The effects of dihydrotestosterone on amino acid transport in ageing mammalian skeletal muscle fibres

衰老导致骨骼肌质量和力量逐渐下降，通常被称为肌肉减少症。骨骼肌减少症主要发生在80岁至80岁的人群中（以及12岁至12岁的啮齿动物中），其后果包括缺乏活动、易跌倒并最终丧失独立性。由于英国80岁以上的人口数量预计在未来20年内将翻一番，从2006年的270万增加到2031年的540万，因此肌肉减少症患者的比例以及这对NHS和整个社会的需求预计将相应增加。尽管它在生理和社会经济方面具有重要意义，但人们对肌肉减少症的病因知之甚少。在这项研究中，我们提出它的发生是因为骨骼肌细胞将氨基酸（蛋白质的组成部分）运输进出细胞的能力下降。我们还认为，这种下降是由于运送氨基酸进出细胞的转运体/泵的数量减少，或者是由于通常刺激这些泵的激素（如男性性激素）浓度的降低。为了验证这些建议，我们计划调查；(1)年龄对小鼠骨骼肌细胞氨基酸转运蛋白数量及其进出氨基酸能力的影响。(2)用睾酮的活性代谢物双氢睾酮（DHT）治疗肌肉细胞是否能逆转这些年龄依赖性变化。(3)最后，我们将测试DHT治疗bb0 ~ 560日龄小鼠4周是否能逆转/减缓肌肉减少症。我们还将研究DHT处理是否会增加转运氨基酸和新蛋白质的合成。本研究将使用三种类型的实验。第一种类型将使用在正常实验室条件下饲养的不同年龄（60-730天）的小鼠。然后，从每个年龄组的edl和比目鱼肌中分离出的小肌纤维束将用于研究年龄对各种氨基酸转运蛋白表达和生理功能的影响，以及这些变化背后的细胞和分子机制。以往的研究大多研究了睾酮治疗对骨骼肌质量和力量的影响，没有研究研究DHT治疗的影响。因此，第二个实验将研究DHT治疗年轻、年老和衰老小鼠至少4周的效果。然后，从治疗和未治疗小鼠的edl和比目鱼肌中分离出的小肌纤维束将用于研究DHT治疗对骨骼肌质量、力量、各种氨基酸转运蛋白的表达和功能、异亮氨酸（Ile）和MeAIB的摄取以及Ile进入蛋白质的影响。在第三个实验中，将研究DHT处理对衰老小鼠edl和比目鱼肌分离的肌纤维束的力产生、氨基酸运输和蛋白质掺入的急性影响。这些效应背后的细胞和分子机制也将被研究。大多数实验将使用CD1小鼠进行，因为它们价格低廉，与正常小鼠和人类种群具有相似的遗传变异，并且在24个月后患有严重的肌肉减少症。这些实验的结果将为氨基酸运输的下降和合成代谢类固醇生物利用度的降低在肌肉减少症中的作用提供重要的信息。它们还将增加我们对骨骼肌随着年龄增长而发生的变化以及这些变化背后的机制的理解。他们还将提供关于DHT治疗是否可以延迟/减缓肌肉减少症的发展及其作用机制的信息。这些发现将对未来治疗肌肉减少症的新方法的发展具有重要意义。

Ageing leads to a gradual decline in skeletal muscle mass and strength, commonly referred to as sarcopenia. Sarcopenia occurs mostly in people >80yrs (and rodents >2 years old) and its consequences include inactivity, increased susceptibility to falls and eventually to loss of independence. As the number of people over 80yrs in the United Kingdom is expected to double in the next 20 years, from 2.7 million in 2006 to 5.4 million by 2031, the percentage of the population suffering from sarcopenia and the demands this imposes on the NHS and society in general is expected to rise accordingly. Despite its physical and socioeconomic importance, little is known about the causes of sarcopenia. In this study we propose that it occurs because of a decrease in the ability of skeletal muscle cells to transport amino acids (the building blocks of proteins ) into and out of the cells. We also suggest that this decline is due to a reduction in the number of the transporters/pumps that move amino acids into and out of the cells or is due to a decrease in the concentration of hormones such as the male sex hormones that normally stimulate these pumps.To test these suggestions we plan to investigate; (1) the effects of age on the number of amino acid transporters and their ability to move amino acids into and out of mouse skeletal muscle cells. (2) Whether treating muscle cells with dihydrotestosterone (DHT), the active metabolite of testosterone, can reverse these age-dependent changes. (3) Finally we will test whether treating mice >560 days old with DHT for 4 weeks reverses/slows down sarcopenia. We will also investigate whether DHT treatment increases the transport amino acids and the synthesis of new proteins.Three types of experiments will be used in the study. The first type will use mice of different ages (range 60-730 days) kept under normal laboratory conditions. Small muscle fibre bundles isolated from the edl and soleus muscles of each age group will then be used to investigate the effects of age on the expression and physiological function of various amino acid transporters and the cellular and molecular mechanisms underlying these changes.Most previous studies have investigated the effects of testosterone treatment on skeletal muscle mass and strength and no studies have investigated those of DHT treatment. Therefore, the second experiment will investigate the effects of treating young, old and senescent mice with DHT for at least 4 weeks. Small muscle fibre bundles isolated from the edl and soleus of treated and untreated mice will then be used to investigate the effects of DHT treatment on skeletal muscle mass, strength, the expression and function of various amino acid transporters, the uptake of isoleucine (Ile) and MeAIB and the incorporation of Ile into proteins.In the third experiment, the acute effects of DHT treatment on force production, amino acid transport and incorporation into proteins in muscle fibre bundles isolated from the edl and soleus of senescent mice will be investigated. The cellular and molecular mechanisms underlying these effects will also be investigated.Most of these experiments will be performed using CD1 mice because they are inexpensive, have similar genetic variations as normal mouse and human populations and suffer from severe sarcopenia from 24 months onwards.The results from these experiments will provide important information on the role a decline in amino acid transport and reduced bioavailability of anabolic steroids, play in sarcopenia. They will also increase our understanding of the changes that occur in skeletal muscle with ageing and the mechanisms underlying these changes. They will also provide information on whether DHT treatment can delay/slow the development of sarcopenia as well as the mechanisms underlying its effects. These findings will be important in the development of new ways of managing sarcopenia in the future.