Fahmi H, Di Battista J A, Pelletier J P, Mineau F, Ranger P, Martel-Pelletier J
H pital Notre-Dame, Montreal, Quebec, Canada.
Arthritis Rheum. 2001 Mar;44(3):595-607. doi: 10.1002/1529-0131(200103)44:3<595::AID-ANR108>3.0.CO;2-8.
To determine the effects of peroxisome proliferator-activated receptor gamma (PPARgamma) agonists on interleukin-1 (IL-1) induction of nitric oxide (NO) and matrix metalloproteinase 13 (MMP-13) in human chondrocytes.
PPARgamma expression and synthesis in human chondrocytes were determined by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry, respectively. Chondrocytes were cultured with IL-1beta, tumor necrosis factor alpha (TNFalpha), and IL-17 in the presence or absence of PPARgamma agonists, and NO and MMP-13 synthesis and expression levels were measured. Transient transfection experiments were performed with the 7-kb inducible NO synthase (iNOS) and 1.6-kb MMP-13 human promoters, as well as with the PPARgamma expression vector and the activator protein 1 (AP-1) and nuclear factor kappaB (NF-kappaB) reporter constructs.
RT-PCR and immunohistochemical analysis revealed that human chondrocytes expressed and produced PPARgamma. Treatment of chondrocytes with PPARgamma ligands BRL 49653 and 15-deoxy-delta12,14-prostaglandin J2 (15d-PGJ2), but not with PPARalpha ligand Wy 14643, decreased IL-1beta-induced NO and MMP-13 production in a dose-dependent manner. In addition, both iNOS and MMP-13 messenger RNA were inhibited in the presence of 15d-PGJ2. The inhibitory effect of PPARgamma activation was not restricted to IL-1beta, since TNFalpha- and IL-17-induced NO and MMP-13 production were also inhibited by 15d-PGJ2. In transient transfection experiments, we showed that a constitutively active form of mitogen-activated protein kinase kinase kinase 1 (AMEKK-1) induced the MMP-13 and iNOS human promoter activity. This process was reduced by 15d-PGJ2 and further inhibited by cotransfection with a PPARgamma expression vector. Similarly, in a PPARgamma-dependent manner, 15d-PGJ2 inhibited deltaMEKK-1-induced AP-1- and NF-kappaB-luciferase reporter plasmid activation.
The findings of this study demonstrate that PPARgamma agonists inhibit IL-1beta induction of both NO and MMP-13 in human chondrocytes. The inhibition occurs at least at the transcriptional level through a PPARgamma-dependent pathway, probably by interfering with the activation of AP-1 and NF-kappaB.
确定过氧化物酶体增殖物激活受体γ（PPARγ）激动剂对人软骨细胞中白细胞介素-1（IL-1）诱导的一氧化氮（NO）和基质金属蛋白酶13（MMP-13）的影响。
分别通过逆转录聚合酶链反应（RT-PCR）和免疫组织化学法测定人软骨细胞中PPARγ的表达和合成。在存在或不存在PPARγ激动剂的情况下，将软骨细胞与IL-1β、肿瘤坏死因子α（TNFα）和IL-17一起培养，并测量NO和MMP-13的合成及表达水平。用7 kb的诱导型一氧化氮合酶（iNOS）和1.6 kb的MMP-13人类启动子，以及PPARγ表达载体、激活蛋白1（AP-1）和核因子κB（NF-κB）报告基因构建体进行瞬时转染实验。
RT-PCR和免疫组织化学分析显示人软骨细胞表达并产生PPARγ。用PPARγ配体BRL 49653和15-脱氧-Δ12,14-前列腺素J2（15d-PGJ2）处理软骨细胞，但不用PPARα配体Wy 14643处理，可剂量依赖性地降低IL-1β诱导的NO和MMP-13产生。此外，在15d-PGJ2存在的情况下，iNOS和MMP-13信使核糖核酸均受到抑制。PPARγ激活的抑制作用不限于IL-1β，因为15d-PGJ2也抑制TNFα和IL-17诱导的NO和MMP-13产生。在瞬时转染实验中，我们表明有丝分裂原激活蛋白激酶激酶激酶1（AMEKK-1）的组成型活性形式诱导了MMP-13和iNOS人类启动子活性。该过程被15d-PGJ2降低，并通过与PPARγ表达载体共转染进一步受到抑制。同样，以PPARγ依赖的方式，15d-PGJ2抑制了δMEKK-1诱导的AP-1和NF-κB荧光素酶报告质粒激活。
本研究结果表明，PPARγ激动剂抑制人软骨细胞中IL-1β诱导的NO和MMP-13产生。这种抑制至少在转录水平通过PPARγ依赖的途径发生，可能是通过干扰AP-1和NF-κB的激活。
