A Metabolic View of Receptor Activation in Cultured Cells Following Cryopreservation

Kuo, Richard C.; Baxter, Gregory T.; Alajoki, Liisa; Miller, Donald L.; Libby, Jeffrey M.; Owicki, John C.

doi:10.1006/cryo.1993.1038

Cited by 6 publications

(4 citation statements)

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“…It should be noted that high correlations have been reported between microphysiometric responses to muscarinic agonists and antagonists in muscarinic receptor transfected cell systems and induced changes in radioligand binding (Wood et al, 1999), Ca 21 mobilization measured using a fluorescent Ca 21 ligand (Ikeda et al, 1999), and phosphoinositide hydrolysis (Baxter et al, 1994). Nonetheless, the results of the following microphysiometric surveys of eigenvector template-generated peptide influences on receptor-mediated function in m 1 AChR and m 2 AChR transfected CHO-K1 cells (Kuo et al, 1993) are to be considered preliminary, awaiting the results of further studies of less ''downstream'' indices of second messenger responses.…”

Section: Locating Putative Allosteric Sites By Representing the M 1 Achr's Leading Eigenvector Modes As Modular Densities In The Morlet Wmentioning

confidence: 90%

“…Muscarinic cholinergic ligand-(carbachol and/or oxotremorine) induced changes in the m 1 and m 2 CHO-K1 cells' IEAR, in the presence and absence of eigenvector templatedesigned peptides, were determined using well-established microphysiometric functional screening methods for m 1 , m 2 , and other muscarinic receptor transfected cell lines (Baxter et al, 1994;Ikeda et al, 1999;Kuo et al, 1993;McConnell et al, 1992). Receptor-mediated changes in overall cellular glycolytic and respiratory metabolism and/or cellular membrane Na 1 /H 1 exchanges indicate receptor-mediated cellular activation, and are reflected in changes in extracellular protonic ([H 1 ]) concentrations (McConnell et al, 1992).…”

Section: Locating Putative Allosteric Sites By Representing the M 1 Achr's Leading Eigenvector Modes As Modular Densities In The Morlet Wmentioning

confidence: 99%

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Designing Human m1 Muscarinic Receptor-Targeted Hydrophobic Eigenmode Matched Peptides as Functional Modulators

Selz

Mandell

Shlesinger

et al. 2004

Biophysical Journal

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A new proprietary de novo peptide design technique generated ten 15-residue peptides targeting and containing the leading nontransmembrane hydrophobic autocorrelation wavelengths, "modes", of the human m(1) muscarinic cholinergic receptor, m(1)AChR. These modes were also shared by the m(4)AChR subtype (but not the m(2), m(3), or m(5) subtypes) and the three-finger snake toxins that pseudoirreversibly bind m(1)AChR. The linear decomposition of the hydrophobically transformed m(1)AChR amino acid sequence yielded ordered eigenvectors of orthogonal hydrophobic variational patterns. The weighted sum of two eigenvectors formed the peptide design template. Amino acids were iteratively assigned to template positions randomly, within hydrophobic groups. One peptide demonstrated significant functional indirect agonist activity, and five produced significant positive allosteric modulation of atropine-reversible, direct-agonist-induced cellular activation in stably m(1)AChR-transfected Chinese hamster ovary cells, reflected in integrated extracellular acidification responses. The peptide positive allosteric ligands produced left-shifts and peptide concentration-response augmentation in integrated extracellular acidification response asymptotic sigmoidal functions and concentration-response behavior in Hill number indices of positive cooperativity. Peptide mode specificity was suggested by negative crossover experiments with human m(2)ACh and D(2) dopamine receptors. Morlet wavelet transformation of the leading eigenvector-derived, m(1)AChR eigenfunctions locates seven hydrophobic transmembrane segments and suggests possible extracellular loop locations for the peptide-receptor mode-matched, modulatory hydrophobic aggregation sites.

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Section: Locating Putative Allosteric Sites By Representing the M 1 Achr's Leading Eigenvector Modes As Modular Densities In The Morlet Wmentioning

confidence: 90%

Section: Locating Putative Allosteric Sites By Representing the M 1 Achr's Leading Eigenvector Modes As Modular Densities In The Morlet Wmentioning

confidence: 99%

Designing Human m1 Muscarinic Receptor-Targeted Hydrophobic Eigenmode Matched Peptides as Functional Modulators

Selz

Mandell

Shlesinger

et al. 2004

Biophysical Journal

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“…Briefly, TF-1 cells were collected from cultures of either 5.0 ϫ 10 3 to 1.0 ϫ 10 4 cells/ml (log growth) or 1.0 -1.2 ϫ 10 6 cells/ml (stationary). Approximately 2.5 ϫ 10 5 cells were confined between two microporous polycarbonate membranes separated by a 50-m-thick annular spacer immobilized within a fibrin clot described previously (20). The microporous membranes and annular spacer form a disk-shaped chamber of 6 mm diameter and 50 m height (volume, ϳ2 l) that, together with the fibrin immobilizing reagent, traps cells during the fluid perfusion of microphysiometer experiments.…”

