Treatment decision tools have been developed in many fields of medicine, including psychiatry, however benefits for patients have not been sustained once the support is withdrawn. We have developed a web-based computerised clinical decision support tool (CDST), which can provide patients and clinicians with continuous, up-to-date, personalised information about the efficacy and tolerability of competing interventions. To test the feasibility and acceptability of the CDST we conducted a focus group study, aimed to explore the views of clinicians, patients and carers.The CDST was developed in Oxford. To tailor treatments at an individual level, the CDST combines the best available evidence from the scientific literature with patient preferences and values, and with patient medical profile to generate personalised clinical recommendations. We conducted three focus groups comprising of three different participant types: consultant psychiatrists, participants with a mental health diagnosis and/or experience of caring for someone with a mental health diagnosis, and primary care practitioners and nurses. Each 1-h focus group started with a short visual demonstration of the CDST. To standardise the discussion during the focus groups, we used the same topic guide that covered themes relating to the acceptability and usability of the CDST. Focus groups were recorded and any identifying participant details were anonymised. Data were analysed thematically and managed using the Framework method and the constant comparative method.The focus groups took place in Oxford between October 2016 and January 2017. Overall 31 participants attended . The main themes that emerged related to CDST applications in clinical practice, communication, conflicting priorities, record keeping and data management. CDST was considered a useful clinical decision support, with recognised value in promoting clinician-patient collaboration and contributing to the development of personalised medicine. One major benefit of the CDST was perceived to be the open discussion about the possible side-effects of medications. Participants from all the three groups, however, universally commented that the terminology and language presented on the CDST were too medicalised, potentially leading to ethical issues around consent to treatment.The CDST can improve communication pathways between patients, carers and clinicians, identifying care priorities and providing an up-to-date platform for implementing evidence-based practice, with regard to prescribing practices. The efficacy of current pharmacological and non-pharmacological interventions in psychiatry is well established and worthwhile, but still far from optimal . DespiteTo help clinicians adhere to evidence-based guidelines and deliver standardized care based on the best-available scientific information, medical algorithms have been developed, such as in the treatment of hypertension, diabetes, high cholesterol, cancer and myocardial infarction. Similarly, in psychiatry the Texas Medication Algorithm Project developed an algorithm to assess its value in managing the pharmacological treatment of patients with schizophrenia, bipolar disorder and major depressive disorder . Even thMoreover, for the algorithm to truly become a clinical tool, innovative methodologies are needed to support probabilistic decision-making and incorporate patients’ views and clinical judgement in a dynamic way . A diffeFor this reason, we have developed the prototype of a new, cloud-based clinical decision support tool (CDST), a treatment algorithm that aims at providing updated and stratified information about interventions for different subgroups of patients. To test the acceptability of the CDST, we designed a focus group study that aimed to explore clinicians’, patients’ and carers’ perspectives on the algorithm prototype. In particular, we wanted to explore whether participants felt it was a useful decision-making tool for improving prescribing practices in routine psychiatric care.The CDST was developed by researchers at the University of Oxford in collaboration with experts working at the University of Tel Aviv Fig.  and is pWe chose to use focus groups to investigate acceptability, to encourage participants to openly explore their own and others’ perspectives collectively, with opportunity for clarification and debate, without using a rigid framework. Three separate focus groups were planned: one for consultant psychiatrists working in the National Health Service (NHS) in the UK, one for patients and carers, and one for general practitioners and nurses. We conducted separate focus groups to allow participants to speak freely without feeling inhibited in the presence of other group members, thus facilitating discussion.We checked with the local Ethics Committee and ethical approvals were not required for the study, however written informed consent was taken from each participant.The study took place in Oxford (United Kingdom) between October 2016 and January 2017. We conducted three focus groups comprising of three different participant types. The first was for Consultant Psychiatrists working in the Oxford Health NHS Foundation Trust each of which received an invitation and participant information sheet via email. The second was for participants who had a prior mental health diagnosis and/or experience of caring for someone with a mental health diagnosis. They received a modified version of the participant information sheet in Plain English to ensure understanding. We managed to have a diverse representation of mental health disorders, including schizophrenia, major depression, bipolar disorder, anorexia and anxiety. Participants for this group were initially identified using a local Patient and the Public Involvement and Engagement (PPI/E) group, followed by purposive sampling to ensure diversity in age, gender, patient and carer experience and technological abilities. The third focus group comprised of primary care general practitioners and nurses, following invitation to participate from a local general practitioner (GP) practice.n = 6; nurses n = 5) and eight were patients or carers. The majority of participants were white British , women  and aged over 45 years   as well as clinicians who were less experienced (fewer than 10 years).Each focus group lasted around one hour and started with a short, interactive, visual demonstration of how the CDST worked, before answering any questions relating to the demonstration. To standardise the procedure, we used the same topic guide that covered themes relating to the acceptability and usability of the CDST Table .Table 2TFocus groups were recorded using a digital voice recorder before being transcribed by a local transcription service. Any identifying participant details were anonymised and the focus group recordings were then removed from the digital voice recorder and transferred to a secure, password protected storage facility in line with the universities’ policies on data storage and protection. Data were analysed thematically and managed using the Framework method . Transcrapplications in clinical practice, communication, conflicting priorities, record keeping and data management. Table The main themes that emerged related to CDST Whilst in general, there was more congruence than divergence in the perspectives of the consultant, primary care and patient and carer focus groups, the views of the groups were sometimes nuanced, based on their contextual needs, values, priorities and concerns. For example, whilst all three groups viewed the CDST as a useful and collaborative learning and decision-making tool, the consultant group expressed concern that it should not be viewed as a replacement for clinical judgement, whilst a prominent concern of Primary Care participants was that patients might become overloaded with worry if provided with too much information. However, the patients and carer group felt the amount of trust they had in their doctor would be a key factor in the shared decision making process. The study findings will now be reported on in more detail.I think it is a useful tool because we’re not perfect so we will forget things and we will also prescribe drugs with interactions that are going to be a real problem.” (Consultant 12).All participants agreed that the CDST has the potential for clinical applications in psychiatry. The majority reported that it was a useful tool to support decision making for prescribing. They felt the tool reminded clinicians to consider all available treatment options . They thought it prompted discussion between clinicians and patients around potential side-effects, best evidence and drug interactions: “For me it almost seems like a formalisation or a regularisation of what should be good practice anyway with the information in there.” (Consultant 12).Most consultants viewed the CDST as an effective way to standardise prescribing practices, by ranking interventions according to their comparative efficacy and side-effect profiles. This was seen as a more favourable method than relying on personally derived drug hierarchies, with some consultants admitting that their prescribing preferences were not always based on the most up-to-date evidence: “This is… a way of quickly reminding us…which [drugs] are better…I always hold in my head actually which one is the best for [patients with] QTc prolongation … It gives you an instant answer.” (Consultant 1).The tool was perceived to be useful by clinicians who had been practicing for a long time as well as clinicians who were relatively new to practice. All clinicians reported the CDST would be useful to help remember and retain information about the severity of different treatment side-effects across the different interventions, especially when there is little difference between certain drugs in terms of efficacy, boosting their prescribing confidence as a result: “You've asked the patient what's important to them…It's come up with that outcome based on that…It's a really useful learning tool for us to be thinking about what our prescribing choices are” (Primary Care 5).Primary care participants suggested that the CDST could inform prescribing practice by providing a history of patients’ previous side-effects. They also felt that the CDST provided a useful learning tool for clinicians if they selected a lower ranking drug than the recommended CDST treatment, by prompting them to question their rationale for doing so: “It helps in making informed decisions because 20 years ago people weren’t being told about side effects necessarily so they couldn’t even make important choices.” (Patient and Carer 7).Patients and carers described the CDST as ‘self-teaching’, helping them make informed choices and allowing them to readily access information than they previously did not have: “When we see someone… it changes completely the context of the conversation…Depending on how much time you have or depending on whether the person is having an attack or not which I did not capture in this [tool].” (Consultant 9).Some consultants were wary of the CDST being used as a replacement for clinical judgement and expressed concern about “becoming blinkered by it”. They reported the tool did not reflect the complex contextual clinical assessment process, being unable to capture detailed information about patient characteristics, time pressures, anxiety, influence of carers, clinician experience and organisational factors. Some clinicians said that this provided a significant limitation to the CDST’s usefulness: “Having this kind of information…Would make you more likely to go and look at the evidence for the drug that's being recommended if it's one that you're not used to.” (Primary Care 9).However, other clinicians thought that the CDST informed and enhanced clinical judgement, by providing unbiased, credible, evidence-based information, thus removing some of the reticence about prescribing drugs that were not routinely used in practice: “That’s what I like about the approach, it does assist judgement, you’re not committing to anything, you’re comparing bits.” (Patient and Carer 3).For patients and carers the CDST encouraged them to consider different factors during the clinical assessment process, rather than just focusing on one or two things: “All participants agreed that the CDST could be widely used for other chronic conditions where medications have to be tailored and adjusted over time such as other psychiatric conditions .How you explain some of these things to patients [will]…very much influence what weight they give to them.” (Consultant 8).One major benefit of the CDST was perceived to be that it opened up discussion about the possible side-effects of medications. Some clinicians commented that the CDST provided opportunity to fully discuss medications’ tolerability profile with patients, which they did not otherwise always do. This was important, as clinicians thought that the way they convey the severity of side-effects to patients is likely to influence their subsequent drug preferences: “‘minefield of discussion’.However, some primary care providers and patients/carers highlighted that there are disadvantages to receiving too much information about possible medication side-effects, as it might cause unnecessary worry and result in clinicians spending considerable time reassuring patients. Whilst it was important they were made aware of the main side-effects from treatment, the majority of patients and carers did not feel they needed to know every detail, so long as their doctor was well informed. This view was echoed by some consultants who reported there was a danger of entering into a Even somebody who’s quite psychotic might have a fairly good idea of what side effect profile they might prefer.” (Consultant 5).Most clinicians said that the CDST could help facilitate the informed consent process, though a few thought it might cause problems for patients with limited intellectual capacity, creating tensions if their preferred treatment choice was at odds with their clinician’s. However, generally it was felt that the CDST could optimise capacity, by teasing out patients’ views and preferences, though the degree of clinician/patient collaboration might vary: “‘series of tutorials’ to explain every acronym: “What concerns me is that it’s very easy for a clinician to lead you at that point if you don’t really understand what it is you’re looking at.” (Patient and Carer 4).Participants from all the three groups universally commented that the terminology and language presented on the CDST were too “medicalised”, making it difficult for lay people to understand and potentially lead to ethical issues around consent to treatment, as patients. There was consensus that simpler language was required and that clinicians needed a pre-determined knowledge of the demographic and level of understanding of patients, to avoid having to provide a The interface needs to be simplified and have lay terms and more explanation…To facilitate the conversation a bit more.” (Primary Care2).Each group made recommendations for improving the way the information was presented . This would facilitate discussions with patients and raise their awareness of different side-effect profiles: “The information must be there because if we’re to be transparent and it’s to be a co-production…To build trust and so forth. The patient should have access to the same information provided it doesn’t override the doctor’s duty of care to that patient.” (Patient and Carer 8).Showing the CDST’s screen to patients during a consultation was a matter of discussion. Many patients said that it could be distressing, causing confusion and uncertainty during the decision-making process. However, some patients and carers felt the CDST should be patient-facing to promote collaboration in decision-making, so long as this did not impinge on their clinician’s duty of care: “It would take quite a lot of knowledge to know which ones to move…It up the top.” (Primary Care 11).A couple of carers thought that having access to the CDST interface might lead to some patients manipulating the tool until they received their preferred drug of choice. However, primary care practitioners reported that patients would require a lot of knowledge for this to happen: “A doctor is not going to respond…well to somebody having done this and then say, actually, what I need is…”. (Patient and Carer 1).One patient commented that some clinicians may not respond well to patients having an increased level of control over the decision-making process, whilst others felt that trust in the clinician-patient relationship was vital for ensuring successful decision-making, something the CDST could not control for: “If you’re helping people to tailor the things that they prefer to avoid, I would hope that that would improve compliance.” (Primary Care 9).Primary care practitioners commented that the CDST could increase patient compliance if it was tailored to their needs: “What the patients come up with…They’ve said what they do or don’t want…Use that as a guide of what we should be thinking of using.” (Primary Care 5).Clinicians, patients and carers reported that prior to using the CDST patients should be encouraged to declare a checklist of their healthcare priorities, to assist with shared decision-making, ensuring mismatched priorities are identified and that their needs, views and perspectives are listened to. This was viewed as a valuable way of ensuring clinicians did not purely rely on their past prescribing preferences: “What would make [the tool] potent is if you could start to include more biological data about patients.” (Consultant 4).Many clinicians, patients and carers agreed that the CDST promoted personalised care. However, they recognised that the CDST was at times too disease-focused and could not compensate for the lifestyle and human factors that influence decision-making. The possibility of incorporating other data into the CDST such as genes and demographics was raised, to increase the CDST’s accuracy: “The cost effectiveness and the limited choice is very important…If something flashes up, potentially it can lead to some problems later down in the relationship that they're not getting the best thing.” (Primary Care 2).Other conflicting priorities were highlighted by primary care practitioners who suggested that increasing pressures on prescribing budgets meant the CDST should only identify medications available from within their own clinical commissioning groups. They commented that conflict could arise if patients felt they were being denied access to recommended CDST treatments which were not commissioned in their local area: “From the education point of view, then more information about more drugs is fine. Because you can go away and talk to your clinical commissioning group about why this one isn't available.” (Primary Care 9).However, from an educational perspective, it was thought helpful to have access to the full range of CDST recommendations, regardless of whether they were available from their clinical commissioning group. This could allow the commissioners to be challenged about why certain medications were not made available, as well as facilitating data collection to compare local prescribing practices against CDST recommendations: “It’s a highly selected group of patients ….” (Consultant 4).According to the majority of clinicians, patients and carers, the CDST could provide a good record of the consultation. However, suggested improvements included linking the CDST to patient clinical records’ database so that individuals’ medication histories could be incorporated. Participants praised the CDST’s ability to continually update its evidence-base, build on existing best available literature and present these outcomes in a quantitative and understandable way. However, some consultants expressed concern about the reliability of data from the network meta-analysis, commenting on the lack of external validity of randomised controlled trials: “This particular tool serves a specific purpose…It’s about deciding on how to treat them pharmaceutically….I think it’s fantastic.” (Patient and Carer 6).Finally, most participants thought that the CDST was easy to use, encouraging clinicians to use it on a daily basis. The CDST was commended for focusing on one specific aspect of the clinical consultation, saving time by collating relevant drug information within one database: “If it comes to enabling people to make a better choice, that's quite a major time saving rather than having them coming back saying, oh I didn't like it.” (Primary Care 11).A few primary care practitioners said that it could be hard to use the CDST within a 10 min consultation period, however they argued that this initial time investment would pay off long term: “The findings from this study provide useful insights into the perspectives of clinicians, patients and carers with regard to the usefulness of clinical support tools. They have led the research team to consider how the CDST might be optimised in the clinical setting to enable it to be mutually beneficial to clinicians and patients. The majority of participants felt the CDST could play a role in regulating and standardising prescribing processes in routine mental health practice.Previous research has shown that computer assisted guidance on drug prescribing can improve patients’ therapeutic outcomes, reduce costs and result in more effective care outcomes . This isAs the findings highlighted, the CDST is not intended to be a replacement for clinical judgement. The complexities surrounding the clinical assessment, management and treatment of mental health patients is multi-factorial. It depends on human, environmental, cultural, social, practical, physical and psychological factors and will not always fit neatly within the medical model of care or the CDST parameters. Although some clinicians expressed concern that the CDST may blinker doctors and stop them from considering other factors, we argue against this. Just as clinicians rely on summaries about patients’ social circumstances, comorbidities and test results to help build a comprehensive history, this background information does not inhibit them from listening and responding to the immediate needs of patients. Similarly, having access to evidence-based medication recommendations should not inhibit clinicians from employing their own clinical judgement. Rather, these recommendations form one piece of a complex jigsaw that requires knowledge, experience and expertise to complete .The findings revealed that clinicians, patients and carers placed value in the CDST’s ability to facilitate discussion about the side-effect profiles of different psychiatric drugs. A study carried out to ascertain whether patients knew their diagnoses, treatment plans and common side effects of prescribed medications, found that less than half were able to list their diagnoses, medication names and purpose, or associated medication side-effects . This suThe issue of informed consent and capacity is integral to decision-making in mental health. Research has shown that deficits in understanding among individuals with psychiatric illness or cognitive impairment can be improved with educational interventions, especially when they are presented in an organised and simplified manner . FurtherThe main limitation of this study is that patients vary substantially in how much they wish to contribute to the decision-making process . Whilst Finally, the CDST was generally considered to be an efficient, time-saving clinical tool and this is important given escalating pressures on healthcare resources making clinicians and patients aware of the brevity on their consultations . To thisThe CDST in the mental health and primary care settings can be a useful clinical decision support, with recognised value in promoting clinician-patient collaboration and contributing to the development of personalised medicine. The CDST can improve communication pathways between patients and clinicians, identifying care priorities and providing an up-to-date platform for implementing evidence-based practice. To further this aim, work is planned with PPI/E representatives to ensure that the medical terminology on the CDST is simplified to make it accessible to lay populations. Whilst the CDST has been designed to focus on prescribing practices within a mental health context, other evidence-based algorithms could be designed to incorporate factors such as demographic variations and non-pharmacological interventions, as well as a variety of mental and physical health disorders. Primary care practitioners may value the CDST for managing and treating patients with long term conditions such as hypertension and diabetes, within short clinic time slots. Primary care practitioners require a broader range of knowledge than practitioners in more specialised secondary and tertiary care services. The CDST may be particularly valuable in these circumstances, by providing up-to-date, evidence based information on a variety of health conditions in a timely manner, something that is essential given the time-limited consultations available.As well as providing a tool to support evidence-based practice, we plan to use the CDST to gather feedback from patients to increase its precision, by effectively incorporating everyday clinical information from ‘real-world’ individuals with the best available evidence that is contained within the CDST. This has the potential not only to improve treatment outcomes, but also to strengthen the evidence base around the interaction of specific drugs with other pharmacological and psychosocial interventions, whilst accounting for individual characteristics and patient preferences.Future research will also involve a pilot randomised controlled trial to test the CDST. If shown to be implementable in routine clinical practice in different settings, the CDST could lead to changes in prescribing practices of clinicians, protecting patient safety, whilst paving the way for precision medicine in every-day mental health care.


