阻斷細胞骨架動態影響腎臟絲球體細胞誘發型一氧化氮合成酶之活化

Perturbance of cytoskeleton dynamics modulate the activation of inducible nitric oxide synthase in glomerular mesangial cells

許淑晴(Shu-Ching Hsu)

指導教授 ： 林庭慧

中山醫學大學/醫學科技學院/生物醫學科學學系碩士班/碩士(2012年)

https://doi.org/10.6834/CSMU.2012.00126

摘要

一氧化氮 (nitric oxide，NO)為一簡單分子，可經由一氧化氮合成酶(nitric oxide synthase，NOS)作用而產生。腎臟絲球體細胞(glomerular mesangial cells，MES-13 cells)經由lipopolysaccharide (LPS)和interferon-γ (IFN-γ)的處理，可活化誘發型一氧化氮合成酶(inducible NOS，iNOS)，進而產生大量的NO。在本篇研究論文中，探討cytochalasin D (微絲阻斷劑)、β,β’-Iminodipropionitrile (IDPN，中間絲阻斷劑)和nocodazole (微管阻斷劑)對LPS/IFN-γ刺激所產生之NO的影響。在不同濃度的cytochalasin D及nocodazole處理下，MES-13細胞在型態上及細胞存活率有明顯的變化，但在IDPN的處理下卻無影響。實驗結果亦顯示，cytochalasin D的處理並不會對NO產量或是iNOS蛋白質有影響；而在nocodazole處理下，細胞存活率、NO產量及iNOS蛋白質皆明顯減少；此外，MES-13細胞處理IDPN後，細胞存活率並無明顯下降，但NO產量、iNOS mRNA和蛋白質表現有隨著IDPN濃度增加而減少的趨勢。進一步實驗結果顯示IDPN可減弱ERK 1/2 MAPK之活化，以及降低NF-κB轉移至細胞核中。因此，我們認為IDPN是藉由抑制NF-κB和ERK的訊息傳遞路徑而抑制iNOS基因的表現及降低NO的產量。

關鍵字

一氧化氮 ； 誘發型一氧化氮合成酶細胞骨架 ； 腎絲球細胞

並列摘要

Nitric oxide (NO) is a simple molecule that produced by nitric oxide synthase (NOS). Lipopolysaccharide (LPS) and interferon-γ (IFN-γ) stimulate the activation of inducible nitric oxide synthase (iNOS) and produce large amount of NO. In this study, the effect of cytochalasin D (an actin filament disrupting agent) , β,β’-Iminodipropionitrile (IDPN, an intermediate filaments disrupting agent), nocodazole (a microtubule disrupting agent) on LPS and IFN-γ-stimulated NO production in MES-13 cells were investigated. At different concentrations tested, the cell morphology and cell viability of LPS/ IFN-γ- treated MES-13 cells were dramatically altered after the treatment of cytochalasin D or nocodazole but not IDPN. NO production and iNOS protein expression showed no significant difference when treated with cytochalasin D. Nocodazole treatment reduced cell viabiblity, NO production and iNOS protein expression. Without affecting cell viability of MES-13 cells, IDPN decresed NO production, iNOS protein expression, and iNOS mRNA expression. IDPN decreased iNOS gene expressions through attenuated ERK1/2 MAPK activation, but had no effect on the p38 and JNK MAPKs, and attenuated nuclear fraction translocation of NF-κB. Thus, our data indicate that IDPN block LPS/IFN-γ- stimulated iNOS expression via inhibition of the ERK and NF-κB signaling pathways in MES-13 cells.

並列關鍵字

nitric oxide ； inducible nitric oxide synthase ； cytoskeleton ； glomerular mesangial cells

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