소교세포에서 heme oxygenase-1 발현 유도를 통한 치자(Gardenia jasminoides)의 항염증 효과 Gardenia jasminoides Exerts Anti-inflammatory Activity via Akt and p38-dependent Heme Oxygenase-1 Upregulation in Microglial Cells원문보기
치자 열매는 아시아에서 음식과 옷의 염료로 사용되어 왔다. 본 연구에서는 BV-2소교세포에서 치자열매 열수추출물(GJ)의 항염증 효과를 관찰하고 그 작용 기전을 연구하였다. GJ는 세포에 독성을 유도하지 않으면서 lipopolysaccharide로 인한 nitric oxide (NO) 분비와 inducible nitric oxide synthase (iNOS) 생성 및 활성산소 생성을 억제하였다. 또한 GJ는 농도의존적으로 heme oxygenase-1 (HO-1)의 발현을 유도하였다. 더군다나 HO-1 siRNA를 처리했을 때는 GJ가 iNOS의 발현을 억제하지 못하였다. GJ는 HO-1의 발현에 관여하는 전사인자인 nuclear factor E2-related factor 2를 핵으로 이동시켰다. 또한 GJ에 의한 HO-1의 발현은 phosphatidylinositol 3-kinase(PI-3K) 및 p38 kinase 억제제에 의해 감소되었으며, GJ가 Akt와 p38 kinase의 인산화를 유도하였다. 이상의 결과를 종합해보면, GJ는 PI3K/Akt 및 p38 신호전달과정을 통해 HO-1의 발현을 유도함으로써 NO와 같은 염증매개물질의 생성을 억제한다는 것을 알 수 있다. 이러한 연구결과는 치자열매가 신경염증을 억제하는 새로운 기전을 밝힌 것이다.
치자 열매는 아시아에서 음식과 옷의 염료로 사용되어 왔다. 본 연구에서는 BV-2 소교세포에서 치자열매 열수추출물(GJ)의 항염증 효과를 관찰하고 그 작용 기전을 연구하였다. GJ는 세포에 독성을 유도하지 않으면서 lipopolysaccharide로 인한 nitric oxide (NO) 분비와 inducible nitric oxide synthase (iNOS) 생성 및 활성산소 생성을 억제하였다. 또한 GJ는 농도의존적으로 heme oxygenase-1 (HO-1)의 발현을 유도하였다. 더군다나 HO-1 siRNA를 처리했을 때는 GJ가 iNOS의 발현을 억제하지 못하였다. GJ는 HO-1의 발현에 관여하는 전사인자인 nuclear factor E2-related factor 2를 핵으로 이동시켰다. 또한 GJ에 의한 HO-1의 발현은 phosphatidylinositol 3-kinase(PI-3K) 및 p38 kinase 억제제에 의해 감소되었으며, GJ가 Akt와 p38 kinase의 인산화를 유도하였다. 이상의 결과를 종합해보면, GJ는 PI3K/Akt 및 p38 신호전달과정을 통해 HO-1의 발현을 유도함으로써 NO와 같은 염증매개물질의 생성을 억제한다는 것을 알 수 있다. 이러한 연구결과는 치자열매가 신경염증을 억제하는 새로운 기전을 밝힌 것이다.
Died Gardenia jasminoides fruit is used as a dye in the food and clothes industries in Asia. The present study investigated the anti-inflammatory effects of aqueous extract of G. jasminoides fruits (GJ) in BV-2 microglial cells. GJ inhibited lipopolysaccharide-induced nitric oxide (NO) secretion, in...
Died Gardenia jasminoides fruit is used as a dye in the food and clothes industries in Asia. The present study investigated the anti-inflammatory effects of aqueous extract of G. jasminoides fruits (GJ) in BV-2 microglial cells. GJ inhibited lipopolysaccharide-induced nitric oxide (NO) secretion, inducible nitric oxide synthase (iNOS) expression, and reactive oxygen species production, without affecting cell viability. Furthermore, GJ increased the expression of heme oxygenase-1 (HO-1) in a dose-dependent manner. Moreover, the inhibitory effect of GJ on iNOS expression was abrogated by small interfering RNA-mediated knock-down of HO-1. In addition, GJ induced nuclear translocation of nuclear factor E2-related factor 2 (Nrf2), a transcription factor that regulates HO-1 expression. GJ-mediated expression of HO-1 was suppressed by LY294002, a phosphoinositide 3-kinase (PI-3K) inhibitor, and SB203580, a p38 kinase inhibitor, but not by the extracellular signal-regulated kinase (ERK) inhibitor PD98059 or c-Jun N-terminal kinase (JNK) inhibitor SP600125. GJ also enhanced the phosphorylation of Akt and p38. These results suggest that GJ suppresses the production of NO, a pro-inflammatory mediator, by inducing HO-1 expression via PI-3K/Akt/p38 signaling. These findings illustrate a novel molecular mechanism by which extract from G. jasminoides fruits inhibits neuroinflammation.
