The effectiveness of traditional and sling exercise strength training in women

Strength training often combines closed-kinetic-chain exercises (CKCEs) and open kinetic-chain exercises (OKCEs). The CKCE may be more effective for improving performance in lower-body training. Recently, we reported upper-body CKCE (using a commercially available system of ropes and slings, Redcord AS, Staubo, Norway) was as effective as OKCE training for strength gains and that CKCE was more effective than OKCE for improving throwing performance. To our knowledge the effectiveness of a strength training program that uses exclusively CKCE is unknown. In this study, we examined the effectiveness of CKCE vs. OKCE strength training programs in women enrolled in an introductory strength training program. Twenty-six participants were randomized to OKCE (traditional exercises) or CKCE (sling-based exercises). Participants completed 6 sets per week for 13 weeks. Pre and posttraining evaluations included the following: 1 repetition maximum (1RM) leg and bench press; sling exercise push-ups; isokinetic dynamometry; lateral step-down test; and the Star Excursion Balance Test. Both groups significantly improved bench press (by an average of 4-6 kg) and leg press (by an average of 23-35 kg) (p < 0.001). There was a significant group × time interaction (p < 0.001) for sling exercise push-ups (OKCE pre = 5.5 ± 8.6, OKCE post = 6.1 ± 8.2, CKCE pre = 6.8 ± 6.0, CKCE post = 16.9 ± 6.6). Isokinetic measures of knee extension, knee flexion, shoulder internal rotation, and shoulder external rotation increased (improvements ranged from 2.7 to 27.7%), with no group differences. Both OKCE and CKCE strength training elicited similar changes in balance. We conclude that CKCE training is equally as effective as OKCE training during the initial phases of a strength training program in women. The fact that only CKCE improved sling exercise push-ups supports previous findings suggesting functional superiority of CKCE.     

Ecological behavior of Lactobacillus reuteri 100-23 is affected by mutation of the luxS gene

The luxS gene of Lactobacillus reuteri 100-23C was amplified by PCR, cloned, and then sequenced. To define a physiological and ecological role for the luxS gene in L. reuteri 100-23C, a luxS mutant was constructed by insertional mutagenesis. The luxS mutant did not produce autoinducers AI-2 or AI-3. Complementation of the luxS mutation by a plasmid construct containing luxS restored AI-2 and AI-3 synthesis. In vitro experiments revealed that neither the growth rate, nor the cell yield, nor cell survival in the stationary phase were compromised in the luxS mutant relative to the wild type and complemented mutant. The ATP content of exponentially growing cells of the luxS mutant was, however, 65% of that of wild-type cells. Biofilms formed by the luxS mutant on plastic surfaces in a bioreactor were thicker than those formed by the wild type. Biofilm thickness was not restored to wild-type values by the addition of purified AI-2 to the culture medium. In vivo experiments, conducted with ex-Lactobacillus-free mice, showed that biofilms formed by the mutant strain on the epithelial surface of the forestomach were approximately twice as thick as those formed by the wild type. The ecological performance of the luxS mutant, when in competition with L. reuteri strain 100-93 in the mouse cecum, was reduced compared to that of a xylA mutant of 100-23C. These results demonstrate that LuxS influences important ecological attributes of L. reuteri 100-23C, the consequences of which are niche specific.     

Specificity patterns indicate that auxin exporters and receptors are the same proteins

A study of transport and action of synthetic auxin analogues can help to identify transporters and receptors of this plant hormone. Both aspects--transportability and action on growth--were tested with 2-naphthoxyacetic acid (2-NOA) and compared across several plant species. 2-NOA stimulates elongation effectively at low concentrations in petioles of the gymnosperm Ginkgo biloba L., in hypocotyls or internodes of the dicot legumes, mung bean (Vigna mungo L.) and pea (Pisum sativum L.), in cotyledons of onion (Allium cepa L.) and in leaf bases of chive (Allium schoenoprasum L.), the latter two of the monocot order Asparagales. In contrast, elongation of coleoptile segments of maize (Zea mays L.) is poorly responsive to 2-NOA. Significant auxin-like transport of 2-NOA was observed in segments of mung bean hypocotyls, pea internodes, and chive leaf bases, but not in segments of the grass coleoptiles. Thus, for the two assays, elongation and polar transportability, the same difference in ligand specificity was observed between the grass and all other species assayed. This finding supports the hypothesis that a common protein mediates auxin efflux as well as auxin action on elongation.     

