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Autophagy mediated CoCrMo particle-induced peri-implant osteolysis by promoting osteoblast apoptosis.

Wang Z, Liu N, Liu K, Zhou G, Gan J, Wang Z, Shi T, He W, Wang L, Guo T, Bao N, Wang R, Huang Z, Chen J, Dong L, Zhao J, Zhang J - Autophagy (2015)

Bottom Line: Both autophagy inhibitor 3-MA (3-methyladenine) and siRNA of Atg5 could dramatically reduce CoPs-induced apoptosis in osteoblasts.Further, inhibition of autophagy with 3-MA ameliorated the severity of osteolysis in PIO animal models.Moreover, 3-MA also prevented osteoblast apoptosis in an antiautophagic way when tested in PIO model.

View Article: PubMed Central - PubMed

Affiliation: a Jinling Hospital; Department of Orthopaedics; State Key Laboratory of Pharmaceutical Biotechnology; Nanjing University ; Nanjing , China.

ABSTRACT
Wear particle-induced osteolysis is the leading cause of aseptic loosening, which is the most common reason for THA (total hip arthroplasty) failure and revision surgery. Although existing studies suggest that osteoblast apoptosis induced by wear debris is involved in aseptic loosening, the underlying mechanism linking wear particles to osteoblast apoptosis remains almost totally unknown. In the present study, we investigated the effect of autophagy on osteoblast apoptosis induced by CoCrMo metal particles (CoPs) in vitro and in a calvarial resorption animal model. Our study demonstrated that CoPs stimulated autophagy in osteoblasts and PIO (particle-induced osteolysis) animal models. Both autophagy inhibitor 3-MA (3-methyladenine) and siRNA of Atg5 could dramatically reduce CoPs-induced apoptosis in osteoblasts. Further, inhibition of autophagy with 3-MA ameliorated the severity of osteolysis in PIO animal models. Moreover, 3-MA also prevented osteoblast apoptosis in an antiautophagic way when tested in PIO model. Collectively, these results suggest that autophagy plays a key role in CoPs-induced osteolysis and that targeting autophagy-related pathways may represent a potential therapeutic approach for treating particle-induced peri-implant osteolysis.

No MeSH data available.


Related in: MedlinePlus

The ERN1-MAPK8 pathway mediated the activation of autophagy induced by CoPs. (A) Western blots performed after osteoblast cells were incubated with CoPs (50 µg/ml) for 12 h. (B) The density of the western blots bands shown in (A) was quantified using ImageJ software. (C) Western blots performed after osteoblast cells were incubated with siCtrl or siErn1 before being stimulated with CoPs (50 µg/ml) for 12 h. siCtrl, siControl. (D) The density of the western blots bands shown in (C) was quantified using ImageJ software. (E) Western blots performed after fibroblast cells were incubated with SP600125 (15 µM) before being stimulated with CoPs (50 µg/ml) for 12 h. (F) The density of the western blots bands shown in (E) was quantified using ImageJ software. Data are presented as means ± S.E.M. from 3 independent experiments. *, P < 0.05 and **, P < 0.01 vs. control; ##, P < 0.01 vs. CoPs group.
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f0002: The ERN1-MAPK8 pathway mediated the activation of autophagy induced by CoPs. (A) Western blots performed after osteoblast cells were incubated with CoPs (50 µg/ml) for 12 h. (B) The density of the western blots bands shown in (A) was quantified using ImageJ software. (C) Western blots performed after osteoblast cells were incubated with siCtrl or siErn1 before being stimulated with CoPs (50 µg/ml) for 12 h. siCtrl, siControl. (D) The density of the western blots bands shown in (C) was quantified using ImageJ software. (E) Western blots performed after fibroblast cells were incubated with SP600125 (15 µM) before being stimulated with CoPs (50 µg/ml) for 12 h. (F) The density of the western blots bands shown in (E) was quantified using ImageJ software. Data are presented as means ± S.E.M. from 3 independent experiments. *, P < 0.05 and **, P < 0.01 vs. control; ##, P < 0.01 vs. CoPs group.

Mentions: Previous studies have suggested that autophagy can be triggered by members of the endoplasmic reticulum (ER) stress signaling pathway including ERN1/IRE1α (endoplasmic reticulum to nucleus signaling 1) and EIF2AK3/PERK (eukaryotic translation initiation factor 2 α kinase 3), and we have also observed that CoPs can induce ER stress in macrophages.15,26,27 Thus, we sought to determine which ER stress signaling pathway(s) is/are involved in CoPs-stimulated autophagy in osteoblasts. CoPs significantly increased the expression of ERN1 and phosphorylated EIF2AK3 (Fig. 2A and B, and Fig. S1A and B). Small interfering RNA against Ern1 (siErn1) significantly decreased the expression of LC3-II (Fig. 2C and D), but no obvious differences between siEif2ak3 and siControl were observed (Fig. S1C to F). Furthermore, we detected the expression of phosphorylated MAPK8/JNK1 (mitogen-activated protein kinase 8) and XBP1 (X-box binding protein 1), both of which are downstream signaling effectors of ERN1.28-30 Interestingly, we observed significantly increased p-MAPK8 levels in cells treated with CoPs (Fig. 2A and B), but not XBP1 (Fig. S1A and B). Next, we asked whether CoPs-induced autophagy was mediated by MAPK8 after activation by ERN1. Osteoblasts were cotreated with CoPs and SP600125, a MAPK8 inhibitor. LC3-II was significantly reduced in cells treated with MAPK8 inhibitors (Fig. 2E and F). Collectively, these results indicated that CoPs-induced autophagy was mediated by the ERN1-MAPK8 pathway.Figure 2.