Hypophthalmichthys molitrix) and bighead (H. nobilis) carp (collectively bigheaded carp) are invasive fish that threaten aquatic ecosystems in the upper Midwest United States and the Laurentian Great Lakes. Controlling bigheaded carp is a priority of fisheries managers and one area of focus involves developing acoustic deterrents to prevent upstream migration. For an acoustic deterrent to be effective however, the hearing ability of bigheaded carp must be characterized. A previous study showed that bigheaded carp detected sound up to 3 kHz but this range is narrower than what has been reported for other ostariophysans. Therefore, silver and bighead carp frequency detection was evaluated in response to 100 Hz to 9 kHz using auditory evoked potentials (AEPs). AEPs were recorded from 100 Hz to 5 kHz. The lowest thresholds were at 500 Hz for both species . These results provide fisheries managers with better insight on effective acoustic stimuli for deterrent systems, however, to fully determine bigheaded carp hearing abilities, these results need to be compared with behavioral assessments.Silver ( Hypophthalmichthys molitrix) and bighead (H. nobilis) carp (collectively bigheaded carp) are invasive to the Mississippi River Drainage and these prodigious filter feeders threaten native species )To determine the relative amplitude for the FFT analyses, the raw voltages (μV2) were normalized based on the highest FFT peak. A repeated measures ANOVA with a Holm-Sidak test was used to compare the sound pressure and acceleration thresholds for all three species at each frequency examined using SigmaPlot (version 12.5). All threshold data were normally distributed (Shapiro-Wilk P > 0.05) and are reported as mean ± 1 SD.rms for all experiments and the baseline particle acceleration level was -96.0 dB re 1 ms-2. The acoustic impedance at all three sound pressure levels examined indicates that there were no major resonances in the tank at the test frequencies  axis had the highest mean particle acceleration across all test frequencies compared with the x (-54.0 ± 9.0 dB re 1 ms-2) and y (-58.7 ± 6.2 dB re 1 ms-2) axes. rms. Auditory evoked potentials were recorded for all carp species from 100 Hz to 5 kHz and the waveforms were similar across species type at each frequency. Figs The ambient sound pressure level was 70 dB re 1 μPa SPLquencies . At 130 rms; rms; rms; rms; rms; 4,38 = 70.46; P < 0.05) thresholds than bighead and common carp at 500, 600, 700, 900, 1000, 1500, and 3000 Hz  mean acceleration thresholds at 500, 600, 700, 900, 1000, 1500, and 3000 Hz  in outdoor concrete ponds but habituated to these pure tone sounds after a few presentations [The need for investigating the upper range of bigheaded carp hearing originates from behavioral studies examining silver and bighead carp responses to pure tones (500–2000 Hz) and broadband sound 0.06–10 kHz) recorded from an outboard motor. Both species showed negative phonotaxis to pure tone stimuli  The tuning curves from the present study and those reported by Lovell et al.  demonstrWhile comparisons between AEP thresholds generated using different setups can be difficult , severalIn addition to reporting AEP-derived threshold curves for the three carp species in regards to sound pressure, the tuning curves for acoustic particle motion were also determined. For all three species, the lowest mean particle acceleration thresholds were at 400 Hz. It is believed that all fish are capable of detecting acoustic particle motion through direct stimulation of the otoliths , 34. HowRecent reviews comparing the AEP and behavioral paradigms have concluded that while the two methods yield similar frequency ranges, the thresholds vary greatly, even among AEP studies , 21. Therms, as other AEP studies have shown that ostariophysans can detect up to ~8 kHz [rms) at frequencies between 6–9 kHz, small peaks on the FFT at two times the stimulus frequency were observed for some fish at 6, 7, 8, and 9 kHz, but these peaks appeared inconsistently and did not meet the established AEP criterion of the present study. However, given the constraints of tank size and speaker output, generating sound pressure levels above 150 dB re 1 μPa SPLrms were not possible. Additional research could examine sensitivity to frequencies > 5 kHz at sound pressure levels above 150 dB re 1 μPa SPLrms in both carp species.Finally, it is possible that bigheaded carp can hear frequencies above 5 kHz at sound pressure levels > 150 dB re 1 μPa SPLo ~8 kHz . When exThe results of the present study provide important insight on the upper range of silver and bighead carp hearing, as they indicate higher frequency hearing than has been previously reported. Together with findings that bigheaded carp behavior can be modified using broadband sound, this research will aid in developing an effective acoustic deterrent. Particularly, the conclusion that bigheaded carp hearing extends up to 5 kHz could allow for refinement of the broadband stimulus to target bigheaded carp. Further research may allow for development of an effective acoustic deterrent primarily comprised of frequencies above the hearing range of non-ostariophysans, however care must to be taken to avoid disturbing native cyprinids.


Ultra-widefield angiography is the latest technology in the evolution of fundus fluorescein angiography. With the ability to capture up to 200° of the fundus in a single image, far peripheral retinal pathology can be imaged. Generally, obtaining high-quality fundus fluorescein angiography in a child without sedation in the outpatient setting is exceedingly challenging. Therefore, there are advantages to imaging platforms that can capture the peripheral retina in young children without anesthesia. Often pediatric retinal diseases have pathology localized to the far periphery, which further validates the utility of ultra-widefield angiography. Ultra-widefield angiography has been successfully used without sedation for evaluation of children with various pediatric retinal diseases such as Coats disease, familial exudative vitreoretinopathy, and retinopathy of prematurity.This non-contact, non-mydriatic modality has been utilized in the evaluation of pediatric retinal diseases and demonstrated to have benefits over conventional fluorescein angiography techniques. For nearly six decades, fluorescein angiography has been a diagnostic staple in the diagnosis and management of vitreoretinal diseases, graphically demonstrating information regarding circulations of the retina, choroid and optic nerve head, and barrier integrity of the neurosensory retina and retina pigment epithelium. Capturing quality angiographic images of the peripheral retina with a 30°–60° field of view of most standard fundus cameras proved difficult . This reTechniques were developed to create montages of images from several fields captured by fundus photography to widen the field of view capable with standard 30 or 50 degree cameras. The 7 Standard Fields (7SF) protocol for fundus photography was developed and popularized by the Diabetic Retinopathy Study group expanded visualization to approximately 75° with 3 photos horizontally across the macula and 4 photos surrounding the optic disc . HoweverWidefield angiography, capturing > 30° to < 200° , was devThe Optos camera  was the first ultra-widefield imaging system, which produces a 200° view of the retina (about 82% of the surface area) , 13. In As discussed above, in the pediatric population, imaging of the peripheral retinal often required general anesthesia. Initially, standard 30° or 45° cameras were used and required positioning of the eye to the periphery in children who were unable to cooperate. Contact cameras were helpful but also required general anesthesia. However, with UWFA, there is the ability to obtain quality images in some children without the need for general anesthesia. We describe several conditions below in which UWFA has provided adequate images for diagnosis and treatment without the need for general anesthesia. The PubMed database was systematically searched using the search terms [(pediatric or pediatric retina or Coats or familiar exudative retinopathy) and (fluorescein angiography or widefield imaging or ultra-widefield imaging)]. To our knowledge, all articles or abstracts describing the use of ultra-widefield fluorescein angiography in the pediatric population were included in this review Table .Table 1SFamilial exudative vitreoretinopathy (FEVR) has a variable presentation, including straightened retinal vessels, temporal dragging of the macula, abnormal retinal vascular permeability Fig. , describIn FEVR, myopia is present and family history should be sought as genetic mutations are recognized in about 50% of cases, but the condition is variable in expressivity so even a normal parent may pass onto offspring a mutation manifesting as disease. Many of the early findings of FEVR occur in the far peripheral retina making the diagnosis challenging. The findings are best appreciated with fluorescein angiography that demonstrates capillary non-perfusion and leakage of the vascular front in the region between vascularized and avascular retina. There is also intravitreal neovascularization in Stage 2 FEVR . Contact Retcam imaging has identified peripheral areas of nonperfused retina in the fellow eye of patients with Coats disease, which is more than three standard deviations greater than the < 1 disc diameter of non-pathologic, nonperfused, temporal retina in normal, young children . Other v0%. ContaUWFA has been extensively utilized in the evaluation and management of Coats disease , 19, 20.Incontinentia pigmenti is an inherited X-linked dominant disease, mainly seen in females as it is lethal in males. Avascular retina is seen and vessels can develop and remodel normally but if intravitreal neovascularization occurs, there is a risk of vision loss from vitreous hemorrhage and tractional retinal detachment. Vision loss also occurs from avascularity in the