KAT1 inactivates at sub-millimolar concentrations of external potassium

Structural analysis of K+ channel pores suggests that the selectivity filter of the pore is an inherent sensor for extracellular K+ (Ko+); channels seem to be inactivated at low Ko+ because of a destabilization of the conducting state and a collapse of the pore. In the present study, the effect of depleting Ko+ on the activity of a plant K+ channel, KAT1, from Arabidopsis thaliana was investigated. This channel is thought to be insensitive to Ko+. The channel was therefore expressed in mammalian HEK293 cells and measured with patch clamp technology in the whole cell configuration. The effect of Ko+ depletion on channel activity was monitored from the tail currents before, during, and after washing Ko+ from the medium. The data show that a depletion of Ko+ results in a decrease in channel conductance, irrespective of whether K+ is simply removed or replaced by either Na+ or Li+. Quantitative analysis suggests that the channel has two binding sites for K+ with the dissociation constant in the order of 20 microM. This high sensitivity of the channel to Ko+ could serve as a safety mechanism, which inactivates the channel at low Ko+ and, in this way, prevents leakage of K+ from the cells via this type of channel.     

A structural basis for the acute effects of HIV protease inhibitors on GLUT4 intrinsic activity

Human immunodeficiency virus (HIV) protease inhibitors (PIs) act as reversible noncompetitive inhibitors of GLUT4 with binding affinities in the low micromolar range and are known to contribute to alterations in glucose homeostasis during treatment of HIV infection. As aspartyl protease inhibitors, these compounds all possess a core peptidomimetic structure together with flanking hydrophobic moieties. To determine the molecular basis for GLUT4 inhibition, a family of related oligopeptides containing structural elements found in PIs was screened for their ability to inhibit 2-deoxyglucose transport in primary rat adipocytes. The peptide oxybenzylcarbonyl-His-Phe-Phe-O-ethyl ester (zHFFe) was identified as a potent inhibitor of zero-trans glucose flux with a K(i) of 26 mum. Similar to PIs, transport inhibition by this peptide was acute, noncompetitive, and reversible. Within a Xenopus oocyte expression system, zHFFe acutely and reversibly inhibited GLUT4-mediated glucose uptake, whereas GLUT1 activity was unaffected at concentrations as high as 1 mm. The related photoactivatable peptide zHFF-p-benzoylphenylalanine-[(125)I]Tyr-O-ethyl ester selectively labeled GLUT4 in rat adipocytes and indinavir effectively protected against photolabeling. Furthermore, GLUT4 bound to a peptide affinity column containing the zHFF sequence and was eluted by indinavir. These data establish a structural basis for PI effects on GLUT4 activity and support the direct binding of PIs to the transport protein as the mechanism for acute inhibition of insulin-stimulated glucose uptake.     

RASCAL Is a New Human Cytomegalovirus-Encoded Protein That Localizes to the Nuclear Lamina and in Cytoplasmic Vesicles at Late Times Postinfection

