title: Inhibition of ERK and GSK3 signalling pathways promotes development and expression of pluripotent markers in bovine blastocysts

authors: Harris, Daina

subjects: bovine

abstract: Embryo-derived pluripotent stem cells (ePSCs) naturally derive from different founder tissues, namely the early and late epiblast, that vary in their cellular characteristics. Recent evidence in mice suggests these sources of starting material correspond with two states of pluripotency referred to as ‘naïve’ and ‘primed’, respectively. Pluripotency is defined as the ability of a cell to give rise to any foetal or subsequent adult cell type. Naïve and primed ePSCs have different fundamental properties and only naïve cells can give rise to all cell types of an animal, including functional gametes (Nichols & Smith 2009). Until recently, naïve ePSCs (also known as embryonic stem cell or ESCs) were only available in mouse. After almost 30 years, the first non-mouse ESCs were derived in rat using a combination of specific inhibitors that block the FGF-ERK differentiation pathway and simultaneously maintain self-renewal through inhibition of GSK3 (Buehr et al. 2008; Ying et al. 2008). The three inhibitor combination, or 3i, included inhibitors of the FGF receptor (PD173074), downstream ERK activity (PD0325901) and the GSK3 pathway (CHIR99021). The dual inhibition of PD0325901 and CHIR99021, termed 2i, was found to be effective for ESC culture; therefore the requirement for PD173074 became dispensable. If pluripotent mechanisms are conserved between species, we hypothesise that these inhibitors would aid the derivation of naïve ESCs in livestock species i.e. cattle.

The aim of this study was to assess the effects of these inhibitors on the developing bovine embryo. We hypothesised that the use of the inhibitors in culture medium would enhance the pluripotent characteristics of blastocysts for subsequent ESC culture. To test this idea we cultured in vitro fertilised bovine embryos in the presence of the inhibitors to initially assess the effect on embryo development. To assess embryo quality, blastocysts were analysed to evaluate the total number of cell nuclei. Additionally, mRNA and protein analyses were undertaken to characterise the expression of core pluripotent genes and assess if the inhibitors had an effect on their expression.

We found that culturing bovine embryos in either 3i or 2i did not influence the total number of embryos that progressed to the blastocyst stage. A significant decrease was observed in the proportion of early vs. late stage blastocysts in 2i, which suggested acceleration in the rate of development. In 2i medium, the number of cell nuclei in D7 blastocysts was significantly increased in both the inner cell mass (ICM) and the outer trophectoderm (TE). Additionally, embryos cultured in 2i showed a >2-fold increase in NANOG, a transcription factor at the core of the pluripotency network. The ICM-specific pluripotency marker SOX2 was also up-regulated in the presence of 2i. Both these genes were found to be significantly enriched in the ICM of bovine blastocysts. Overall, we conclude that bovine blastocysts differ from mouse in their response to 2i signal inhibition and molecular control of pluripotency.

text: Inhibition of ERK and GSK3 signalling pathways promotes development and expression of pluripotent markers in bovine blastocysts Embryo-derived pluripotent stem cells (ePSCs) naturally derive from different founder tissues, namely the early and late epiblast, that vary in their cellular characteristics. Recent evidence in mice suggests these sources of starting material correspond with two states of pluripotency referred to as ‘naïve’ and ‘primed’, respectively. Pluripotency is defined as the ability of a cell to give rise to any foetal or subsequent adult cell type. Naïve and primed ePSCs have different fundamental properties and only naïve cells can give rise to all cell types of an animal, including functional gametes (Nichols & Smith 2009). Until recently, naïve ePSCs (also known as embryonic stem cell or ESCs) were only available in mouse. After almost 30 years, the first non-mouse ESCs were derived in rat using a combination of specific inhibitors that block the FGF-ERK differentiation pathway and simultaneously maintain self-renewal through inhibition of GSK3 (Buehr et al. 2008; Ying et al. 2008). The three inhibitor combination, or 3i, included inhibitors of the FGF receptor (PD173074), downstream ERK activity (PD0325901) and the GSK3 pathway (CHIR99021). The dual inhibition of PD0325901 and CHIR99021, termed 2i, was found to be effective for ESC culture; therefore the requirement for PD173074 became dispensable. If pluripotent mechanisms are conserved between species, we hypothesise that these inhibitors would aid the derivation of naïve ESCs in livestock species i.e. cattle.

