Control of breathing during sleep assessed by proportional assist ventilation

Meza, S., E. Giannouli, and M. Younes. Control ofbreathing during sleep assessed by proportional assist ventilation. J. Appl. Physiol. 84(1): 3-12, 1998.We used proportional assist ventilation (PAV) to evaluate thesources of respiratory drive during sleep. PAV increases the slope ofthe relation between tidal volume(VT) andrespiratory muscle pressure output (Pmus). We reasoned that ifrespiratory drive is dominated by chemical factors, progressiveincrease of PAV gain should result in only a small increase inVT because Pmus would bedownregulated substantially as a result of small decreases inPCO₂. In the presence of substantialnonchemical sources of drive [believed to be the case inrapid-eye-movement (REM) sleep] PAV should result in a substantial increase in minute ventilation and reductionin PCO₂ as the output related to thechemically insensitive drive source is amplified severalfold. Twelvenormal subjects underwent polysomnography while connected to a PAVventilator. Continuous positive air pressure (5.2 ± 2.0 cmH₂O) was administered tostabilize the upper airway. PAV was increased in 2-min steps from 0 to20, 40, 60, 80, and 90% of the subject's elastance and resistance.VT, respiratory rate, minuteventilation, and end-tidal CO₂pressure were measured at the different levels, and Pmus wascalculated. Observations were obtained in stage 2 sleep (n = 12), slow-wave sleep(n = 11), and REM sleep(n = 7). In all cases, Pmus wassubstantially downregulated with increase in assist so that theincrease in VT, althoughsignificant (P < 0.05), was small(0.08 liter at the highest assist). There was no difference in responsebetween REM and non-REM sleep. We conclude that respiratory driveduring sleep is dominated by chemical control and that there is nofundamental difference between REM and non-REM sleep in this regard.REM sleep appears to simply add bidirectional noise to what isbasically a chemically controlled respiratory output.

     

A single tRNA base pair mediates bacterial tRNA-dependent biosynthesis of asparagine

In many prokaryotes and in organelles asparagine and glutamine are formed by a tRNA-dependent amidotransferase (AdT) that catalyzes amidation of aspartate and glutamate, respectively, mischarged on tRNA^Asn and tRNA^Gln. These pathways supply the deficiency of the organism in asparaginyl- and glutaminyl-tRNA synthtetases and provide the translational machinery with Asn-tRNA^Asn and Gln-tRNA^Gln. So far, nothing is known about the structural elements that confer to tRNA the role of a specific cofactor in the formation of the cognate amino acid. We show herein, using aspartylated tRNA^Asn and tRNA^Asp variants, that amidation of Asp acylating tRNA^Asn is promoted by the base pair U₁–A₇₂ whereas the G₁–C₇₂ pair and presence of the supernumerary nucleotide U_20A in the D-loop of tRNA^Asp prevent amidation. We predict, based on comparison of tRNA^Gln and tRNA^Glu sequence alignments from bacteria using the AdT-dependent pathway to form Gln-tRNA^Gln, that the same combination of nucleotides also rules specific tRNA-dependent formation of Gln. In contrast, we show that the tRNA-dependent conversion of Asp into Asn by archaeal AdT is mainly mediated by nucleotides G₄₆ and U₄₇ of the variable region. In the light of these results we propose that bacterial and archaeal AdTs use kingdom-specific signals to catalyze the tRNA-dependent formations of Asn and Gln.     

Gelation of high methoxy pectin by acidification with d-glucono-δ-lactone (GDL) at room temperature

