• Asian-Australasian Journal of Animal Sciences
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• 제31권4호
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• pp.595-606
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• 2018

Objective: The Fusarium mycotoxins of deoxynivalenol (DON) and zerolenone (ZEN) cause health hazards for both humans and farm animals. Therefore, the main intention of this study was to reveal DON and ZEN effects on the mRNA expression of pro-inflammatory cytokines and other immune related genes in the liver of piglets. Methods: In the present study, 15 six-week-old piglets were randomly assigned to the following three different dietary treatments for 4 weeks: control diet, diet containing 8 mg DON/kg feed, and diet containing 0.8 mg ZEN/kg feed. After 4 weeks, liver samples were collected and sequenced using RNA-Seq to investigate the effects of the mycotoxins on genes and gene networks associated with the immune systems of the piglets. Results: Our analysis identified a total of 249 differentially expressed genes (DEGs), which included 99 upregulated and 150 downregulated genes in both the DON and ZEN dietary treatment groups. After biological pathway analysis, the DEGs were determined to be significantly enriched in gene ontology terms associated with many biological pathways, including immune response and cellular and metabolic processes. Consistent with inflammatory stimulation due to the mycotoxin-contaminated diet, the following Kyoto encyclopedia of genes and genomes pathways, which were related to disease and immune responses, were found to be enriched in the DEGs: allograft rejection pathway, cell adhesion molecules, graft-versus-host disease, autoimmune thyroid disease (AITD), type I diabetes mellitus, human T-cell leukemia lymphoma virus infection, and viral carcinogenesis. Genome-wide expression analysis revealed that DON and ZEN treatments downregulated the expression of the majority of the DEGs that were associated with inflammatory cytokines (interleukin 10 receptor, beta, chemokine [C-X-C motif] ligand 9), proliferation (insulin-like growth factor 1, major facilitator superfamily domain containing 2A, insulin-like growth factor binding protein 2, lipase G, and salt inducible kinase 1), and other immune response networks (paired immunoglobulin-like type 2 receptor beta, Src-like-adaptor-1 [SLA1], SLA3, SLA5, SLA7, claudin 4, nicotinamide N-methyltransferase, thyrotropin-releasing hormone degrading enzyme, ubiquitin D, histone $H_2B$ type 1, and serum amyloid A). Conclusion: In summary, our results demonstrated that high concentrations DON and ZEN disrupt immune-related processes in the liver.

• Reproductive and Developmental Biology
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• 제36권1호
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• pp.33-37
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• 2012

Differential capacity of the parthenogenetic embryonic stem cells (PESCs) is still under controversy and the mechanisms of its neural induction are yet poorly understood. Here we demonstrated neural lineage induction of PESCs by addition of insulin-like growth factor-2 (Igf2), which is an important factor for embryo organ development and a paternally expressed imprinting gene. Murine PESCs were aggregated to embryoid bodies (EBs) by suspension culture under the leukemia inhibitory factor-free condition for 4 days. To test the effect of exogenous Igf2, 30 ng/ml of Igf2 was supplemented to EBs induction medium. Then neural induction was carried out with serum-free medium containing insulin, transferrin, selenium, and fibronectin complex (ITSFn) for 12 days. Normal murine embryonic stem cells derived from fertilized embryos (ESCs) were used as the control group. Neural potential of differentiated PESCs and ESCs were analyzed by immunofluorescent labeling and real-time PCR assay (Nestin, neural progenitor marker; Tuj1, neuronal cell marker; GFAP, glial cell marker). The differentiated cells from both ESC and PESC showed heterogeneous population of Nestin, Tuj1, and GFAP positive cells. In terms of the level of gene expression, PESC showed 4 times higher level of GFAP expression than ESCs. After exposure to Igf2, the expression level of GFAP decreased both in derivatives of PESCs and ESCs. Interestingly, the expression level of $Tuj1$ increased only in ESCs, not in PESCs. The results show that IGF2 is a positive effector for suppressing over-expressed glial differentiation during neural induction of PESCs and for promoting neuronal differentiation of ESCs, while exogenous Igf2 could not accelerate the neuronal differentiation of PESCs. Although exogenous Igf2 promotes neuronal differentiation of normal ESCs, expression of endogenous $Igf2$ may be critical for initiating neuronal differentiation of pluripotent stem cells. The findings may contribute to understanding of the relationship between imprinting mechanism and neural differentiation and its application to neural tissue repair in the future.

