7 results on '"Pittman, Laura M."'
Search Results
2. Coordinating epidermal growth factor-induced motility promotes efficient wound closure
- Author
-
Kurten, Richard C., primary, Chowdhury, Parag, additional, Sanders, Ronald C., additional, Pittman, Laura M., additional, Sessions, Laura W., additional, Chambers, Timothy C., additional, Lyle, Christopher S., additional, Schnackenberg, Bradley J., additional, and Jones, Stacie M., additional
- Published
- 2005
- Full Text
- View/download PDF
3. Enhanced β2-adrenergic receptor (β2AR) signaling by adeno-associated viral (AAV)-mediated gene transfer
- Author
-
Jones, Stacie M, Hiller, F Charles, Jacobi, Sandie E, Foreman, Susan K, Pittman, Laura M, and Cornett, Lawrence E
- Subjects
Luminescent Proteins ,Genetic Vectors ,Green Fluorescent Proteins ,Cyclic AMP ,Gene Transfer Techniques ,Gene Expression ,Humans ,Receptors, Adrenergic, beta-2 ,Cells, Cultured ,Recombinant Proteins ,Research Article ,Adenoviridae ,Signal Transduction - Abstract
Beta2-adrenergic receptors (beta2AR) play important regulatory roles in a variety of cells and organ systems and are important therapeutic targets in the treatment of airway and cardiovascular disease. Prolonged use of beta-agonists results in tolerance secondary to receptor down-regulation resulting in reduced therapeutic efficiency. The purpose of this work is to evaluate the signaling capabilities of the beta2AR expressed by a recombinant adeno-associated viral (AAV) vector that also included an enhanced green fluorescent protein (EGFP) gene (AAV-beta2AR/EGFP).By epifluorescence microscopy, approximately 40% of infected HEK 293 cells demonstrated EGFP expression. beta2AR density measured with [3H]dihydroalprenolol ([3H]DHA) increased either 13- or 77-fold in infected cells compared to mock infected controls depending on the culture conditions used. The [3H]DHA binding was to a single receptor population with a dissociation constant of 0.42 nM, as would be expected for wild-type beta2AR. Agonist competition assays with [3H]DHA showed the following rank order of potency: isoproterenolepinephrinenorepinephrine, consistent with beta2AR interaction. Isoproterenol-stimulated cyclic AMP levels were 5-fold higher in infected cells compared to controls (314 +/- 43 vs. 63.4 +/- 9.6 nmol/dish; n = 3). Receptor trafficking demonstrated surface expression of beta2AR with vehicle treatment and internalization following isoproterenol treatment.We conclude that HEK 293 cells infected with AAV-beta2AR/EGFP effectively express beta2AR and that increased expression of these receptors results in enhanced beta2AR signaling. This method of gene transfer may provide an important means to enhance function in in vivo systems.
- Published
- 2003
4. Enhanced β2-adrenergic receptor (β2AR) signaling by adeno-associated viral (AAV)-mediated gene transfer.
- Author
-
Jones, Stacie M., Hiller, F. Charles, Jacobi, Sandie E., Foreman, Susan K., Pittman, Laura M., and Cornett, Lawrence E.