The products of numerous open reading frames (ORFs) present in the genome of human cytomegalovirus (CMV) have not been characterized. Here, we describe the identification of a new CMV protein localizing to the nuclear envelope and in cytoplasmic vesicles at late times postinfection. Based on this distinctive localization pattern, we called this new protein nuclear rim-associated cytomegaloviral protein, or RASCAL. Two RASCAL isoforms exist, a short version of 97 amino acids encoded by the majority of CMV strains and a longer version of 176 amino acids encoded by the Towne, Toledo, HAN20, and HAN38 strains. Both isoforms colocalize with lamin B in deep intranuclear invaginations of the inner nuclear membrane (INM) and in novel cytoplasmic vesicular structures possibly derived from the nuclear envelope. INM infoldings have been previously described as sites of nucleocapsid egress, which is mediated by the localized disruption of the nuclear lamina, promoted by the activities of viral and cellular kinases recruited by the lamina-associated proteins UL50 and UL53. RASCAL accumulation at the nuclear membrane required the presence of UL50 but not of UL53. RASCAL and UL50 also appeared to specifically interact, suggesting that RASCAL is a new component of the nuclear egress complex (NEC) and possibly involved in mediating nucleocapsid egress from the nucleus. Finally, the presence of RASCAL within cytoplasmic vesicles raises the intriguing possibility that this protein might participate in additional steps of virion maturation occurring after capsid release from the nucleus.Cytomegalovirus (CMV) is a highly prevalent betaherpesvirus that can cause severe multiorgan disease in immunocompromised individuals (45). The ability of this virus to infect an exceptionally wide variety of different cell types substantially contributes to pathogenesis (5, 68). CMV tropism is largely determined by a finely tuned interplay between cellular and viral factors, many of which act at the earliest stages of infection (30, 68). We recently showed that the cellular protein vimentin is required for efficient onset of infection in primary human foreskin fibroblasts (HF). Interestingly, the degree of reliance on the presence and integrity of vimentin intermediate filaments is dependent on the virus strain, with the broadly tropic strain TB40/E being more negatively affected than the HF-adapted, attenuated strain AD169 (44).Serial passage of clinical isolates in HF or in endothelial cells (EC) has produced strains with different tropisms. The attenuated strains AD169 and Towne were developed as vaccine candidates by propagation in HF for more than 50 (AD169) and 125 (Towne) serial passages (19, 53, 61). During this process, both strains, compared to clinical isolates, accumulated multiple mutations and genomic deletions, resulting in the loss of more than 19 open reading frames (ORFs) (8). The number of serial passages in HF of another commonly used strain, Toledo, has been more moderate (19, 54, 58). This, however, did not prevent the emergence of numerous genomic mutations, including the inversion of an ∼15-kb fragment (8, 16, 56). As a consequence of these changes, productive infections by AD169, Towne, and Toledo are largely restricted to HF. In contrast, propagation of clinical isolates in EC has yielded a series of strains with more-intact genomes and broader tropisms, such as TB40/E, VHL/E, and FIX (VR1814) (25, 60, 71). These strains retain the ability to grow in a wider variety of cell types, including EC, epithelial cells, and dendritic cells (DC), in addition to HF (23, 28, 59, 60, 68).The UL128, UL130, and UL131A gene products were recently identified as essential mediators of CMV infection of EC and epithelial cells (26, 72, 73) and of virus transfer from infected EC to monocyte-derived DC (23). Each of these proteins is independently required for the broader tropisms of EC-propagated CMV isolates (63, 64), and the presence of mutations affecting their functionality has been directly linked to the inability of AD169, Towne, and Toledo to initiate productive infections in EC and epithelial cells (26, 72, 73).We have shown that mature Langerhans-type DC differentiated in vitro from CD34⁺ hematopoietic progenitor cells are highly permissive to direct infection with TB40/E or VHL/E, with 48 to 72% of cells in culture expressing the viral immediate-early genes IE1 and IE2 at 48 h postinfection (hpi) (28). In contrast, only 2 to 5% and 0% of mature Langerhans cells were IE1/IE2 positive after exposure to Towne and Toledo, respectively. However, productive infection was detected in 12 to 17% of cells infected with AD169, despite the fact that this strain lacks expression of the UL131A gene as a consequence of a frameshift mutation (26). These results suggested the existence of additional viral genes with products involved in mediating tropisms for specific cell types, such as DC. To identify possible candidates, we compared the amino acid sequence of each ORF found in the genome of TB40-BAC4, a sequenced clone of the TB40/E strain in a bacterial artificial chromosome (BAC) (GenBank accession number {"type":"entrez-nucleotide","attrs":{"text":"EF999921","term_id":"157779983","term_text":"EF999921"}}EF999921) (69), to the sequence of each ORF found in AD169 and AD169-BAC (accession numbers {"type":"entrez-nucleotide","attrs":{"text":"X17403","term_id":"59591","term_text":"X17403"}}X17403 and {"type":"entrez-nucleotide","attrs":{"text":"AC146999","term_id":"38093733","term_text":"AC146999"}}AC146999) (10, 49), Towne and Towne-BAC (accession numbers {"type":"entrez-nucleotide","attrs":{"text":"FJ616285","term_id":"222354438","term_text":"FJ616285"}}FJ616285, {"type":"entrez-nucleotide","attrs":{"text":"AC146851","term_id":"37654163","term_text":"AC146851"}}AC146851, and {"type":"entrez-nucleotide","attrs":{"text":"AY315197","term_id":"124303521","term_text":"AY315197"}}AY315197) (17, 18, 49), and Toledo-BAC (accession number {"type":"entrez-nucleotide","attrs":{"text":"AC146905","term_id":"37777314","term_text":"AC146905"}}AC146905) (49). The product of a putative ORF, originally identified by Murphy et al. and named conserved ORF 29 (c-ORF29) (49), was considered of particular interest because the amino acid sequence of the putative protein encoded by Toledo and Towne was extended by 79 residues compared to the putative protein encoded by TB40/E and AD169. This led to our speculation that that the extended version might result in a nonfunctional version of the c-ORF29-encoded protein. We thus focused our studies on the products of this ORF.Here, we show for the first time that CMV c-ORF29 encodes a protein expressed at early to late times postinfection (p.i.) and localizes to the nuclear rim in peculiar invaginations of the nuclear lamina and in cytoplasmic vesicular structures at late times p.i. Based on this localization pattern, we named this gene product nuclear rim-associated cytomegaloviral protein, or RASCAL. Surprisingly, no difference was observed in the distributions of RASCAL during infection of HF with TB40/E or Towne, suggesting that the intracellular trafficking of this protein is not affected by the presence of the additional residues at the C terminus of RASCAL from strain Towne (RASCAL_Towne). Ectopic expression of RASCAL in human embryo kidney 293T (HEK293T) cells further revealed that this protein requires the presence of the nuclear egress complex (NEC) member UL50 to reach the nuclear rim, while coimmunoprecipitation (co-IP) assays provided evidence for the existence of an interaction between RASCAL and UL50. These findings suggest that RASCAL may be a new component of the NEC with possible roles in remodeling the nuclear lamina during nucleocapsid egress from the nucleus.     

