Identification of a Novel β-Cell Glucokinase (GCK) Promoter Mutation (−71G>C) That Modulates GCK Gene Expression Through Loss of Allele-Specific Sp1 Binding Causing Mild Fasting Hyperglycemia in Humans

OBJECTIVE: Inactivating mutations in glucokinase (GCK) cause mild fasting hyperglycemia. Identification of a GCK mutation has implications for treatment and prognosis; therefore, it is important to identify these individuals. A significant number of patients have a phenotype suggesting a defect in glucokinase but no abnormality of GCK. We hypothesized that the GCK beta-cell promoter region, which currently is not routinely screened, could contain pathogenic mutations; therefore, we sequenced this region in 60 such probands. RESEARCH DESIGN AND METHODS: The beta-cell GCK promoter was sequenced in patient DNA. The effect of the identified novel mutation on GCK promoter activity was assessed using a luciferase reporter gene expression system. Electrophoretic mobility shift assays (EMSAs) were used to determine the impact of the mutation on Sp1 binding. RESULTS: A novel -71G>C mutation was identified in a nonconserved region of the human promoter sequence in six apparently unrelated probands. Family testing established cosegregation with fasting hyperglycemia (> or = 5.5 mmol/l) in 39 affected individuals. Haplotype analysis in the U.K. family and four of the Slovakian families demonstrated that the mutation had arisen independently. The mutation maps to a potential transcriptional activator binding site for Sp1. Reporter assays demonstrated that the mutation reduces promoter activity by up to fourfold. EMSAs demonstrated a dramatic reduction in Sp1 binding to the promoter sequence corresponding to the mutant allele. CONCLUSIONS: A novel beta-cell GCK promoter mutation was identified that significantly reduces gene expression in vitro through loss of regulation by Sp1. To ensure correct diagnosis of potential GCK-MODY (maturity-onset diabetes of the young) cases, analysis of the beta-cell GCK promoter should be included

Barriers and facilitators to using NHS Direct: a qualitative study of ‘users’ and ‘non-users’

Background NHS Direct, introduced in 1998, has provided 24/7 telephone-based healthcare advice and information to the public in England and Wales. National studies have suggested variation in the uptake of this service amongst the UK¿s diverse population. This study provides the first exploration of the barriers and facilitators that impact upon the uptake of this service from the perspectives of both `users¿ and `non- users¿. Methods Focus groups were held with NHS Direct `users¿ (N?=?2) from Bedfordshire alongside `non-users¿ from Manchester (N?=?3) and Mendip, Somerset (N?=?4). Each focus group had between five to eight participants. A total of eighty one people aged between 21 and 94 years old (M: 58.90, SD: 22.70) took part in this research. Each focus group discussion lasted approximately 90 minutes and was audiotape-recorded with participants¿ permission. The recordings were transcribed verbatim. A framework approach was used to analyse the transcripts. Results The findings from this research uncovered a range of barriers and facilitators that impact upon the uptake of NHS Direct. `Non-users¿ were unaware of the range of services that NHS Direct provided. Furthermore, `non-users¿ highlighted a preference for face-to face communication, identifying a lack of confidence in discussing healthcare over the telephone. This was particularly evident among older people with cognitive difficulties. The cost to telephone a `0845¿ number from a mobile was also viewed to be a barrier to access NHS Direct, expressed more often by `non-users¿ from deprived communities. NHS Direct `users¿ identified that awareness, ease of use and convenience were facilitators which influenced their decision to use the service. Conclusions An understanding of the barriers and facilitators which impact on the access and uptake of telephone-based healthcare is essential to move patients towards the self-care model. This research has highlighted the need for telephone-based healthcare services to increase public awareness; through the delivery of more targeted advertising to promote the service provision availabl