- Subjects
BETA adrenoceptors ,GENETIC transformation ,CELLS ,GREEN fluorescent protein ,GENE expression - Abstract
Background: β
2 -Adrenergic receptors (β2 AR) play important regulatory roles in a variety of cells and organ systems and are important therapeutic targets in the treatment of airway and cardiovascular disease. Prolonged use of β-agonists results in tolerance secondary to receptor down-regulation resulting in reduced therapeutic efficiency. The purpose of this work is to evaluate the signaling capabilities of the β2 AR expressed by a recombinant adeno-associated viral (AAV) vector that also included an enhanced green fluorescent protein (EGFP) gene (AAV-β2 AR/EGFP). Results: By epifluorescence microscopy, ~40% of infected HEK 293 cells demonstrated EGFP expression. β2 AR density measured with [³H]dihydroalprenolol ([³H]DHA) increased either 13- or 77-fold in infected cells compared to mock infected controls depending on the culture conditions used. The [3H]DHA binding was to a single receptor population with a dissociation constant of 0.42 nM, as would be expected for wild-type β2 AR. Agonist competition assays with [3H]DHA showed the following rank order of potency: isoproterenol>epinephrine> norepinephrine, consistent with β2 AR interaction. Isoproterenol-stimulated cyclic AMP levels were 5-fold higher in infected cells compared to controls (314 ± 43 vs. 63.4 ± 9.6 nmol/dish; n = 3). Receptor trafficking demonstrated surface expression of β2 AR with vehicle treatment and internalization following isoproterenol treatment. Conclusions: We conclude that HEK 293 cells infected with AAV-β2 AR/EGFP effectively express β2 AR and that increased expression of these receptors results in enhanced β2 AR signaling. This method of gene transfer may provide an important means to enhance function in in vivo systems. [ABSTRACT FROM AUTHOR]- Published
- 2003
5. The beta-agonist isoproterenol attenuates EGF-stimulated wound closure in human airway epithelial cells.
- Author
-
Schnackenberg BJ, Jones SM, Pate C, Shank B, Sessions L, Pittman LM, Cornett LE, and Kurten RC
- Subjects
- Bacterial Proteins genetics, Bacterial Proteins metabolism, Bronchi cytology, Bronchi metabolism, Cells, Cultured, Colforsin pharmacology, Cyclic AMP metabolism, Drug Combinations, Epithelial Cells metabolism, ErbB Receptors metabolism, Humans, Luminescent Proteins genetics, Luminescent Proteins metabolism, Mitogen-Activated Protein Kinases metabolism, Receptors, Adrenergic, beta-2 genetics, Receptors, Adrenergic, beta-2 metabolism, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Signal Transduction, Adrenergic beta-Agonists pharmacology, Bronchi drug effects, Epidermal Growth Factor pharmacology, Epithelial Cells drug effects, Isoproterenol pharmacology, Wound Healing drug effects
- Abstract
Asthma is a disease characterized by reversible airway obstruction. An additional hallmark of chronic asthma is altered wound healing that leads to airway remodeling. Although beta-agonists are effective in treating the bronchospasm associated with asthma, their effects on airway wound healing, which are related to airway remodeling, are unknown. It has been demonstrated that beta-agonists can alter the signaling of epidermal growth factor (EGF) receptors, which are important in timely wound healing. Therefore, we hypothesized that the beta-agonist isoproterenol would affect wound healing. Using an in vitro scrape wound assay, we demonstrated that isoproterenol attenuates EGF-stimulated wound healing in 16HBE airway epithelial cell cultures. Through experiments with forskolin and cells overexpressing beta2-adrenergic receptor-yellow fluorescent protein, we show that attenuation is due to the accumulation of cAMP and the involvement of at least one additional pathway. Furthermore, attenuation is not due to a direct effect on the EGF receptor or to an alteration of the ERK/MAPK signaling cascade. Based on these results, we propose that isoproterenol may exert its effects through other MAPK signaling pathways (JNK and/or p38) or through parallel mechanisms. These results also demonstrate a problem of potential therapeutic relevance in which a commonly prescribed medication may alter wound healing and contribute to the remodeling of asthmatic airways.
