides. J Immunol. 2001; 167(6): 3324-8
46. Hacker H, Mischak H, Miethke T, Liptay S, Schmid R, Sparwasser T, Heeg K, Lipford GB, Wagner H. CpG-DNA-specific activation of antigen-presenting cells requires stress kinase activity and is preceded by non-specific endocytosis and endosomal maturation. EMBO J. 1998; 17(21): 6230-40
47. Hadziyannis SJ, Sette H Jr, Morgan TR, Balan V, Diago M, Marcellin P, Ramadori G, Bodenheimer H Jr, Bernstein D, Rizzetto M, Zeuzem S, Pockros PJ, Lin A, Ackrill AM; PEGASYS International Study Group. Peginterferon-alpha2a and ribavirin combination therapy in chronic hepatitis C: a randomized study of treatment duration and ribavirin dose. Ann Intern Med. 2004; 140(5): 346-55
48. Halperin SA, Van Nest G, Smith B, Abtahi S, Whiley H, Eiden JJ. A phase I study of the safety and immunogenicity of recombinant hepatitis B surface antigen co-administered with an immunostimulatory phosphorothioate oligonucleotide adjuvant. Vaccine. 2003; 21(19-20): 2461-7
49. Harandi AM, Eriksson K, Holmgren J. A Protective Role of Locally Administered Immunostimulatory CpG Oligodeoxynucleotide in a Mouse Model of Genital Herpes Infection. Journal of virology. 2003; 77(2): 953–962.
50. Hartmann G, Krieg AM. CpG DNA and LPS induce distinct patterns of activation in human monocytes. Gene Ther. 1999; 6(5): 893-903
51. Hartmann G, Krieg AM. Mechanism and function of a newly identified CpG DNA motif in human primary B cells. J Immunol. 2000; 164(2): 944-53
52. Hartmann G, Weeratna RD, Ballas ZK, Payette P, Blackwell S, Suparto I, Rasmussen WL, Waldschmidt M, Sajuthi D, Purcell RH, Davis HL, Krieg AM. Delineation of a CpG Phosphorothioate Oligodeoxynucleotide for Activating Primate Immune Responses In Vitro and In Vivo. J Immunol. 2000; 164: 1617–1624
53. Hayashi F, Means TK, Luster AD. Toll-like receptors stimulate human neutrophil function. Blood. 2003; 102(7): 2660-9
54. Hayashi M, Satou E, Ueki R, Yano M, Miyano-Kurosaki N, Fujii M, Takaku H. Resistance to influenza A virus infection by antigen-conjugated CpG oligonucleotides, a novel antigen-specific immunomodulator. Biochem Biophys Res Commun. 2005; 329(1): 230-6
55. He XS, Draghi M, Mahmood K, Holmes TH, Kemble GW, Dekker CL, Arvin AM, Parham P, Greenberg HB. T cell-dependent production of IFN-gamma by NK cells in response to influenza A virus. J Clin Invest. 2004; 114(12): 1812-9
56. Heikenwalder M, Polymenidou M, Junt T, Sigurdson C, Wagner H, Akira S, Zinkernagel R, Aguzzi A. Lymphoid follicle destruction and immunosuppression after repeated CpG oligodeoxynucleotide administration. Nat Med. 2004; 10(2): 187-92
57. Heil F, Hemmi H, Hochrein H, Ampenberger F, Kirschning C, Akira S, Lipford G, Wagner H, Bauer S. Species-specific recognition of single-stranded RNA via toll-like receptor 7 and 8. Science. 2004; 303(5663): 1526-9
58. Heller T, Saito S, Auerbach J, Williams T, Moreen TR, Jazwinski A, Cruz B, Jeurkar N, Sapp R, Luo G, Liang TJ. An in vitro model of hepatitis C virion production. Proc Natl Acad Sci U S A. 2005; 102(7): 2579-83
59. Hemmi H, Takeuchi O, Kawai T, Kaisho T, Sato S, Sanjo H, Matsumoto M, Hoshino K, Wagner H, Takeda K, Akira S. A Toll-like receptor recognizes bacterial DNA. Nature. 2000; 408(6813): 740-5
60. Hengel H, Brune W, Koszinowski UH. Immune evasion by cytomegalovirus--survival strategies of a highly adapted opportunist. Trends Microbiol. 1998; 6(5): 190-7
