• BMB Reports
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• 제44권11호
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• pp.758-763
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• 2011

Immunostimulatory CpG-DNA targeting TLR9 is one of the most extensively evaluated vaccine adjuvants. Previously, we found that a particular form of natural phosphodiester bond CpG-DNA (PO-ODN) encapsulated in a phosphatidyl-${\beta}$-oleoyl-${\gamma}$-palmitoyl ethanolamine (DOPE) : cholesterol hemisuccinate (CHEMS) (1 : 1 ratio) complex (Lipoplex(O)) is a potent adjuvant. Complexes containing peptide and Lipoplex(O) are extremely useful for B cell epitope screening and antibody production without carriers. Here, we showed that IL-12 production was increased in bone marrow derived dendritic cells in a CpG sequence-dependent manner when PO-ODN was encapsulated in Lipoplex(O), DOTAP or lipofectamine. However, the effects of Lipoplex(O) surpassed those of PO-ODN encapsulated in DOTAP or lipofectamine and also other various forms of liposome-encapsulated CpG-DNA in terms of potency for protein antigen-specific IgG production and Th1- associated IgG2a production. Therefore, Lipoplex(O) may have a unique potent immunoadjuvant activity which can be useful for various applications involving protein antigens as well as peptides.

• BMB Reports
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• 제39권6호
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• pp.788-793
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• 2006

Bacterial DNA containing immunostimulatory CpG motifs can stimulate antigen-presenting cells to express co-stimulatory molecules and to produce various cytokines in vivo and in vitro. In this study, we fragmented macromolecular E.coli genomic DNA with DNase I, and analyzed the ability of the resulting DNA fragments to induce the NF-${\kappa}B$ activation and humoral immune response. Furthermore, using computational analysis and luciferase assay for synthetic ODNs based on the sequence of the immunostimulatory DNA fragments (DF-ODNs), an active component of DF-ODNs sequences was investigated. Experimental results demonstrated that DF-ODN is optimal for the NF-${\kappa}B$-responsive promoter activation in the mouse macrophage cell line and the humoral immune response in vivo. In agreement with the activity of the DF-ODNs processed by DNase I, a synthetic ODN based on the DF-ODN sequences is potent at inducing IL-12 mRNA expression in primary dendritic cells. These results suggest that the discovery and characterization of a highly active natural CpG-ODN may be achieved by the analyses of bacterial DNA fragments generated by a nuclease activity.

• Journal of Animal Science and Technology
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• 제61권5호
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• pp.245-253
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• 2019

Pathogen-associated Molecular Patterns (PAMPs) are highly conserved structural motifs that are recognized by Pathogen Recognition receptors (PRRs) to initiate immune responses. Infection by these pathogens and the immune response to PAMPS such as lipopolysaccharide (LPS), Peptidoglycan (PGN), bacterial oligodeoxynucleotides [CpG oligodeoxynucleotides 2006 (CpG ODN2006) and CpG oligodeoxynucleotides 2216 (CpG ODN2216)], and viral RNA Polyinosinic-Polycytidylic Acid (Poly I:C), are associated with infectious and metabolic diseases in animals impacting health and production. It is established that PAMPs mediate the production of cytokines by binding to PRRs such as Toll-like receptors (TLR) on immune cells. Galectins (Gal) are carbohydrate-binding proteins that when expressed play essential roles in the resolution of infectious and metabolic diseases. Thus it is important to determine if the expression of galectin gene (LGALS) and Gal secretion in blood are affected by exposure to LPS and PGN, PolyI:C and bacterial CpG ODNs. LPS increased transcription of LGALS4 and 12 (2.5 and 2.02 folds respectively) and decreased secretion of Gal 4 (p < 0.05). PGN increased transcription of LGALS-1, -2, -3, -4, -7, and -12 (3.0, 2.3, 2.0, 4.1, 3.3, and 2.4 folds respectively) and secretion of Gal-8 and Gal-9 (p < 0.05). Poly I:C tended to increase the transcription of LGALS1, LGALS4, and LGALS8 (1.78, 1.88, and 1.73 folds respectively). Secretion of Gal-1, -3, -8 and nine were significantly increased in treated samples compared to control (p < 0.05). CpG ODN2006 did not cause any significant fold changes in LGALS transcription (FC < 2) but increased secretion of Gal-1, and-3 (p < 0.05) in plasma compared to control. Gal-4 was however reduced in plasma (p < 0.05). CpG ODN2216 increased transcription of LGALS1 and LGALS3 (3.8 and 1.6 folds respectively), but reduced LGALS2, LGALS4, LGALS7, and LGALS12 (-1.9, -2.0, -2.0 and; -2.7 folds respectively). Secretion of Gal-2 and -3 in plasma was increased compared to control (p < 0.05). Gal-4 secretion was reduced in plasma (p < 0.05). The results demonstrate that PAMPs differentially modulate galectin transcription and translation of galectins in cow blood.

