BtuM is a bacterial cobalamin transporter that binds the transported substrate in the base-off state, with a cysteine residue providing the α-axial coordination of the central cobalt ion via a ...sulfur-cobalt bond. Binding leads to decyanation of cobalamin variants with a cyano group as the β-axial ligand. Here, we report the crystal structures of untagged BtuM bound to two variants of cobalamin, hydroxycobalamin and cyanocobalamin, and unveil the native residue responsible for the β-axial coordination, His28. This coordination had previously been obscured by non-native histidines of His-tagged BtuM. A model in which BtuM initially binds cobinamide reversibly with low affinity (K
= 4.0 μM), followed by the formation of a covalent bond (rate constant of 0.163 s
), fits the kinetics data of substrate binding and decyanation of the cobalamin precursor cobinamide by BtuM. The covalent binding mode suggests a mechanism not used by any other transport protein.
We perform molecular dynamics (MD) simulations of nanoscale water capillary bridges (WCBs) expanding between two parallel walls and determine the smallest separation between the walls above which the ...capillarity theory (CT) remains valid. We consider silica-based walls with tuned surface partial charges that expand from hydrophobic to hydrophilic. We find that the CT is valid (i.e., it predicts successfully the WCB geometry and forces induced on the walls) for, approximately, wall separations h ≥ h 0 = 3.0 nm for all surfaces considered. At these separations, the CT holds without including any line tension and the results are robust relative to the method employed to obtain the WCB profile from MD simulations. At approximately 2.0 nm ≤ h < h 0, the contact angle of water θ varies with h, suggesting that at such wall separations the CT requires the inclusion of a line tension τ. However, we find that the specific behavior of θ(h) and the associated value of τ are inherently dependent on the method employed to calculate the WCB profile from MD simulations. Interestingly, the forces induced by the WCBs on the walls obey the prediction of the CT without the need to include a line tension for approximately h ≥ 2.5 nm for all surfaces considered. Our results are interpreted in terms of the rearrangement of water molecules within the WCBs and show that the CT breaks down at h < h 0 because its assumptions, that the WCB is a bulklike water volume confined by solid–liquid and liquid–vapor interfaces, do not hold.
Snoring is extremely common in the general population and may indicate OSA. However, snoring is not objectively measured during polysomnography, and no standard treatment is available for primary ...snoring or when snoring is associated with mild forms of OSA. This study determined the effects of oropharyngeal exercises on snoring in minimally symptomatic patients with a primary complaint of snoring and diagnosis of primary snoring or mild to moderate OSA.
Patients were randomized for 3 months of treatment with nasal dilator strips plus respiratory exercises (control) or daily oropharyngeal exercises (therapy). Patients were evaluated at study entry and end by sleep questionnaires (Epworth Sleepiness Scale, Pittsburgh Sleep Quality Index) and full polysomnography with objective measurements of snoring.
We studied 39 patients (age, 46 ± 13 years; BMI, 28.2 ± 3.1 kg/m2; apnea-hypopnea index (AHI), 15.3 ± 9.3 events/h; Epworth Sleepiness Scale, 9.2 ± 4.9; Pittsburgh Sleep Quality Index, 6.4 ± 3.3). Control (n = 20) and therapy (n = 19) groups were similar at study entry. One patient from each group dropped out. Intention-to-treat analysis was used. No significant changes occurred in the control group. In contrast, patients randomized to therapy experienced a significant decrease in the snore index (snores > 36 dB/h), 99.5 (49.6-221.3) vs 48.2 (25.5-219.2); P = .017 and total snore index (total power of snore/h), 60.4 (21.8-220.6) vs 31.0 (10.1-146.5); P = .033.
Oropharyngeal exercises are effective in reducing objectively measured snoring and are a possible treatment of a large population suffering from snoring.
ClinicalTrials.gov; No.: NCT01636856; URL: www.clinicaltrials.gov.
