Kim Yoon Seong, Kim Sung Soo, Cho Jeong Je, Choi Dong Hee, Hwang Onyou, Shin Dong Hoon, Chun Hong Sung, Beal M Flint, Joh Tong H
Burke Medical Research Institute, Weill Medical College and Graduate School of Medical Sciences of Cornell University, White Plains, New York 10605, USA.
J Neurosci. 2005 Apr 6;25(14):3701-11. doi: 10.1523/JNEUROSCI.4346-04.2005.
Microglial activation and inflammation are associated with progressive neuronal apoptosis in neurodegenerative human brain disorders. We sought to investigate molecular signaling mechanisms that govern activation of microglia in apoptotic neuronal degeneration. We report here that the active form of matrix metalloproteinase-3 (MMP-3) was released into the serum-deprived media (SDM) of PC12 cells and other media of apoptotic neuronal cells within 2-6 h of treatment of the cells, and SDM and catalytic domain of recombinant MMP-3 (cMMP-3) activated microglia in primary microglia cultures as well as BV2 cells, a mouse microglia cell line. Both SDM and cMMP-3 induced generation of tumor necrosis factor alpha (TNF-alpha), interleukin-6 (IL-6), IL-1beta, and interleukin-1 receptor antagonist but not IL-12 and inducible nitric oxide synthase, which are readily induced by lipopolysaccharide, in microglia, suggesting that there is a characteristic pattern of microglial cytokine induction by apoptotic neurons. Neither glial cell line-derived neurotrophic factor nor anti-inflammatory cytokines, such as IL-10 and transforming growth factor-beta1, were induced. SDM and cMMP-3 extensively released TNF-alpha from microglia and activated the nuclear factor-kappaB pathway, and these microglial responses were totally abolished by preincubation with an MMP-3 inhibitor, NNGH [N-isobutyl-N-(4-methoxyphenylsulfonyl)-glycylhydroxamic acid]. MMP-3-mediated microglial activation mostly depended on ERK (extracellular signal-regulated kinase) phosphorylation but not much on either JNK (c-Jun N-terminal protein kinase) or p38 activation. Conditioned medium of SDM- or cMMP-3-activated BV2 cells caused apoptosis of PC12 cells. These results strongly suggest that the distinctive signal of neuronal apoptosis is the release of active form of MMP-3 that activates microglia and subsequently exacerbates neuronal degeneration. Therefore, the release of MMP-3 from apoptotic neurons may play a major role in degenerative human brain disorders, such as Parkinson's disease.
小胶质细胞激活和炎症与神经退行性人脑疾病中神经元的进行性凋亡相关。我们试图研究在凋亡性神经元变性中调控小胶质细胞激活的分子信号机制。我们在此报告，基质金属蛋白酶-3（MMP-3）的活性形式在处理细胞2 - 6小时内释放到PC12细胞的血清饥饿培养基（SDM）以及凋亡性神经元细胞的其他培养基中，并且SDM和重组MMP-3的催化结构域（cMMP-3）在原代小胶质细胞培养物以及BV2细胞（一种小鼠小胶质细胞系）中激活了小胶质细胞。SDM和cMMP-3均诱导小胶质细胞产生肿瘤坏死因子α（TNF-α）、白细胞介素-6（IL-6）、IL-1β和白细胞介素-1受体拮抗剂，但不诱导脂多糖容易诱导的IL-12和诱导型一氧化氮合酶，这表明凋亡神经元诱导小胶质细胞产生细胞因子存在一种特征模式。胶质细胞源性神经营养因子和抗炎细胞因子，如IL-10和转化生长因子-β1均未被诱导。SDM和cMMP-3从小胶质细胞中大量释放TNF-α并激活核因子-κB途径，并且这些小胶质细胞反应通过用MMP-3抑制剂NNGH [N-异丁基-N-(4-甲氧基苯基磺酰基)-甘氨酰羟肟酸]预孵育而完全消除。MMP-3介导的小胶质细胞激活主要依赖于细胞外信号调节激酶（ERK）磷酸化，而对c-Jun N端蛋白激酶（JNK）或p38激活的依赖性不大。SDM或cMMP-3激活的BV2细胞的条件培养基导致PC12细胞凋亡。这些结果强烈表明，神经元凋亡的独特信号是活性形式的MMP-3的释放，其激活小胶质细胞并随后加剧神经元变性。因此，凋亡神经元中MMP-3的释放可能在退行性人脑疾病如帕金森病中起主要作用。