macula . Laser tRetinopathy of prematurity (ROP) is a leading cause of blindness and vision loss worldwide in premature infants. ROP is identified by screening infants at risk and monitoring until physiologic vascularization has extended to the ora serrata. Type I ROP is treated with laser photocoagulation or intravitreal anti-VEGF agents. In the past, fluorescein angiography was not standard care for ROP , 24. HowUWFA has also been successfully used in the evaluation of children with X-linked retinoschisis, Stargardt disease, Best’s disease, toxoplasmosis chorioretinitis, juvenile sarcoidosis, choroidal melanoma, juvenile idiopathic arthritis, pars planitis, and traumatic retinal detachment . Ages raUltra-widefield fluorescein angiography has established itself as a useful tool to the vitreoretinal specialist and is proving to be valuable in the care of pediatric retina patients. There is mounting evidence that UWFA may be a valuable adjunct to identify peripheral retinal pathology undetected by the conventional 75° field from 7 Standard Fields in the evaluation of pediatric retinal diseases . There iHowever, there are definite limitations of UWFA. Poor image quality, distortion, and artifact may affect up to 5–10% of images in some studies , 34. NatUltra-widefield angiography is a valuable adjunct to traditional forms of fluorescein angiography as the technique proves to be less invasive and better tolerated in pediatric patients. The greater ability of UWFA to identify peripheral retinal pathology over conventional angiography platforms is especially suited to pediatric retinal conditions such as Coats disease and FEVR. The future will hold better and less invasive methods of imaging the peripheral retina vasculature such as ultra-widefield optical coherence tomography (OCT) angiography . Further


A growing number of students with hearing loss are being granted access to higher education in South Africa due to the adoption of inclusive educational policies. However, available statistics indicate that participation by students with hearing impairments in higher education remains low and research suggests that support provisioning for those who do gain access is inadequate.This article aims to illustrate that the assumed self-identity of students with hearing impairment influences their choice to disclose their disability. The choice not to disclose their hearing loss prevents them from accessing the necessary reasonable accommodations and this in turn may affect their eventual educational success.Reported here is a qualitative descriptive case study at a South African university. Purposive sampling methods were employed. Data were gathered from in-depth interviews with seven students with hearing impairment ranging from moderate to profound, using spoken language. Constructivist grounded theory was used as an approach to the process of generating and transforming the data, as well as the construction of theory.All the student participants identified as having a hearing rather than a D/deaf identity cultural paradigm and viewed themselves as ‘normal’. Linked to this was their unwillingness to disclose their hearing impairment and thus access support.It is crucially important for academic, support and administrative staff to be aware of both the assumed ‘hearing’ identity and therefore subsequent non-disclosure practices of students with a hearing impairment using the oral method of communication. Universities need to put measures in place to encourage students to voluntarily disclose their hearing impairment in order to provide more targeted teaching and learning support. This could lead to improved educational outcomes for students. Hearing impairment is recognised as a global pandemic  as a primary medium of communication, who use various means of communication and assistive hearing technologies. These include speech, speech/lip reading, hearing aid systems, cochlear implants, bone anchored hearing aid and applicable assistive listening devices, etc., or a combination thereof Nair .The issue of identity is crucial for academic success as students’ self-perception and their perception of how others view them play a pivotal role in students’ interactions with both institutional processes and structures, which has important implications for accessing support through personal disclosure as well as teaching and learning experiences, and possibly educational outcomes. Due to the invisibility of hearing impairment, the assumed identity of the student with a hearing impairment is open to perpetual negotiation . By avoiding confronting their hearing impairment through self-disclosure and seeking reasonable accommodations, these students may not be able to enjoy full and equal participation in academic life Hindhede .This article aims to illustrate that the assumed self-identity of students with hearing impairment influences their choice to disclose their disability. This choice may in turn negatively affect their eventual educational success. It is important for academic, support and administrative staff to be aware of both the assumed identity and non-disclosure practices of students with a hearing impairment using the oral method of communication in order to put measures in place to encourage disclosure and to provide more targeted teaching and learning support.The next few sections of this article will focus firstly on the theoretical perspectives and then the empirical study. The theoretical aspects are:an exploration of the constructs of disability as these constructs tend to frame our thinking and thus our responses to persons with disabilities;the two main ways in which deafness is conceptualised;how the assumed identity of a student with a hearing impairment as either Culturally Deaf or Hearing contributes to positive aspects of his or her social identity and finally;how this can be measured via the Deaf Identity Development Scale (DIDS)  and note-takers. As a move away from the previously dominant medical model , the social model of disability (where persons with disabilities are ‘disabled’ by society) is firmly entrenched (more in words than in practice) in higher education in South Africa (CHE 2005). Although support services for students with disabilities are currently based on this model, it seems that Reindal’s social-relational model 2008) could be more suited to inclusion as it retains the main tenets of the social model of disability, such as the effects of impairment and the phenomenon of disability, elaborated as a ‘social relational phenomenon’ but it also maintains the perspective of ‘oppression and discrimination’ in contrast to ‘disadvantage due to restriction of activity’ , but additionally, his or her personal experiences of being a person living with reduced function plus discrimination and oppression would be encapsulated Bell . The finIn society, there are two main constructions of deafness: one construes ‘deaf’ as a category of disability while the other construes ‘Deaf’ as being a member of a linguistic minority group with its associated culture Lane . Large dAccording to social identity theory Tajfel , an indiet al.  at a young age, which significantly impacts language development, but this is regionally dependent. Thus, student communities from the different universities in South Africa would differ significantly.A purposeful sampling procedure Patton  was usedhad to have hearing impairments, regardless of the degree of hearing impairment or the age of onset;needed to be registered students at the selected university (either undergraduate or postgraduate); andhad to make use of spoken language (either English or Afrikaans as their home language) rather than Sign Language.With the required permission obtained, students who had disclosed their hearing loss to the disability unit at the selected university were invited by email to participate in the research. Seven out of a possible 13 students volunteered to participate in the study refer to .In-depth interviews were used to generate the data. I arranged an initial meeting with each student, a ‘get-to-know-each-other’ session, to build a relationship. At this meeting, I explained the nature and aim of the research project, requested their participation and asked them to complete a biographical questionnaire, which provided a large amount of background information, for example, type of hearing loss, age of onset, use of assistive technologies. A follow-up meeting was scheduled with each student at which the individual in-depth interview took place.The interviews were conducted in a quiet location on campus, to facilitate barrier-free communication. An interview guide refer to  was desiBOX 1Interview guide.Interviews were transcribed and then ATLAS.ti was used to code the data, form categories and themes and build network views Charmaz . I folloInitial coding phase: involves naming each word, line or segment of data.Focused coding phase: uses the most significant or frequent initial codes to sort, synthesise, integrate and organise large amounts of data.Theoretical coding phase: a sophisticated level where the theoretical codes specify possible relationships between categories developed during the focused coding.Memos (within ATLAS.ti) were used throughout the data transformation process to assist with data interpretation while transforming the data. The measures used to ensure trustworthiness of the data were crystallisation ‘I really don’t see myself as being disabled … I have never been treated as someone who is deaf … and with me they won’t see immediately, they will assume that I am a normal person. That is how it has always been.’ (Merle)I would assert that there are three main reasons for the participants in this study assuming a hearing identity: oral communication tradition, previous experience and invisible nature of hearing equipment. The first reason could be that they come from an oral communication tradition where lip- and speech-reading as well as the use of their residual hearing are valued. Except for one student, all seven participants came from hearing families. The participants also only generally interacted with hearing friends and peers, feeling that their self-identity should depend on personal rather than audiological definitions, and consequently contact with hearing peers was valued Leigh . In suchThis characteristic of normalisation was a recurring theme throughout the data. The participants did not want to be seen as ‘different’, but rather viewed themselves as normal hearing university students. Noelene expressed her self-identity as follows:‘… and with me they won’t see immediately, they will assume that I am a normal person. That is how it has always been … I have always been a normal student.’ (Noelene)The stigma of being labelled as disabled was strongly rejected by the participants. Astrid explained her experiences at school as positive as she was not viewed as being disabled, while Paul shared how he tried to cope on his own without disclosing his hearing impairment as he did not want any special concessions. The comments of this participant illustrate the rejection of labels.‘I had a lot of teachers that would never treat me as a disabled student in the first place, but they would also forget about it [my hearing impairment] sometimes and when I think back now that was really good – that you are not different from anyone else, so just get on with it [life].’ (Astrid)Part of their hearing identity was taking on the responsibility to fit in and cope in a hearing world in order to gain employment and eventually be economically independent. One of the participants (Barry) mentioned that he viewed the use of oral communication as extremely important and that the use of Sign Language was severely limiting due to special schools being under-resourced and therefore often viewed as having lower academic standards than mainstream public and private schools. None of the seven participants had ever been exposed to Deaf culture or Deaf communities and felt that the use of Sign Language was not supported in higher education or society at large and was therefore never an option for consideration. It also seemed that in a ‘hearing’ academic setting, which does not support the use of SASL, students with hearing impairment do not have a choice, in any event, but to assume a Deaf identity.The second possible reason for the participants having a hearing identity is previous experience, namely the fact that they all came through mainstream primary and secondary schools, except for one student who attended a special primary school for learners with hearing impairment. In the mainstream environment, they reported that they were not treated as learners with a disability.Thirdly, because of the often ‘invisible’ nature of a hearing impairment, especially for girls with long hair covering their hearing aids (speech processors), students with hearing impairment are often seen as ‘normal’ as the sensory impairment is not easily visible. In some cases, participants reported that they purposefully hid their hearing instruments in order to avoid being labelled and stigmatised as ‘deaf’ or ‘disabled’. If is often for this same reason, namely to remain ‘invisible’, that students with hearing impairment refuse to make use of any assistive listening devices which could make them ‘identifiable’ or ‘extra-visible’ Goode .The issue of identity is critical as students’ self-perception/self-identity and their perception of how others view them play a pivotal role in their interactions with both institutional processes and structures, and this may have important implications for their academic success. For example, if a student with hearing impairment has a self-perception of being ‘normal’ or non-disabled, then his or her interaction with the institutional processes will be as a hearing student, without disclosing or requesting any learning support, potentially resulting in poorer academic results. The outcomes of this study support the findings by Hindhede , indicatDisclosure of hearing impairment refers to the willingness The participants also referred to their hearing impairment as being ‘not so visible’ and they reported that friends often did not realise that they have a hearing impairment due to its ‘invisibility’. As Merle said:‘Very few of them [friends], because it is not so visible, many people don’t know, some of my friends don’t even know.’ (Merle)Some disclosed their hearing impairment to their lecturers, but generally only if compelled to do so through circumstances, and one of the participants deliberately used his ‘disabled’ status to secure a place in the university residence. Disclosure at university seems to be a reactive action in most cases, as expressed by Noelene:‘I never went to tell them of my disability … I feel it is not necessary to tell them unless I have a problem.’ (Noelene)This phenomenon also made it difficult in the early phases of the research study to identify the student participants for this investigation.The most logical explanation for participants’ non-disclosure could be linked to their ‘hearing identity’ and their rejection of being labelled or stigmatised as ‘disabled’ Watson . In SoutIt was clear that the participants in this study did not want to define themselves or their relationships with others based on their hearing impairment. They tried to assimilate as much as possible into the hearing world, identified as culturally hearing, and thus chose not to disclose their hearing loss.Despite increased access for students with hearing impairment into higher education, such students generally remain under-represented, unaccounted for and under-supported; experiencing many barriers and having to develop personalised coping strategies. One reason for this could be linked to their self-identity as the issue of assumed identity plays a crucial role in their personal choice to disclose (or not disclose) their disability which in turn affects their access to teaching and learning support.All the participants in this study came from hearing families and they made use of spoken language. Factors such as choice of communication, family environment, attending mainstream schools and a focus on the person as opposed to the impairment led the students to assuming a hearing cultural identity rather than a D/deaf cultural identity and electing not to disclose their hearing loss for fear of stigmatisation. In this way, they believed that other people would see them as normal and not as disabled. In addition to assuming a hearing identity, the participants did not self-advocate in order to negotiate for their communication needs (full accessibility in the teaching and learning environment). This is directly related to their hearing self-identity and the level of importance they attached to being invisible as opposed to extra-visible and drawing attention to themselves. The need to blend in and to be seen as normal seemed to be high. Their personal choice of non-disclosure may be detrimental to their academic success as it limits the extent to which the university/disability unit is able to provide appropriate support. If the communication barriers that they experience in the teaching and learning environment become insurmountable or if their personalised coping strategies are insufficient or ineffective, this may lead to poor academic outcomes, affecting their future economic well-being.These findings offer practical value for the individual, the disability unit staff and the university as a whole:Suggestions for the individual with a hearing impairment:Develop and make use of skills to self-advocate by informing significant role players in their university education of their hearing impairment and be able to negotiate for the necessary accommodations.Build effective relationships with lecturers, peers and staff from the disability unit to be able to interact appropriately and confidently and negotiate communication access.Suggestions for the disability unit staff and the university:Communicate to students with hearing impairment the benefits of and reasons for disclosure.Ensure that students are fully informed regarding all aspects of supporting the communication and accessibility needs of students with hearing impairment, including up-to-date knowledge of the latest available assistive technologies, and remain sensitive to their needs.Ensure that students with hearing impairment have as much knowledge as possible (concerning their rights as students with disabilities and the availability of support) to assist them to make good decisions about their communication and other support needs.Have in place clear institutional guidelines concerning disclosure and confidentiality.Make available financial and human resources to support the access needs of students with hearing impairment.The education of students with hearing impairment, using the oral approach for communication at mainstream schools, colleges and universities in South Africa is an under-researched area. The following recommendations for future research are suggested:to identify the skills, strategies and awareness necessary for increased disclosure and self-advocacy andto identify the specific types of support that students with hearing impairment are receiving at universities throughout South Africa, as well as their experiences, both negative and positive, in relation to these.It is of vital importance that students with a hearing impairment who have a hearing cultural identity be taught the skill of self-advocacy and self-representation from an early age so that they become comfortable and confident in disclosing their disability and not feel ashamed or stigmatised. In this way, they also learn to self-advocate for their communication accessibility needs, not having to rely on their personal coping strategies but on the support  provided by the institution, the disability unit, their lecturers, their tutors and their peers, which could lead to improved educational outcomes.