Rheological comparisons have been made between preparations of high methoxy pectin (DE ≈ 70%) gelled by acidification with d-glucono-δ-lactone (GDL) on holding for 16 h at 25 °C in the presence of 60 wt% sucrose, and otherwise identical preparations gelled by acidification with citric acid at high temperature and cooling from 90 to 25 °C at 1 °C/min. Two series of experiments were carried out for both methods of acidification. In the first series, the concentration of pectin (c) was held constant at 1.0 wt% and the final pH attained after holding (with GDL) or cooling (with citric acid) was varied from 3.75 to 2.25. In the second series, the final pH was held constant at 3.0 and c was varied from 0.25 to 2.00 wt%. All samples were then heated (1 °C/min) from 25 to 90 °C. Rheological changes on cooling/holding and heating were characterised by low-amplitude oscillatory measurements of storage modulus (G′) and loss modulus (G′′) at 1 rad s^?1 and 0.5% strain, and mechanical spectra were recorded at 25 °C. Selected samples, gelled with GDL, were also characterised by compression testing (at 25 °C), and a direct linear relationship was found between the logarithm of yield stress and log G′.The concentration-dependence of moduli for the samples acidified to pH 3.0 with GDL had the form typical of biopolymer gels, with log G′ versus log c approaching a limiting slope of 2 as c was raised above the minimum critical gelling concentration (c_o ≈ 0.3 wt%). Under all conditions of pH and pectin concentration studied, the values of G′′ (at 25 °C) for the samples acidified with citric acid were higher than those of the corresponding GDL-induced networks. The values of G′ were also higher, except at very low pH (below ～2.7 at c = 1.0 wt%) or very high concentrations of pectin. At pectin concentrations above ～1.5 wt%, the moduli of the samples gelled with citric acid (at pH 3.0) levelled out, or decreased slightly, with the values of G′ dropping below those of the GDL-induced networks towards the end of the concentration range studied (at c ≈ 2 wt%). All samples acidified with citric acid showed gel-like response (G′ > G′′) at 90 °C, attributed to hydrophobic association. The downturn in moduli at 25 °C for high concentrations of pectin is attributed to formation and disruption of strong networks during mixing with citric acid at high temperature (“pregelation”). It is suggested, however, that “weak gels” formed at lower concentrations or at pH values above ～2.7 may enhance gel properties by preserving a continuous network as hydrophobic junctions dissociate on cooling and are replaced by hydrogen-bonded junctions, in contrast to random percolation during gelation with GDL at 25 °C. On re-heating from 25 to 90 °C, the reverse processes (dissociation of hydrogen-bonded structures and formation of hydrophobic associations) were evident in an initial reduction and subsequent increase in moduli, as observed in previous studies. Similar heating traces were obtained for samples acidified with GDL to pH values above ～3.0 (at c = 1.0 wt%) or with pectin concentrations below ～1.0 wt% (at pH 3.0). However, at higher concentrations or lower values of pH (i.e. conditions favourable to extensive intermolecular association) an abrupt decrease in G′, with an accompanying maximum in G′′, was observed on heating through the temperature range ～60–80 °C. This is attributed to excessive hydrophobic association, causing collapse of network structure. It is further suggested that, for samples acidified with citric acid, there is preferential association of chain sequences of high ester content into hydrophobic junctions at 90 °C, leaving sequences with a high content of unesterified carboxyl groups available to form long hydrogen-bonded junctions during cooling, and thus giving gels that are stronger and more resistant to network collapse.     

TGF-beta regulates differentially the proliferation of fetal and adult human skin fibroblasts via the activation of PKA and the autocrine action of FGF-2

Transforming growth factor-beta (TGF-beta) is a potent regulator of cell proliferation; interestingly its action is clearly cell type-dependent. In particular, it inhibits epithelial and endothelial cells' proliferation, while its action on many mesenchymal cells has been reported to be stimulatory. In this direction, we have recently shown that TGF-beta regulates the proliferation of normal human skin fibroblasts according to their developmental origin: i.e. it inhibits fetal fibroblasts, while it stimulates the proliferation of adult ones. Here, we present evidence on the mechanisms underlying this differential action. Concerning fetal fibroblasts, we have found that TGF-beta activates Protein Kinase A (PKA) and induces the expression of the cyclin-dependent kinase inhibitors (CKIs) p21(CIP1/WAF1) and p15(INK4B). Moreover, the specific PKA inhibitor H-89 blocks the induction of both CKIs and annuls the TGF-beta-mediated inhibitory effect, indicating the central role of PKA in this process. In contrast, in adult cells no PKA activation is observed. Moreover, TGF-beta stimulates cell proliferation by activating the MEK-ERK pathway, as the MEK inhibitor PD98059 blocks this effect. A specific neutralizing antibody against Fibroblast Growth Factor-2 (FGF-2) inhibits both ERK activation and the mitogenic activity of TGF-beta, indicating that the latter establishes an autocrine loop, via FGF-2, leading to cell proliferation. This loop requires FGF receptor-1 (FGFR-1), as its down-regulation by siRNA approach prevents TGF-beta from stimulating ERK-1/2 activation and DNA synthesis. In conclusion, the differential proliferative response of fetal and adult normal human skin fibroblasts to TGF-beta is regulated by distinct signaling pathways and furthermore it may provide information on the bimodal effect of this factor on cell proliferation, in general.     

