Sitte N, Merker K, Von Zglinicki T, Grune T, Davies K J
Clinics of Physical Medicine and Rehabilitation, Humboldt University Berlin, Germany.
FASEB J. 2000 Dec;14(15):2495-502. doi: 10.1096/fj.00-0209com.
Oxidized and cross-linked proteins tend to accumulate in aging cells. Declining activity of proteolytic enzymes, particularly the proteasome, has been proposed as a possible explanation for this phenomenon, and direct inhibition of the proteasome by oxidized and cross-linked proteins has been demonstrated in vitro. We have further examined this hypothesis during both proliferative senescence (this paper) and postmitotic senescence (see the accompanying paper, ref 1 ) of human BJ fibroblasts. During proliferative senescence, we found a marked decline in all proteasome activities (trypsin-like activity, chymotrypsin-like activity, and peptidyl-glutamyl-hydrolyzing activity) and in lysosomal cathepsin activity. Despite the loss of proteasome activity, there was no concomitant change in cellular levels of actual proteasome protein (immunoassays) or in the steady-state levels of mRNAs for essential proteasome subunits. The decline in proteasome activities and lysosomal cathepsin activities was accompanied by dramatic increases in the accumulation of oxidized and cross-linked proteins. Furthermore, as proliferation stage increased, cells exhibited a decreasing ability to degrade the oxidatively damaged proteins generated by an acute, experimentally applied oxidative stress. Thus, oxidized and cross-linked proteins accumulated rapidly in cells of higher proliferation stages. Our data are consistent with the hypothesis that proteasome is progressively inhibited by small accumulations of oxidized and cross-linked proteins during proliferative senescence until late proliferation stages, when so much proteasome activity has been lost that oxidized proteins accumulate at ever-increasing rates. Lysosomes attempt to deal with the accumulating oxidized and cross-linked proteins, but declining lysosomal cathepsin activity apparently limits their effectiveness. This hypothesis, which may explain the progressive intracellular accumulation of oxidized and cross-linked proteins in aging, is further explored during postmitotic senescence in the accompanying paper (1).
氧化和交联蛋白往往在衰老细胞中积累。蛋白水解酶活性下降，尤其是蛋白酶体，被认为是这一现象的一种可能解释，并且在体外已证实氧化和交联蛋白可直接抑制蛋白酶体。我们在人BJ成纤维细胞的增殖性衰老（本文）和有丝分裂后衰老（见随附论文，参考文献1）过程中进一步研究了这一假设。在增殖性衰老过程中，我们发现所有蛋白酶体活性（胰蛋白酶样活性、糜蛋白酶样活性和肽基 - 谷氨酰水解活性）以及溶酶体组织蛋白酶活性均显著下降。尽管蛋白酶体活性丧失，但实际蛋白酶体蛋白的细胞水平（免疫测定）或必需蛋白酶体亚基的mRNA稳态水平并未随之改变。蛋白酶体活性和溶酶体组织蛋白酶活性的下降伴随着氧化和交联蛋白积累的显著增加。此外，随着增殖阶段增加，细胞降解由急性实验性施加的氧化应激产生的氧化损伤蛋白的能力下降。因此，氧化和交联蛋白在较高增殖阶段的细胞中迅速积累。我们的数据与以下假设一致：在增殖性衰老过程中，蛋白酶体逐渐被少量积累的氧化和交联蛋白抑制，直至增殖后期，此时大量蛋白酶体活性丧失，氧化蛋白以不断增加的速率积累。溶酶体试图处理积累的氧化和交联蛋白，但溶酶体组织蛋白酶活性下降显然限制了它们的有效性。这一假设可能解释了衰老过程中氧化和交联蛋白在细胞内的逐渐积累，随附论文（1）在有丝分裂后衰老过程中对这一假设进行了进一步探讨。
