title: Grainyhead-like 2 is required for morphological integrity of mouse embryonic stem cells and orderly formation of inner ear-like organoids

authors: Forrester-Gauntlett, Blaise

abstract: Mutations in the transcription factor gene grainyhead-like 2 (GRHL2) are associated with progressive non-syndromic sensorineural deafness autosomal dominant type 28 (DFNA28) in humans. Since complete loss of Grhl2 is lethal in mouse embryos, we studied its role during inner ear pathology and hearing loss in vitro. To this end, we generated different homozygous deletions to knockout Grhl2 in mouse embryonic stem cells (Grhl2-KO ESCs), including some mimicking naturally occurring truncations in the dimerisation domain related to human DFNA28. Under naïve culture conditions, Grhl2-KO cells in suspension were more heterogenous in size and larger than wild-type controls. Adherent Grhl2-KO cells were also larger, with a less uniform shape, flattened, less circular morphology, forming loose monolayer colonies with poorly defined edges. These changes correlated with lower expression of epithelial cadherin Cdh1 but no changes in tight junction markers (Ocln, Tjp2) or other Grhl isoforms (Grhl1, Grhl3). Clonogenicity from single cells, proliferation rates of cell populations and proliferation markers were reduced in Grhl2-KO ESCs. We next induced stepwise directed differentiation of Grhl2-KO ESCs along an otic pathway, giving rise to three-dimensional inner ear-like organoids (IELOs). Quantitative morphometry revealed that Grhl2-KO cells initially formed larger IELOs with a less compacted structure, more eccentric shape and increased surface area. These morphological changes persisted for up to one week. They were partially rescued by forced cell aggregation and fully restored by stably overexpressing exogenous Grhl2 in Grhl2-KO ESCs, indicating that Grhl2 alters cell-cell interactions. On day 8, aggregates were transferred into minimal maturation medium to allow self-guided organogenesis for another two weeks. During this period, Grhl2-KO cells and wild-type controls developed similarly, expressing neural, neuronal and sensory hair cell markers, while maintaining their initial differences in size and shape. In summary, Grhl2 is required for morphological maintenance of ESCs and orderly formation of IELOs, consistent with an essential role in organising epithelial integrity during inner ear development. Our findings validate quantitative morphometry as a useful, non-invasive screening method for molecular phenotyping of candidate mutations during organoid development.

text: Grainyhead-like 2 is required for morphological integrity of mouse embryonic stem cells and orderly formation of inner ear-like organoids Mutations in the transcription factor gene grainyhead-like 2 (GRHL2) are associated with progressive non-syndromic sensorineural deafness autosomal dominant type 28 (DFNA28) in humans. Since complete loss of Grhl2 is lethal in mouse embryos, we studied its role during inner ear pathology and hearing loss in vitro. To this end, we generated different homozygous deletions to knockout Grhl2 in mouse embryonic stem cells (Grhl2-KO ESCs), including some mimicking naturally occurring truncations in the dimerisation domain related to human DFNA28. Under naïve culture conditions, Grhl2-KO cells in suspension were more heterogenous in size and larger than wild-type controls. Adherent Grhl2-KO cells were also larger, with a less uniform shape, flattened, less circular morphology, forming loose monolayer colonies with poorly defined edges. These changes correlated with lower expression of epithelial cadherin Cdh1 but no changes in tight junction markers (Ocln, Tjp2) or other Grhl isoforms (Grhl1, Grhl3). Clonogenicity from single cells, proliferation rates of cell populations and proliferation markers were reduced in Grhl2-KO ESCs. We next induced stepwise directed differentiation of Grhl2-KO ESCs along an otic pathway, giving rise to three-dimensional inner ear-like organoids (IELOs). Quantitative morphometry revealed that Grhl2-KO cells initially formed larger IELOs with a less compacted structure, more eccentric shape and increased surface area. These morphological changes persisted for up to one week. They were partially rescued by forced cell aggregation and fully restored by stably overexpressing exogenous Grhl2 in Grhl2-KO ESCs, indicating that Grhl2 alters cell-cell interactions. On day 8, aggregates were transferred into minimal maturation medium to allow self-guided organogenesis for another two weeks. During this period, Grhl2-KO cells and wild-type controls developed similarly, expressing neural, neuronal and sensory hair cell markers, while maintaining their initial differences in size and shape. In summary, Grhl2 is required for morphological maintenance of ESCs and orderly formation of IELOs, consistent with an essential role in organising epithelial integrity during inner ear development. Our findings validate quantitative morphometry as a useful, non-invasive screening method for molecular phenotyping of candidate mutations during organoid development.

year: 2023-01-01

title: Statistical representations of the housing problem in briefings to incoming ministers, 2008–2020: The politics of housing numbers

authors: Barrett, Patrick

abstract: An interpretive policy studies perspective on what is understood to be the ‘facts’ about the housing problem in New Zealand has potential to uncover the way statistical representations of problems are embedded in larger normative narratives, and the consequential implications for housing policy and governance. This paper analyses such representations as they are evident in policy briefings to incoming Ministers of Housing between 2008 and 2020 by Housing New Zealand (HNZ) and its successor Kāinga Ora, and the way these reflect and reinforce a neoliberal political rationality and an agenda to residualise and marketise state housing support. The briefings represent authoritative accounts by a key bureaucratic advisory agency of the significant issues and priorities in the housing portfolio. The period prior to 2017 saw the emergence of new articulations of housing problems relating to affordability, declining rates of home ownership, an increasing rental population, and increasing homelessness. In the briefings to ministers, these problems were routinely framed as issues of supply and demand and market adjustment, and this has continued following the change of government in 2017. The focus of the paper is on identifying statistical representations within the discursive context that give weight to particular policy choices.

