• BMB Reports
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• 제46권9호
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• pp.448-453
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• 2013

CpG-DNA has various immunomodulatory effects in dendritic cells, B cells, and macrophages. While induction of cytokines by CpG-DNA has been well documented in macrophages, the expression of costimulatory molecules in CpG-DNA treated macrophages has not yet been defined. Therefore, we investigated the effects of CpG-DNA on the expression of costimulatory molecules in RAW 264.7 cells. The surface expression of CD80 was slightly increased and CD83 expression was significantly increased in response to CpG-DNA. However, the expression of CD86 and MHC class II was not changed. As expression of CD83 mRNA was also increased by CpG-DNA, CD83 expression is regulated at a transcriptional level. To understand the contribution of signaling pathways to CD83 induction, we used pathway specific inhibitors. The NF-${\kappa}B$ inhibitor significantly reduced surface expression of CD83 as well as phagocytic activity of RAW 264.7 cells. Therefore, CD83 expression may contribute to the immunostimulatory effects of CpG-DNA in macrophage cells.

• BMB Reports
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• 제52권11호
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• pp.635-640
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• 2019

CpG-DNA triggers the proliferation and differentiation of B cells which results in the increased production of antibodies. The presence of bacteria-reactive IgM in normal serum was reported; however, the relevance of CpG-DNA with the production of bacteria-reactive IgM has not been investigated. Here, we proved the function of CpG-DNA for the production of bacteria-reactive IgM. CpG-DNA administration led to increased production of bacteria-reactive IgM both in the peritoneal fluid and serum through TLR9 signaling pathway. When we stimulated B cells with CpG-DNA, production of bacteria-reactive IgM was reproduced in vitro. We established a bacteria-reactive monoclonal IgM antibody using CpG-DNA stimulated-peritoneal B cells. The monoclonal IgM antibody enhanced the phagocytic activity of RAW 264.7 cells against S. aureus MW2 infection. Therefore, we suggest that CpG-DNA enhances the antibacterial activity of the immune system by triggering the production of bacteria-reactive IgM. We also suggest the possible application of the antibodies for the treatment of antibiotics-resistant bacterial infections.

• Journal of Animal Science and Technology
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• 제47권5호
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• pp.783-788
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• 2005

본 시험은 사료의 조단백질 수준이 육성기 흑염소의 발육과 육질에 미치는 영향을 구명하기 하기 위하여 흑염소 육성축 수컷 40두를 공시하여 조단백질 수준을 각각 달리하여 2004년 4월 19일부터 2004년 9월 13일까지 시험을 실시하였으며 그 결과는 다음과 같다. 일당증체량은 CP 14%, CP 18%구가 각각 84.0, 83.0g으로 CP 12%구 69.2g 보다 높았으며(P<0.05) CP 16%구는 80.9g으로 비슷한 경향이었다. 1일 사료섭취량은 농후사료와 볏짚 각각 590g, 45g 내외로 처리구간에 비슷하였고 증체 g당 사료요구율은 CP 14%, 16%, 18%구간에는 7.0-7.3으로 비슷하였으나 CP 12%구는 다소 높은 경향이었다. 도체율은 CP 12%구가 61.7%로 CP 14와 CP 18%구 보다 높았으나(P<0.05) CP 16%구와는 비슷하였다. 정육율은 CP 16%가 51.7%로 CP 12와 CP 14%구 보다 높았다(P<0.05). 지방율은 CP 12%구가 12.8% 높았으며(P<0.05) 뼈율은 17.0% 내외로 처리구간에 비슷하였다. 고기의 물리적 특성인 전단력과 가열 감량은 CP 16%구와 CP 14%구가 다른 처리구보다 다소 낮은 경향이었고 보수력은 CP 16%구가 CP 18%구 보다 높았다(P<0.05). 관능검사결과 다즙성은 CP 16%구가 CP 12%구 보다 우수하였으며(P<0.05) 연도도 다른 처리구에 비해 우수한 경향이었다. 이러한 결과를 종합적으로 고려할 때 육성기 흑염소의 발육과 육질개선을 위한 사료의 적정 조단백질 수준은 14-16%가 적합한 것으로 사료된다.

