OBJECTIVE:To investigate the effect of constant compressive stress induced by imitating Tuina stimulation with various durations on the cell cycle,cellular secretion,apoptosis,and expression of myogenic regulatory fac...OBJECTIVE:To investigate the effect of constant compressive stress induced by imitating Tuina stimulation with various durations on the cell cycle,cellular secretion,apoptosis,and expression of myogenic regulatory factors(MRFs),myogenic factor 5(Myf5)and myogenic differentiation(MyoD)of rat skeletal muscle cells(RSkMCs)in vitro.METHODS:Third passage RSkMCs were subjected to constant compressive stresses with various durations at 2000μstrain for 15,30,60,90,and 120 min via a four-point bending system.The control group(CG)was cultured in the absence of mechanical loading.Alterations of the cell cycle and apoptosis rate were detected by flow cytometry(FCM).The concentrations of interleukin 6(IL-6)/prostaglandin E2(PGE2)and nitric oxide(NO)in supernatants were determined by enzyme-linked immunosorbent assays and the nitrate reductase method,respectively.Expression of Myf5 and MyoD was detected by immunohistochemistry.RESULTS:Compared with the CG,a significant alteration was observed in the synthesis phase fraction(SPF)(P<0.01).The SPF and proliferation index(PI)were reduced from 15 to 90 min,but reached levels similar to those at 120 min.Apoptosis was increased significantly at 30 min(P<0.05)and especially at 90 and 120 min(P<0.01).Expression of MyoD and Myf5 was increased significantly at 15,30,and 90 min(P<0.01).Compared with 15 and 30 min,MyoD and Myf5 expression at 60 and 120 min was decreased significantly(P<0.01).Compared with 60 min,M yoD expression at 90 min was increased significantly(P<0.05),whereas MyoD and Myf5 expression at 120 min was significantly lower(P<0.05).The IL-6 concentration was increased at 60 min compared with the CG and 15 min(P<0.05),whereas the concentrations of PGE2 and NO were the highest at 15 and 30 min,respectively,compared with the CG and other time points(P<0.05).CONCLUSION:The cell cycle,secretion,apoptosis,and Myf5 and MyoD expression of RSkMCs were regulated by compressive stress in a time-dependent manner.SPF and PI were inhibited at short durations(<90 min),but NO and PGE2 secretion was the highest at shorter durations(<30 min).With the prolongation of stimulation time,SPF,PI,and apoptosis were increased,but Myf5 and MyoD expression was decreased gradually at 15-30 min.展开更多
Myogenic Regulatory Factors(MRFs)is involved in the muscle growth and differentiation.In this study,the cDNA sequence of yellowtail kingfish MRFs genes were cloned by rapid amplification of cDNA ends(RACE)method;then,...Myogenic Regulatory Factors(MRFs)is involved in the muscle growth and differentiation.In this study,the cDNA sequence of yellowtail kingfish MRFs genes were cloned by rapid amplification of cDNA ends(RACE)method;then,the character-istics of these genes and the predicted protein sequences were analyzed by bioinformatics methods,the tissue and embryonic stages differential expression pattern were detected by the quantitative real-time PCR.Our results showed that the yellowtail kingfish(YTK)Myf5 cDNA has a full length of 951 bp,encoding 266 amino acids.The yellowtail kingfish Myf6 cDNA has a full length of 1105bp,encoding 250 amino acids.The proteins containα-helix,β-strand,and loops.The Neighbour-joining tree revealed that YTK Myf5 and Myf6 are closely related to Seriola dumerili.The yellowtail kingfish Myf5 and Myf6 gene expressed significantly higher in mus-cle than in other tissues(P<0.05).In addition,Myf5 and Myf6 in muscle was significantly expressed in 400g and 500 g fish but not in 50 g,suggesting that myogenic regulatory factors expression had a great relationship with the fish size.Our results also indicated that Myf5 and Myf6 have different functions during embryonic development,because Myf5 showed highest expression level at the neuroembryo period,but Myf6 had the highest expression level at embryo coverage yolk 70%stage.Myf5 gene showed highest ex-pression at 30 d of age,suggesting it played key roles in myogenic period.However,the Myf6 gene was significantly highly ex-pressed at 60 d,revealing this gene functioned in the later muscle formation period.展开更多
