Loss of β-arrestin2 aggravated condylar cartilage degeneration at the early stage of temporomandibular joint osteoarthritis

Abstract

Objective: Temporomandibular joint osteoarthritis (TMJOA) is a chronic degenerative joint disorder characterized by extracellular matrix degeneration and inflammatory response of condylar cartilage. β-arrestin2 is an important regulator of inflammation response, while its role in TMJOA remains unknown. The objective of this study was to investigate the role of β-arrestin2 in the development of TMJOA at the early stage and the underlying mechanism.

Methods: A unilateral anterior crossbite (UAC) model was established on eight-week-old wild-type (WT) and β-arrestin2 deficiency mice to simulate the progression of TMJOA. Hematoxylin-eosin (HE) staining and microcomputed tomography (micro-CT) analysis were used for histological and radiographic assessment. Immunohistochemistry was performed to detect the expression of inflammatory and degradative cytokines, as well as autophagy related factors. Terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) assay was carried out to assess chondrocyte apoptosis.

Results: The loss of β-arrestin2 aggravated cartilage degeneration and subchondral bone destruction in the model of TMJOA at the early stage. Furthermore, in UAC groups, the expressions of degradative (Col-X) and inflammatory (TNF-α and IL-1β) factors in condylar cartilage were increased in β-arrestin2 null mice compared with WT mice. Moreover, the loss of β-arrestin2 promoted apoptosis and autophagic process of chondrocytes at the early stage of TMJOA.

Conclusion: In conclusion, we demonstrated for the first time that β-arrestin2 plays a protective role in the development of TMJOA at the early stage, probably by inhibiting apoptosis and autophagic process of chondrocytes. Therefore, β-arrestin2 might be a potential therapeutic target for TMJOA, providing a new insight for the treatment of TMJOA at the early stage.

Keywords: Apoptosis; Autophagy; Cartilage degeneration; Inflammation; Temporomandibular joint osteoarthritis; β-arrestin2.

Fig. 1

Effects of β-arrestin2 on condylar cartilage thickness at the early stage of TMJOA. (A) Schematic diagram of the animal experiment and main methodological steps. (B) Immunohistochemistry of β-arrestin2 in condylar cartilage of WT mice at 3 weeks of sham or UAC operation. (C) Quantitative analysis of positive cell rate of β-arrestin2 shown in 1 A (n = 6). (D) HE-stained sections of condylar cartilage at 3 weeks. (E) Quantification of cartilage thickness of the samples shown in 1 C (n = 6). Data were presented as mean ± SD. *P < 0.05

Fig. 2

Loss of β-arrestin2 aggravated cartilage matrix degradation at the early stage of TMJOA. (A) Safranin O staining of condylar cartilage of different groups at 3 weeks. (B) Quantitative data of the percentage of Safranin O positive area (n = 6). (C) Immunohistochemistry of COL-II in condylar cartilage at 3 weeks. (D) Quantitative analysis of positive cell rate of COL-II shown in 2 C (n = 6). (E) Immunohistochemistry of COL-X in condylar cartilage at 3 weeks. (F) Quantitative analysis of positive cell rate of COL-X shown in 2 E (n = 6). Data were presented as mean ± SD. *P < 0.05, **P < 0.01

Fig. 3

Loss of β-arrestin2 exacerbated subchondral bone destruction at the early stage of TMJOA. (A) Representative images showing trabecular architecture by micro-CT reconstruction in the subchondral bone at 3 weeks. (B) Quantitative analysis of BV/TV in the subchondral bone (n = 6). (C) Quantitative analysis of Tb.N in the subchondral bone (n = 6). (D) Quantitative analysis of Tb.Sp in the subchondral bone (n = 6). (E) Quantitative analysis of Tb.Th in the subchondral bone (n = 6). (F) Representative images of TRAP staining of the subchondral bone. (G) Quantitative analysis of TRAP positive cell numbers (n = 6). Data were presented as mean ± SD. *P < 0.05, **P < 0.01

Fig. 4

Loss of β-arrestin2 promotes chondrocyte inflammation at the early stage of TMJOA. (A) Immunohistochemistry of TNF-α in condylar cartilage at 3 weeks. (B) Quantitative analysis of positive cell rate of TNF-α shown in 4 A (n = 6). (C) Immunohistochemistry of IL-1β in condylar cartilage at 3 weeks. (D) Quantitative analysis of positive cell rate of IL-1β shown in 4 C (n = 6). Data were presented as mean ± SD. *P < 0.05, **P < 0.01

Fig. 5

Loss of β-arrestin2 promoted apoptosis and autophagic process of chondrocytes at the early-stage TMJOA. (A) Representative images of TUNEL staining of the condylar cartilage. (B) Quantitative analysis of TUNEL positive cell numbers shown in 5 A (n = 6). (C) Immunohistochemistry of BECN in condylar cartilage at 3 weeks. (D) Quantitative analysis of positive cell rate of BECN shown in 5 C (n = 6). (E) Immunohistochemistry of LC3B-II in condylar cartilage at 3 weeks. (F) Quantitative analysis of positive cell rate of LC3B-II shown in 5E (n = 6). Data were presented as mean ± SD. *P < 0.05, **P < 0.01