慢性睡眠剥夺致大鼠咬肌肌球重链蛋白异构型改建

目的：观察慢性睡眠剥夺（ CSD）因素作用下咬肌肌纤维肌球重链蛋白（ MyHC）异构型的变化，探讨慢性睡眠剥夺引起颞下颌关节紊乱病（ TMD）的致病机制。方法180只雄性Wistar大鼠随机分为3组：慢性睡眠剥夺组（ CSD）、正常笼养组（ CC）、大平台对照组（ TC），每组60只。每组根据实验时间不同分别分为3个亚组：7 d、14 d、21 d组，每个亚组20只。建立大鼠慢性睡眠剥夺模型。通过免疫组织化学DAB法检测咬肌肌纤维MyHC异构型构成比；应用Westem Blot技术和实时荧光定量PCR定量检测咬肌组织MyHC异构型蛋白和mRNA的表达水平。结果 CSD 7 d组MyHC-Ⅰ型咬肌深浅部肌纤维（0．314±0．005，0．134±0．005），MyHC-ⅡA型咬肌深浅部肌纤维（7．960±0．465，7．090±0．564），MyHC-ⅡB型咬肌深浅部肌纤维（2．840±0．054，2．580±0．054）的表达与对照组相比均差异有统计学意义（均P＜0．05）。 CSD 14 d组咬肌深浅部MyHC-Ⅰ型肌纤维（0．284±0．005，0．106±0．015）与CSD14d组MyHC-ⅡA型咬肌深浅部肌纤维（7．030±1．045，6．050±0．976），MyHC-ⅡB型深浅部肌纤维（3．680±0．548，3．850±0．457）的表达与对照组相比均差异有统计学意义（均P＜0．05）。 CSD 7 d组大鼠咬肌MyHC-Ⅱ型肌纤维明显减少，而MyHC-Ⅰ型增加，随着CSD时间的继续延长，大鼠咬肌MyHC-Ⅱ型肌纤维数量会有所增加，MyHC-Ⅰ型数量则减少。 CSD 21 d组咬肌深部MyHC-Ⅱ型和MyHC-Ⅰ型肌纤维数量与正常水平仍差异有统计学意义（P＜0．05），CSD 21 d组咬肌浅部两种肌纤维数量与正常水平差异无统计学意义（P＞0．05）。结论睡眠剥夺可使咬肌肌纤维MyHC表型发生改变，肌纤维MyHC异构型的改变可能是睡眠剥夺致TMD中咀嚼肌功能紊乱的发病机制之一。

烟酰胺对牛骨骼肌卫星细胞增殖与分化的影响

为了探究烟酰胺(NAM)对牛骨骼肌卫星细胞(BSMSCs)增殖与分化的影响,试验以体外培养的BSMSCs成肌分化模型为研究对象,用0,1,2,4,8 mmol/L烟酰胺处理BSMSCs(对照组及1,2,4,8 mmol/L烟酰胺组),采用CCK-8法筛选烟酰胺对BSMSCs作用的最适浓度,后续试验均以不添加烟酰胺作为对照组,最适浓度作为试验组,采用EdU试剂盒检测最适浓度烟酰胺对BSMSCs增殖的影响;光学显微镜下观察最适浓度烟酰胺处理的BSMSCs分化24,48,72小时时的肌管形态,采用Western-blot检测BSMSCs中肌球重链蛋白(MyHC)和肌原蛋白(MyOG)的表达水平,分析烟酰胺对BSMSCs分化的影响。结果表明:与对照组比较,BSMSCs的OD450值随着烟酰胺浓度的上升而下降,4 mmol/L烟酰胺组OD450值显著降低(P<0.05),8 mmol/L烟酰胺组OD450值极显著降低(P<0.01),但8 mmol/L烟酰胺组的细胞存活率低于50%,超过药物对细胞的半数致死浓度,最终以4 mmol/L作为烟酰胺作用于BSMSCs的最适浓度。4 mmol/L烟酰胺处理BSMSCs后,增殖率没有发生显著变化(P>0.05)。与对照组比较,4 mmol/L烟酰胺组BSMSCs融合形成的肌管数量增多且肌管直径变大,MyHC和MyOG蛋白的相对表达量显著或极显著上升(P<0.05或P<0.01)。说明烟酰胺对BSMSCs增殖没有影响但可促进其分化。

Molecular cloning and mRNA expression analysis of myosin heavy chain(MyHC)from fast skeletal muscle of grass carp,Ctenopharyngodon idella

The myosin heavy chain(MyHC)is one of the major structural and contracting proteins of muscle.We have isolated the cDNA clone encoding MyHC of the grass carp,Ctenopharyngodon idella. The sequence comprises 5 934 bp,including a 5 814 bp open reading frame encoding an amino acid sequence of 1 937 residues.The deduced amino acid sequence showed 69%homology to rabbit fast skeletal MyHC and 73%–76%homology to the MyHCs from the mandarin fish,walleye pollack,white croaker,chum salmon,and carp.The putative sequences of subfragment-1 and the light meromyosin region showed 61.4%–80%homology to the corresponding regions of other fish MyHCs.The tissue-specific and developmental stage-specific expressions of the MyHC gene were analyzed by quantitative real-time PCR.The MyHC gene showed the highest expression in the muscles compared with the kidney,spleen and intestine.Developmentally,there was a gradual increase in MyHC mRNA expression from the neural formation stage to the tail bud stage.The highest expression was detected in hatching larva.Our work on the MyHC gene from the grass carp has provided useful information for fish molecular biology and fish genomics.

