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At least 1,171 records · Page 65Linked to original sources
PubMed1999

Activation of cAMP-dependent protein kinase A in prefrontal cortex impairs working memory performance.

Activation of the adenylyl cyclase-cAMP-protein kinase A (PKA) intracellular signaling cascade is necessary for long-term memory consolidation in brain regions such as the hippocampus. However, the role of the PKA cascade in the working memory functions of the prefrontal cortex (PFC) is unknown. The present study examined the effects of manipulating PKA activity in the PFC using the cAMP stereoisomers Sp-cAMPS and Rp-cAMPS, which activate and inhibit PKA, respectively. Animals received bilateral infusions of Sp-cAMPS and/or Rp-cAMPS into the PFC immediately before testing on the delayed alternation task, a test of spatial working memory that depends on the integrity of the PFC. Low doses of Sp-cAMPS (0.21, 2. 1, or 21 nmol/0.5 microl) produced a marked, dose-dependent impairment in working memory performance. The impairment produced by infusion of Sp-cAMPS (21 nmol/0.5 microl) was fully reversed by co-infusion of Rp-cAMPS (21 nmol/0.5 microl), consistent with actions on PKA. Rp-cAMPS (21 or 42 nmol/0.5 microl) by itself had no effect on performance. These results indicate that activation of the PKA intracellular signaling cascade in the PFC impairs working memory performance. The current findings contrast with studies of long-term memory consolidation, in which inhibition of PKA with agents such as Rp-cAMPS impaired memory consolidation (Bernabeu et al., 1997; Bourtchouladze et al., 1998), whereas enhancement of the PKA pathway improved memory (Bernabeu et al., 1997; Barad et al., 1998). These results demonstrate that discrete cognitive processes subserved by different cortical regions are mediated by distinct intracellular mechanisms.

Animals↗
PubMed2006

Oxytocin alleviates oxidative renal injury in pyelonephritic rats via a neutrophil-dependent mechanism.

BACKGROUND: Urinary tract infection (UTI) may cause inflammation of the renal parenchyma and may lead to impairment in renal function and scar formation. Oxidant injury and reactive oxygen species (ROS) have been found responsible in the pathogenesis of UTI. The neurohypophyseal hormone oxytocin (OT) facilitates wound healing and is involved in the modulation of immune and inflammatory processes. We investigated the possible therapeutic effects of OT against Escherichia coli induced pyelonephritis in rats both in the acute and chronic setting. METHODS: Twenty-four Wistar rats were injected 0.1 ml solution containing E. coli ATCC 25922 10(10) colony forming units/ml into left renal medullae. Six rats were designed as sham group and were given 0.1 ml 0.9% NaCl. Pyelonephritic rats were treated with either saline or OT immediately after surgery and at daily intervals. Half of the pyelonephritic rats were decapitated at the 24th hour of E. coli infection, and the rest were followed for 7 days. Renal function tests (urea, creatinine), systemic inflammation markers [lactate dehydrogenase (LDH) and tumor necrosis factor alpha (TNF-alpha)] and renal tissue malondialdehyde (MDA) as an end product of lipid peroxidation, glutathione (GSH) as an antioxidant parameter and myeloperoxidase (MPO) as an indirect index of neutrophil infiltration were studied. RESULTS: Blood urea, creatinine, and TNF-alpha levels were increased, renal tissue MDA and MPO levels were elevated and GSH levels were decreased in both of the pyelonephritic (acute and chronic) rats. All of these parameters and elevation of LDH at the late phase were all reversed to normal levels by OT treatment. CONCLUSION: OT alleviates oxidant renal injury in pyelonephritic rats by its anti-oxidant actions and by preventing free radical damaging cascades that involves excessive infiltration of neutrophils.

Animals↗
PubMedDate not supplied

Paraspinal denervation and the spinal degenerative cascade.

