Lung fluid absorption is induced in preterm guinea pigs ventilated with low tidal volumes.
Time Factors; Animals; Gestational Age; Guinea Pigs; *Premature Birth; Phosphorylation; Enzyme Activation; Permeability; Extracellular Signal-Regulated MAP Kinases/metabolism; Intubation; Epithelial Sodium Channels/metabolism; Absorption; Sodium-Potassium-Exchanging ATPase/metabolism; Fetal Organ Maturity; Epinephrine/blood; Extravascular Lung Water/*metabolism; *Tidal Volume; Albumins/administration & dosage/*metabolism; Hydrocortisone/blood; Lung/embryology/*metabolism/physiopathology; MAP Kinase Kinase Kinases/metabolism; Ventilator-Induced Lung Injury/metabolism/physiopathology/*prevention & control; Newborn; Intratracheal; Artificial/adverse effects/*methods; Respiration
The objective of this study was to determine if low tidal volume (V(t)) ventilation was beneficial when ventilating preterm fetuses. The authors ventilated preterm guinea pig fetuses at gestation day (GD) 67, 3 days before birth, newborn, and 10-day-old (PD10) guinea pigs with low V(t) (6 mL/kg body weight [bw]) and compared them to age-matched fetuses/animals ventilated with higher potentially injurious V(t) (12 mL/kg bw). Lung fluid absorption was measured after intratracheal instillation of 5% albumin in 0.9% NaCl. Low V(t) ventilation stimulated lung fluid absorption when compared to higher V(t) in all groups. The increased lung fluid absorption in low V(t)-ventilated fetuses was associated with increased alpha epithelial Na channel (alphaEnaC) mRNA. However, alphaENaC and betaENaC protein was unchanged over the 1-hour study. Because stretch induces mitogen-activated protein (MAP) kinase expression and MAP kinases may affect lung fluid absorption, the authors investigated if MAP kinase (MAPK) expression was affected by V(t). Extracellular signal-regulated kinase (ERK) and MAPK/ERK kinase (MEK) were phosphorylated in the higher V(t)-ventilated guinea pig fetuses. This suggested that a reduced activation of MAP kinases might explain the increased lung fluid absorption in the low V(t)-ventilated fetuses. Thus these data suggest that low V(t) ventilation increases fetal lung fluid absorption and thus may be preferential to use clinically.
Koshy Shyny; Beard LaMonta L; Kuzenko Stephanie R; Li Tianbo; Folkesson Hans G
Experimental lung research
2011
2011-02
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.3109/01902148.2010.514024" target="_blank" rel="noreferrer noopener">10.3109/01902148.2010.514024</a>
Angiotensin II-induced extracellular signal-regulated kinase 1/2 activation is mediated by protein kinase Cdelta and intracellular calcium in adult rat cardiac fibroblasts.
Acetophenones/pharmacology; Angiotensin II/*physiology; Animals; Benzopyrans/pharmacology; Calcium/*metabolism; Cell Proliferation; Cells; Cultured; Enzyme Activation; ErbB Receptors/metabolism; Fibroblasts/*metabolism; Male; Mitogen-Activated Protein Kinase 1/*metabolism; Mitogen-Activated Protein Kinase 3/*metabolism; Myocardium/*cytology/metabolism; Phorbol Esters/pharmacology; Phosphorylation; Protein Kinase C-delta/genetics/*metabolism; Rats; Signal Transduction/physiology; Sprague-Dawley
Angiotensin II (Ang II)-induced proliferation of cardiac fibroblasts is a major contributing factor to the pathogenesis of cardiac fibrosis. Ang II activates extracellular signal-regulated kinase (ERK) 1/2 to induce cardiac fibroblast proliferation, but the signaling pathways leading to ERK 1/2 activation have not been elucidated in these cells. The goal of the current study was to identify the intracellular mediators of Ang II-induced ERK 1/2 activation in adult rat cardiac fibroblasts. We determined that 100 nmol/L of Ang II-induced ERK 1/2 phosphorylation is inhibited by simultaneous chelation of cytosolic calcium and downregulation of protein kinase C (PKC) by phorbol ester or by the specific PKCdelta inhibitor rottlerin, as well as PKCdelta small interfering RNA, but not by inhibition of 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetate, phorbol ester, rottlerin, or PKCdelta small interfering RNA alone. We also found that Ang II does not transactivate the epidermal growth factor receptor in adult cardiac fibroblasts, because pretreatment with 1 mumol/L of AG 1478 did not significantly inhibit [(3)H]-thymidine incorporation or ERK 1/2 activation. In addition, immunoprecipitation of the epidermal growth factor receptor demonstrated no significant Ang II-induced phosphorylation of tyrosine residues. Inhibition of phosphatidylinositide 3-kinase, PKCzeta, and src tyrosine kinase had no effect on Ang II-induced ERK 1/2 activation. Collectively, these data demonstrate that Ang II does not transactivate the epidermal growth factor receptor in adult rat cardiac fibroblasts to activate ERK 1/2, a common pathway described in vascular smooth muscle and other cell types, but rather occurs via activation of distinct parallel signaling pathways mechanistically controlled by intracellular Ca(2+) and PKCdelta.
