The mutation rates of di-, tri- and tetranucleotide repeats in Drosophila melanogaster

In a recent study, we reported that the combined average mutation rate of 10 di-, 6 tri-, and 8 tetranucleotide repeats in Drosophila melanogaster was 6.3 x 10(-6) mutations per locus per generation, a rate substantially below that of microsatellite repeat units in mammals studied to date (range = 10(-2)-10(-5) per locus per generation). To obtain a more precise estimate of mutation rate for dinucleotide repeat motifs alone, we assayed 39 new dinucleotide repeat microsatellite loci in the mutation accumulation lines from our earlier study. Our estimate of mutation rate for a total of 49 dinucleotide repeats is 9.3 x 10(-6) per locus per generation, only slightly higher than the estimate from our earlier study. We also estimated the relative difference in microsatellite mutation rate among di-, tri-, and tetranucleotide repeats in the genome of D. melanogaster using a method based on population variation, and we found that tri- and tetranucleotide repeats mutate at rates 6.4 and 8.4 times slower than that of dinucleotide repeats, respectively. The slower mutation rates of tri- and tetranucleotide repeats appear to be associated with a relatively short repeat unit length of these repeat motifs in the genome of D. melanogaster. A positive correlation between repeat unit length and allelic variation suggests that mutation rate increases as the repeat unit lengths of microsatellites increase.     

Accuracy of the conductance catheter for measurement of ventricular volumes seen clinically: effects of electric field homogeneity and parallel conductance

Ultrasound returns from tissue display variations in amplitude on several spatial scales. Although large-scale variations result from factors such as attenuation, variations on smaller scales are caused by tissue characteristics such as variations in scatterer spacing and reflectance. These small scale variations cause a corresponding variation in the amplitude of the ultrasound return. A simple and direct method for detecting and quantifying periodicity in these variations in the presence of attenuation is described. The radiofrequency ultrasound return is first demodulated by full-wave rectification. The normalized power spectrum of the demodulated return then yields an index that we call the relative Fourier energy. Both computer simulations and in vitro experiments were performed in order to study how relative Fourier energy performed in discriminating between periodic and random scatterer distributions. Computer simulations demonstrated significant differences between the returns from periodic and random scatterer distributions. Ultrasound returns from aortic tissue yielded a relative Fourier energy index that was significantly different between normal vs. atherosclerotic tissue (normal: 0.868 +/- 0.076, mean +/- s.d., fibrofatty plaque: 0.705 +/- 0.109, p < 0.01 vs. normal, calcified plaque: 0.753 +/- 0.078, p < 0.01 vs. normal). In contrast, no difference was found in comparisons of overall reflectance.     

Exogenous fatty acids modulate the functional and cytotoxic responses of cultured pulmonary artery endothelial cells to oxidant stress

We previously reported that supplementation with exogenous fatty acids modulated the susceptibility of cultured pulmonary artery endothelial cells (PAEC) to oxidant-mediated cytotoxicity. The current study investigates the effects of fatty acids with increasing degrees of unsaturation on oxidant-mediated dysfunction and cytotoxicity in cultured porcine pulmonary artery and aortic endothelial cells (AEC). Monolayers supplemented with 0.1 mmol/L oleic (18:1), linoleic (18:2), or gamma-linolenic (18:3) acids were exposed to oxidant stress (100 mumol/L hydrogen peroxide (H2O2)) or to control conditions for 30 minutes. Gas chromatographic analysis of the PAEC fatty acids confirmed incorporation of supplemental fatty acids into PAEC lipids. Cytotoxicity, measured as the release of intracellular lactate dehydrogenase (LDH), and PAEC monolayer barrier function, assessed by measuring the monolayer clearance of Evans blue dye bound to albumin, were determined for 1 to 3 hours after oxidant stress. The PAEC and AEC demonstrated comparable responses to H2O2. Hydrogen peroxide caused increases in monolayer permeability and detachment of cells from the monolayer that were most attenuated by supplementation with 18:2 or 18:3, and to a lesser degree with 18:1. In contrast, H2O2-mediated LDH release was attenuated by supplementation with 18:1, whereas 18:2 and 18:3 potentiated cytotoxicity after exposure to H2O2. These results indicate that the relationship between PAEC lipid composition and oxidant susceptibility is complex and that the extent of fatty acid unsaturation does not predict the functional or cytotoxic responses of PAEC to oxidant stress. Furthermore, these results suggest that functional derangements may not correlate with traditional assays of cytotoxicity induced by oxidant injury in cultured endothelium.     

