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PubMed2006

Activation pathways of alpha4beta1 integrin leading to distinct T-cell cytoskeleton reorganization, Rac1 regulation and Pyk2 phosphorylation.

Alpha4beta1 integrin is highly expressed in lymphocytes and is essential in hematopoiesis, extravasation, and the inflammatory response. Alpha4beta1 can be activated by intracellular signals elicited upon T-cell activation by phorbol esters, CD3 crosslinking, or certain chemokine/receptor interactions (inside-out activation). Divalent cations or certain anti-beta1 mAbs (i.e., TS2/16) can also bind and activate integrins directly (outside-in activation). In both cases, activation results in increased adhesion and/or affinity for ligands. It is not known if these various stimuli produce the same or different post-adhesion events. To address this, we have studied the cytoskeleton organization and intracellular signaling following activation of 41 in Jurkat cells and in human T-lymphoblasts. Treatment with Mn2+, alpha-CD3 mAb or the chemokine SDF-1alpha followed by attachment to the fibronectin fragment H89 or the endothelial molecule VCAM-1 (alpha4beta1 ligands), resulted in cell polarization and migration. In contrast, activation with PMA or TS2/16 induced cell spreading and strong adherence. Video microscopy and Transwell analyses confirmed these results, which correlated with different resistance to detachment under flow. Activation of the small GTPase RhoA or transfection with the constitutively active mutants V14RhoA or V12Rac1, abolished the alpha4beta1-induced cell polarization but did not affect cell spreading. Moreover, Rac1 activity was distinctly modulated by agents that induce a polarized or spread phenotype. The tyrosine kinase Pyk2 was highly phosphorylated upon induction of cell polarity but not during cell spreading. These results reveal novel properties of alpha4beta1 integrin, namely the ability to trigger two types of T-cell cytoskeletal response with different signaling requirements.

Cell Adhesion↗

PubMed1994

P-selectin-dependent leukocyte recruitment and intestinal mucosal injury induced by lactoferrin.

Plasma concentrations of lactoferrin relevant to an inflammatory response are known to elicit leukocyte-endothelial cell adhesion in mesenteric venules. The objectives of this study were (1) to determine whether exogenously administered lactoferrin causes microvascular and mucosal injury in rat intestine and (2) to assess the contribution of adherent leukocytes to a lactoferrin-mediated injury process. Mucosal myeloperoxidase (MPO) activity and vascular protein clearance were monitored in the distal intestine of male Sprague-Dawley rats. Macroscopic erosive lesions of the mucosa and increases in mucosal MPO and intestinal vascular protein were observed 2 h following the lactoferrin infusion, results consistent with granulocyte accumulation and microvascular protein leakage. These lactoferrin-induced alterations were significantly attenuated in animals pretreated with a monoclonal antibody (mAb) directed against P-selectin but not by an E-selectin-specific mAb. In another series of experiments, leukocyte adherence/emigration and leakage of fluorescein isothiocyanate (FITC)-labeled albumin were measured in rat mesenteric venules using intravital video microscopy. Lactoferrin elicited increases in both leukocyte adhesion/emigration and albumin extravasation, which were attenuated by mAbs directed against P-selectin but not E-selectin. These observations indicate that (1) the lactoferrin released by activated neutrophils may lead to significant microvascular and mucosal injury or dysfunction and (2) the lactoferrin-induced injury is related to P-selectin-mediated adhesion of leukocytes to microvascular endothelium. Our results raise the possibility that neutrophil-derived lactoferrin contributes to the inflammatory response by promoting further granulocyte accumulation and activation and that mAbs to P-selectin may be therapeutically beneficial in inflammatory disorders.

Animals↗

PubMed2004

Larval development of Phoronis pallida (Phoronida): implications for morphological convergence and divergence among larval body plans.