Globular adiponectin induces differentiation and fusion of skeletal muscle cells

The growing interest in skeletal muscle regeneration is associated with the opening of new therapeutic strategies for muscle injury after trauma, as well as several muscular degenerative pathologies, including dystrophies, muscu- lar atrophy, and cachexia. Studies focused on the ability of extracellular factors to promote myogenesis are therefore highly promising. We now report that an adipocyte-derived factor, globular adiponectin （gAd）, is able to induce mus- cle gene expression and cell differentiation, gAd, besides its well-known ability to regulate several metabolic func- tions in muscle, including glucose uptake and consumption and fatty acid catabolism, is able to block cell cycle entry of myoblasts, to induce the expression of specific skeletal muscle markers such as myosin heavy chain or caveolin-3, as well as to provoke cell fusion into multinucleated syncytia and, finally, muscle fibre formation, gAd exerts its pro- differentiative activity through redox-dependent activation of p38, Akt and 5＇-AMP-activated protein kinase path- ways. Interestingly, differentiating myoblasts are autocrine for adiponectin, and the mimicking of pro-inflammatory settings or exposure to oxidative stress strongly increases the production of the hormone from differentiating cells. These data suggest a novel function of adiponectin, directly coordinating the myogenic differentiation program and serving an autocrine function during skeletal myogenesis.

AMPK Subunit Expression Regulates Intramuscular Fat Content and Muscle Fiber Type in Chickens

The objective of this study was to assess the role of AMPK in intramuscular fat(IMF) and fiber type in chicken muscle. The chickens were slaughtered and their muscles were collected at the ages of 4, 8, and 16 weeks so as to determine the IMF contents, as well as the expression levels of AMPK subunits, regulators of adipogenesis. In addition, the myosin heavy chains(My HCs) in thigh muscle tissues were also measured. The results showed that the IMF contents in 16-week old chickens were higher than those in 4 and 8-week-old chickens(P<0.05).The expression levels of fatty acid synthase(FAS) and fatty aicd translocase CD36(FAT/CD36) m RNA were increased significantly in samples collected at the ages of4 and 16 weeks(P<0.05). The expression levels of My HC IIa and IIb differed significantly among all the developmental stages(P <0.05). The AMPKα2, AMPKγ1,and AMPKγ3 m RNA levels were dramatically decreased with the increase of age(P <0.05). To examine the role of AMPK in adipogenesis regulation, the SV cells were cultured in an adipogenesis medium and treated with AICAR and Compound C respectively, the specific activator and inhibit of AMPK. The Compound C induced dramatically a greater expression of C/EBPβ, SREBP1 and PPARγ(P <0.05). In conclusion, the expression of AMPKα2, AMPKγ1, and AMPKγ3 m RNA is significantly correlated with the adipogenesis in skeletal muscle of chickens.

Effects and mechanism of different adrenergic receptor antagonists on left ventricular hypertrophy subsequent to coarctation of abdominal aorta in rats

Objective: To study the changes of a collagen-binding protein (Colligin) and myosin heavy chain isoform (α/β-MHC) gene and protein in left ventricular hypertrophy subsequent to coarctation of abdominal aorta in rats and the effects of three kinds of adrenergic receptor blockers: Carvedilol (CAR) , Metoprolol (MET) and Terazosin (TER) on these changes, and to elucidate the effects and new mechanism of CAR on left ventricular hypertrophy regression. Methods: A model of hypertrophy induced by coarctation of abdominal aorta(CAA)was used in this study. Thirty two male wistar rats were divided randomly into four groups 4 weeks after CAA operation: CAA, CAR, MET and TER. Hemodynamics, ventricular remodeling parameters, expressions of Colligin and α/β-MHC mRNA, protein expressions of Collagen Ⅰ / Ⅲ and Colligin were investigated in the four groups and sham operation group. Results: Left ventricle hypertrophy was observed clearly 16 weeks after operation. The ratio of α/β-MHC mRNA decreased, while expressions of Collagen Ⅰ /Ⅲ proteins and Colligin mRNA/protein increased( P < 0.05). CAR could ameliorate left ventricle hypertrophy prior to MET and TER. CAR could also change the expressions of α/β-MHC, Collagen Ⅰ /Ⅲ and Colligin in both gene and protein levels ( P < 0.05), while MET and TER have no effect on them ( P > 0.05). Conclusion: The effects of CAR on extracellular matrix proteins and MHC isoform shift regression of left ventricle may be due to antiproliferative or antioxidative mechanism, which was independent of beta-adrenergic receptor antagonist.