The Role Of Endoplasmic Reticulum Stress On Rat Mandibular Condylar Chondrocytes And Its Regulation By Mechanical Stress

Part1:The study on protective effects of salubrinal on mandibular condylar chondrocytes apoptosis induced by tunicamycinObjective:To investigate the protective effects of salubrinal on mandibular condylar chondrocytes apoptosis induced by tunicamycin.Methods:1. Mandibular condylar chondrocytes from3-week-old Sprague-Dawley rats were isolated and cultured in vitro. Third-generation condylar chondrocytes on slides were validated by type II collagen immunohistochemistry staining.2. Chondrocytes were exposed to40μM Salubrinal for30minutes followed by4μg/ml Tunicamycin treatment for24hours. Chondrocytes were divided into four groups: Control, Tunicamycin, Salubrinal, Salubrinal+Tunicamycin. Flow cytometer was used to detect cell apoptosis after24h treatment. The expression of GRP78, GRP94, CHOP, Caspase-12was detected by real-time quantitive PCR and Western Blot.Results:1. Chondrocytes apoptosis increased (P<0.05) after24hours treatment of tunicamycin. Salubrinal has no effect on chondrocytes apoptosis. Pretreatment of salubrinal could partically decreased chondrocytes apoptosis induced by tunicamycin.2. Tunicamycin increased the expression of GRP78, GRP94, CHOP, Caspase-12on both transcription and protein translation level in mandibular condylar chondrocytes. Salubrinal+Tunicamycin down regulated their expression.Conclusion:1. Tunicamycin induced ER stress in mandibular condylar chondrocytes and triggered apoptosis.2. Salubrinal protected mandibular condylar chondrocytes apoptosis induced by Tunicamycin through down regulated ER stress marker proteins. Part2:The role of endoplasmic reticulum stress in mandibular condylar chondrocytes under cyclic tensile stretchObjective:To investigate whether cyclic tensile stress could induce endoplasmic reticulum stress in mandibular condylar chondrocytes.Methods:1. Mandibular condylar chondrocytes from3-week-old Sprague-Dawley rats were isolated and cultured in vitro. Third-generation condylar chondrocytes on slides were validated by type Ⅱ collagen immunohistochemistry staining.2. Mandibular condylar chondrocytes were subjected to12%elongation of tensile stess for24hours, collagen Ⅱ expression was detected by real-time quantitive PCR and cell apoptosis was detected by flow cytometry.3. Chondrocytes were subjected to12%elongation of tensile stress for24hours or with a combination of4μg/ml Tunicamycin. Chondrocytes were divided into four groups: Control, Tunicamycin, cyclic tensile stress, cyclic tensile stress+Tunicamycin. The expression of GRP78, GRP94, CHOP, Caspase-12was detected by real-time quantitive PCR and Western Blot. Results:1. The expression of collagen II was decreased after12%cyclic tensile stress for24h.12%cyclic tensile stress for24h did not induce mandibular condylar chondrocytes apoptosis.2. After24hours12%cyclic tensile stress, the expression of GRP78, GRP94, CHOP, Caspase-12did not increase on both transcription and translation level in mandibular condylar chondrocytes. In the presence of12%cyclic tensile stress and tunicamycin, the expression of GRP78, GRP94, CHOP were higher than treated by tunicamycin alone, the expression of CHOP was statistically significant (P<0.05).Conclusion:1.12%cyclic tensile stress for24h did not induce ERS and apoptosis in mandibular condylar chondrocytes significantly.2.12%cyclic tensile stress+Tunicamycin for24h can aggravate ERS in mandibular condylar chondrocytes.