• 한국식품과학회지
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• 제30권3호
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• pp.702-708
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• 1998

연구는 버섯을 이용한 기능성식품 개발 및 버섯가공 산업 육성에 기초자료를 제공하고자 표고버섯 및 느타리버섯 자실체, 균사체 및 폐상의 조단백다당체 분말을 0, 5, 25 mg/kg 농도로 생리식염수에 용해시켜 마우스(DBA/2와 ICR)에 주사하여 i) 백혈병$(L_{1210})$, 간암$(H_{22}),\;mouser{\;}sarcoma180\;(S_{180})$에 대한 항암효과, ii) 면역기전에 관련된 장기중량변화, 용혈반 형성 세포수 변화 등 조단백다당이 면역에 미치는 영향, 그리고 in vitro 세포독성실험을 수행하였다. 본 실험에서 사용한 모든 조단백다당류는 백혈병에 대해 항암효과를 보였는데 그 중 표고버섯자실체 25 mg/kg 처리구가 가장 높은 저지율(86%)을 보였다. 간암$(H_{22})$에 대한 항암효과는 표고버섯균사체 5 mg/kg 처리구를 제외하고 모든 처리구에서 저지효과를 보였는데, 그 중 느타리버섯 폐상 25 mg/kg 처리구가 가장 높은 저지율(87.6%)을 보였으며, 다음은 표고버섯자실체 25 mg/kg 처리구(71%)였다. Sarcoma180에 대한 항암효과는 표고버섯균사체와 자실체 25 mg/kg 처리구에서 각각 30.9,와 33.9%의 저지율을 보였다. In vitro 세포독성검사에서 각 시료의 최종농도 $50,\;100,\;200,\;400\;{\mu}g/{\mu}L$에서 $L_{1210}$에 대한 유의적 세포사망률은 보이지 않았다. 면역효과 실험에서 간장과 비장중량은 농도증가에 따라 증가하는 추세이나 현저한 차이는 없었다. 표고버섯균사체와 자실체 처리시 항체생성능력을 지닌 용혈반 형성 세포수는 대조군에 비해 현저히 높게 나타났다.

• Archives of Pharmacal Research
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• 제21권5호
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• pp.581-590
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• 1998

We developed a novel water-soluble camptothecin analobue, CKD602, and evaluated the inhibition of topoisomerase I and the antitumor activities against mammalian tumor cells and human tumor xenografts. CKD602 was a nanomolar inhibitor of the topoisomerase I enzyme in the cleavable complex assay. CKD602 was found to be 3 times and slightly more potent than topotecan and camptothecin as inhibitors of topoisomerase, respecitively. In tumor cell cytotoxicity, CKD602 was more potent than topotecan in 14 out of 26 human cancer cell lines tested, while it was comparable to camptothecin. CKD602 was tested for the in vivo antitumor activity against the human tumor xenograft models. CKD602 was able to imduce regression of established HT-29, WIDR and CX-1 colon tumors, LX-1 lung tumor, MX-1 breast tumor and SKOV-3 ovarian tumor as much as 80, 94, 76, 67, 87% and 88%, respectively, with comparable body weight changes to those of topotecan. Also the therapeutic margin (R/Emax: maximum tolerance dose/$ED-{58}$) of CKD602 was significantly higher than that of topotecan by 4 times. Efficacy was determined at the maximal tolerated dose levels using schedule dependent i.p. administration in mice bearing L1210 leukemia. On a Q4dx4 (every 4 day for 4 doses) schedule, the maximum tolerated dose (MTD) was 25 mg/kg per administration, which caused great weight loss and lethality in <5% tumor bearing mouse. this schedule brought significant increase in life span (ILS), 212%, with 33% of long-term survivals. The ex vivo antitumor activity of CKD602 was compared with that of topotecan and the mean antitumor index (ATI) values recorded for CKD602 were significantly higher than that noted for topotecan. From these results, CKD602 warrants further clinical investigations as a potent inhibitor of topoisomerase I.