The growth of children in relation to the timing of obesity development

Objective: To examine whether there is an association between the timing of the development of obesity and children's growth. Research Methods and Procedures: This study investigated 141 prepubertal obese children (76 girls) and 72 healthy non‐obese children (39 girls). The target height standard deviation score (SDS), the percentage weight for height, and the height SDS (H‐SDS) at presentation and at the age of 2 years were calculated. Patients were classified, according to whether obesity developed before or after the age of 3 years, as presenting with early‐onset or late‐onset obesity, respectively. Results: Mean age (±SD) at presentation was 9.4 (2.1) years. At the age of 2 years, the H‐SDS of the children with early‐onset obesity was 1.3 (1.0) vs. 0.9 (1.3) for the late‐onset obese (p > 0.5) and 0.4 (1.0) for controls (p < 0.001), and the children with late‐onset obesity were also significantly taller than controls (p < 0.005). At presentation, children with early‐onset obesity were significantly taller than children with late‐onset obesity [1.1 (0.8) vs. 0.6 (1.0); p < 0.001] and controls [0.2 (0.8); p < 0.001]. There was no increase in H‐SDS after the age of 2 years in the late‐onset obese children (p > 0.05). H‐SDS values were below average in 21% of the children with late‐onset obesity and in only 4% of the children with early‐onset obesity. Discussion: These findings indicate that late development of obesity is not associated with increased stature in prepubertal children; however, it may be preceded by growth acceleration in the early years of life. Growth acceleration in early life may be a predictor for future obesity.     

“Melt-in-the-mouth” gels from mixtures of xanthan and konjac glucomannan under acidic conditions: A rheological and calorimetric study of the mechanism of synergistic gelation

The effect of acidification on a typical commercial xanthan and on pyruvate-free xanthan (PFX), alone and in gelling mixtures with konjac glucomannan (KGM), has been studied by differential scanning calorimetry (DSC) and small-deformation oscillatory measurements of storage modulus (G′) and loss modulus (G″). For both xanthan samples, progressive reduction in pH caused a progressive increase in temperature of the disorder–order transition in DSC, and a progressive reduction in gelation temperature with KGM. This inverse correlation is interpreted as showing that synergistic gelation involves disruption of the xanthan 5-fold helix, probably by attachment of KGM to the cellulosic backbone of the xanthan molecule (as proposed previously by a research group in the Institute of Food Research, Norwich, UK). Higher transition temperature accompanied by lower gelation temperature for PFX in comparison with commercial xanthan at neutral pH is explained in the same way. However, an additional postulate from the Norwich group, that attachment of KGM (or galactomannans) can occur only when the xanthan molecule is disordered, is inconsistent with the observation that gelation of acidified mixtures of KGM with PFX can occur at temperatures more than 60 °C below completion of conformational ordering of the PFX component (as characterised by DSC). Increase in G′ on cooling for mixtures of commercial xanthan with KGM at pH values of 4.5 and 4.25 occurred in two discrete steps, the first following the temperature-course observed for the same mixtures at neutral pH and the second occurring over the lower temperatures observed for mixtures of KGM with PFX at the same values of pH. These two “waves” of gel formation are attributed to interaction of KGM with, respectively, xanthan sequences that had retained a high content of pyruvate substituents, and sequences depleted in pyruvate by acid hydrolysis. At pH values of 4.0 and lower, gelation of mixtures of KGM with commercial xanthan followed essentially the same temperature-course as for mixtures with PFX, indicating extensive loss of pyruvate under these more strongly acidic conditions. Mixtures prepared at pH values in the range 4.0–3.5 gave comparable moduli at room temperature (20 °C) to those obtained at neutral pH, but showed substantial softening on heating to body temperature, suggesting possible applications in replacement of gelatin in products where “melt-in-the-mouth” characteristics are important for acceptability to the consumer.     