Autophagy mediated CoCrMo particle-induced peri-implant osteolysis by promoting osteoblast apoptosis.

Wang Z, Liu N, Liu K, Zhou G, Gan J, Wang Z, Shi T, He W, Wang L, Guo T, Bao N, Wang R, Huang Z, Chen J, Dong L, Zhao J, Zhang J - Autophagy (2015)

The ERN1-MAPK8 pathway mediated the activation of autophagy induced by CoPs. (A) Western blots performed after osteoblast cells were incubated with CoPs (50 µg/ml) for 12 h. (B) The density of the western blots bands shown in (A) was quantified using ImageJ software. (C) Western blots performed after osteoblast cells were incubated with siCtrl or siErn1 before being stimulated with CoPs (50 µg/ml) for 12 h. siCtrl, siControl. (D) The density of the western blots bands shown in (C) was quantified using ImageJ software. (E) Western blots performed after fibroblast cells were incubated with SP600125 (15 µM) before being stimulated with CoPs (50 µg/ml) for 12 h. (F) The density of the western blots bands shown in (E) was quantified using ImageJ software. Data are presented as means ± S.E.M. from 3 independent experiments. *, P < 0.05 and **, P < 0.01 vs. control; ##, P < 0.01 vs. CoPs group.
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Related In: Results  -  Collection

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f0002: The ERN1-MAPK8 pathway mediated the activation of autophagy induced by CoPs. (A) Western blots performed after osteoblast cells were incubated with CoPs (50 µg/ml) for 12 h. (B) The density of the western blots bands shown in (A) was quantified using ImageJ software. (C) Western blots performed after osteoblast cells were incubated with siCtrl or siErn1 before being stimulated with CoPs (50 µg/ml) for 12 h. siCtrl, siControl. (D) The density of the western blots bands shown in (C) was quantified using ImageJ software. (E) Western blots performed after fibroblast cells were incubated with SP600125 (15 µM) before being stimulated with CoPs (50 µg/ml) for 12 h. (F) The density of the western blots bands shown in (E) was quantified using ImageJ software. Data are presented as means ± S.E.M. from 3 independent experiments. *, P < 0.05 and **, P < 0.01 vs. control; ##, P < 0.01 vs. CoPs group.
Mentions: Previous studies have suggested that autophagy can be triggered by members of the endoplasmic reticulum (ER) stress signaling pathway including ERN1/IRE1α (endoplasmic reticulum to nucleus signaling 1) and EIF2AK3/PERK (eukaryotic translation initiation factor 2 α kinase 3), and we have also observed that CoPs can induce ER stress in macrophages.15,26,27 Thus, we sought to determine which ER stress signaling pathway(s) is/are involved in CoPs-stimulated autophagy in osteoblasts. CoPs significantly increased the expression of ERN1 and phosphorylated EIF2AK3 (Fig. 2A and B, and Fig. S1A and B). Small interfering RNA against Ern1 (siErn1) significantly decreased the expression of LC3-II (Fig. 2C and D), but no obvious differences between siEif2ak3 and siControl were observed (Fig. S1C to F). Furthermore, we detected the expression of phosphorylated MAPK8/JNK1 (mitogen-activated protein kinase 8) and XBP1 (X-box binding protein 1), both of which are downstream signaling effectors of ERN1.28-30 Interestingly, we observed significantly increased p-MAPK8 levels in cells treated with CoPs (Fig. 2A and B), but not XBP1 (Fig. S1A and B). Next, we asked whether CoPs-induced autophagy was mediated by MAPK8 after activation by ERN1. Osteoblasts were cotreated with CoPs and SP600125, a MAPK8 inhibitor. LC3-II was significantly reduced in cells treated with MAPK8 inhibitors (Fig. 2E and F). Collectively, these results indicated that CoPs-induced autophagy was mediated by the ERN1-MAPK8 pathway.Figure 2.

Bottom Line: Both autophagy inhibitor 3-MA (3-methyladenine) and siRNA of Atg5 could dramatically reduce CoPs-induced apoptosis in osteoblasts.Further, inhibition of autophagy with 3-MA ameliorated the severity of osteolysis in PIO animal models.Moreover, 3-MA also prevented osteoblast apoptosis in an antiautophagic way when tested in PIO model.

View Article: PubMed Central - PubMed

Affiliation: a Jinling Hospital; Department of Orthopaedics; State Key Laboratory of Pharmaceutical Biotechnology; Nanjing University ; Nanjing , China.

ABSTRACT
Wear particle-induced osteolysis is the leading cause of aseptic loosening, which is the most common reason for THA (total hip arthroplasty) failure and revision surgery. Although existing studies suggest that osteoblast apoptosis induced by wear debris is involved in aseptic loosening, the underlying mechanism linking wear particles to osteoblast apoptosis remains almost totally unknown. In the present study, we investigated the effect of autophagy on osteoblast apoptosis induced by CoCrMo metal particles (CoPs) in vitro and in a calvarial resorption animal model. Our study demonstrated that CoPs stimulated autophagy in osteoblasts and PIO (particle-induced osteolysis) animal models. Both autophagy inhibitor 3-MA (3-methyladenine) and siRNA of Atg5 could dramatically reduce CoPs-induced apoptosis in osteoblasts. Further, inhibition of autophagy with 3-MA ameliorated the severity of osteolysis in PIO animal models. Moreover, 3-MA also prevented osteoblast apoptosis in an antiautophagic way when tested in PIO model. Collectively, these results suggest that autophagy plays a key role in CoPs-induced osteolysis and that targeting autophagy-related pathways may represent a potential therapeutic approach for treating particle-induced peri-implant osteolysis.

No MeSH data available.


Related in: MedlinePlus