• Tuberculosis and Respiratory Diseases
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• 제44권6호
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• pp.1353-1365
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• 1997

연구배경 : 종양괴사인자(tumor necrosis factor ; TNF)는 다양한 생물학적 기능을 가지고 있는 바, 그 중 생체 외에서 증명된 뚜렷한 항암 효과로 말미암아 최근 항암 유전자요법의 중요한 대상으로 관심을 모으고 있다. 현재 유전자 이입의 기술적 문제로 생체 외에서 암세포에 유전자 이입을 시행한 후 이를 다시 환자의 생체내로 이식하는 방법이 연구의 주종을 이루고 있다. 그러나 저자들의 과거의 연구를 포함한 여러 연구에서 TNF가 이입된 암세포는 TNF에 대해 내성을 보이는 이에는 새로이 방어 단백질을 합성하는 것이 관여할 것이라는 시사가 있었다. 이 획득내성의 기전을 밝히는 것이 종양생물학의 이해를 넓히고 보다 효과적인 항암 유전자요법을 개발하기위한 매우 중요한 과제로 생각된다. 저자들은 TNF 유전자 이입에 따른 암세포의 TNF에 대한 획득내성에, 일부 세포에서 TNF에 의해 발현이 유도된다는 것이 밝혀진, 항산화효소의 하나인 MnSOD의 발현의 변화가 관여하는 지를 규명하고자 본 실험을 수행하였다. 방 법 : TNF에 다양한 감수성을 보이는 인체 및 생쥐 기원의 4가지 암세포주(WEHI164, NCI-H2058, A549, ME180)에 TNF-$\alpha$ 유전자를 retroviral 이용하여 이입하고 TNF의 발현을 시도하여 PCR, ELISA, MTT assay로 확인하였고, TNF 유전자가 이입된 세포(WEHI164-TNF, NCI-H2058-TNF, A549-TNF, ME180-TNF)는 TNF에 내성을 보이는지 역시 MTT assay로 검증하였다. TNF 유전자 이입 전후의 MnSOD mRNA 발현의 차이는 Northern blot analysis를 통하여 비교하였다. 결 과 : 1) TNF-$\alpha$ 유천자 이입 및 발현 확인 PCR을 시행한 결과 TNF 유전자가 이입된 각 세포 790 base pair 표기의 진한 DNA band를 보인 반면 모세포주는 보이지 않아서 retroviral vector를 이용한 유전자 이입이 DNA 수준에서 이루어 있었다. 그리고 TNF 유전자가 이입된 세포의 배양상층액에서 TNF양을 ELISA로 측정한 결과 TNF를 세포에 따라 1.91ng/24hr/$10^6\;cells$에서 3.91ng/24hr/$10^6\;cells$ 생산함을 알 수 있었다. 2) TNF 유전자 이입 전후, 암세포의 TNF에 대한 감수성 비교 TNF 농도 100ng/ml에서 WEHI164-TNF와 ME180-TNF 세포는 통계적으로 유의하게 (p<0.01) TNF에 대한 내성을 획득함을 알 수 있었다. 3) TNF 유전자 이입 전후외 MnSOD mRNA 발현양상 TNF에 감수성을 보이는 WEHl164와 ME180세포는 TNF 유전자 이입 후에 MnSOD mRNA 발현이 증가되지 않았으며, TNF에 내성을 보이는 NCIH2058과 A549 세포는 TNF 유전자 이입 후에 MnSOD mRNA 발현의 증가가 관찰되었으나 이는 모세포에 외부에서 TNF를 주었을 때도 관찰되었다. 결 론 : 세포주에 TNF 유전자를 이입하여 TNF를 발현하게 하였을때 세포 자신은 TNF에 대해 내성을 하게 되는데, 이 획득내성은 MnSOD 발현 능력의 향상에 의한 것은 아닐 것으로 판단되나 각 세포주의 자연 상태의 TNF에 대한 감수성 여부는 MnSOD의 발현 차이가 관련될 것으로 생각된다.