Neurally Adjusted Ventilatory Assist (NAVA) is a proportional ventilatory mode that uses the electrical activity of the diaphragm (EAdi) to offer ventilatory assistance in proportion to patient ...effort. NAVA has been increasingly used for critically ill patients, but it has not been evaluated during spontaneous breathing trials (SBT). We designed a pilot trial to assess the feasibility of using NAVA during SBTs, and to compare the breathing pattern and patient-ventilator asynchrony of NAVA with Pressure Support (PSV) during SBTs.
We conducted a crossover trial in the ICU of a university hospital in Brazil and included mechanically ventilated patients considered ready to undergo an SBT on the day of the study. Patients underwent two SBTs in randomized order: 30 min in PSV of 5 cmH
O or NAVA titrated to generate equivalent peak airway pressure (Paw), with a positive end-expiratory pressure of 5 cmH
O. The ICU team, blinded to ventilatory mode, evaluated whether patients passed each SBT. We captured flow, Paw and electrical activity of the diaphragm (EAdi) from the ventilator and used it to calculate respiratory rate (RR), tidal volume (VT), and EAdi. Detection of asynchrony events used waveform analysis and we calculated the asynchrony index as the number of asynchrony events divided by the number of neural cycles.
We included 20 patients in the study. All patients passed the SBT in PSV, and three failed the SBT in NAVA. Five patients were reintubated and the extubation failure rate was 25% (95% CI 9-49%). Respiratory parameters were similar in the two modes: VT = 6.1 (5.5-6.5) mL/Kg in NAVA vs. 5.5 (4.8-6.1) mL/Kg in PSV (p = 0.076) and RR = 27 (17-30) rpm in NAVA vs. 26 (20-30) rpm in PSV, p = 0.55. NAVA reduced AI, with a median of 11.5% (4.2-19.7) compared to 24.3% (6.3-34.3) in PSV (p = 0.033).
NAVA reduces patient-ventilator asynchrony index and generates a respiratory pattern similar to PSV during SBTs. Patients considered ready for mechanical ventilation liberation may be submitted to an SBT in NAVA using the same objective criteria used for SBTs in PSV.
ClinicalTrials.gov ( NCT01337271 ), registered April 12, 2011.
We perform molecular dynamics (MD) simulations of water capillary bridges formed between parallel walls. The underlying structure of the walls corresponds to hydroxilated (crystalline) ...β-cristobalite, modified to cover a wide range of hydrophobicity/hydrophilicity. The capillary bridges are stretched during the MD simulations, from wall–wall separation h = 5 nm up to h ≈ 7.5 nm, until they become unstable and break. During the stretching process, we calculate the profiles of capillary bridges as well as the force and pressure induced on the walls, among other properties. We find that, for all walls separations and surface hydrophobicity/hydrophilicity considered, the results from MD simulations are in excellent agreement with the predictions from capillarity theory (CT). In addition, we find that CT is able to predict very closely the limit of stability of the capillary bridges, i.e., the value of h at which the bridges break. We also confirm that CT predicts correctly the relationship between the surface hydrophobicity/hydrophilicity and the resulting droplets of the capillary bridge rupture. Depending on the contact angle of water with the corresponding surface, the rupture of the capillary bridges results in (i) a single droplet attached to one of the walls, (ii) two identical, or (iii) two different droplets, one attached to each wall. This work expands upon a previous study of nanoscale droplets and (stable) capillary bridges where CT was validated at the nanoscale using MD simulations. The validation of CT at such small scales is remarkable, since CT is a macroscopic theory that is expected to fail at <10 nm scales, where molecular details may become relevant. In particular, we find that CT works for capillary bridges that are ≈2-nm thick, comparable to the thickness of the water–vapor interface.
