Use of buccal cells collected in mouthwash as a source of DNA for clinical testing

CONTEXT: To maximize the participation rate in population genetic studies, alternatives to invasive whole blood collection are increasing. One such alternative is buccal epithelial cell collection, which, in contrast to venipuncture and finger sticks, is painless. Buccal cells, if collected and purified efficiently, offer an acceptable source for DNA to be used in research and clinical applications. OBJECTIVE: To develop a noninvasive sampling method for collecting cells for routine DNA testing in a clinical laboratory setting. DESIGN: Five factors were used to evaluate several brands of mouthwash: (1) compatibility with the DNA purification chemistry, (2) DNA yield, (3) DNA quality, (4) DNA stability at room temperature, and (5) mouthwash taste. Next, an optimization study was undertaken to maximize DNA yield. Finally, a validation study was undertaken with the optimized protocol to test a panel of 14 donors for DNA yield and performance and to test for the stability of DNA held in mouthwash. SETTING: Industrial research and development laboratory. RESULTS: Of 5 mouthwashes tested, Scope brand mouthwash received the highest overall ranking. The addition of proteinase K and glycogen to the protocol significantly enhanced DNA yields, with a test panel (n = 14) giving a range of 12 to 60 microg of DNA per donor. In a 4-week room temperature stability study, the DNA in mouthwash samples was found to be stable for at least 2 weeks. CONCLUSION: A clinically validated DNA purification chemistry was adapted to a noninvasive specimen collection method. This method used a commercially available mouthwash, Scope, to collect buccal epithelial cells for the preparation of high-quality DNA in high yield.     

Relation between gastric histology and gastric juice pH and nitrite and N-nitroso compound concentrations in the stomach after surgery for duodenal ulcer

Formation of N-nitroso compounds in gastric juice has been implicated in the pathogenesis of cancer in the stomach after operation. Gastric juice was aspirated from 85 subjects: 23 were controls, 51 had previously undergone vagotomy and gastrojejunostomy, and 11 had previously undergone vagotomy and pyloroplasty. The gastric juice samples were analysed for pH, nitrite, and total N-nitroso compounds. A significant correlation was found between pH and nitrite concentration (p less than 0.01). No significant correlation was found between pH and total N-nitroso compound concentration or between nitrite and N-nitroso compound concentration. The vagotomy and gastrojejunostomy patients had higher pH values and higher concentrations of nitrites and N-nitroso compounds than controls (p = 0.01 in all cases). The 51 vagotomy and gastrojejunostomy patients also underwent endoscopy and biopsy. They were divided into three groups: group 1 (21 patients) had no intestinal metaplasia and no more than mild dysplasia; group 2 (20 patients) had intestinal metaplasia; and group 3 (10 patients) had moderate or severe dysplasia. Groups 2 and 3 both had higher pH values and higher nitrite concentrations than group 1 (p = 0.01 in all cases). There was no significant difference, however, between either group 2 or 3 and group 1 for total N-nitroso compound concentration. Since there was no simple linear relation between pH and N-nitroso compound concentration, it was concluded that formation of N-nitroso compounds at high pH was unlikely to be involved in the pathogenesis of gastric cancer in the hypochlorhydric stomach after operation. The relation between nitrite and histological abnormality was not associated with a similar relation between N-nitroso compounds and histological abnormality. It therefore appears that there is no simple relation between N-nitroso compounds and the pathogenesis of premalignant gastric mucosal changes.     

Heat shock protein 70 (HSP70) and complement C4 genotypes in patients with hyperthyroid Graves'' disease.

The genetic polymorphisms of the heat shock protein 70 (HSP70) and complement component C4 were investigated in 90 patients with hyperthyroid Graves' disease and 92 normal control subjects. The 8.5-kb PstI HSP70 allele was strongly associated with Graves' disease when compared with controls (P less than 0.001). The presence of the 8.5-kb PstI HSP70 allele was strongly associated with a deletion of the C4A gene in both patients and controls (P less than 0.0003 and P less than 0.00005 respectively). However, in the absence of C4A gene deletion, the frequency of the 8.5-kb PstI HSP70 allele was still significantly higher in patients when compared with controls (P less than 0.04). These results suggest that the HSP70 locus may have an immunological role to play in autoimmune hyperthyroid Graves' disease.     

