Metabolic behavior of immobilized aggregates of Escherichia coli under conditions of varying mechanical stress

Experiments were conducted on immobilized aggregates of Escherichia coli cells. Mechanical stress was applied by forcing a convective stream of nutrient medium through the aggregate. It was shown to be possible to maintain uniform exponential growth with this convective supply of nutrients. Analysis of effluent from the system allowed investigation of metabolic responses unambiguously attributable to mechanical stress. A reversible increase in catabolic activity was observed after an increase in mechanical stress. Changes in the level of catabolism were accompanied by an alteration in the total acid yield on glucose and in the spectrum of organic acids produced during glucose fermentation. The behavior observed here was likely due to an osmoregulatory response induced by the mechanically stressed bacteria to counteract changes in shape.     

Red blood cells under mechanical stress

The effect of mechanical stress on erythrocytes suspended in various media was studied. The ability of the cells to increase their glucose consumption was found to be the major criterion allowing to divide the media into two groups. In plasma, serum or in Ringer's solution supplemented with albumin and glucose the energy consumption by mechanically stressed erythrocytes increased 20 to 50%; no morphological changes of the cells were observed either in suspension or on Giemsa smears. The cells behaved in the same way in Mg2(+)-free medium. The other group included protein-free medium (Ringer's solution supplemented with glucose) and Ca2(+)-free Ringer's solution supplemented with albumin and glucose; under these conditions erythrocytes were unable to raise their energy consumption in response to mechanical stress, and after some period structural impairment of the membrane could be observed on Giemsa smears. No differences in metabolism-associated nucleotide concentrations (ATP, ADP, NAD, NADP) were observed between the samples. Resealed red cell ghosts with high concentrations of intracellular components were prepared as a model of cells with damaged membrane. In these ghosts (with low ATP concentration) mechanical stress produced increased proportions of echinocytes, even in the "native" suspension. These results have confirmed the vital role of the energy-consuming contractile apparatus in the erythrocyte membrane, and supplied a clue to the role of Ca2+ in its activation and to the influence of extracellular proteins on the maintenance of in red cell shape.     

Insights into interstitial flow,shear stress,and mass transport effects on ECM heterogeneity in bioreactor-cultivated engineered cartilage hydrogels

Interstitial flow in articular cartilage is secondary to compressive and shear deformations during joint motion and has been linked with the well-characterized heterogeneity in structure and composition of its extracellular matrix. In this study, we investigated the effects of introducing gradients of interstitial flow on the evolution of compositional heterogeneity in engineered cartilage. Using a parallel-plate bioreactor, we observed that Poiseuille flow stimulation of chondrocyte-seeded agarose hydrogels led to an increase in glycosaminoglycan and type II collagen deposition in the surface region of the hydrogel exposed to flow. Experimental measurements of the interstitial flow fields based on the fluorescence recovery after photobleaching technique suggested that the observed heterogeneity in composition is associated with gradients in interstitial flow in a boundary layer at the hydrogel surface. Interestingly, the interstitial flow velocity profiles were nonlinearly influenced by flow rate, which upon closer examination led us to the original observation that the apparent hydrogel permeability decreased exponentially with increased interfacial shear stress. We also observed that interstitial flow enhances convective mass transport irrespective of molecular size within the boundary layer near the hydrogel surface and that the convective contribution to transport diminishes with depth in association with interstitial flow gradients. The implications of the nonlinearly inverse relationship between the interfacial shear stress and the interstitial flux and permeability and its consequences for convective transport are important for tissue engineering, since porous scaffolds comprise networks of Poiseuille channels (pores) through which interstitial flow must navigate under mechanical stimulation or direct perfusion.     

Experimental and theoretical evidence for convective nutrient transport in an immobilized cell support.

