Cell Volume Regulation in Response to Hypotonicity Is Impaired in HeLa Cells Expressing a Protein Kinase C α Mutant Lacking Kinase Activity

Hermoso, Marcela A.; Olivero, Pablo; Torres, Rubén; Riveros, Ana; Quest, Andrew F. G.; Stutzin, Andrés

doi:10.1074/jbc.m304506200

Cited by 25 publications

(26 citation statements)

References 52 publications

(61 reference statements)

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“…The reduced PKC activity was associated with decreased ganglion cell size, which may possibly be related to distal fiber atrophy in this model, as demonstrated in our previous studies (11). A relationship of the reduced PKC-␣ expression with the ganglion cell atrophy accords with the recent data that PKC-␣ is crucial for the regulation of slow axonal transport (25) as well as the maintenance of cell size in an altered osmotic environment, a condition that may occur in hyperglycemic milieu (26). The ganglion cell atrophy is common with other diabetic animal models (27,28) and may be closely related to metabolic imbalance through polyol pathway hyperactivity as well as other metabolic cascade (29).…”

Section: Discussionsupporting

confidence: 88%

Effects of Polyol Pathway Hyperactivity on Protein Kinase C Activity, Nociceptive Peptide Expression, and Neuronal Structure in Dorsal Root Ganglia in Diabetic Mice

et al. 2004

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We explored the specific impact of polyol pathway hyperactivity on dorsal root ganglia (DRG) using transgenic mice that overexpress human aldose reductase because DRG changes are crucial for the development of diabetic sensory neuropathy. Littermate mice served as controls. Half of the animals were made diabetic by streptozotocin injection and followed for 12 weeks. After diabetes onset, diabetic transgenic mice showed a significant elevation of pain sensation threshold after transient decrease and marked slowing of motor and sensory nerve conduction at the end of the study, while these changes were modest in diabetic littermate mice. Protein kinase C (PKC) activities were markedly reduced in diabetic transgenic mice, and the changes were associated with reduced expression of membrane PKC-␣ isoform that was translocated to cytosol. Membrane PKC-␤II isoform expression was contrariwise increased. Calcitonin gene-related peptide-and substance P-positive neurons were reduced in diabetic transgenic mice and less severely so in diabetic littermate mice. Morphometric analysis disclosed neuronal atrophy only in diabetic transgenic mice. Treatment with an aldose reductase inhibitor (fidarestat 4 mg ⅐ kg -1 ⅐ day -1, orally) corrected all of the changes detected in diabetic transgenic mice. These findings underscore the pathogenic role of aldose reductase in diabetic sensory neuropathy through the altered cellular signaling and peptide expressions in DRG neurons. Diabetes 53:3239 -3247, 2004 P olyol pathway hyperactivity has been extensively studied for the mechanisms of diabetic neuropathy, where aldose reductase is a key regulating enzyme (1,2). In animal models, diabetes-induced peripheral nerve conduction deficits, neurometabolic imbalances, altered nerve blood flow, and morphologic abnormalities are prevented by structurally diverse aldose reductase inhibitors (ARIs) (3-5). Past clinical trials of ARIs were not, however, convincingly successful (6,7), and specific effects of polyol pathway hyperactivity on the clinical and structural aspects of diabetic sensory neuropathy is yet to be clear (8,9). The development of a transgenic animal model provides an ideal tool to identify the specific role of single molecules in disease mechanisms. We have established a strain of transgenic mice that overexpress human aldose reductase (10 -12). In this model, we confirmed that polyol pathway hyperactivity was indeed related to the severity of motor nerve conduction delay and nerve fiber atrophy. More recently, a new transgenic model that overexpresses aldose reductase, specifically in Schwann cells by use of myelin protein Po promoter, has enabled researchers to address more precise mechanisms of polyol pathway in the development of neuropathy in diabetes (13). Nevertheless, from these studies, it was not shown which measures were comparable with those found in human diabetic neuropathy, and it still remains obscure what could be the most appropriate target for the intervention with a potent and specific inhibitor for human aldos...