BACKGROUND CONTEXT: The aging of the spine has been reported to follow a degenerative cascade described by Kirkaldy-Willis. This cascade does not take into account the paraspinal muscles. Substantial recent knowledge of the anatomy, physiology and pathophysiology of the paraspinal muscles suggests that they should play a role in the degenerative cascade. Pain relates to paraspinal denervation in a number of disorders. Using new needle electromyographic technique, MiniPM, we may be able to quantify the extent of paraspinal muscle denervation. PURPOSE: To explore the potential interactions between paraspinal muscle denervation and the degenerative cascade. STUDY DESIGN: Literature review, reanalysis of a blinded electromyographic study and case reports. PATIENT SAMPLE: A previously reported group of 35 asymptomatic paid volunteers, ages 18 to 58 years. Two elderly volunteers with moderate to severe spinal stenosis on magnetic resonance imaging (MRI), one significantly disabled, one asymptomatic. OUTCOME MEASURES: Blinded electromyographic testing, including MiniPM, a codified, quantified needle electromyography technique for the paraspinal muscles. In the two case reports, magnetic resonance imaging and 15-minute timed ambulation. METHODS: The 35 volunteers were tested by a blinded electromyographer using the MiniPM technique. The two older subjects underwent MRI, MiniPM and extensive electrodiagnostic evaluations, functional testing, complete history and physical examination. RESULTS: The patient with symptomatic stenosis had a MiniPM score of 6 on each side, and the asymptomatic volunteer had a score of 0. In the asymptomatic subject group, reproducible evidence for denervation was found in 20 subjects (57%), sparsely distributed, but more caudal than cranial (p<.0001). Based on conservative estimates of nerve regeneration rates, the population averaged a detected axonotemesis denervating event approximately once every 120 days. CONCLUSIONS: Paraspinal denervation may be proven to be a better marker than MRI findings for symptomatic spinal stenosis. Subtle paraspinal muscle denervation occurs frequently in asymptomatic persons. Whether a cause or an effect, paraspinal denervation might lead to hypermobility and thus progression along the degenerative cascade.

Adolescent↗
PubMed2005

The steroid hormone-regulated gene Broad Complex is required for dendritic growth of motoneurons during metamorphosis of Drosophila.

Dendrites are subject to subtle modifications as well as extensive remodeling during the assembly and maturation of neural circuits in a wide variety of organisms. During metamorphosis, Drosophila flight motoneurons MN1-MN4 undergo dendritic regression, followed by regrowth, whereas MN5 differentiates de novo (Consoulas et al. [2002] J. Neurosci. 22:4906-4917). Many cellular changes during metamorphosis are triggered and orchestrated by the steroid hormone 20-hydroxyecdysone, which initiates a cascade of coordinated gene expression. Broad Complex (BRC), a primary response gene in the ecdysone cascade, encodes a family of transcription factors (BRC-Z1-Z4) that are essential for metamorphic reorganization of the central nervous system (CNS). Using neuron-filling techniques that reveal cellular morphology with very high resolution, we tested the hypothesis that BRC is required for metamorphic development of MN1-MN5. Through a combination of loss-of-function mutant analyses, genetic mapping, and transgenic rescue experiments, we found that 2Bc function, mediated by BRC-Z3, is required selectively for motoneuron dendritic regrowth (MN1-MN4) and de novo outgrowth (MN5), as well as for soma expansion of MN5. In contrast, larval development and dendritic regression of MN1-MN4 are BRC-independent. Surprisingly, BRC proteins are not expressed in the motoneurons, suggesting that BRC-Z3 exerts its effect in a non-cell-autonomous manner. The 2Bc mutants display no gross defects in overall thoracic CNS structure, or in peripheral structures such as target muscles or sensory neurons. Candidates for mediating the effect of BRC-Z3 on dendritic growth of MN1-MN5 include their synaptic inputs and non-neuronal CNS cells that interact with them through direct contact or diffusible factors.

Animals↗
PubMed1987

Mechanism of complement-mediated thromboxane production by the perfused rabbit liver: lack of effect of dantrolene sodium.

