Dipyridamole synergizes with nitric oxide to prolong inhibition of thrombin-induced platelet shape change

We and others have previously demonstrated that nitric oxide (NO)-induced inhibition of platelet shape change is important in regulating platelet adhesion and aggregation, and therapeutic intervention of this pathway is clinically relevant for secondary prevention of stroke with dipyridamole. In the present study, we investigated whether dipyridamole affected the shape change of aspirinated platelets. Platelet shape change was inhibited using both authentic NO and sodium nitroprusside, as monitored by light scattering and mean platelet volume measurements. Dipyridamole synergized with NO, even at supra-therapeutic levels, to inhibit thrombin-induced shape change and further potentiated cAMP dependent protein kinase (PKA) mediated phosphorylation of vasodilator stimulated phosphoprotein (VASP) Ser157, even without altered levels of platelet cAMP. The effect of dipyridamole on NO-inhibited shape change depended on cGMP synthesis as evaluated by inhibition of soluble guanylyl cyclase. Measured increases in cGMP levels by dipyridamole and NO was assessed by mathematical modeling and found to be consistent with inhibition of phosphodiesterase 5 (PDE5). The model could explain the unexpected efficiency of dipyridamole in inhibiting PDE5 at the measured cGMP levels, by the majority of cGMP being bound to cGMP-dependent protein kinase (PKG). Still, selective activators of PKG failed to extend NO-mediated inhibition of the thrombin-induced platelet shape change, suggesting that PKG was not responsible for the inhibitory effect of NO and dipyridamole on shape change. The effects of dipyridamole were independent of the prostanoid and ADP pathways. Thus, the effect of dipyridamole on NO-mediated inhibition of platelet shape change may be an important and additional beneficial therapeutic effect of dipyridamole, which we suggest, is acting though localized amplification of the NO/cGMP/Phosphodiesterase3/cAMP/PKA-pathway. Probably, the efficiency of dipyridamole could be amplified clinically with NO donors.     

Loss of astrocyte polarization in the tg-ArcSwe mouse model of Alzheimer's disease

Aquaporin-4 (AQP4) is the predominant water channel in brain and is selectively expressed in astrocytes. Astrocytic endfoot membranes exhibit tenfold higher densities of AQP4 than non-endfoot membranes, making AQP4 an excellent marker of astrocyte polarization. Loss of astrocyte polarization is known to compromise astrocytic function and to be associated with impaired water and K+ homeostasis. Here we investigate by a combination of light and electron microscopic immunocytochemistry whether amyloid deposition is associated with a loss of astrocyte polarization, using AQP4 as a marker. We used the tg-ArcSwe mouse model of Alzheimer's disease, as this model displays perivascular plaques as well as plaques confined to the neuropil. 3D reconstructions were done to establish the spatial relation between plaques and astrocytic endfeet, the latter known to contain the perivascular pool of AQP4. Changes in AQP4 expression emerge just after the appearance of the first plaques. Typically, there is a loss of AQP4 from endfoot membranes at sites of perivascular amyloid deposits, combined with an upregulation of AQP4 in the neuropil surrounding plaques. By electron microscopy it could be verified that the upregulation reflects an increased concentration of AQP4 in those delicate astrocytic processes that abound in synaptic regions. Thus, astrocytes exhibit a redistribution of AQP4 from endfoot membranes to non-endfoot membrane domains. The present data suggest that the development of amyloid deposits is associated with a loss of astrocyte polarization. The possible perturbation of water and K+ homeostasis could contribute to cognitive decline and seizure propensity in patients with Alzheimer's disease.     

Incomplete pregnancy is not associated with breast cancer risk: the California Teachers Study

BACKGROUND: Early studies of incomplete pregnancy and development of breast cancer suggested that induced abortion might increase risk. Several large prospective studies, which eliminate recall bias, did not detect associations, but this relationship continues to be debated. STUDY DESIGN: To further inform this important question, we examined invasive breast cancer as it relates to incomplete pregnancy, including total number of induced abortions, age at first induced abortion and total number of miscarriages among women participating in the ongoing California Teachers Study (CTS) cohort. Incomplete pregnancy was self-reported on the CTS baseline questionnaire in 1995-1996. Incident breast cancers were ascertained in 3324 women through 2004 via linkage with the California Cancer Registry. RESULTS: Using Cox multivariable regression, we found no statistically significant association between any measure of incomplete pregnancy and breast cancer risk among nulliparous or parous women. CONCLUSION: These results provide strong evidence that there is no relationship between incomplete pregnancy and breast cancer risk.     

