mtDNA diversity in rhesus monkeys reveals overestimates of divergence time and paraphyly with neighboring species

Reconstructions of the human-African great ape phylogeny by using mitochondrial DNA (mtDNA) have been subject to considerable debate. One confounding factor may be the lack of data on intraspecific variation. To test this hypothesis, we examined the effect of intraspecific mtDNA diversity on the phylogenetic reconstruction of another Plio- Pleistocene radiation of higher primates, the fascicularis group of macaque (Macaca) monkey species. Fifteen endonucleases were used to identify 10 haplotypes of 40-47 restriction sites in M. mulatta, which were compared with similar data for the other members of this species group. Interpopulational, intraspecific mtDNA diversity was large (0.5%- 4.5%), and estimates of divergence time and branching order incorporating this variation were substantially different from those based on single representatives of each species. We conclude that intraspecific mtDNA diversity is substantial in at least some primate species. Consequently, without prior information on the extent of genetic diversity within a particular species, intraspecific variation must be assessed and accounted for when reconstructing primate phylogenies. Further, we question the reliability of hominoid mtDNA phylogenies, based as they are on one or a few representatives of each species, in an already depauperate superfamily of primates.      

Characterization of phosphate oxygen exchange reactions catalyzed by myosin through measurement of the distribution of 18-O-labeled species

The change in the distribution of the phosphate species containing 0 to 4 18O oxygens per Pi was investigated during medium Pi equilibrium HOH exchange catalyzed by myosin subfragment 1. At 25 degrees C, a Pi molecule once bound loses an average of 3.9 of its original 4 oxygens prior to release which means that at least 100 reversals of the exchange reaction must have occurred. At 0 degrees C, only 3.4 of the 4 oxygens are lost prior to release indicating an average of 17 reversals. Distribution patterns are consistent with equivalent participation in the exchange reactions of all 4 oxygens of bound Pi. The intermediate exchange of Pi oxygens during hydrolysis of 18O-labeled ATP by myosin has also been investigated. The distribution of the product Pi species shows that there is an ATPase component in myosin preparations which hydrolyzes ATP without intermediate exchange. Presence of this component, which is likely a contaminating ATPase, provides a simple explanation of the apparent nonequivalence of phosphate oxygens which has been observed. When correction is made for this contaminant, characteristics of the myosin intermediate Pi equilibrium HOH exchange are similar to those of myosin subfragment 1 medium exchange, and intermediate exchange data are in much closer agreement with other kinetic measurements.     

Effect of actin concentration on the intermediate oxygen exchange of myosin; relation to the refractory state and the mechanism of exchange.

The effect of actin concentration on the myosin catalyzed exchange of phosphate oxygens with water accompanying ATP hydrolysis has been investigated. The extent of exchange was found to extrapolate to zero at infinite actin concentration at 23 and 0 degrees C for myosin subfragments S1(A1) and S1(A2). This result is consistent with actin associating directly with the product of the hydrolysis step and is not readily consistent with refractory state schemes in which the entire flow goes via a dissociating pathway. The possibility of a refractory state in the form of a phosphorylated intermediate or a bound metaphosphate state with hydrolysis occurring in the transition to the refractory state merits consideration. A full analysis of the dependence of intermediate exchange on the rate constants of the acto-S1 scheme is given and the errors arising from other methods of analysis are discussed. The rate of oxygen exchange was measured as 10 s-1 (23 degrees C) a value comparable with but slightly lower than the rate of reversal of the ATP cleavage step.     

Actin mediated release of ATP from a myosin-ATP complex.

The apparent second-order rate constant, ka-2, of actin binding to a myosin-ATP state (M*.ATP) and releasing ATP to the medium has been determined by two methods. The first was the measurement of the amount of ATP released when actin was added to the intermediate state, M*.ATP; the second was the measurement of oxygen exchange between ATP and HOH. A quantitative treatment of ATP in equilibrium HOH exchange is given to allow extraction of elementary rate constants from the data. Agreement between the two methods was good and at low ionic strength and 23 degrees C, ka-2 is 6 X 10(5) M-1 s-1 which is about one-third the value of the apparent second-order rate constant, ka4, of actin binding to the myosin product state (M**.ADP.Pi). The determination of ka-2 allows a lower limit of 6 s-1 to be placed upon the first-order rate of ATP release from AM.ATP. This is to be compared with a value of less than or equal to 1.5 X 10(-4) s-1 for the equivalent steps of the myosin scheme; thus actin enhances the rate by a factor of 4 X 10(4) or more. A greater proportion of the bound ATP is released to the medium as ATP with increasing actin concentration. This reflects the contribution to rate limitation at saturating actin concentration of steps between myosin states dissociated from actin.     