61. Higgins PG, Phillpotts RJ, Scott GM, Wallace J, Bernhardt LL, Tyrrell DA. Intranasal interferon as protection against experimental respiratory coronavirus infection in volunteers. Antimicrob Agents Chemother. 1983; 24(5): 713-5
62. Hofmann WP, Zeuzem S, Sarrazin C. Hepatitis C virus-related resistance mechanisms to interferon alpha-based antiviral therapy. J Clin Virol. 2005; 32(2): 86-91
63. Horner AA, Raz E. Immunostimulatory sequence oligodeoxynucleotide: A novel mucosal adjuvant. Clin Immunol. 2000; 95(1 Pt 2): S19-29
64. Hornung V, Rothenfusser S, Britsch S, Krug A, Jahrsdorfer B, Giese T, Endres S, Hartmann G. Quantitative expression of toll-like receptor 1-10 mRNA in cellular subsets of human peripheral blood mononuclear cells and sensitivity to CpG oligodeoxynucleotides. J Immunol. 2002; 168(9): 4531-7
65. Iho S, Yamamoto T, Takahashi T, Yamamoto S. Oligodeoxynucleotides containing palindrome sequences with internal 5'-CpG-3' act directly on human NK and activated T cells to induce IFN-gamma production in vitro. J Immunol. 1999; 163(7): 3642-52
66. Jacobs S, Grussendorf-Conen EI, Rosener I, Rubben A. Molecular analysis of the effect of topical imiquimod treatment of HPV 2/27/57-induced common warts. Skin Pharmacol Physiol. 2004; 17(5): 258-66
67. Jefferson T, Deeks JJ, Demicheli V, Rivetti D, Rudin M. Amantadine and rimantadine for preventing and treating influenza A in adults. Cochrane Database Syst Rev. 2004; (3): CD001169
68. Jeong SH, Qiao M, Nascimbeni M, Hu Z, Rehermann B, Murthy K, Liang TJ. Immunization with hepatitis C virus-like particles induces humoral and cellular immune responses in nonhuman primates. J Virol. 2004; 78(13): 6995-7003
69. Jiang JQ, Patrick A, Moss RB, Rosenthal KL. CD8+ T-cell-mediated cross-clade protection in the genital tract following intranasal immunization with inactivated human immunodeficiency virus antigen plus CpG oligodeoxynucleotides. J Virol. 2005 Jan;79(1):393-400
70. Jiao X, Wang RY, Qiu Q, Alter HJ, Shih JW. Enhanced hepatitis C virus NS3 specific Th1 immune responses induced by co-delivery of protein antigen and CpG with cationic liposomes. J Gen Virol. 2004; 85 (Pt 6): 1545-53
71. Jinushi M, Takehara T, Tatsumi T, Kanto T, Miyagi T, Suzuki T, Kanazawa Y, Hiramatsu N, Hayashi N. Negative regulation of NK cell activities by inhibitory receptor CD94/NKG2A leads to altered NK cell-induced modulation of dendritic cell functions in chronic hepatitis C virus infection. J Immunol. 2004; 173(10): 6072-81
72. Johnson DC, Hegde NR. Inhibition of the MHC class II antigen presentation pathway by human cytomegalovirus. Curr Top Microbiol Immunol. 2002; 269: 101-15
73. Joseph A, Louria-Hayon I, Plis-Finarov A, Zeira E, Zakay-Rones Z, Raz E, Hayashi T, Takabayashi K, Barenholz Y, Kedar E. Liposomal immunostimulatory DNA sequence (ISS-ODN): an efficient parenteral and mucosal adjuvant for influenza and hepatitis B vaccines. Vaccine. 2002; 20(27-28): 3342-54
74. Jozsef L, Khreiss T, Filep JG. CpG motifs in bacterial DNA delay apoptosis of neutrophil granulocytes. FASEB J. 2004; 18(14): 1776-8
75. Jung J, Yi AK, Zhang X, Choe J, Li L, Choi YS. Distinct response of human B cell subpopulations in recognition of an innate immune signal, CpG DNA. J Immunol. 2002; 169(5): 2368-73
