Abstract
Aging is a major risk factor for the development of osteoarthritis (OA). One hallmark of aging is loss of proteostasis resulting in increased cellular stress and cell death. However, its effect on the development of OA is not clear. Here, using knee articular cartilage tissue from young and old cynomolgus monkeys (Macaca fascicularis), we demonstrate that with aging there is loss of molecular chaperone expression resulting in endoplasmic reticulum (ER) stress and cell death. Chondrocytes from aged articular cartilage showed decreased expression of molecular chaperones, including protein disulfide isomerase, calnexin, and Ero1-like protein alpha, and increased immunohistochemical staining for ER stress markers (phosphorylated IRE1 alpha, spliced X-box binding protein-1, activating transcription factor 4 and C/EBP homologous protein), and apoptotic markers [cleaved caspase 3 and cleaved poly(ADP-ribose) polymerase], suggesting that decreased expression of molecular chaperone during aging induces ER stress and chondrocyte apoptosis in monkey articular cartilage. Apoptosis induced by aging-associated ER stress was further confirmed by TUNEL staining. Aged monkey cartilage also showed increased expression of nuclear protein 1 (Nupr1) and tribbles related protein-3 (TRB3), known regulators of apoptosis and cell survival pathways. Treatment of cultured monkey chondrocytes with a small molecule chemical chaperone, 4-phenylbutyric acid (PBA, a general ER stress inhibitor) or PERK Inhibitor I (an ER stress inhibitor specifically targeting the PERK branch of the unfolded protein response pathway), decreased the expression of ER stress and apoptotic markers and reduced the expression of Nupr1 and TRB3. Consistent with the above finding, knockdown of calnexin expression induces ER stress and apoptotic markers in normal human chondrocytes in vitro. Taken together, our study clearly demonstrates that aging promotes loss of proteostasis and induces ER stress and chondrocyte apoptosis in articular cartilage. Thus, restoring proteostasis using chemical/molecular chaperone or ER stress inhibitor could be a therapeutic option to treat aged-linked OA.
Highlights
Aging is a major risk factor for the development of osteoarthritis (OA)
Our data showed significantly increased expression of endoplasmic reticulum (ER) stress markers, including BIP, P-IRE1α, X-box binding protein-1 (XBP1), activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP), in the articular cartilage of the old monkey compared to young monkeys (Fig. 1), demonstrating that aging induces ER stress in articular cartilage
We investigated whether the aginginduced ER stress triggers caspase-mediated apoptosis in articular cartilage
Summary
Aging is a major risk factor for the development of osteoarthritis (OA). One hallmark of aging is loss of proteostasis resulting in increased cellular stress and cell death. The capacity of chondrocytes to restore cartilage homeostasis declines due to aging-related cellular changes, including cellular senescence [9], oxidative stress [10], increased inflammation [11], autophagy defects [12] and mitochondrial dysfunction [13] Taken together, these changes promote cartilage destruction and development of OA; the mechanisms involved are not clearly understood. Molecular chaperones are broadly categorized into multiple protein families: 1) heat shock proteins, 2) lectins (calreticulin and calnexin), 3) oxidoreductases or protein disulfide isomerases, and 4) co-chaperones that are members of ERp57 family that are associated with calnexin and calreticulin [19] These chaperones play a very specific function in protein folding/proteostasis. They bind to misfolded proteins and target them for degradation [26]
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