Oral cavity as natural reservoir for intestinal lactobacilli

Ecological studies indicate that most Lactobacillus species found in the human gastrointestinal tract are likely to be transient (allochthonous), originating from either the oral cavity or food. In order to investigate if oral lactobacilli constitute a part of the faecal Lactobacillus biota, the Lactobacillus biota of saliva and faeces of three human subjects were investigated and compared at two time-points in a 3-months interval. Bacteriological culture, performed by incubation under standard (37 degrees C, anaerobic) and alternative (30 degrees C, microaerobic) conditions, as well as PCR-DGGE with group-specific primers were used to characterize the predominant lactobacilli. Cell counts varied among the subjects and over time, reaching up to 10(7)CFU/ml in saliva and 5 x 10(6)CFU/g in faecal samples. The species composition of the Lactobacillus biota of human saliva and faeces was found to be subject-specific and fluctuated to some degree, but the species Lactobacillus gasseri, Lactobacillus paracasei, Lactobacillus rhamnosus and Lactobacillus vaginalis were detected at both time-points in saliva and faecal samples of individual subjects. RAPD-PCR analysis indicated that several strains of these species were present both in the oral cavity and in the faecal samples of the same subject. Oral isolates of the species L. gasseri and L. vaginalis showing identical RAPD types were found to persist over time, suggesting that these species are autochthonous to the oral cavity. Our results together with recent published data give strong evidence that some lactobacilli found in human faeces are allochthonous to the intestine and originate from the oral cavity.     

Spatial Differentiation in the Characterisation of Photochemical Ozone Formation: The EDIP2003 Methodology

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DOI: http://dx.doi.org/10.1065/lca2006.04.014

Background, Aims and Scope

In the life cycle of a product, emissions take place at many different locations. The location of the source and its surrounding conditions influence the fate of the emitted pollutant and the subsequent exposure it causes. This source of variation is normally neglected in Life Cycle Impact Assessment (LCIA), although it is well known that the impacts predicted by site-generic LCIA in some cases differ significantly from the actual impacts. Environmental impacts of photochemical ozone (ground-level ozone) depend on parameters with a considerable geographical variability (like emission patterns and population densities). A spatially differentiated characterisation model thus seems relevant.

