- Email: [email protected]
Autophagy in osteoarthritis
G Model
ARTICLE IN PRESS
BONSOI-4208; No. of Pages 6
Joint Bone Spine xxx (2015) xxx–xxx
Available online at
ScienceDirect www.sciencedirect.com
Review
Autophagy in osteoarthritis Yu-Sheng Li a , Fang-Jie Zhang b , Chao Zeng a , Wei Luo a , Wen-Feng Xiao a , Shu-Guang Gao a , Guang-Hua Lei a,∗ a b
Department of Orthopaedics, Xiangya Hospital, Central South University, No. 87 Xiangya Road, Changsha, Hunan 410008, PR China Department of Emergency Medicine, Xiangya Hospital, Central South University, No. 87 Xiangya Road, Changsha, Hunan 410008, PR China
a r t i c l e
i n f o
Article history: Accepted 21 June 2015 Available online xxx Keywords: Autophagy Chondrocyte Osteoarthritis Rapamycin
a b s t r a c t Degradation of the articular cartilage is at the centre of the pathogenesis of osteoarthritis (OA), for which age is the major risk factor. Maintaining the chondrocytes in a healthy condition appears to be an important factor for preservation of the entire cartilage and preventing its degeneration. Autophagy, which is an essential cellular homeostatic mechanism for the removal of dysfunctional cellular organelles and macromolecules, is increased by catabolic and nutritional stresses. Autophagy is increased in OA chondrocytes and cartilage, particularly during the initial degenerative phase, to regulate changes in OA-like gene expression through modulation of apoptosis and reactive oxygen species (ROS). In this way, autophagy acts as an adaptive response to protect chondrocytes from various environmental changes, while with gradual cartilage degradation, decreased autophagy is linked with cell death. Rapamycin, which is a specific inhibitor of the mTOR signaling pathway, enhances expression of autophagy regulators and prevents chondrocyte death. In the future, pharmacological activation of autophagy may be an effective therapeutic approach for OA. © 2015 Société franc¸aise de rhumatologie. Published by Elsevier Masson SAS. All rights reserved.
1. Introduction Osteoarthritis (OA), the most common age-related joint pathology, is a degenerative disease affecting all the structures of the joints and is characterized by articular cartilage destruction along with changes in other joint components, including bone, meniscus, synovium, ligament, capsule, and muscle [1]. Multiple factors have been implicated in the pathogenesis of OA, including mechanical, genetic, and age-associated factors, with age being the major risk factor for OA [2]. Chondrocytes are capable of responding to structural changes in the surrounding cartilage matrix although the capacity of adult articular chondrocytes to regenerate normal cartilage matrix architecture is limited and declines with age, due to cell death and abnormal responsiveness to anabolic stimuli [3]. Therefore, maintaining the chondrocytes in a healthy condition appears to be an important factor for preservation of the entire cartilage and preventing its degeneration. Autophagy (from auto-phagos: self-eating) is an essential cellular homeostatic mechanism for the removal of dysfunctional cellular organelles and macromolecules [4]. The autophagy pathway is integrated with multiple signal transduction pathways that respond to nutrient supply, energy balance, cytokines, and growth factors [4]. Here, we
∗ Corresponding author. E-mail address: [email protected] (G.-H. Lei).
summarize the current understanding of the molecular mechanism of autophagy, focusing on recent studies that have examined the role of autophagy in the pathogenesis of OA. 2. Definition of autophagy All living organisms undergo continuous renovation. Cells constantly adapt to changing environmental conditions by adjusting their content to prevailing needs. This process, known as homeostasis, involves continuous biosynthesis and turnover of cellular components. Eukaryotic cells have two major degradation systems: the lysosome and the proteasome [5]. Proteasomal degradation has high selectivity; the proteasome generally recognizes only ubiquitinated substrates, which are primarily short-lived proteins. In contrast, degradation in the lysosome does not follow such a simple pattern. Extracellular material and plasma membrane proteins can be delivered to lysosomes for degradation via the endocytic pathway. Furthermore, cytosolic components and organelles can also be delivered to the lysosome by autophagy [5]. Autophagy is a mechanism of cellular homeostasis that is strongly induced in a wide range of eukaryotic organisms and in multiple different cell types under conditions of nutrient starvation. This leads to bulk degradation of cytoplasmic components (proteins, organelles), the building blocks of which are used as an energy supply and the synthesis of components essential for survival [6,7]. In cells defective in autophagy, the total intracellular pool of amino
http://dx.doi.org/10.1016/j.jbspin.2015.06.009 1297-319X/© 2015 Société franc¸aise de rhumatologie. Published by Elsevier Masson SAS. All rights reserved.
Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
2
acids is greatly reduced, leading to the inability to synthesize proteins that are essential for cellular survival [8]. Autophagy may also serve other functions and is important for cellular housekeeping by removing exhausted, redundant or unnecessary components. Therefore, autophagy is implicated in the retardation of aging and in the prevention and therapy of age-associated diseases [9], as well as in support of cell remodeling during development [10] or contributing to cellular defence against pathogens [11]. Autophagy is a generic term for all pathways by which cytoplasmic materials are delivered to the lysosome in animal cells or the vacuole in plant and yeast cells [5]. The currently known autophagy processes are subdivided into three types depending on the route of cargo delivery: chaperone-mediated autophagy, macroautophagy and microautophagy [12]. Of these, chaperonemediated autophagy is, as yet, described only in mammals and is involved in the degradation of individual, soluble proteins. In contrast, macroautophagy and microautophagy occur in a wide range of eukaryotes, including mammals, plants and fungi, leading to the degradation of portions of the cytoplasm, which may include cellular organelles [12]. Macroautophagy is thought to be the major type of autophagy and is mediated by double (or multiple) membranebound vacuoles, the formation of which is highly conserved from yeast to humans [13]. Macroautophagy is the most extensively studied form of autophagy and is, therefore, referred to simply as “autophagy” hereafter. Fig. 1. The regulation factors in autophagy progression. : interaction effect. fx2: negative effect;
: positive effect;
3. The basic process of autophagy The main purpose of autophagy in yeast is to degrade the cytoplasm and recycle the resulting macromolecules for use in the synthesis of essential components during nutrient stress [14]. Although autophagy and autophagy-related processes are dynamic, they can be broken down into several discrete steps [14,15]: • induction: in mammalian cells, autophagy occurs at a basal level under vegetative growth conditions. Accordingly, there must be a mechanism for sensing the extracellular milieu and transducing an appropriate signal to regulatory elements that allow the induction of autophagy. One of the major regulatory components is the protein kinase TOR (target of rapamycin), which inhibits autophagy under basal or nutrient-rich conditions [16]; • cargo selection and packaging: autophagy is generally considered to be a non-specific process in which bulk cytoplasm is randomly sequestered into the cytosolic autophagosome; • nucleation of vesicle formation: this is the initial step that recruits proteins and lipids for autophagosome construction; • vesicle expansion and completion: autophagosome formation is a de novo process, in which membranes emerging at the preautophagosomal structure (PAS) expand, either by direct flow from a source, such as the endoplasmic reticulum (ER) or by vesicular addition, and then seal to enclose the cytosolic cargo; • retrieval: the process by which certain components are retrieved for reuse; • targeting: docking and fusion of the completed vesicle with the lysosome/vacuole; • breakdown of the intraluminal vesicle (either the Cvt or autophagic bodies) and its cargo and recycling of the macromolecular constituents. 4. Regulatory factors Several protein kinases and signaling pathways are involved in autophagy and autophagy-related processes. A number of critical autophagy-related genes (ATG or Atg) have been identified, the gene products of which regulate distinct steps in the induction Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
or progression of autophagy [17]. The Ser/Thr protein kinase TOR, which plays a key role in signaling of nutrient limitation [18], resides in a multiprotein complex 1, mTORC1 [19]. In response to stimulation by nutrients or growth factors, mTORC1 negatively regulates the mammalian uncoordinated-51-like protein kinase (ULK1) complex that includes ULK1, ATG13, ATG101, and FIP200 (RB1CC1), which results in autophagy suppression [20,21]. Under glucose starvation, AMP-activated protein kinase (AMPK) promotes autophagy by activating ULK1 directly. Under nutrient sufficiency, high mTOR activity prevents ULK1 activation and disrupts the interaction between ULK1 and AMPK [22]. In addition, autophagy is negatively regulated by the phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathway [23], which contains three highly conserved proteins, namely the protein kinase Vps15, phosphatidylinositol-3-kinase Vps34 [24] and Atg6 [25]. Autophagy is also regulated by the Beclin-1 complex, consisting of Beclin-1, class III phosphatidylinositol-3kinase (VPS34 or PI3KC3) and ATG14L or UVRAG. This is subject to negative regulation by the PI3K/Akt pathway [23] as well as by binding interactions with the anti-apoptotic Bcl-2 family proteins [26]. Stimulation of the Beclin-1 complex generates phosphatidylinositol-3-phosphate (PI3P) [27], which triggers autophagosomal nucleation. Following phagophore formation, elongation of the autophagosome membrane requires the action of two ubiquitin-like conjugation systems, the Atg5–Atg12 conjugation system and the microtubule-associated protein-1 light chain 3 (LC3, Atg8) conjugation system [28,29]. Atg4B converts the proform of LC3B to its cytosolic free form (LC3-I). In mammals, the conversion of LC3-I (and other Atg8 homologues) to its phosphatidylethanolamine-conjugated and autophagosome membrane-associated form (i.e., LC3-II) is an initiating step in autophagy [30,31]. Fig. 1 depicts a schematic flow chart describing the regulation factors in autophagy progression. 5. Expression of autophagy in OA The articular cartilage is an avascular, aneural, alymphatic and viscoelastic connective tissue that derives its nutrition and oxygen Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
