BACKGROUND:Due to its unknown etiology and the lack of definitive curative treatments,management of temporomandibular joint osteoarthritis primarily focuses on symptom relief.Therefore,the identification of effective early molecular diagnostic biomarkers or potential therapeutic targets holds great significance.OBJECTIVE:To investigate the expression of mitochondrial creatine kinase 2 in temporomandibular joint osteoarthritis in rats and its role in the progression of inflammation.METHODS:(1)Animal experiment:Twenty Sprague-Dawley rats were randomly divided into control and unilateral anterior crossbite groups(n=10).A rat model of temporomandibular joint osteoarthritis was made in the unilateral anterior crossbite group.Four weeks after modeling,histological evaluations,including hematoxylin-eosin and Safranin O-fast green staining,were performed to assess pathological changes in the cartilage and subchondral bone of the mandibular condyle.Quantitative real-time PCR and immunohistochemical staining were utilized to detect the mRNA and protein expression levels of interleukin-1β,matrix metalloproteinase 13,type II collagen,aggrecan,and mitochondrial creatine kinase 2 in condylar cartilage.(2)Cell experiment:Passage 3 condylar cartilage cells from Sprague-Dawley rats were divided into control group and model group.Cells in the control group were routinely cultured,while an inflammation model of condylar cartilage cells was established with interleukin 1β in the model group were treated with interleukin-1β to induce inflammatory cell models.After 24 hours of interleukin-1β treatment,western blot was used to evaluate the expression of matrix metalloproteinase 13,type II collagen proteins in chondrocytes.Western blot was also used to detect the protein expression of mitochondrial creatine kinase 2 in the model group at 0,12,24,48,and 72 hours after treatment with interleukin-1β.RESULTS AND CONCLUSION:(1)Animal experiment:The results of hematoxylin-eosin staining showed that the unilateral anterior crossbite group exhibited disintegration of the fibrous layer in the cartilage of the mandibular condyle,disorganization of chondrocyte hierarchy,dense cellularity of the proliferative layer,obvious cell clustering,and infiltration of inflammatory cells in the subchondral bone.The results of Safranin O-fast green staining showed that in the cartilage matrix in the unilateral anterior crossbite group,the red stain became less extensive and lighter in color,and the tidal line was discontinuous.The mRNA and protein expression levels of interleukin-1β and matrix metalloproteinase 13 were elevated in the unilateral anterior crossbite group compared with the control group(P<0.05).Conversely,the relative expression of type II collagen,aggrecan,and mitochondrial creatine kinase 2 mRNA and protein decreased(P<0.05).(2)Cell experiment:Compared with the control group,the protein expression of matrix metalloproteinase 13 in the experimental group was significantly increased(P<0.05),while the protein expression of type II collagen was decreased(P<0.05).With prolonged interleukin-1β treatment,the protein expression of mitochondrial creatine kinase 2 in condylar chondrocytes of the model group gradually decreased.All the results indicate that the relative expression level of mitochondrial creatine kinase 2 was negatively correlated with the degree of chondrocyte inflammation in the rat model of temporomandibular joint osteoarthritis.Therefore,it is reasonable to infer that mitochondrial creatine kinase 2 is an important indicator for assessing the progression of inflammation in temporomandibular joint osteoarthritis.

BACKGROUND:Due to its unknown etiology and the lack of definitive curative treatments,management of temporomandibular joint osteoarthritis primarily focuses on symptom relief.Therefore,the identification of effective early molecular diagnostic biomarkers or potential therapeutic targets holds great significance.OBJECTIVE:To investigate the expression of mitochondrial creatine kinase 2 in temporomandibular joint osteoarthritis in rats and its role in the progression of inflammation.METHODS:(1)Animal experiment:Twenty Sprague-Dawley rats were randomly divided into control and unilateral anterior crossbite groups(n=10).A rat model of temporomandibular joint osteoarthritis was made in the unilateral anterior crossbite group.Four weeks after modeling,histological evaluations,including hematoxylin-eosin and Safranin O-fast green staining,were performed to assess pathological changes in the cartilage and subchondral bone of the mandibular condyle.Quantitative real-time PCR and immunohistochemical staining were utilized to detect the mRNA and protein expression levels of interleukin-1β,matrix metalloproteinase 13,type II collagen,aggrecan,and mitochondrial creatine kinase 2 in condylar cartilage.(2)Cell experiment:Passage 3 condylar cartilage cells from Sprague-Dawley rats were divided into control group and model group.Cells in the control group were routinely cultured,while an inflammation model of condylar cartilage cells was established with interleukin 1β in the model group were treated with interleukin-1β to induce inflammatory cell models.After 24 hours of interleukin-1β treatment,western blot was used to evaluate the expression of matrix metalloproteinase 13,type II collagen proteins in chondrocytes.Western blot was also used to detect the protein expression of mitochondrial creatine kinase 2 in the model group at 0,12,24,48,and 72 hours after treatment with interleukin-1β.RESULTS AND CONCLUSION:(1)Animal experiment:The results of hematoxylin-eosin staining showed that the unilateral anterior crossbite group exhibited disintegration of the fibrous layer in the cartilage of the mandibular condyle,disorganization of chondrocyte hierarchy,dense cellularity of the proliferative layer,obvious cell clustering,and infiltration of inflammatory cells in the subchondral bone.The results of Safranin O-fast green staining showed that in the cartilage matrix in the unilateral anterior crossbite group,the red stain became less extensive and lighter in color,and the tidal line was discontinuous.The mRNA and protein expression levels of interleukin-1β and matrix metalloproteinase 13 were elevated in the unilateral anterior crossbite group compared with the control group(P<0.05).Conversely,the relative expression of type II collagen,aggrecan,and mitochondrial creatine kinase 2 mRNA and protein decreased(P<0.05).(2)Cell experiment:Compared with the control group,the protein expression of matrix metalloproteinase 13 in the experimental group was significantly increased(P<0.05),while the protein expression of type II collagen was decreased(P<0.05).With prolonged interleukin-1β treatment,the protein expression of mitochondrial creatine kinase 2 in condylar chondrocytes of the model group gradually decreased.All the results indicate that the relative expression level of mitochondrial creatine kinase 2 was negatively correlated with the degree of chondrocyte inflammation in the rat model of temporomandibular joint osteoarthritis.Therefore,it is reasonable to infer that mitochondrial creatine kinase 2 is an important indicator for assessing the progression of inflammation in temporomandibular joint osteoarthritis.