OBJECTIVE: To investigate the effects of Gyejibokryeong-Hwan(Guizhifuling-wan, GBH) on muscle injury in a mouse model of muscle contusion.METHODS: C57/BL6 mouse biceps femoris muscles were injured using the drop-mass ...OBJECTIVE: To investigate the effects of Gyejibokryeong-Hwan(Guizhifuling-wan, GBH) on muscle injury in a mouse model of muscle contusion.METHODS: C57/BL6 mouse biceps femoris muscles were injured using the drop-mass method and injured animals were treated orally with GBH(50,100, or 500 mg/kg) once a day for 7 d. Open field and treadmill running tests were performed to assess functional recovery from muscle injury. The production of pro-inflammatory cytokines was examined by enzyme-linked immunosorbent assay and Western blotting analysis. Expression of the muscle regeneration biomarkers, myoblast determination(Myo D), myogenic factor 5(Myf5), and smooth muscle actin(α-SMA), in the biceps femoris muscle was investigated at the protein and m RNA level by Western blotting and real time-PCR, respectively. Histological analysis was performed using hematoxylin and eosin staining. Finally, myosin heavy chain production was investigated in differentiated C2C12 myoblasts in the presence of GBH.RESULTS: GBH treatment markedly improved locomotion and running behavior. GBH significantly inhibited the secretion of monocyte chemoattractant protein-1 into the bloodstream in muscle-contused animals. The levels of Myo D, Myf5, and α-SMA protein and m RNA were significantly up-regulated by GBH in injured muscle tissue. Histological studies suggested that GBH facilitated recovery from muscle damage. However, GBH did not induce the production of myosin heavy chain in vitro.CONCLUSION: Overall, the present study suggested that GBH improves the recovery of the injured muscles in the mouse model of muscle contusion.展开更多
基金a Grant from the National Natural Science Foundation of China:the Research on Biomechanical Parameters of Kneading Manipulation and Their Optimization on FEM of Skeletal Muscle Cells(No.81373760)。
文摘OBJECTIVE:To investigate the effect of constant compressive stress induced by imitating Tuina stimulation with various durations on the cell cycle,cellular secretion,apoptosis,and expression of myogenic regulatory factors(MRFs),myogenic factor 5(Myf5)and myogenic differentiation(MyoD)of rat skeletal muscle cells(RSkMCs)in vitro.METHODS:Third passage RSkMCs were subjected to constant compressive stresses with various durations at 2000μstrain for 15,30,60,90,and 120 min via a four-point bending system.The control group(CG)was cultured in the absence of mechanical loading.Alterations of the cell cycle and apoptosis rate were detected by flow cytometry(FCM).The concentrations of interleukin 6(IL-6)/prostaglandin E2(PGE2)and nitric oxide(NO)in supernatants were determined by enzyme-linked immunosorbent assays and the nitrate reductase method,respectively.Expression of Myf5 and MyoD was detected by immunohistochemistry.RESULTS:Compared with the CG,a significant alteration was observed in the synthesis phase fraction(SPF)(P<0.01).The SPF and proliferation index(PI)were reduced from 15 to 90 min,but reached levels similar to those at 120 min.Apoptosis was increased significantly at 30 min(P<0.05)and especially at 90 and 120 min(P<0.01).Expression of MyoD and Myf5 was increased significantly at 15,30,and 90 min(P<0.01).Compared with 15 and 30 min,MyoD and Myf5 expression at 60 and 120 min was decreased significantly(P<0.01).Compared with 60 min,M yoD expression at 90 min was increased significantly(P<0.05),whereas MyoD and Myf5 expression at 120 min was significantly lower(P<0.05).The IL-6 concentration was increased at 60 min compared with the CG and 15 min(P<0.05),whereas the concentrations of PGE2 and NO were the highest at 15 and 30 min,respectively,compared with the CG and other time points(P<0.05).CONCLUSION:The cell cycle,secretion,apoptosis,and Myf5 and MyoD expression of RSkMCs were regulated by compressive stress in a time-dependent manner.SPF and PI were inhibited at short durations(<90 min),but NO and PGE2 secretion was the highest at shorter durations(<30 min).With the prolongation of stimulation time,SPF,PI,and apoptosis were increased,but Myf5 and MyoD expression was decreased gradually at 15-30 min.