text: Statistical representations of the housing problem in briefings to incoming ministers, 2008–2020: The politics of housing numbers An interpretive policy studies perspective on what is understood to be the ‘facts’ about the housing problem in New Zealand has potential to uncover the way statistical representations of problems are embedded in larger normative narratives, and the consequential implications for housing policy and governance. This paper analyses such representations as they are evident in policy briefings to incoming Ministers of Housing between 2008 and 2020 by Housing New Zealand (HNZ) and its successor Kāinga Ora, and the way these reflect and reinforce a neoliberal political rationality and an agenda to residualise and marketise state housing support. The briefings represent authoritative accounts by a key bureaucratic advisory agency of the significant issues and priorities in the housing portfolio. The period prior to 2017 saw the emergence of new articulations of housing problems relating to affordability, declining rates of home ownership, an increasing rental population, and increasing homelessness. In the briefings to ministers, these problems were routinely framed as issues of supply and demand and market adjustment, and this has continued following the change of government in 2017. The focus of the paper is on identifying statistical representations within the discursive context that give weight to particular policy choices.

year: 2021

title: Sport in the Waikato c.1897-1974: Narratives of Play, Identity and Belonging

authors: Buckley, Karen

subjects: sport

abstract: Regional identities and allegiances in New Zealand have most often been revealed at sporting events. From the colours worn, the slogans chanted, and even the ringing of cowbells, players, teams and supporters have presented facets of their collective character. This thesis engages with narratives of sport in the Waikato region over a period of eighty years to examine these representations as they produced meanings of sporting activities for local people. Informed by competing ideas about historical truth, the thesis interprets printed media as reconstructions of both recurring events, and belonging and membership, to investigate the place of sport in the region, and to decode the ways these stories of drama and contest contribute to the development of some components of a local identity.
Historically, the area now known as the ‘Waikato’ was part of the Auckland Province. The thesis argues that sport is at the centre of a notion of separation from the urban metropolis, and provided opportunities to demonstrate a growing independence. By examining examples of recurring sporting events such as the annual Ngāruawāhia Regatta, regular Auckland/Waikato rugby matches, Empire and Commonwealth Games participation, and regular lawn bowls tournaments, the place and function of sport in a regional space is able to be, in part, exposed. Uneven, often subtle, changes both in aspects of these events and in the representations of them, reveal underlying tensions and power relationships.
Local, if not exclusive, understandings of the place and role of Māori, and depictions of normative gender roles, are central to the analysis of these relationships. Sport is most often a social activity, and dissection of the newspaper texts has enabled the formation of ideas about the ways sports-men, and sometimes women, are collectively presented and organised. In much of the material used, these men and women are also the most recognisable and celebrated ‘heroes’. Consideration of the role of heroes in the formation of culturally constructed group identities informs the thesis, and the relationships between local sporting participants and groups are also important. Divisions, dissension, and competition for resources are all revealed in the reported activities of sports organisations, and changes in the importance afforded these endeavours are also apparent.
The focussed critical use of a local media source provides a consistent, if sometimes homogenous, group of historical narratives. The inclusion of local club and association histories, often produced with extensive use of newspaper archives, changes the perspective and enables a more nuanced examination of the place of different types of heroes, of gender norms and, most importantly, of the ways sporting organisations see and project themselves as part of the region. The stories they collectively choose to remember and tell are central to this narrative of the place of sport in the Waikato.

text: Sport in the Waikato c.1897-1974: Narratives of Play, Identity and Belonging Regional identities and allegiances in New Zealand have most often been revealed at sporting events. From the colours worn, the slogans chanted, and even the ringing of cowbells, players, teams and supporters have presented facets of their collective character. This thesis engages with narratives of sport in the Waikato region over a period of eighty years to examine these representations as they produced meanings of sporting activities for local people. Informed by competing ideas about historical truth, the thesis interprets printed media as reconstructions of both recurring events, and belonging and membership, to investigate the place of sport in the region, and to decode the ways these stories of drama and contest contribute to the development of some components of a local identity.
Historically, the area now known as the ‘Waikato’ was part of the Auckland Province. The thesis argues that sport is at the centre of a notion of separation from the urban metropolis, and provided opportunities to demonstrate a growing independence. By examining examples of recurring sporting events such as the annual Ngāruawāhia Regatta, regular Auckland/Waikato rugby matches, Empire and Commonwealth Games participation, and regular lawn bowls tournaments, the place and function of sport in a regional space is able to be, in part, exposed. Uneven, often subtle, changes both in aspects of these events and in the representations of them, reveal underlying tensions and power relationships.
Local, if not exclusive, understandings of the place and role of Māori, and depictions of normative gender roles, are central to the analysis of these relationships. Sport is most often a social activity, and dissection of the newspaper texts has enabled the formation of ideas about the ways sports-men, and sometimes women, are collectively presented and organised. In much of the material used, these men and women are also the most recognisable and celebrated ‘heroes’. Consideration of the role of heroes in the formation of culturally constructed group identities informs the thesis, and the relationships between local sporting participants and groups are also important. Divisions, dissension, and competition for resources are all revealed in the reported activities of sports organisations, and changes in the importance afforded these endeavours are also apparent.
The focussed critical use of a local media source provides a consistent, if sometimes homogenous, group of historical narratives. The inclusion of local club and association histories, often produced with extensive use of newspaper archives, changes the perspective and enables a more nuanced examination of the place of different types of heroes, of gender norms and, most importantly, of the ways sporting organisations see and project themselves as part of the region. The stories they collectively choose to remember and tell are central to this narrative of the place of sport in the Waikato.

year: 2016