title: Targets of Elf5 in Mouse Trophoblast Stem Cells

authors: Deane, Jessica Robyn

subjects: Elf5

abstract: The Placenta is an essential organ for all mammalian embryonic development as it provides the nutritional link between maternal and foetal blood streams. The cells which go on to proliferate and contribute to all the major cell types of the embryo derived placenta have been located to the trophoblast (TE) cells overlying the Inner Cell Mass (ICM) of the embryo. Immortal cell lines have been subsequentially derived from this tissue and called Trophoblast Stem (TS) cells. In parallel with their  in vivo  counterparts they are also reliant on Fibroblast growth factor 4 (FgF4) (Tanaka et al., 1998) and Activin/Nodal signalling (Guzman-Ayala et al., 2004). 

The Ets family transcription factor, Elf5, has been shown to be specifically expressed in the early placental trophoblast and subsequent derived tissues. Mice deficient in Elf5 failed to form a placenta post implantation. Furthermore TS cells were unable to be derived from Elf5 knockout embryos (Donnison et al., 2005). This work suggested that Elf5 plays an essential role in TS cells and their differentiation. The aim of this study was to determine the downstream target genes of Elf5 in mouse TS cells. The target genes of Fgf4 and Activin/Nodal signalling in TS cells were also investigated.  This work is hoped to contribute to an overall greater understanding of the molecular networks underlying TS cell maintenance and to contribute to our knowledge of early placental development. 

Small interfering RNA (siRNA) targeted reduction of Elf5 mRNA expression in mTS cells was achieved using two independent siRNAs; with Elf5 reduction exceeding 80%. The resulting changes in gene expression were measured in order to determine the downstream targets of Elf5. Selected genes known to be important for trophoblast differentiation and maintenance were measured using real-time PCR in a candidate gene approach .Global changes in gene expression as a consequence of Elf5 silencing were measured using an Affymetrix microarray. Global changes in gene expression due to growth factor (Fgf4 and/or Activin) removal were also measured. Expression of 22 genes was changed using either of the Elf5 siRNA oligonucleotides. Of these, 9 were also significantly changed by growth factor removal. Included in this set were Synopl, Hst3st3b1, Cyr61 and Sox2. In the overall analysis, many genes whose expression changed upon loss of Elf5 are known to play important roles in trophectoderm cell specification. Real-time PCR validation agreed closely with the up or down regulation measured using the microarray. This work has thus led to the discovery of sets of Elf5 target genes potentially involved in trophoblast stem cell function and has provided the foundation for future work exploring the molecular pathways of trophoblast development.

text: Targets of Elf5 in Mouse Trophoblast Stem Cells The Placenta is an essential organ for all mammalian embryonic development as it provides the nutritional link between maternal and foetal blood streams. The cells which go on to proliferate and contribute to all the major cell types of the embryo derived placenta have been located to the trophoblast (TE) cells overlying the Inner Cell Mass (ICM) of the embryo. Immortal cell lines have been subsequentially derived from this tissue and called Trophoblast Stem (TS) cells. In parallel with their  in vivo  counterparts they are also reliant on Fibroblast growth factor 4 (FgF4) (Tanaka et al., 1998) and Activin/Nodal signalling (Guzman-Ayala et al., 2004). 

The Ets family transcription factor, Elf5, has been shown to be specifically expressed in the early placental trophoblast and subsequent derived tissues. Mice deficient in Elf5 failed to form a placenta post implantation. Furthermore TS cells were unable to be derived from Elf5 knockout embryos (Donnison et al., 2005). This work suggested that Elf5 plays an essential role in TS cells and their differentiation. The aim of this study was to determine the downstream target genes of Elf5 in mouse TS cells. The target genes of Fgf4 and Activin/Nodal signalling in TS cells were also investigated.  This work is hoped to contribute to an overall greater understanding of the molecular networks underlying TS cell maintenance and to contribute to our knowledge of early placental development. 