• Clinical and Experimental Pediatrics
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• 제52권9호
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• pp.1015-1020
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• 2009

목 적 : 알레르기 환자와 정상인에서 형질세포양 수지상세포의 분포, TLR9 mRNA 양과 $IFN-{\alpha}$의 분비능에 차이가 있는지 알아보고자 하였다. 방 법 : 19명의 알레르기 환자와 17명의 건강한 성인을 대상으로 하였다. 말초혈액단핵세포를 채취하여 Lineage Cocktail(CD3, CD14, CD16, CD19, CD20, CD56)음성, HLA-DR 양성이면서 CD123양성을 유세포 분석기로 분석하였다. 말초혈액단핵세포에 TLR9 작용제(agonists)인 CpG-ODN 2216과 음성 대조를 위해 CpG-ODN 2206으로 자극하고 24시간 후 상청액을 추출하여 $IFN-{\alpha}$의 농도를 측정하였다. 또한 real time RT-PCR을 이용하여 TLR9 mRNA 정량분석을 시행하였다. 결 과 : 말초혈액단핵세포에서 형질세포양 수지상세포의 분포는 알레르기 환자가 평균 $0.1{\pm}0.04%$, 대조군이 평균 $0.25{\pm}0.23%$이었다. TLR9 mRNA 상대적인 양을 나타내는 ${\Delta}{\Delta}Ct$는 알레르기 환자에서 $1.29{\pm}0.41$이었고 대조군은 $1.25{\pm}0.23$이었다. TLR9 리간드인 CpG-ODN 2216 자극에 따른 $IFN-{\alpha}$의 분비능은 알레르기 환자에서 $911{\pm}829pg/mL$ 이었고 대조군에서 $1,095{\pm}888pg/mL$ 이었다. 이 세 결과에서 통계적인 차이는 없었다. 결 론 : TLR9을 통한 신호전달이 알레르기 환자의 면역반응을 대표하지는 않는 것으로 보이며, 향후 더 자세한 TLR9의 역할에 대한 연구가 필요하다.

• IMMUNE NETWORK
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• 제15권1호
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• pp.9-15
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• 2015

Streptococcus pneumoniae (the pneumococcus) causes a major upper respiratory tract infection often leading to severe illness and death in the elderly. Thus, it is important to induce safe and effective mucosal immunity against this pathogen in order to prevent pnuemocaccal infection. However, this is a very difficult task to elicit protective mucosal IgA antibody responses in older individuals. A combind nasal adjuvant consisting of a plasmid encoding the Flt3 ligand cDNA (pFL) and CpG oligonucleotide (CpG ODN) successfully enhanced S. pneumoniae-specific mucosal immunity in aged mice. In particular, a pneumococcal surface protein A-based nasal vaccine given with pFL and CpG ODN induced complete protection from S. pneumoniae infection. These results show that nasal delivery of a combined DNA adjuvant offers an attractive potential for protection against the pneumococcus in the elderly.

• 한국어병학회지
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• 제16권2호
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• pp.103-110
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• 2003

• The Korean Journal of Physiology and Pharmacology
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• 제18권3호
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• pp.225-231
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• 2014

In this study we aim to extensively investigate the anti-influenza virus immune responses in human pharyngeal epithelial cell line (Hep-2) and evaluate the protective role of Toll-like receptor (TLR) ligands in seasonal influenza A H1N1 (sH1N1) infections in vitro. We first investigated the expression of the TLRs and cytokines genes in resting and sH1N1 infected Hep-2 cells. Clear expressions of TLR3, TLR9, interleukin (IL)-6, tumour necrosis factor (TNF)-${\alpha}$ and interferon (IFN)-${\beta}$ were detected in resting Hep-2 cells. After sH1N1 infection, a ten-fold of TLR3 and TLR9 were elicited. Concomitant with the TLRs activation, transcriptional expression of IL-6, TNF-${\alpha}$ and IFN-${\beta}$ were significantly induced in sH1N1-infected cells. Pre-treatment of cells with poly I:C (an analog of viral double-stranded RNA) and CpG-ODN (a CpG-motif containing oligodeoxydinucleotide) resulted in a strong reduction of viral and cytokines mRNA expression. The results presented indicated the innate immune response activation in Hep-2 cells and affirm the antiviral role of Poly I:C and CpG-ODN in the protection against seasonal influenza A viruses.