76. Kadowaki N, Antonenko S, Liu YJ. Distinct CpG DNA and polyinosinic-polycytidylic acid double-stranded RNA, respectively, stimulate CD11c-type 2 dendritic cell precursors and CD11c+ dendritic cells to produce type I IFN. J Immunol. 2001; 166(4): 2291-5
77. Kadowaki N, Ho S, Antonenko S, Malefyt RW, Kastelein RA, Bazan F, Liu YJ. Subsets of human dendritic cell precursors express different toll-like receptors and respond to different microbial antigens. J Exp Med. 2001; 194(6): 863-9
78. Kaiserlian D, Dubois B. Dendritic cells and viral immunity: friends or foes? Semin Immunol. 2001; 13(5): 303-10
79. Kamstrup S, Verthelyi D, Klinman DM. Response of porcine peripheral blood mononuclear cells to CpG-containing oligodeoxynucleotides. Vet Microbiol. 2001; 78(4): 353-62
80. Kandimalla ER, Bhagat L, Cong YP, Pandey RK, Yu D, Zhao Q, Agrawal S. Secondary structures in CpG oligonucleotides affect immunostimulatory activity. Biochemical and Biophysical Research Communications. 2003; 306: 948–953
81. Kerkmann M, Rothenfusser S, Hornung V, Towarowski A, Wagner M, Sarris A, Giese T, Endres S, Hartmann G. Activation with CpG-A and CpG-B Oligonucleotides Reveals Two Distinct Regulatory Pathways of Type I IFN Synthesis in Human Plasmacytoid Dendritic Cells. J Immunol. 2003; 170(9): 4465-74
82. Kleymann G. Agents and strategies in development for improved management of herpes simplex virus infection and disease. Expert Opin Investig Drugs. 2005; 14(2): 135-61
83. Klinman DM. Immunotherapeutic uses of CpG oligodeoxynucleotides. Nat Rev Immunol. 2004; 4(4): 249-58
84. Klinman DM, Currie D. Hierarchical recognition of CpG motifs expressed by immunostimulatory oligodeoxynucleotides. Clin Exp Immunol. 2003; 133: 227–232
85. Kohl S, Thomas JW, Loo LS. Defective production of anti-herpes simplex virus antibody by neonatal mice. Reconstitution with Ia+ macrophages and T helper lymphocytes from nonimmune adult syngeneic mice. J Immunol. 1986; 136(8): 3038-44
86. Kopf M, Baumann H, Freer G, Freudenberg M, Lamers M, Kishimoto T, Zinkernagel R, Bluethmann H, Kohler G. Impaired immune and acute-phase responses in interleukin-6-deficient mice. Nature. 1994; 368(6469): 339-42
87. Kotwal GJ. Poxviral mimicry of complement and chemokine system components: what's the end game? Immunol Today. 2000; 21(5): 242-8
88. Kranzer K, Bauer M, Lipford GB, Heeg K, Wagner H, Lang R. CpG-oligodeoxynucleotides enhance T-cell receptor-triggered interferon-gamma production and up-regulation of CD69 via induction of antigen-presenting cell-derived interferon type I and interleukin-12. Immunology. 2000; 99(2): 170-8
89. Krieg AM. CpG motifs in bacterial DNA and their immune effects. Annu Rev Immunol. 2002; 20: 709-60
90. Krieg AM, Yi AK, Matson S, Waldschmidt TJ, Bishop GA, Teasdale R, Koretzky GA, Klinman DM. CpG motifs in bacterial DNA trigger direct B-cell activation. Nature. 1995; 374: 546-549
91. Krug A, Rothenfusser S, Hornung V, Jahrsdorfer B, Blackwell S, Ballas ZK, Endres S, Krieg AM, Hartmann G. Identification of CpG oligonucleotide sequences with high induction of IFN-α/βin plasmacytoid dendritic cells. Eur. J. Immunol. 2001; 31: 2154-2163
92. Krug A, Rothenfusser S, Selinger S, Bock C, Kerkmann M, Battiany J, Sarris A, Giese T, Speiser D, Endres S, Hartmann G. CpG-A oligonucleotides induce a monocyte-derived dendritic cell-like phenotype that preferentially activates CD8 T cells. J Immunol. 2003; 170(7): 3468-77