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Section: Discussionsupporting

confidence: 88%

Effects of Polyol Pathway Hyperactivity on Protein Kinase C Activity, Nociceptive Peptide Expression, and Neuronal Structure in Dorsal Root Ganglia in Diabetic Mice

et al. 2004

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“…We also examined the potential role of PKC because this enzyme is also activated by H 2 O 2 (43,44) and has been shown to regulate VRAC activity (26,45). The potent PKC inhibitors 1 M Go6983, 1 M bisindolylmaleimide I, and 5 M Ro32-0432 (46 -48) were ineffective in blocking the H 2 O 2 -enhanced EAA release during cell swelling (Fig.…”

Section: Hydrogen Peroxide Potentiatesmentioning

confidence: 99%

Hydrogen Peroxide Potentiates Volume-sensitive Excitatory Amino Acid Release via a Mechanism Involving Ca2+/Calmodulin-dependent Protein Kinase II

Haskew-Layton

Mongin

Kimelberg³

2005

Journal of Biological Chemistry

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“…Among conventional isoforms, expression of , I, II, but not has been detected (Slepko et al 1999). Since two previous studies implicated PKC in VRAC regulation (Roman et al 1998;Hermoso et al 2004), we initially used a commercially available mammalian plasmid pKD-PKC -v4 expressing shRNA that targets PKC (Millipore). In these experiments, PKC protein levels were reduced by ~70%, while the ATP-induced regulation of D-[ 3 H] aspartate release was completely unaffected (Supplemental figure 1, n=5, p>0.9 for both maximal and integral release values).…”

Section: Functional Effects Of Sirna-mediated Downregulation Of the Pmentioning

confidence: 99%

“…In HeLa cells, HTC rat hepatoma cells, and Mz-ChA-1 human cholangiocarcinoma cells, PKC activity was found to be critical for hypotonic (i.e. non-GPCR-related) activation of VRAC (Roman et al 1998;Hermoso et al 2004). The study by A. Stutzin and colleagues (Hermoso et al 2004) employed the dominant-negative PKC expression.…”

Section: Conventional Pkc Isoforms Are Major Contributors To Vrac Regmentioning

confidence: 99%

“…non-GPCR-related) activation of VRAC (Roman et al 1998;Hermoso et al 2004). The study by A. Stutzin and colleagues (Hermoso et al 2004) employed the dominant-negative PKC expression. The authors found that DN-PKC blocks swelling activated Cl − conductance and suppresses the process of regulatory volume decrease under hypoosmotic conditions.…”

Section: Conventional Pkc Isoforms Are Major Contributors To Vrac Regmentioning

confidence: 99%

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Two conventional protein kinase C isoforms, α and βI, are involved in the ATP‐induced activation of volume‐regulated anion channel and glutamate release in cultured astrocytes

Rudkouskaya

Chernoguz

Haskew-Layton

et al. 2008

Journal of Neurochemistry

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Volume-regulated anion channels (VRACs) are activated by cell swelling and are permeable to inorganic and small organic anions, including the excitatory amino acids glutamate and aspartate. In astrocytes, ATP potently enhances VRAC activity and glutamate release via a P2Y receptordependent mechanism. Our previous pharmacological study identified protein kinase C (PKC) as a major signaling enzyme in VRAC regulation by ATP. However, conflicting results obtained with potent PKC blockers prompted us to re-evaluate PKC involvement in regulation of astrocytic VRAC using siRNA and pharmacological inhibitors that selectively target individual PKC isoforms. In primary rat astrocyte cultures, application of hypoosmotic medium (30% reduction in osmolarity) and 20 M ATP synergistically increased the release of excitatory amino acids, measured with non-metabolized analogue of L-glutamate, D-[ 3 H]aspartate. Both Go6976, the selective inhibitor of Ca 2+ -sensitive PKC , I/II, and , and MP-20-28, a cell permeable pseudosubstrate inhibitory peptide of PKC and I/II, reduced the effects of ATP on D-[ 3 H]aspartate release by ~45-55%. Similar results were obtained with a mixture of siRNAs targeting rat PKC and I. Surprisingly, downregulation of individual and I PKC isozymes by siRNA was completely ineffective. These data suggest that ATP regulates VRAC activity and volume-sensitive excitatory amino acid release via cooperative activation of PKC and I.

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Cell Volume Regulation in Response to Hypotonicity Is Impaired in HeLa Cells Expressing a Protein Kinase C α Mutant Lacking Kinase Activity

Cited by 25 publications

References 52 publications

Effects of Polyol Pathway Hyperactivity on Protein Kinase C Activity, Nociceptive Peptide Expression, and Neuronal Structure in Dorsal Root Ganglia in Diabetic Mice

Effects of Polyol Pathway Hyperactivity on Protein Kinase C Activity, Nociceptive Peptide Expression, and Neuronal Structure in Dorsal Root Ganglia in Diabetic Mice

Hydrogen Peroxide Potentiates Volume-sensitive Excitatory Amino Acid Release via a Mechanism Involving Ca2+/Calmodulin-dependent Protein Kinase II

Two conventional protein kinase C isoforms, α and βI, are involved in the ATP‐induced activation of volume‐regulated anion channel and glutamate release in cultured astrocytes

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