supply by diffusion from the synovial fluid and the subchondral bone [32]. Chondrocytes are responsible for both synthesis and turnover of the abundant extracellular matrix (ECM) [3]; therefore, maintaining the chondrocytes in a healthy condition appears to be an important factor for maintaining the integrity of the entire cartilage. Autophagy has been suggested to be an important cell survival mechanism under various forms of stress [4]. Autophagy serves not only to regulate the final stages of the chondrocyte lifecycle, but also the rate at which chondrocytes enter the maturation process [33]. Autophagy in normal adult articular cartilage is an important mechanism for cellular homeostasis [34]; thus, cells in the superficial zone express high levels of the autophagy proteins BECN1, ATG5, and MAP1LC3 [34]. Catabolic and nutritional stresses increase autophagy in OA, and during the initial degenerative phase at least, autophagy is increased in OA chondrocytes and cartilage, with increased expression of LC3 and Beclin-1 messenger RNA (mRNA) in OA chondrocytes [35]. Furthermore, autophagy regulates changes in OA-like gene expression through the modulation of apoptosis and reactive oxygen species (ROS) [35], which promote oxidative stress resulting in the assembly of multiprotein inflammatory complexes, called the inflammasome [36]. Disruption of the interplay between autophagy and inflammasome contributes to the age-related increase in the prevalence of OA and decreased efficacy of articular cartilage repair [37]. Enhanced autophagy, suppressed mTOR and reduced MAP4K3 activity are considered to constitute an early response in the biomechanicallyinduced degeneration of the temporomandibular joint cartilage, especially in premature chondrocytes [38]. Transiently increased autophagy represents a compensatory response to cellular stress, with damage occurring when prolonged stress exceeds the capacity of this mechanism [39] and failure to mount an autophagic response may lead to further degeneration [33]. In cartilage with mild OA, ULK1, Beclin-1, and LC3 protein expression are reduced in the cartilage superficial zone, although these three proteins are strongly expressed in the OA cell clusters as well as the middle and deep zones [40]. In mouse OA knee joints, expression of ULK1, Beclin-1, and LC3 is decreased together with glycosaminoglycan (GAG) loss, while PARP p85 expression is increased with the gradual cartilage degradation [40]. Furthermore, the reduction in these key regulators of autophagy is accompanied by increased apoptosis. Aging of mouse knee joints is associated with reduced autophagy and cellularity along with increased apoptotic cell death [41]. mTOR is overexpressed in human OA cartilage as well as mouse and dog experimental OA. Upregulated mTOR expression in OA correlates with increased chondrocyte apoptosis and reduced expression of key autophagy genes, including ATG3, ATG5, ATG12, ULK1, LC3B, Beclin-1 and BNIP3 [42]. In the cartilage-specific ablation of mTOR knockout mice, autophagy signaling is increased with a significant reduction in articular cartilage degradation, apoptosis and synovial fibrosis [42]. Stress conditions in the epiphyseal growth plate can promote the autophagic response through the modulation of genes controlling metabolite utilization to achieve survival of the terminally differentiated cells exposed to the brief rigors of the harsh local microenvironment [43]. In a mouse model consisting of the green fluorescent protein (GFP) fused to light chain 3 (LC3) on the C57BL/6J genetic background (GFP-LC3 transgenic mice) generated to activate autophagy in normal cartilage and monitor autophagy changes in age-related cartilage degradation, the number and area of autophagosomes in chondrocytes of older mice were significantly decreased compared to those in younger mice. Furthermore, the average number and size of vesicles in osteocytes in subchondral bone decreased in old mice [41]. In GFP-LC3 transgenic mice, the highest levels of GFP-LC3 and the key autophagy proteins (Atg5 and LC3) were observed in chondrocytes in the superficial and upper mid-zones of articular cartilage in young mice, while the levels were much lower in deep zone cells and also in old mice [41]. Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
3
These observations in cartilage are consistent with the notion that basal autophagic activity decreases with age [41,44] and that the marked inhibition of autophagy would have a negative impact on chondrocyte survival and differentiation [33], thus contributing to the accumulation of damaged macromolecules and susceptibility to age-related OA [44]. 6. Hypoxia, mechanical injury and autophagy Articular cartilage is maintained in a low oxygen environment throughout life [32]. It has been demonstrated that an oxygen gradient exists in cartilage from around 6% at the joint surface to 1% in the deep layers [45]. Chondrocytes are therefore adapted to these hypoxic conditions. Hypoxia-inducible factors (HIFs) are transcription factors that respond to decreases in the level of available oxygen in the cellular environment. Two main HIF isoforms (HIF1␣ and HIF-2␣) mediate the response of all cells to hypoxia [46,47]. HIF-1␣ may protect articular cartilage by promoting the chondrocyte phenotype, maintaining chondrocyte viability, and supporting metabolic adaptation to a hypoxic environment. In contrast, HIF2␣ is a catabolic factor in the pathogenesis of OA [48]. HIF-1␣, which is expressed in normal and OA human articular chondrocytes cultured under normoxic conditions [49] can be further induced or stabilized by hypoxia. The relatively high constitutive expression of HIF-1␣ in chondrocytes may be an important adaptation to survival in the avascular-hypoxic environment of cartilage [49]. HIF-2␣ expression is also higher in OA cartilages versus normal cartilage in mice and humans [50]. HIF-1␣ and HIF-2␣ play critical roles in the survival, development, and differentiation of chondrocytes in vascular hypoxia [51]. Under hypoxic conditions, both HIF-1␣ and HIF-2␣ are more strongly associated with the nuclei in the middle/deep cells, compared with the full-thickness and superficial zone chondrocytes [52]. A relationship between HIF-1 ␣ and increased accumulation of autophagic proteins, such as Beclin-1, has been reported [53], while HIF-2␣ has been shown to be a suppressor of autophagy in vitro [54]. In chondrocytes, AMPK (the enzyme that monitors the cellular energy status) is activated in a HIF-1-dependent manner. This enzyme inhibits the effects of mTOR on development of the autophagic state [55]. Studies have shown that 1% O2 significantly increased phospho-AMPK (pAMPK) protein expression compared with the effects of 5% or 20% O2 [56]. Suppression of AMPK activates mTOR and survival signaling mediated by the Akt pathway. The relationship between HIF-1, AMPK, mTOR, and autophagy involves HIF-1 activation in response to a reduction in the cellular energy charge during tissue hypoxia, which then results in the phosphorylation and activation of AMPK. Once activated, AMPK suppresses mTOR, thereby activating autophagic flux, which is also activated directly by HIF-1 [55]. In growth plate chondrocytes, increased HIF-1-mediated glycolytic activity elevates AMP levels. The resulting AMP kinase activation and suppression of mTOR causes increased autophagy. Once autophagy is activated, the post-mitotic chondrocytes remain viable in their unique microenvironment and complete their lifecycle [57]. Moreover, HIF-2 is robustly expressed in the pre-hypertrophic cells of the mouse growth cartilage and suppresses chondrocyte autophagy, while elevated autophagic responses are observed throughout the growth plate in HIF-2␣-knockout mice [54]. Silenced HIF-2␣ in chondrocytes leads to decreased Akt-1 and mTOR activities, reduced Bcl-XL expression and a robust autophagic response [54]. Mechanical load is the other main risk factor for OA besides aging. Mechanical injury also leads to suppression of autophagy [58] and has been shown to induce cell death and loss of sGAG, in association with significantly decreased ULK1, Beclin-1 and LC3 expression in the cartilage superficial zone at 48 hours post-injury. Rapamycin enhances the expression of autophagy regulators and prevents cell death and sGAG loss in mechanically injured explants [58] Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
4
Surgically induced OA in mice lacking the von Hippel-Lindau gene (Vhl) in osteochonadral progenitor cells (referred to as Vhl cKO mice) resulted in HIF-2␣ upregulation, increased chondrocyte apoptosis as well as decreased autophagy with cartilage destruction and proteoglycan loss during OA development [59]. In surgically induced OA in PPAR␥ KO mice, increased cartilage degradation and chondrocyte apoptosis was observed, as well as the overproduction of OA inflammatory/catabolic factors associated with the increased expression of mTOR and the suppression of key autophagy markers [60]. 7. Inflammation and autophagy The inflammation of the synovial membrane that occurs in both the early and late phases of OA is associated with alterations in the adjacent cartilage [61]. Increased expression of proinflammatory cytokines in cartilage, synovial membrane and subchondral bone are believed to be linked to the development and progression of structural changes in the OA joint [62]. Inflammatory signals activate inflammasome-dependent responses and caspases, predominantly caspase-1, which cleave the inactive precursors of IL-1 and IL-18 and stimulate their secretion and activity. Consequently, these cytokines provoke inflammatory responses and accelerate the aging process by inhibiting autophagy [63]. Increased expression of mTOR in peripheral blood mononuclear cells of OA patients is related to the disease activity associated with synovitis [64]. IL-1 treatment of human OA chondrocytes results in a significant increase in mTOR expression [42]. Blocking autophagy potentiated inflammasome activity, whereas stimulating autophagy had the opposite effect. Assembled inflammasomes undergo ubiquitination and recruit the autophagic adaptor p62, which assists their delivery to autophagosomes. Thus, autophagy accompanies inflammasome activation to moderate inflammation by eliminating active inflammasomes [65]. PPAR␥ deficiency in chondrocytes results in the aberrant expression of inflammatory markers (iNOS and COX-2) associated with the dysregulated expression of mTOR and inhibition of critical autophagy markers, ultimately resulting in increased inflammatory activity within the articular cartilage leading to severe/accelerated OA [60]. Therefore, mTOR is an important link between synovitis and structural damage in inflammatory arthritis. Inhibition of mTOR reduces synovial osteoclast formation and protects against local bone erosion and cartilage loss. Clinical signs of arthritis are improved after mTOR inhibition, and histologic evaluation showed a decrease in synovitis [66]. Rapamycin treatment of C57Bl/6 OA mice also maintains cartilage cellularity, and decreases IL-1 expression in articular cartilage [67]. 