93. Krug A, Towarowski A, Britsch S, Rothenfusser S, Hornung V, Bals R, Giese T, Engelmann H, Endres S, Krieg AM, Hartmann G. Toll-like receptor expression reveals CpG DNA as a unique microbial stimulus for plasmacytoid dendritic cells which synergizes with CD40 ligand to induce high amounts of IL-12. Eur J Immunol. 2001; 31(10): 3026-37
94. Latz E, Schoenemeyer A, Visintin A, Fitzgerald KA, Monks BG, Knetter CF, Lien E, Nilsen NJ, Espevik T, Golenbock DT. TLR9 signals after translocating from the ER to CpG DNA in the lysosome. Nat Immunol. 2004; 5(2): 190-8
95. Le Bon A, Tough DF. Links between innate and adaptive immunity via type I interferon. Curr Opin Immunol. 2002; 14(4): 432-6
96. Lee SW, Song MK, Baek KH, Park Y, Kim JK, Lee CH, Cheong HK, Cheong C, Sung YC. Effects of a hexameric deoxyriboguanosine run conjugation into CpG oligodeoxynucleotides on their immunostimulatory potentials. J Immunol. 2000; 165(7): 3631-9
97. Leifer CA, Verthelyi D, Klinman DM. Heterogeneity in the human response to immunostimulatory CpG oligodeoxynucleotides. J Immunother. 2003; 26(4): 313-9
98. Leydet A, Moullet C, Roque JP, Witvrouw M, Pannecouque C, Andrei G, Snoeck R, Neyts J, Schols D, De Clercq E. Polyanion inhibitors of HIV
and other viruses. 7. Polyanionic compounds and polyzwitterionic
compounds derived from cyclodextrins as inhibitors of HIV
transmission. J Med Chem. 1998; 41(25): 4927-32
99. Li S, Labrecque S, Gauzzi MC, Cuddihy AR, Wong AH, Pellegrini S, Matlashewski GJ, Koromilas AE. The human papilloma virus (HPV)-18 E6 oncoprotein physically associates with Tyk2 and impairs Jak-STAT activation by interferon-alpha. Oncogene. 1999; 18(42): 5727-37
100. Liang H, Reich CF, Pisetsky DS, Lipsky PE. The Role of Surface Ig Binding in the Activation of Human B Cells by Phosphorothioate Oligodeoxynucleotides. Scand J Immunol. 2001; 54: 551-563
101. Liu B, Mori I, Hossain MJ, Dong L, Takeda K, Kimura Y. Interleukin-18 improves the early defence system against influenza virus infection by augmenting natural killer cell-mediated cytotoxicity. J Gen Virol. 2004; 85(Pt 2): 423-8
102. Malmgaard L. Induction and regulation of IFNs during viral infections. J Interferon Cytokine Res. 2004; 24(8): 439-54
103. Manzel L, Macfarlane DE. Lack of immune stimulation by immobilized CpG-oligodeoxynucleotide. Antisense Nucleic Acid Drug Dev. 1999; 9(5): 459-64
104. Mao QG, Luo KX, Zhang MX, Liu D, Hou JL. Generation of neutralizing anti-interferon antibodies and factors influencing their production in chronic hepatitis B patients receiving recombinant interferon-alpha treatment Di Yi Jun Yi Da Xue Xue Bao. 2003; 23(12): 1319-22
105. Marshall JD, Fearon K, Abbate C, Subramanian S, Yee P, Gregorio J, Coffman RL, Van Nest G. Identification of a novel CpG DNA class and motif that optimally stimulate B cell and plasmacytoid dendritic cell functions. J Leukoc Biol. 2003; 73(6): 781-92
106. McMichael AJ. Environmental and social influences on emerging infectious diseases: past, present and future. Philos Trans R Soc Lond B Biol Sci. 2004; 359(1447): 1049-58
107. Messina JP, Gilkeson GS, Pisetsky DS. Stimulation of in vitro murine lymphocyte proliferation by bacterial DNA. J Immunol. 1991; 147(6): 1759-64.