Even though immobilized-cell reactors possess several engineering advantages over free-cell reactors, their full potential has not been realized because mass transfer often limits the rate of nutrient supply and product removal from immobilized cell supports. We studied the interaction between mass transfer and reaction kinetics in the anaerobic conversion of glucose to CO2 and ethanol by yeast immobilized in a porous rotating disk on the agitator shaft of a conventional CSTR. A Sherwood number correlation was used to show that external mass-transfer resistances were negligible under typical operating conditions. The modulus of Weisz based on observable reaction parameters was used to gauge the importance of pore diffusion limitations. Under conditions for which significant pore diffusion effects and hence low effectiveness factors (eta = ca. 0.1) would be predicted, the observed reaction rates were much higher than expected (eta = ca. 1), suggesting that pore diffusion limitations were at least partially relieved by convective transport of glucose into the support. Two possible mechanisms of convective transport are discussed. We hypothesize that gas evolution was responsible for the convective enhancement of glucose supply.     

Psychological stress impairs Na+-dependent glucose absorption and increases GLUT2 expression in the rat jejunal brush-border membrane

Chronic psychological stress impacts many functions of the gastrointestinal tract. However, the effect of stress on nutrient absorption is poorly documented. This study was designed to investigate glucose transporters in rats submitted to different periods of water-avoidance stress (WAS). Rats were subjected to WAS (1 h/day) for 1, 5, or 10 consecutive days. Four hours after the last WAS session, rats were killed and segments of jejunum were mounted in Ussing chambers to study electrophysiological properties of the jejunum and Na+-dependent glucose absorption kinetics. Mucosa was obtained to prepare brush-border membrane vesicles (BBMV) used to measure [14C]fructose uptake as well as sodium-glucose transporter 1 (SGLT-1) and GLUT2 expression by Western blot analysis. Exposure of animals to WAS induced a decrease in Na+-dependent glucose absorption Vmax after 1, 5, and 10 days without any change in SGLT-1 expression. Potential difference across the jejunum was decreased for all stressed groups. Furthermore, we observed an increase in phloretin-sensitive uptake of [14C]fructose by BBMV after 1, 5, or 10 days of WAS, which was not present in control animals. This suggested the abnormal appearance of GLUT2 in the brush border, which was confirmed by Western blot analysis. We concluded that psychological stress induces major changes in glucose transport with a decrease in Na+-dependent glucose absorption and an increase in GLUT2 expression at the brush-border membrane level.     

Role of paxillin in metabolic oxidative stress-induced cytoskeletal reorganization: involvement of SAPK signal transduction pathway and PTP-PEST gene expression

Previous studies have shown that glucose deprivation-induced cell death is associated with apoptosis, which is characterized by cellular membrane blebbing in multi-drug-resistant human breast carcinoma MCF-7/ADR cells. In this study, we investigated the mechanism of glucose deprivation-induced cytoskeletal reorganization, which is known to be responsible for the morphological alterations. An increase in the formation of focal adhesion and stress fibers was observed during the early period of glucose deprivation (1-2 h). However, a disappearance of focal adhesion complexes and a loss of stress fiber formation along with membrane blebbing were observed when glucose deprivation continued. These alterations were delayed in MCF-7/ADR cells transfected with bcl-2 and completely suppressed by treatment with an antioxidant, N-acetyl-L-cysteine. These results indicated that glucose deprivation-induced oxidative stress caused the cytoskeletal reorganization. The glucose deprivation-induced alteration of cytoskeletal organization was further investigated by studying a modification of paxillin, one of the focal adhesion proteins. Immunoblotting with anti-paxillin antibody showed that the paxillin band shifted from 68 kDa to about 80 kDa during 1-4 h of glucose deprivation. The mobility shift indicated the modification of paxillin. This possibility was further studied by an immunoprecipitation assay with anti-paxillin/anti-phosphotyrosine antibody and phosphoamino acid analysis (PAA). The immunoprecipitation study revealed that the level of tyrosine phosphorylation of paxillin was maintained for 2 h and then markedly decreased without a change in the total level of paxillin. The PAA study showed that paxillin is dephosphorylated on tyrosine concurrent with phosphorylation on serine/threonine. Expression of a dominant-negative mutant of c-Jun NH(2)-terminal kinase (JNK1) suppressed glucose deprivation-induced JNK1 activation, PTP-PEST gene expression, and alteration of paxillin. Taken together, these results suggest that the alteration of the phosphorylation/dephosphorylation of paxillin may be related to the cytoskeletal reorganization and these events are mediated by glucose deprivation-induced oxidative stress and the stress-activated protein kinase signal transduction pathway.     