• 한방안이비인후피부과학회지
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• 제25권3호
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• pp.1-12
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• 2012

Objectives : Fagopyrum esculentum(FE) has been used for removal of inflammation of internal organs and treatment of sore and ulcer by heat toxin in Korean herbal medicines. In this study, To investigated the protective effect of FE on allergic response, we determined whether FE inhibits allergic response. Methods : The effect of FE was analyzed by ELISA, RT-PCR and Western blot in RBL-2H3 cells. We investigated cell viability, ${\beta}$-hexosaminidase, as a marker of degranulation, cytokne, and intracellular ROS and MAPK and NF-${\kappa}B$ signaling. Results : We found that FE suppressed ${\beta}$-hexosaminidase release, the production of IL-4 and TNF-${\alpha}$ and intracellular ROS level in RBL-2H3 by the anti-DNP IgE plus DNP-HSA stimulation. FE also significantly inhibited cytokine mRNA expressions, such as IL-$1{\beta}$, IL-2, IL-3, IL-4, IL-5, IL-6, IL-13, TNF-${\alpha}$ and GM-CSF in RBL-2H3. In addition, PF suppressed the phospholyation of ERK1/2, JNK1/2, p38 and $I{\kappa}B{\alpha}$ and NF-${\kappa}B$ signal transduction pathway. Conclusions : Our results indicate that FE protects against allergic response and exerts an anti-inflammatory effect through the inhibition of degranulation and production of cytokines and ROS via the suppression MAPK and NF-${\kappa}B$ of signal transduction. Abbrevations : FE, Fagopyrum esculentum; RBL-2H3, rat basophilic leukemia cell line; ROS, reactive oxygen species; MAPK, Mitogen-activated protein kinase; $NF{\kappa}B$, nuclear factor ${\kappa}B$; $TNF{\alpha}$, Tumor necrosis factor alpha; GM-CSF, Granulocyte macrophage colony-stimulating factor; ERK, extracellular-signal-regulated kinase; JNK, c-Jun NH2-terminal kinase; p38, p38 MAP kinase; $I{\kappa}B{\alpha}$, inhibitory-kappa B alpha.

• 한국약용작물학회지
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• 제11권1호
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• pp.31-39
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• 2003

바위솔의 추출물로부터 항산화 활성 및 동물 항암세포를 이용한 생물학적 특성을 조사 해 본 결과는 다음과 같다. 생육지별 DPPH 프리라디칼 소거 및 xanthine/xanthine oxidase 억제 활성비교에서 재배지에서 수확한 시료의 활성이 $6.07(IC_{50}:{\mu}g/m{\ell})$로 가장 높았다 생육시기별 DPPH 프리라디칼 소거 및 xanthine/xanthine oxidase 억제의 활성은 9월 수확물이 가장 활성이 높았다. 실리카겔 컬럼크로마토그래피를 이용한 항산화물질 분리결과에서 S-4 분획물의 DPPH와 xanthine oxidase 억제활성이 $5.02(IC_{50}\;:\;{\mu}g/m{\ell})$와 $6.18(IC_{50}:{\mu}g/m{\ell})$을 나타냄으로써 높은 분리효율를 보였다. 항산화활성이 가장 높았던 LH-4 분획물의 주요화합물은 GC/MS에 의하여 지방산, 폴리페놀화합물 및 페놀유도체로 동정되었으며 alpha-androsta-7,14-diene과 1,2,3-benzenetriol이 주 물질이었다. POD 및 SOD 활성은 생육지간에 재배지, 산, 바닷가 순으로, 생육시기간에는 시기가 늦을수록 높게 나타났다 SOD 동위효소의 밴드수는 전 생육지, 전 생육시기에서 공히 2개의 밴드가 나타났으나, 그 세기는 재배지와 9월의 수확물이 가장 높은 활성을 나타내었다. 정제된 LH-4 분획물은 HL-60 세포의 종양증식을 억제시켰으며 종양유발유전자와 IPTG로 콜로니 형성을 유도한 암세포(2-12 cells)에서 분획물 400ppm 처리와 negative control 처리간의 colony형성은 유의차가 없었다.

