Rheological characterization of neutral and anionic polysaccharides with reduced mucociliary transport rates

The purpose of this research was to compare the viscoelastic properties of several neutral and anionic polysaccharide polymers with their mucociliary transport rates (MTR) across explants of ciliated bovine tracheal tissue to identify rheologic parameters capable of predicting the extent of reduction in mucociliary transport. The viscoelastic properties of the polymer gels and gels mixed with mucus were quantified using controlled stress rheometry. In general, the anionic polysaccharides were more efficient at decreasing the mucociliary transport rate than were the neutral polymers, and a concentration threshold, where no further decreases in mucociliary transport occurred with increasing polymer concentration, was observed for several of the neutral polysaccharides. No single rheologic parameter (ν, G′, G″, tan δ, G^*) was a good predictor of the extent of mucociliary transport reduction, but a combination of the apparent viscosity (ν), tangent to the phase angle (tan δ), and complex modulus (G^*) was found to be useful in the identification of formulations capable of decreasing MTR. The relative values of each of the rheologic parameters were unique for each polymer, yet once the relationships between the rheologic parameters and mucociliary transport rate reduction were determined, formulations capable of resisting mucociliary clearance could be rapidly optimized. Published: April 20, 2007     

Head-to-tail regulation is critical for the in vivo function of myosin V

Cell organization requires regulated cargo transport along cytoskeletal elements. Myosin V motors are among the most conserved organelle motors and have been well characterized in both yeast and mammalian systems. Biochemical data for mammalian myosin V suggest that a head-to-tail autoinhibitory interaction is a primary means of regulation, but the in vivo significance of this interaction has not been studied. Here we generated and characterized mutations in the yeast myosin V Myo2p to reveal that it is regulated by a head-to-tail interaction and that loss of regulation renders the myosin V constitutively active. We show that an unregulated motor is very deleterious for growth, resulting in severe defects in Myo2-mediated transport processes, including secretory vesicle transport, mitochondrial inheritance, and nuclear orientation. All of the defects associated with motor misregulation could be rescued by artificially restoring regulation. Thus, spatial and temporal regulation of myosin V in vivo by a head-to-tail interaction is critical for the normal delivery functions of the motor.     

Engineering ethanologenic Escherichia coli for levoglucosan utilization

Levoglucosan is a major product of biomass pyrolysis. While this pyrolyzed biomass, also known as bio-oil, contains sugars that are an attractive fermentation substrate, commonly-used biocatalysts, such as Escherichia coli, lack the ability to metabolize this anhydrosugar. It has previously been shown that recombinant expression of the levoglucosan kinase enzyme enables use of levoglucosan as carbon and energy source. Here, ethanologenic E. coli KO11 was engineered for levoglucosan utilization by recombinant expression of levoglucosan kinase from Lipomyces starkeyi. Our engineering strategy uses a codon-optimized gene that has been chromosomally integrated within the pyruvate to ethanol (PET) operon and does not require additional antibiotics or inducers. Not only does this engineered strain use levoglucosan as sole carbon source, but it also ferments levoglucosan to ethanol. This work demonstrates that existing biocatalysts can be easily modified for levoglucosan utilization.     

Key issues in recruitment to randomised controlled trials with very different interventions: a qualitative investigation of recruitment to the SPARE trial (CRUK/07/011)

Background

Low back pain (LBP) is a recognized public health problem, impacting up to 80% of US adults at some point in their lives. Patients with LBP are utilizing integrative health care such as spinal manipulation (SM). SM is the therapeutic application of a load to specific body tissues or structures and can be divided into two broad categories: SM with a high-velocity low-amplitude load, or an impulse "thrust", (HVLA-SM) and SM with a low-velocity variable-amplitude load (LVVA-SM). There is evidence that sensorimotor function in people with LBP is altered. This study evaluates the sensorimotor function in the lumbopelvic region, as measured by postural sway, response to sudden load and repositioning accuracy, following SM to the lumbar and pelvic region when compared to a sham treatment.

Methods/Design

A total of 219 participants with acute, subacute or chronic low back pain are being recruited from the Quad Cities area located in Iowa and Illinois. They are allocated through a minimization algorithm in a 1:1:1 ratio to receive either 13 HVLA-SM treatments over 6 weeks, 13 LVVA-SM treatments over 6 weeks or 2 weeks of a sham treatment followed by 4 weeks of full spine "doctor's choice" SM. Sensorimotor function tests are performed before and immediately after treatment at baseline, week 2 and week 6. Self-report outcome assessments are also collected. The primary aims of this study are to 1) determine immediate pre to post changes in sensorimotor function as measured by postural sway following delivery of a single HVLA-SM or LVVA-SM treatment when compared to a sham treatment and 2) to determine changes from baseline to 2 weeks (4 treatments) of HVLA-SM or LVVA-SM compared to a sham treatment. Secondary aims include changes in response to sudden loads and lumbar repositioning accuracy at these endpoints, estimating sensorimotor function in the SM groups after 6 weeks of treatment, and exploring if changes in sensorimotor function are associated with changes in self-report outcome assessments.