Disassembly of the Cdc45-MCM-GINS (CMG) DNA helicase is the key regulated step during DNA replication termination in eukaryotes, involving ubiquitylation of the Mcm7 helicase subunit, leading to a disassembly process that requires the Cdc48 “segregase”. Here, we employ a screen to identify partners of budding yeast Cdc48 that are important for disassembly of ubiquitylated CMG helicase at the end of chromosome replication. We demonstrate that the ubiquitin-binding Ufd1-Npl4 complex recruits Cdc48 to ubiquitylated CMG. Ubiquitylation of CMG in yeast cell extracts is dependent upon lysine 29 of Mcm7, which is the only detectable site of ubiquitylation both in vitro and in vivo (though in vivo other sites can be modified when K29 is mutated). Mutation of K29 abrogates in vitro recruitment of Ufd1-Npl4-Cdc48 to the CMG helicase, supporting a model whereby Ufd1-Npl4 recruits Cdc48 to ubiquitylated CMG at the end of chromosome replication, thereby driving the disassembly reaction.  •Ufd1-Npl4 are required for CMG helicase disassembly during DNA replication termination•Ufd1-Npl4 recruit Cdc48 to ubiquitylated CMG helicase•Budding yeast CMG helicase is ubiquitylated on K29 of its Mcm7 subunit•Ubiquitylation of Mcm7-K29 is required in vitro for recruitment of Ufd1-Npl4-Cdc48 Disassembly of the CMG helicase is the key regulated step during DNA replication termination in eukaryotes, driven by CMG ubiquitylation and the Cdc48 segregase. Maric et al. find that the Ufd1-Npl4 heterodimer is essential for CMG disassembly in budding yeast and is required to recruit Cdc48 to the ubiquitylated helicase. Regulated unwinding of the parental DNA duplex ensures that eukaryotic cells make a single copy of their chromosomes during each cell cycle . The essDia2 is required to ubiquitylate CMG at the end of chromosome replication . Correspondingly, inactivation of the Ufd1-Npl4 heterodimer led to accumulation of the CMG helicase with ubiquitylated Mcm7 subunit .Whereas CMG disassembly was partially defective when synchronized cultures of  subunit D and S2C subunit . These fth dia2Δ E, mirrorutations . Moreoveth dia2Δ E. In conDia2-dependent ubiquitylation of CMG in yeast cell extracts . These data indicate that the Ufd1-Npl4 heterodimer is required for recruitment of Cdc48 to the CMG helicase in budding yeast.To establish that the Ufd1-Npl4-Cdc48 complex plays a direct role in CMG disassembly, we examined the association of all three factors with the helicase, using an in vitro system for SCFextracts . Whereasextracts A and 2B, S phase C. As desDia2-dependent ubiquitylation by mass spectrometry, we first developed a strategy for enrichment of ubiquitylated Mcm7 from the isolated CMG complex after in vitro ubiquitylation of the helicase in extracts of S phase cells .Whereas degradation of Mcm7-td was sufficient to block CMG assembly in the subsequent S phase A. In conDia2-dependent ubiquitylation of the 11-subunit CMG helicase is remarkably specific both in vivo and in yeast cell extracts.Lysine 29 is located within the longest tryptic peptide of Mcm7, probably contributing to the initial difficulty in detecting the sites of Mcm7 ubiquitylation by mass spectrometry. Therefore, we combined trypsin with the endoproteinase GluC that cuts after glutamate residues to reduce the size of the cleaved peptide containing K29. After partial purification of Mcm7 from isolated CMG helicase that had been ubiquitylated in yeast cell extracts A, subseqMCM7 locus was modified in order to introduce the mcm7-K29A or mcm7-K29R mutations (see MCM7), this association was greatly diminished when in vitro ubiquitylation of CMG was blocked by mutation of K29 of Mcm7 (mcm7-K29A). These findings indicate that ubiquitylation of Mcm7 plays an important role in recruiting Ufd1-Npl4-Cdc48 to the CMG helicase.Mapping and mutating the site of Mcm7 modification allowed us to explore in vitro the role of ubiquitylation in recruitment of Ufd1-Npl4-Cdc48 to the CMG helicase. We generated haploid yeast strains in which the endogenous ions see . In extrions see B, MCM7, mcm7-K29A cells complete S phase . Moreover, in vivo ubiquitylation of Mcm7-K29A occurs with comparable efficiency to CMG ubiquitylation on lysine 29 of Mcm7 in wild-type cells (MCM7 with mcm7-K29A). Interestingly, therefore, the regulation of CMG ubiquitylation ensures that it is highly specific in wild-type yeast cells, being limited to a single lysine residue on Mcm7, and yet is still able to switch elsewhere on Mcm7 with high efficiency when K29 of Mcm7 is mutated . Ubiquitylation of CMG in yeast cell extracts is more constrained, at least under our in vitro conditions, such that K29 of Mcm7 is the only possible acceptor site for ubiquitin.Lysine 29 of Mcm7 is the only detectable site of CMG ubiquitylation in wild-type yeast cells both in vivo and in vitro , and mutmcm7-K29A cells complete S phase  appears to be functionally important during DNA replication termination, because CMG disassembly is highly efficient when  S phase F and S4G2Δ cells , such as2Δ cells H.Dia2  when they meet the 5′ end of the opposing fork’s lagging strand, as CMG disassembly has been shown to occur after all DNA intermediates have been ligated and DNA synthesis is complete . The chahila CMG  indicateDia2.It remains to be determined how ubiquitylation of CMG is restricted to a specific lysine residue in wild-type cells, and yet is able to switch in vivo (but not in cell extracts) to other sites upon mutation of lysine 29 of Mcm7. It will be important in future studies to map and mutate the alternative sites of ubiquitylation of Mcm7-K29A. The ultimate goal would be to create a non-ubiquitylatable version of the CMG helicase, in order to test directly the significance of CMG ubiquitylation for helicase disassembly at the end of chromosome replication and explore the importance of CMG ubiquitylation for genome integrity in yeast cells that still contain active SCFTET-shp1-aid allele was constructed using a modified version of the auxin-inducible degron system  containing 2% glucose (YPD). For the cultures involving  strains , cells wdia2Δ cells, as well as for ufd1-2 and npl4-1 cells, the additional supplements of alpha factor were increased to 5 μg/mL.Other yeast cultures were grown at 24°C or 30°C. In order to arrest cells in the G1 phase of the cell cycle, the mating pheromone alpha factor  was initially added to cultures at 7.5 μg/mL and then supplemented by an additional 2.5 μg/mL every 20 min after 60 min from initial incubation (for cultures grown at 30°C or every 30 min after 90 min if the cultures were grown at 24°C) until 90% or more of cells formed “shmoos”. For GAL-UBR1 in YPGal medium, cells were washed into YPGal medium containing 10 μg/mL alpha factor, and additional aliquots of 2.5 μg/mL alpha factor were then added after 30 min and then again every 15 min.For the experiments shown in To arrest cells in early S phase, cells were grown in the presence of 0.2 M hydroxyurea (HU).Degradation of the Shp1-aid protein .For the experiment shown in The content of DNA in cells was monitored by flow cytometry of fixed cells stained with propidium iodide . Flow cyThe isolation of protein complexes from budding yeast cell extracts was performed as described previously . In the After in vitro ubiquitylation in low-salt extracts (100 mM potassium acetate) or in vivo ubiquitylation after inactivation of Cdc48-aid , we isolated the Mcm7 subunit of the CMG helicase using the two-step purification procedure that we described previously .Peptides were extracted using the in-gel tryptic digestion protocol described previously . The extm/z 300–1,800) were acquired in the Orbitrap with resolution R = 70,000 at m/z 200 . The ten most intense ions were fragmented by “higher energy collisional dissociation” . Precursor ions of unassigned charge, +1, or >+8 charge were excluded from the MS/MS analysis. The intensity threshold was set to 2 × 104. To obtain higher quality spectra of GlyGly-K-containing peptides, further MS runs were performed on the doubly digested peptides (trypsin + GluC) using the same parameters as described above, with the exception of selecting only the top seven peptides for MS/MS analysis at a resolution of 35,000, with a target value of 1,000,000 ions or a fill time of 150 ms.Mass spectrometry analysis was performed by liquid chromatography-tandem mass spectrometry (LC-MS/MS)  equipped with an integrated nano-electrospray  coupled to a nano-uHPLC system . Peptides were fractionated on a 50 cm × 75 μm internal diameter, PepMap RSLC C18, 2 μm reverse-phase column heated to 45°C. For initial analysis, between 2 and 6 μL peptide samples were loaded and fractionated over a 60-min gradient. Data were acquired in the data-dependent mode. Full scan spectra  for peptide masses and 0.5 Da for MS/MS peaks. The minimum peptide length was set to seven amino acids and a maximum of two missed cleavages (Trypsin-P) or three missed cleavages (Trypsin-P + Glu-C). A 1% false discovery rate (FDR) was required at both the protein and peptide level. In addition to the FDR threshold, proteins were considered as “identified,” if they had at least one unique peptide. Protein identifications and intensity values based on extracted ion chromatograms were reported for each slice from each gel lane. Spectral annotations were performed automatically in MaxQuant.All the raw MS data files were analyzed together using the quantitative MS software MaxQuant  incorporhttps://mrcppureagents.dundee.ac.uk) at the University of Dundee.Polyclonal antibodies for the detection of replisome components and Cdc48 by immunoblotting were described previously . PolycloM.M. performed all experiments except for the following: P.M. performed the experiments in