Morphological variation among larval body plans must be placed into a phylogenetic and ecological context to assess whether similar morphologies are the result of phylogenetic constraints or convergent selective pressures. Investigations are needed of the diverse larval forms within the Lophotrochozoa, especially the larvae of phoronids and brachiopods. The actinotroch larva of Phoronis pallida (Phoronida) was reared in the laboratory to metamorphic competence. Larval development and growth were followed with video microscopy, SEM, and confocal microscopy. Early developmental features were similar to other phoronid species. Gastrulation was accomplished by embolic invagination of the vegetal hemisphere. Mesenchymal cells were found in the remaining blastocoelic space after invagination began. Mesenchymal cells formed the body wall musculature during the differentiation of larval features. Body wall musculature served as the framework from which all other larval muscles proliferated. Larval growth correlated best with developmental stage rather than age. Consistent with other phoronid species, differentiation of juvenile tissues occurred most rapidly at the latest stages of larval development. The minimum precompetency period of P. pallida was estimated to be approximately 4-6 weeks. Previously published studies have documented that the planktonic embryos of P. pallida develop faster than the brooded embryos of P. vancouverensis. However, these data showed that the difference in developmental rate between the two species decreased in succeeding larval stages. There may be convergent selective pressures that result in similar timing to metamorphic competence among phoronid and brachiopod planktotrophic larval types. Morphological differences between these larval types result from heterochronic developmental shifts in the differentiation of juvenile tissue. Similarities in the larval morphology of phoronids and basal deuterostomes are likely the result of functional and developmental constraints rather than a shared (recent) evolutionary origin. These constraints are imposed by the functional design of embryological stages, feeding structures, and swimming structures.

Animals↗

PubMed2003

Three-dimensional method for comparing in vivo interventional MR images of thermally ablated tissue with tissue response.

PURPOSE: To investigate the ability of magnetic resonance (MR) to monitor radio-frequency (RF) ablation treatments by comparing MR images of thermal lesions to histologically assayed cellular damage. We developed a new methodology using three-dimensional registration for making spatial correlations. MATERIALS AND METHODS: A low-field, open MRI system was used to guide an ablation probe into rabbit thigh muscle and acquire MR volumes after ablation. After fixation, we sliced and photographed the tissue at 3-mm intervals, using a specially designed apparatus, to obtain a volume of tissue images. Histologic samples were digitized using a video microscopy system. For our three-dimensional registration method, we used the tissue images as the reference, and registered histology and MR images to them using two different computer alignment steps. First, the MR volume was aligned to the volume of tissue images by registering needle fiducials placed near the tissue of interest. Second, we registered the histology images with the tissue images using a two-dimensional warping technique that aligned internal features and the outside boundary of histology and tissue images. RESULTS: The MR and histology images were very well aligned, and registration accuracy, determined from displacement of needle fiducials, was 1.32 +/- 0.39 mm (mean +/- SD), which compared favorably to the MR voxel dimensions (0.70 mm in-plane and 3.0 mm thick). A preliminary comparison of MR and tissue response showed that the region inside the elliptical hyperintense rim in MR closely corresponds to the region of necrosis as established by histology, with a mean absolute distance between MR and histology boundaries of 1.17 mm, slightly smaller than the mean registration error. The MR region slightly overestimated the region of necrosis, with a mean signed distance between boundaries of 0.85 mm. CONCLUSION: Our results suggest that our methodology can be used to achieve three-dimensional registration of histology and in vivo MR images. In MR lesion images, the inner border of the hyperintense region corresponds to the border of irreversible cell damage. This is good evidence that during RF ablation treatments, iMRI lesion images can be used for real-time feedback.

Animals↗

PubMed2004

Radiofrequency thermal ablation: correlation of hyperacute MR lesion images with tissue response.

