The importance of the Thr17 residue of phospholamban as a phosphorylation site under physiological and pathological conditions

The sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA2a) is under the control of an SR protein named phospholamban (PLN). Dephosphorylated PLN inhibits SERCA2a, whereas phosphorylation of PLN at either the Ser16 site by PKA or the Thr17 site by CaMKII reverses this inhibition, thus increasing SERCA2a activity and the rate of Ca2+ uptake by the SR. This leads to an increase in the velocity of relaxation, SR Ca2+ load and myocardial contractility. In the intact heart, ß-adrenoceptor stimulation results in phosphorylation of PLN at both Ser16 and Thr17 residues. Phosphorylation of the Thr17 residue requires both stimulation of the CaMKII signaling pathways and inhibition of PP1, the major phosphatase that dephosphorylates PLN. These two prerequisites appear to be fulfilled by ß-adrenoceptor stimulation, which as a result of PKA activation, triggers the activation of CaMKII by increasing intracellular Ca2+, and inhibits PP1. Several pathological situations such as ischemia-reperfusion injury or hypercapnic acidosis provide the required conditions for the phosphorylation of the Thr17 residue of PLN, independently of the increase in PKA activity, i.e., increased intracellular Ca2+ and acidosis-induced phosphatase inhibition. Our results indicated that PLN was phosphorylated at Thr17 at the onset of reflow and immediately after hypercapnia was established, and that this phosphorylation contributes to the mechanical recovery after both the ischemic and acidic insults. Studies on transgenic mice with Thr17 mutated to Ala (PLN-T17A) are consistent with these results. Thus, phosphorylation of the Thr17 residue of PLN probably participates in a protective mechanism that favors Ca2+ handling and limits intracellular Ca2+ overload in pathological situations.

Understanding the mechanisms of lung mechanical stress

Physical forces affect both the function and phenotype of cells in the lung. Bronchial, alveolar, and other parenchymal cells, as well as fibroblasts and macrophages, are normally subjected to a variety of passive and active mechanical forces associated with lung inflation and vascular perfusion as a result of the dynamic nature of lung function. These forces include changes in stress (force per unit area) or strain (any forced change in length in relation to the initial length) and shear stress (the stress component parallel to a given surface). The responses of cells to mechanical forces are the result of the cell's ability to sense and transduce these stimuli into intracellular signaling pathways able to communicate the information to its interior. This review will focus on the modulation of intracellular pathways by lung mechanical forces and the intercellular signaling. A better understanding of the mechanisms by which lung cells transduce physical forces into biochemical and biological signals is of key importance for identifying targets for the treatment and prevention of physical force-related disorders.

13CO2 recovery fraction in expired air of septic patients under mechanical ventilation

The continuous intravenous administration of isotopic bicarbonate (NaH13CO2) has been used for the determination of the retention of the 13CO2 fraction or the 13CO2 recovered in expired air. This determination is important for the calculation of substrate oxidation. The aim of the present study was to evaluate, in critically ill patients with sepsis under mechanical ventilation, the 13CO2 recovery fraction in expired air after continuous intravenous infusion of NaH13CO2 (3.8 µmol/kg diluted in 0.9% saline in ddH2O). A prospective study was conducted on 10 patients with septic shock between the second and fifth day of sepsis evolution (APACHE II, 25.9 ± 7.4). Initially, baseline CO2 was collected and indirect calorimetry was also performed. A primer of 5 mL NaH13CO2 was administered followed by continuous infusion of 5 mL/h for 6 h. Six CO2 production (VCO2) measurements (30 min each) were made with a portable metabolic cart connected to a respirator and hourly samples of expired air were obtained using a 750-mL gas collecting bag attached to the outlet of the respirator. 13CO2 enrichment in expired air was determined with a mass spectrometer. The patients presented a mean value of VCO2 of 182 ± 52 mL/min during the steady-state phase. The mean recovery fraction was 0.68 ± 0.06%, which is less than that reported in the literature (0.82 ± 0.03%). This suggests that the 13CO2 recovery fraction in septic patients following enteral feeding is incomplete, indicating retention of 13CO2 in the organism. The severity of septic shock in terms of the prognostic index APACHE II and the sepsis score was not associated with the 13CO2 recovery fraction in expired air.