Olson Erik R; Shamhart Patricia E; Naugle Jennifer E; Meszaros J Gary
Hypertension (Dallas, Tex. : 1979)
2008
2008-03
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1161/HYPERTENSIONAHA.107.098459" target="_blank" rel="noreferrer noopener">10.1161/HYPERTENSIONAHA.107.098459</a>
Redox-dependent coronary metabolic dilation.
*Paracrine Communication; *Vasodilation/drug effects; Acetylcysteine/pharmacology; Animals; Antioxidants/pharmacology; Cardiac/drug effects/*metabolism; Conditioned/metabolism; Coronary Vessels/drug effects/enzymology/*metabolism; Culture Media; Dithiothreitol/pharmacology; Enzyme Activation; Fluorescence; Hydrogen Peroxide/metabolism; Imidazoles/pharmacology; In Vitro Techniques; Microscopy; Myocytes; Nitroprusside/pharmacology; Oxidation-Reduction; p38 Mitogen-Activated Protein Kinases/antagonists & inhibitors/metabolism; Phosphorylation; Protein Kinase Inhibitors/pharmacology; Pyridines/pharmacology; Rats; Reactive Oxygen Species/*metabolism; Reducing Agents/pharmacology; Sulfhydryl Compounds/*metabolism; Vasodilator Agents/pharmacology; Wistar
We have observed that hydrogen peroxide (H2O2), the dismutated product of superoxide, is a coronary metabolic dilator and couples myocardial oxygen consumption to coronary blood flow. Because the chemical activity of H2O2 favors its role as an oxidant, and thiol groups are susceptible to oxidation, we hypothesized that coronary metabolic dilation occurs via a redox mechanism involving thiol oxidation. To test this hypothesis, we studied the mechanisms of dilation of isolated coronary arterioles to metabolites released by metabolically active (paced at 400 min) isolated cardiac myocytes and directly compared these responses with authentic H2O2. Studies were performed under control conditions and using interventions designed to reduce oxidized thiols [0.1 microM dithiothreitol (DTT) and 10 mM N-acetyl-L-cysteine (NAC)]. Aliquots of the conditioned buffer from paced myocytes produced vasodilation of isolated arterioles (peak response, 71% +/- 6% of maximal dilation), whereas H2O2 produced complete dilation (92% +/- 7%). Dilation to either the conditioned buffer or to H2O2 was significantly reduced by the administration of either NAC or DTT. The location of the thiols oxidized by the conditioned buffer or of H2O2 was determined by the administration of the fluorochromes monochlorobimane (20 microM) or monobromotrimethylammoniobimane (20 microM), which covalently label the reduced total or extracellular-reduced thiols, respectively. H2O2 or the conditioned buffer predominantly oxidized intracellular thiols since the fluorescent signal from monochlorobimane was reduced more than that of monobromotrimethylammoniobimane. To determine whether one of the intracellular targets of thiol oxidation that leads to dilation is the redox-sensitive kinase p38 mitogen-activated protein (MAP) kinase, we evaluated dilation following the administration of the p38 inhibitor SB-203580 (10 microM). The inhibition of p38 attenuated dilation to either H2O2 or to the conditioned buffer from stimulated myocytes by a similar degree, but SB-203580 did not attenuate dilation to nitroprusside. Western blot analysis for the activated form of p38 (phospho-p38) in the isolated aortae revealed robust activation of this enzyme by H2O2. Taken together, our results show that an active component of cardiac metabolic dilation, like that of H2O2, produces dilation by the oxidation of thiols, which are predominantly intracellular and dependent activation on the p38 MAP kinase. Thus coronary metabolic dilation appears to be mediated by redox-dependent signals.
Saitoh Shu-ichi; Kiyooka Takahiko; Rocic Petra; Rogers Paul A; Zhang Cuihua; Swafford Albert; Dick Gregory M; Viswanathan Chandrasekar; Park Yoonjung; Chilian William M
American journal of physiology. Heart and circulatory physiology
2007
2007-12
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1152/ajpheart.00436.2007" target="_blank" rel="noreferrer noopener">10.1152/ajpheart.00436.2007</a>
PKCalpha mediates acetylcholine-induced activation of TRPV4-dependent calcium influx in endothelial cells.