PURPOSE: To investigate the hypothesis that the outer boundary of the hyperintense region observed in hyperacute (several minutes post-ablation) T2 and gadolinium contrast-enhanced (CE) T1-weighted magnetic resonance (MR) lesion images is an accurate predictor of eventual cell death from radiofrequency (RF) thermal ablation. MATERIALS AND METHODS: A low-field, open MR imaging system was used to guide an ablation electrode into a thigh muscle of five rabbits and acquire in vivo T2 and CE T1-weighted MR volumes. Ablation occurred by applying RF current for two minutes with the electrode's temperature maintained at 90 degrees +/- 2 degrees C. After fixation, we sliced and photographed the tissue at 3 mm intervals, using a specially designed apparatus, to obtain a volume of tissue images. Digital images of hematoxylin and eosin (H&E) and Masson trichrome-stained histologic samples were obtained, and distinct regions of tissue damage were labeled using a video microscopy system. After the MR and histology images were aligned using a three-dimensional registration method, we compared tissue damage boundaries identified in histology with boundaries marked in MR images. RESULTS: The lesions have distinct zones of tissue damage histologically: a central zone of necrotic cells surrounded by an outer zone with cells that appeared non-viable and associated with marked interstitial edema. In 14 histology images from five lesions, the inner and outer boundaries of the outer zone were compared with the boundaries of a hyperintense rim that surrounds a central hypointense region in the T2 and CE T1-weighted MR images. For T2 and CE T1-weighted MR images, respectively, the mean absolute distance was 1.04 +/- 0.30 mm (mean +/- SD) and 1.00 +/- 0.34 mm for the inner boundaries, and 0.96 +/- 0.34 mm and 0.94 +/- 0.44 mm for the outer boundaries. The mean absolute distances for T2 and CE T1-weighted MR images were not sufficiently different to achieve statistical significance (P = 0.745, 0.818, for the inner and outer boundary, respectively). CONCLUSION: In hyperacute T2 and CE T1-weighted MR lesion images, observations strongly suggest that the outer boundary of the hyperintense rim corresponds to the region of eventual cell necrosis within a distance comparable to our ability to measure. This is good evidence that during RF ablation procedures, MR lesion images can be used to accurately localize the zone of irreversible tissue damage at the lesion margin.

Animals↗

PubMed2001

Laser pulse impact on rat mesenteric blood vessels in relation to laser treatment of port wine stain.

BACKGROUND AND OBJECTIVE: To study the impact of laser pulses on animal microvasculature as a model for laser treatment of port wine stains. STUDY DESIGN/MATERIALS AND METHODS: Rat mesenteric blood vessels were irradiated with a laser pulse (585 nm, 0.2-0.6 ms pulse duration, 0.5-30 J/cm(2) radiant exposure). Video microscopy was used to assess vessel dilation, formation of intravascular thrombi, bubble formation, and vessel rupture. Changes in reflection during a laser pulse were measured by simultaneously recording the temporal behavior of the incident and reflected signals. RESULTS: A threshold radiant exposure of approximately 3 J/cm(2) was found for changes in optical properties of blood in vivo, confirming previous in vitro results. Often, laser exposure induced a significant increase in vessel diameter, up to three times the initial diameter for venules and four times for arterioles, within 200 ms after laser exposure. Arterioles were more likely to dilate than venules. Sometimes, immediately after the pulse, round structures, interpreted as being gas bubbles, were seen within the vessel lumen. CONCLUSIONS: A variety of phenomena can occur when blood vessels of sizes comparable to those in port wine stains are irradiated with laser pulses as used in port wine stain treatment. Thrombus formation and vessel rupture have been described before from histological sections of laser-irradiated port wine stains. However, vessel dilation and formation of non-transient gas bubbles as found in this study have not been described before.

Animals↗

PubMed1989

Intravascular thrombosis in skeletal muscle microcirculation after ischemia.