Streptozotocin-induced mechanical hypernociception is not dependent on hyperglycemia

Since streptozotocin (STZ)-induced diabetes is a widely used model of painful diabetic neuropathy, the aim of the present study was to design a rational protocol to investigate whether the development of mechanical hypernociception induced by STZ depends exclusively on hyperglycemia. Male Wistar rats (180-200 g; N = 6-7 per group) received a single intravenous injection of STZ at three different doses (10, 20, or 40 mg/kg). Only the higher dose (40 mg/kg) induced a significant increase in blood glucose levels, glucose tolerance and deficiency in weight gain. However, all STZ-treated rats (hyperglycemic or not) developed persistent (for at least 20 days) and indistinguishable bilateral mechanical hypernociception that was not prevented by daily insulin treatment (2 IU twice a day, sc). Systemic morphine (2 mg/kg) but not local (intraplantar) morphine treatment (8 µg/paw) significantly inhibited the mechanical hypernociception induced by STZ (10 or 40 mg/kg). In addition, intraplantar injection of STZ at doses that did not cause hyperglycemia (30, 100 or 300 µg/paw) induced ipsilateral mechanical hypernociception for at least 8 h that was inhibited by local and systemic morphine treatment (8 µg/paw or 2 mg/kg, respectively), but not by dexamethasone (1 mg/kg, sc). The results of this study demonstrate that systemic administration of STZ induces mechanical hypernociception that does not depend on hyperglycemia and intraplantar STZ induces mechanical sensitization of primary sensory neurons responsive to local morphine treatment.

Mechanical cardiac remodeling and new-onset atrial fibrillation in long-term follow-up of subjects with chronic Chagas' disease

Atrial fibrillation (AF) affects subjects with Chagas' disease and is an indicator of poor prognosis. We investigated clinical, echocardiographic and electrocardiographic variables of Chagas' disease in a long-term longitudinal study as predictors of a new-onset AF episode lasting >24 h, nonfatal embolic stroke and cardiac death. Fifty adult outpatients (34 to 74 years old, 62% females) staged according to the Los Andes classification were enrolled. During a follow-up of (mean ± SD) 84.2 ± 39.0 months, 9 subjects developed AF (incidence: 3.3 ± 1.0%/year), 5 had nonfatal stroke (incidence: 1.3 ± 1.0%/year), and nine died (mortality rate: 2.3 ± 0.8%/year). The progression rate of left ventricular mass and left ventricular ejection fraction was significantly greater in subjects who experienced AF (16.4 ± 20.0 g/year and -8.6 ± 7.6%/year, respectively) than in those who did not (8.2 ± 8.4 g/year; P = 0.03, and -3.0 ± 2.5%/year; P = 0.04, respectively). In univariate analysis, left atrial diameter ≥3.2 cm (P = 0.002), pulmonary arterial hypertension (P = 0.035), frequent premature supraventricular and ventricular contraction counts/24 h (P = 0.005 and P = 0.007, respectively), ventricular couplets/24 h (P = 0.002), and ventricular tachycardia (P = 0.004) were long-term predictors of AF. P-wave signal-averaged ECG revealed a limited long-term predictive value for AF. In chronic Chagas' disease, large left atrial diameter, pulmonary arterial hypertension, frequent supraventricular and ventricular premature beats, and ventricular tachycardia are long-term predictors of AF. The rate of left ventricular mass enlargement and systolic function deterioration impact AF incidence in this population.

Exercise training and detraining modify the morphological and mechanical properties of single cardiac myocytes obtained from spontaneously hypertensive rats

We determined the effects of exercise training and detraining on the morphological and mechanical properties of left ventricular myocytes in 4-month-old spontaneously hypertensive rats (SHR) randomly divided into the following groups: sedentary for 8 weeks (SED-8), sedentary for 12 weeks (SED-12), treadmill-running trained for 8 weeks (TRA, 16 m/min, 60 min/day, 5 days/week), and treadmill-running trained for 8 weeks followed by 4 weeks of detraining (DET). At sacrifice, left ventricular myocytes were isolated enzymatically, and resting cell length, width, and cell shortening after stimulation at a frequency of 1 Hz (~25°C) were measured. Cell length was greater in TRA than in SED-8 (161.30 ± 1.01 vs 156.10 ± 1.02 μm, P < 0.05, 667 vs 618 cells, respectively) and remained larger after detraining. Cell width and volume were unaffected by either exercise training or detraining. Cell length to width ratio was higher in TRA than in SED-8 (8.50 ± 0.08 vs 8.22 ± 0.10, P < 0.05) and was maintained after detraining. Exercise training did not affect cell shortening, which was unchanged with detraining. TRA cells exhibited higher maximum velocity of shortening than SED-8 (102.01 ± 4.50 vs 82.01 ± 5.30 μm/s, P < 0.05, 70 cells per group), with almost complete regression after detraining. The maximum velocity of relengthening was higher in TRA cells than in SED-8 (88.20 ± 4.01 vs70.01 ± 4.80 μm/s, P < 0.05), returning to sedentary values with detraining. Therefore, exercise training affected left ventricle remodeling in SHR towards eccentric hypertrophy, which remained after detraining. It also improved single left ventricular myocyte contractile function, which was reversed by detraining.