• BMB Reports
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• 제43권3호
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• pp.164-169
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• 2010

CpG-DNA, which contains unmethylated CpG dinucleotides in the context of specific sequences, has remarkable and diverse immunological effects, including induction of proinflammatory cytokine expression and regulation of the Th1/Th2 immune response. Here, we examined the immunostimulatory activities of double-stranded (ds) CpG-DNA in the human B cell line RPMI8226. To investigate whether dsCpG-DNA stimulates immune cells, we constructed a plasmid containing repeated dsCpG-DNA and produced dsCpG-DNA by PCR amplification and EcoR I digestion. PCR-amplified dsCpG-DNA alone did not have immmunostimulatory activity. However, dsCpGDNA encapsulated with lipofectin induced IL-8 promoter activation, HLA-DRA expression, and IL-8 expression in a CG sequence-independent manner. The effects of encapsulated dsCpGDNA were independent of minor endotoxin contamination. These findings suggest the potential use of dsCpG-DNA as a therapy for immune response regulation.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제33권6호
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• pp.636-642
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• 2007

Background: CpG DNA plays an important role in immune cell function. This study examined whether the temporal control of toll-like receptor (TLR)9 by CpG DNA can regulate the expression of matrix metalloproteinase-9(MMP-9). Methods and materials: Macrophages were cultured in the presence of 10% FBS. For the various MMP genes analysis, RT-PCR and real-time PCR were performed. In addition, zymography assay performed for the MMP activity. The phosphorylation assay did for the ERK1/2 and NF${\kappa}B$ activation, and luciferase promoter assay was for the NF${\kappa}B$ activity. Results: CpG DNA induced the mRNA expression of MMP-2, MMP-9, and MMP-13, but not of MMP-7, MMP-8, and MMP-12, in a time-dependent manner. Especially, the mRNA expression of MMP-9 was strongly induced by CpG DNA using real-time RT-PCR. The TLR9 inhibitor, chloroquine, suppressed CpG DNA-induced MMP-9 expression and its activity. Moreover, CpG DNA induced the phosphorylation of ERK and the inhibition of ERK by U0126 suppressed CpG DNA-induced MMP-9 expression and its activity. CpG DNA stimulated $I{\kappa}B-{\alpha}$ degradation and luciferase activity. In addition, pretreatment of SN-50, the inhibitor of NF${\kappa}B$, strongly blocked the CpG DNA-induced MMP-9 expression and activity. Conclusion: These observations suggest that CpG DNA may play important roles in the activation of macrophages by regulating the production of MMP-9 via the sequential TLR9-ERK-NF${\kappa}B$ signaling pathway.

• Reproductive and Developmental Biology
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• 제35권1호
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• pp.93-97
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• 2011

Previously, we reported that the osmolarity conditions in the satellite region were affected CpG DNA methylation status while Pre-1 sequence was not affected CpG DNA methylation in pNT blastocyst stage. This study was conducted to investigate the DNA methylation status of repeat sequences in pig nuclear transfer (pNT) embryos produced under different osmolarity culture conditions. Control group of pNT embryos was cultured in PZM-3 for six days. Other two treatment groups of pNT embryos were cultured in modified PZM-3 with 138 mM NaCl or 0.05 M sucrose (mPZM-3, 320 mOsmol) for two days, and then cultured in PZM-3 (270 mOsmol) for four days. The DNA methylation status of the Pre-1 sequences in blastocysts was characterized using a bisulfite-sequencing method. Intriguingly, in the present study, we found the unique DNA methylation at several non-CpG sequences at the Pre-1 sequences in all groups. The non-CpG methylation was hypermethylated in all three groups, including in vivo group (86.90% of PZM-3; 83.87% of NaCl; 84.82% of sucrose; 90.94% of in vivo embryos). To determine whether certain non-CpG methylated sites were preferentially methylated, we also investigated the methylation degree of CpA, CpT and CpC. Excepting in vivo group, preference of methylation was CpT>CpC>CpA in all three groups investigated. These results indicate that DNA methylation of Pre-1 sequences was hypermethylated in CpG as well as non-CpG site, regardless modification of osmolarity in a culture media.