Early short-term intensive insulin (STII) therapy can induce drug-free glycemic remission for up to 1 year in half of newly diagnosed type 2 diabetic mellitus (T2DM) patients. Whether exenatide following STII therapy will induce higher long-term glycaemic remission is currently unknown. To assess the effect of STII+ exenatide therapy, compared with STII only, on maintenance of glycaemic remission in newly diagnosed T2DM patients. In this randomized, parallel-group, open-label, controlled trial, 129 patients (66 in STII+ exenatide group and 63 in STII only group) firstly completed 3-week STII therapy, then STII+ exenatide group was treated with exenatide for 12 weeks further. The cumulative probabilities of 1-year and 2-year glycaemic remission in STII+ exenatide group were 68.2 ± 5.7% and 53.0 ± 6.1%, which were significantly higher than STII only group (36.5 ± 6.1% and 31.8 ± 5.9%) . Patients in STII+ exenatide group, compared with STII only group, showed significantly decreased levels of waist  cm v.s. 84.2  cm, p = 0.048) and HbA1c % v.s. 6.49 %, p < 0.001) after 12-week exenatide treatment, but these differences disappeared after 1-year and 2-year follow-up. As conclusions, Improved effect of sequential exenatide after STII therapy on maintenance of glycaemic remission only occurred during exenatide treatment and lost upon treatment cessation. Increasing data have shown that the early use of short-term intensive insulin (STII) therapy can improve β-cell function and induce long-term glycaemic remission in patients with newly diagnosed T2DM without further antidiabetic medication, and thus STII therapy has become a strategy of interest5. In a meta-analysis, pooling data from four studies showed that early STII therapy in the newly diagnosed T2DM patients resulted in 46% of patients remaining in 1-year drug-free remission6. Despite these promising results, more than half of patients receiving STII therapy will suffer hyperglycaemic relapse within 1 year.Pancreatic β-cell dysfunction is the critical pathologic process resulting in the progressive deterioration of glucose control and the complications in the natural history of type 2 diabetes mellitus (T2DM)7. However, the generally applied STII therapy lasting 2–3 weeks is limited to correct these pathophysiological deficiencies for a long term. Therefore, new therapeutic approaches after STII therapy in the newly diagnosed T2DM patients which may improve longer term glycaemic remission are eagerly awaited.The ideas of early STII therapy inducing long-term drug free remission in the newly diagnosed T2DM patients are to reverse β-cell deficiency and insulin resistance by rapid correction of glucotoxicity and lipotoxicity11. Whether exenatide following STII therapy, compared with STII only therapy, will induce higher long-term glycaemic remission rate in newly diagnosed T2DM patients is currently unknown. Available evidence also shows that after stopping GLP-1RA treatment, their enhancement effects on β-cell function will disappear14; however, there is no evidence available on whether their effect on glycaemic remission will lose upon cessation of GLP-1RA therapy. We therefore conducted a randomized parallel-group trial with the primary aim to assess the effect of STII+ exenatide therapy, compared with STII only therapy, on maintenance of 1-year and 2-year glycaemic remission rates in newly diagnosed and drug-naive T2DM patients in China. We also aimed to test if the effect of exenatide after STII therapy on maintenance of glycaemic remission will lose upon cessation of exenatide therapy.Exenatide is the first-in-class glucagon-like peptide-1 receptor agonist (GLP-1RA) and has been proved to lower blood glucose by enhancing glucose-dependent insulin secretion of β-cells in T2DM patients2, FPG of 11.9 ± 3.6 mmol/L and HbA1c of 10.3 ± 2.8%, all patients in STII+ exenatide group (n = 66) and STII only group (n = 63) reached the glycaemic remission goal during STII therapy and were included for intention-to-treat analyses  clinical characteristics between STII+ exenatide group and STII only group shows that glucose levels, lipid profiles, and indicates of β-cell function (HOMA-B) and insulin resistance (HOMA-IR) were comparable between the two groups  Table , but morWithin 12-month follow-up period after STII therapy, 47 patients in STII+ exenatide group and 25 patients in STII only group sustained in glycaemic remission. The cumulative probability of maintenance of 1-year glycaemic remission in the STII+ exenatide group was 68.2 ± 5.7%, which was significantly higher than that of STII only group (36.5 ± 6.1%) (log-rank test: p < 0.001). During 24-month follow-up, 37 patients in STII+ exenatide group and 22 patients in STII only group maintained glycaemic remission, and the cumulative probability of maintenance of 2-year glycaemic remission in the STII+ exenatide group was 53.0 ± 6.1%, which was also significantly higher than that of STII only group (31.8 ± 5.9%) (log-rank test: p < 0.001).th week (after cessation of exenatide treatment for STII+ exenatide group) to the end of 24 months of follow-up. During Part I follow-up, the cumulative probabilities of glycaemic remission maintenances were 90.9 ± 3.5% for the STII+ exenatide group and 60.3 ± 6.2% for the STII only group (log-rank test: p < 0.001)  Fig. . And FigRepeated measurements on clinical parameters, blood pressure, lipid profiles, FPG, HbA1c, insulin resistance (HOMA-IR) and basal β-cell function (HOMA-B) were conducted for patients sustained in glycaemic remission only. After 2 days of exenatide cessation in STII+ exenatide group and 12 weeks after insulin cessation in STII only group, patients in both groups, compared with their baseline levels, had significantly decreased levels of body mass index, waist, FPG, HbA1c, triglycerides, LDL-C and significantly increased levels of HOMA-B (data not shown). As for differences between the two groups, analyses of covariance with adjustment for corresponding values at baseline showed that patients in STII+ exenatide group (N = 60), compared with those in STII only group (N = 37), had significantly decreased levels of waist , LDL-c  and HbA1c  . For exenatide treatment, the most frequently observed adverse event was mild-to-moderate nausea (48.4%); and the other gastrointestinal adverse events included vomiting and fatigue. Two patients (3.0%) withdrew as a result of nausea or vomiting. One patient in STII+ exenatide group developed pancreatitis, which was resolved after withdrawal of the exenatide medication. One patient in STII only group developed esophagus cancer and was given surgical operation after withdrawal.The present study found that, for newly diagnosed and drug-naive T2DM patients who reached glycaemic remission by STII therapy, sequential exenatide therapy for 12 weeks induced significantly higher 1-year and 2-year maintenance of glycaemic remission rates compared with STII only therapy. However, this improvement on maintenance of glycaemic remission only occurred during the 12-week exenatide treatment and was completely lost after stopping exenatide treatment. Furthermore, the effects of exenatide on lowering waist and HbA1c during exenatide therapy totally disappeared upon exenatide cessation. Thus, the beneficial effects of sequential exenatide therapy for 12 weeks after STII on maintenance of glycaemic remission and lowering waist and HbA1c did not sustain upon cessation of exenatide treatment.15. Insulin has been shown to protect the β-cell by inducing rapid reversal of glucotoxicity and lipotoxicity16, and short-term treatment with insulin therapy has been proposed for eliminating glucolipotoxicity and thus has been become a strategy of interest. Chinese patients with T2DM generally have severe β-cell dysfunction at diagnosis, and preservation of β-cell function is an more important treatment goal in newly diagnosed Chinese T2DM patients17. Although STII therapy has been shown to improve β-cell function and induce long-term glycaemic remission6, there were still more than half of patients who suffered hyperglycemic relapse within 1 year, and new strategies aimed to improve longer term maintenance of drug-free glycaemic remission after STII are therefore warranted.T2DM is characterised by β-cell dysfunction and insulin resistance, and glucotoxicity and lipotoxicity, which are effects of elevated glucose and free fatty acids on β-cell mass and function, have been proved to have deleterious effect on both β-cell function and insulin action19.Studies in cultured β-cells and rodent models have consistently shown that GLP-1RA treatment may help preserve beta-cells function. However, evidence about the effect of GLP-1RA, e.g. exenatide, following STII therapy on long term glycaemic remission rate is not available. Our results demonstrate that sequential exenatide treatment for 12 weeks after STII therapy can induce significantly higher maintenance of 1-year and 2-year glycaemic remission rates as compared to STII only therapy. The present study also found that, during exenatide treatment, patients in STII+ exenatide group had significantly decreased levels of HbA1c, LDL-c and waist as compared with those in STII only group. Therefore, one possible reason for the higher maintenance of 1-year and 2-year glycaemic remission rates in the STII+ exenatide group compared with STII only group is that STII can eliminate glucolipotoxicity, improve β-cell function and thus achieve optimum glycaemic control, and sequential exenatide treatment following STII may maintain or further enhance β-cell function and then improve the maintenance of glycaemic remission. Reduction of waist during exenatide therapy may contribute to another possible reason for the improved maintenance of glycaemic remission in STII+ exenatide group, because body fat reduction per se has been shown to improve β-cell function in subjects with and without type 2 diabetes13. Bunck MC and co-workers also found that exenatide significantly improved β-cell function during 1-year of treatment in T2DM patients, but β-cell function and glycemic control returned to pretreatment values after cessation of therapy20. However, after following a longer period (3-year) treatment with exenatide, they further reported that the disposition index, another β-cell function index, was sustained after a 4-week off-drug period21, which may imply that a sustained beneficial effect on β-cell preservation may be induced with longer treatment with exenatide. However, evidence on whether the effect of GLP-1RA on glycaemic remission will disappear upon treatment cessation is not available now. The present study found that the improvement of 1-year and 2-year maintenance of glycaemic remission rates in STII+ exenatide group mainly occurred during the 12-week exenatide treatment. After cessation of exenatide treatment, the cumulative probabilities of maintenance of glycaemic remission between the two groups were not statistically significant. Furthermore, we also found the effects of exenatide on lowering waist and HbA1c for patients in STII+ exenatide group during exenatide treatment disappeared upon cessation of exenatide therapy. Therefore, our data, together with others, suggests that sequential exenatide for 12 weeks after STII therapy may not be enough to maintain beneficial effect on glycaemic remission and that ongoing treatment of GLP-1RA, e.g. exenatide, or further treatment with other methods should be explored and emphasized.Retnakaran and colleagues reported that after the initial improvement in β-cell function achieved with pre-randomisation intensive insulin therapy, GLP-1RA liraglutide induced a robust further enhancement of β-cell function that was sustained for 48 weeks in early T2DM but lost upon cessation of therapySeveral limitations of the present study need to be recognized. First, our sample size was