108. Miller DM, Cebulla CM, Sedmak DD. Human cytomegalovirus inhibition of major histocompatibility complex transcription and interferon signal transduction. Curr Top Microbiol Immunol. 2002; 269: 153-70
109. Moldoveanu Z, Love-Homan L, Huang WQ, Krieg AM. CpG DNA, a novel immune enhancer for systemic and mucosal immunization with influenza virus. Vaccine. 1998; 16(11-12): 1216-24
110. Mutwiri G, Pontarollo R, Babiuk S, Griebel P, van Drunen Littel-van den Hurk S, Mena A, Tsang C, Alcon V, Nichani A, Ioannou X, Gomis S, Townsend H, Hecker R, Potter A, Babiuk LA. Biological activity of immunostimulatory CpG DNA motifs in domestic animals. Vet Immunol Immunopathol. 2003; 91(2): 89-103
111. Muzio M, Bosisio D, Polentarutti N, D'amico G, Stoppacciaro A, Mancinelli R, van't Veer C, Penton-Rol G, Ruco LP, Allavena P, Mantovani A. Differential expression and regulation of toll-like receptors (TLR) in human leukocytes: selective expression of TLR3 in dendritic cells. J
Immunol. 2000; 164(11): 5998-6004
112. Neyts J, Reymen D, Letourneur D, Jozefonvicz J, Schols D, Este J, Andrei G, McKenna P, Witvrouw M, Ikeda S, et al. Differential antiviral activity of derivatized dextrans. Biochem Pharmacol. 1995; 50(6): 743-51
113. Nichani AK, Kaushik RS, Mena A, Popowych Y, Dent D, Townsend HG, Mutwiri G, Hecker R, Babiuk LA, Griebel PJ. CpG oligodeoxynucleotide induction of antiviral effector molecules in sheep. Cell Immunol. 2004; 227(1): 24-37
114. Nicholson KG, Wood JM, Zambon M. Influenza. Lancet. 2003; 362(9397): 1733-45
115. Nomura H, Tanimoto H, Kajiwara E, Shimono J, Maruyama T, Yamashita N, Nagano M, Higashi M, Mukai T, Matsui Y, Hayashi J, Kashiwagi S, Ishibashi H. Factors contributing to ribavirin-induced anemia. J Gastroenterol Hepatol. 2004; 19(11): 1312-7
116. Oaks JL, McGuire TC, Ulibarri C, Crawford TB. Equine infectious anemia virus is found in tissue macrophages during subclinical infection. J Virol. 1998; 72(9): 7263-9
117. Olbrich AR, Schimmer S, Dittmer U. Preinfection treatment of resistant mice with CpG oligodeoxynucleotides renders them susceptible to friend retrovirus-induced leukemia. J Virol. 2003; 77(19): 10658-62
118. Olbrich AR, Schimmer S, Heeg K, Schepers K, Schumacher TN, Dittmer U. Effective postexposure treatment of retrovirus-induced disease with immunostimulatory DNA containing CpG motifs. J Virol. 2002; 76(22): 11397-404
119. Ozato K, Tsujimura H, Tamura T. Toll-like receptor signaling and regulation of cytokine gene expression in the immune system. Biotechniques 2002; 66-68,70,72
120. Oxford JS. Influenza A pandemics of the 20th century with special reference to 1918: virology, pathology and epidemiology. Rev Med Virol. 2000; 10(2): 119-33
121. Park W, Keeffe EB. Diagnosis and treatment of chronic hepatitis B. Minerva Gastroenterol Dietol. 2004; 50(4): 289-303
122. Park Y, Lee SW, Sung YC. Cutting Edge: CpG DNA inhibits dendritic cell apoptosis by up-regulating cellular inhibitor of apoptosis proteins through the phosphatidylinositide-3'-OH kinase pathway. J Immunol. 2002; 168(1): 5-8
123. Pestka S, Krause CD, Walter MR. Interferons, interferon-like cytokines, and their receptors. Immunol Rev. 2004; 202: 8-32