Impaired myocardial metabolic reserve and substrate selection flexibility during stress in patients with idiopathic dilated cardiomyopathy

Under resting conditions, the failing heart shifts fuel use toward greater glucose and lower free fatty acid (FFA) oxidation. We hypothesized that chronic metabolic abnormalities in patients with dilated cardiomyopathy (DCM) are associated with the absence of the normal increase in myocardial glucose uptake and maintenance of cardiac mechanical efficiency in response to pacing stress. In 10 DCM patients and 6 control subjects, we measured coronary flow by intravascular ultrasonometry and sampled arterial and coronary sinus blood. Myocardial metabolism was determined at baseline, during atrial pacing at 130 beats/min, and at 15 min of recovery by infusion of [(3)H]oleate and [(13)C]lactate and measurement of transmyocardial arteriovenous differences of oxygen and metabolites. At baseline, DCM patients showed depressed coronary flow, reduced uptake and oxidation of FFA, and preferential utilization of carbohydrates. During pacing, glucose uptake increased by 106% in control subjects but did not change from baseline in DCM patients. Lactate release increased by 122% in DCM patients but not in control subjects. Cardiac mechanical efficiency in DCM patients was not different compared with control subjects at baseline but was 34% lower during stress. Fatty acid uptake and oxidation did not change with pacing in either group. Our results show that in DCM there is preferential utilization of carbohydrates, which is associated with reduced flow and oxygen consumption at rest and an impaired ability to increase glucose uptake during stress. These metabolic abnormalities might contribute to progressive cardiac deterioration and represent a target for therapeutic strategies aimed at modulating cardiac substrate utilization.     

Effects of the exogenous application of auxin and cytokinin on carbohydrate accumulation in grains of rice under salt stress

Phytohormones, such as auxin and cytokinin, are known to be involved in the regulation of plant responses to salinity stress and counteract the adverse effect of stress conditions. This work investigated the effects of the exogenous spraying of indole-3-acetic acid (IAA) and kinetin (KIN) during the reproductive phase on grain yield by examining the 1000-grain weight and filled-grain percentage as well as the changes in starch, total soluble sugars, sucrose, glucose and fructose concentrations in the grains of two rice cultivars under salt stress. The results indicated that the applied IAA and KIN led to an increased grain yield, 1000-grain weight and filled-grain percentage for both rice cultivars under salt stress. The storage starch content in the grain of the salt-sensitive cultivar was more than that in the salt-tolerant cultivar under IAA application compared with KIN, whereas a decrease in the total soluble sugar content was observed with both IAA and KIN treatments, in comparison to the non-hormone treatment. Interestingly, this study showed that IAA led to a much higher increase in the sucrose content in grain, as compared to the KIN. Furthermore, this experiment suggests that glucose and fructose may play important roles during salt stress because there were clearly higher concentrations of these sugars in the grain of the stressed cultivars under IAA and KIN application: it appears that their accumulation was the earliest response detected during the grain-filling period in rice. Finally, this work indicated that an increase in the rice grain yield, 1000-grain weight and filled-grain percentage are associated with an increase in the contents of starch, sucrose, glucose and fructose in grain caused by the application of IAA and KIN.     

Interaction of the effects of glucose and stress hormones on the activity of key enzymes of liver glycogen metabolism in rats

Endogenous corticosterone released in protracted immobilization stress fails to increase the activity of liver glycogen synthase, perhaps because of the inhibition of synthase phosphatase by phosphorylase a. It was also found, that in rats subjected to acute immobilization stress the stimulation of the activity of both synthase a and total forms by glucose administered i.v. is depressed. Finally, in rats fasting for 24 h a paradoxical augmentation by glucose of the stimulatory effect of glycogenolytic hormones released in acute immobilization stress on phosphorylase a activity was observed.     