• 약학회지
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• 제43권4호
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• pp.474-480
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• 1999

The purpose of this study is to define whether Asterina pectinifera Lectin (APL) is effective on the cytokine production. Isolated mRNA from hPBMC (human peripheral blood mononuclear cells) stimulated with APL for various reaction times (1 to 96 hours) was detected by RT-PCR. The intensity of band for IL-1 and $IFN{\gamma}$ mRNA was markedly increased at l hour, and IL-2 mRNA was strongly expressed at 4 hours. The mRNA band of APL-induced IL-2 and $IFN{\gamma}$ was weaker than that of IL-1, IL-6 and $TNF{\alpha}$. The mRNA expression of 4 cytokines (IL-1, IL-2, $IFN{\gamma}$ and $TNF{\alpha}$) was detected up to 48 hours, and that of IL-6 was detected until 72 hours. ELISA was used to look protein secretion of the cytokine gene with IL-1, IL-2 and TNF$\alpha$expressed strongly in RT-PCR. The highest protein secretion was at 4 hours with IL-1, at 8 hours with IL-2 and at 4 hours with $TNF{\alpha}$. These results suggest that APL can induce the production of some cytokines and the immune response from PBMC was done within the first few hours of stimulation with APL.

• Korean Journal of Acupuncture
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• 제36권3호
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• pp.162-170
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• 2019

Objectives : Increasing evidence suggests that parasympathetic vagus nerve activity plays a role in modulating acupuncture-induced anti-inflammatory reaction, but the function of sympathetic nerve is not known. Here, we investigated whether splanchnic sympathetic nerve activity was involved in the regulation of splenic expression of $TNF-{\alpha}$ mRNA by electroacupuncture (EA) in LPS-injected animals. Methods : DiI was injected into the stomach or celiac ganglion (CG) for retrograde labeling of the target tissues. EA was given at ST36 and the electrical stimulation on the sciatic nerve in LPS-injected mice. c-Fos signals in the tissues were analyzed by immunofluorescence staining, and $TNF-{\alpha}$ mRNA was analyzed by real-time PCR. Results : Application of EA at ST36 or electrical stimulation on the sciatic nerve induced c-Fos expression in neurons of the spinal cord and celiac ganglion (CG). Then, the vagotomy reduced c-Fos levels in CG neurons but not in the spinal cord in animals given EA. Expression of $TNF-{\alpha}$ mRNA which was induced in the spleen after LPS was significantly inhibited by EA, then the vagotomy elevated $TNF-{\alpha}$ mRNA level similar to that in LPS-injected animals. Splanchnectomy in animals given LPS and EA also increased $TNF-{\alpha}$ mRNA though it was less effective than vagotomy. Conclusions : Our data suggest that EA delivered to the spleen via the splanchnic sympathetic nerve may be involved in attenuating splenic inflammatory responses in LPS-injected animals.

• 대한한방부인과학회지
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• 제21권4호
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• pp.49-68
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• 2008

Purpose: This study was performed to evaluate anti-inflammatory effects of Cheongyeoljohyeoltangkamibang water extract (CYJHT). Methods: In the study of anti-inflammatory effects. CYJHT was investigated using cultured cells and murine models. As for the parameters of inflammation. levels of several inflammatory cytokines and chemical mediators which are known to be related to inflammation were determined in mouse lung fibroblast cells(mLFCs). RAW 264.7 cells and acute inflammation-induced mice. Results: 1. CYJHT showed a safety in cytotoxicity and toxicity of liver. 2. CYJHT effected scavenging activity on 2.2-diphenyl-1-picrylhydrazyl(DPPH) free radical, superoxide dismutase(SOD) and superoxide anion radical(SAR). 3. CYJHT in RAW 264.7 cell decreased IL-l$\beta$ mRNA expression at 100, 50 ${\mu}g$/ml and also decreased TNF-$\alpha$ mRNA expression at 100 ${\mu}g/ml$ and decreased COX-2. NOS-II mRNA expression and decreased IL-6 mRNA expression in a concentration-dependent manner. 4. CYJHT in RAW 264.7 cell decreased IL-l$\beta$ significantly at 100, 50 ${\mu}g$/ml and decreased IL-6. TNF-$\alpha$ significantly at 100 ${\mu}g$/ml. 5. CYJHT inhibited IL-l1$\beta$, IL-6 and TNF-$\alpha$ production significantly in serum of acute inflammation-induced mice. 6. CYJHT decreased IL-1$\beta$, IL-6 and TNF-$\alpha$ mRNA production significantly in spleen tissue. and also decreased IL-l$\beta$. TNF-$\alpha$ mRNA production significantly in liver tissue of acute inflammation-induced mice. Conclusion: These results suggest that CYJHT can be useful in treating diverse female diseases caused by inflammation such as menstrual pain. menstrual disorder. leukorrhea. pelvic inflammatory disease and so on.