124. Pfeffer K, Matsuyama T, Kundig TM, Wakeham A, Kishihara K, Shahinian A, Wiegmann K, Ohashi PS, Kronke M, Mak TW. Mice deficient for the 55 kd tumor necrosis factor receptor are resistant to endotoxic shock, yet succumb to L. monocytogenes infection. Cell. 1993; 73(3): 457-67
125. Pyles RB, Higgins D, Chalk C, Zalar A, Eiden J, Brown C, Van Nest G, Stanberry LR. Use of immunostimulatory sequence-containing oligonucleotides as topical therapy for genital herpes simplex virus type 2 infection. J Virol. 2002; 76(22): 11387-96
126. Qiao M, Murata K, Davis AR, Jeong SH, Liang TJ. Hepatitis C virus-like particles combined with novel adjuvant systems enhance virus-specific immune responses. Hepatology. 2003; 37(1): 52-9
127. Rehermann B, Nascimbeni M. Immunology of hepatitis B virus and hepatitis C virus infection. Nat Rev Immunol. 2005; 5(3): 215-29
128. Ronco LV, Karpova AY, Vidal M, Howley PM. Human papillomavirus 16 E6 oncoprotein binds to interferon regulatory factor-3 and inhibits its transcriptional activity. Genes Dev. 1998; 12(13): 2061-72
129. Rossol S, Marinos G, Carucci P, Singer MV, Williams R, Naoumov NV. Interleukin-12 induction of Th1 cytokines is important for viral clearance in chronic hepatitis B. J Clin Invest. 1997; 99(12): 3025-33
130. Rothenfusser S, Hornung V, Krug A, Towarowski A, Krieg AM, Endres S, Hartmann G. Distinct CpG oligonucleotide sequences activate human gamma delta T cells via interferon-alpha/-beta. Eur J Immunol. 2001; 31(12): 3525-34.
131. Schattenberg D, Schott M, Reindl G, Krueger T, Tschoepe D, Feldkamp J, Scherbaum WA, Seissler J. Response of human monocyte-derived dendritic cells to immunostimulatory DNA. Eur J Immunol. 2000; 30(10): 2824-31
132. Sha Q, Truong-Tran AQ, Plitt JR, Beck LA, Schleimer RP. Activation of airway epithelial cells by toll-like receptor agonists. Am J Respir Cell Mol Biol. 2004; 31(3): 358-64
133. Siegal FP, Kadowaki N, Shodell M, Fitzgerald-Bocarsly PA, Shah K, Ho S, Antonenko S, Liu YJ. The nature of the principal type 1 interferon-producing cells in human blood. Science. 1999; 284(5421): 1835-7
134. Sikorski M, Zrubek H. Long-term follow-up of patients treated with recombinant human interferon gamma for cervical intraepithelial neoplasia. Int J Gynaecol Obstet. 2003; 82(2): 179-85
135. Silvera P, Savary JR, Livingston V, White J, Manson KH, Wyand MH, Salk PL, Moss RB, Lewis MG. Vaccination with gp120-depleted HIV-1 plus immunostimulatory CpG oligodeoxynucleotides in incomplete Freund's adjuvant stimulates cellular and humoral immunity in rhesus macaques. Vaccine. 2004; 23(6): 827-39
136. Skowronski DM, Astell C, Brunham RC, Low DE, Petric M, Roper RL, Talbot PJ, Tam T, Babiuk L. Severe acute respiratory syndrome (SARS): a year in review. Annu Rev Med. 2005; 56: 357-81
137. Sparwasser T, Koch ES, Vabulas RM, Heeg K, Lipford GB, Ellwart JW, Wagner H. Bacterial DNA and immunostimulatory CpG oligonucleotides trigger maturation and activation of murine dendritic cells. Eur J Immunol. 1998; 28(6): 2045-54
138. Sun J, Wang Z, Ma S, Zeng G, Zhou Z, Luo K, Hou J. Clinical and virological characteristics of lamivudine resistance in chronic hepatitis B patients: a single center experience. J Med Virol. 2005; 75(3): 391-8