Changes in non-structural carbohydrates in olive (Olea europaea) leaves during root zone salinity stress

Self-rooted olive ( Olea europaea L.) plants were grown in hydroponics at various NaCl concentrations (from 0 to 200m M ) for 28 to 32 days followed by 28 to 30 days of relief from salinity over two growing seasons. Olive leaves accumulated both glucose and mannitol during the period of salinity stress. The concentrations of fructose, myo -inositol, galactose, galactinol, sucrose, raffinose, and stachyose were not significantly affected by salinity. Starch content was decreased by salinity. The mannitol/glucose and mannitol/soluble carbohydrates ratios increased as the external NaCl concentration was increased, but returned to the control levels during the relief period. The increase in mannitol or glucose molar concentrations, expressed on a leaf tissue water basis, was partially due to a reduction in leaf tissue water content under salinity stress. However, an increase in mannitol concentration was also observed when expressed on a dry weight basis. The accumulation of mannitol in leaf tissue preceded any reduction in leaf area rate or net assimilation rate. The increase in leaf mannitol or glucose concentration was positively correlated with the increasing level of salinity at the root zone, but not with the accumulation of Na⁺ in the shoot. The role of mannitol. a potential osmoregulator in leaf mesophyll during salinity stress, is discussed in relation to the complex carbohydrate composition of olive leaves.     

Comparative analysis of diffusive and stress induced nutrient transport efficiency in the lacunar-canalicular system of osteons

Marker migration experiments suggest that cyclic mechanical loading of cortical bone in vivo increases marker penetration into bone. Is this a result of stress induced fluid flow or of stress stimulation of active transport processes? Active lacunar-canalicular transport of nutrients was suggested by Ham in 1979 on the basis of the presence of actin filaments in osteocyte processes and their suspected role in cell motility. In addition, Tanaka in 1984 observed active transport of microperoxidase in bone and Tanaka-Kamioka et al. in 1998 observed experimentally that osteocyte processes are able to actively change their form. In this study we performed parametric and comparative analyses of the transport efficiencies of diffusion and stress generated fluid flow of (glucose) nutrients in lacunar-canalicular systems in cortical bone. The result obtained is that neither diffusion nor stress induced fluid flow is capable of sustaining osteocyte viability. It is possible that cyclic stress stimulates an active nutrient transport mechanism to supplement stress flows.     

Quantitative genetic analysis of the physiological stress response in three strains of brook charr Salvelinus fontinalis and their hybrids

Three strains [domestic (D), Laval (L) and Rupert (R)] of brook charr Salvelinus fontinalis and their reciprocal hybrids were submitted to transport stress to measure stress resistance. Primary (cortisol) and secondary (glucose, osmolality and haematocrit) stress responses were measured for each cross. Significant heritabilities were observed for both levels of stress response, with mean ± S.E. heritability (h(2)) = 0.60 ± 0.20 for plasma cortisol and 0.61 ± 0.20 for plasma glucose. There were strain differences whereby the R strain was the least sensitive to stress at the primary and secondary levels. No heterosis was detected, and only one case of outbreeding depression was present. The outbreeding depression was observed in the D(♀) R(♂) hybrid, which had a 27% increase of plasma glucose compared to parental strains. The D(♀) R(♂) and R(♀) L(♂) hybrids had more pronounced variations (increase or decrease) in plasma osmolality than their respective parental strains, but these variations were difficult to relate definitively with the potential secondary stress response. These results indicate a strong potential for genetic improvement in the stress response to transport with the use of purebred crosses while hybridization has little value in this regard.     

Stimulation of glucose transport in L6 muscle cells by long-term intermittent stretch-relaxation.

