宰后牦牛肉骨骼肌细胞凋亡过程研究

An Investigation of Skeletal Muscle Apoptosis during Postmortem Aging of Yak Meat

为研究宰后牦牛肉在成熟过程中前驱肱三头肌(triceps brachi,TB)、中部背最长肌(musculus longissimus,ML)、后驱半膜肌(semimembranosus,SM)中骨骼肌细胞凋亡过程的发生,选取24头甘南黑牦牛按照伊斯兰屠宰方式进行屠宰,于0～4℃、风速0.5 m/s条件下成熟,在宰后成熟过程中的不同时间点(1、3、5、7 d)进行细胞核的He染色观察、原位末端法检测骨骼肌细胞凋亡率以及caspase-3活力测定。宰后1 d,TB、ML和SM的骨骼肌细胞核结构完整,并且轮廓清晰,核质均匀分布;随着成熟时间的延长核染色质凝聚在核膜下呈月牙状,细胞核浓缩破裂形成凋亡小体,细胞体积皱缩变小。在宰后前3 d的ML和SM中几乎检测不到阳性细胞核;宰后5 d后TB和SM中阳性细胞核显著增多(P<0.01);宰后7 d,TB和ML中阳性细胞核数量差异极显著(P<0.01);宰后12 h caspase-3活力显著高于宰后0.5 h的活力(TB,P<0.01;ML,P<0.001;SM,P<0.01);宰后12 h ML的caspase-3达到最高活力,较宰后0.5 h酶活力升高273.01%;TB和SM在宰后1 d达到最高活力,较宰后0.5 h酶活力分别升高了273.93%和386.17%;宰后5 d TB、ML和SM的caspase-3活力下降到最小值。结果表明:宰后成熟过程中牦牛肉的骨骼肌细胞核浓缩破裂形成凋亡小体和caspase-3活力的激活,同时阳性细胞核的检出并且数量随成熟时间增大,这一切都说明宰后牦牛肉骨骼肌细胞中发生了细胞凋亡,为宰后牦牛肉成熟机制提供一个更加合理、系统的解释途径。

The purpose of this study was to shed light on the occurrence of skeletal muscle apoptosis in triceps brachi （TB）, musculus longissimus （ML）, semimembranosus （SM） muscles from yak meat during postmortem aging. Totally 24 Ganan black yaks were selected and slaughtered in the Islamic way, and different muscles were excised from each carcass and aged under the conditions of 0-4℃and 0.5 m/s for air lfow rate. Morphological observation of the cell nucleus was performed using He staining at different time points during the aging process, and apoptotic index was measured by in situ end-labeling. Meanwhile, caspase-3 activity was assayed. At 1 d postmortem, the structure of the cell nucleus in the skeletal muscles TB, ML and SM was complete and apparent with evenly distributed nucleoplasm. As the aging progressed, the chromatin inside the nucleus was agglomerated to form a crescent-like shape, the cell nucleus was concentrated and broken, forming apoptotic bodies, and as a result, the cells were shrunk. Almost no positive cell nuclei were observed in ML and SM muscles within the ifrst 3 d premortem. At 5 d postmortem, TB and SM muscles exhibited signiifcantly increased counts of TUNEL-positive nuclei （P〈.01）, and at 7 d, a signiifcant difference in the count TUNEL-positive nuclei was noticed between TB and ML muscles （P〈.01）. The caspase-3 activity was signiifcantly higher at 12 h postmortem than at 0.5 h （TB, P〈.01;ML, P〈.001;SM, P〈.01）. In ML muscle, this enzyme activity reached the maximum level at 12 h postmortem, showing a 273.01%increase over that observed at 0.5 h. In contrast, the enzyme activity reached the maximum level at 1 h postmortem, which was higher by 273.93%and 386.17%than at 0.5 h, respectively. At 5 h, the minimum levels of caspase-3 activity were detected in all the three muscles. In conclusion, during postmortem aging, the concentration and breaking of the cell nuclei in yak skeletal muscles to form apoptotic bodies, increased levels of caspase-3 activity, and the detectable presence of TUNEL-positive nuclei, positively dependent on aging time, together conifrm the occurrence of skeletal muscle apoptosis, which offers a systematic and reasonable way to interpret the mechanism of postmortem aging of yak meat.