基金the National Key Research and Development Program(Nos.2018YFD0901204 and 2019YFD0900503)the National Natural Science Foun-dation of China(No.31772829)+4 种基金the Laboratory for Ma-rine Fisheries Science and Food Production Processes,Qingdao National Laboratory for Marine Science and Technology,P.R.China(No.2017-3A01)Central Pub-lic-interest Scientific Institution Basal Research,CAFS&Key Laboratory of Sustainable Development of Marine Fisheries,Ministry of Agriculture and Rural Affairs,P.R.China(No.2019HY-XKQ01)the Central Public-interest Scientific Institution Basal Research Fund,YSFRI,CAFS(No.20603022021004)the Central Public-interest Scien-tific Institution Basal Research fund,CAFS(No.47)the China Agriculture Research System(No.CARS-47).
文摘Myogenic Regulatory Factors(MRFs)is involved in the muscle growth and differentiation.In this study,the cDNA sequence of yellowtail kingfish MRFs genes were cloned by rapid amplification of cDNA ends(RACE)method;then,the character-istics of these genes and the predicted protein sequences were analyzed by bioinformatics methods,the tissue and embryonic stages differential expression pattern were detected by the quantitative real-time PCR.Our results showed that the yellowtail kingfish(YTK)Myf5 cDNA has a full length of 951 bp,encoding 266 amino acids.The yellowtail kingfish Myf6 cDNA has a full length of 1105bp,encoding 250 amino acids.The proteins containα-helix,β-strand,and loops.The Neighbour-joining tree revealed that YTK Myf5 and Myf6 are closely related to Seriola dumerili.The yellowtail kingfish Myf5 and Myf6 gene expressed significantly higher in mus-cle than in other tissues(P<0.05).In addition,Myf5 and Myf6 in muscle was significantly expressed in 400g and 500 g fish but not in 50 g,suggesting that myogenic regulatory factors expression had a great relationship with the fish size.Our results also indicated that Myf5 and Myf6 have different functions during embryonic development,because Myf5 showed highest expression level at the neuroembryo period,but Myf6 had the highest expression level at embryo coverage yolk 70%stage.Myf5 gene showed highest ex-pression at 30 d of age,suggesting it played key roles in myogenic period.However,the Myf6 gene was significantly highly ex-pressed at 60 d,revealing this gene functioned in the later muscle formation period.
基金Supported by Daejeon University Research Grants (2017)。
文摘OBJECTIVE: To investigate the effects of Gyejibokryeong-Hwan(Guizhifuling-wan, GBH) on muscle injury in a mouse model of muscle contusion.METHODS: C57/BL6 mouse biceps femoris muscles were injured using the drop-mass method and injured animals were treated orally with GBH(50,100, or 500 mg/kg) once a day for 7 d. Open field and treadmill running tests were performed to assess functional recovery from muscle injury. The production of pro-inflammatory cytokines was examined by enzyme-linked immunosorbent assay and Western blotting analysis. Expression of the muscle regeneration biomarkers, myoblast determination(Myo D), myogenic factor 5(Myf5), and smooth muscle actin(α-SMA), in the biceps femoris muscle was investigated at the protein and m RNA level by Western blotting and real time-PCR, respectively. Histological analysis was performed using hematoxylin and eosin staining. Finally, myosin heavy chain production was investigated in differentiated C2C12 myoblasts in the presence of GBH.RESULTS: GBH treatment markedly improved locomotion and running behavior. GBH significantly inhibited the secretion of monocyte chemoattractant protein-1 into the bloodstream in muscle-contused animals. The levels of Myo D, Myf5, and α-SMA protein and m RNA were significantly up-regulated by GBH in injured muscle tissue. Histological studies suggested that GBH facilitated recovery from muscle damage. However, GBH did not induce the production of myosin heavy chain in vitro.CONCLUSION: Overall, the present study suggested that GBH improves the recovery of the injured muscles in the mouse model of muscle contusion.