Skeletal muscle stretch increases resting metabolism and causes hypertrophy. We have examined the effect of mechanical stretch in vitro on glucose transport activity and transporter contents in L6 muscle cells. Long-term (24-48 h) stretch-relaxation (25% maximal elongation at 30 cycles per min) of cell monolayers significantly increased glucose uptake by 1.6- to 2-fold in myotubes but not in myoblasts. The presence of serum was required for the stretch-relaxation induced increase in glucose uptake. Cycloheximide inhibited the mechanical stimulation of glucose uptake, and the latter response was not additive to the stimulatory effect of long-term exposure to insulin. GLUT1 and GLUT4 glucose transporter contents were not changed in total cell membranes from mechanically stimulated cells relative to controls. These results indicate that mechanical stimulation through passive stretch may be an important regulation of nutrient uptake in fetal myotubes independent of innervation.     

Detachment of multi species biofilm in circulating fluidized bed bioreactor

In this study, the detachment rates of various microbial species from the aerobic and anoxic biofilms in a circulating fluidized bed bioreactor (CFBB) with two entirely separate aerobic and anoxic beds were investigated. Overall detachment rate coefficients for biomass, determined on the basis of volatile suspended solids (VSS), glucose and protein as well as for specific microbial groups, i.e., for nitrifiers, denitrifiers, and phosphorous accumulating organisms (PAOs), were established. Biomass detachment rates were found to increase with biomass attachment on carrier media in both beds. The detachment rate coefficients based on VSS were significantly affected by shear stress, whereas for protein, glucose and specific microbial groups, no significant effect of shear stress was observed. High detachment rates were observed for the more porous biofilm structure. The presence of nitrifiers in the anoxic biofilm and denitrifiers in the aerobic biofilm was established by the specific activity measurements. Detachment rates of PAOs in aerobic and anoxic biofilms were evaluated.     

Increased gluconeogenesis in rats exposed to hyper-G stress

The role of gluconeogenesis on the increase in plasma glucose and liver glycogen of rats exposed to hyper-G (radial acceleration) stress was determined. Overnight-fasted, male Sprague-Dawley rats (250-300 g) were injected i.p. with uniformly labeled 1 4C lactate, alanine, or glycerol (5 microCi/rat) and immediately exposed to 3.1G for 0.25, 0.50, and 1.0 hr. 1 4C incorporation of the labeled substrates into plasma glucose and liver glycogen was measured and compared to uncentrifuged control rats injected in a similar manner. Significant increases in 1 4C incorporation of all three labeled substrates into plasma glucose were observed in centrifuged rats at all exposure periods; 1 4C incorporation into liver glycogen was significantly increased only at 0.50 and 1.0 hr. The i.p. administration (5 mg/100-g body wt) of 5-methoxyindole-2-carboxylic acid, a potent gluconeogenesis inhibitor, prior to centrifugation blocked the increase in plasma glucose and liver glycogen during the first hour of centrifugation. The increase in plasma glucose and liver glycogen was also abolished in adreno-demedullated rats exposed to centrifugation for 1.0 hr. Propranolol, a beta-adrenergic blocker, suppressed the increase in plasma glucose of rats exposed to centrifugation for 0.25 hr. From the results of this study, it is concluded that the initial, rapid rise in plasma glucose as well as the increase in liver glycogen of rats exposed to hyper-G stress can be attributed to an increased rate of gluconeogenesis, and that epinephrine plays a dominant role during the early stages of exposure to centrifugation.     

A continuum model for root growth

This paper presents a study of a simple one-dimensional continuum model for growth of the plant root. A fundamental constitutive equation is derived. The model is studied by means of various special cases of increasing complexity. Asymptotic expansions are used to derive approximate solutions to the equation of the model under the fundamental assumption that cell wall thickness is small in comparison with the diameter of the cell. The basic results of the study may be summarized as follows. The observed growth pattern of the root cannot be modelled by a mechanical system whose properties are independent of position on the root. The observed pattern can be modelled by a simple mechanical system in which, for example, cell wall yield stress first decreases and then increases. Two fundamental observations are made based on the modelling study. The first is that any mechanical model must take into account the convective displacement from the tip of points along the root. The second is that in describing growth, data on cell wall mechanical properties are meaningless without corresponding data on cell water potential, and vice versa.     