- Published
- 2006
- Full Text
- View/download PDF
6. Coordinating epidermal growth factor-induced motility promotes efficient wound closure.
- Author
-
Kurten RC, Chowdhury P, Sanders RC Jr, Pittman LM, Sessions LW, Chambers TC, Lyle CS, Schnackenberg BJ, and Jones SM
- Subjects
- Animals, Blood Proteins pharmacology, Cell Differentiation drug effects, Cells, Cultured, Chlorocebus aethiops, Epithelial Cells cytology, Fibroblasts cytology, Kidney cytology, Cell Movement drug effects, Epidermal Growth Factor pharmacology, Wound Healing drug effects
- Abstract
Wound healing is a response to injury that is initiated to reconstruct damaged tissue. In skin, reepithelialization involves both epithelial cells and fibroblasts and contributes to the reformation of a barrier between the external environment and internal milieu. Growth factors including epidermal growth factor (EGF) play important roles in promoting this process. In the present studies we employed CV-1 fibroblasts in a tissue culture model of reepithelialization to develop strategies for optimizing wound closure stimulated by EGF. We found that EGF enhanced cell motility within 6-8 h of EGF treatment in serum-free medium but wounds failed to close within 24 h. However, if medium on these cultures was exchanged for medium containing serum, cells pretreated with EGF closed new scrape wounds more rapidly than did cells that were not pretreated. These results indicate that serum factors work in concert with EGF to coordinate cell motility for efficient wound closure. Indeed, EGF enhanced the rate of wound closure in the presence of serum, and this effect also persisted for at least 24 h after EGF was removed. This coordination of EGF-induced cell motility was accompanied by an increase in the transient phosphorylation of ERK1 and ERK2. The persistent effects of EGF were blocked by transient exposure to reversible inhibitors of transcription and translation, indicating that the expression of new proteins mediated this response. We propose that EGF-stimulated CV-1 fibroblast motility is coordinated by a serum component that induces cell-cell adhesive properties consistent with an epithelial phenotype, thereby enhancing the reepithelialization process.
- Published
- 2005
- Full Text
- View/download PDF
7. Enhanced beta2-adrenergic receptor (beta2AR) signaling by adeno-associated viral (AAV)-mediated gene transfer.
- Author
-
Jones SM, Hiller FC, Jacobi SE, Foreman SK, Pittman LM, and Cornett LE
- Subjects
- Adenoviridae genetics, Cells, Cultured, Cyclic AMP metabolism, Gene Expression, Gene Transfer Techniques, Genetic Vectors genetics, Green Fluorescent Proteins, Humans, Luminescent Proteins metabolism, Recombinant Proteins metabolism, Receptors, Adrenergic, beta-2 physiology, Signal Transduction physiology
- Abstract
Background: Beta2-adrenergic receptors (beta2AR) play important regulatory roles in a variety of cells and organ systems and are important therapeutic targets in the treatment of airway and cardiovascular disease. Prolonged use of beta-agonists results in tolerance secondary to receptor down-regulation resulting in reduced therapeutic efficiency. The purpose of this work is to evaluate the signaling capabilities of the beta2AR expressed by a recombinant adeno-associated viral (AAV) vector that also included an enhanced green fluorescent protein (EGFP) gene (AAV-beta2AR/EGFP)., Results: By epifluorescence microscopy, approximately 40% of infected HEK 293 cells demonstrated EGFP expression. beta2AR density measured with [3H]dihydroalprenolol ([3H]DHA) increased either 13- or 77-fold in infected cells compared to mock infected controls depending on the culture conditions used. The [3H]DHA binding was to a single receptor population with a dissociation constant of 0.42 nM, as would be expected for wild-type beta2AR. Agonist competition assays with [3H]DHA showed the following rank order of potency: isoproterenol>epinephrine> norepinephrine, consistent with beta2AR interaction. Isoproterenol-stimulated cyclic AMP levels were 5-fold higher in infected cells compared to controls (314 +/- 43 vs. 63.4 +/- 9.6 nmol/dish; n = 3). Receptor trafficking demonstrated surface expression of beta2AR with vehicle treatment and internalization following isoproterenol treatment., Conclusions: We conclude that HEK 293 cells infected with AAV-beta2AR/EGFP effectively express beta2AR and that increased expression of these receptors results in enhanced beta2AR signaling. This method of gene transfer may provide an important means to enhance function in in vivo systems.
- Published
- 2003
- Full Text
- View/download PDF
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.