Small interfering RNA (siRNA) targeted reduction of Elf5 mRNA expression in mTS cells was achieved using two independent siRNAs; with Elf5 reduction exceeding 80%. The resulting changes in gene expression were measured in order to determine the downstream targets of Elf5. Selected genes known to be important for trophoblast differentiation and maintenance were measured using real-time PCR in a candidate gene approach .Global changes in gene expression as a consequence of Elf5 silencing were measured using an Affymetrix microarray. Global changes in gene expression due to growth factor (Fgf4 and/or Activin) removal were also measured. Expression of 22 genes was changed using either of the Elf5 siRNA oligonucleotides. Of these, 9 were also significantly changed by growth factor removal. Included in this set were Synopl, Hst3st3b1, Cyr61 and Sox2. In the overall analysis, many genes whose expression changed upon loss of Elf5 are known to play important roles in trophectoderm cell specification. Real-time PCR validation agreed closely with the up or down regulation measured using the microarray. This work has thus led to the discovery of sets of Elf5 target genes potentially involved in trophoblast stem cell function and has provided the foundation for future work exploring the molecular pathways of trophoblast development.

year: 2007

title: White-noise susceptibility and critical slowing in neurons near spiking threshold

authors: Steyn-Ross, D. Alistair

subjects: neuron

abstract: We present mathematical and simulation analyses of the below-threshold noisy response of two biophysically motivated models for excitable membrane due to H. R. Wilson: a squid axon (“resonator”) and a human cortical neuron (“integrator”). When stimulated with a low-intensity white noise superimposed on a dc control current, both membrane types generate voltage fluctuations that exhibit critical slowing down—that is, the voltage responsiveness to noisy input currents grows in amplitude while slowing in frequency—as the membrane approaches spiking threshold from below. We define threshold unambiguously as that dc current that renders a zero real eigenvalue for the Jacobian matrix for the integrator neuron, and, for the resonator neuron, as the dc current that gives a complex eigenvalue pair whose real part is zero. Using a linear Ornstein-Uhlenbeck analysis, we give exact small-noise expressions for the variance, power spectrum, and correlation function of the voltage fluctuations, and we derive the scaling laws for the divergence of susceptibility and correlation times for approach to threshold. We compare these predictions with numerical simulations of the nonlinear stochastic equations, and demonstrate that, provided the white-noise perturbations are kept sufficiently small, the linearized theory works well. These predictions should be testable in the laboratory using a current-clamped cell configuration. If confirmed, then the proximity of a neuron to its spike-transition point can be judged by measuring its subthreshold susceptibility to white-noise stimulation. We postulate that such temporally correlated fluctuations could provide a means of subthreshold signaling via gap-junction connections with neighboring neurons.

text: White-noise susceptibility and critical slowing in neurons near spiking threshold We present mathematical and simulation analyses of the below-threshold noisy response of two biophysically motivated models for excitable membrane due to H. R. Wilson: a squid axon (“resonator”) and a human cortical neuron (“integrator”). When stimulated with a low-intensity white noise superimposed on a dc control current, both membrane types generate voltage fluctuations that exhibit critical slowing down—that is, the voltage responsiveness to noisy input currents grows in amplitude while slowing in frequency—as the membrane approaches spiking threshold from below. We define threshold unambiguously as that dc current that renders a zero real eigenvalue for the Jacobian matrix for the integrator neuron, and, for the resonator neuron, as the dc current that gives a complex eigenvalue pair whose real part is zero. Using a linear Ornstein-Uhlenbeck analysis, we give exact small-noise expressions for the variance, power spectrum, and correlation function of the voltage fluctuations, and we derive the scaling laws for the divergence of susceptibility and correlation times for approach to threshold. We compare these predictions with numerical simulations of the nonlinear stochastic equations, and demonstrate that, provided the white-noise perturbations are kept sufficiently small, the linearized theory works well. These predictions should be testable in the laboratory using a current-clamped cell configuration. If confirmed, then the proximity of a neuron to its spike-transition point can be judged by measuring its subthreshold susceptibility to white-noise stimulation. We postulate that such temporally correlated fluctuations could provide a means of subthreshold signaling via gap-junction connections with neighboring neurons.

