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《The Journal of Traditional Chinese Orthopedics and Traumatology》 2018-08
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Effect of Duhuo Jisheng Tang(独活寄生汤) medicated serum on protein kinase R-like endoplasmic reticulum kinase/immunoglobulin-binding protein signaling pathway in degenerated chondrocytes of rats

CHEN Jun;WU Guangwen;XU Huifeng;ZHENG Chunsong;DAI Yichen;QIU Jianqing;LIU Shuru;CHEN Zhenyuan;LIU Xianxiang;Fujian University of Traditional Chinese Medicine;Fujian Key Laboratory of Integrated Medicine on Geriatrics;  
Objective: To observe the effect of Duhuo Jisheng Tang(独活寄生汤,DHJST) medicated serum on protein kinase R-like endoplasmic reticulum kinase( PERK)/immunoglobulin-binding protein( BiP) signaling pathway in degenerated chondrocytes of rats.Methods: Thirty-six 2-month-old clean-grade male SD rats were randomly divided into medicated serum group and blank serum group,18 rats in each group. The rats in medicated serum group were intragastric administrated with DHJST in dosage of 9. 3 g/kg,while the others in blank serum group were intragastric administrated with the normal saline in dosage of 10 m L/kg,once a day for consecutive 7 days. After the last intragastric administration,their blood were fetched out from abdominal aorta under anesthesia and were made into DHJST medicated serum and blank serum respectively. The knee joints of 4-week-old SD rats were fetched out and the chondrocytes were isolated from knee articular cartilage and were cultured. The third-generation chondrocytes were intervented by interleukin-1β( IL-1β) for 24 hours and then the degenerated chondrocytes were obtained. The obtained degenerated chondrocytes were divided into 2 groups and were intervened by DHJST medicated serum( DHJST group) and blank serum( blank group) respectively. The degenerated chondrocytes apoptosis rate,the contents of PERK mRNA,Bip mRNA,eukaryotic initiation factor 2α( e IF-2α) mRNA,activating transcription factor 4( ATF-4) mRNA and growth arrest and DNA damage-inducible 153( GADD153) mRNA and the activities of cysteine containing aspartate specific protease( Caspase)-9 and Caspase-3 in degenerated chondrocytes were detected at 24,48 and 72 hours after the intervention respectively. Results: There was interaction between time factor and group factor in degenerated chondrocytes apoptosis rate( F = 9. 022,P =0. 038). There was statistical difference in degenerated chondrocytes apoptosis rate between the 2 groups in general,in other words,there was group effect( F = 325. 246,P = 0. 000). There was statistical difference in chondrocytes apoptosis rate between different timepoints after the intervention,in other words,there was time effect( F = 389. 768,P = 0. 000). The chondrocytes apoptosis rate presented a time-dependent decreasing trend in both of the 2 groups. The chondrocytes apoptosis rate was lower in DHJST group compared to blank group at different timepoints after intervention( t = 14. 055,P = 0. 000; t = 13. 264,P = 0. 000; t = 12. 183,P = 0. 000). There was no interaction between time factor and group factor in the content of PERK mRNA in degenerated chondrocytes( F = 3. 002,P = 0. 157). The content of PERK mRNA in chondrocytes was lower in DHJST group compared to blank group in general,in other words,there was group effect( F =17. 833,P = 0. 013). There was statistical difference in content of PERK mRNA in chondrocytes between different timepoints after intervention,in other words,there was time effect( F = 43. 060,P = 0. 003). The content of PERK mRNA in chondrocytes presented a time-dependent decreasing trend in both of the 2 groups. There was no interaction between time factor and group factor in the content of Bip mRNA in degenerated chondrocytes( F = 4. 092,P = 0. 106). The content of Bip mRNA in chondrocytes was lower in DHJST group compared to blank group in general,in other words,there was group effect( F = 15. 136,P = 0. 018). There was statistical difference in content of Bip mRNA in chondrocytes between different timepoints after intervention,in other words,there was time effect( F = 62. 