Rajagopal Senthilkumar, Nalli Ancy D, Kumar Divya P, Bhattacharya Sayak, Hu Wenhui, Mahavadi Sunila, Grider John R, Murthy Karnam S
Department of Physiology and Biophysics, VCU Program in Enteric Neuromuscular Sciences, Virginia Commonwealth University, Richmond, Virginia.
Department of Physiology and Biophysics, VCU Program in Enteric Neuromuscular Sciences, Virginia Commonwealth University, Richmond, Virginia
J Pharmacol Exp Ther. 2015 Mar;352(3):509-18. doi: 10.1124/jpet.114.221929. Epub 2014 Dec 30.
The effect of proinflammatory cytokines on the expression and activity of soluble guanylyl cyclase (sGC) and cGMP-phosphodiesterases (PDEs) was determined in intestinal longitudinal smooth muscle. In control muscle cells, cGMP levels are regulated via activation of sGC and PDE5; the activity of the latter is regulated via feedback phosphorylation by cGMP-dependent protein kinase. In muscle cells isolated from muscle strips cultured with interleukin-1β (IL-1β) or tumor necrosis factor α (TNF-α) or obtained from the colon of TNBS (2,4,6-trinitrobenzene sulfonic acid)-treated mice, expression of inducible nitric oxide synthase (iNOS) was induced and sGC was S-nitrosylated, resulting in attenuation of nitric oxide (NO)-induced sGC activity and cGMP formation. The effect of cytokines on sGC S-nitrosylation and activity was blocked by the iNOS inhibitor 1400W [N-([3-(aminomethyl)phenyl]methyl)ethanimidamide dihydrochloride]. The effect of cytokines on cGMP levels measured in the absence of IBMX (3-isobutyl-1-methylxanthine), however, was partly reversed by 1400W or PDE1 inhibitor vinpocetine and completely reversed by a combination of 1400W and vinpocetine. Expression of PDE1A was induced and was accompanied by an increase in PDE1A activity in muscle cells isolated from muscle strips cultured with IL-1β or TNF-α or obtained from the colon of TNBS-treated mice; the effect of cytokines on PDE1 expression and activity was blocked by MG132 (benzyl N-[(2S)-4-methyl-1-[[(2S)-4-methyl-1-[[(2S)-4-methyl-1-oxopentan-2-yl]amino]-1-oxopentan-2-yl]amino]-1-oxopentan-2-yl]carbamate), an inhibitor of nuclear factor κB activity. NO-induced muscle relaxation was inhibited in longitudinal muscle cells isolated from muscle strips cultured with IL-1β or TNF-α or obtained from the colon of TNBS-treated mice, and this inhibition was completely reversed by the combination of both 1400W and vinpocetine. Inhibition of smooth muscle relaxation during inflammation reflects the combined effects of decreased sGC activity via S-nitrosylation and increased cGMP hydrolysis via PDE1 expression.
在肠道纵行平滑肌中测定了促炎细胞因子对可溶性鸟苷酸环化酶（sGC）和环鸟苷酸磷酸二酯酶（PDEs）表达及活性的影响。在对照肌细胞中，cGMP水平通过sGC和PDE5的激活来调节；后者的活性通过依赖cGMP的蛋白激酶的反馈磷酸化来调节。在用白细胞介素-1β（IL-1β）或肿瘤坏死因子α（TNF-α）培养的肌条中分离得到的肌细胞，或从经2,4,6-三硝基苯磺酸（TNBS）处理的小鼠结肠中获取的肌细胞，诱导型一氧化氮合酶（iNOS）的表达被诱导，且sGC发生S-亚硝基化，导致一氧化氮（NO）诱导的sGC活性和cGMP生成减弱。iNOS抑制剂1400W [N-([3-(氨甲基)苯基]甲基)乙亚胺酰胺二盐酸盐]可阻断细胞因子对sGC S-亚硝基化和活性的影响。然而，在不存在异丁基甲基黄嘌呤（IBMX）的情况下测定细胞因子对cGMP水平的影响时，1400W或PDE1抑制剂长春西汀可部分逆转该影响，而1400W和长春西汀联合使用则可完全逆转。在用IL-1β或TNF-α培养的肌条中分离得到的肌细胞，或从经TNBS处理的小鼠结肠中获取的肌细胞中，PDE1A的表达被诱导，同时PDE1A活性增加；细胞因子对PDE1表达和活性的影响可被核因子κB活性抑制剂MG132（苄基N-[(2S)-4-甲基-1-[[(2S)-4-甲基-1-[[(2S)-4-甲基-1-氧代戊烷-2-基]氨基]-1-氧代戊烷-2-基]氨基]-1-氧代戊烷-2-基]氨基甲酸酯）阻断。在用IL-1β或TNF-α培养的肌条中分离得到的肌细胞，或从经TNBS处理的小鼠结肠中获取的纵行肌细胞中，NO诱导的肌肉舒张受到抑制，而1400W和长春西汀联合使用可完全逆转这种抑制。炎症期间平滑肌舒张的抑制反映了通过S-亚硝基化导致的sGC活性降低以及通过PDE1表达导致的cGMP水解增加的综合作用。