Activated components of the complement system have been shown to stimulate the arachidonic acid cascade. We have reported that hepatic thromboxane production in response to plasma activated with zymosan is self-limiting, not affected by nifedipine, but inhibited by mepacrine, a phospholipase inhibitor. To further study this relationship, we have tested the effects of dantrolene sodium, an agent reported to immobilize intracellular calcium. Control group livers were perfused with Krebs-Henseleit bicarbonate buffer at a rate of 120 ml/min in a nonrecirculating perfusion system and administered 1 ml/min of normal rabbit plasma for 10 minutes. This group of livers demonstrated stable wet weight, perfusion pressure, and rates of release of lactic dehydrogenase, thromboxane B2, and prostacyclin over a 150 minute experimental period. In contrast, the administration of 1 ml/min of zymosan-activated plasma resulted in significant increases in the rate of thromboxane B2 release at 1, 3, and 5 minutes after the start of the infusion. The rate of thromboxane production then returned to baseline values. Neither prostacyclin nor lactic dehydrogenase release changed significantly after ZAP. A similar change in thromboxane production following ZAP administration was seen in livers being continually perfused with 10 microM dantrolene sodium. Perfusion pressure was significantly elevated in this group during the ZAP infusion period. These results confirm complement-mediated thromboxane production in the isolated rabbit liver model but do not describe a definitive role of dantrolene-sensitive intracellular calcium release in the mechanism of ZAP-mediated thromboxane production.

6-Ketoprostaglandin F1 alpha↗
PubMed2004

Inhibition of phosphatidylinositol 3'-kinase induces preferentially killing of PTEN-null T leukemias through AKT pathway.

We examined the functional role of the phosphatidylinositol 3'-kinase pathway in the growth and survival of cell lines of T-cell origin. Pharmacological inhibition of PI3'-kinase using LY294002 resulted in apoptosis of acute lymphoblastic T-cell leukemia (T-ALL) cell lines including CEM, Jurkat, and MOLT-4. On the other hand, the cutaneous T-cell lymphoma cell line HUT-78 was found to be refractory to LY294002- inducible apoptosis. Sensitivity or resistance to pharmacological inhibitors of PI3'-kinase correlated with tumor suppressor PTEN gene expression, as sensitive T-ALL cells do not express PTEN and have high level of activated AKT, in contrast to HUT-78 cells. Our data demonstrate that inhibition of PI3'-kinase results in dephosphorylation of AKT and partial inhibition of Bcl-xL expression in T-ALL cells, but not in HUT-78 cells. Interestingly, HUT-78 cells were also found to express higher levels of Bcl-xL protein as compared to T-ALL cells. Inhibition of PI3'-kinase also induces release of cytochrome c from mitochondria and activation of caspase-3 and PARP in all T-ALL cell lines tested, but not in HUT-78 cells. Taken altogether, our data demonstrate that the PI3'-kinase/AKT pathway plays a major role in the growth and survival of PTEN-null T-ALL cells, and identify this cascade as promising target for therapeutic intervention in acute T-cell leukemias.

Apoptosis↗
PubMed2005

Temporal development of hippocampal cell death is dependent on tissue strain but not strain rate.