Effect of polymeric cosolutes on calcium pectinate gelation. Part 2. Dextrans and inulin

In a continuation of the work reported in the preceding paper, the effect of chicory root inulin (M_r≈4.5 kD) and branched dextrans of M_r 67, 464 and 2000 kD on gelation of low methoxy pectin (DE 31; 2.0 wt%; pH≈2.9–3.0) on cooling from 90 to 5 °C in the presence of stoichiometric Ca²⁺ has been characterised by low amplitude oscillatory measurements of G′ and G″. As found for other polymeric cosolutes studied previously (oxidised starch, potato maltodextrin, guar gum and locust bean gum), increasing concentrations of dextran or inulin caused a progressive increase of G′ and G″ in the solution state at 90 °C (attributed to segregative interactions promoting formation of calcium-mediated ‘egg-box’ junctions between pectin chains) and a progressive reduction in final moduli at 5 °C (attributed to excessive association of calcium pectinate into large aggregated bundles), with cooling curves crossing those for calcium pectinate alone at 55 °C. For the dextran samples, the ability to promote initial association of pectin chains at high temperature decreased with increasing molecular weight (i.e. with the smallest molecules having the greatest effect, as was also found for guar gum), but the reduction in final modulus on completion of cooling to 5 °C became more pronounced. By the criterion of the decrease in the change in log G′ during cooling, the three dextran samples studied seem roughly comparable in their overall effectiveness in promoting self-association of pectin, with inulin having a substantially smaller effect. Both materials, however, are less effective than the partially depolymerised starches and galactomannans studied previously. It is tentatively suggested that this may be associated with the flexibility introduced by the presence of three single bonds in the predominant glycosidic linkages of both dextran and inulin.     

Visualizing S1P-directed cellular egress by intravital imaging

Sphingosine-1-phosphate (S1P) is a bioactive lipid that provides cellular signals through plasma membrane G protein-coupled receptors. The S1P receptor signaling system has a fundamental and widespread function in licensing the exit and release of hematopoietically derived cells from various tissues into the circulation. Although the outlines of the mechanism have been established through genetic and pharmacologic perturbations, the temporal and spatial dynamics of the cellular events involved have been unclear. Recently, two-photon intravital imaging has been applied to living tissues to visualize the cellular movements and interactions that occur during egress processes. Here we discuss how some of these recent findings provide a clearer picture regarding S1P receptor signaling in modulating cell egress into the circulation. This article is part of a Special Issue entitled New Frontiers in Sphingolipid Biology.     

PGE2 inhibits apoptosis in human adenocarcinoma Caco-2 cell line through Ras-PI3K association and cAMP-dependent kinase A activation

PGE2 plays a critical role in colorectal carcinogenesis. We have previously shown that COX-2 expression and PGE2 synthesis are mediated by IGF-II/IGF-I receptor signaling in the Caco-2 cell line and that the pathway of phosphatidylinositol 3-kinase (PI3K)/Akt protects the cell from apoptosis. In the present study, we demonstrate that PGE2 has the ability to increase Ras and PI3K association and decrease the level of apoptosis in the same experimental system. The effect of PGE2 on PI3K/Ras association is dependent on the activation of EP4 receptor, the increase of cAMP levels, and the activation of PKA. In fact, treatment of cells with the PKA inhibitor H89 decreases the association of Ras and PI3K and Ras-associated PI3K activity. PKA inhibitor H89 is able to decrease threonine phosphorylation of Akt and to increase serine phosphorylation of Akt by p38 MAPK and counteracts the cytoprotective effect induced by PGE2. In addition, PGE2 is able to activate p38 MAPK and the inhibition of p38 MAPK, with SB203580 specific inhibitor or with dominant negative MKK6 kinase, is able to revert the apoptotic effect of H89 and serine phosphorylation of Akt. The effect of PGE2 on Caco-2 cell survival through PKA activation is mediated and regulated by the balance of threonine/serine phosphorylation of Akt by p38 kinase and PI3K. In conclusion, our data elucidate a novel mechanism for regulation of colon cancer cell survival and provide evidences for new combinatory treatments of colon cancer.     

Genome sequences of three Acinetobacter baumannii strains assigned to the multilocus sequence typing genotypes ST2, ST25, and ST78

Acinetobacter baumannii is an emerging opportunistic gram-negative pathogen responsible for hospital-acquired infections. A. baumannii epidemics described in Europe and worldwide were caused by a limited number of genotypic clusters of multidrug-resistant strains. Here, we report the availability of draft genome sequences for three multidrug-resistant A. baumannii strains assigned to multilocus sequence typing genotypes ST2, ST25, and ST78 that were more frequently isolated during outbreaks occurred in Greece, Italy, Lebanon, and Turkey.