In vivo porcine left atrial wall stress: Effect of ventricular tachypacing on spatial and temporal stress distribution

Animal models of ventricular tachypacing (VTP) have been successfully used to reproduce the relevant features observed in patients with atrial fibrillation, such as increased atrial pressure and volume, ion-channel alterations and fibrosis. After performing VTP on a healthy Yorkshire pig, we measured an increase in volume of 60%, a two-fold rise in pressure, and a complex pattern of local mechanical, histological and biochemical changes, including a generalized stiffening of the wall. A protocol recently developed was employed to generate computational models of the porcine left atrium mechanics in healthy conditions and after VTP. Comparison of the stress distribution in the healthy vs. VTP case provided a map of how pressure overload affects and modifies left atrium mechanics. Overall, a positive increase in stress was computed after the VTP treatment. Regions of large increase in the stresses post-VTP were the appendage boundaries, the area around the lower pulmonary vein and the area in the front of the atrium towards the appendage. Due to the elevated stress, the back of the atrium mainly modified its mechanical response, while the appendage remodeled both its shape and its mechanical properties. Large changes in the shape of the mitral valve annulus could be observed as a consequence of the remodeling in the front of the atrium. The relation between local mechanical stress and remodeling that emerges from the results is in agreement with our hypothesis that the structural changes in the atrium are a consequence of a stress-mediated mechanism.     

Heme oxygenase-1 is an important modulator in limiting glucose-induced apoptosis in human umbilical vein endothelial cells

Hyperglycaemia is associated with oxidative stress. The inducible isoform of heme oxygenase (HO-1) is an effective system to counteract oxidative stress, yet it is unclear how hyperglycaemia affects HO-1. In this study, we explored: 1) the HO-1 protein content and HO activity in human umbilical vein endothelial cells (HUVECs) exposed to different glucose concentrations, and 2) the mechanisms which account for the high glucose-induced effects on HO-1. We evaluated HO-1 protein expression, HO activity, apoptosis and reactive oxygen species (ROS) in HUVECs treated for 48 h with 5.5, 10 and 20 mM glucose. A dose-dependent production of reactive oxygen species was observed. At 10 mM glucose, an increase of HO-1 protein expression and HO activity was observed, whereas at 20 mM, there was no change in protein content and activity relative to at 5.5 mM glucose. HO-1 protein expression in HUVECs exposed to 20 mM of glucose was increased in the presence of 20 U/ml superoxide dismutase (SOD). HO-1 gene silencing augments ROS production both at 5.5 and 10 mM glucose, leading to an increased apoptosis. We conclude that, in endothelial cells, the regulation of HO-1 by glucose is dependent upon levels of glucose itself. Lack of homeostatic HO-1 upregulation fails to protect from oxidative damage and results in a higher rate of apoptotic cell death.     

虫草发酵菌丝体对胰岛素抵抗大鼠氧化应激的缓解作用

探究虫草发酵菌丝体对胰岛素抵抗(IR)大鼠氧化应激的影响。采用低中高剂量虫草发酵菌丝体(剂量分别为1.65g/kg饲料,3.30g/kg饲料和6.60g/kg饲料)分别干预IR大鼠。口服葡萄糖耐量实验观察不同剂量虫草发酵菌丝体的降糖效果,同时观察血脂、氧化应激水平的变化情况。RT-PCR法测肌肉、肝脏组织氧化应激相关基因Nrf2、HO-1和NQO1mRNA相对表达量,Westernblot法测其蛋白相对表达量。结果发现CM干预能够显著改善IR大鼠空腹血糖、血脂和机体氧化应激水平,肌肉、肝脏组织Nrf2、HO-1和NQO1mRNA相对表达量显著上升,肝脏Nrf2和NQO1蛋白相对表达量显著上调,并且具有剂量依赖性。说明CM具有缓解IR大鼠氧化应激的作用。