8. Pharmacological targets of autophagy Augmentation of autophagy via inhibition of the TOR signaling pathway by genetic or pharmacological intervention extends lifespan, indicating that the further development of interventions targeting mTOR represents a strategy for the treatment and prevention of age-related diseases [68]. Moreover, since autophagy is a critical protective mechanism against mitochondrial dysfunction, pharmacologic interventions that enhance autophagy may have chondroprotective activity in cartilage degenerative processes in OA [69]. In experimental OA mice, rapamycin treatment maintains cartilage cellularity and decreases ADAMTS-5 and interleukin-1 expression in articular cartilage. Rapamycin also reduces the severity of synovitis and the synovial tissue level of IL-1 in the knee joints in murine experimental OA by inhibiting ribosomal protein S6 phosphorylation [67]. Clinical signs of arthritis are improved and histological signs decreased in synovitis after mTOR inhibition by rapamycin or everolimus, and in vitro, mTOR inhibition Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
downregulates the expression of digestive enzymes and leads to osteoclast apoptosis [66]; thus, when autophagy is activated, the severity of experimental OA is improved [67]. Recently, a study indicated that glucosamine, which is effective in animal models of OA and has anti-inflammatory and anabolic effects on cartilage cells, modulates molecular targets of the autophagy pathway both in vitro and in vivo. Furthermore, the enhancement of autophagy by increased LC3-II levels, formation of LC3 puncta, and increased LC3 turnover is mainly dependent on the Akt/FoxO/mTOR pathway [70]. Glucosamine protects nucleus pulposus (NP) cells and upregulates autophagy in a dose-dependent manner within 24 hours by inhibiting the mTOR pathway by reducing mTOR phosphorylation [71]. Similarly, glucosamine protects PPAR␥-mTOR double KO mice against DMM-induced OA. This effect was associated with significant protection from cartilage destruction, proteoglycan loss and loss of chondrocytes in both the medial tibial plateau and the medial femoral condyle, with a significant reduction in chondrocyte cell death and the percentage of MMP13-positive cells, as well as enhanced LC3B II expression in double KO OA chondrocytes [60]. Local intra-articular injection of rapamycin in a murine model of OA indicated that rapamycin significantly reduced the severity of articular cartilage degradation, with a reduction in mTOR expression and activation of LC3 in the chondrocytes. Furthermore, rapamycin treatment also reduced VEGF, COL10A1, and MMP13 expression after DMM surgery [72]. In a murine OA model, after intra-articular administration of gelatin hydrogels incorporating rapamycin micelles, OA mediator genes (IL-1 and IL-6, MMP9, MMP13, C/EBP, and mTOR) were downregulated while Col2a1 was upregulated in the rapamycin-treated mice [73]. Thus, intraarticular administration of rapamycin could be a novel therapeutic approach for OA [72,73]. In conclusion, age is the major risk factor for OA. The capacity of adult articular chondrocytes to regenerate the normal cartilage matrix architecture is limited and declines with age. During the development of OA, autophagy may increase as an adaptive response to protect chondrocytes from various environmental changes, while failure of the adaptation may lead to progression in cartilage degradation. Thus, pharmacological activation of autophagy may be an appropriate therapeutic approach for OA. Disclosure of interest The authors declare that they have no conflicts of interest concerning this article. Acknowledgments This work was supported by the National Natural Science Foundation of China (No.81272034, 81402224 and 81501923), the Provincial Science Foundation of Hunan (No.2015JJ3139), the Scientific Research Project of Science and Technology Bureau of Hunan Province (2012FJ6001), the Scientific Research Project of Science and Technology Office of Changsha City (K1203040-31), the Scientific Research Project of Health and Family Planning Commission of Hunan Province (B2014-12), the Hunan Provincial Innovation Foundation for Postgraduates (CX2012B086), the Fundamental Research Funds for the Central Universities of Central South University (2013zzts081, 2013zzts319) and the College Student’s Innovation and Entrepreneurship Project of Central South University (DL14505). References [1] Hunter DJ, Felson DT. Osteoarthritis. BMJ 2006;332:639–42. [2] Shane AA, Loeser RF. Why is osteoarthritis an age-related disease? Best Pract Res Clin Rheumatol 2010;24:15–26.
Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
[3] Loeser RF. Aging and osteoarthritis: the role of chondrocyte senescence and aging changes in the cartilage matrix. Osteoarthritis Cartilage 2009;17: 971–9. [4] Mizushima N. Physiological functions of autophagy. Curr Top Microbiol Immunol 2009;335:71–84. [5] Mizushima N, Komatsu M. Autophagy: renovation of cells and tissues. Cell 2011;147:728–41. [6] Tsukada M, Ohsumi Y. Isolation and characterization of autophagy-defective mutants of Saccharomyces cerevisiae. Febs Lett 1993;333:169–74. [7] Levine B, Kroemer G. Autophagy in the pathogenesis of disease. Cell 2008;132:27–42. [8] Onodera J, Ohsumi Y. Autophagy is required for maintenance of amino acid levels and protein synthesis under nitrogen starvation. J Biol Chem 2005;280:31582–6. [9] Bergamini E. Autophagy: a cell repair mechanism that retards ageing and ageassociated diseases and can be intensified pharmacologically. Mol Aspects Med 2006;27:403–10. [10] Levine B, Klionsky DJ. Development by self-digestion: molecular mechanisms and biological functions of autophagy. Dev Cell 2004;6:463–77. [11] Mizushima N, Levine B, Cuervo AM, et al. Autophagy fights disease through cellular self-digestion. Nature 2008;451:1069–75. [12] Todde V, Veenhuis M, van der Klei IJ. Autophagy: principles and significance in health and disease. Biochim Biophys Acta 2009;1792:3–13. [13] Klionsky DJ, Abeliovich H, Agostinis P, et al. Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes. Autophagy 2008;4:151–75. [14] Klionsky DJ. The molecular machinery of autophagy: unanswered questions. J Cell Sci 2005;118:7–18. [15] Chen Y, Klionsky DJ. The regulation of autophagy – unanswered questions. J Cell Sci 2011;124:161–70. [16] Carrera AC. TOR signaling in mammals. J Cell Sci 2004;117:4615–6. [17] Klionsky DJ. Autophagy. Curr Biol 2005;15:R282–3. [18] Pattingre S, Espert L, Biard-Piechaczyk M, et al. Regulation of macroautophagy by mTOR and Beclin 1 complexes. Biochimie 2008;90:313–23. [19] Jung CH, Ro SH, Cao J, et al. mTOR regulation of autophagy. Febs Lett 2010;584:1287–95. [20] Mizushima N. The role of the Atg1/ULK1 complex in autophagy regulation. Curr Opin Cell Biol 2010;22:132–9. [21] Ganley IG, Lam DH, Wang J, et al. ULK1.ATG13.FIP200 complex mediates mTOR signaling and is essential for autophagy. J Biol Chem 2009;284:12297–305. [22] Kim J, Kundu M, Viollet B, et al. AMPK and mTOR regulate autophagy through direct phosphorylation of Ulk1. Nat Cell Biol 2011;13:132–41. [23] Wang RC, Wei Y, An Z, et al. Akt-mediated regulation of autophagy and tumorigenesis through Beclin 1 phosphorylation. Science 2012;338:956–9. [24] Kihara A, Noda T, Ishihara N, et al. Two distinct Vps34 phosphatidylinositol 3-kinase complexes function in autophagy and carboxypeptidase Y sorting in Saccharomyces cerevisiae. J Cell Biol 2001;152:519–30. [25] Kametaka S, Okano T, Ohsumi M, et al. Apg14p and Apg6/Vps30p form a protein complex essential for autophagy in the yeast, Saccharomyces cerevisiae. J Biol Chem 1998;273:22284–91. [26] Pattingre S, Tassa A, Qu X, et al. Bcl-2 antiapoptotic proteins inhibit Beclin 1dependent autophagy. Cell 2005;122:927–39. [27] He C, Levine B. The Beclin 1 interactome. Curr Opin Cell Biol 2010;22:140–9. [28] Tanida I, Ueno T, Kominami E. LC3 conjugation system in mammalian autophagy. Int J Biochem Cell Biol 2004;36:2503–18. [29] Ohsumi Y. Molecular dissection of autophagy: two ubiquitin-like systems. Nat Rev Mol Cell Biol 2001;2:211–6. [30] He H, Dang Y, Dai F, et al. Post-translational modifications of three members of the human MAP1LC3 family and detection of a novel type of modification for MAP1LC3B. J Biol Chem 2003;278:29278–87. [31] Satoo K, Noda NN, Kumeta H, et al. The structure of Atg4B-LC3 complex reveals the mechanism of LC3 processing and delipidation during autophagy. Embo J 2009;28:1341–50. [32] Milner PI, Fairfax TP, Browning JA, et al. The effect of O2 tension on pH homeostasis in equine articular chondrocytes. Arthritis Rheum 2006;54:3523–32. [33] Shapiro IM, Layfield R, Lotz M, et al. Boning up on autophagy: the role of autophagy in skeletal biology. Autophagy 2014;10:7–19. [34] Almonte-Becerril M, Navarro-Garcia F, Gonzalez-Robles A, et al. Cell death of chondrocytes is a combination between apoptosis and autophagy during the pathogenesis of osteoarthritis within an experimental model. Apoptosis 2010;15:631–8. [35] Sasaki H, Takayama K, Matsushita T, et al. Autophagy modulates osteoarthritis-related gene expression in human chondrocytes. Arthritis Rheum 2012;64:1920–8. [36] Salminen A, Ojala J, Kaarniranta K, et al. Mitochondrial dysfunction and oxidative stress activate inflammasomes: impact on the aging process and age-related diseases. Cell Mol Life Sci 2012;69:2999–3013. [37] Mobasheri A, Matta C, Zakany R, et al. Chondrosenescence: definition, hallmarks and potential role in the pathogenesis of osteoarthritis. Maturitas 2015;80:237–44. [38] Zhang M, Zhang J, Lu L, et al. Enhancement of chondrocyte autophagy is an early response in the degenerative cartilage of the temporomandibular joint to biomechanical dental stimulation. Apoptosis 2013;18: 423–34. [39] Barranco C, Osteoarthritis:. Activate autophagy to prevent cartilage degeneration? Nat Rev Rheumatol 2015;11:127.
Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
5
[40] Carames B, Taniguchi N, Otsuki S, et al. Autophagy is a protective mechanism in normal cartilage, and its aging-related loss is linked with cell death and osteoarthritis. Arthritis Rheum 2010;62:791–801. [41] Carames B, Olmer M, Kiosses WB, et al. The relationship of autophagy defects and cartilage damage during joint aging in a mouse model. Arthritis Rheumatol 2015;67:1568–76. [42] Zhang Y, Vasheghani F, Li YH, et al. Cartilage-specific deletion of mTOR upregulates autophagy and protects mice from osteoarthritis. Ann Rheum Dis 2015;74:1432–40. [43] Srinivas V, Shapiro IM. Chondrocytes embedded in the epiphyseal growth plates of long bones undergo autophagy prior to the induction of osteogenesis. Autophagy 2006;2:215–6. [44] Cuervo AM, Dice JF. Age-related decline in chaperone-mediated autophagy. J Biol Chem 2000;275:31505–13. [45] Silver IA. Measurement of pH and ionic composition of pericellular sites. Philos Trans R Soc Lond B Biol Sci 1975;271:261–72. [46] Semenza GL. Hypoxia-inducible factors in physiology and medicine. Cell 2012;148:399–408. [47] Smith TG, Robbins PA, Ratcliffe PJ. The human side of hypoxia-inducible factor. Br J Haematol 2008;141:325–34. [48] Zhang FJ, Luo W, Lei GH. Role of HIF-1alpha and HIF-2alpha in osteoarthritis. Joint Bone Spine 2015;82:144–7. [49] Coimbra IB, Jimenez SA, Hawkins DF, et al. Hypoxia inducible factor-1 alpha expression in human normal and osteoarthritic chondrocytes. Osteoarthritis Cartilage 2004;12:336–45. [50] Saito T, Fukai A, Mabuchi A, et al. Transcriptional regulation of endochondral ossification by HIF-2alpha during skeletal growth and osteoarthritis development. Nat Med 2010;16:678–86. [51] Aro E, Khatri R, Gerard-O’Riley R, et al. Hypoxia-inducible factor-1 (HIF-1) but not HIF-2 is essential for hypoxic induction of collagen prolyl 4-hydroxylases in primary newborn mouse epiphyseal growth plate chondrocytes. J Biol Chem 2012;287:37134–44. [52] Schrobback K, Malda J, Crawford RW, et al. Effects of oxygen on zonal marker expression in human articular chondrocytes. Tissue Eng Part A 2012;18:920–33. [53] Bohensky J, Shapiro IM, Leshinsky S, et al. HIF-1 regulation of chondrocyte apoptosis: induction of the autophagic pathway. Autophagy 2007;3: 207–14. [54] Bohensky J, Terkhorn SP, Freeman TA, et al. Regulation of autophagy in human and murine cartilage: hypoxia-inducible factor 2 suppresses chondrocyte autophagy. Arthritis Rheum 2009;60:1406–15. [55] Bohensky J, Leshinsky S, Srinivas V, et al. Chondrocyte autophagy is stimulated by HIF-1 dependent AMPK activation and mTOR suppression. Pediatr Nephrol 2010;25:633–42. [56] Fermor B, Gurumurthy A, Diekman BO. Hypoxia, RONS and energy metabolism in articular cartilage. Osteoarthritis Cartilage 2010;18:1167–73. [57] Srinivas V, Bohensky J, Shapiro IM. Autophagy: a new phase in the maturation of growth plate chondrocytes is regulated by HIF, mTOR and AMP kinase. Cells Tissues Organs 2009;189:88–92. [58] Carames B, Taniguchi N, Seino D, et al. Mechanical injury suppresses autophagy regulators and pharmacologic activation of autophagy results in chondroprotection. Arthritis Rheum 2012;64:1182–92. [59] Weng T, Xie Y, Yi L, et al. Loss of Vhl in cartilage accelerated the progression of age-associated and surgically induced murine osteoarthritis. Osteoarthritis Cartilage 2014;22:1197–205. [60] Vasheghani F, Zhang Y, Li YH, et al. PPARgamma deficiency results in severe, accelerated osteoarthritis associated with aberrant mTOR signalling in the articular cartilage. Ann Rheum Dis 2015;74:569–78. [61] Sellam J, Berenbaum F. The role of synovitis in pathophysiology and clinical symptoms of osteoarthritis. Nat Rev Rheumatol 2010;6:625–35. [62] Kapoor M, Martel-Pelletier J, Lajeunesse D, et al. Role of proinflammatory cytokines in the pathophysiology of osteoarthritis. Nat Rev Rheumatol 2011;7:33–42. [63] Salminen A, Kaarniranta K, Kauppinen A. Inflammaging: disturbed interplay between autophagy and inflammasomes. Aging 2012;4:166–75. [64] Tchetina EV, Poole AR, Zaitseva EM, et al. Differences in mammalian target of rapamycin gene expression in the peripheral blood and articular cartilages of osteoarthritic patients and disease activity. Arthritis 2013;2013: 461486. [65] Shi CS, Shenderov K, Huang NN, et al. Activation of autophagy by inflammatory signals limits IL-1beta production by targeting ubiquitinated inflammasomes for destruction. Nat Immunol 2012;13:255–63. [66] Cejka D, Hayer S, Niederreiter B, et al. Mammalian target of rapamycin signaling is crucial for joint destruction in experimental arthritis and is activated in osteoclasts from patients with rheumatoid arthritis. Arthritis Rheum 2010;62:2294–302. [67] Carames B, Hasegawa A, Taniguchi N, et al. Autophagy activation by rapamycin reduces severity of experimental osteoarthritis. Ann Rheum Dis 2012;71:575–81. [68] Harrison DE, Strong R, Sharp ZD, et al. Rapamycin fed late in life extends lifespan in genetically heterogeneous mice. Nature 2009;460:392–5. [69] Lopez DFP, Lotz MK, Blanco FJ, et al. Autophagy activation and protection from mitochondrial dysfunction in human chondrocytes. Arthritis Rheumatol 2015;67:966–76. [70] Carames B, Kiosses WB, Akasaki Y, et al. Glucosamine activates autophagy in vitro and in vivo. Arthritis Rheum 2013;65:1843–52.
Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
[71] Jiang L, Jin Y, Wang H, et al. Glucosamine protects nucleus pulposus cells and induces autophagy via the mTOR-dependent pathway. J Orthop Res 2014;32:1532–42. [72] Takayama K, Kawakami Y, Kobayashi M, et al. Local intra-articular injection of rapamycin delays articular cartilage degeneration in a murine model of osteoarthritis. Arthritis Res Ther 2014;16:482.
Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
[73] Matsuzaki T, Matsushita T, Tabata Y, et al. Intra-articular administration of gelatin hydrogels incorporating rapamycin-micelles reduces the development of experimental osteoarthritis in a murine model. Biomaterials 2014;35:9904–11.
Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
ARTICLE IN PRESS
BONSOI-4208; No. of Pages 6
Joint Bone Spine xxx (2015) xxx–xxx
Available online at
ScienceDirect www.sciencedirect.com
Review
Autophagy in osteoarthritis Yu-Sheng Li a , Fang-Jie Zhang b , Chao Zeng a , Wei Luo a , Wen-Feng Xiao a , Shu-Guang Gao a , Guang-Hua Lei a,∗ a b
Department of Orthopaedics, Xiangya Hospital, Central South University, No. 87 Xiangya Road, Changsha, Hunan 410008, PR China Department of Emergency Medicine, Xiangya Hospital, Central South University, No. 87 Xiangya Road, Changsha, Hunan 410008, PR China
a r t i c l e
i n f o
Article history: Accepted 21 June 2015 Available online xxx Keywords: Autophagy Chondrocyte Osteoarthritis Rapamycin
a b s t r a c t Degradation of the articular cartilage is at the centre of the pathogenesis of osteoarthritis (OA), for which age is the major risk factor. Maintaining the chondrocytes in a healthy condition appears to be an important factor for preservation of the entire cartilage and preventing its degeneration. Autophagy, which is an essential cellular homeostatic mechanism for the removal of dysfunctional cellular organelles and macromolecules, is increased by catabolic and nutritional stresses. Autophagy is increased in OA chondrocytes and cartilage, particularly during the initial degenerative phase, to regulate changes in OA-like gene expression through modulation of apoptosis and reactive oxygen species (ROS). In this way, autophagy acts as an adaptive response to protect chondrocytes from various environmental changes, while with gradual cartilage degradation, decreased autophagy is linked with cell death. Rapamycin, which is a specific inhibitor of the mTOR signaling pathway, enhances expression of autophagy regulators and prevents chondrocyte death. In the future, pharmacological activation of autophagy may be an effective therapeutic approach for OA. © 2015 Société franc¸aise de rhumatologie. Published by Elsevier Masson SAS. All rights reserved.
1. Introduction Osteoarthritis (OA), the most common age-related joint pathology, is a degenerative disease affecting all the structures of the joints and is characterized by articular cartilage destruction along with changes in other joint components, including bone, meniscus, synovium, ligament, capsule, and muscle [1]. Multiple factors have been implicated in the pathogenesis of OA, including mechanical, genetic, and age-associated factors, with age being the major risk factor for OA [2]. Chondrocytes are capable of responding to structural changes in the surrounding cartilage matrix although the capacity of adult articular chondrocytes to regenerate normal cartilage matrix architecture is limited and declines with age, due to cell death and abnormal responsiveness to anabolic stimuli [3]. Therefore, maintaining the chondrocytes in a healthy condition appears to be an important factor for preservation of the entire cartilage and preventing its degeneration. Autophagy (from auto-phagos: self-eating) is an essential cellular homeostatic mechanism for the removal of dysfunctional cellular organelles and macromolecules [4]. The autophagy pathway is integrated with multiple signal transduction pathways that respond to nutrient supply, energy balance, cytokines, and growth factors [4]. Here, we
∗ Corresponding author. E-mail address: [email protected] (G.-H. Lei).
summarize the current understanding of the molecular mechanism of autophagy, focusing on recent studies that have examined the role of autophagy in the pathogenesis of OA. 2. Definition of autophagy All living organisms undergo continuous renovation. Cells constantly adapt to changing environmental conditions by adjusting their content to prevailing needs. This process, known as homeostasis, involves continuous biosynthesis and turnover of cellular components. Eukaryotic cells have two major degradation systems: the lysosome and the proteasome [5]. Proteasomal degradation has high selectivity; the proteasome generally recognizes only ubiquitinated substrates, which are primarily short-lived proteins. In contrast, degradation in the lysosome does not follow such a simple pattern. Extracellular material and plasma membrane proteins can be delivered to lysosomes for degradation via the endocytic pathway. Furthermore, cytosolic components and organelles can also be delivered to the lysosome by autophagy [5]. Autophagy is a mechanism of cellular homeostasis that is strongly induced in a wide range of eukaryotic organisms and in multiple different cell types under conditions of nutrient starvation. This leads to bulk degradation of cytoplasmic components (proteins, organelles), the building blocks of which are used as an energy supply and the synthesis of components essential for survival [6,7]. In cells defective in autophagy, the total intracellular pool of amino
http://dx.doi.org/10.1016/j.jbspin.2015.06.009 1297-319X/© 2015 Société franc¸aise de rhumatologie. Published by Elsevier Masson SAS. All rights reserved.
Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
2
acids is greatly reduced, leading to the inability to synthesize proteins that are essential for cellular survival [8]. Autophagy may also serve other functions and is important for cellular housekeeping by removing exhausted, redundant or unnecessary components. Therefore, autophagy is implicated in the retardation of aging and in the prevention and therapy of age-associated diseases [9], as well as in support of cell remodeling during development [10] or contributing to cellular defence against pathogens [11]. Autophagy is a generic term for all pathways by which cytoplasmic materials are delivered to the lysosome in animal cells or the vacuole in plant and yeast cells [5]. The currently known autophagy processes are subdivided into three types depending on the route of cargo delivery: chaperone-mediated autophagy, macroautophagy and microautophagy [12]. Of these, chaperonemediated autophagy is, as yet, described only in mammals and is involved in the degradation of individual, soluble proteins. In contrast, macroautophagy and microautophagy occur in a wide range of eukaryotes, including mammals, plants and fungi, leading to the degradation of portions of the cytoplasm, which may include cellular organelles [12]. Macroautophagy is thought to be the major type of autophagy and is mediated by double (or multiple) membranebound vacuoles, the formation of which is highly conserved from yeast to humans [13]. Macroautophagy is the most extensively studied form of autophagy and is, therefore, referred to simply as “autophagy” hereafter. Fig. 1. The regulation factors in autophagy progression. : interaction effect. fx2: negative effect;
: positive effect;
3. The basic process of autophagy The main purpose of autophagy in yeast is to degrade the cytoplasm and recycle the resulting macromolecules for use in the synthesis of essential components during nutrient stress [14]. Although autophagy and autophagy-related processes are dynamic, they can be broken down into several discrete steps [14,15]: • induction: in mammalian cells, autophagy occurs at a basal level under vegetative growth conditions. Accordingly, there must be a mechanism for sensing the extracellular milieu and transducing an appropriate signal to regulatory elements that allow the induction of autophagy. One of the major regulatory components is the protein kinase TOR (target of rapamycin), which inhibits autophagy under basal or nutrient-rich conditions [16]; • cargo selection and packaging: autophagy is generally considered to be a non-specific process in which bulk cytoplasm is randomly sequestered into the cytosolic autophagosome; • nucleation of vesicle formation: this is the initial step that recruits proteins and lipids for autophagosome construction; • vesicle expansion and completion: autophagosome formation is a de novo process, in which membranes emerging at the preautophagosomal structure (PAS) expand, either by direct flow from a source, such as the endoplasmic reticulum (ER) or by vesicular addition, and then seal to enclose the cytosolic cargo; • retrieval: the process by which certain components are retrieved for reuse; • targeting: docking and fusion of the completed vesicle with the lysosome/vacuole; • breakdown of the intraluminal vesicle (either the Cvt or autophagic bodies) and its cargo and recycling of the macromolecular constituents. 4. Regulatory factors Several protein kinases and signaling pathways are involved in autophagy and autophagy-related processes. A number of critical autophagy-related genes (ATG or Atg) have been identified, the gene products of which regulate distinct steps in the induction Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
or progression of autophagy [17]. The Ser/Thr protein kinase TOR, which plays a key role in signaling of nutrient limitation [18], resides in a multiprotein complex 1, mTORC1 [19]. In response to stimulation by nutrients or growth factors, mTORC1 negatively regulates the mammalian uncoordinated-51-like protein kinase (ULK1) complex that includes ULK1, ATG13, ATG101, and FIP200 (RB1CC1), which results in autophagy suppression [20,21]. Under glucose starvation, AMP-activated protein kinase (AMPK) promotes autophagy by activating ULK1 directly. Under nutrient sufficiency, high mTOR activity prevents ULK1 activation and disrupts the interaction between ULK1 and AMPK [22]. In addition, autophagy is negatively regulated by the phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathway [23], which contains three highly conserved proteins, namely the protein kinase Vps15, phosphatidylinositol-3-kinase Vps34 [24] and Atg6 [25]. Autophagy is also regulated by the Beclin-1 complex, consisting of Beclin-1, class III phosphatidylinositol-3kinase (VPS34 or PI3KC3) and ATG14L or UVRAG. This is subject to negative regulation by the PI3K/Akt pathway [23] as well as by binding interactions with the anti-apoptotic Bcl-2 family proteins [26]. Stimulation of the Beclin-1 complex generates phosphatidylinositol-3-phosphate (PI3P) [27], which triggers autophagosomal nucleation. Following phagophore formation, elongation of the autophagosome membrane requires the action of two ubiquitin-like conjugation systems, the Atg5–Atg12 conjugation system and the microtubule-associated protein-1 light chain 3 (LC3, Atg8) conjugation system [28,29]. Atg4B converts the proform of LC3B to its cytosolic free form (LC3-I). In mammals, the conversion of LC3-I (and other Atg8 homologues) to its phosphatidylethanolamine-conjugated and autophagosome membrane-associated form (i.e., LC3-II) is an initiating step in autophagy [30,31]. Fig. 1 depicts a schematic flow chart describing the regulation factors in autophagy progression. 5. Expression of autophagy in OA The articular cartilage is an avascular, aneural, alymphatic and viscoelastic connective tissue that derives its nutrition and oxygen Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