• 한국약용작물학회지
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• 제15권1호
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• pp.30-37
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• 2007

한국산 겨우살이 (Viscum album)는 면역 활성효과와 항암효과가 있다는 것으로 잘 알려져 있다. 본 연구에서는 한국산 겨우살이 추출물 M11C (비렉틴 구성물질)가 사람의 말초혈액 단구를 활성화시켜 tumor necrosis factor-alpha $(TNF-\alpha)$를 생산 분비하게 하는지를 규명하기 위해 실험에 이용되었다. 단구로부터 $TNF-\alpha$의 생산에 있어 M11C의 효과를 알기 위해 단구를 여러 농도 $(0-2000{\mu}g/m\ell)$의 M11C로 0.5-24시간 동안 자극한 후 배양액 MCM을 수거했다. 수거한 배양액 MCM을 $TNF-\alpha$에 민감한 L929 세포에 첨가한 후 L929 세포의 독성 정도를 MTT 기법으로 검사하였는데, 배양액 MCM이 L929 독성효과를 가졌으며, 이 MCM의 L929 세포 독성효과는 $TNF-\alpha$ 항체에 의해 거의 완전하게 억제되었다. 이러한 결과는 배양액 MCM에 $TNF-\alpha$가 존재함을 지적했으며, 또한 M11C가 단구를 자극해 $TNF-\alpha$를 생산하리라고 암시했었다. 이러한 암시를 확인하기 위해 immunoblotting 기법을 사용해 배양액 MCM에 존재하는 $TNF-\alpha$을 검사를 수행했다. 검사결과 배양액 MCM에 많은 량의 $TNF-\alpha$가 검출되었다. 이러한 $TNF-\alpha$ 검출량은 단구를 자극한 M11C의 농도와 자극시간에 비례해서 증가했다. 즉, 단구로부터 $TNF-\alpha$의 생산 분비량은 M11C의 농도와 자극시간에 의존적이었다. 이러한 M11C의 농도와 자극시간의 의존적 효과는 단구로부터 $TNF-\alpha$ mRNA 발현에도 같은 경향을 보였다. 결론적으로 한국산 겨우살이로부터 열탕 추출한 M11C가 면역활성제로 작용할 수 있음을 시사함과 동시에 한방탕제로 사용되는 한국산 겨우살이가 면역학적 효능을 가지고 있음을 말해주고 있다.

• 대한본초학회지
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• 제28권2호
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• pp.93-101
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• 2013

Objects : Baicalein is a major bioactive flavonoid component of Scutellaria baicalensis Georgi that shows a wide range of biological activities, including neuroprotections and anti-inflammatory actions. Hence it is a potential therapeutic material for the treatment of neuroinflammation. In this study, we investigated the modulatory effect of baicalein on neuroinflammation. Method : Pro-inflammatory cytokines (TNF-${\alpha}$, IL-$1{\beta}$ and IL-6 mRNA), COX-2 mRNA expression and microglial activation in the brain tissue is induced by systemic lipopolysaccharide (LPS) treatment in C57BL/6 mice. Baicalein was treated orally with 10, 20, and 30 mg/kg 1 hour prior to the LPS (3 mg/kg, i.p.) injection. TNF-${\alpha}$, IL-$1{\beta}$, IL-6 and COX-2 mRNA expression in the brain tissue was measured by the quantitative real-time polymerase chain reaction(PCR) method. Iba1 expression in the brain was measured by western blotting method. Microglia was observed with immunohistochemistry. Results : Baicalein 30 mg/kg significantly attenuated the expression of TNF-${\alpha}$, IL-$1{\beta}$, IL-6 and COX-2 mRNA in the brain tissue. Baicalein 20 mg/kg significantly attenuated the expression of IL-6 mRNA in the brain tissue. Baicalein 30 mg/kg significantly attenuated the expression of Iba1 protein expression in the brain tissue. Baicalein 30 mg/kg significantly decreased the number and cell size of microglia in the cerebral cortex and hypothalamic region and the area percentage of Iba1-expressed microglia in the hippocampus. Conclusion : These results demonstrated that baicalein attenuates LPS induced neuroinflammation in the mice via reduction of pro-inflammatory cytokines (TNF-${\alpha}$, IL-$1{\beta}$, IL-6), COX-2 mRNA expression and microglial activation.