The mean linear intercept (L(m)) can be used to estimate the surface area for gas exchange in the lung. However, in recent years, it is most commonly used as an index for characterizing the ...enlargement of airspaces in emphysema and the associated severity of structural destruction in the lung. Specifically, an increase in L(m) is thought to result from an increase in airspace sizes. In this paper, we examined how accurately L(m) measures the linear dimensions of airspaces from histological sections and a variety of computer-generated test images. To this end, we developed an automated method for measuring linear intercepts from digitized images of tissue sections and calculate L(m) as their mean. We examined how the shape of airspaces and the variability of their sizes influence L(m) as well as the distribution of linear intercepts. We found that, for a relatively homogeneous enlargement of airspaces, L(m) was a reliable index for detecting emphysema. However, in the presence of spatial heterogeneities with a large variability of airspace sizes, L(m) did not significantly increase and sometimes even decreased compared with its value in normal tissue. We also developed an automated method for measuring the area and computed an equivalent diameter of each individual airspace that is independent of shape. Finally, we introduced new indexes based on the moments of diameter that we found to be more reliable than L(m) to characterize airspace enlargement in the presence of heterogeneities.
Mapping the cytoskeletal prestress Park, Chan Young; Tambe, Dhananjay; Alencar, Adriano M ...
American Journal of Physiology: Cell Physiology,
05/2010, Letnik:
298, Številka:
5
Journal Article
Recenzirano
Odprti dostop
Cell mechanical properties on a whole cell basis have been widely studied, whereas local intracellular variations have been less well characterized and are poorly understood. To fill this gap, here ...we provide detailed intracellular maps of regional cytoskeleton (CSK) stiffness, loss tangent, and rate of structural rearrangements, as well as their relationships to the underlying regional F-actin density and the local cytoskeletal prestress. In the human airway smooth muscle cell, we used micropatterning to minimize geometric variation. We measured the local cell stiffness and loss tangent with optical magnetic twisting cytometry and the local rate of CSK remodeling with spontaneous displacements of a CSK-bound bead. We also measured traction distributions with traction microscopy and cell geometry with atomic force microscopy. On the basis of these experimental observations, we used finite element methods to map for the first time the regional distribution of intracellular prestress. Compared with the cell center or edges, cell corners were systematically stiffer and more fluidlike and supported higher traction forces, and at the same time had slower remodeling dynamics. Local remodeling dynamics had a close inverse relationship with local cell stiffness. The principal finding, however, is that systematic regional variations of CSK stiffness correlated only poorly with regional F-actin density but strongly and linearly with the regional prestress. Taken together, these findings in the intact cell comprise the most comprehensive characterization to date of regional variations of cytoskeletal mechanical properties and their determinants.
Although redox processes closely interplay with mechanoresponses to control vascular remodeling, redox pathways coupling mechanostimulation to cellular cytoskeletal organization remain unclear. The ...peri/epicellular pool of protein disulfide isomerase-A1 (pecPDIA1) supports postinjury vessel remodeling. Using distinct models, we investigated whether pecPDIA1 could work as a redox-dependent organizer of cytoskeletal mechanoresponses. In vascular smooth muscle cells (VSMCs), pecPDIA1 immunoneutralization impaired stress fiber assembly in response to equibiaxial stretch and, under uniaxial stretch, significantly perturbed cell repositioning perpendicularly to stretch orientation. During cyclic stretch, pecPDIA1 supported thiol oxidation of the known mechanosensor β