Methods

and Results. The European RAINS model is applied for calculation of site-dependent characterisation factors for Non-Methane Volatile Organic Compounds (NMVOCs) and nitrogen oxides (NOx) for 41 countries or regions within Europe, and compatible characterisation factors for carbon monoxide (CO) are developed based on expert judgement. These factors are presented for three emission years (1990, 1995 and 2010), and they address human health impacts and vegetation impacts in two separate impacts categories, derived from AOT40 and AOT60 values respectively. Compatible site-generic characterisation factors for NMVOC, NOx, CO and methane (CH4) are calculated as emission-weighted European averages to be applied on emissions for which the location is unknown. The site-generic and site-dependent characterisation factors are part of the EDIP2003 LCIA methodology. The factors are applied in a specific case study, and it is demonstrated how the inclusion of spatial differentiation may alter the results of the photochemical ozone characterisation of life cycle impact assessment.

Discussion

and Conclusions. Compared to traditional midpoint characterisation modelling, this novel approach is spatially resolved and comprises a larger part of the cause-effect chain including exposure assessment and exceeding of threshold values. This positions it closer to endpoint modelling and makes the results easier to interpret. In addition, the developed model allows inclusion of the contributions from NOx, which are ne- glected when applying the traditional approaches based on Photochemical Ozone Creation Potentials (POCPs). The variation in site-dependent characterisation factors is far larger than the variation in POCP factors. It thus seems more important to represent the spatially determined variation in exposure than the difference in POCP among the substances.

     

Genomic splice-site analysis reveals frequent alternative splicing close to the dominant splice site

Alternative pre-mRNA splicing may be the most efficient and widespread mechanism to generate multiple protein isoforms from single genes. Here, we describe the genomic analysis of one of the most frequent types of alternative pre-mRNA splicing, alternative 5'- and 3'-splice-site selection. Using an EST-based alternative splicing database recording >47,000 alternative splicing events, we determined the frequency and location of alternative 5'- and 3'-splice sites within the human genome. The most common alternative splice sites used in the human genome are located within 6 nucleotides (nt) of the dominant splice site. We show that the EST database overrepresents alternative splicing events that maintain the reading frame, thus supporting the concept that RNA quality-control steps ensure that mRNAs that encode for potentially harmful protein products are destroyed and do not serve as templates for translation. The most frequent location for alternative 5'-splice sites is 4 nt upstream or downstream from the dominant splice site. Sequence analysis suggests that this preference is a consequence of the U1 snRNP binding sequence at the 5'-splice site, which frequently contains a GU dinucleotide 4 nt downstream from the dominant splice site. Surprisingly, approximately 50% of duplicated 3'-YAG splice junctions are subject to alternative splicing. This high probability of alternative 3'-splice-site activation in close proximity of the dominant 3'-splice site suggests that the second step of the splicing may be prone to violate splicing fidelity.     

ERCC1, XPD and RAI mRNA levels in lymphocytes are not associated with lung cancer risk in a prospective study of Danes

Low DNA-repair capacity has been associated with increased risk of several types of cancer. mRNA levels of the nucleotide excision repair genes ERCC1 and XPD have been shown to correlate with the DNA-repair capacity. Likewise, mRNA levels of several DNA-repair genes including ERCC1 have been shown to be lower in lymphocytes from patients with lung cancer and head and neck cancer compared with healthy persons. In these studies, the low DNA-repair gene expression levels could be either a risk factor for disease or a consequence of the same. In this nested case-cohort study, which to our knowledge, is the first prospective study of DNA-repair gene mRNA levels as predictors of lung cancer, we have investigated the occurrence of lung cancer in relation to the mRNA level of the two DNA-repair genes ERCC1 and XPD and the NF kappaB inhibitor RAI in blood samples prior to disease. Among 54,220 members of a Danish prospective cohort study, 265 lung cancer cases were identified and a sub-cohort comprising 272 individuals was used for comparison. The expression levels of the three adjacent genes were found to be highly inter-correlated, to be higher in women compared to men and to be lower in older individuals. The incidence rate ratios for lung cancer in association with one log-unit increase (natural logarithm) in mRNA levels were 1.12 (CI=0.89-1.41) for ERCC1, 1.00 (CI=0.83-1.21) for XPD and 1.25 (0.89-1.74) for RAI. In conclusion, this study indicated no association between mRNA expression of the DNA-repair genes ERCC1 and XPD and risk of subsequent development of lung cancer.