Virulence Differences in Mice of Type A and B Histoplasma capsulatum Yeasts Grown in Continuous Light and Total Darkness

Type B yeasts were more virulent for mice than type A under most experimental conditions. Mice infected with type B yeasts grown in the light lived significantly longer than those with type B yeasts grown in the dark. Virulence differences of type A yeasts grown in continuous fluorescent light versus total darkness were not statistically significant.     

Collagen-induced arthritis in C57BL/6 (H-2b) mice: new insights into an important disease model of rheumatoid arthritis

Collagen-induced arthritis (CIA) is a widely used model of rheumatoid arthritis (RA) and has been important for understanding autoimmunity. CIA is purportedly restricted to mice bearing the MHC class II H-2q or H-2r haplotypes. In this study, we re-examined established concepts regarding susceptibility to CIA. We found mice derived from the C57BU6 (B6) (H-2b) background can develop CIA with high incidence (60-70%), and sustained severity by using an immunization procedure modified for optimum response in DBA/1 (D1) (H-2q) mice. Clinically and histologically the B6 disease resembles that of D1 mice and is dependent on immunization with type II collagen, as well as on B and CD4+ T cells. In contrast, 129/Sv mice, which share H-2b, are resistant to CIA. We conclude that susceptibility to CIA may reflect immunization conditions and/or important contributions from non-MHC genes, revealed by different immunization protocols. A practical outcome is that CIA can be directly applied to gene knockout mice generated from B6 embryonic stem cells without need for backcross onto the D1 background. This model may lead to improved understanding of autoimmunity in CIA and RA and may provide a platform for analysis of the contribution of non-MHC genes to CIA.     

Depressive symptoms predict medical care utilization in a population-based sample

BACKGROUND: Several examinations have detected a relation between depressive symptoms and medical utilization. However, selection biases have been involved in most previous examinations. We sought to test the association between depressive symptoms and prospective, increased medical care utilization, in a population-based Canadian sample, while controlling for utilization due to medical illness and controlling for selection bias. METHODS: Data from the Nova Scotia Health Survey 1995, an age- and sex-stratified random sampling of 3227 Nova Scotian adults, included the Center for Epidemiological Studies-Depression scale and items assessing chronic medical conditions and current limitations in daily activities resulting from medical illness. We linked survey data with medical care utilization measures for the year following the survey, including out-patient visits, reimbursement for out-patient services, hospitalizations, and hospitalization days. RESULTS: After controlling for age, sex, count of medical diagnoses and current medical severity, those with a greater level of depressive symptoms were at greater risk of having increased medical care utilization in the following year. These results remained after removing mental health care utilization costs. CONCLUSIONS: In a population-based sample, depressive symptoms predicted greater medical care utilization, independent of a number of medical severity measures. Whether depressive symptoms are a risk marker or a causal risk factor for increased medical utilization remains to be explored.     

Influences on prescribing in non-fundholding general practices.

BACKGROUND: The experience from general practice fundholding suggests that financial incentives may influence prescribing; guidelines and hospital prescribing are two other suggested influences. AIM: A study was undertaken to establish general practitioners' attitudes to a financial prescribing incentive scheme, the presence and use of guidelines, and the influence of prescribing initiated within secondary care. METHOD: A postal questionnaire survey of non-fundholding general practices in the former Northern Region was conducted. RESULTS: Practices' thinking and subsequent decisions about the incentive prescribing scheme were most often influenced by discussions within the practice (45%). Those practices that achieved their savings under the incentive scheme were less likely than those not achieving savings to feel that the target was not achievable, the time scale was unacceptable, and that the philosophy behind the scheme was unacceptable. Forty-five per cent of practices received advice from neither a medical nor a pharmaceutical adviser; 27% of practices received advice from both, 12% from a medical adviser only and 16% from a pharmaceutical adviser only. Of the practices that tried to make their target savings, 91% intended to increase generic prescribing; fewer than one-third of practices mentioned any other measure. Prescribing guidelines were reported by a minority of practices, although reported rates of use were high when these were present. Clinical guidelines for three conditions, asthma, diabetes and hypertension, were present in more than 50% of practices; 25% of practices had no clinical guidelines. Hospital prescribing was reported as 'always' or 'usually' influencing prescribing for diabetes by 57% of respondents, ischaemic heart disease by 55%, peptic ulceration by 49%, asthma by 42% and hypertension by 39%. CONCLUSIONS: General practitioner prescribing is influenced by a complex web of factors, with no single factor pre-eminent. To understand this area further, there is a need to take each of these areas and ascertain the match between doctors' perceptions and actual practice.