Generating biomarkers of human Nek8 kinase activity

Nek8 is a member of the human NIMA-related serine/threonine protein kinase family that has roles in cell cycle progression, primary cilia function and the DNA damage response. Mutations in the human nek8 gene cause Nephronophthisis (NPHP), an autosomal recessive polycystic kidney disease in which defects in primary cilia lead to a broad range of often severe symptoms that affect many organs of the body. In the kidneys, NPHP causes the development of cortico-medullary cysts that impair kidney function and ultimately lead to kidney failure. Nek8 localises to the primary cilium where it interacts with Inversin and PC-2, products of genes that are also mutated in inherited cystic kidney diseases. Nek8 also plays a role in the intra-S phase DNA damage checkpoint where it contributes to cell cycle arrest by inhibiting CDK2 activity to allow time for repair of stalled replication forks. The purpose of this study was to generate novel biomarkers of Nek8 kinase activity that could be used to shed light on its role at the primary cilium and in the replication stress response. Our aim was therefore to identify phosphosites of Nek8 in its potential substrate proteins, Inversin and PC-2, and to generate phospho-specific antibodies against these sites. We chose three sites in the Inversin N-terminus for phospho-specific antibody generation and showed that these were capable of detecting purified, phosphorylated Inversin upon incubation with Nek8. Second, we characterised a phospho-Nek8 antibody that recognises a phosphorylation site in the activation loop of the kinase. This antibody was capable of detecting autophosphorylated Nek8 by Western blot and immunofluorescence microscopy as confirmed using two, newly identified chemical inhibitors of Nek8. Localisation studies with this antibody revealed novel data on the presence of active Nek8 at centrosomes, cilia and sites of DNA damage. Finally, we found that Nek8 inhibition was associated with generation of enlarged multinucleated cells and accumulation of DNA damage foci. Together, these data support a role for Nek8 in linking ciliary signalling pathways and the DNA damage response, while the phospho-specific antibodies represent a new set of tools that can be used to explore Nek8 function in normal and pathological states

Challenges and solutions: surveying researchers on what type of community engagement and involvement activities are feasible in low and middle income countries during the COVID-19 pandemic

OBJECTIVES: Measures to limit the spread of infection during the COVID-19 global pandemic have made engaging and involving members of the community in global health research more challenging. This research aimed to explore how global health researchers adapted to the imposed pandemic measures in low and middle income countries (LMICs) and how they overcame challenges to effective community engagement and involvement (CEI).DESIGN: A qualitative two-stage mixed-methods study involving an online survey and a virtual round table.SETTING: The survey and round table were completed online.PARTICIPANTS: Of 53 participants, 43 were LMIC-based or UK-based global health researchers and/or CEI professionals, and 10 worked for the National Institute for Health Research or UK Government's Department of Health and Social Care.OUTCOME MEASURES: This study aimed to capture data on: the number of CEI activities halted and adapted because of the COVID-19 pandemic; where CEI is possible; how it has been adapted; what the challenges and successes were; and the potential impact of adapted or halted CEI on global health research.RESULTS: Pandemic control measures forced the majority of researchers to stop or amend their planned CEI activities. Most face-to-face CEI activities were replaced with remote methods, such as online communication. Virtual engagement enabled researchers to maintain already established relationships with community members, but was less effective when developing new relationships or addressing challenges around the inclusion of marginalised community groups.CONCLUSIONS: COVID-19 has highlighted the need for contingency planning and flexibility in CEI. The redesigning and adopting of remote methods has come with both advantages and disadvantages, and required new skills, access to technology, funding, reliable services and enthusiasm from stakeholders. The methods suggested have the potential to augment or substitute previously preferred CEI activities. The effectiveness and impact of these remote CEI activities need to be assessed.</p

Community engagement and involvement in Ghana:conversations with community stakeholders to inform surgical research