Deformation of brain tissue in response to mechanical loading of the head is the root-cause of traumatic brain injury (TBI). Even below ultimate failure limits, deformation activates pathophysiological cascades resulting in delayed cell death. Injury response of soft tissues, such as the chest and spinal cord, is dependent on the product of deformation and velocity, a parameter termed the viscous criterion. We set out to test if hippocampal cell death could be predicted by a similar combination of strain and strain rate and if the viscous criterion was valid for hippocampus. Quantitative prediction of the brain's biological response to mechanical stimuli is difficult to achieve in animal models of TBI, so we utilized an in vitro model of TBI based on hippocampal slice cultures. We quantified the temporal development of cell death after precisely controlled deformations for 30 combinations of strain (0.05-0.50) and strain rate (0.1-50s(-1)) relevant to TBI. Loading conditions for a subset of cultures were verified by analysis of high-speed video. Cell death was found to be significantly dependent on time-post injury, on strain magnitude, and to a lesser extent, on anatomical region by a repeated-measures, three-way ANOVA. The responses of the CA1 and CA3 regions of the hippocampus were not statistically different in contrast to some in vivo TBI studies. Surprisingly, cell death was not dependent on strain rate leading us to conclude that the viscous criterion is not a valid predictor for hippocampal tissue injury. Given the large data set and extensive combinations of biomechanical parameters, predictive mathematical functions relating independent variables (strain, region, and time post-injury) to the resultant cell death were defined. These functions can be used as tolerance criteria to equip finite element models of TBI with the added capability to predict biological consequences.

Analysis of Variance↗
PubMed2002

Volumetric determination of the wear of ceramics for hip joints.

Osteolysis relating to the reaction cascade to wear debris is the main cause of the failure of arthroprostheses. New materials are still under development to minimise the wear of joints and to improve in this way the performance of total joint replacements. Testing the wear performance of very low wear materials is a rather sophisticated technique. Currently, the worn volume is calculated from the weight variations of the sample, observed from wear testing. This method may be limited especially when very low weight differences are to be evaluated on high hardness, high-density materials like CoCr alloys or ceramics like alumina or zirconia. In the present work, the tribological behaviour of ceramic biomaterials like alumina, Y-TZP and alumina-zirconia composites was evaluated by a pin-on-disc apparatus under different testing conditions. The worn volume was calculated by the use of 3-D (three-dimensional) optical profilometry data. Comparison between 3-D optical profilometry and gravimetric wear data shows the better accuracy of the profilometric method in the determination of wear rate in the range of 10(-7) mm3 (mN(-1)) or lower.

Biomechanical Phenomena↗
PubMedDate not supplied

SSRI antidepressant use patterns and their relation to clinical global impression scores: a naturalistic study.

BACKGROUND: A cascade of events follows initial antidepressant selection which includes the subsequent antidepressant use pattern, resultant clinical outcomes, and associated health care expenditures. PURPOSE: The purpose of this study using data from a clinical practice setting was to test whether the pattern of antidepressant use was correlated with patients' treatment response as measured by the score on the Clinical Global Impression-Improvement scale. DATA AND METHODS: A retrospective dataset of patients who initiated therapy on fluoxetine, fluvoxamine, paroxetine, or sertraline in a primary care setting in Spain was used. A Cox proportional hazard analysis was used to predict the likelihood of treatment response based upon the pattern of initial antidepressant use, while minimizing the influence of other factors. RESULTS: After controlling for other observed baseline characteristics including initial disease severity, (a) patients who remained on their initial antidepressant therapy for at least 2 months with no switching, augmentation, or upward dose titration were 1.63 times more likely to experience a treatment response than patients who had an adjustment to therapy; and (b) patients who initiated therapy on sertraline were 0.46 times as likely to experience a treatment response as patients who initiated therapy on the most common study antidepressant, fluoxetine. CONCLUSION: The pattern of antidepressant use is an important determinant of treatment response among patients initiating therapy on the newer antidepressants in clinical practice.

Adult↗
PubMed2002

Characterization of inhalation aerosols: a critical evaluation of cascade impactor analysis and laser diffraction technique.