• BMB Reports
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• 제44권9호
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• pp.607-612
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• 2011

Several investigators have shown that CpG-DNA has outstanding effects as a Th1-responsive adjuvant and that its potent adjuvant effects are enhanced by encapsulation with a liposome of proper composition. In this study, we showed that encapsulation with phosphatidyl-${\beta}$-oleoyl-${\gamma}$-palmitoyl ethanolamine (DOPE): cholesterol hemisuccinate (CHEMS) complex enhances the immunostimulatory activity of CpG DNA and the binding of CpG-DNA to TLR9. We also examined involvement of myeloid differentiation protein (MyD88) and NF-${\kappa}B$ activation in liposome-encapsulated CpG-DNA-induced IL-8 promoter activation. In this manuscript, the natural phosphodiester bond CpG-DNA encapsulated by DOPE : CHEMS complex is designated as Lipoplex(O). Importantly, we successfully screened B cell epitopes of envelope protein (E protein) of hepatitis C virus (HCV-E) and attachment glycoprotein G of human respiratory syncytial virus (HRSV-G) by immunization with complexes of several peptides and Lipoplex(O) without carriers. Therefore, Lipoplex(O) is potentially applicable as a universal adjuvant for peptide-based epitope screening and antibody production.

• Tuberculosis and Respiratory Diseases
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• 제57권6호
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• pp.543-552
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• 2004

연구배경 : 급성천식 동물모델에 투여할 경우 특징적인 기도과민성이나 호산구성 염증 등이 호전되는 것으로 알려져 있으나, 만성천식 모델에서 기도의 만성염증 및 기도재구성을 억제할 수 있다는 연구는 많지 않다. CpG-ODN이 기도의 만성염증 및 기도재구성을 예방할 수 있는 지 알기위하여, 만성천식 마우스모델에서 CpG-ODN의 기도의 만성염증 및 기도재구성에 대한 영향을 연구하고자 하였다. 방 법 : BALB/C 마우스를 ovalbumin으로 감작시킨뒤 7주간 만성적인 자극을 주어 만성 천식모델을 만들고, CpG-ODN은 감작시에 복강내에 주사하였다. 7주후기 도저항(기도과민성)의 측정, 혈청 IgE의 측정, 기관지 폐포세척액에서 염증세포분획을 검사하였고, 폐조직검사로 염증세포의 침윤 및 기저막하부의 섬유화의 정도를 관찰하였고, OVA군과 CpG-ODN군과 비교하였다. 결 과 : 1) 기관지폐포세척액내 호산구는 CpG-ODN군은 OVA군에 비해 통계적으로 유의하게 감소하였다($0.9{\pm}0.8$ ; $9.7{\pm}5.8{\times}10^4/ml$, P<0.01). 2) 기도과민성은 CpG-ODN군은 OVA군에 비해서 기도과민성(Penh)이 $50mg/m{\ell}$를 제외한 농도에서 모두 유의하게 감소한 소견을 보였다(P<0.01). 3) 혈청 총 IgE는 CpG-ODN군이 OVA군에 비해 감소한 소견을 보였고($0.21{\pm}0.11$ ; $1.16{\pm}0.39{\mu}g/ml$, P<0.01), 혈청 OVA specific IgE도 CpG-ODN군이 OVA군에 비해 감소한 소견을 보였다($1.1{\pm}0.5$ ; $1.7{\pm}0.6$ OD units, P<0.01). 4) 배상세포의 증식정도는 CpG-ODN군은 OVA군에 감소된(P<0.01) 소견을 보였다. 5) 기저막하부의 섬유화의 정도는 CpG-ODN군이 OVA군에 비해서 유의하게 감소된(P<0.01) 소견을 보였다. 결 론 : CpG-ODN은 기도의 급성염증 및 기도과민성을 억제할 뿐만 아니라 만성천식 마우스모델에서 기도의 만성염증 및 기도재구성을 억제시킴을 알 수 있었다.