The aim of this study was to assess the effects of these inhibitors on the developing bovine embryo. We hypothesised that the use of the inhibitors in culture medium would enhance the pluripotent characteristics of blastocysts for subsequent ESC culture. To test this idea we cultured in vitro fertilised bovine embryos in the presence of the inhibitors to initially assess the effect on embryo development. To assess embryo quality, blastocysts were analysed to evaluate the total number of cell nuclei. Additionally, mRNA and protein analyses were undertaken to characterise the expression of core pluripotent genes and assess if the inhibitors had an effect on their expression.

We found that culturing bovine embryos in either 3i or 2i did not influence the total number of embryos that progressed to the blastocyst stage. A significant decrease was observed in the proportion of early vs. late stage blastocysts in 2i, which suggested acceleration in the rate of development. In 2i medium, the number of cell nuclei in D7 blastocysts was significantly increased in both the inner cell mass (ICM) and the outer trophectoderm (TE). Additionally, embryos cultured in 2i showed a >2-fold increase in NANOG, a transcription factor at the core of the pluripotency network. The ICM-specific pluripotency marker SOX2 was also up-regulated in the presence of 2i. Both these genes were found to be significantly enriched in the ICM of bovine blastocysts. Overall, we conclude that bovine blastocysts differ from mouse in their response to 2i signal inhibition and molecular control of pluripotency.

year: 2012

title: Making sense of leadership in early childhood education: Tensions and complexities between concepts and practices

authors: Klevering, Nicki

subjects: Leadership

abstract: Effective leadership within early childhood settings is aligned with the perceived successful implementation of high quality care and education programmes (Thornton, Tamati, Clarkin-Philips, Aitken & Wansbrough, 2009). With growing attention on the role early childhood education (ECE) plays in preparing children to be successful in their lives, it is not surprising that there is increased focus on the work and impact of educational leaders in this endeavour. An expanding body of research specifically exploring leadership within ECE settings illustrates how much of the educational leadership theory corpus lacks contextual relevance and fails to recognise the complexities and realities of leading in early years contexts (Rodd, 2013). The qualitative research reported on in this article examines the leadership understandings and perceptions of five qualified, registered early childhood leaders. The findings illustrated that whilst participants were cognisant of the role effective leadership plays in providing quality care and education, there was much confusion about what leadership entailed and how leadership differentiated from management in this context. The contextual complexities of the ECE sector were a significant influence on each participant’s opportunity to learn about, and practise leadership. Findings also revealed a need for contextually relevant and progressive approaches to leadership learning to support early childhood leaders and teachers in their leadership work.

text: Making sense of leadership in early childhood education: Tensions and complexities between concepts and practices Effective leadership within early childhood settings is aligned with the perceived successful implementation of high quality care and education programmes (Thornton, Tamati, Clarkin-Philips, Aitken & Wansbrough, 2009). With growing attention on the role early childhood education (ECE) plays in preparing children to be successful in their lives, it is not surprising that there is increased focus on the work and impact of educational leaders in this endeavour. An expanding body of research specifically exploring leadership within ECE settings illustrates how much of the educational leadership theory corpus lacks contextual relevance and fails to recognise the complexities and realities of leading in early years contexts (Rodd, 2013). The qualitative research reported on in this article examines the leadership understandings and perceptions of five qualified, registered early childhood leaders. The findings illustrated that whilst participants were cognisant of the role effective leadership plays in providing quality care and education, there was much confusion about what leadership entailed and how leadership differentiated from management in this context. The contextual complexities of the ECE sector were a significant influence on each participant’s opportunity to learn about, and practise leadership. Findings also revealed a need for contextually relevant and progressive approaches to leadership learning to support early childhood leaders and teachers in their leadership work.

year: 2019

title: Mouri Tu, Mouri Moko, Mouri Ora! Moko as a wellbeing strategy

authors: Penehira, Mera

subjects: Maori

abstract: This research has been undertaken within the Health Research Council funded International Collaborative Indigenous Health Research Partnership (ICIHRP) program, “The role of resiliency in responding to blood borne viral and sexually transmitted infections in Indigenous communities”. Some Indigenous communities in Australia, New Zealand and Canada have been shown to experience higher rates of blood borne viral infections. There are a number of categories of health intervention that have been shown to be effective in prevention and enhancing access to treatment for blood borne viral infections, including various forms of health promotion, enhanced diagnostic and treatment services, and harm reduction measures related to injecting drug use. The present research was undertaken with the understanding that development of effective responses of this kind among Indigenous people would benefit from a better understanding of the social and cultural factors that might protect against these infections and their consequences. It is argued that such factors are linked in various ways to Indigenous resistance and resiliency, which is described as the means by which people choose to make use of individual and community strengths to protect themselves against adverse health outcomes and enhance their health and wellbeing.