• Molecular & Cellular Toxicology
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• 제4권2호
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• pp.106-112
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• 2008

Parkinson's disease (PD) progresses severely by a gradual loss of dopaminergic neurons in the substantia nigra (SN). Epidemiological studies showed that the incidences of PD were reduced by smoking of which the major component, nicotine might be neuroprotective. But the function of nicotine, which might suppress the incidences of PD, is still unknown. Fortunately, recently it was reported that a glial reaction and inflammatory processes might participate in a selective loss of dopaminergic neurons in the SN. The levels of tumour necrosis factor (TNF)-${\alpha}$ synthesised by astrocytes and microglia are elevated in striatum and cerebrospinal fluid (CSF) in PD. TNF-${\alpha}$ kills the cultured dopaminergic neurons through the apoptosis mechanism. TNF-${\alpha}$ release from glial cells may mediate progression of nigral degeneration in PD. Nicotine pretreatment considerably decreases microglial activation with significant reduction of TNF-${\alpha}$ mRNA expression and TNF-${\alpha}$ release induced by lipopholysaccharide (LPS) stimulation. Thus, this study was intended to explore the role of nicotine pretreatment to inhibit the expressions of TNF-${\alpha}$ mRNA in human fetal astrocytes (HFA) stimulated with IL-$1{\beta}$. The results are as follows: HFA were pretreated with 0.1, 1, and $10{\mu}g/mL$ of nicotine and then stimulated with IL-$1{\beta}$ (100 pg/mL) for 2h. The inhibitory effect of nicotine on expressions of TNF-${\alpha}$ mRNA in HFA with pretreated $0.1{\mu}g/mL$ of nicotine was first noted at 8hr, and the inhibitory effect was maximal at 12 h. The inhibitory effect at $1{\mu}g/mL$ of nicotine was inhibited maximal at 24 h. Cytotoxic effects of nicotine were noted above $10{\mu}g/mL$ of nicotine. Moreover, Nicotine at 0.1, 1 and $10{\mu}g/mL$concentrations significantly inhibited IL-$1{\beta}$-induced TF-${\kappa}B$ activation. Collectively, these results indicate that in activated HFA, nicotine may inhibit the expression of TNF-${\alpha}$ mRNA through the pathway which suppresses the NF-${\kappa}B$ activation. This study suggests that nicotine might be neuroprotective to dopaminergic neurons in the SN and reduce the incidences of PD.

• 한국식품과학회지
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• 제36권6호
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• pp.986-990
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• 2004

대식세포는 IL-1, IL-6, $TNF-{\alpha},\;IFN-{\gamma}$의 중요한 근원이고, 전염성의 병원체와 종양 세포에 대해 저항하는 주요한 effector 세포이다. 이것은 cytokine signal에 의해 세포파괴가 되도록 활성화 된다. 이번 연구에서 우리는 산더덕의 물 추출물이 cytokine, nitric oxide와 같은 effector 분자를 유도하기 위해 대식세포를 활성화하는지를 알아보기 위하여 실험하였다. 산더덕 추출물에 의한 대식세포의 NO 생성은 농도 의존적이고, 시간 의존적이었다. iNOS는 시간이 지날수록 발현이 유도되었다. 산더덕 추출물에 의한 IL-1과 IL-6 mRNA 유도는 시간 의존적으로 증가 되었고, 처리 후 24시간에 최고점에 도달하였다. 이것은 활성화된 대식세포가 종양세포를 죽일 수 있음을 말한다. $TNF-{\alpha}\;mRNA$의 양은 시간이 지나도 일정하였고, $IFN-{\gamma}\;mRNA$의 양은 산더덕 추출물의 처리 1시간 후에 빠르게 강화되었다. 이러한 결과로부터 산더덕은 효과적인 면역조절자이고 대식세포의 항종양활성을 강화함을 알 수 있다.