Long-term trends in tuberculosis. Comparison of age-cohort data between Hong Kong and England and Wales

Apoptosis is cellular suicide functionally opposite of mitosis. It plays an important role in tissue growth control and removal of damaged and premalignant cells. The decrease in death suppressor Bcl-2 protein level was implicated in the many types of apoptotic cell death. Because Bcl-2 protein was recently found to be cleaved during apoptosis induced by Fas ligation, IL-3 withdrawal, and alphavirus infection, we assessed whether Bcl-2 protein was also cleaved during the anticancer drug (VP-16)-induced apoptotic cell death in U937 cells. We found that Bcl-2 protein was cleaved in vivo and in vitro after the treatment of VP-16. We also found that caspase-3/CPP32, which was activated after VP-16 treatment, was responsible for the direct cleavage of Bcl-2 protein. The overexpression of the cleaved Bcl-2 fragment increased the sensitivity to VP-16 and promoted apoptotic cell death. Therefore, caspase-3/CPP32 accelerates VP-16-induced U937 cell apoptosis by cleaving death suppressor Bcl-2 protein to produce a death promoter Bcl-2 fragment.     

Chromosome translocations: segregation modes and strategies for preimplantation genetic diagnosis

Preimplantation genetic diagnosis (PGD) offers polymerase chain reaction tests for an increasing range of single gene defects, and fluorescence in situ hybridization tests for sex determination (for X-linked conditions) and for aneuploidy detection. Patients carrying chromosome translocations with a high reproductive risk are increasingly seeking to increase their chances of a normal pregnancy with the help of PGD, for which they present a special challenge. This paper describes the behaviour of reciprocal translocations at meiosis, discusses current methods of detecting meiotic outcomes at the preimplantation stage and outlines ways forward for preimplantation diagnosis of these common rearrangements. We also propose a more general strategy using recently developed chromosome-specific sub-telomeric probes, combined, if possible, with proximal probes, to form a strong diagnostic tool.     

Direct force measurements of insulin monomer-monomer interactions

Direct measurement of the forces involved in protein-protein and protein-receptor interactions can, in principle, provide insight necessary for the advancement of structural biology, molecular biology, and the development of therapeutic proteins. The protein insulin is illustrative in this respect as the mechanisms of insulin dimer dissociation and insulin-insulin receptor binding are crucial to the efficacy of insulin medications for the control of diabetes. Insulin molecules, modified with a photochemically active azido functionality on specific residues, were attached to force microscope tips and opposing mica surfaces in configurations that would either favor or disfavor dimer formation. Force curve measurements performed in buffer solution revealed the complexity of the insulin monomer-monomer interaction with multiple unbinding events occurring upon tip retraction, suggesting disruption of discrete molecular bonds at the monomer-monomer interface. Furthermore, the force curves exhibit long-range unbinding events, consistent with considerable elongation of the insulin molecule prior to dissociation. The unbinding forces observed in this study are the result of a combination of molecular disentanglement and dimer dissociation processes.     

Erythrocyte glutathione balance and membrane stability during preeclampsia

OBJECTIVE: To assess the risk of borderline ovarian cancer among infertile women treated with fertility drugs. DESIGN: Case-control study. SETTING: Nationwide data obtained from public registers and postal questionnaires. PATIENT(S): All Danish women <60 years old with borderline ovarian cancer during the period 1989-1994 and randomly selected population controls. The analysis included 231 cases and 1,721 controls. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Influence of parity, infertility, and fertility drugs on the risk of borderline ovarian cancer after multivariate confounder control. RESULT(S): The odds ratio (OR) for borderline ovarian cancer among infertile untreated nulliparous women compared with fertile nulliparous women was 1.9. The OR for borderline ovarian cancer among treated nulliparous women compared with untreated infertile nulliparous women was 1.5, and the OR among treated parous women compared with untreated infertile parous women was 1.5. CONCLUSION(S): Among fertile women, the difference in the risk of borderline ovarian cancer between nulliparous women and parous women was not statistically significant. Nulliparous women who were infertile and who did not receive medical treatment had a twofold higher risk of borderline ovarian cancer than fertile nulliparous women. There was no statistically significant increase in the risk of borderline ovarian cancer among nulliparous women who were treated with fertility drugs compared with nulliparous untreated infertile women or among parous women who were treated with fertility drugs compared with parous untreated infertile women.     