• 생명과학회지
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• 제27권4호
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• pp.482-499
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• 2017

Allergy 질환은 지난 십여년 동안 개발도상국을 포함해서 전 세계적으로 증가하고 있다. Allergy 염증 반응은 수지상 세포와 같은 항원제시 세포에 의한 allergy 항원섭취를 시작으로 하여 Th2 면역 반응에 의해서 일어난다. 장내 미생물은 신체의 대사나 생리적 기능을 조절하고, 생애 초기의 면역 체계 성숙과 일생 동안 면역 체계 항상성 및 상피세포 총체성에 기여한다. Bifidobacteria는 Th1/Th2 balance에 strain-specific 한 면역 자극 특성을 가지며, TSLP와 IgE 발현을 억제 시키고 Flg과 FoxP3 발현을 촉진 시켜 allergy를 완화시킨다. 또한 Unmethylated CpG motif ODN은 B 세포와 수지상 세포의 TLR9에 의해 인식 되어 선천성과 적응성 면역 반응을 유도하고, Clostridium butyricum에 의해서 생산된 butyrate는 수지상 세포의 anti-inflammatory 유전자의 발현을 유도하기 위해 GPR109a signaling pathway를 활성화시키고, GPR43 활성화를 통하여 tTreg 세포 proliferation을 직접 자극하거나 HADC 활성을 억제시켜 Foxp3 gene intronic enhancer의 histone H3 acetylation을 통해 naive $CD4^+$ T 세포를 pTreg 세포로 분화시킨다.

• Journal of Ginseng Research
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• 제31권4호
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• pp.181-190
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• 2007

본 연구에서는 고려홍삼으로부터 새로 분리한 CK 함유분획을 이용하여 마우스 대식세포에 대한 선천면역반응 조절에 미치는 영향을 조사하였다. 본 연구에서 사용된 농도의 CK 함유분획에서는 세포독성 효과가 관찰되지 않았으며 CK함유 분획의 전처리에 의하여 그람음성세균의 LPS, 또는 CpG-ODN에 의해 유도되는 NF-${\kappa}B$와 MAPK 활성 및 전염증성 사이토카인, NO의 분비가 TLR4 및 TLR9 특이적으로 억제되었다. 이와 같은 결과는 CK 함유분획이 TLR4을 매개로하는 염증반응뿐만 아니라 TLR9을 통한 염증반응에도 영향을 미치는 것으로 해석된다. 따라서 앞으로 CK 함유 분획에 포함된 개별 사포닌 등 시료 성분에 대한 면밀한 분석, 그리고 이들 개별 물질이 각각의 신호전달 체계에 미치는 영향과 그 기작에 대한 연구가 더욱 필요할 것으로 사료되며 염증억제제로서의 개발 가능성을 탐구하기 위한 생체 내에서의 효능 및 작용기전 분석이 요구된다.

• IMMUNE NETWORK
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• 제24권4호
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• pp.25.1-25.14
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• 2024

Lupus is characterized by the autoantibodies against nuclear Ags, underscoring the importance of identifying the B cell subsets driving autoimmunity. Our research focused on the mitochondrial activity and CXCR4 expression in CD11c⁺ B cells from lupus patients after ex vivo stimulation with a TLR9 agonist, CpG-oligodeoxyribonucleotide (ODN). We also evaluated the response of CD11c⁺ B cells in ODN-injected mice. Post-ex vivo ODN stimulation, we observed an increase in the proportion of CD11c^hi cells, with elevated mitochondrial activity and CXCR4 expression in CD11c⁺ B cells from lupus patients. In vivo experiments showed similar patterns, with TLR9 stimulation enhancing mitochondrial and CXCR4 activities in CD11c^hi B cells, leading to the generation of anti-dsDNA plasmablasts. The CXCR4 inhibitor AMD3100 and the mitochondrial complex I inhibitor IM156 significantly reduced the proportion of CD11c⁺ B cells and autoreactive plasmablasts. These results underscore the pivotal roles of mitochondria and CXCR4 in the production of autoreactive plasmablasts.