Low PIP(2) molar fractions induce nanometer size clustering in giant unilamellar vesicles.

Phosphatidylinositol (4,5) bisphosphate (PIP2) is an important signaling molecule located on the inner leaflet of the cell membrane. In order to perform its various signaling functions, it is suggested that PIP2 must be able to form localized clusters. In this study, we have used LAURDAN generalized polarization function (GP) with unlabeled PIP2 and single point fluorescence correlation spectroscopy and brightness analysis of various BODIPY labeled PIP2 to determine the presence of clusters in the membrane of giant unilamellar vesicles (GUVs) made of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) or a mixture of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), sphingomyelin and cholesterol. We determined the number of freely diffusing fluorescent BODIPY molecules in the membrane and found that in GUVs containing various amounts of labeled PIP2, this number was significantly lower than in GUVs made with the control BODIPY labeled hexadecyl phosphatidylcholine (BODIPY-HPC). Also, we noted an increase in brightness of the labeled PIP2 particles with increasing labeled PIP2 molar fraction. Together with the observed change in LAURDAN GP with increasing molar fraction of unlabeled PIP2, these results demonstrate the presence of PIP2 enriched clusters that are smaller than the resolution limit of the fluorescent microscope. In addition, we report the presence of a hypsochromic shift of the fluorescence for the BODIPY labeled lipids that we attributed to clustering. This clustering result in a change in the partitioning of the lipids with the BODIPY labeled PIP2 lipids able to move between the liquid ordered and liquid disordered phase.

Green fluorescent protein expression triggers proteome changes in breast cancer cells.

Green fluorescent protein (GFP) is the most commonly used reporter of expression in cell biology despite evidence that it affects the cell physiology. The molecular mechanism of GFP-associated modifications has been largely unexplored. In this paper we investigated the proteome modifications following stable expression of GFP in breast cancer cells (MDA-MB-231). A combination of three different proteome analysis methods (2-DE, iTRAQ, label-free) was used to maximise proteome coverage. We found that GFP expression induces changes in expression of proteins that are associated with protein folding, cytoskeletal organisation and cellular immune response. In view of these findings, the use of GFP as a cell reporter should be carefully monitored.

Osteoporosis and genetic influence: a three-generation study.

We have studied 27 triads of mother, daughter and grandmother for possible genetic influence on distal and proximal forearm bone density, measured by single photon absorptiometry. We found a significant correlation of bone density at the proximal forearm between the mothers and grandmothers (r = 0.499, P < 0.01). There was also a weak correlation between proximal forearm bone densities of mothers and daughters (r = 0.327, P < 0.1). Significant correlations were found between the three generations for grip strength, pedometry, height and triceps skinfold thickness. There was also significant correlation between mother and grandmother for alcohol intake. There was no correlation for contraceptive pill use, smoking, dietary calcium intake, body weight or body mass index. The study concludes that, although there are similarities in bone mineral content between the three generations, genetic factors cannot be conclusively proven to be the major determinant of bone density. Lifestyle and environmental factors may have a bearing on achieving the peak bone mass and subsequent development of osteoporosis.

Climacteric symptoms in healthy middle-aged women.

The incidence of climacteric symptoms was determined in 247 healthy premenopausal women in a community setting. These volunteers had been recruited to a longitudinal study of bone density. Of these subjects, 46 ceased to menstruate during the study, and in this subgroup symptoms were compared before and after cessation of menstruation. Only hot flushes increased after cessation of menstruation in the longitudinal study and showed age correlation in the cross-sectional study. Hot flushes thus emerged as a true menopausal symptom. Although evidence for this is weaker, cold sweats and suffocation seem likely to be genuinely menopausal. Breast discomfort and the four mood symptoms of irritability, excitability, depression and poor concentration improved after cessation of menstruation, and this study gives no support for their being part of the menopausal syndrome; it suggests that these symptoms are more likely to be related to menstruation than to the menopause.

Determinants of forearm bone density in premenopausal women: a study in one general practice.

Predictors of distal and proximal forearm bone density, measured by photon absorbtiometry, were investigated in 248 premenopausal women aged 39-56 years. Only one strong predictor of lower bone density was found--history of previous fracture at any site (P less than 0.001). Two other factors showed a weaker association with density, but only at the distal site--history of diuretic use showed a positive association (P less than 0.02) whereas alkaline phosphatase level was inversely correlated with density (P less than 0.01). Other factors were not significant predictors: these included age, calcium intake, level of exercise, anthropometric measures of obesity, serum calcium level, parity, lactation history, a menopausal symptom history, use of the contraceptive pill, smoking and alcohol intake. These results contrast with the far stronger predictors found for postmenopausal women and suggest that genetic endowment rather than lifestyle may be the major determinant of bone density before the menopause.

Death certification by British doctors: a demographic analysis.

Regional differences in mortality from cardiovascular diseases, respiratory diseases, strokes, and other causes have been observed in England and Wales. To determine to what extent the death certification practices of doctors influence these variations, we surveyed 123 British general practitioners and housemen. Each doctor completed death certificates based on fictitious case histories. We found that diagnostic groupings did not vary significantly according to the doctors' region, type of practice, place of medical training, sex, or year of qualification. Doctors qualifying before 1955, however, appeared more inclined than their younger colleagues to list stomach cancer on the death certificate.

Women's attitudes to screening for neural tube defects.

The attitudes of 2254 pregnant and 262 other women to serum alpha-fetoprotein (AFP) screening for neural tube defects (NTDs) were investigated, the former by interview and the latter by questionnaires to which 228 replies were received. Over 97 per cent of the pregnant women wanted to be included in such a screening programme. Ninety-six per cent of other women who completed the questionnaire were in favour of national screening for NTDs but 14 per cent of them were uncertain about the action they would take if the test were positive.

Establishment of two group practice centres by conversion of existing houses.

A practice of five (later six) doctors established their own group practice centre by purchase and conversion of an existing house. Later a second, smaller centre was established in the same way to replace an existing branch surgery.The planning, financing, and execution of these projects is described.

The prognostic value of human placental lactogen levels in threatened abortion in general practice.

We report the first use of human placental lactogen as an indicator of the outcome of threatened abortion in general practice. The results were not found to be helpful in clinical management.