-integrin and promoted polarized compartmentalization of sulfenylated proteins. Using traction force microscopy, we showed that pecPDIA1 organizes intracellular force distribution. The net contractile moment ratio of platelet-derived growth factor-exposed to basal VSMCs decreased from 0.90 ± 0.09 (IgG-exposed controls) to 0.70 ± 0.08 after pecPDI neutralization ( P < 0.05), together with an enhanced coefficient of variation for distribution of force modules, suggesting increased noise. Moreover, in a single cell model, pecPDIA1 neutralization impaired migration persistence without affecting total distance or velocity, whereas siRNA-mediated total PDIA1 silencing disabled all such variables of VSMC migration. Neither expression nor total activity of the master mechanotransmitter/regulator RhoA was affected by pecPDIA1 neutralization. However, cyclic stretch-induced focal distribution of membrane-bound RhoA was disrupted by pecPDI inhibition, which promoted a nonpolarized pattern of RhoA/caveolin-3 cluster colocalization. Accordingly, FRET biosensors showed that pecPDIA1 supports localized RhoA activity at cell protrusions versus perinuclear regions. Thus, pecPDI acts as a thiol redox-dependent organizer and noise reducer mechanism of cytoskeletal repositioning, oxidant generation, and localized RhoA activation during a variety of VSMC mechanoresponses. NEW & NOTEWORTHY Effects of a peri/epicellular pool of protein disulfide isomerase-A1 (pecPDIA1) during mechanoregulation in vascular smooth muscle cells (VSMCs) were highlighted using approaches such as equibiaxial and uniaxial stretch, random single cell migration, and traction force microscopy. pecPDIA1 regulates organization of the cytoskeleton and minimizes the noise of cell alignment, migration directionality, and persistence. pecPDIA1 mechanisms involve redox control of β
-integrin and localized RhoA activation. pecPDIA1 acts as a novel organizer of mechanoadaptation responses in VSMCs.
Background
Protective mechanical ventilation is recommended for patients with acute respiratory distress syndrome (ARDS), but it usually requires controlled ventilation and sedation. Using neurally ...adjusted ventilatory assist (NAVA) or pressure support ventilation (PSV) could have additional benefits, including the use of lower sedative doses, improved patient–ventilator interaction and shortened duration of mechanical ventilation. We designed a pilot study to assess the feasibility of keeping tidal volume (
V
T
) at protective levels with NAVA and PSV in patients with ARDS.
Methods
We conducted a prospective randomized crossover trial in five ICUs from a university hospital in Brazil and included patients with ARDS transitioning from controlled ventilation to partial ventilatory support. NAVA and PSV were applied in random order, for 15 min each, followed by 3 h in NAVA. Flow, peak airway pressure (Paw) and electrical activity of the diaphragm (EAdi) were captured from the ventilator, and a software (Matlab, Mathworks, USA), automatically detected inspiratory efforts and calculated respiratory rate (RR) and
V
T
. Asynchrony events detection was based on waveform analysis.
Results
We randomized 20 patients, but the protocol was interrupted for five (25%) patients for whom we were unable to maintain
V
T
below 6.5 mL/kg in PSV due to strong inspiratory efforts and for one patient for whom we could not detect EAdi signal. For the 14 patients who completed the protocol,
V
T
was 5.8 ± 1.1 mL/kg for NAVA and 5.6 ± 1.0 mL/kg for PSV (
p
= 0.455) and there were no differences in RR (24 ± 7 for NAVA and 23 ± 7 for PSV,
p
= 0.661). Paw was greater in NAVA (21 ± 3 cmH
2
O) than in PSV (19 ± 3 cmH
2
O,
p
= 0.001). Most patients were under continuous sedation during the study. NAVA reduced triggering delay compared to PSV (
p
= 0.020) and the median asynchrony Index was 0.7% (0–2.7) in PSV and 0% (0–2.2) in NAVA (
p
= 0.6835).
Conclusions
It was feasible to keep
V
T
in protective levels with NAVA and PSV for 75% of the patients. NAVA resulted in similar
V
T
, RR and Paw compared to PSV. Our findings suggest that partial ventilatory assistance with NAVA and PSV is feasible as a protective ventilation strategy in selected ARDS patients under continuous sedation.
Trial registration
ClinicalTrials.gov (NCT01519258). Registered 26 January 2012,
https://clinicaltrials.gov/ct2/show/NCT01519258
Random-walk model of cotransport Barreto, Yan B.; Suki, Béla; Alencar, Adriano M.
Physical review. E,
08/2020, Letnik:
102, Številka:
2
Journal Article
PDF