Discussion

This study may provide clues to the sensorimotor mechanisms that explain observed functional deficits associated with LBP, as well as the mechanism of action of SM.

Trial registration

This trial is registered in ClinicalTrials.gov, with the ID number of NCT00830596, registered on January 27, 2009. The first participant was allocated on 30 January 2009 and the final participant was allocated on 17 March 2011.     

Differences between Belclare and Suffolk ewes in fertilization rate, embryo quality and accessory sperm number after cervical or laparoscopic artificial insemination

Ewe breed has been shown to have a major effect on pregnancy rates following cervical AI using frozen-thawed semen. The main objective of this study was to examine the differences between purebred Belclare and Suffolk ewes (multiparous) in fertilization rate, number of accessory sperm and stage of embryo development on day 6 after cervical or laparoscopic AI with frozen-thawed semen. In experiment 1, Belclare and Suffolk ewes were synchronized for 12 days and were either cervically inseminated (year 1: n=28 and 31; year 2: n=16 and 15, respectively) or laparoscopically inseminated (year 2: n=13 and 14). In experiment 2, superovulated Belclare (n=4) and Suffolk (n=13) ewes were laparoscopically inseminated. All ewes were slaughtered 6 days after AI; oocytes/embryos were recovered, morphologically graded and stained to assess the number of cells and accessory spermatozoa. Data from both experiments were combined for statistical analysis. The proportion of ewes with fertilized oocytes was significantly higher following laparoscopic AI compared with cervical AI (54% versus 19%). More Belclare than Suffolk ewes yielded fertilized oocyte(s) after cervical AI (34% versus 10%, P<0.02) but there was no difference after laparoscopic AI (62% versus 60%). From the ewes that yielded at least one fertilized oocyte the proportion of Belclare ewes with embryos at the morula/blastocyst stage was significantly greater than for Suffolk ewes (94% versus 59%, P<0.02). A higher proportion of Belclare than Suffolk ewes had evidence of sperm reaching the site of fertilization following cervical AI (39% versus 15%, P<0.02) but there was no difference after laparoscopic AI (62% versus 64%, P>0.8). Amongst the ewes with evidence of sperm at the site of fertilization, laparoscopic AI resulted in a higher number of sperm per oocyte/embryo or per ewe than cervical AI (P<0.01). These results suggested that the difference in pregnancy rate between Suffolk and Belclare ewes following cervical AI was due to: (i) sperm traversing the cervix and uterus in a higher proportion of Belclare than Suffolk ewes, leading to a higher incidence of fertilization and (ii) the lower developmental competence of fertilized oocytes from Suffolk ewes.     

Regulation of proliferation during central nervous system development

The retina is one of the best-characterized regions of the central nervous system (CNS) and has served as a model for many of the principles that now form the foundation for CNS development. In the past several years, a number of advances have been made in our understanding of the coordination of proliferation and cell fate specification during retinal development. In this review, we will draw on findings from studies of the retina and highlight similarities and differences in other regions in the CNS, namely the cerebellum and cortex. We will present a framework in which to pose challenges and outstanding questions for future studies on the coordination of proliferation and cell fate specification in the developing CNS.     

Genetics of species differences in the wild annual sunflowers, Helianthus annuus and H. petiolaris

Much of our knowledge of speciation genetics stems from quantitative trait locus (QTL) studies. However, interpretations of the size and distribution of QTL underlying species differences are complicated by differences in the way QTL magnitudes are estimated. Also, many studies fail to exploit information about QTL directions or to compare inter- and intraspecific QTL variation. Here, we comprehensively analyze an extensive QTL data set for an interspecific backcross between two wild annual sunflowers, Helianthus annuus and H. petiolaris, interpret different estimates of QTL magnitudes, identify trait groups that have diverged through selection, and compare inter- and intraspecific QTL magnitudes. Our results indicate that even minor QTL (in terms of backcross variance) may be surprisingly large compared to levels of standing variation in the parental species or phenotypic differences between them. Morphological traits, particularly flower morphology, were more strongly or consistently selected than life history or physiological traits. Also, intraspecific QTL were generally smaller than interspecific ones, consistent with the prediction that larger QTL are more likely to spread to fixation across a subdivided population. Our results inform the genetics of species differences in Helianthus and suggest an approach for the simultaneous mapping of inter- and intraspecific QTL.     