Impact of Obstructive Sleep Apnea on the Levels of Placental Growth Factor (PlGF) and Their Value for Predicting Short-Term Adverse Outcomes in Patients with Acute Coronary Syndrome

Background

Placental growth factor (PlGF) induces angiogenesis and promotes tissue repair, and plasma PlGF levels change markedly during acute myocardial infarction (AMI). Currently, the impact of obstructive sleep apnea (OSA) in patients with AMI is a subject of debate. Our objective was to evaluate the relationships between PlGF levels and both the severity of acute coronary syndrome (ACS) and short-term outcomes after ACS in patients with and without OSA.

Methods

A total of 538 consecutive patients (312 OSA patients and 226 controls) admitted for ACS were included in this study. All patients underwent polygraphy in the first 72 hours after hospital admission. The severity of disease and short-term prognoses were evaluated during the hospitalization period. Plasma PlGF levels were measured using an electrochemiluminescence immunoassay.

Results

Patients with OSA were significantly older and more frequently hypertensive and had higher BMIs than those without OSA. After adjusting for age, smoking status, BMI and hypertension, PlGF levels were significantly elevated in patients with OSA compared with patients without OSA (19.9 pg/mL, interquartile range: 16.6–24.5 pg/mL; 18.5 pg/mL, interquartile range: 14.7–22.7 pg/mL; p<0.001), and a higher apnea-hypopnea index (AHI) was associated with higher PlGF concentrations (p<0.003). Patients with higher levels of PlGF had also an increased odds ratio for the presence of 3 or more diseased vessels and for a Killip score>1, even after adjustment.

Conclusions

The results of this study show that in patients with ACS, elevated plasma levels of PlGF are associated with the presence of OSA and with adverse outcomes during short-term follow-up.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01335087","term_id":"NCT01335087"}}NCT01335087     

Hidden-Markov methods for the analysis of single-molecule actomyosin displacement data: the variance-Hidden-Markov method

In single-molecule experiments on the interaction between myosin and actin, mechanical events are embedded in Brownian noise. Methods of detecting events have progressed from simple manual detection of shifts in the position record to threshold-based selection of intermittent periods of reduction in noise. However, none of these methods provides a "best fit" to the data. We have developed a Hidden-Markov algorithm that assumes a simple kinetic model for the actin-myosin interaction and provides automatic, threshold-free, maximum-likelihood detection of events. The method is developed for the case of a weakly trapped actin-bead dumbbell interacting with a stationary myosin molecule (Finer, J. T., R. M. Simmons, and J. A. Spudich. 1994. Nature. 368:113-119). The algorithm operates on the variance of bead position signals in a running window, and is tested using Monte Carlo simulations to formulate ways of determining the optimum window width. The working stroke is derived and corrected for actin-bead link compliance. With experimental data, we find that modulation of myosin binding by the helical structure of the actin filament complicates the determination of the working stroke; however, under conditions that produce a Gaussian distribution of bound levels (cf. Molloy, J. E., J. E. Burns, J. Kendrick-Jones, R. T. Tregear, and D. C. S. White. 1995. Nature. 378:209-212), four experiments gave working strokes in the range 5.4-6.3 nm for rabbit skeletal muscle myosin S1.     

Serosurvey of Brucella spp. infection in the Kafue lechwe (Kobus leche kafuensis) of the Kafue flats in Zambia

One of the diseases of veterinary and public health importance affecting the Kafue lechwe (Kobus leche kafuensis) on the Kafue flats is brucellosis, for which only scant information is available. During the 2003 (October), 2004 (December), and 2008 (July-December) hunting seasons in the Kafue flats, we conducted a study to determine the seroprevalence of Brucella spp. in the Kafue lechwe and to evaluate serologic tests for detection of Brucella spp. antibodies in lechwe. The Rose Bengal Test (RBT), competitive enzyme-linked immunosorbent assay (cELISA), and fluorescence polarization assay (FPA) were used. A total of 121 Kafue lechwe were hunted for disease investigations in 2003, 2004, and 2008 in the Kafue Flat Game Management Area. Of these, 21.6%, (95% confidence interval [CI]: 14.2-29.1%) had detectable antibodies to Brucella spp. The Kafue lechwe in Lochnivar National Park had higher antibody results than those in Blue Lagoon National Park (odds ratio=3.0; 95% CI: 0.94-9.4). Infection levels were similar in females (21.6%) and males (21.7%). Results were similar among RBT, FPA, cELISA tests, suggesting that these could effectively be used in diagnosing brucellosis in the Kafue lechwe. Our study demonstrates the presence of Brucella infections in the Kafue lechwe in two national parks located in the Kafue flats and further highlights the suitability of serologic assays for testing the Kafue lechwe. Because the Kafue lechwe is the most hunted wildlife species in Zambia, hunters need to be informed of the public health risk of Brucella spp. infection.     