Acetylcholine/*pharmacology; Animals; Calcium Signaling/*drug effects; Carbazoles/pharmacology; Cells; Cultured; Endothelial Cells/*drug effects/enzymology; Enzyme Activation; Inbred C57BL; Knockout; Male; Mesenteric Arteries/drug effects/enzymology; Mice; Mutation; Nitric Oxide Synthase Type III/metabolism; Phosphorylation; Protein Kinase C-alpha/genetics/*metabolism; Protein Kinase Inhibitors/pharmacology; Tetradecanoylphorbol Acetate/pharmacology; Time Factors; Transfection; TRPV Cation Channels/*agonists/deficiency/genetics/metabolism; Vasodilation/*drug effects; Vasodilator Agents/*pharmacology
Transient receptor potential vanilloid channel 4 (TRPV4) is a polymodally activated nonselective cationic channel implicated in the regulation of vasodilation and hypertension. We and others have recently shown that cyclic stretch and shear stress activate TRPV4-mediated calcium influx in endothelial cells (EC). In addition to the mechanical forces, acetylcholine (ACh) was shown to activate TRPV4-mediated calcium influx in endothelial cells, which is important for nitric oxide-dependent vasodilation. However, the molecular mechanism through which ACh activates TRPV4 is not known. Here, we show that ACh-induced calcium influx and endothelial nitric oxide synthase (eNOS) phosphorylation but not calcium release from intracellular stores is inhibited by a specific TRPV4 antagonist, AB-159908. Importantly, activation of store-operated calcium influx was not altered in the TRPV4 null EC, suggesting that
Adapala Ravi K; Talasila Phani K; Bratz Ian N; Zhang David X; Suzuki Makoto; Meszaros J Gary; Thodeti Charles K
American journal of physiology. Heart and circulatory physiology
2011
2011-09
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1152/ajpheart.00142.2011" target="_blank" rel="noreferrer noopener">10.1152/ajpheart.00142.2011</a>
Prostate apoptosis response-4 enhances secretion of amyloid beta peptide 1-42 in human neuroblastoma IMR-32 cells by a caspase-dependent pathway.
*Intracellular Signaling Peptides and Proteins; Amyloid beta-Peptides/*biosynthesis/metabolism; Apoptosis Regulatory Proteins; Apoptosis/*physiology; Carrier Proteins/genetics/*physiology; Caspases/*metabolism; Cultured; Enzyme Activation; Humans; Kinetics; Leucine Zippers; Neuroblastoma; Peptide Fragments/*biosynthesis/metabolism; Recombinant Proteins/metabolism; Time Factors; Transfection; Tumor Cells
Prostate apoptosis response-4 (Par-4) is a leucine zipper protein that promotes neuronal cell death in Alzheimer's disease (AD). Neuronal degeneration in AD may result from extracellular accumulation of amyloid beta peptide (Abeta) 1-42. To examine the effect of Par-4 on Abeta secretion and to reconcile amyloid/apoptosis hypotheses of AD, we generated IMR-32 cell lines that overexpress Par-4 and/or its leucine zipper domain. Overexpression of Par-4 did not significantly affect levels of the endogenously expressed beta amyloid precursor protein but drastically increased the Abeta(1-42)/Abeta(total) ratio in the conditioned media about 6-8 h after trophic factor withdrawal. Time course analysis of caspase activation reveals that Par-4 overexpression exacerbated caspase activation, which is detectable within 2 h after trophic factor withdrawal. Furthermore, inhibition of caspase activity by the broad spectrum caspase inhibitor BD-fmk significantly attenuated the Par-4-induced increase in Abeta 1-42 production. In addition, the effects of Par-4 on secretion of Abeta 1-42 were consistently blocked by co-expression of the leucine zipper domain, indicating that the effect of Par-4 on Abeta secretion may require its interaction with other protein(s). These results suggest that Par-4 increases secretion of Abeta 1-42 largely through a caspase-dependent pathway after apoptotic cascades are initiated.
Guo Q; Xie J; Chang X; Du H
The Journal of biological chemistry
2001
2001-05
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1074/jbc.M010996200" target="_blank" rel="noreferrer noopener">10.1074/jbc.M010996200</a>
The large protein 'L' of Peste-des-petits-ruminants virus exhibits RNA triphosphatase activity, the first enzyme in mRNA capping pathway.