968,P = 0. 001). The content of Bip mRNA in chondrocytes presented a time-dependent decreasing trend in both of the 2 groups. There was no interaction between time factor and group factor in the content of e IF-2α mRNA in degenerated chondrocytes( F = 2. 138,P = 0. 201). The content of e IF-2α mRNA in chondrocytes was lower in DHJST group compared to blank group in general,in other words,there was group effect( F =155. 852,P = 0. 000). There was statistical difference in content of e IF-2α mRNA in chondrocytes between different timepoints after intervention,in other words,there was time effect( F = 50. 328,P = 0. 000). The content of e IF-2α mRNA in chondrocytes presented a time-dependent decreasing trend in both of the 2 groups,and the 2 groups were consistent with each other in the decreasing trend of content of e IF-2α mRNA in chondrocytes. There was interaction between time factor and group factor in the content of ATF-4 mRNA in degenerated chondrocytes( F = 14. 612,P = 0. 017). There was statistical difference in content of ATF-4 mRNA in chondrocytes between the 2 groups in general,in other words,there was group effect( F = 33. 703,P = 0. 004). There was statistical difference in content of ATF-4 mRNA in chondrocytes between different timepoints after intervention,in other words,there was time effect( F = 187. 700,P = 0. 000). The content of ATF-4 mRNA in chondrocytes presented a time-dependent decreasing trend in both of the 2 groups. The content of ATF-4 mRNA in chondrocytes was lower in DHJST group compared to blank group at different timepoints after intervention( t = 4. 343,P = 0. 012;t = 3. 915,P = 0. 017; t = 10. 719,P = 0. 000). There was no interaction between time factor and group factor in content of GADD153 mRNA in degenerated chondrocytes( F = 3. 053,P = 0. 116). The content of GADD153 mRNA in chondrocytes was lower in DHJST group compared to blank group in general,in other words,there was group effect( F = 16. 946,P = 0. 015). There was statistical difference in content of GADD153 mRNA in chondrocytes between different timepoints after intervention,in other words,there was time effect( F = 88. 206,P =0. 000). The content of GADD153 mRNA in chondrocytes presented a time-dependent decreasing trend in both of the 2 groups. There was no interaction between time factor and group factor in the activity of Caspase-9 in degenerated chondrocytes( F = 1. 266,P = 0. 327). The activity of Caspase-9 in chondrocytes was lower in DHJST group compared to blank group in general,in other words,there was group effect( F = 21. 164,P = 0. 010). There was statistical difference in activity of Caspase-9 in chondrocytes between different timepoints after intervention,in other words,there was time effect( F = 263. 384,P = 0. 000). The activity of Caspase-9 in chondrocytes presented a time-dependent decreasing trend in both of the 2 groups,and the 2 groups were consistent with each other in the decreasing trend of activity of Caspase-9 in chondrocytes. There was no interaction between time factor and group factor in activity of Caspase-3 in degenerated chondrocytes( F = 1. 346,P = 0. 314). The activity of Caspase-3 in chondrocytes was lower in DHJST group compared to blank group in general,in other words,there was group effect( F = 19. 422,P = 0. 012). There was statistical difference in activity of Caspase-3 in chondrocytes between different timepoints after intervention,in other words,there was time effect( F = 130. 649,P = 0. 000). The activity of Caspase-3 in chondrocytes presented a time-dependent decreasing trend in both of the 2 groups,and the 2 groups were consistent with each other in the decreasing trend of activity of Caspase-3 in chondrocytes. Conclusion: DHJST medicated serum can inhibit the chondrocyte apoptosis caused by endoplasmic reticulum stress reaction through regulating PERK/Bi P signaling pathway in the degenerated chondrocytes of rats.
【Fund】: 国家自然科学基金项目(81573801);; 福建省自然科学基金杰青项目(2017J06018);; 福建省卫生计生科研人才培养项目(2017-ZQN-62)
【CateGory Index】: R285.5
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