BACKGROUND: Involving patients and communities with health research in low- and middle-income countries (LMICs) contributes to increasing the likelihood that research is relevant in local context and caters to the needs of the population, including vulnerable and marginalised groups. When done right, it can also support empowerment of wider communities in taking ownership of their own health, lead to increased access and uptake of health services and generally improve the wellbeing of individuals. However, the evidence base of how to undertake successful community engagement and involvement (CEI) activities in LMICs is sparse. This paper aims to add to the available literature and describes how the Global Health Research Unit on Global Surgery’s (GSU) team in Ghana worked collaboratively with the Unit’s team in the UK and a UK-based Public Advisory Group to involve community stakeholders in rural Ghana with surgical research. The aim was to explore ways of reaching out to patients and community leaders in rural Ghana to have conversations that inform the relevance, acceptability, and feasibility of a clinical trial, called TIGER. METHODS: As this kind of larger scale involvement of community stakeholders with research was a novel way of working for the team in Ghana, a reflective approach was taken to outline step-by-step how the GSU team planned and undertook these involvement activities with 31 hernia patients, two Chiefs (community leaders), a community finance officer and a local politician in various locations in Ghana. The barriers that were experienced and the benefits of involving community stakeholders are highlighted with the aim to add to the evidence base of CEI in LMICs. RESULTS: GSU members from the UK and Ghana planned and organised successful involvement activities that focused on establishing the best way to talk to patients and other community stakeholders about their experiences of living with hernias and undergoing hernia repairs, and their perceptions of the impact of hernias on the wider community. The Ghanaian team suggested 1:1 conversations in easily accessible locations for rural patient contributors, creating a welcoming environment and addressing contributors in their local dialects. A UK-based Public Advisory Group helped in the initial stages of planning these conversations by highlighting potential barriers when approaching rural communities and advising on how to phrase questions around personal experiences. Conversations mainly focused on understanding the needs of hernia patients in rural Ghana to then incorporate these in the design of the TIGER trial to ensure its relevance, acceptability and feasibility. When talking to patient contributors, the GSU teams found that they were more likely to open up when they knew members of the team and the opportunity to speak to local leaders only arose because of the Ghanaian team members being well-respected amongst communities. The experiences of the patient and community contributors led to changes in the study protocol, such as including women in the patient cohort for the trial, and allowed the GSU teams to confirm the relevance and acceptability of this trial. These conversations also taught the team a lot about perceptions of health in rural communities, allowed the Ghanaian team to establish relationships with community leaders that can be utilised when future studies need input from the public, and has changed the minds of the Ghanaian research team about the importance of involving patients with research. CONCLUSION: This paper contributes to the evidence base on successful CEI activities in LMICs by providing an example of how CEI can be planned and organised, and the benefits this provides. The conversations the teams had with patient contributors in Ghana are an example of successful patient consultations. Even though there are certain limitations to the extent of these involvement activities, a solid foundation has been built for researchers and community stakeholders to establish relationships for ongoing involvement. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s40900-021-00270-5

Adaptation of the Wound Healing Questionnaire universal-reporter outcome measure for use in global surgery trials (TALON-1 study): mixed-methods study and Rasch analysis

BackgroundThe Bluebelle Wound Healing Questionnaire (WHQ) is a universal-reporter outcome measure developed in the UK for remote detection of surgical-site infection after abdominal surgery. This study aimed to explore cross-cultural equivalence, acceptability, and content validity of the WHQ for use across low- and middle-income countries, and to make recommendations for its adaptation.MethodsThis was a mixed-methods study within a trial (SWAT) embedded in an international randomized trial, conducted according to best practice guidelines, and co-produced with community and patient partners (TALON-1). Structured interviews and focus groups were used to gather data regarding cross-cultural, cross-contextual equivalence of the individual items and scale, and conduct a translatability assessment. Translation was completed into five languages in accordance with Mapi recommendations. Next, data from a prospective cohort (SWAT) were interpreted using Rasch analysis to explore scaling and measurement properties of the WHQ. Finally, qualitative and quantitative data were triangulated using a modified, exploratory, instrumental design model.ResultsIn the qualitative phase, 10 structured interviews and six focus groups took place with a total of 47 investigators across six countries. Themes related to comprehension, response mapping, retrieval, and judgement were identified with rich cross-cultural insights. In the quantitative phase, an exploratory Rasch model was fitted to data from 537 patients (369 excluding extremes). Owing to the number of extreme (floor) values, the overall level of power was low. The single WHQ scale satisfied tests of unidimensionality, indicating validity of the ordinal total WHQ score. There was significant overall model misfit of five items (5, 9, 14, 15, 16) and local dependency in 11 item pairs. The person separation index was estimated as 0.48 suggesting weak discrimination between classes, whereas Cronbach's α was high at 0.86. Triangulation of qualitative data with the Rasch analysis supported recommendations for cross-cultural adaptation of the WHQ items 1 (redness), 3 (clear fluid), 7 (deep wound opening), 10 (pain), 11 (fever), 15 (antibiotics), 16 (debridement), 18 (drainage), and 19 (reoperation). Changes to three item response categories (1, not at all; 2, a little; 3, a lot) were adopted for symptom items 1 to 10, and two categories (0, no; 1, yes) for item 11 (fever).ConclusionThis study made recommendations for cross-cultural adaptation of the WHQ for use in global surgical research and practice, using co-produced mixed-methods data from three continents. Translations are now available for implementation into remote wound assessment pathways