Evaluation of established breast cancer risk factors as modifiers of BRCA1 or BRCA2: a multi-center case-only analysis

The incomplete penetrance of mutations in BRCA1 and BRCA2 suggests that some combination of environmental and genetic factors modifies the risk of breast cancer in mutation carriers. This study sought to identify possible interactions between established breast cancer risk factors and BRCA1 or BRCA2 mutations using a case-only study design. Breast cancer cases that had been tested for BRCA1 and BRCA2 mutations were identified from 11 collaborating centers. Comparisons of reproductive and lifestyle risk factors were made between women with breast cancer who were positive for BRCA1 mutations (n = 283), BRCA2 mutations (n = 204), or negative for both BRCA1 and BRCA2 mutations (n = 894). Interaction risk ratios (IRRs) were calculated using multinominal logistic regression models. Compared with non-carriers, statistically significant IRRs were observed for later age at menarche among BRCA2 mutation carriers, for a greater number of pregnancies among both BRCA1 and BRCA2 mutation carriers, and for alcohol use among BRCA1 mutation carriers. Our data suggest that the risk for breast cancer among BRCA1 or BRCA2 carriers may be modified by reproductive characteristics and alcohol use. However, our results should be interpreted cautiously given the overall inconsistency in the epidemiologic literature on modifiers of BRCA1 and BRCA2.     

Expression levels of uridine 5'-diphospho-glucuronosyltransferase genes in breast tissue from healthy women are associated with mammographic density

Introduction

Mammographic density (MD), as assessed from film screen mammograms, is determined by the relative content of adipose, connective and epithelial tissue in the female breast. In epidemiological studies, a high percentage of MD confers a four to six fold risk elevation of developing breast cancer, even after adjustment for other known breast cancer risk factors. However, the biologic correlates of density are little known.

Methods

Gene expression analysis using whole genome arrays was performed on breast biopsies from 143 women; 79 women with no malignancy (healthy women) and 64 newly diagnosed breast cancer patients, both included from mammographic centres. Percent MD was determined using a previously validated, computerized method on scanned mammograms. Significance analysis of microarrays (SAM) was performed to identify genes influencing MD and a linear regression model was used to assess the independent contribution from different variables to MD.

Results

SAM-analysis identified 24 genes differentially expressed between samples from breasts with high and low MD. These genes included three uridine 5''-diphospho-glucuronosyltransferase (UGT) genes and the oestrogen receptor gene (ESR1). These genes were down-regulated in samples with high MD compared to those with low MD. The UGT gene products, which are known to inactivate oestrogen metabolites, were also down-regulated in tumour samples compared to samples from healthy individuals. Several single nucleotide polymorphisms (SNPs) in the UGT genes associated with the expression of UGT and other genes in their vicinity were identified.

Conclusions

Three UGT enzymes were lower expressed both in breast tissue biopsies from healthy women with high MD and in biopsies from newly diagnosed breast cancers. The association was strongest amongst young women and women using hormonal therapy. UGT2B10 predicts MD independently of age, hormone therapy and parity. Our results indicate that down-regulation of UGT genes in women exposed to female sex hormones is associated with high MD and might increase the risk of breast cancer.     

A multi-state model for sick-leave data applied to a randomized control trial study of low back pain

AIMS: Analysing and presenting data on different outcomes after sick-leave is challenging. The use of extended statistical methods supplies additional information and allows further exploitation of data. METHODS: Four hundred and fifty-seven patients, sick-listed for 8-12 weeks for low back pain, were randomized to intervention (n=237) or control (n=220). Outcome was measured as: "sick-listed', "returned to work', or "disability pension'. The individuals shifted between the three states between one and 22 times (mean 6.4 times). In a multi-state model, shifting between the states was set up in a transition intensity matrix. The probability of being in any of the states was calculated as a transition probability matrix. The effects of the intervention were modelled using a non-parametric model. RESULTS: There was an effect of the intervention for leaving the state sick-listed and shifting to returned to work (relative risk (RR)=1.27, 95% confidence interval (CI) 1.09- 1.47). The nonparametric estimates showed an effect of the intervention for leaving sick-listed and shifting to returned to work in the first 6 months. We found a protective effect of the intervention for shifting back to sick-listed between 6 and 18 months. The analyses showed that the probability of staying in the state returned to work was not different between the intervention and control groups at the end of the follow-up (3 years). CONCLUSIONS: We demonstrate that these alternative analyses give additional results and increase the strength of the analyses. The simple intervention did not decrease the probability of being on sick-leave in the long term; however, it decreased the time that individuals were on sick-leave.     

Mammographic Density and Intake of Selected Nutrients and Vitamins in Norwegian Women

Investigating the association between dietary factors and mammographic density (MD) could shed light on the relationship between diet and breast cancer risk. We took advantage of a national mammographic screening program to study the association between intake of nutrients and MD. In this study, we analyzed data of 2,252 postmenopausal women aged 50–69 yr who participated in the Norwegian Breast Cancer Screening Program in 2004. MD was assessed on digitized mammograms using a computer-assisted method. We used multivariate linear regression models to determine least square means of percent and absolute MD. Overall, we observed no associations between MD and intake of total calories, protein, carbohydrates, cholesterol, and dietary fiber. There was a positive borderline statistically significant association between absolute MD and total fat intake (P = 0.10) and between percent MD and intake of saturated fat (P = 0.06). There was no association between MD and intake of calcium, retinol, vitamins A, B12, C, or D, or combined intake of vitamin D and calcium. This study provides some evidence of an association between MD and dietary intake. Our study highlights the importance of adequate adjustments for BMI in studies of diet and MD.