supply by diffusion from the synovial fluid and the subchondral bone [32]. Chondrocytes are responsible for both synthesis and turnover of the abundant extracellular matrix (ECM) [3]; therefore, maintaining the chondrocytes in a healthy condition appears to be an important factor for maintaining the integrity of the entire cartilage. Autophagy has been suggested to be an important cell survival mechanism under various forms of stress [4]. Autophagy serves not only to regulate the final stages of the chondrocyte lifecycle, but also the rate at which chondrocytes enter the maturation process [33]. Autophagy in normal adult articular cartilage is an important mechanism for cellular homeostasis [34]; thus, cells in the superficial zone express high levels of the autophagy proteins BECN1, ATG5, and MAP1LC3 [34]. Catabolic and nutritional stresses increase autophagy in OA, and during the initial degenerative phase at least, autophagy is increased in OA chondrocytes and cartilage, with increased expression of LC3 and Beclin-1 messenger RNA (mRNA) in OA chondrocytes [35]. Furthermore, autophagy regulates changes in OA-like gene expression through the modulation of apoptosis and reactive oxygen species (ROS) [35], which promote oxidative stress resulting in the assembly of multiprotein inflammatory complexes, called the inflammasome [36]. Disruption of the interplay between autophagy and inflammasome contributes to the age-related increase in the prevalence of OA and decreased efficacy of articular cartilage repair [37]. Enhanced autophagy, suppressed mTOR and reduced MAP4K3 activity are considered to constitute an early response in the biomechanicallyinduced degeneration of the temporomandibular joint cartilage, especially in premature chondrocytes [38]. Transiently increased autophagy represents a compensatory response to cellular stress, with damage occurring when prolonged stress exceeds the capacity of this mechanism [39] and failure to mount an autophagic response may lead to further degeneration [33]. In cartilage with mild OA, ULK1, Beclin-1, and LC3 protein expression are reduced in the cartilage superficial zone, although these three proteins are strongly expressed in the OA cell clusters as well as the middle and deep zones [40]. In mouse OA knee joints, expression of ULK1, Beclin-1, and LC3 is decreased together with glycosaminoglycan (GAG) loss, while PARP p85 expression is increased with the gradual cartilage degradation [40]. Furthermore, the reduction in these key regulators of autophagy is accompanied by increased apoptosis. Aging of mouse knee joints is associated with reduced autophagy and cellularity along with increased apoptotic cell death [41]. mTOR is overexpressed in human OA cartilage as well as mouse and dog experimental OA. Upregulated mTOR expression in OA correlates with increased chondrocyte apoptosis and reduced expression of key autophagy genes, including ATG3, ATG5, ATG12, ULK1, LC3B, Beclin-1 and BNIP3 [42]. In the cartilage-specific ablation of mTOR knockout mice, autophagy signaling is increased with a significant reduction in articular cartilage degradation, apoptosis and synovial fibrosis [42]. Stress conditions in the epiphyseal growth plate can promote the autophagic response through the modulation of genes controlling metabolite utilization to achieve survival of the terminally differentiated cells exposed to the brief rigors of the harsh local microenvironment [43]. In a mouse model consisting of the green fluorescent protein (GFP) fused to light chain 3 (LC3) on the C57BL/6J genetic background (GFP-LC3 transgenic mice) generated to activate autophagy in normal cartilage and monitor autophagy changes in age-related cartilage degradation, the number and area of autophagosomes in chondrocytes of older mice were significantly decreased compared to those in younger mice. Furthermore, the average number and size of vesicles in osteocytes in subchondral bone decreased in old mice [41]. In GFP-LC3 transgenic mice, the highest levels of GFP-LC3 and the key autophagy proteins (Atg5 and LC3) were observed in chondrocytes in the superficial and upper mid-zones of articular cartilage in young mice, while the levels were much lower in deep zone cells and also in old mice [41]. Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
3
These observations in cartilage are consistent with the notion that basal autophagic activity decreases with age [41,44] and that the marked inhibition of autophagy would have a negative impact on chondrocyte survival and differentiation [33], thus contributing to the accumulation of damaged macromolecules and susceptibility to age-related OA [44]. 6. Hypoxia, mechanical injury and autophagy Articular cartilage is maintained in a low oxygen environment throughout life [32]. It has been demonstrated that an oxygen gradient exists in cartilage from around 6% at the joint surface to 1% in the deep layers [45]. Chondrocytes are therefore adapted to these hypoxic conditions. Hypoxia-inducible factors (HIFs) are transcription factors that respond to decreases in the level of available oxygen in the cellular environment. Two main HIF isoforms (HIF1␣ and HIF-2␣) mediate the response of all cells to hypoxia [46,47]. HIF-1␣ may protect articular cartilage by promoting the chondrocyte phenotype, maintaining chondrocyte viability, and supporting metabolic adaptation to a hypoxic environment. In contrast, HIF2␣ is a catabolic factor in the pathogenesis of OA [48]. HIF-1␣, which is expressed in normal and OA human articular chondrocytes cultured under normoxic conditions [49] can be further induced or stabilized by hypoxia. The relatively high constitutive expression of HIF-1␣ in chondrocytes may be an important adaptation to survival in the avascular-hypoxic environment of cartilage [49]. HIF-2␣ expression is also higher in OA cartilages versus normal cartilage in mice and humans [50]. HIF-1␣ and HIF-2␣ play critical roles in the survival, development, and differentiation of chondrocytes in vascular hypoxia [51]. Under hypoxic conditions, both HIF-1␣ and HIF-2␣ are more strongly associated with the nuclei in the middle/deep cells, compared with the full-thickness and superficial zone chondrocytes [52]. A relationship between HIF-1 ␣ and increased accumulation of autophagic proteins, such as Beclin-1, has been reported [53], while HIF-2␣ has been shown to be a suppressor of autophagy in vitro [54]. In chondrocytes, AMPK (the enzyme that monitors the cellular energy status) is activated in a HIF-1-dependent manner. This enzyme inhibits the effects of mTOR on development of the autophagic state [55]. Studies have shown that 1% O2 significantly increased phospho-AMPK (pAMPK) protein expression compared with the effects of 5% or 20% O2 [56]. Suppression of AMPK activates mTOR and survival signaling mediated by the Akt pathway. The relationship between HIF-1, AMPK, mTOR, and autophagy involves HIF-1 activation in response to a reduction in the cellular energy charge during tissue hypoxia, which then results in the phosphorylation and activation of AMPK. Once activated, AMPK suppresses mTOR, thereby activating autophagic flux, which is also activated directly by HIF-1 [55]. In growth plate chondrocytes, increased HIF-1-mediated glycolytic activity elevates AMP levels. The resulting AMP kinase activation and suppression of mTOR causes increased autophagy. Once autophagy is activated, the post-mitotic chondrocytes remain viable in their unique microenvironment and complete their lifecycle [57]. Moreover, HIF-2 is robustly expressed in the pre-hypertrophic cells of the mouse growth cartilage and suppresses chondrocyte autophagy, while elevated autophagic responses are observed throughout the growth plate in HIF-2␣-knockout mice [54]. Silenced HIF-2␣ in chondrocytes leads to decreased Akt-1 and mTOR activities, reduced Bcl-XL expression and a robust autophagic response [54]. Mechanical load is the other main risk factor for OA besides aging. Mechanical injury also leads to suppression of autophagy [58] and has been shown to induce cell death and loss of sGAG, in association with significantly decreased ULK1, Beclin-1 and LC3 expression in the cartilage superficial zone at 48 hours post-injury. Rapamycin enhances the expression of autophagy regulators and prevents cell death and sGAG loss in mechanically injured explants [58] Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
4
Surgically induced OA in mice lacking the von Hippel-Lindau gene (Vhl) in osteochonadral progenitor cells (referred to as Vhl cKO mice) resulted in HIF-2␣ upregulation, increased chondrocyte apoptosis as well as decreased autophagy with cartilage destruction and proteoglycan loss during OA development [59]. In surgically induced OA in PPAR␥ KO mice, increased cartilage degradation and chondrocyte apoptosis was observed, as well as the overproduction of OA inflammatory/catabolic factors associated with the increased expression of mTOR and the suppression of key autophagy markers [60]. 7. Inflammation and autophagy The inflammation of the synovial membrane that occurs in both the early and late phases of OA is associated with alterations in the adjacent cartilage [61]. Increased expression of proinflammatory cytokines in cartilage, synovial membrane and subchondral bone are believed to be linked to the development and progression of structural changes in the OA joint [62]. Inflammatory signals activate inflammasome-dependent responses and caspases, predominantly caspase-1, which cleave the inactive precursors of IL-1 and IL-18 and stimulate their secretion and activity. Consequently, these cytokines provoke inflammatory responses and accelerate the aging process by inhibiting autophagy [63]. Increased expression of mTOR in peripheral blood mononuclear cells of OA patients is related to the disease activity associated with synovitis [64]. IL-1 treatment of human OA chondrocytes results in a significant increase in mTOR expression [42]. Blocking autophagy potentiated inflammasome activity, whereas stimulating autophagy had the opposite effect. Assembled inflammasomes undergo ubiquitination and recruit the autophagic adaptor p62, which assists their delivery to autophagosomes. Thus, autophagy accompanies inflammasome activation to moderate inflammation by eliminating active inflammasomes [65]. PPAR␥ deficiency in chondrocytes results in the aberrant expression of inflammatory markers (iNOS and COX-2) associated with the dysregulated expression of mTOR and inhibition of critical autophagy markers, ultimately resulting in increased inflammatory activity within the articular cartilage leading to severe/accelerated OA [60]. Therefore, mTOR is an important link between synovitis and structural damage in inflammatory arthritis. Inhibition of mTOR reduces synovial osteoclast formation and protects against local bone erosion and cartilage loss. Clinical signs of arthritis are improved after mTOR inhibition, and histologic evaluation showed a decrease in synovitis [66]. Rapamycin treatment of C57Bl/6 OA mice also maintains cartilage cellularity, and decreases IL-1 expression in articular cartilage [67]. 