Section: Methodsmentioning

confidence: 99%

Tumor Necrosis Factor-α Mediates Both Apoptotic Cell Death and Cell Proliferation in a Human Hematopoietic Cell Line Dependent on Mitotic Activity and Receptor Subtype Expression

Baxter

Kuo

Jupp

et al. 1999

Journal of Biological Chemistry

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The TF-1 human erythroleukemic cell line exhibits opposing physiological responses toward tumor necrosis factor-␣ (TNF) treatment, dependent upon the mitotic state of the cells. Mitotically active cells in log growth respond to TNF by rapidly undergoing apoptosis whereas TNF exposure stimulates cellular proliferation in mitotically quiescent cells. The concentrationdependent TNF-induced apoptosis was monitored by cellular metabolic activity and confirmed by both DNA epifluorescence and DNA fragmentation. Moreover, these responses could be detected by measuring extracellular acidification activity, enabling rapid prediction (within ϳ 1.5 h of TNF treatment) of the fate of the cell in response to TNF. Growth factor resupplementation of quiescent cells, resulting in reactivation of cell cycling, altered TNF action from a proliferative stimulus to an apoptotic signal. Expression levels of the type II TNF receptor subtype (p75TNFR) were found to correlate with sensitivity to TNF-induced apoptosis. Pretreatment of log growth TF-1 cells with a neutralizing antip75TNFR monoclonal antibody inhibited TNF-induced apoptosis by greater than 80%. Studies utilizing TNF receptor subtype-specific TNF mutants and neutralizing antisera implicated p75TNFR in TNF-dependent apoptotic signaling. These data show a bifunctional physiological role for TNF in TF-1 cells that is dependent on mitotic activity and controlled by the p75TNFR.Cells have the capability of responding to a multitude of signals that it encounters in its extracellular environment. One such signal with widespread pleiotropic actions is the cytokine tumor necrosis factor-␣ (TNF) 1 (1). TNF has been shown to modulate proliferation, differentiation, and apoptotic or necrotic cell death in a number of different cell types (2-4). These disparate responses to TNF are mediated by TNF binding to specific cell surface receptors. Two distinct TNF receptors, type I (p55TNFR) and type II (p75TNFR) (M r 55,000 -60,000 and 70,000 -80,000 in human cells, respectively), have been identified (5, 6), although it remains unclear which of the many responses reported for TNF can be attributed to a specific receptor subtype (4). Moreover, the precise signal transduction pathways for each of these receptor subtypes have yet to be fully delineated. One action of TNF, the induction of apoptosis, is characterized by a discrete set of cellular events regulated by gene expression (7,8). The physiological events accompanying apoptosis include condensation of the chromatin, degradation of DNA through the activation of endogenous nucleases, and dissolution of the cell into small membrane-bound apoptotic vesicles (9, 10). In vivo, these vesicles are phagocytosed by macrophages or other phagocytic cells. Cell death by apoptosis is essential in many physiological processes, including embryonic development of the nervous system (11), oncogenic pathology (12), and clonal selection of hematopoietic cells (13).Conversely, TNF has also been shown to stimulate cellular proliferation in a variety of systems...

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“…The determination of EAR by microphysiometry involves a proton sensitive, photo current driven silicon semiconductor sensor that measures changes in the EAR resulting from effectorevoked alterations in cellular glycolytic and respiratory energy metabolism and/or alterations in sodium-hydrogen exchanges across cellular membranes (Kuo et al, 1993;McConnell et al, 1992;Neve et al, 1992). Protonic H + neutralizes the charge on the surface of the semiconductor, reducing the produced photocurrent at a rate linearly related to H + production.…”

Section: Brief Description Of Representative Studies Demonstrating Thmentioning

confidence: 99%

Algorithmically designed peptides ameliorate behavioral defects in animal model of ADHD by an allosteric mechanism

Kinkead

Selz

Owens

et al. 2006

Journal of Neuroscience Methods

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A Metabolic View of Receptor Activation in Cultured Cells Following Cryopreservation

Cited by 6 publications

References 0 publications

Designing Human m1 Muscarinic Receptor-Targeted Hydrophobic Eigenmode Matched Peptides as Functional Modulators

Designing Human m1 Muscarinic Receptor-Targeted Hydrophobic Eigenmode Matched Peptides as Functional Modulators

Tumor Necrosis Factor-α Mediates Both Apoptotic Cell Death and Cell Proliferation in a Human Hematopoietic Cell Line Dependent on Mitotic Activity and Receptor Subtype Expression

Algorithmically designed peptides ameliorate behavioral defects in animal model of ADHD by an allosteric mechanism

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