The Efficiency of Selenocysteine Incorporation Is Regulated by Translation Initiation Factors

Selenocysteine (Sec) incorporation is an essential process required for the production of at least 25 human selenoproteins. This unique amino acid is co-translationally incorporated at specific UGA codons that normally serve as termination signals. Recoding from stop to Sec involves a cis-acting Sec insertion sequence element in the 3′ untranslated region of selenoprotein mRNAs as well as Sec insertion sequence binding protein 2, Sec-tRNA^Sec, and the Sec-specific elongation factor, eEFSec. The interplay between recoding and termination at Sec codons has served as a focal point in researching the mechanism of Sec insertion, but the role of translation initiation has not been addressed. In this report, we show that the cricket paralysis virus intergenic internal ribosome entry site is able to support Sec incorporation, thus providing evidence that the canonical functions of translation initiation factors are not required. Additionally, we show that neither a 5′ cap nor a 3′ poly(A) tail enhances Sec incorporation. Interestingly, however, the presence of the internal ribosome entry site significantly decreases Sec incorporation efficiency, suggesting a role for translation initiation in regulating the efficiency of UGA recoding.     

Swimming in the sea hare Aplysia brasiliana: Cost of transport, parapodial morphometry, and swimming behavior

The energetics and behavior of the parapodial-swimming Aplysia brasiliana were investigated in order to compare net cost of transport (COT_net) between swimming and crawling, and to compare transport costs with other swimmers. Oxygen consumption (V_O2) increased with increasing animal mass for resting, crawling, and swimming animals. Slopes of the regressions of log V_O2 on log mass were 0.90, 0.91, and 0.89 for resting, crawling, and swimming, respectively. The regression for resting V_O2 on mass was significantly lower than regressions of crawling and swimming on mass, which fell into a statistically homogenous subgroup. During 4-h swimming bouts, parapodial beat frequency dropped by less than 10% of starting values after 2 h and then stabilized for the remainder of the trial, whereas velocity steadily decreased to about 70% of starting values over the 4-h period. Initial beat frequency (at the start of a swimming bout) was negatively related to body mass, varying from 1.1 beat s^− 1 for a 34 g individual to 0.7 beats s^− 1 for a 500 g individual. Final beat frequency (at the end of a swimming bout) was also negatively related to body mass, but had a significantly lower intercept than initial beat frequency. Neither initial swimming velocity nor final swimming velocity was related to mass, but final velocity was significantly lower than initial velocity. A 250 g A. brasiliana swam at 345 m h^− 1 and crawled at 7 m h^− 1. Swimming COT_net (0.1 ml O₂ kg^− 1 m^− 1) for a 250 g A. brasiliana was 50 times less than crawling COT_net (5.3 ml O₂ kg^− 1 m^− 1). While the crawling COT_net for A. brasiliana fell within the range of other marine gastropods, swimming COT_net was less than that of swimming crustaceans, and much less than another gastropod, Melibe leonina, that uses lateral bending to swim.     

Comparative studies of duodenal and macrophage ferroportin proteins

Intestinal epithelial cells and reticuloendothelial macrophages are, respectively, involved in diet iron absorption and heme iron recycling from senescent erythrocytes, two critical processes of iron homeostasis. These cells appear to use the same transporter, ferroportin (Slc40a1), to export iron. The aim of this study was to compare the localization, expression, and regulation of ferroportin in both duodenal and macrophage cells. Using a high-affinity purified polyclonal antibody, we analyzed the localization and expression of ferroportin protein in the spleen, liver, and duodenum isolated from normal mice as well as from well-characterized mouse models of altered iron homeostasis. Ferroportin was found to be predominantly expressed in enterocytes of the duodenum, in splenic macrophages, and in liver Kupffer cells. Interestingly, the protein species detected in these cells migrated differently on SDS-PAGE. These differences in apparent molecular masses were partly explained by posttranslational complex N-linked glycosylations. In addition, in enterocytes, the transporter was mostly expressed at the basolateral membrane, whereas in bone marrow-derived macrophages, ferroportin was found predominantly localized in the intracellular vesicular compartment. However, some microdomains positive for ferroportin were also detected at the plasma membrane of macrophages. Despite these differences, we observed a parallel upregulation of ferroportin expression in tissue macrophages and enterocytes in response to iron-restricted erythropoiesis, suggesting that iron homeostasis is likely maintained through coordinate expression of the iron exporter in both intestinal and phagocytic cells. Our data also confirm a predominant regulation of ferroportin through systemic regulator(s) likely including hepcidin.