Died Gardenia jasminoides fruit is used as a dye in the food and clothes industries in Asia. The present study investigated the anti-inflammatory effects of aqueous extract of G. jasminoides fruits (GJ) in BV-2 microglial cells. GJ inhibited lipopolysaccharide-induced nitric oxide (NO) secretion, inducible nitric oxide synthase (iNOS) expression, and reactive oxygen species production, without affecting cell viability. Furthermore, GJ increased the expression of heme oxygenase-1 (HO-1) in a dose-dependent manner. Moreover, the inhibitory effect of GJ on iNOS expression was abrogated by small interfering RNA-mediated knock-down of HO-1. In addition, GJ induced nuclear translocation of nuclear factor E2-related factor 2 (Nrf2), a transcription factor that regulates HO-1 expression. GJ-mediated expression of HO-1 was suppressed by LY294002, a phosphoinositide 3-kinase (PI-3K) inhibitor, and SB203580, a p38 kinase inhibitor, but not by the extracellular signal-regulated kinase (ERK) inhibitor PD98059 or c-Jun N-terminal kinase (JNK) inhibitor SP600125. GJ also enhanced the phosphorylation of Akt and p38. These results suggest that GJ suppresses the production of NO, a pro-inflammatory mediator, by inducing HO-1 expression via PI-3K/Akt/p38 signaling. These findings illustrate a novel molecular mechanism by which extract from G. jasminoides fruits inhibits neuroinflammation.
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문제 정의
jasminoides have been used as a natural color additive in food and known to have anti-inflammatory effects. In this study, we assessed the possible molecular mechanism underlying its anti-inflammatory effect. We found that GJ significantly inhibited LPS-induced NO synthesis and iNOS expression as well as ROS production in BV-2 microglial cells without appreciable cytotoxic effects.
제안 방법
To elucidate the molecular target of GJ in further upstream signaling pathway of HO-1 expression, we examined the effect of pharmaceutical protein kinase inhibitors, LY294002 (PI-3K inhibitor), SP600125 (JNK inhibitor), PD 98059 (ERK inhibitor), and SB203580 (p38 kinase inhibitor). As shown in Fig.
To investigate whether GJ induces HO-1 expression in microglia, BV-2 cells were incubated with various concentrations of GJ. The HO-1 protein level was significantly increased by GJ in a dose-dependent manner (Fig.
대상 데이터
Antibodies against phosphorylated Akt (p-Akt) and p-p38 were purchased from Cell Signaling Technology (Beverly, MA, USA). Actinomycin D (Act. D) and cycloheximide (CHX) were purchased from Sigma-Aldrich Co. (St. Louis, MO). CM-H2DCFDA and fetal bovine serum (FBS) were purchased from Invitrogen Corporation (San Diego, CA, USA).
Cobalt protoporphyrin (CoPP), HO-1 siRNA, and antibodies for iNOS, HO-1, Nrf-2, Akt, p38, histone deacetylase (HDAC), and α-tubulin were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).
The siRNAs for HO-1 (GenBank accession no. NM 010442.1) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Cells were transfected with HO-1 siRNA or negative control siRNA using INTERFERin (Polyplus transfection, France).
데이터처리
Statistical significances were compared between each treated group and analyzed by the Student’s t-test.
이론/모형
NO synthesis in cell cultures was measured by a microplate assay method. To measure nitrite, 100 μl aliquots were removed from conditioned medium and incubated with an equal volume of the Griess reagent (1% sulfanilamide/0.
BV-2 cells were incubated with GJ for 3 hr and stimulated with LPS for 24 hr. The amount of NO released into culture medium was measured by the method of Griess. As shown in Fig.
(B) Cells were incubated with various concentrations of GJ for 3 hr and then stimulated with LPS (1 μg/ml) for 24 hr. The amount of nitrite released was measured by the method of Griess. * p<0.
The cytotoxicity of GJ was assessed using the microculture tetrazolium (MTT)-based colorimetric assay. The remaining cells after Griess reaction were used for MTT assay.
성능/효과
In conclusion, we demonstrated that GJ inhibited NO release and iNOS expression in LPS-stimulated macrophages, and these effects are mediated by PI-3K/Akt and p38-dependent HO-1 expression. Our finding could help us to understand the molecular mechanism of anti-inflammatory action of GJ, although it was examined in vitro, and suggest that GJ may have therapeutic potential for treatment of neuroinflammatory diseases.
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