No association between leptin levels and sleep duration or quality in obese adults

Previous research in lean subjects has found lower leptin levels associated with shorter sleep duration. Since leptin levels are higher and some of the actions of leptin are impaired in obese individuals, one cannot assume that sleep will be similarly associated with leptin in obese individuals. The aim of this paper was to examine the cross-sectional association between habitual sleep duration and quality and plasma leptin levels in a sample of 80 obese men and premenopausal women aged 18-50 years. Leptin levels (ng/ml) were assayed on a fasting blood sample taken in the morning. We calculated a relative leptin level by dividing leptin by body fat percentage. Sleep duration and sleep efficiency were measured by 2 weeks of wrist actigraphy and respiratory disturbance index (RDI), a measure of sleep disordered breathing, was assessed by a portable screening device on a single night. Mean leptin levels and body fat percentage were higher in women than men (P < 0.001), however, mean RDI was higher in men (P = 0.01). There were no significant associations between relative leptin level and any of the sleep measures, including sleep duration, sleep efficiency, and sleep disordered breathing. There was also no difference between men and women in the association between sleep and leptin. In conclusion, contrary to what has been reported in other studies, measures of sleep duration and quality were not associated with leptin levels in our sample of obese adults.     

Multiplexed expression and screening for recombinant protein production in mammalian cells

Background  

A variety of approaches to understanding protein structure and function require production of recombinant protein. Mammalian based expression systems have advantages over bacterial systems for certain classes of protein but can be slower and more laborious. Thus the availability of a simple system for production and rapid screening of constructs or conditions for mammalian expression would be of great benefit. To this end we have coupled an efficient recombinant protein production system based on transient transfection in HEK-293 EBNA1 (HEK-293E) suspension cells with a dot blot method allowing pre-screening of proteins expressed in cells in a high throughput manner.     

A new brace treatment similar for adolescent scoliosis and kyphosis based on restoration of thoracolumbar lordosis. Radiological and subjective clinical results after at least one year of treatment

Study design

A prospective treatment study with a new brace was conducted Objective. To evaluate radiological and subjective clinical results after one year conservative brace treatment with pressure onto lordosis at the thoracolumbar joint in children with scoliosis and kyphosis.

Summary of background data

Conservative brace treatment of adolescent scoliosis is not proven to be effective in terms of lasting correction. Conservative treatment in kyphotic deformities may lead to satisfactory correction. None of the brace or casting techniques is based on sagittal forces only applied at the thoracolumbar spine (TLI= thoracolumbar lordotic intervention). Previously we showed in patients with scoliosis after forced lordosis at the thoracolumbar spine a radiological instantaneous reduction in both coronal curves of double major scoliosis.

Methods

A consecutive series of 91 children with adolescent scoliosis and kyphosis were treated with a modified symmetric 30 degrees Boston brace to ensure only forced lordosis at the thoracolumbar spine. Scoliosis was defined with a Cobb angle of at least one of the curves [greater than or equal to] 25 degrees and kyphosis with or without a curve <25 degrees in the coronal plane. Standing radiographs were made i) at start, ii) in brace at beginning and iii) after one year treatment without brace.

Results

Before treatment start ??in brace?? radiographs showed a strong reduction of the Cobb angles in different curves in kyphosis and scoliosis groups (sagittal n = 5 all p < 0.001, pelvic obliquity p < 0.001). After one year of brace treatment in scoliosis and kyphosis group the measurements on radiographs made without brace revealed an improvement in 3 Cobb angles each.

Conclusion

Conservative treatment using thoracolumbar lordotic intervention in scoliotic and kyphotic deformities in adolescence demonstrates a marked improvement after one year also in clinical and postural criteria. An effect not obtained with current brace techniques.