• 동의생리학회지
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• 제14권2호통권20호
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• pp.229-237
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• 1999

Hwanhonsan has been used for curing tumor as a Oriental medicine without any experimental evidence to support the rational basis for their clinical use. This experiment was carried out to evaluate the possible therapeutic or antitumoral effects of Hwanhonsan extract against cancer, and to study some mechanisms responsible for its effect. Some kind of tumors were induced by the typical application of 3-methylcholanthrene(MCA) or by the implantation of malignant tumor cells such as leukemia cells(3LL cells) or sarcoma cells(S180 cells) and FasII cells. Treatment of the Hwanhonsan extract(daily 1 mg/mouse, i.p.) was continued for 7 days prior to tumor induction and after that the treatment was lasted for 20 hrs. Against squamous cell carcinoma induced by MCA, Hwanhonsan decreased. not only the frequency of tumor production but also the number and weight of tumors per tumor bearing mice(TBM). Hwanhonsan also significantly suppressed the development of 3LL cells and S180 cells implanted tumors by frequency and their size, and some developed tumors were regressed by the continuous treatment of Hwanhonsan extract into TBM. However, when tumor was induced by FsaII cells implantation, the growth of implanted cells in mice was delayed by the water extract of Hwanhonsan until 7 days and then rapid growth ensued. In vitro treatment of Hwanhonsan extract had no inhibitory effect on the tumor induced by some kind of cell lines such as A431 cells strain but it significantly inhibited the proliferation of 3LL cells, S180 cells. These results suggested that Hwanhonsan extract exhibited a significant prophylactic benefits against tumors and its antitumor activity was manifested depending on the type of tumor cells.

• 한국식품과학회지
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• 제34권3호
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• pp.505-509
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• 2002

천마의 methanol 추출물로부터 VHR DS-PTPase 저해 물질을 분리하여 이를 HREI-MS와 $^1H-NMR$, $^{13}C-NMR$, DEPT 등의 기기 분석 자료에 의하여 baicalein으로 구조를 동정 하였다. 이 물질은 VHR에 대하여 $2.4{\mu}M$의 $IC_{50}$값을 나타내었고 T-cell PTPase나 PPase 1과 같은 다른 단백질 탈인산화 효소에 대하여는 저해 활성을 나타내지 않았다. 또한 7종류의 인간 암세포주(흑색종 세포주인 LOX-IMVI, 폐암 세포주인 NCI H23과 A549, 대장암 세포주인 HCT 116와 SW 620, 전립선암 세포주인 PC-3와 백혈병 세포주인 MOLT 4F)에 대한 세포독성을 조사하여 본 결과 $5.26{\sim}12.93\;{\mu}g/mL$에서 $GI_{50}$값을 나타내었다.

• Clinical and Experimental Pediatrics
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• 제49권4호
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• pp.439-445
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• 2006