• Fisheries and Aquatic Sciences
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• 제24권3호
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• pp.129-137
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• 2021

Dietary protein requirements of abalone (Haliotis discus) depending on abalone size were determined and compared. One thousand and fifty small abalone (initial weight of 2.7 g) and five hundred forty large one (initial weight of 16.0 g) were distributed into 15 and 18 containers in Trial 1 and 2, respectively. Five and six experimental diets containing crude protein level from 20% to 40% and 20% to 45% with 5% increment of protein level for the small and large abalone were prepared and referred to as the CP20, CP25, CP30, CP35, CP40, and CP45 diets, respectively. The experimental diets were fed to abalone for 16 weeks in Trials 1 and 2. Specific growth rate (SGR) of the small abalone fed the CP20 diet was lower compared to that of abalone fed all other diets in Trial 1. Growth performance (weight gain and SGR) of the large abalone fed the CP30, CP35, and CP40 diets were greater than that of abalone fed the CP20, CP25, and CP45 diets in Trial 2. Dietary protein requirements were estimated to be 33.0% and 33.5% for the small and large abalone based on the 2^nd order polynomial analysis, respectively. Dietary protein requirements for the small abalone grown from 2.7 g to 7.4 g and the large one grown from 16 g to 21 g were estimated to be 33.0% and 33.5%, respectively. Size differences in abalone did not affect dietary protein requirement under this experimental conditions.

• Asian-Australasian Journal of Animal Sciences
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• 제31권8호
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• pp.1141-1149
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• 2018

Objective: Cluster of differentiation 4 protein (CD4) gene is an important immune related gene which plays a significant role in T cell development and host resistance during viral infection. Methods: In order to unravel the relationship of CpG island methylation level of CD4 gene with its gene expression and T lymphocyte subpopulation traits, we used one typical Chinese indigenous breed (Dapulian, DP) and one commercial breed (Landrace), then predicted the CpG island of CD4 gene, determined the methylation status of CpG sites by bisulfite sequencing polymerase chain reaction (BSP), and carried out the correlation analyses of methylation frequencies of CpG sites with mRNA expression and T lymphocyte subpopulation traits. Results: There was one CpG island predicted in the upstream -2 kb region and exon one of porcine CD4 gene, which located 333 bp upstream from the start site of gene and contained nine CpG sites. The correlation analysis results indicated that the methylation frequency of CpG_2 significantly correlated with CD4 mRNA expression in the DP and Landrace combined population, though it did not reach significance level in DP and Landrace separately. Additionally, 15 potential binding transcription factors (TFs) were predicted within the CpG island, and one of them (Jumonji) contained CpG_2 site, suggesting that it may influence the CD4 gene expression through the potential binding TFs. We also found methylation frequency of CpG_2 negatively correlated with T lymphocyte subpopulation traits CD4+CD8-CD3-, CD4-CD8+CD3- and CD4+/CD8+, and positively correlated with CD4-CD8+CD3+ and CD4+CD8+CD3+ (for all correlation, p<0.01) in DP and Landrace combined population. Thus, the CpG_2 was a critical methylation site for porcine CD4 gene expression and T lymphocyte subpopulation traits. Conclusion: We speculated that increased methylation frequency of CpG_2 may lead to the decreased expression of CD4, which may have some kind of influence on T lymphocyte subpopulation traits and the immunity of DP population.