Assessment of the functional impact of germline BRCA1/2 variants located in non-coding regions in families with breast and/or ovarian cancer predisposition.

PURPOSE: The molecular mechanism of breast and/or ovarian cancer susceptibility remains unclear in the majority of patients. While germline mutations in the regulatory non-coding regions of BRCA1 and BRCA2 genes have been described, screening has generally been limited to coding regions. The aim of this study was to evaluate the contribution of BRCA1/2 non-coding variants. METHODS: Four BRCA1/2 non-coding regions were screened using high-resolution melting analysis/Sanger sequencing or next-generation sequencing on DNA extracted from index cases with breast and ovarian cancer predisposition (3926 for BRCA1 and 3910 for BRCA2). The impact of a set of variants on BRCA1/2 gene regulation was evaluated by site-directed mutagenesis, transfection, followed by Luciferase gene reporter assay. RESULTS: We identified a total of 117 variants and tested twelve BRCA1 and 8 BRCA2 variants mapping to promoter and intronic regions. We highlighted two neighboring BRCA1 promoter variants (c.-130del; c.-125C > T) and one BRCA2 promoter variants (c.-296C > T) inhibiting significantly the promoter activity. In the functional assays, a regulating region within the intron 12 was found with the same enhancing impact as within the intron 2. Furthermore, the variants c.81-3980A > G and c.4186-2022C > T suppress the positive effect of the introns 2 and 12, respectively, on the BRCA1 promoter activity. We also found some variants inducing the promoter activities. CONCLUSION: In this study, we highlighted some variants among many, modulating negatively the promoter activity of BRCA1 or 2 and thus having a potential impact on the risk of developing cancer. This selection makes it possible to conduct future validation studies on a limited number of variants.

Validation of patient reports, automated pharmacy records, and pill counts with electronic monitoring of adherence to antihypertensive therapy.

OBJECTIVES: To evaluate the validity of patient report, pharmacy dispensing records, and pill counts as measures of antihypertensive adherence using electronic monitoring as the validation standard. METHODS: The study was conducted among 286 members of Harvard Pilgrim Health Care, a managed care organization, who were at least 18 years of age, on monotherapy for hypertension, and had prescription drug coverage. Prescription refill adherence during the 12 months before enrollment was determined from their automated pharmacy dispensing records. Participants were interviewed about their medication adherence before and after a 3-month electronic monitoring period during which pill counts were also performed. Adherence to both recommended number and timing of doses was estimated from electronic monitoring data. Data analysis was based on statistical correlation and analysis of variance. RESULTS: Electronic adherence monitoring revealed that the proportion of prescribed doses consumed was higher (0.92) than the proportion of doses taken on time (0.63). The correlation between adherence to quantity and timing of doses was 0.32. Concurrent pill counts and earlier refilling patterns were moderately correlated with electronic monitoring (pill count: r = .52 with quantity and r = .17 with timing; refill adherence r = .32 with quantity and r = .22 with timing). There was considerable misclassification of adherence reported by patients, although nonadherence was generally accurately reported. CONCLUSIONS: Deviation from recommended timing of doses appears to be greater than from prescribed number of doses. Pharmacy dispensing records demonstrate predictive validity as measures of cumulative exposure and gaps in medication supply. Adherence levels determined from pill counts and pharmacy dispensing records correlate more closely with quantity than with timing of doses. Nonadherence reported by patients can serve as a qualitative indicator and predictor of reduced adherence.

Regenerative therapy. The future of periodontics.

The role of periodontics in dentistry is changing subtly from primarily resective procedures to regenerative techniques that are becoming increasingly more predictable. The periodontal focus is shifting to regeneration of the attachment apparatus, guided bone regeneration and more desirable implant placement. However, certain techniques and materials may not be as predictable as others and therein lies the predicament for the practicing clinician.

Occlusion of arterial flow in the extremities at subsystolic pressures through the use of wide tourniquet cuffs.

Tourniquet-induced peripheral neuropathy is at least partially attributable to excessive forces applied to the nerves beneath cuffs inflated to high pressures. Lowering the inflation pressure to the minimum necessary to obtain an effective arrest of blood flow distal to the tourniquet cuff should increase the safety of these systems. Tourniquet cuffs with widths varying from 4.5 cm to 80 cm were applied to the upper and lower extremities of 34 healthy, normotensive volunteers. Occlusion pressure for the arterial system under study was estimated by determining that level of cuff inflation at which the distal pulse became detectable by ultrasonic flowmetry. The occlusion pressure was inversely proportional to the ratio of tourniquet cuff width to limb circumference and was in the subsystolic range at a cuff width to limb circumference ratio above 0.5. Wide tourniquet cuffs can achieve an effective arrest of the regional arterial circulation at subsystolic pressures of inflation. Wide cuffs may reduce the risk of tourniquet-induced injury to underlying soft tissues by lowering the inflation pressure required to secure a bloodless field.

Perineural pressures under the pneumatic tourniquet in the upper extremity.

The literature indicates that tourniquet-induced neurological injuries are relatively common and frequently occur at a subclinical level. In order to evaluate the pressure transmitted to the major peripheral nerves of the arm by an externally applied pneumatic tourniquet, a fully implantable biomedical pressure transducer was placed adjacent the radial, median and ulnar nerves in six cadaver upper extremities of average dimensions. This sensor allowed accurate, reproducible measurements of perineural pressures without requiring significant disruption of the normal anatomical structures of the test limb for its installation. At levels of tourniquet cuff inflation which are commonly used in clinical practice, there was little or no decrease in the pressure detected in the perineural regions over that applied to the surface of the limb. In addition, there was a steep gradient of perineural pressure between locations beneath the edge of the cuff and those under its midpoint. This was most marked at the highest levels of tourniquet inflation. At presently accepted levels of inflation, pneumatic tourniquet cuffs transmit high pressures to the peripheral nerves without any significant attenuation by the intervening soft tissues. The distribution of these forces is one which may subject the underlying nerves to deleterious shear forces, especially at higher levels of inflation.

Alpha 1-adrenergic stimulation of ACTH secretion in vivo in the rat.

The alpha-adrenergic agonist phenylephrine (100 micrograms, i.v.) causes a rapid 5- to 10-fold elevation of plasma ACTH levels 5 min after its administration in adult ovariectomized rats, the concentration of the hormone remaining elevated up to at least 2h. Epinephrine (10 micrograms) causes also a rapid but shorter-lived stimulation of ACTH secretion. While having no effect alone under basal conditions (conscious freely-moving animals), the highly specific alpha 1-adrenergic antagonist prazosin (0.25 micrograms) almost completely reverses the stimulatory effect of phenylephrine. Pretreatment with dexamethasone inhibits basal plasma ACTH levels by 70% and almost completely prevents the stimulatory effect of phenylephrine on this parameter. Plasma levels of alpha-MSH, on the other hand, are only stimulated 1-fold above control 5 min after the administration of phenylephrine and are insensitive to corticosteroid treatment. Based on the specificity of action of prazosin on postsynaptic alpha 1-adrenergic receptors and of dexamethasone on the anterior lobe of the pituitary gland, the present data indicate that phenylephrine is a potent stimulator of ACTH secretion by a direct action on an alpha 1-adrenergic receptor in corticotrophs of the adenohypophysis. They also support the suggestion that epinephrine and/or norepinephrine could be involved as physiological corticotropin-releasing factor(s).