year: 2006

title: A critical interruption in the governance of a New Zealand state high school

authors: L'Huillier, Barbara Marie

subjects: governance

abstract: The governance of contemporary Western education aspires to serve the intertwining values of human emancipation and social inclusiveness in both the organisational form of educational institutions and their pedagogical direction. In New Zealand (NZ), what is understood as ‘good’ governance of education has varied over time according to economic conditions and political interests. The fourth Labour government (1984-1990), which undertook a widespread, rapid commitment to neo-liberal principles, reorganised the provision and governance of education along quasi public-private sector lines, explicitly using a business approach to guide practice, yet retaining hegemonic characteristics. This commitment to neo-liberal principles has been continued by all subsequent NZ governments. The intended transformation of the education system from a centrally-governed bureaucracy to a community-governed process has not been easy. Underpinning the new governance regime is an implied notion that school communities share a common understanding of what is meant by education, processes of representation, and of ‘good’ governance. However, due to the different aspirations members of school communities’ can hold for their respective schools this commonality of meaning can be a false dichotomy resulting in schools being governed in a state of tension. This tension serves to highlight contradictions and disconnections between the espoused aims of devolved governance and a State-mandated process that works to exclude certain factions of a school community. Increasing numbers of NZ school Boards of Trustees are experiencing these tensions and contradictions resulting in an interruption in school governance. The interruption I am interested in takes the form of the commodification of education that has been taking place in NZ since the mid 1980s. I argue this to be an interruption with profound effects on both the flourishing of individuals and the cohesion of communities and usefully serves to draw attention to the disconnection between the broad ideals of education, as articulated by the early promoters of a State-funded education system, and the neo-liberal re-visioning of State education. Fairfield College is a NZ State high school where the tensions and contradictions surrounding the meaning of education, processes of representation, and of good governance have generated a crisis resulting in a critical interruption in school governance. The school’s Board of Trustees received considerable media coverage between 2008 and 2009 regarding a critical interruption to their governance. This interruption presented a unique research opportunity to explore how the words used to frame the problems facing the Board of Trustees, the words used to explain and justify State intervention in the governance of the school, and the words used to propose solutions to perceived problems were invested with meaning. With my interest in connotative meaning, I illustrate how these words were used to encourage and endorse actions that comply with prevailing public policy. In this case study I examined how the school’s direction has been reconfigured according to the principles of corporate governance using a companion lexicon of business. Directives have been devised by various NZ governments to align education with the neo-liberal principles that have prevailed in the governance of this country since 1984. The multiple meanings attributed to ‘governance’ and ‘corporate governance’ in the relevant literatures, and the various implications for applied governance processes and dynamics, influenced my decision to adopt the dual orientation of social constructionism and critical theory to underpin this research. Social constructionists focus on how individuals and groups produce, disseminate, and validate their perceptions of reality. Critical theorists aim to create awareness of constraints to emancipatory potential and inclusiveness. They seek to expose the contradictions between espoused aims of justice and the flourishing of individuals and that which is practised, with the intention to contribute to changes that would enhance justice and the flourishing of individuals. One such contribution is to give voice to those who feel their voices are not being heard. Both theoretical schools regard the use of language as pivotal in the construction of social processes in which meaning is continually negotiated and re-negotiated by the stakeholders. In this research I engaged in audio-recorded semi-structured interviews with members from four stakeholder groups involved in the governance of Fairfield College: the Whānau Support Group (a predominantly Māori special interest group), parents and caregivers, teachers, and those previously or currently charged with governance responsibilities for the school. These narratives were then analysed using thematic analysis to provide important insights into how ‘governance’ of the school was interpreted, challenged, and/or endorsed. My research at Fairfield College indicates that the different constructions of ‘governance’ are associated with identifiable overlapping issues arranged in a hierarchy. The foundational tier represents the outcome of various NZ governments’ preference for market-generated principles of organisation at a macro State level, namely the commodification of public services – in this instance education. The middle tier addresses concerns raised by some stakeholders regarding their notion of inclusiveness and the vulnerability of the democratic process as experienced at community level. The top tier encompasses the twin issues of communication and consultation processes at a management level that appear to demonstrate a conflict between the State’s espoused principles of good governance and their application at Fairfield College. As a result of examining the critical interruption to the governance of Fairfield College I conclude that when viewed through the lens of a social constructionist orientation informed by critical theory, ‘governance’ in the education sector can be interpreted as a process supporting neo-liberal preferences for market-orientated approaches to education. Such preferences overshadow any other ideals held by members of the school’s community and, according to a critical understanding of neo-liberalism, weaken the principles of emancipation and inclusiveness. The insights generated from this research provide a basis for further research into the governance of other organisations with emancipatory and inclusive aspirations.