139. Sun S, Zhang X, Tough D, Sprent J. Multiple effects of immunostimulatory DNA on T cells and the role of type I interferons. Springer Semin Immunopathol. 2000; 22(1-2): 77-84
140. Svelander L, Erlandsson Harris H, Lorentzen JC, Trollmo C, Klareskog L, Bucht A. Oligodeoxynucleotides containing CpG motifs can induce T cell-dependent arthritis in rats. Arthritis Rheum. 2004 Jan;50(1):297-304
141. Takeshita A, Imai K, Hanazawa S. CpG motifs in Porphyromonas gingivalis DNA stimulate interleukin-6 expression in human gingival
fibroblasts. Infect Immun. 1999; 67(9): 4340-5
142. Takeshita F, Gursel I, Ishii KJ, Suzuki K, Gursel M, Klinman DM. Signal transduction pathways mediated by the interaction of CpG DNA with Toll-like receptor 9. Semin Immunol. 2004; 16(1): 17-22
143. Talon J, Horvath CM, Polley R, Basler CF, Muster T, Palese P, Garcia-Sastre A. Activation of interferon regulatory factor 3 is inhibited by the influenza A virus NS1 protein. J Virol. 2000; 74(17): 7989-96
144. Thimme R, Bukh J, Spangenberg HC, Wieland S, Pemberton J, Steiger C, Govindarajan S, Purcell RH, Chisari FV. Viral and immunological determinants of hepatitis C virus clearance, persistence, and disease. Proc Natl Acad Sci U S A. 2002; 99(24): 15661-8
145. Tokunaga T, Yamamoto H, Shimada S, Abe H., Fukuda T, Fujisawa Y, Furutani Y, Yano O, Kataoka T, Sudo T, Makiguchi N and Suganuma T. Antitumor activity of deoxyribonucleic acid fraction from Mycobacterium bovis BCG. L. Isolation, physicochemical characterization, and antitimor activity. J. Natl. Cancer Inst., 1984; 72: 955-62
146. Tortorella D, Gewurz BE, Furman MH, Schust DJ, Ploegh HL. Viral subversion of the immune system. Annu Rev Immunol. 2000; 18: 861-926
147. Trevani AS, Chorny A, Salamone G, Vermeulen M, Gamberale R, Schettini J, Raiden S, Geffner J. Bacterial DNA activates human neutrophils by a CpG-independent pathway. Eur J Immunol. 2003; 33(11): 3164-74
148. Tsai SL, Sheen IS, Chien RN, Chu CM, Huang HC, Chuang YL, Lee TH, Liao SK, Lin CL, Kuo GC, Liaw YF. Activation of Th1 immunity is a common immune mechanism for the successful treatment of hepatitis B and C: tetramer assay and therapeutic implications. J Biomed Sci. 2003; 10(1): 120-35
149. Tseng CT, Klimpel GR. Binding of the hepatitis C virus envelope protein E2 to CD81 inhibits natural killer cell functions. J Exp Med. 2002; 195(1): 43-9
150. Turner RB, Felton A, Kosak K, Kelsey DK, Meschievitz CK. Prevention of experimental coronavirus colds with intranasal alpha-2b interferon. J Infect Dis. 1986; 154(3): 443-7
151. Verani A, Gras G, Pancino G. Macrophages and HIV-1: dangerous liaisons. Mol Immunol. 2005; 42(2): 195-212
152. Verthelyi D, Ishii KJ, Gursel M, Takeshita F, Klinman DM. Human peripheral blood cells differentially recognize and respond to two distinct CpG motifs. J Immunol. 2001; 166(4): 2372-7
153. Verthelyi D, Klinman DM. Immunoregulatory activity of CpG oligonucleotides in humans and nonhuman primates. Clin Immunol. 2003; 109(1): 64-71
