title: Role of growth factors in the regulation of skeletal muscle mass, fibre type and fibre size

authors: Martyn, Julie Anne Kerr

abstract: Growth factors play an important role in the regulation of the growth and development of many tissues including muscle. Muscle is derived from somitic cells that differentiate to form myoblasts. These myoblasts then fuse to form myofibres. As these myofibres mature they take on the morphological, biochemical and physiological properties of mature muscle fibres. It is the plasticity of muscle fibres with respect to these properties that gives skeletal muscle its remarkable ability to adapt to different conditions of loading and usage. The aim of this thesis was to determine the role of growth factors in regulating muscle fibre type and fibre size in two different models of altered muscle mass.

Bovine muscle development was examined in normal foetuses (NM) and in double-muscled Belgian Blue foetuses (DM) carrying a mutation in the gene for myostatin. Muscle fibre type and fibre size were characterised using semi-automated computer-assisted image analysis after myofibrillar ATPase histochemical staining. Plasma IGF levels were measured, IGF-1 mRNA was measured using in situ hybridisation and IGF receptor binding was analysed by receptor autoradiography. Myostatin mRNA was analysed using semi-quantitative RT-PCR.

Skeletal muscle hypertrophy and atrophy were induced m adult rabbits by immobilisation of the hind limb with muscles in a lengthened and shortened position respectively. Fibre type and fibre size changes were analysed as described above and myostatin mRNA expression levels were measured using Northern blot analysis and semi-quantitative RT-PCR.

Results showed there was a biphasic change in the percentage of type 1 muscle fibres in both NM and DM, with the percentage initially declining, then increasing again. The percentage of type 1 fibres was consistently lower in DM. Circulating levels of IGF-1 increased with gestational age and levels of IGF-1 and IGF-2 were not different between NM and DM. IGF-1 mRNA expression in skeletal muscle declined with age in a pattern that was unrelated to changes in fibre type. Levels of IGF-1 mRNA were lower in DM during late gestation and this decrease was temporally linked to a reduced area of type 1 fibres in DM at this stage of development. Myostatin mRNA declined with gestational age, and levels of the mutant mRNA were higher in DM.

In induced hypertrophy, myostatin mRNA expression was lower than in controls and in atrophy, myostatin mRNA was elevated at 2 days but not at 6 days. There were no changes in fibre type or fibre size over the treatment period. Myostatin protein was evenly localised to all muscle fibre types.

In conclusion, IGF-1 appears to be unrelated to changes in fibre type during bovine muscle development, but may play a role in the regulation of fibre size. The DM condition is associated with a change in fibre type, but the role of myostatin in mediating this change remains unknown. Changes in myostatin mRNA in response to induced hypertrophy and atrophy strongly suggest an important role for myostatin as a negative regulator of skeletal muscle mass in postnatal animals.

text: Role of growth factors in the regulation of skeletal muscle mass, fibre type and fibre size Growth factors play an important role in the regulation of the growth and development of many tissues including muscle. Muscle is derived from somitic cells that differentiate to form myoblasts. These myoblasts then fuse to form myofibres. As these myofibres mature they take on the morphological, biochemical and physiological properties of mature muscle fibres. It is the plasticity of muscle fibres with respect to these properties that gives skeletal muscle its remarkable ability to adapt to different conditions of loading and usage. The aim of this thesis was to determine the role of growth factors in regulating muscle fibre type and fibre size in two different models of altered muscle mass.

Bovine muscle development was examined in normal foetuses (NM) and in double-muscled Belgian Blue foetuses (DM) carrying a mutation in the gene for myostatin. Muscle fibre type and fibre size were characterised using semi-automated computer-assisted image analysis after myofibrillar ATPase histochemical staining. Plasma IGF levels were measured, IGF-1 mRNA was measured using in situ hybridisation and IGF receptor binding was analysed by receptor autoradiography. Myostatin mRNA was analysed using semi-quantitative RT-PCR.

Skeletal muscle hypertrophy and atrophy were induced m adult rabbits by immobilisation of the hind limb with muscles in a lengthened and shortened position respectively. Fibre type and fibre size changes were analysed as described above and myostatin mRNA expression levels were measured using Northern blot analysis and semi-quantitative RT-PCR.