text: A critical interruption in the governance of a New Zealand state high school The governance of contemporary Western education aspires to serve the intertwining values of human emancipation and social inclusiveness in both the organisational form of educational institutions and their pedagogical direction. In New Zealand (NZ), what is understood as ‘good’ governance of education has varied over time according to economic conditions and political interests. The fourth Labour government (1984-1990), which undertook a widespread, rapid commitment to neo-liberal principles, reorganised the provision and governance of education along quasi public-private sector lines, explicitly using a business approach to guide practice, yet retaining hegemonic characteristics. This commitment to neo-liberal principles has been continued by all subsequent NZ governments. The intended transformation of the education system from a centrally-governed bureaucracy to a community-governed process has not been easy. Underpinning the new governance regime is an implied notion that school communities share a common understanding of what is meant by education, processes of representation, and of ‘good’ governance. However, due to the different aspirations members of school communities’ can hold for their respective schools this commonality of meaning can be a false dichotomy resulting in schools being governed in a state of tension. This tension serves to highlight contradictions and disconnections between the espoused aims of devolved governance and a State-mandated process that works to exclude certain factions of a school community. Increasing numbers of NZ school Boards of Trustees are experiencing these tensions and contradictions resulting in an interruption in school governance. The interruption I am interested in takes the form of the commodification of education that has been taking place in NZ since the mid 1980s. I argue this to be an interruption with profound effects on both the flourishing of individuals and the cohesion of communities and usefully serves to draw attention to the disconnection between the broad ideals of education, as articulated by the early promoters of a State-funded education system, and the neo-liberal re-visioning of State education. Fairfield College is a NZ State high school where the tensions and contradictions surrounding the meaning of education, processes of representation, and of good governance have generated a crisis resulting in a critical interruption in school governance. The school’s Board of Trustees received considerable media coverage between 2008 and 2009 regarding a critical interruption to their governance. This interruption presented a unique research opportunity to explore how the words used to frame the problems facing the Board of Trustees, the words used to explain and justify State intervention in the governance of the school, and the words used to propose solutions to perceived problems were invested with meaning. With my interest in connotative meaning, I illustrate how these words were used to encourage and endorse actions that comply with prevailing public policy. In this case study I examined how the school’s direction has been reconfigured according to the principles of corporate governance using a companion lexicon of business. Directives have been devised by various NZ governments to align education with the neo-liberal principles that have prevailed in the governance of this country since 1984. The multiple meanings attributed to ‘governance’ and ‘corporate governance’ in the relevant literatures, and the various implications for applied governance processes and dynamics, influenced my decision to adopt the dual orientation of social constructionism and critical theory to underpin this research. Social constructionists focus on how individuals and groups produce, disseminate, and validate their perceptions of reality. Critical theorists aim to create awareness of constraints to emancipatory potential and inclusiveness. They seek to expose the contradictions between espoused aims of justice and the flourishing of individuals and that which is practised, with the intention to contribute to changes that would enhance justice and the flourishing of individuals. One such contribution is to give voice to those who feel their voices are not being heard. Both theoretical schools regard the use of language as pivotal in the construction of social processes in which meaning is continually negotiated and re-negotiated by the stakeholders. In this research I engaged in audio-recorded semi-structured interviews with members from four stakeholder groups involved in the governance of Fairfield College: the Whānau Support Group (a predominantly Māori special interest group), parents and caregivers, teachers, and those previously or currently charged with governance responsibilities for the school. These narratives were then analysed using thematic analysis to provide important insights into how ‘governance’ of the school was interpreted, challenged, and/or endorsed. My research at Fairfield College indicates that the different constructions of ‘governance’ are associated with identifiable overlapping issues arranged in a hierarchy. The foundational tier represents the outcome of various NZ governments’ preference for market-generated principles of organisation at a macro State level, namely the commodification of public services – in this instance education. The middle tier addresses concerns raised by some stakeholders regarding their notion of inclusiveness and the vulnerability of the democratic process as experienced at community level. The top tier encompasses the twin issues of communication and consultation processes at a management level that appear to demonstrate a conflict between the State’s espoused principles of good governance and their application at Fairfield College. As a result of examining the critical interruption to the governance of Fairfield College I conclude that when viewed through the lens of a social constructionist orientation informed by critical theory, ‘governance’ in the education sector can be interpreted as a process supporting neo-liberal preferences for market-orientated approaches to education. Such preferences overshadow any other ideals held by members of the school’s community and, according to a critical understanding of neo-liberalism, weaken the principles of emancipation and inclusiveness. The insights generated from this research provide a basis for further research into the governance of other organisations with emancipatory and inclusive aspirations.

year: 2011