not enough  and all subjects were recruited from one hospital only, which may hampered the generalization of our findings. Second, repeat measurement on clinical and laboratory tests were available for patients sustained in glycaemic remission only but not for those with hyperglycemic relapse, which resulted in uncompleted data, and therefore we might underestimate differences of repeat measurements on clinical parameters during follow-up between these two groups. Third, these patients had relatively higher baseline HbA1c levels (around 10%), representing a group with poor glycaemia control. So our results may not necessarily be translated to the common patients whose HbA1c levels are lower than 10%. Last but not least, our results may be biased due to the following limitations: (a) more males patients were allocated into STII+ exenatide group, which may imply our randomization procedure might not be conducted adequately; (b) our study was not blinded to patients, although the nurses and medical students who collected data were not aware of patients’ allocation; (c) patients may change their life style habits during the 2-year follow-up period, but this may not confound our results much. These limitations, however, do not diminish the value of this study. For example, our study is, to the best of our knowledge, the first to apply a therapeutic strategy of sequential exenatide after STII therapy in newly diagnosed and drug-naive T2DM patients. Second, the total 2 years of follow-up duration was relatively long enough.22 or the sodium-glucose cotransporter 2 (SGLT2) inhibitor23, actively compared with metformin or placebo, should be explored.In conclusion, the present study demonstrated that sequential exenatide therapy for 12 weeks for newly diagnosed and drug-naive T2DM patients who reached glycaemic remission goal by STII therapy can induce significantly higher maintenance of 1-year and 2-year glycaemic remission rates as compared to STII only therapy, but the improvement on maintenance of glycaemic remission mainly occurred during the 12-week exenatide treatment and was completely lost after stopping exenatide treatment. Therefore, our results suggest that effect of exenatide after STII therapy for 12 weeks on maintenance of glycaemic remission did not sustain upon cessation of exenatide treatment and that ongoing treatment of exenatide or further treatment with other methods should be emphasized. For future study that may address further the remission effect of STII therapy in newly diagnosed T2DM patients, sequential treatment with incretin-related agent (GLP-1RA and dipeptidyl peptidase- 4 (DPP- 4) inhibitors2. Patients with the following conditions were excluded: acute or severe chronic diabetic complications, maturity onset diabetes in youth, mitochondria diabetes mellitus, severe intercurrent illness, or positive for glutamic acid decarboxylase antibody.This study was designed as a randomized, parallel-group, open-label, controlled trial. One hundred and fifty-seven patients, who were newly diagnosed with T2DM according to WHO criteria (1999) and had not received any antihyperglycaemic therapy, were screened in 2013 from the First Affiliated Hospital of Xiamen University, Xiamen, China. Eligibility criteria for this trial included age 30–70 years, fasting plasma glucose (FPG) 7.0–16.7 mmol/L, and body mass index (BMI) 20.0~35.0 kg/mwww.chictr.org.cn, number ChiCTR-IPR-15006298). Written informed consent was obtained from all patients before undergoing randomisation. The study was approved by the human ethics committees of the First Affiliated Hospital of Xiamen University and conducted in compliance with Good Clinical Practice (GCP) guidelines and the Declaration of Helsinki.The study aimed to explore effect of exenatide after STII therapy on maintenance of 1-year glycaemic remission was registered with Clinical Trials. gov (number NCT01776788) with 1 year of follow up after STII therapy. To explore effect of exenatide after STII therapy on longer time glycaemic remission, extension of follow-up period for another 1-year after STII therapy was registered with Chinese Clinical Trial Registry  or STII only group . The randomisation was conducted independently at a central office using a computer-generated random allocation sequence table. Allocation concealment was performed by enclosing assignments in sequentially numbered, opaque, closed envelopes.There was a 7-day run-in period of diet and physical exercise only. Then all patients received STII treatment in the hospital: insulin  with an insulin pump (Medtronic Minimed Paradigm Insulin Pump). Initial insulin doses were 0.4–0.5 IU/kg and total daily doses were divided into 50% of basal and 50% of bolus injection. Patients were monitored of capillary blood glucose levels at least six times a day, enabling titration of insulin doses to attain the glycaemic remission goal, which was defined as a fasting capillary blood glucose of 4.0–6.0 mmol/L and capillary blood glucose at 2-h after each of three meals of less than 8.0 mmol/L throughout the period of STII therapy. One patient who did not reach glycaemic remission goal was regarded as requiring additional or different therapy and was excluded from the efficacy analysis. All of the left 129 patients in two groups (n = 66 in STII+ exenatide group and n = 63 in STII only group) reached the glycaemic remission goal, and STII treatment was maintained for 2 weeks further. Then patients were discharged from the hospital, and patients in the STII+ exenatide group were trained and treated sequentially with exenatide  at home. Exenatide treatment was initiated at a dose of 5 ug b.i.d. (injected 30 min before breakfast and dinner) for 4 weeks, and followed at a dose of 10 ug b.i.d. for 8 weeks. Patients in the STII only group did not receive any further treatment after STII therapy.Baseline characteristics, including socio-demographic status, present and previous health history and medications, were collected during face-to-face interview by the trained nurses and medical students who were blinded to study objectives. Anthropometric measurements were obtained using standard protocols and techniques. After removal of shoes and heavy clothing, each subject underwent weight, height and waist circumference measurements, using a calibrated scale. Arterial blood pressure was measured with a mercury sphygmomanometer after sitting for at least 15 minutes. Three readings were taken at 5-min intervals. The mean of the three measurements was recorded.Fasting blood samples after a 10-h overnight fasting were used to measure FPG, glycosylated hemoglobin A1c (HbA1c), fasting insulin and lipid profiles. For conducting 2-h postprandial plasma glucose (2h-PPG) test, all patients were provided with the standardized breakfast which contained 439 kcal  after a 10-h overnight fasting. All laboratory tests were done in the central laboratory of the First Affiliated Hospital of Xiamen University. FPG and 2h-PPG were measured by using the hexokinase method, insulin by radioimmunoassay , and HbA1c by the Bio-Rad Variant Hemoglobin A1c assay. Homoeostasis model assessment (HOMA) was used to estimate basal β-cell function (HOMA-B) and insulin resistance (HOMA-IR) with the following equations: HOMA-B = 20*fasting serum insulin /-3.5), and HOMA-IR = fasting serum insulin  *fasting blood glucose /22.5.Follow-up for maintenance of glycaemic remission was initiated after STII therapy for both groups. After exenatide treatment for STII+ exenatide group and insulin treatment for STII only group, patients in both groups were instructed to continue diet and physical exercise only. Maintenance of glycaemic remission was ascertained by systematic glycaemic monitoring (FPG and 2h-PPG) monthly during the initial 3 months and at a 3-month interval thereafter. All patients were asked to conduct self-monitoring of blood glucose at home and encouraged to contact the medical staff with any intercurrent problem. Maintenance of glycaemic remission was defined as FPG < = 7.0 mmol/L or 2h-PPG < = 10.0 mmol/L. Hyperglycaemia relapse was defined as either FPG > 7.0 mmol/L or 2h-PPG > 10.0 mmol/L, and confirmation of hyperglycaemia relapse was conducted two week later. The time of hyperglycaemic relapse was recorded and patients with hyperglycaemia relapse were treated with metformin and then gliclazide-MR for the remaining study period according to the guideline of Chinese Diabetes Society.The primary endpoints were the maintenance of 1-year and 2-year glycaemic remission rates which were calculated from the time point after STII therapy for the original study and the extension study, respectively. Secondary endpoints were repeat measurements of clinical parameters , insulin resistance (HOMA-IR), etc) during follow-up.6. The margin of an acceptable difference of 1-year drug-free remission rate between two groups was set as 25%. The criterion for establishing superiority was that the lower bound of the two side 95% confidence interval (CI) for the difference between the two groups must exceed the predefined superiority margin of 5%. Under these assumptions, a sample size of 65 patients in each of the 2 treatment groups was needed to yield 80% power to conclude that STII+ exenatide group was superior to the STII only group (control group) with α = 0.05. Further taking account of the dropout rate as around 10%, then 72 patients was needed for each group. And the extension study was further extended for another 1-year of follow-up after STII therapy.The original study with 1-year duration of follow-up was designed as a superiority trial aimed to show STII+ exenatide group was superior to the STII only group on maintenance of 1-year glycaemic remission. Our previously pilot study in 2010 showed that around 70% (11 of 15) patients who received exenatide after STII achieved 1-year remission; and existing evidence showed that 45% patients who received STII only achieved 1-year remissionData were analyzed using Stata14.0  and performed on the intention-to-treat population. Differences of baseline characteristics between the two groups were compared using analyses of variance for continuous variables and chi-square test for categorical variables. For differences of repeat measurements between these two groups after treatment , analyses of covariance were performed with adjustment for corresponding values at baseline. Kaplan-Meier plots were used to analyse the time for the 1-year and 2-year maintenance of glycaemic remission from the time point after STII therapy, which were compared between two groups with the use of log-rank test. Significance levels were defined as p < 0.05.www.ClinicalTrials.gov, Registration date: January 22, 2013; ChiCTR-IPR-15006298, www.chictr.org.cn, Registration date: April 21, 2015.NCT01776788,