Periodic mechanical stress activates MEK1/2-ERK1/2 mitogenic signals in rat chondrocytes through Src and PLCγ1

The mitogenic effects of periodic mechanical stress on chondrocytes have been studied extensively but the mechanisms whereby chondrocytes sense and respond to periodic mechanical stress remain a matter of debate. We explored the signal transduction pathways of chondrocyte proliferation and matrix synthesis under periodic mechanical stress. In particular, we sought to identify the role of the MEK1/2-ERK1/2 signaling pathway in chondrocyte proliferation and matrix synthesis following cyclic physiologic mechanical compression. Under periodic mechanical stress, both rat chondrocyte proliferation and matrix synthesis were significantly increased (P < 0.05) and were associated with increases in the phosphorylation of Src, PLCγ1, MEK1/2, and ERK1/2 (P < 0.05). Pretreatment with the MEK1/2-ERK1/2 selective inhibitor, PD98059, and shRNA targeted to ERK1/2 reduced periodic mechanical stress-induced chondrocyte proliferation and matrix synthesis (P < 0.05), while the phosphorylation levels of Src-Tyr418 and PLCγ1-Tyr783 were not inhibited. Proliferation, matrix synthesis and phosphorylation of MEK1/2-Ser217/221 and ERK1/2-Thr202/Tyr204 were inhibited after pretreatment with the PLCγ1 inhibitor U73122 in chondrocytes in response to periodic mechanical stress (P < 0.05), while the phosphorylation site of Src-Tyr418 was not affected. Inhibition of Src activity with PP2 and shRNA targeted to Src abrogated chondrocyte proliferation and matrix synthesis (P < 0.05) and attenuated PLCγ1, MEK1/2 and ERK1/2 activation in chondrocytes subjected to periodic mechanical stress (P < 0.05). These findings suggest that periodic mechanical stress promotes chondrocyte proliferation and matrix synthesis in part through the Src-PLCγ1-MEK1/2-ERK1/2 signaling pathway, which links these three important signaling molecules into a mitogenic cascade.

Implications of extubation failure and prolonged mechanical ventilation in the postoperative period following elective intracranial surgery

Patients undergoing neurosurgery are predisposed to a variety of complications related to mechanical ventilation (MV). There is an increased incidence of extubation failure, pneumonia, and prolonged MV among such patients. The aim of the present study was to assess the influence of extubation failure and prolonged MV on the following variables: postoperative pulmonary complications (PPC), mortality, reoperation, tracheostomy, and duration of postoperative hospitalization following elective intra-cranial surgery. The study involved a prospective observational cohort of 317 patients submitted to elective intracranial surgery for tumors, aneurysms and arteriovenous malformation. Preoperative assessment was performed and patients were followed up for the determination of extubation failure and prolonged MV (>48 h) until discharge from the hospital or death. The occurrence of PPC, incidence of death, the need for reoperation and tracheostomy, and the length of hospitalization were assessed during the postoperative period. Twenty-six patients (8.2%) experienced extubation failure and 30 (9.5%) needed prolonged MV after surgery. Multivariate analysis showed that extubation failure was significant for the occurrence of death (OR = 8.05 [1.88; 34.36]), PPC (OR = 11.18 [2.27; 55.02]) and tracheostomy (OR = 7.8 [1.12; 55.07]). Prolonged MV was significant only for the occurrence of PPC (OR = 4.87 [1.3; 18.18]). Elective intracranial surgery patients who experienced extubation failure or required prolonged MV had a higher incidence of PPC, reoperation and tracheostomy and required a longer period of time in the ICU. Level of consciousness and extubation failure were associated with death and PPC. Patients who required prolonged MV had a higher incidence of extubation failure.

Brazilian roasted coffee oil obtained by mechanical expelling: compositional analysis by GC-MS

A preliminary analysis by GC-MS comparing the mass spectrum of the compounds with the Wiley 275 L mass spectral data base was used to identify the fatty acids and mainly, some volatile compounds responsible for the flavor of the roasted coffee oil. The oil was obtained by mechanical expelling of Brazilian beans (Coffea arabica) roasted at 238ºC for 10 minutes. Different sample preparation methodologies such as headspace, adsorbent suction trapping and esterification were used. It was possible to identify pyrazines, pyridines, furan derivatives and other compounds not reported in the literature.