Cascade impactor analysis is the standard technique for in vitro characterization of aerosol clouds generated by medical aerosol generators. One important reason for using this inertial separation principle is that drug fractions are classified into aerodynamic size ranges that are relevant to the deposition in the respiratory tract. Measurement of these fractions with chemical detection methods enables establishment of the particle size distribution of the drug in the presence of excipients. However, the technique is laborious and time consuming and most of the devices used for inhaler evaluation lack sufficient possibilities for automation. In addition to that, impactors often have to be operated under conditions for which they were not designed and calibrated. Particularly, flow rates through impactors are increased to values at which the flow through the nozzles is highly turbulent. This has an uncontrolled influence on the collection efficiencies and cut-off curves of these nozzles. Moreover, the cut-off value varies with the flow rate through an impactor nozzle. On the other hand, the high air flow resistances of most impactors are rather restricting to the attainable (fixed) inspiratory flow curves through these devices. Especially for breath actuated dry powder inhalers, higher flow rates and flow increase rates may be desirable than can be achieved in combination with a particular type of impactor. In this paper, the applicability of laser diffraction technology is evaluated as a very fast and highly reliable alternative for cascade impactor analysis. With this technique, aerodynamic diameters cannot be measured, but for comparative evaluation and development, comprising most in vitro applications, this is not necessary. Laser diffraction has excellent possibilities for automated recording of data and testing conditions, and the size classes are independent of the flow rate. Practical limitations can be overcome by using a special inhaler adapter which enables control of the inspiratory flow curve through the inhaler, analysis of the emitted fine particle mass fraction and pre-separation of large particles during testing of dry powder inhalers containing adhesive mixtures.

Administration, Inhalation↗
PubMed1999

Anti-GAD monoclonal antibody delays the onset of diabetes mellitus in NOD mice.

Insulin Dependent Diabetes Mellitus (IDDM type I) is the result of autoimmune destruction of insulin producing pancreatic beta-cells by the cellular immune system, specifically, autoreactive T cells. Disease progression is evident by multiple autoantibodies responding to self-antigens in a cascade mechanism, wherein the first self-antigen induces the activation of the immune system, leading to the destruction of beta-cells and consequently, exposure of other antigens. Glutamic Acid Decarboxylase (GAD) is recognized in the literature as a primary autoantigen involved in the cascade. We questioned the immunological involvement of this autoantigen in the overall progression of the disease, specifically if antigen recognition by the cellular immune system (T cells) is necessary for organ specific autoimmunity and cellular toxicity. We tested this hypothesis by isolating, purifying and injecting monoclonal antibodies against GAD (anti-GAD Ab; 0.1 mg or 0.3 mg) into non-obese diabetic (NOD) mice on a weekly basis. We suggest that the anti-GAD Ab will bind to the GAD antigen, or perhaps bind to the epitope presented in association with APC-MHC and prevent T cell recognition, thereby delaying disease onset. Our results demonstrate a delay in the onset of diabetes and a decrease in the severity of insulitis in our test animals, when compared to controls. The mechanism of action of the anti-GAD Ab may be associated with a passive protection mechanism, as evidenced by the fact that splenocytes transferred from anti-GAD Ab treated mice did not prevent or delay diabetes in syngeneic irradiated NOD mice. The mechanism of diabetes prevention by administration of anti-GAD antibody could be associated with an interference in recognition of GAD by T cells, and continuing research will be perform to investigate this hypothesis.

Animals↗
PubMed2000

Ras/MEK signaling suppresses Myc-dependent apoptosis in cells transformed by c-myc and activated ras.

Cooperation of myc and activated ras has been suggested to cause malignant cell transformation but the mechanism is still unknown. Here we isolated a transformed cell line in which activation of c-Myc and Ras are independently controllable, and show that after establishment of the transformed state by c-myc and activated ras, removal of activated Ras initiates apoptosis that is dependent on c-Myc activity. Apoptosis is also initiated by an inhibitor of MEK (MAPK/ERK kinase), a kinase downstream of Ras, and apoptosis is blocked by activated Mek1. These results suggest that one of the conditions required for establishment of the transformed state is a block of apoptosis involving MEK activity. We tested the effect of MEK inhibition on cells transformed by various oncogenes. Suppression of apoptosis by MEK is not critical in general, but in cells transformed by c-myc plus a gene that activates the MAPK cascade it is necessary to avoid cell death. Activated Ras/MEK did not suppress c-myc-dependent apoptosis due to serum-limitation. Overexpression of chicken bcl-xL suppressed apoptosis under serum-limiting conditions, but not apoptosis initiated by Ras/MEK inhibition in cells transformed by myc and activated ras. Altogether, these results suggest the existence of a novel regulatory mechanism for myc-dependent apoptosis in certain transformed cells.