8. Pharmacological targets of autophagy Augmentation of autophagy via inhibition of the TOR signaling pathway by genetic or pharmacological intervention extends lifespan, indicating that the further development of interventions targeting mTOR represents a strategy for the treatment and prevention of age-related diseases [68]. Moreover, since autophagy is a critical protective mechanism against mitochondrial dysfunction, pharmacologic interventions that enhance autophagy may have chondroprotective activity in cartilage degenerative processes in OA [69]. In experimental OA mice, rapamycin treatment maintains cartilage cellularity and decreases ADAMTS-5 and interleukin-1 expression in articular cartilage. Rapamycin also reduces the severity of synovitis and the synovial tissue level of IL-1 in the knee joints in murine experimental OA by inhibiting ribosomal protein S6 phosphorylation [67]. Clinical signs of arthritis are improved and histological signs decreased in synovitis after mTOR inhibition by rapamycin or everolimus, and in vitro, mTOR inhibition Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
downregulates the expression of digestive enzymes and leads to osteoclast apoptosis [66]; thus, when autophagy is activated, the severity of experimental OA is improved [67]. Recently, a study indicated that glucosamine, which is effective in animal models of OA and has anti-inflammatory and anabolic effects on cartilage cells, modulates molecular targets of the autophagy pathway both in vitro and in vivo. Furthermore, the enhancement of autophagy by increased LC3-II levels, formation of LC3 puncta, and increased LC3 turnover is mainly dependent on the Akt/FoxO/mTOR pathway [70]. Glucosamine protects nucleus pulposus (NP) cells and upregulates autophagy in a dose-dependent manner within 24 hours by inhibiting the mTOR pathway by reducing mTOR phosphorylation [71]. Similarly, glucosamine protects PPAR␥-mTOR double KO mice against DMM-induced OA. This effect was associated with significant protection from cartilage destruction, proteoglycan loss and loss of chondrocytes in both the medial tibial plateau and the medial femoral condyle, with a significant reduction in chondrocyte cell death and the percentage of MMP13-positive cells, as well as enhanced LC3B II expression in double KO OA chondrocytes [60]. Local intra-articular injection of rapamycin in a murine model of OA indicated that rapamycin significantly reduced the severity of articular cartilage degradation, with a reduction in mTOR expression and activation of LC3 in the chondrocytes. Furthermore, rapamycin treatment also reduced VEGF, COL10A1, and MMP13 expression after DMM surgery [72]. In a murine OA model, after intra-articular administration of gelatin hydrogels incorporating rapamycin micelles, OA mediator genes (IL-1 and IL-6, MMP9, MMP13, C/EBP, and mTOR) were downregulated while Col2a1 was upregulated in the rapamycin-treated mice [73]. Thus, intraarticular administration of rapamycin could be a novel therapeutic approach for OA [72,73]. In conclusion, age is the major risk factor for OA. The capacity of adult articular chondrocytes to regenerate the normal cartilage matrix architecture is limited and declines with age. During the development of OA, autophagy may increase as an adaptive response to protect chondrocytes from various environmental changes, while failure of the adaptation may lead to progression in cartilage degradation. Thus, pharmacological activation of autophagy may be an appropriate therapeutic approach for OA. Disclosure of interest The authors declare that they have no conflicts of interest concerning this article. Acknowledgments This work was supported by the National Natural Science Foundation of China (No.81272034, 81402224 and 81501923), the Provincial Science Foundation of Hunan (No.2015JJ3139), the Scientific Research Project of Science and Technology Bureau of Hunan Province (2012FJ6001), the Scientific Research Project of Science and Technology Office of Changsha City (K1203040-31), the Scientific Research Project of Health and Family Planning Commission of Hunan Province (B2014-12), the Hunan Provincial Innovation Foundation for Postgraduates (CX2012B086), the Fundamental Research Funds for the Central Universities of Central South University (2013zzts081, 2013zzts319) and the College Student’s Innovation and Entrepreneurship Project of Central South University (DL14505). References [1] Hunter DJ, Felson DT. Osteoarthritis. BMJ 2006;332:639–42. [2] Shane AA, Loeser RF. Why is osteoarthritis an age-related disease? Best Pract Res Clin Rheumatol 2010;24:15–26.
Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
[3] Loeser RF. Aging and osteoarthritis: the role of chondrocyte senescence and aging changes in the cartilage matrix. Osteoarthritis Cartilage 2009;17: 971–9. [4] Mizushima N. Physiological functions of autophagy. Curr Top Microbiol Immunol 2009;335:71–84. [5] Mizushima N, Komatsu M. Autophagy: renovation of cells and tissues. Cell 2011;147:728–41. [6] Tsukada M, Ohsumi Y. Isolation and characterization of autophagy-defective mutants of Saccharomyces cerevisiae. Febs Lett 1993;333:169–74. [7] Levine B, Kroemer G. Autophagy in the pathogenesis of disease. Cell 2008;132:27–42. [8] Onodera J, Ohsumi Y. Autophagy is required for maintenance of amino acid levels and protein synthesis under nitrogen starvation. J Biol Chem 2005;280:31582–6. [9] Bergamini E. Autophagy: a cell repair mechanism that retards ageing and ageassociated diseases and can be intensified pharmacologically. Mol Aspects Med 2006;27:403–10. [10] Levine B, Klionsky DJ. Development by self-digestion: molecular mechanisms and biological functions of autophagy. Dev Cell 2004;6:463–77. [11] Mizushima N, Levine B, Cuervo AM, et al. Autophagy fights disease through cellular self-digestion. Nature 2008;451:1069–75. [12] Todde V, Veenhuis M, van der Klei IJ. Autophagy: principles and significance in health and disease. Biochim Biophys Acta 2009;1792:3–13. [13] Klionsky DJ, Abeliovich H, Agostinis P, et al. Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes. Autophagy 2008;4:151–75. [14] Klionsky DJ. The molecular machinery of autophagy: unanswered questions. J Cell Sci 2005;118:7–18. [15] Chen Y, Klionsky DJ. The regulation of autophagy – unanswered questions. J Cell Sci 2011;124:161–70. [16] Carrera AC. TOR signaling in mammals. J Cell Sci 2004;117:4615–6. [17] Klionsky DJ. Autophagy. Curr Biol 2005;15:R282–3. [18] Pattingre S, Espert L, Biard-Piechaczyk M, et al. Regulation of macroautophagy by mTOR and Beclin 1 complexes. Biochimie 2008;90:313–23. [19] Jung CH, Ro SH, Cao J, et al. mTOR regulation of autophagy. Febs Lett 2010;584:1287–95. [20] Mizushima N. The role of the Atg1/ULK1 complex in autophagy regulation. Curr Opin Cell Biol 2010;22:132–9. [21] Ganley IG, Lam DH, Wang J, et al. ULK1.ATG13.FIP200 complex mediates mTOR signaling and is essential for autophagy. J Biol Chem 2009;284:12297–305. [22] Kim J, Kundu M, Viollet B, et al. AMPK and mTOR regulate autophagy through direct phosphorylation of Ulk1. Nat Cell Biol 2011;13:132–41. [23] Wang RC, Wei Y, An Z, et al. Akt-mediated regulation of autophagy and tumorigenesis through Beclin 1 phosphorylation. Science 2012;338:956–9. [24] Kihara A, Noda T, Ishihara N, et al. Two distinct Vps34 phosphatidylinositol 3-kinase complexes function in autophagy and carboxypeptidase Y sorting in Saccharomyces cerevisiae. J Cell Biol 2001;152:519–30. [25] Kametaka S, Okano T, Ohsumi M, et al. Apg14p and Apg6/Vps30p form a protein complex essential for autophagy in the yeast, Saccharomyces cerevisiae. J Biol Chem 1998;273:22284–91. [26] Pattingre S, Tassa A, Qu X, et al. Bcl-2 antiapoptotic proteins inhibit Beclin 1dependent autophagy. Cell 2005;122:927–39. [27] He C, Levine B. The Beclin 1 interactome. Curr Opin Cell Biol 2010;22:140–9. [28] Tanida I, Ueno T, Kominami E. LC3 conjugation system in mammalian autophagy. Int J Biochem Cell Biol 2004;36:2503–18. [29] Ohsumi Y. Molecular dissection of autophagy: two ubiquitin-like systems. Nat Rev Mol Cell Biol 2001;2:211–6. [30] He H, Dang Y, Dai F, et al. Post-translational modifications of three members of the human MAP1LC3 family and detection of a novel type of modification for MAP1LC3B. J Biol Chem 2003;278:29278–87. [31] Satoo K, Noda NN, Kumeta H, et al. The structure of Atg4B-LC3 complex reveals the mechanism of LC3 processing and delipidation during autophagy. Embo J 2009;28:1341–50. [32] Milner PI, Fairfax TP, Browning JA, et al. The effect of O2 tension on pH homeostasis in equine articular chondrocytes. Arthritis Rheum 2006;54:3523–32. [33] Shapiro IM, Layfield R, Lotz M, et al. Boning up on autophagy: the role of autophagy in skeletal biology. Autophagy 2014;10:7–19. [34] Almonte-Becerril M, Navarro-Garcia F, Gonzalez-Robles A, et al. Cell death of chondrocytes is a combination between apoptosis and autophagy during the pathogenesis of osteoarthritis within an experimental model. Apoptosis 2010;15:631–8. [35] Sasaki H, Takayama K, Matsushita T, et al. Autophagy modulates osteoarthritis-related gene expression in human chondrocytes. Arthritis Rheum 2012;64:1920–8. [36] Salminen A, Ojala J, Kaarniranta K, et al. Mitochondrial dysfunction and oxidative stress activate inflammasomes: impact on the aging process and age-related diseases. Cell Mol Life Sci 2012;69:2999–3013. [37] Mobasheri A, Matta C, Zakany R, et al. Chondrosenescence: definition, hallmarks and potential role in the pathogenesis of osteoarthritis. Maturitas 2015;80:237–44. [38] Zhang M, Zhang J, Lu L, et al. Enhancement of chondrocyte autophagy is an early response in the degenerative cartilage of the temporomandibular joint to biomechanical dental stimulation. Apoptosis 2013;18: 423–34. [39] Barranco C, Osteoarthritis:. Activate autophagy to prevent cartilage degeneration? Nat Rev Rheumatol 2015;11:127.
Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
5
[40] Carames B, Taniguchi N, Otsuki S, et al. Autophagy is a protective mechanism in normal cartilage, and its aging-related loss is linked with cell death and osteoarthritis. Arthritis Rheum 2010;62:791–801. [41] Carames B, Olmer M, Kiosses WB, et al. The relationship of autophagy defects and cartilage damage during joint aging in a mouse model. Arthritis Rheumatol 2015;67:1568–76. [42] Zhang Y, Vasheghani F, Li YH, et al. Cartilage-specific deletion of mTOR upregulates autophagy and protects mice from osteoarthritis. Ann Rheum Dis 2015;74:1432–40. [43] Srinivas V, Shapiro IM. Chondrocytes embedded in the epiphyseal growth plates of long bones undergo autophagy prior to the induction of osteogenesis. Autophagy 2006;2:215–6. [44] Cuervo AM, Dice JF. Age-related decline in chaperone-mediated autophagy. J Biol Chem 2000;275:31505–13. [45] Silver IA. Measurement of pH and ionic composition of pericellular sites. Philos Trans R Soc Lond B Biol Sci 1975;271:261–72. [46] Semenza GL. Hypoxia-inducible factors in physiology and medicine. Cell 2012;148:399–408. [47] Smith TG, Robbins PA, Ratcliffe PJ. The human side of hypoxia-inducible factor. Br J Haematol 2008;141:325–34. [48] Zhang FJ, Luo W, Lei GH. Role of HIF-1alpha and HIF-2alpha in osteoarthritis. Joint Bone Spine 2015;82:144–7. [49] Coimbra IB, Jimenez SA, Hawkins DF, et al. Hypoxia inducible factor-1 alpha expression in human normal and osteoarthritic chondrocytes. Osteoarthritis Cartilage 2004;12:336–45. [50] Saito T, Fukai A, Mabuchi A, et al. Transcriptional regulation of endochondral ossification by HIF-2alpha during skeletal growth and osteoarthritis development. Nat Med 2010;16:678–86. [51] Aro E, Khatri R, Gerard-O’Riley R, et al. Hypoxia-inducible factor-1 (HIF-1) but not HIF-2 is essential for hypoxic induction of collagen prolyl 4-hydroxylases in primary newborn mouse epiphyseal growth plate chondrocytes. J Biol Chem 2012;287:37134–44. [52] Schrobback K, Malda J, Crawford RW, et al. Effects of oxygen on zonal marker expression in human articular chondrocytes. Tissue Eng Part A 2012;18:920–33. [53] Bohensky J, Shapiro IM, Leshinsky S, et al. HIF-1 regulation of chondrocyte apoptosis: induction of the autophagic pathway. Autophagy 2007;3: 207–14. [54] Bohensky J, Terkhorn SP, Freeman TA, et al. Regulation of autophagy in human and murine cartilage: hypoxia-inducible factor 2 suppresses chondrocyte autophagy. Arthritis Rheum 2009;60:1406–15. [55] Bohensky J, Leshinsky S, Srinivas V, et al. Chondrocyte autophagy is stimulated by HIF-1 dependent AMPK activation and mTOR suppression. Pediatr Nephrol 2010;25:633–42. [56] Fermor B, Gurumurthy A, Diekman BO. Hypoxia, RONS and energy metabolism in articular cartilage. Osteoarthritis Cartilage 2010;18:1167–73. [57] Srinivas V, Bohensky J, Shapiro IM. Autophagy: a new phase in the maturation of growth plate chondrocytes is regulated by HIF, mTOR and AMP kinase. Cells Tissues Organs 2009;189:88–92. [58] Carames B, Taniguchi N, Seino D, et al. Mechanical injury suppresses autophagy regulators and pharmacologic activation of autophagy results in chondroprotection. Arthritis Rheum 2012;64:1182–92. [59] Weng T, Xie Y, Yi L, et al. Loss of Vhl in cartilage accelerated the progression of age-associated and surgically induced murine osteoarthritis. Osteoarthritis Cartilage 2014;22:1197–205. [60] Vasheghani F, Zhang Y, Li YH, et al. PPARgamma deficiency results in severe, accelerated osteoarthritis associated with aberrant mTOR signalling in the articular cartilage. Ann Rheum Dis 2015;74:569–78. [61] Sellam J, Berenbaum F. The role of synovitis in pathophysiology and clinical symptoms of osteoarthritis. Nat Rev Rheumatol 2010;6:625–35. [62] Kapoor M, Martel-Pelletier J, Lajeunesse D, et al. Role of proinflammatory cytokines in the pathophysiology of osteoarthritis. Nat Rev Rheumatol 2011;7:33–42. [63] Salminen A, Kaarniranta K, Kauppinen A. Inflammaging: disturbed interplay between autophagy and inflammasomes. Aging 2012;4:166–75. [64] Tchetina EV, Poole AR, Zaitseva EM, et al. Differences in mammalian target of rapamycin gene expression in the peripheral blood and articular cartilages of osteoarthritic patients and disease activity. Arthritis 2013;2013: 461486. [65] Shi CS, Shenderov K, Huang NN, et al. Activation of autophagy by inflammatory signals limits IL-1beta production by targeting ubiquitinated inflammasomes for destruction. Nat Immunol 2012;13:255–63. [66] Cejka D, Hayer S, Niederreiter B, et al. Mammalian target of rapamycin signaling is crucial for joint destruction in experimental arthritis and is activated in osteoclasts from patients with rheumatoid arthritis. Arthritis Rheum 2010;62:2294–302. [67] Carames B, Hasegawa A, Taniguchi N, et al. Autophagy activation by rapamycin reduces severity of experimental osteoarthritis. Ann Rheum Dis 2012;71:575–81. [68] Harrison DE, Strong R, Sharp ZD, et al. Rapamycin fed late in life extends lifespan in genetically heterogeneous mice. Nature 2009;460:392–5. [69] Lopez DFP, Lotz MK, Blanco FJ, et al. Autophagy activation and protection from mitochondrial dysfunction in human chondrocytes. Arthritis Rheumatol 2015;67:966–76. [70] Carames B, Kiosses WB, Akasaki Y, et al. Glucosamine activates autophagy in vitro and in vivo. Arthritis Rheum 2013;65:1843–52.
Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),
G Model BONSOI-4208; No. of Pages 6 6
ARTICLE IN PRESS Y.-S. Li et al. / Joint Bone Spine xxx (2015) xxx–xxx
[71] Jiang L, Jin Y, Wang H, et al. Glucosamine protects nucleus pulposus cells and induces autophagy via the mTOR-dependent pathway. J Orthop Res 2014;32:1532–42. [72] Takayama K, Kawakami Y, Kobayashi M, et al. Local intra-articular injection of rapamycin delays articular cartilage degeneration in a murine model of osteoarthritis. Arthritis Res Ther 2014;16:482.
Please cite this article in press as: Li http://dx.doi.org/10.1016/j.jbspin.2015.06.009
Y-S,
et
al.
[73] Matsuzaki T, Matsushita T, Tabata Y, et al. Intra-articular administration of gelatin hydrogels incorporating rapamycin-micelles reduces the development of experimental osteoarthritis in a murine model. Biomaterials 2014;35:9904–11.
Autophagy
in
osteoarthritis.
Joint
Bone
Spine
(2015),