The effect of supervision of residents on quality of care in five university-affiliated emergency departments

BACKGROUND: The purpose of the study was to compare the analgesic and side effects of two epidurally administered mixtures of bupivacaine and fentanyl with the same drug ratios. METHODS: One hundred patients scheduled for colorectal surgery were randomized to receive a thoracic epidural infusion of either bupivacaine 0.12% with fentanyl 2 micrograms/ml or bupivacaine 0.24% with fentanyl 4 micrograms/ml during 48 h postoperatively. The pumps were adjusted to keep the visual analogue scale (VAS) pain score at 3 or less (on a scale of 0-10) with a minimum of adverse effects. RESULTS: There were no statistically significantly differences between the two groups in VAS pain scores. The average VAS pain score resting varied between 0.5 and 1, and coughing between 1.9 and 3.4. One case of respiratory depression with breathing frequency 7 occurred in each group, but none of the patients required naloxone. One patient in the low concentration group developed partial motor weakness in both legs 36 h postoperatively. Equal drug amounts--bupivacaine 10.8-11 mg/h and fentanyl 18-18.4 micrograms/h--were given in both groups throughout the study. CONCLUSIONS: Both groups had low pain scores with few and comparable adverse effects. It thus seems that the volume is not important when mixtures of bupivacaine and fentanyl in the studies concentrations are infused epidurally at a low thoracic level. Practical reasons favour the higher concentration mixture.     

Function of the c-Myc antagonist Mad1 during a molecular switch from proliferation to differentiation

Mad-Max heterodimers have been shown to antagonize Myc transforming activity by a mechanism requiring multiple protein-protein and protein-DNA interactions. However, the mechanism by which Mad functions in differentiation is unknown. Here, we present evidence that Mad functions by an active repression mechanism to antagonize the growth-promoting function(s) of Myc and bring about a transition from cellular proliferation to differentiation. We demonstrate that exogenously expressed c-Myc blocks inducer-mediated differentiation of murine erythroleukemia cells without disrupting the induction of endogenous Mad; rather, high levels of c-Myc prevent a heterocomplex switch from growth-promoting Myc-Max to growth-inhibitory Mad-Max. Cotransfection of a constitutive c-myc with a zinc-inducible mad1 results in clones expressing both genes, whereby a switch from proliferation to differentiation can be modulated. Whereas cells grown in N'N'-hexamethylene bisacetamide in the absence of zinc fail to differentiate, addition of zinc up-regulates Mad expression by severalfold and differentiation proceeds normally. Coimmunoprecipitation analysis reveals that Mad-Max complexes are in excess of Myc-Max in these cotransfectants. Moreover, we show that the Sin-binding, basic region, and leucine zipper motifs are required for Mad to function during a molecular switch from proliferation to differentiation.     

Determinants of the pulmonary artery pressure rise in left ventricular dysfunction

Pulmonary artery hypertension in patients with left ventricular dysfunction is related to poor outcome but the role of cardiac functional abnormalities in the genesis of pulmonary hypertension remains unknown. The aim of this prospective study was to identify the determinants of pulmonary hypertension in 102 consecutive patients with primary left ventricular dysfunction (ejection fraction < 50%). Systolic pulmonary artery pressure was measured by continuous wave Doppler. Left ventricular systolic and diastolic function, severity of functional mitral regurgitation, cardiac output, and left atrial volume were assessed using Doppler echocardiography. In patients with left ventricular dysfunction, systolic pulmonary artery pressure was increased (51 +/- 14 mmHg, range 23 to 87 mmHg). Mitral deceleration time (r = -0.61; p = 0.0001) and mitral effective regurgitant orifice (r = 0.50; p = 0.0001) were the strongest parameters related to systolic pulmonary artery pressure. Multivariate analysis identified these two variables as the strongest predictors of systolic pulmonary artery pressure in association with the mitral E/A ratio (p = 0.006) and age (p = 0.005). In conclusion, pulmonary hypertension is common and variable in patients with left ventricular dysfunction. It is closely related to diastolic dysfunction and severity of functional mitral regurgitation but not independently to the degree of left ventricular systolic dysfunction. These findings underline the importance of assessing diastolic function and quantifying mitral regurgitation in patients with left ventricular dysfunction.