This study was undertaken to elucidate the role of thrombosis in blood vessels less than 0.1 mm in diameter in skeletal muscle after an ischemic episode. Capillaries were examined after normothermic ischemia using intravital video microscopy of the cremaster muscle of an anesthetized rat. Histologic sections of the cremaster muscle and silicone rubber intravascular casts were also analyzed. Our model demonstrates that, after 3 hours of warm ischemia, up to two-thirds of capillaries are clotted, as is most of the venous system. Our findings also indicate that capillaries may reopen during 30 minutes of reperfusion. These findings suggest that ischemia may cause thrombosis in the microvasculature of skeletal muscle but the thrombosis appears to be partially reversible.

Animals↗

PubMed2005

Allurin, a 21 kD sperm chemoattractant, is rapidly released from the outermost jelly layer of the Xenopus egg by diffusion and medium convection.

Allurin, a 21 kD protein from Xenopus laevis egg jelly, has been demonstrated to attract sperm by video microscopy and by quantitative chemotaxis chamber assays. Here, we use immunocytochemistry to demonstrate that this sperm chemoattractant is located in the outermost layer of egg jelly (J3) and is rapidly released into the surrounding medium. SDS-PAGE analysis and Western blotting confirm the appearance of allurin in the medium within 1.5 min and separation of proteins in the medium by anion exchange FPLC, shows that nearly half of the allurin released over a 12-hr period is discharged in the first 5 min. The kinetics of allurin release from J3 and its appearance in the medium were quantitatively accounted for, by computer simulation of mathematical diffusion and convection models. Comparison of simulation data to quantitative measurements of allurin appearance in the medium suggests that allurin, although larger than most chemoattractants, is effectively dispersed by a combination of diffusion and medium mixing at the jelly surface during spawning. Our model further predicts that the innermost jelly layer, J1, is less permeable to allurin than the other layers, allowing it to act as a "reflector" to speed up allurin discharge.

Algorithms↗

PubMed2002

Survival of bundleless hair cells and subsequent bundle replacement in the bullfrog's saccule.

Our senses of hearing and balance depend upon hair cells, the sensory receptors of the inner ear. Millions of people suffer from hearing and balance deficits caused by damage to hair cells as a result of exposure to noise, aminoglycoside antibiotics, and antitumor drugs. In some species such damage can be reversed through the production of new cells. This proliferative response is limited in mammals but it has been hypothesized that damaged hair cells might survive and undergo intracellular repair. We examined the fate of bullfrog saccular hair cells after exposure to a low dose of the aminoglycoside antibiotic gentamicin to determine whether hair cells could survive such treatment and subsequently be repaired. In organ cultures of the bullfrog saccule a combination of time-lapse video microscopy, two-photon microscopy, electron microscopy, and immunocytochemistry showed that hair cells can lose their hair bundle and survive as bundleless cells for at least 1 week. Time-lapse and electron microscopy revealed stages in the separation of the bundle from the cell body. Scanning electron microscopy (SEM) of cultures fixed 2, 4, and 7 days after antibiotic treatment showed that numerous new hair bundles were produced between 4 and 7 days of culture. Further examination revealed hair cells with small repaired hair bundles alongside damaged remnants of larger surviving bundles. The results indicate that sensory hair cells can undergo intracellular self-repair in the absence of mitosis, offering new possibilities for functional hair cell recovery and an explanation for non-proliferative recovery.

Animals↗

PubMed2003

Methylene blue blocks cGMP production and disrupts directed migration of microglia to nerve lesions in the leech CNS.

Migration and accumulation of microglial cells at sites of injury are important for nerve repair. Recent studies on the leech central nervous system (CNS), in which synapse regeneration is successful, have shown that nitric oxide (NO) generated immediately after injury by endothelial nitric oxide synthase (eNOS) stops migrating microglia at the lesion. The present study obtained results indicating that NO may act earlier, on microglia migration, and aimed to determine mechanisms underlying NO's effects. Injury induced cGMP immunoreactivity at the lesion in a pattern similar to that of eNOS activity, immunoreactivity, and microglial cell accumulation, which were all focused there. The soluble guanylate cyclase (sGC) inhibitor methylene blue (MB) at 60 microM abolished cGMP immunoreactivity at lesions and blocked microglial cell migration and accumulation without interfering with axon conduction. Time-lapse video microscopy of microglia in living nerve cords showed MB did not reduce cell movement but reduced directed movement, with significantly more cells moving away from the lesion or reversing direction and fewer cells moving toward the lesion. The results indicate a new role for NO, directing the microglial cell migration as well as stopping it, and show that NO's action may be mediated by cGMP.