154. Vollmer J, Weeratna R, Payette P, Jurk M, Schetter C, Laucht M, Wader T, Tluk S, Liu M, Davis HL, Krieg AM. Characterization of three CpG oligodeoxynucleotide classes with distinct immunostimulatory activities. Eur. J. Immunol. 2004; 34: 251–262
155. Wagner M, Poeck H, Jahrsdoerfer B, Rothenfusser S, Prell D, Bohle B, Tuma E, Giese T, Ellwart JW, Endres S, Hartmann G. IL-12p70-dependent Th1 induction by human B cells requires combined activation with CD40 ligand and CpG DNA. J Immunol. 2004; 172(2): 954-63
156. Weeratna RD, McCluskie MJ, Xu Y, Davis HL. CpG DNA induces stronger immune responses with less toxicity than other adjuvants. Vaccine. 2000; 18(17): 1755-62
157. Weighardt H, Feterowski C, Veit M, Rump M, Wagner H, Holzmann B. Increased resistance against acute polymicrobial sepsis in mice challenged with immunostimulatory CpG oligodeoxynucleotides is related to an enhanced innate effector cell response. J Immunol. 2000; 165(8): 4537-43
158. Weiner GJ. The immunobiology and clinical potential of immunostimulatory CpG oligodeoxynucleotides. J Leukoc Biol. 2000; 68(4): 455-63
159. Wenzel J, Uerlich M, Haller O, Bieber T, Tueting T. Enhanced type I interferon signaling and recruitment of chemokine receptor CXCR3-expressing lymphocytes into the skin following treatment with the TLR7-agonist imiquimod. J Cutan Pathol. 2005; 32(4): 257-62
160. Wieland S, Thimme R, Purcell RH, Chisari FV. Genomic analysis of the host response to hepatitis B virus infection. Proc Natl Acad Sci U S A. 2004; 101(17): 6669-74
161. 许正锯,王崇国,李旭红,杨红,张启华。干扰素抗体与干扰素α-2a 治疗慢性乙型肝炎疗效的关系。中华传染病杂志。1998;16(3):26-27
162. Yamamoto S, Kuramoto E, Shimada S, Tokunaga T. In vitro augmentation of natural killer cell activity and production of interferon-alpha/beta and -gamma with deoxyribonucleic acid fraction from Mycobacterium bovis BCG. Jpn J Cancer Res. 1988; 79(7): 866-73
163. Yamamoto S, Yamamoto T, Kataoka T, Kuramoto E, Yano O, Tokunaga T. Unique palindromic sequences in synthetic oligonucleotides are required to induce IFN [correction of INF] and augment IFN-mediated [correction of INF] natural killer activity. J Immunol. 1992; 148(12):4072-6
164. Yamamoto T, Yamamoto S, Kataoka T, Tokunaga T. Lipofection of synthetic oligodeoxyribonucleotide having a palindromic sequence of AACGTT to murine splenocytes enhances interferon production and natural killer activity. Microbiol. Immunol. 1994; 38(10): 831–836
165. Yi AK, Klinman DM, Martin TL, Matson S, Krieg AM. Rapid immune activation by CpG motifs in bacterial DNA. Systemic induction of IL-6 transcription through an antioxidant-sensitive pathway. J Immunol. 1996; 157(12): 5394-402
166. Yi AK, Yoon JG, Krieg AM. Convergence of CpG DNA-and BCR-mediated signals at the c-Jun N-terminal kinase and NF-kappaB activation pathways: regulation by mitogen-activated protein kinases. Int Immunol. 2003; 15(5): 577-91
167. Yu D, Zhu FG, Bhagat L, Wang H, Kandimalla ER, Zhang R, Agrawal S. Potent CpG oligonucleotides containing phosphodiester linkages: in vitro and in vivo immunostimulatory properties. Biochem Biophys Res Commun. 2002; 297(1): 83-90
168. Yu H, Babiuk LA, van Drunen Littel-van den Hurk S. Priming with CpG-enriched plasmid and boosting with protein formulated with CpG