Animals↗
PubMed1989

Interleukin 2 acts as an adjuvant to increase the potency of inactivated rabies virus vaccine.

Interleukin 2 (IL-2) occupies a central position in the cascade of events involved in the immune response. We were interested in determining whether IL-2 could function as an adjuvant to vaccination, to increase the immune response to vaccine immunogens. Using the National Institutes of Health test for rabies vaccine potency, we found that daily systemic administration of IL-2 in conjunction with inactivated rabies virus can increase the potency of vaccination in outbred mice at least 25-fold, as measured by survival following challenge with virulent rabies virus. Enhanced protection is not correlated with an increase in virus-neutralizing antibody titers, and we suggest that IL-2 acts to increase the cellular immune response to vaccination.

Adjuvants, Immunologic↗
PubMed2006

Early diagnosis of early gastric cancer.

The prognosis of gastric cancer is closely related to the stage of disease at diagnosis. Early gastric cancer, whereby disease is limited to mucosa and submucosa, confers a survival rate of greater than 90% in 5 years in many centres. Gastric cancer is still a major cause of cancer mortality worldwide. In high incidence areas such as Japan, screening of asymptomatic population has been advocated. However, in Western countries, mass screening is not cost-effective. Hence, strategy has been directed to screen symptomatic individuals who are at higher risk of gastric cancer. Most patients with early gastric cancer present with symptoms indistinguishable from benign peptic ulcer disease. Screening for this group of patients improves detection rate of early gastric cancer and therefore its prognosis. Endoscopy for surveillance of premalignant lesions has been explored with this objective in mind. Serology testing for biomarkers such as pepsinogen, anti-Helicobacter pylori antibody and gastrin has been studied as an alternative to endoscopy. There is compelling evidence for the role of H. pylori in the initiation of Correa's cascade (stepwise progression from chronic active gastritis, atrophic gastritis, intestinal metaplasia, dysplasia and finally adenocarcinoma). Regression of premalignant lesions has been demonstrated with H. pylori eradication. However, it is not known whether this might effectively prevent gastric cancer in either low or high-risk population.

Early Diagnosis↗
PubMed1985

Non-steroid anti-inflammatory drugs in asthma: dangerous or useful therapy?

The therapeutic potential of non-steroid anti-inflammatory drugs in clinical asthma is offset by the real possibility of hypersensitivity and induction of severe airways obstruction. The influence of indomethacin on the antigen-induced asthmatic response was tested. Early and delayed asthmatic responses were recorded after antigen challenge in 13 subjects. Indomethacin pretreatment totally or partially inhibited the delayed asthmatic response in 10 of 11 subjects. Inhibition by indomethacin of products of the arachidonic cascade which participate in the pathogenesis of the delayed asthmatic response could explain this phenomenon. A similar therapeutic response was documented without adverse drug reactions when five subjects were restudied after several months. In the same group the early asthmatic response was suppressed in six, enhanced in two and unchanged in four of 12-subjects. This variable response indicates that spasmogenic prostaglandin breakdown products may be important for certain individuals, but are generally of less importance in the early asthmatic response. Clinical trials with indomethacin as a steroid saving agent in allergic asthma appear feasible and can be conducted safely.

Adolescent↗
PubMed2000

Feature selection and classifier performance in computer-aided diagnosis: the effect of finite sample size.