Results showed there was a biphasic change in the percentage of type 1 muscle fibres in both NM and DM, with the percentage initially declining, then increasing again. The percentage of type 1 fibres was consistently lower in DM. Circulating levels of IGF-1 increased with gestational age and levels of IGF-1 and IGF-2 were not different between NM and DM. IGF-1 mRNA expression in skeletal muscle declined with age in a pattern that was unrelated to changes in fibre type. Levels of IGF-1 mRNA were lower in DM during late gestation and this decrease was temporally linked to a reduced area of type 1 fibres in DM at this stage of development. Myostatin mRNA declined with gestational age, and levels of the mutant mRNA were higher in DM.

In induced hypertrophy, myostatin mRNA expression was lower than in controls and in atrophy, myostatin mRNA was elevated at 2 days but not at 6 days. There were no changes in fibre type or fibre size over the treatment period. Myostatin protein was evenly localised to all muscle fibre types.

In conclusion, IGF-1 appears to be unrelated to changes in fibre type during bovine muscle development, but may play a role in the regulation of fibre size. The DM condition is associated with a change in fibre type, but the role of myostatin in mediating this change remains unknown. Changes in myostatin mRNA in response to induced hypertrophy and atrophy strongly suggest an important role for myostatin as a negative regulator of skeletal muscle mass in postnatal animals.

year: 1999

title: The year after: sustaining the effects of literacy professional development in New Zealand secondary schools

authors: Whitehead, David

subjects: literacy practices

abstract: This paper describes some key characteristics of two secondary schools that were able to sustain benefits accrued from funded, externally facilitated literacy professional development intervention, during the year after facilitation had ceased. Characteristics that, together, signal a school with the potential to sustain gains in understanding about learning and literacy included the role of key people in building a critical mass community of practice, the theoretical and learner-focused nature of professional learning sessions built around an inquiry cycle, the use of intellectual resources focused more broadly on learning rather than literacy and the use of literacy data to facilitate evidence-based teaching.

text: The year after: sustaining the effects of literacy professional development in New Zealand secondary schools This paper describes some key characteristics of two secondary schools that were able to sustain benefits accrued from funded, externally facilitated literacy professional development intervention, during the year after facilitation had ceased. Characteristics that, together, signal a school with the potential to sustain gains in understanding about learning and literacy included the role of key people in building a critical mass community of practice, the theoretical and learner-focused nature of professional learning sessions built around an inquiry cycle, the use of intellectual resources focused more broadly on learning rather than literacy and the use of literacy data to facilitate evidence-based teaching.

year: 2010

title: He Piko, He Repo: Environmental DNA as a biomonitoring tool for wetland restoration

authors: Bird, Starsha

subjects: colonisation

abstract: Wetlands are biodiversity hotspots, harbouring distinctive species while providing numerous ecosystem services. Unfortunately, throughout human civilisation, they have been systematically degraded, resulting in a dramatic global loss in wetland extent, the establishment of invasive species and the extinction of many natives and, in Aotearoa New Zealand, impediment of the special connection Māori share with their repo (wetlands). Monitoring wetland biodiversity to take appropriate restorative action and minimise further degradation is imperative, particularly in the face of climate change. Environmental DNA (eDNA; genetic material released by organisms into the environment) is an increasingly popular non-invasive biomonitoring method that provides information about the presence or potential absence of species inhabiting a given ecosystem, but has yet to be optimised for wetland environments. My thesis aimed to address the consequences of colonisation on the relationship between Māori and their wetlands, and to explore the value eDNA holds for wetland biomonitoring. The first analysis (Chapter 2) delved into the past (post-European settlement; 1840s), the present (2023), and the future (2073) of Opuatia Wetland (Waikato, Aotearoa New Zealand) through a series of interviews with three individuals who whakapapa to (are connected with) Opuatia Wetland and its surroundings. I found that Opuatia Wetland once flourished with native biodiversity, and kai (food) was readily available. However, land confiscation by European settlers marked the downfall of this once productive and connected catchment such that it is presently in a fairly degraded state, dominated by exotic species, with Māori feeling disconnected from their whenua (land). Considering the future, the interviewees envisioned a revived abundance of native species and desire for restoration efforts to be facilitated by both mātauranga Māori (Māori knowledge) and science. The second analysis (Chapter 3) examined a publicly available eDNA database to retrieve data from 26 wetland sites across Aotearoa New Zealand to understand taxonomic diversity patterns. Here, I showed that DNA sequence composition varied across the motu (country), illustrating that varying environmental conditions among wetland types influences biodiversity. Notably, most sites were dominated by >50% exotic species. These findings highlight the value of publicly shared data for generating new insights, and the need for resource allocations to tackle the persistence of exotic species in wetland ecosystems. The third analysis (Chapter 4) focused on analysing biodiversity dynamics at a single wetland and tested various eDNA sampling techniques. I sampled four spatially distant sites across Opuatia Wetland at three time points during an austral spring and simultaneously obtained data from conventional taxonomic surveys, tested three different filter sizes, and assessed DNA degradation rates using foreign DNA. I found significant differences in biodiversity across time and space, and when using different eDNA collection filters. Foreign DNA persisted for up to one week post-release within a 10 m radius, and combining conventional and eDNA methods provided a more comprehensive overview of biodiversity patterns. These findings collectively showcased key changes in biodiversity, even over short spatial and temporal scales, and identified parameters to consider for wetland biodiversity monitoring and associated data interpretation. It is only a matter of time until the impacts of climate change and invasive species take firmer hold of Aotearoa New Zealand's distinctive and fragile environment. Undoubtedly, eDNA can be part of the solution to mitigate these challenges, owing to its versatility across ecosystem types, capacity to extend to various substrates, and accessibility for researchers and citizen scientists alike. As eDNA methods continue to improve and become integrated into ongoing biomonitoring schemes, building equitable and empowering relationships between Māori and researchers/government bodies will best ensure that Aotearoa New Zealand repo and other environments are maintained and conserved for future generations.