In this thesis I explore how people and their identity are affected when you are part of a marginalised or vulnerable population – namely, Māori women who have contracted the Hepatitis C Virus (HCV). I argue that traditional knowledge and healing practices are central to Māori getting well and keeping well, and that the use of cultural frameworks and practices have potentially restorative, therapeutic and healing values that are not yet researched or understood by the health field. I argue that a Māori framework of wellbeing, namely ‘Mana Kaitiakitanga’, provides the context in which tā moko (Māori traditional tattoo) fits naturally as a healing intervention. I share the stories of Māori women with HCV who have applied this (tā moko and other forms of tattoo) in their lives and in their journeys back to wellness. Tā moko is a process that penetrates the flesh and marks the skin; it is a process that involves both blood and pain, which may seem incongruous with healing. It is argued however that through pain comes understanding; through pain comes a RE-membering of strength; through pain comes joy; and finally through marking comes identity of who we are and how well we have been and can be again.

Issues and intersections of identity, marginalisation, gender, health, and wellbeing are at the forefront of this research story and of the journeys of the three women whose case studies are presented in this thesis. What makes this thesis unique is that it researches all three of these potential cornerstones of health: identity; wellbeing (spiritual and emotional), and physical wellbeing, in relation to a specific health problem, namely HCV. It is intended that this work will add power to what might be viewed as a particularly Indigenous solution to a virus that disproportionately affects our Indigenous population. This is not about rejecting Western health solutions, but it is about recognising and returning to Indigenous health solutions.

text: Mouri Tu, Mouri Moko, Mouri Ora! Moko as a wellbeing strategy This research has been undertaken within the Health Research Council funded International Collaborative Indigenous Health Research Partnership (ICIHRP) program, “The role of resiliency in responding to blood borne viral and sexually transmitted infections in Indigenous communities”. Some Indigenous communities in Australia, New Zealand and Canada have been shown to experience higher rates of blood borne viral infections. There are a number of categories of health intervention that have been shown to be effective in prevention and enhancing access to treatment for blood borne viral infections, including various forms of health promotion, enhanced diagnostic and treatment services, and harm reduction measures related to injecting drug use. The present research was undertaken with the understanding that development of effective responses of this kind among Indigenous people would benefit from a better understanding of the social and cultural factors that might protect against these infections and their consequences. It is argued that such factors are linked in various ways to Indigenous resistance and resiliency, which is described as the means by which people choose to make use of individual and community strengths to protect themselves against adverse health outcomes and enhance their health and wellbeing.

In this thesis I explore how people and their identity are affected when you are part of a marginalised or vulnerable population – namely, Māori women who have contracted the Hepatitis C Virus (HCV). I argue that traditional knowledge and healing practices are central to Māori getting well and keeping well, and that the use of cultural frameworks and practices have potentially restorative, therapeutic and healing values that are not yet researched or understood by the health field. I argue that a Māori framework of wellbeing, namely ‘Mana Kaitiakitanga’, provides the context in which tā moko (Māori traditional tattoo) fits naturally as a healing intervention. I share the stories of Māori women with HCV who have applied this (tā moko and other forms of tattoo) in their lives and in their journeys back to wellness. Tā moko is a process that penetrates the flesh and marks the skin; it is a process that involves both blood and pain, which may seem incongruous with healing. It is argued however that through pain comes understanding; through pain comes a RE-membering of strength; through pain comes joy; and finally through marking comes identity of who we are and how well we have been and can be again.

Issues and intersections of identity, marginalisation, gender, health, and wellbeing are at the forefront of this research story and of the journeys of the three women whose case studies are presented in this thesis. What makes this thesis unique is that it researches all three of these potential cornerstones of health: identity; wellbeing (spiritual and emotional), and physical wellbeing, in relation to a specific health problem, namely HCV. It is intended that this work will add power to what might be viewed as a particularly Indigenous solution to a virus that disproportionately affects our Indigenous population. This is not about rejecting Western health solutions, but it is about recognising and returning to Indigenous health solutions.

year: 2011