Animals↗

PubMed1992

Neurite outgrowth on a step gradient of chondroitin sulfate proteoglycan (CS-PG).

Sulfated proteoglycans (PGs) may play a significant role in the regulation of neurite outgrowth. They are present in axon-free regions of the developing nervous system and repel elongating neurites in a concentration-dependent manner in vitro. The addition of growth-promoting molecules, such as laminin, can modify the inhibitory effect of PGs on neurite outgrowth (Snow, Steindler, and Silver, 1990b). Substrata containing a high-PG/low-laminin ratio completely inhibit neurite outgrowth, while normal, unimpeded outgrowth is observed on low-PG/high-laminin substrata. Therefore, different patterns of neurite outgrowth may result from regulation of the ratio of growth-promoting molecules to growth-inhibiting molecules. Using video microscopy, embryonic chicken dorsal root ganglia neurons (DRG), chicken retinal ganglia neurons (RGC), and rat forebrain neurons (FB) were analyzed as they extended processes from a substratum consisting of laminin alone onto a step gradient of increasing concentrations of chondroitin sulfate proteoglycan (CS-PG) bound to laminin. In contrast to neurite outgrowth inhibition that occurs at the border of a single stripe of high concentration of CS-PG (Snow et al., 1990b and this study), growth cones grew onto and up CS-PG presented in a step-wise graded distribution. Although the behavior of the different cell types was unique, a common behavior of each cell type was a decrease in the rate of neurite outgrowth with increasing CS-PG concentration. These data suggest that appropriate concentrations of growth-promoting molecules combined with growth-inhibiting molecules may regulate the direction and possibly the timing of neurite outgrowth in vivo. The different responses of different neuronal types suggest that the presence of sulfated PG may have varying effects on different aspects of neuronal development.

Aggrecans↗

PubMed1994

Growth behavior of retinotectal axons in live zebrafish embryos under TTX-induced neural impulse blockade.

The growth dynamics of individual DiO-labeled retinal axons deprived of normal neural impulse activity by TTX was monitored in the tectum of living zebrafish embryos with time-lapse video microscopy and compared with normal active axons. Growth cones of TTX-blocked axons advance intermittently with an average velocity similar to normal axons. While exploring their local environment, they are broadened and bear ruffling lamellipodia and filopodia, but become streamlined when advancing. The activity-deprived axons grow directly towards their retinotopic target sites in the tectum as do their normal counterparts and very rarely extend branches en route. Much like normal axons, TTX-blocked axons begin to branch and develop their terminal arbors only at their retinotopic target area. They emit and retract numerous short side branches over a period of several hours. The area they contact (the "exploration field") is of similar dimension as that of active axons, covering from 1% to 7.4% of the tectal neuropil surface, but the final arbors cover an area only one-half to one-sixth as large. TTX arbors are as small as arbors of normal active axons and retinotopically correct. Thus, the typical exploratory growth behavior of developing retinal axons in the tectum, the dynamics of terminal arbor formation at retinotopically correct sites, the dimension of the exploration field, and the shaping of the arbors in zebrafish embryos are unaffected by TTX-induced neural impulse blockade.

Animals↗

PubMed1994

Pharmacological characterization of a serotonin receptor involved in an early embryonic behavior of Helisoma trivolvis.