text: He Piko, He Repo: Environmental DNA as a biomonitoring tool for wetland restoration Wetlands are biodiversity hotspots, harbouring distinctive species while providing numerous ecosystem services. Unfortunately, throughout human civilisation, they have been systematically degraded, resulting in a dramatic global loss in wetland extent, the establishment of invasive species and the extinction of many natives and, in Aotearoa New Zealand, impediment of the special connection Māori share with their repo (wetlands). Monitoring wetland biodiversity to take appropriate restorative action and minimise further degradation is imperative, particularly in the face of climate change. Environmental DNA (eDNA; genetic material released by organisms into the environment) is an increasingly popular non-invasive biomonitoring method that provides information about the presence or potential absence of species inhabiting a given ecosystem, but has yet to be optimised for wetland environments. My thesis aimed to address the consequences of colonisation on the relationship between Māori and their wetlands, and to explore the value eDNA holds for wetland biomonitoring. The first analysis (Chapter 2) delved into the past (post-European settlement; 1840s), the present (2023), and the future (2073) of Opuatia Wetland (Waikato, Aotearoa New Zealand) through a series of interviews with three individuals who whakapapa to (are connected with) Opuatia Wetland and its surroundings. I found that Opuatia Wetland once flourished with native biodiversity, and kai (food) was readily available. However, land confiscation by European settlers marked the downfall of this once productive and connected catchment such that it is presently in a fairly degraded state, dominated by exotic species, with Māori feeling disconnected from their whenua (land). Considering the future, the interviewees envisioned a revived abundance of native species and desire for restoration efforts to be facilitated by both mātauranga Māori (Māori knowledge) and science. The second analysis (Chapter 3) examined a publicly available eDNA database to retrieve data from 26 wetland sites across Aotearoa New Zealand to understand taxonomic diversity patterns. Here, I showed that DNA sequence composition varied across the motu (country), illustrating that varying environmental conditions among wetland types influences biodiversity. Notably, most sites were dominated by >50% exotic species. These findings highlight the value of publicly shared data for generating new insights, and the need for resource allocations to tackle the persistence of exotic species in wetland ecosystems. The third analysis (Chapter 4) focused on analysing biodiversity dynamics at a single wetland and tested various eDNA sampling techniques. I sampled four spatially distant sites across Opuatia Wetland at three time points during an austral spring and simultaneously obtained data from conventional taxonomic surveys, tested three different filter sizes, and assessed DNA degradation rates using foreign DNA. I found significant differences in biodiversity across time and space, and when using different eDNA collection filters. Foreign DNA persisted for up to one week post-release within a 10 m radius, and combining conventional and eDNA methods provided a more comprehensive overview of biodiversity patterns. These findings collectively showcased key changes in biodiversity, even over short spatial and temporal scales, and identified parameters to consider for wetland biodiversity monitoring and associated data interpretation. It is only a matter of time until the impacts of climate change and invasive species take firmer hold of Aotearoa New Zealand's distinctive and fragile environment. Undoubtedly, eDNA can be part of the solution to mitigate these challenges, owing to its versatility across ecosystem types, capacity to extend to various substrates, and accessibility for researchers and citizen scientists alike. As eDNA methods continue to improve and become integrated into ongoing biomonitoring schemes, building equitable and empowering relationships between Māori and researchers/government bodies will best ensure that Aotearoa New Zealand repo and other environments are maintained and conserved for future generations.

year: 2023