목 적 : ${\alpha}$-Galactosylceramide (GalCer)로 자극한 자연살상 T 세포(NKT)는 CD1d 및 T 세포 수용체(T cell receptor) 의존적으로 일부 백혈병에서 항암효과를 발현하나, CD1d음성인 신경모세포종에서는 세포독성을 유도할 수 없다. 이들 NKT세포의 활성화 시 분비되는 많은 양의 사이토카인의 직접적인 항암효과에 대해서는 소수의 보고가 있다. 본 연구에서는 hCD1d/${\alpha}$-GalCer tetramer로 NKT세포를 자극하여 얻은 상청액(supernatant)을 이용하여 NKT세포에 의한 신경모세포종의 치료적 접근의 가능성을 알아보았다. 방 법 : 신경모세포종 세포 주를 IMDM 배지에 배양하였고, NKT세포에서 분비되는 사이토카인 양은 cytometric bead array (CBA)분석으로 측정하였다. 세포 생존율은 calcein-AM 형광물질을 이용하여 digital image microscopy scanning (DIMSCAN)으로 측정하였고 특이 세포고사(specific apoptosis)는 annexin V and 7-AAD 염색 후 유식세포분석기를 통하여 산출하였다. 결 과 : 활성화된 NKT세포는 많은 양의 IL-2, IL-4, INF-${\gamma}$와 TNF-${\alpha}$을 분비하였다. NKT 자극 후 얻어진 상청액은 8개의 신경모세포종 세포 주 중 4개에서 의미있는 세포독성을 나타냈으며, 그 기전은 annexin-V 염색이나 pancaspase 억제제의 전 처치 실험으로 세포고사을 통하여 유도됨을 알 수 있었다. 그리고 이들 세포고사 유도는 anti-TNF-${\alpha}$, anti-IFN-${\gamma}$ 중화항체의 단독투여 시 현저히 감소하였고 동시투여 시에는 완전하게 억제되었다. 결 론 : NKT 세포의 활성화에 의해 분비된 IFN-${\gamma}$와 TNF${\alpha}$가 일부 신경모세포종 세포 주에서 협동적 세포 독성을 유도하였다.

• IMMUNE NETWORK
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• 제1권1호
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• pp.87-94
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• 2001

목적 : 제대혈의 조혈모세포 체외확장 시 조혈세포 증폭과 더불어 조혈미세환경의 변화가 일어난다. 이때 제대혈 $CD34^+$ 세포에서 유래되는 지지세포의 계열 분석조혈성장인자 분비능력을 알아보고 지지세포 증식 조건을 확립하여 효과적인 제대혈의 체외증폭을 제시하고자 하였다. 방법 : 제대혈부터 $CD34^+$ 세포를 분리하여 실험에 사용하였다. 무혈청배지에서 각종 조혈성장인자를 다양한 조합으로 첨가하여 배양하였고 증식정도는 현미경으로 관찰하여 배양용기를 점유한 면적 비율로 계산하였다. 세포외간질 단백의 효과를 분석하기 위하여 collagen S, fibronectin, laminin 및 poly-L-ly sine를 미리 coating한 용기에 배양하여 분석하였다. 제대혈 $CD34^+$ 세포를 조혈성장인자의 첨가 없이 3주간 액체배양하였다. 배양 시, 1주, 2주 및 3주에 상층액을 얻어 $-80^{\circ}C$에 보관하였다가 한꺼번에 IL-3, IL-6, GM-CSF, IL-$1{\beta}$ 및 TNF-$\alpha$등을 ELISA 방법으로 내부적으로 분비되는 량을 측정하였다. 분화된 지지세포의 계열을 분석하기 위해 E-selectin, VCAM-1, ICAM-1, PECAM-1, vWF, vimentin 및 CD 14 항체를 이용하여 면역화학염색 후 형광현미경으로 관찰하였다. 결과 : 제대혈 $CD34^+$ 세포 체외증폭시키는 과정에서 배양 4일에 지지세포가 출현하기 시작하여 7-10일이 지나면서 증식하기 시작하였고 14-2 1일 경에 서로 뭉치는 양상을 보여주었다. 제대혈 $CD34^+$ 세포 배양하면서 내부적으로 분비되는 GM-CSF, IL-6의 측정치는 시간이 지남에 따라 증가되었다. 제대혈 $CD34^+$ 세포 체외확장 시 지지세포의 증식 정도는 TPO+FL+SCF+LIF의 조합의 조혈성장인자가 첨가되었을 때 그리고 세포외간질 단백 성분 중 1% poly-L-lysine으로 처리한 경우 가장 효과적이었다. 결론 : 체외 증폭시 제대혈 $CD34^+$ 세포로부터 지지세포가 나타났으며 적절한 조혈성장인자의 첨가나 세포외간질 단백의 첨가에 의해 증폭될 수 있다.