In contrast to the abundance of information on the many physiological and developmental actions of serotonin in molluscan nervous systems, comparatively little is known about the serotonin receptors involved in these responses. Embryos of the pulmonate gastropod, Helisoma trivolvis, display a cilia-driven rotational behavior that is regulated by endogenous serotonin. In the present study, two functional assays were used to determine some of the pharmacological properties of the receptors that mediate the cilio-excitatory action of serotonin. Time-lapse video microscopy was used to measure whole embryo rotation rate and cilia beat frequency in isolated cells. In dose-response experiments, serotonin was approximately 10 times more potent in stimulating cilia beat frequency over embryo rotation. In rotation experiments, 5-carboxyamidotryptamine and methysergide had effective agonist activity in dose ranges similar to that of serotonin (1 to 100 microM). In contrast, 8-hydroxydipropylaminotetralin HBr (8-OH-DPAT) displayed agonist activity of lower potency and effectiveness. Several compounds displayed antagonist activity in the 1 to 100 microM dose range, including mianserin, spiperone, ritanserin, 1-(1-naphthyl)piperazine, and propranolol. alpha-Methylserotonin had mixed agonist-antagonist activity, and metoclopramide, MDL-72222, and ketanserin were inactive. Experiments on isolated cells suggested that the extremely effective antagonism displayed by mianserin in the embryo rotation assay was due to its specific activity at ciliary serotonin receptors. These results implicate the presence of a novel serotonin receptor on embryonic ciliated cells that is pharmacologically distinct from those previously characterized in vertebrate or invertebrate systems.

Animals↗

PubMed1995

Inhibition of formation of filopodia after axotomy by inhibitors of protein tyrosine kinases.

The activity of motile protrusions of the growth cone--filopodia, veils, and lamellipodia--is essential for directed growth of a neuronal process. The regulation of the formation of these protrusions is not well understood. Numerous filopodia and veils or lamellipodia form within minutes of transection of an Aplysia axon in culture, as the initial components of growth cones of regenerating neurites. Axotomy, therefore, provides a robust and reliable protocol for analyzing the formation of these protrusions. We evaluated the involvement of protein phosphorylation in the regulation of protrusive activity. Of the inhibitors of protein kinases assayed, only the inhibitors of protein tyrosine kinases--genistein, lavendustin A, herbimycin A, and erbstatin analogue--suppressed the formation of protrusions, as assessed by high magnification video microscopy. These drugs did not work by preventing resealing of the axon, as evident from visual inspection and by the unimpaired effectiveness of genistein or lavendustin in preventing formation of filopodia when applied after resealing. Inhibition of protein tyrosine kinases not only prevented the formation of actin-based protrusions, but also caused deterioration of the actin network underlying the protrusive area of preexisting growth cones. Consistent with an involvement of protein tyrosine phosphorylation in the generation of protrusive structures, immunocytochemistry revealed that aggregates of phosphotyrosine appeared at the margins of the axon, from which protrusions emerge shortly after axotomy. These results suggest a role for protein tyrosine phosphorylation in the formation and maintenance of actin-based protrusive structures.

Actins↗

PubMed1985

The effects of calcium regulating hormones on bone resorption by isolated human osteoclastoma cells.

Cells were disaggregated from osteoclastomas, and the response of the giant cells to calcium-regulating hormones, prostaglandin (PG)E1 and dibutyryl cyclic AMP (dbcAMP) was observed by phase-contrast time-lapse video microscopy. The pattern and nature of their response was very similar to that previously found to be characteristic of osteoclasts: calcitonin (CT), PGE1 and dbcAMP induced cytoplasmic quiescence, while parathyroid hormone (PTH) showed no influence on cytoplasmic motility or behaviour. The cells were also cultured on slices of devitalized cortical bone for 5 or 18 h. After this time the giant cells were associated with the appearance in the scanning electron microscope of characteristic resorption pits, the volume of which was calculated by computer-assisted morphometric and stereophotogrammetric techniques after removal of cells. Calcitonin caused a dramatic reduction in the volume of bone resorbed by these isolated cells compared with control cultures, while PTH was without significant effect. This result supports the view that PTH does not increase bone resorption in intact bone through a direct effect on osteoclasts. PGE1, which stimulates bone resorption when added to intact bone, paradoxically reduced resorption in our cultures. It thus appears possible that PGE1 acts as a direct inhibitor of osteoclastic bone resorption but has an additional effect on other cells in bone, which are induced by PGE1 to cause osteoclastic stimulation.