Acid Anhydride Hydrolases/*metabolism; Animals; Baculoviridae/genetics; Cercopithecus aethiops; Cloning; Conventional mRNA capping; Enzyme Activation; Gene Expression; Genetic Vectors/genetics; Messenger/*genetics/*metabolism; Molecular; Morbillivirus; mRNA capping; Peste-des-petits-ruminants virus L protein; Peste-des-petits-ruminants virus/*physiology; Peste-des-Petits-Ruminants/*virology; PPRV; RNA; RNA Caps/*metabolism; RNA triphosphatase; Vero Cells; Viral Proteins/*metabolism
Peste-des-petits-ruminants is a highly contagious and fatal disease of goats and sheep caused by non-segmented, negative strand RNA virus belonging to the Morbillivirus genus-Peste-des-petits-ruminants virus (PPRV) which is evolutionarily closely related to Rinderpest virus (RPV). The large protein 'L' of the members of this genus is a multifunctional catalytic protein, which transcribes and replicates the viral genomic RNA as well as possesses mRNA capping, methylation and polyadenylation activities; however, the detailed mechanism of mRNA capping by PPRV L protein has not been studied. We have found earlier that the L protein of RPV has RNA triphosphatase (RTPase), guanylyltransferase (GTase) and methyltransferase activities, and unlike vesicular stomatitis virus (VSV), follows the conventional pathway of mRNA capping. In the present work, using a 5'-end labelled viral RNA as substrate, we demonstrate that PPRV L protein has RTPase activity when present in the ribonucleoprotein complex of purified virus as well as recombinant L-P complex expressed in insect cells. Further, a minimal domain in the C-terminal region (aa1640-1840) of the L protein has been expressed in E. coli and shown to exhibit RTPase activity. The RTPase activity of PPRV L protein is metal-dependent and functions with a divalent cation, either magnesium or manganese. In addition, RTPase associated nucleotide triphosphatase activity (NTPase) of PPRV L protein is also demonstrated. This work provides the first detailed study of RTPase activity and identifies the RTPase domain of PPRV L protein.
Ansari Mohammad Yunus; Singh Piyush Kumar; Rajagopalan Deepa; Shanmugam Purnima; Bellur Asutosh; Shaila Melkote Subbarao
Virus Genes
2019
2019-02
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1007/s11262-018-1617-5" target="_blank" rel="noreferrer noopener">10.1007/s11262-018-1617-5</a>
Osteoactivin Promotes Migration of Oral Squamous Cell Carcinomas.
*Cell Movement; Carcinoma; Cell Adhesion; Cell Line; Cell Proliferation; Cell Survival; Enzyme Activation; Gene Expression Regulation; Head and Neck Neoplasms/genetics/*metabolism/pathology; Humans; Integrin beta1/metabolism; Membrane Glycoproteins/genetics/*metabolism; Messenger/metabolism; Mitogen-Activated Protein Kinases/metabolism; Mouth Neoplasms/genetics/*metabolism/pathology; Neoplasm Invasiveness; Neoplastic; Protein Binding; RNA; RNA Interference; Signal Transduction; Squamous Cell Carcinoma of Head and Neck; Squamous Cell/genetics/*metabolism/pathology; Time Factors; Transfection; Tumor
Nearly 50% of patients with oral squamous cell carcinoma (OSCC) die of metastases or locoregional recurrence. Metastasis is mediated by cancer cell adhesion, migration, and invasion. Osteoactivin (OA) overexpression plays a role in metastases in several malignancies. The aims were to determine how integrin interactions modulate OA-induced OSCC cell migration; and to investigate OA effects on cell survival and proliferation. We confirmed OA mRNA and protein overexpression in OSCC cell lines. We assessed OA's interactions with integrins using adhesion inhibition assays, fluorescent immunocytochemistry and co-immunoprecipitation. We investigated OA-mediated activation of mitogen-activated protein kinases (MAPKs) and cell survival. Integrin inhibition effects on OA-mediated cell migration were determined. We assessed effects of OA knock-down on cell migration and proliferation. OA is overexpressed in OSCC cell lines, and serves as a migration-promoting adhesion molecule. OA co-localized with integrin subunits, and co-immunoprecipitated with the subunits. Integrin blocking antibodies, especially those directed against the beta1 subunit, inhibited cell adhesion (P = 0.03 for SCC15 cells). Adhesion to OA activated MAPKs in UMSCC14a cells and OA treatment promoted survival of SCC15 cells. Integrin-neutralizing antibodies enhanced cell migration with OA in the extracellular matrix. OA knock-down resulted in decreased proliferation of SCC15 and SCC25 cells, but did not inhibit cell migration. OA in the extracellular matrix promotes OSCC cell adhesion and migration, and may be a novel target in the prevention of HNSCC spread. J. Cell. Physiol. 231: 1761-1770, 2016. (c) 2015 Wiley Periodicals, Inc.
Arosarena Oneida A; Dela Cadena Raul A; Denny Michael F; Bryant Evan; Barr Eric W; Thorpe Ryan; Safadi Fayez F
Journal of cellular physiology
2016
2016-08
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1002/jcp.25279" target="_blank" rel="noreferrer noopener">10.1002/jcp.25279</a>