In computer-aided diagnosis (CAD), a frequently used approach for distinguishing normal and abnormal cases is first to extract potentially useful features for the classification task. Effective features are then selected from this entire pool of available features. Finally, a classifier is designed using the selected features. In this study, we investigated the effect of finite sample size on classification accuracy when classifier design involves stepwise feature selection in linear discriminant analysis, which is the most commonly used feature selection algorithm for linear classifiers. The feature selection and the classifier coefficient estimation steps were considered to be cascading stages in the classifier design process. We compared the performance of the classifier when feature selection was performed on the design samples alone and on the entire set of available samples, which consisted of design and test samples. The area Az under the receiver operating characteristic curve was used as our performance measure. After linear classifier coefficient estimation using the design samples, we studied the hold-out and resubstitution performance estimates. The two classes were assumed to have multidimensional Gaussian distributions, with a large number of features available for feature selection. We investigated the dependence of feature selection performance on the covariance matrices and means for the two classes, and examined the effects of sample size, number of available features, and parameters of stepwise feature selection on classifier bias. Our results indicated that the resubstitution estimate was always optimistically biased, except in cases where the parameters of stepwise feature selection were chosen such that too few features were selected by the stepwise procedure. When feature selection was performed using only the design samples, the hold-out estimate was always pessimistically biased. When feature selection was performed using the entire finite sample space, the hold-out estimates could be pessimistically or optimistically biased, depending on the number of features available for selection, the number of available samples, and their statistical distribution. For our simulation conditions, these estimates were always pessimistically (conservatively) biased if the ratio of the total number of available samples per class to the number of available features was greater than five.

Algorithms↗
PubMed1991

ACTH and cortisol secretion in patients with Alzheimer's disease.

The "glucocorticoid cascade hypothesis" for pathological ageing of the brain is supported by strong experimental data, but the clinical correlates are far less clear. The basal ACTH and cortisol secretion have been studied before and after the dexamethasone suppression test in patients in the early stages of clinically probable Alzheimer's disease and in controls, and the results were all normal. These findings do not support the hypothesis that the pathological brain ageing of Alzheimer's type is caused by hyperactivity of the pituitary-adrenal axis.

Adrenocorticotropic Hormone↗
PubMed2006

Stretch-induced phosphorylation of focal adhesion kinase in endothelial cells: role of mitochondrial oxidants.

Mechanical stretch activates a number of signaling pathways in endothelial cells, and it elicits a variety of functional responses including increases in the phosphorylation of focal adhesion kinase (FAK), a nonreceptor tyrosine kinase involved in integrin-mediated signal transduction. Stretch also triggers an increase in the generation of reactive oxygen species (ROS), which may function as second messengers in the signal transduction cascades that activate cellular responses to strain. Mitochondria represent an important source of ROS in the cell, and these organelles may release ROS in response to strain by virtue of their attachment to cytoskeletal proteins. We therefore tested whether cyclic stretch increases FAK phosphorylation at Tyr397 through a mitochondrial ROS signaling pathway in bovine pulmonary artery endothelial cells (BPAEC). Oxidant signaling, measured using 2'7'-dichlorofluorescin (DCFH), increased 152 +/- 16% during 1.5 h of cyclic strain relative to unstrained controls. The mitochondrial inhibitors diphenylene iodonium (5 microM) or rotenone (2 microM) attenuated this increase, whereas L-nitroarginine (100 microM), allopurinol (100 microM), or apocynin (30 microM) had no effect. The antioxidants ebselen (5 microM) and dithiodidiethyldithiocarbamate (1 mM) inhibited the strain-induced increase in oxidant signaling, but Hb (5 microM) had no effect. These results indicate that strain induces oxidant release from mitochondria. Treatment with cytochalasin D (5 microM) abrogated strain-induced DCFH oxidation in BPAEC, indicating that actin filaments were required for stretch-induced mitochondrial ROS generation. Cyclic strain increased FAK phosphorylation at Tyr397, but this was abolished by mitochondrial inhibitors as well as by antioxidants. Strain-induced FAK phosphorylation was abrogated by inhibition of protein kinase C (PKC) with Ro-31-8220 or Gö-6976. These findings indicate that mitochondrial oxidants generated in response to endothelial strain trigger FAK phosphorylation through a signaling pathway that involves PKC.

Animals↗