Alprostadil↗

PubMed2001

A quantitative histochemistry technique for measuring regional distribution of acetylcholinesterase in the brain using digital scanning densitometry.

Studies of brain acetylcholinesterase (AChE) are traditionally based on biochemical assays, immunoreactivity, and histochemistry. Conventional histochemistry yields rich morphological data from tissue sections but yields quantitative results only with great difficulty. Several histochemical methods developed in recent years, including microdensitometry, microphotometry, and video-based histochemistry, are effective in quantitative and detailed study of AChE in tissue sections. However, they are usually time-consuming. As we report here, we adapted digital scanning densitometry to quantitate AChE histochemical staining in brain sections. The AChE and butyrylcholinesterase (BuChE), as measured by the method, were heterogeneously distributed throughout the brain, results that are consistent with those obtained by biochemical methods. The staining intensity is dependent on section thickness, substrate concentration, and reaction time. The cholinesterase inhibitor methyl paraoxon significantly decreased AChE staining intensity. Furthermore, data acquired from densitometry are similar to those obtained by video-based microscopy or by spectrophotometry. The advantage of the densitometric measurements compared to other quantitative histochemical methods is that it is very rapid while collecting data that are equivalent in quality. Because the digital scanning densitometers provide high quality and sensitive imaging, wide dynamic ranges, and convenient image analysis software, they are very useful tools in quantitative histochemistry.

Acetylcholinesterase↗

PubMed1993

Modulation of airway epithelial cell ciliary beat frequency by nitric oxide.

NOS activity has been recently described in airway epithelial cells. Because these cells are often ciliated we hypothesized that NO modulates airway ciliary beating. CBF was measured in cultured BBECs using video microscopy. L-NMMA, a NOS inhibitor, caused a 40% decrease in CBF following pre-stimulation with isoproterenol (8.5 +/- 0.3 Hz vs 14.6 +/- 0.2 Hz; p < 0.0001) which lasted approximately 60 minutes. Similar attenuation in CBF after isoproterenol pre-treatment was observed with another NOS inhibitor, L-NAME. NOS inhibitor-induced CBF slowing was also observed when cells were pre-stimulated with either bradykinin or substance P and was completely reversed by L-arginine or SNP but not by D-arginine. These observations demonstrate a novel NO-dependent mechanism that upregulates ciliary motility in response to stimulation.

Amino Acid Oxidoreductases↗

PubMed1993

Fluid shear stress induces synthesis and nuclear localization of c-fos in cultured human endothelial cells.

Quiescent cultured primary human umbilical vein endothelial cells were exposed to low levels (4 dynes/cm2) or arterial levels (25 dynes/cm2) of steady laminar shear stress for one hour. Intracellular c-fos protein was measured by immunocytochemistry and quantitative fluorescence video microscopy. The nuclear-localized c-fos protein level was 5.4 +/- 2.0 fold higher (p < 0.01) in the cells exposed to arterial levels of shear stress as compared to the cells maintained in stationary culture. In contrast, the cells exposed to low levels of shear stress showed diffuse but slightly elevated levels of c-fos (2.4 +/- 0.73 times higher than control; p < 0.01) without preferential nuclear localization. The protein kinase C inhibitor, H7 (10 microM) significantly attenuated the induction of c-fos by 50% in cells exposed to arterial shear stress for 1 hour.

Cell Compartmentation↗