Determination of structural and mechanical properties, diffractometry, and thermal analysis of chitosan and hydroxypropylmethylcellulose (HPMC) films plasticized with sorbitol

In this work, the structural, mechanical, diffractometric, and thermal parameters of chitosan-hydroxypropylmethylcellulose (HPMC) films plasticized with sorbitol were studied. Solutions of HPMC (2% w/v) in water and chitosan (2% w/v) in 2% acetic acid solution were prepared. The concentration of sorbitol used was 10% (w/w) to both polymers. This solutions were mixed at different proportions (100/0; 70/30; 50/50; 30/70, and 0/100) of chitosan and HPMC, respectively, and 20 mL was cast in Petri dishes for further analysis of dried films. The miscibility of polymers was assessed by X-ray diffraction, scanning electronic microscopy (SEM), differential scanning calorimetry (DSC), and thermal gravimetric analysis (TGA). The results obtained indicate that the films are not fully miscible at a dry state despite the weak hydrogen bonding between the polymer functional groups.

Mechanical and water vapor permeability properties of biodegradables films based on methylcellulose, glucomannan, pectin and gelatin

Mimic biological structures such as the cell wall of plant tissues may be an alternative to obtain biodegradable films with improved mechanical and water vapor barrier properties. This study aims to evaluate the mechanical properties and water vapor permeability (WVP) of films produced by using the solvent-casting technique from blended methylcellulose, glucomannan, pectin and gelatin. First, films from polysaccharides at pH 4 were produced. The film with the best mechanical performance (tensile strength = 72.63 MPa; elongation = 9.85%) was obtained from methylcellulose-glucomannan-pectin at ratio 1:4:1, respectively. Then, gelatin was added to this polysaccharide blend and the pH was adjusted to 4, 5 and 6. Results showed significant improvement in WVP when films were made at pH 5 and at polysaccharides/gelatin ratio of 90/10 and 10/90, reaching 0.094 and 0.118 g.mm/h.m².kPa as values, respectively. Films with the best mechanical properties were obtained from the blend of polysaccharides, whereas WVP was improved from the blend of polysaccharides and gelatin at pH 5.

Effects of the mechanical damage on the water absorption process by corn kernel

The purpose of this study was to investigate and model the water absorption process by corn kernels with different levels of mechanical damage Corn kernels of AG 1510 variety with moisture content of 14.2 (% d.b.) were used. Different mechanical damage levels were indirectly evaluated by electrical conductivity measurements. The absorption process was based on the industrial corn wet milling process, in which the product was soaked with a 0.2% sulfur dioxide (SO2) solution and 0.55% lactic acid (C3H6O3) in distilled water, under controlled temperatures of 40, 50, 60, and 70 ºC and different mechanical damage levels. The Peleg model was used for the analysis and modeling of water absorption process. The conclusion is that the structural changes caused by the mechanical damage to the corn kernels influenced the initial rates of water absorption, which were higher for the most damaged kernels, and they also changed the equilibrium moisture contents of the kernels. The Peleg model was well adjusted to the experimental data presenting satisfactory values for the analyzed statistic parameters for all temperatures regardless of the damage level of the corn kernels.

Effect of incorporation of nutraceutical capsule waste of safflower oil in the mechanical characteristics of corn starch films

Abstract Biodegradable films blends made of safflower oil nutraceutical capsules waste corn starch (20:4, 30:4, 40:4 and 50:4) were prepared. The objective of this study was to evaluate the influence of addition of different concentrations of safflower oil nutraceutical capsule waste in the mechanical properties (tensile strength, elongation at break, Young’s modulus) and thickness of corn starch films. A decrease in tensile strength and Young’s modulus and an increase in elongation at break were observed with the increase in the content of the nutraceutical capsule waste. The results showed that the blends of safflower oil capsules waste-corn starch films demonstrated promising characteristics to form biodegradable films with different mechanical characteristics.

X-Ray analysis to assess mechanical damage in sweet corn seeds

The X-ray test is a precise, fast and non-destructive method to detect mechanical damage in seeds. In the present study, the efficiency of X-ray analysis in identifying the extent of mechanical damage in sweet corn seeds and its relationship with germination and vigor was evaluated. Hybrid 'SWB 551' (sh2) seeds with round (R) and flat (F) shapes were classified as large (L), medium (M1, M2 and M3) and small (S), using sieves with round and oblong screens. After artificial exposure to different levels of damage (0, 1, 3, 5 and 7 impacts), seeds were X-rayed (15 kV, 5 min) and submitted to germination (25 °C/5 days) and cold (10 °C/7 days) tests. Digital images of normal and abnormal seedlings and ungerminated seeds from germination and cold tests were jointly analyzed with the seed X-ray images. Results showed that damage affecting the embryonic axis resulted in abnormal seedlings or dead seeds in the germination and cold tests. The X-ray analysis is efficient for identifying mechanical damage in sweet corn seeds, allowing damage severity to be associated with losses in germination and vigor.