An investigation of IGF-I isoforms in human skeletal muscle. The effects of age and exercise.

This thesis has focussed on the expression of the different insulin like growth factor-I (IGF-I) isoforms (IGF-IEa, IGF-IEb and MGF) in human skeletal muscle, with specific reference to exercise and the ageing process. To enable this, a method of real time quantitative PCR was developed for the accurate detection and quantification of these splice variants. Studies were performed both in our laboratory and at the Copenhagen Muscle Research Centre, Denmark. The acute response of IGF-IEa and MGF to a single bout of high resistance exercise was studied in young (n 8, age 29.5 1.5 years) and old (n 7, age 74.4 1.8 years) subjects. At rest there were no significant differences between resting levels of either isoform between the young and old subjects. The study showed that when measured 2.5 hours after the end of the exercise bout, the mechano-sensitive isoform, MGF, was significantly upregulated in the young but not the old subjects (P 0.05). No change was observed after exercise in the IGF-IEa iso form in either age group suggesting that the isoforms were differentially regulated. Furthermore, in situ hybridisation using oligo probes specific for MGF mRNA confirmed its localisation within the muscle fibres. No significant correlation was observed between the change in MGF expression and the muscle myosin heavy chain isoform composition was observed. A second study in young (n 10, age 20-27 years) and old subjects (n 10, age 67-75 years) evaluated the effects of a single bout of prolonged (1 hour) eccentric muscle damaging cycling exercise. In this study, and in contrary to the weightlifting study, a significant increase was observed in MGF mRNA levels in both the young and old subjects after the end of the exercise challenge (P 0.05), whereas levels of IGF-IEa again showed no significant increase. Thus suggesting that older muscle might require some myofibrillar disruption and/or sarcolemmal damage caused by eccentric exercise as a prerequisite for stimulating MGF expression. The effects of a period of resistance training in older people (n 19, age 70-82 years) performed over 12 weeks revealed a significant increase in the mRNA of both MGF and IGF-IEa and also the third isoform detected, IGF-IEb (P 0.05). In addition, the role that growth hormone (GH) might play in regulating the IGF-I spice variants in muscle with training was also investigated in this study. Recombinant GH administered to subjects without exercise, favoured expression of the IGF-IEa isoform. When resistance training was combined with GH administration, enhanced levels of MGF were observed but not IGF-IEa. This suggests that GH administration may lead to an overall upregulation of the primary IGF-I transcript, which is spliced towards MGF when mechanical activity in the form of resistance training is imposed. The results of the work in this thesis have shown that it possible to detect and quantify IGF-I mRNA transcripts in human skeletal muscle, even those expressed at low levels. It has also shown that there may be differential regulation of the different isoforms in young and old muscle in response to exercise, which would support the idea that these isoforms of IGF-I have different physiological actions.