• 융합정보논문지
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• 제9권6호
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• pp.218-226
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• 2019

본 연구의 목적은 삼내자(Kaempferia Galanga) 열수추출물의 항산화, 세포독성, 항염 효능을 조사하여 화장품 성분으로서 활용 가능성을 알아보고자 하였다. 항산화 활성 평가를 위해 DPPH, ABTS 라디컬 소거능 활성, FRAP 환원력, 총폴리페놀 함량을 측정하였다. 쥐의 대식세포인 RAW264.7 세포를 대상으로 MTT assay를 수행하여 세포독성을 확인하였다. 항염 효능을 알아보기 위해 LPS로 유도된 RAW264.7 세포에서 nitric oxide(NO), $TNF-{\alpha}$ 분비 및 iNOS, $TNF-{\alpha}$ mRNA 발현 수준을 조사하였다. 그 결과 DPPH와 ABTS 라디칼 소거능은 농도 의존적으로 라디칼 소거 활성이 증가하였다. FRAP 값은 5 mg/mL에서 24.5 uM로 가장 높게 나타났으며, 총폴리페놀 함량은 $1.28{\pm}0.064mg\;GAE/g$로 나타났다. KG 추출물 농도 0.625~2.5 mg/mL에서 세포독성은 보이지 않았다. 또한 RAW264.7 세포에서 LPS 유도에 의한 NO, $TNF-{\alpha}$ 분비 그리고 iNOS, $TNF-{\alpha}$의 mRNA 발현이 KG 추출물에 의해 유의하게 감소하였다. 이러한 결과는 삼내자 열수추출물이 안전하고 효과적인 화장품 원료로서의 활용 가능성이 있음을 보여준다.

• Biomolecules & Therapeutics
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• 제20권3호
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• pp.293-298
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• 2012

The purpose of this study was to investigate the modification of expression and functionality of the drug transporter P-glycoprotein (P-gp) by tumor necrosis factor-alpha (TNF-${\alpha}$) and interferon-gamma (IFN-${\gamma}$) at the blood-brain barrier (BBB). We used immortalized human brain microvessel endothelial cells (iHBMEC) and primary human brain microvessel endothelial cells (pHBMEC) as in vitro BBB model. To investigate the change of p-gp expression, we carried out real time PCR analysis and Western blotting. To test the change of p-gp activity, we performed rhodamin123 (Rh123) accumulation study in the cells. In results of real time PCR analysis, the P-gp mRNA expression was increased by TNF-${\alpha}$ or IFN-${\gamma}$ treatment for 24 hr in both cell types. However, 48 hr treatment of TNF-${\alpha}$ or IFN-${\gamma}$ did not affect P-gp mRNA expression. In addition, co-treatment of TNF-${\alpha}$ and IFN-${\gamma}$ markedly increased the P-gp mRNA expression in both cells. TNF-${\alpha}$ or IFN-${\gamma}$ did not influence P-gp protein expression whatever the concentration of cytokines or duration of treatment in both cells. However, P-gp expression was increased after treatments of both cytokines together in iHBMEC cells only compared with untreated control. Furthermore, in both cell lines, TNF-${\alpha}$ or IFN-${\gamma}$ induced significant decrease of P-gp activity for 24 hr treatment. And, both cytokines combination treatment also decreased significantly P-gp activity. These results suggest that P-gp expression and function at the BBB is modulated by TNF-${\alpha}$ or/and IFN-${\gamma}$. Therefore, the distribution of P-gp depending drugs in the central nervous system can be modulated by neurological inflammatory diseases.