Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Although several studies indicated that parathyroid hormone (PTH) exerted anabolic action on bone, its precise mechanisms have been unknown. On the other hand, transforming growth factor beta (TGF-beta), abundantly stored in bone matrix, stimulates bone formation with a local injection in rodents. Although our previous study suggested that Smad3 is an important molecule for the stimulation of bone formation, no reports have been available about the effects of PTH on Smad3. In this present study, we examined the effects of PTH on Smad3 and the physiological significance in mouse osteoblastic cells. PTH promoted the expression of Smad3 mRNA within 10 min and the protein level in a dose-dependent manner in MC3T3-E1 and rat osteoblastic UMR-106 cells. Protein kinase A (PKA) activator as well as protein kinase C (PKC) activators increased Smad3 protein level, and both PKA and PKC inhibitors antagonized PTH-induced Smad3, indicating that PTH promotes the production of Smad3 through both PKA and PKC pathways. Next, we examined anti-apoptotic effects of PTH and Smad3 in these cells, employing trypan blue, transferase-mediated nick end labeling, and Hoechst staining. Pretreatment with PTH or overexpression of Smad3 decreased the number of apoptotic cells induced by dexamethasone and etoposide. Moreover, a dominant negative mutant, Smad3DeltaC, abrogated PTH-induced anti-apoptotic effects. On the other hand, PTH augmented TGF-beta-induced transcriptional activity. Furthermore, PTH enhanced TGF-beta-induced production of type I collagen, whereas it did not affect TGF-beta-reduced proliferation in MC3T3-E1 cells. These observations indicated that PTH amplified the anabolic effects of TGF-beta by accelerating the transcriptional activity of Smad3. In conclusion, we first demonstrated that PTH-Smad3 axis exerts anti-apoptotic effects in osteoblasts and reinforces the anabolic action by TGF-beta in osteoblasts. Hence, PTH-Smad3 axis might be involved in the bone anabolic action of PTH.
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PMID:Parathyroid hormone-Smad3 axis exerts anti-apoptotic action and augments anabolic action of transforming growth factor beta in osteoblasts. 1451 10

Chondrocytes undergo phenotypic alterations following extended periods in monolayer culture, i.e., they become bipolar and flattened, proliferate, and synthesise type I as opposed to type II collagen. This process has been termed chondrocyte dedifferentiation. Bistratene A is a macrolide polyether that specifically activates the delta isoform of protein kinase C (PKCdelta) in some cell types. Here, we show that dedifferentiated human articular chondrocytes became rounded and underwent cell growth arrest after treatment with bistratene A. In addition, bistratene A-treated chondrocytes became more immunopositive for type II collagen, but less immunopositive for type I collagen. These phenotypic changes were associated with a prior and extensive disruption of actin microfilaments and translocation of PKCdelta to the nuclear membrane. Concurrent treatments of chondrocytes with a specific inhibitor of PKCdelta, rottlerin, partially blocked the morphological effects of bistratene A.
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PMID:Phenotypic modulation of human articular chondrocytes by bistratene A. 1456 53

The role of protein kinase C (PKC) in the regulation of contraction has been controversial. Recently, CPI-17, a PKC-potentiated inhibitor protein of PP1, has been cloned and shown to be specifically expressed in SMC. In this study, we over-expressed CPI-17 and its mutants in NIH3T3 cells, which do not express CPI-17, and examined its effect on the contractile property. For the measurement of tension, NIH3T3 cells were collected by trypsinization, mixed with type I collagen and made into a ring preparation (reconstituted ring: RR). The isometric tension developments were measured using this RR and a force transducer. The application of phorbol dibutyrate (PDBu; 0.3 microM) to RR transfected by CPI-17 induced a contraction that reached 2-3 times greater that the 10% FBS-induced contraction, while PDBu relaxed the RR transfected with vector alone (control) or CPI-17 mutants (T38A and T38E). The PDBu-induced contraction of the CPI-17 transfected RR could be inhibited by 3 microM GF109203X (PKC inhibitor) but not by 3 microM Y27632 (rho kinase inhibitor). The application of PDBu during contraction induced by 1 microM bradykinin induced further contraction in CPI-17 transfected RR, while it induced a complete relaxation in control RR. These results indicated that CPI-17 is a molecular switch that reverses the PKC mediated effect on contraction. The limited expression of CPI-17 to SMC may explain the previous observation that the effects of PKC stimulation were quite different in SMC from those of other cell types.
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PMID:[Changes in the contractility of the NIH3T3 fibroblast induced by the over-expression of CPI-17]. 1472 19

In long-term space travel, the crew is exposed to microgravity and radiation that invoke potential hazards to the immune system. T cell activation is a critical step in the immune response. Receptor-mediated signaling is inhibited in both microgravity and modeled microgravity (MMG) as reflected by diminished DNA synthesis in peripheral blood lymphocytes and their locomotion through gelled type I collagen. Direct activation of protein kinase C (PKC) bypassing cell surface events using the phorbol ester PMA rescues MMG-inhibited lymphocyte activation and locomotion, whereas the calcium ionophore ionomycin had no rescue effect. Thus calcium-independent PKC isoforms may be affected in MMG-induced locomotion inhibition and rescue. Both calcium-dependent isoforms and calcium-independent PKC isoforms were investigated to assess their expression in lymphocytes in 1 g and MMG culture. Human lymphocytes were cultured and harvested at 24, 48, 72, and 96 h, and serial samples were assessed for locomotion by using type I collagen and expression of PKC isoforms. Expression of PKC-alpha, -delta, and -epsilon was assessed by RT-PCR, flow cytometry, and immunoblotting. Results indicated that PKC isoforms delta and epsilon were downregulated by >50% at the transcriptional and translational levels in MMG-cultured lymphocytes compared with 1-g controls. Events upstream of PKC, such as phosphorylation of phospholipase Cgamma in MMG, revealed accumulation of inactive enzyme. Depressed calcium-independent PKC isoforms may be a consequence of an upstream lesion in the signal transduction pathway. The differential response among calcium-dependent and calcium-independent isoforms may actually result from MMG intrusion events earlier than PKC, but after ligand-receptor interaction.
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PMID:Modeled microgravity-induced protein kinase C isoform expression in human lymphocytes. 1496 22

PKC is a family of 12 serine/threonine isoenzymes that plays a pivotal role in signal transduction in a large number of biological processes. In the present work we have investigated the expression of PKC (alpha, delta, epsilon, zeta) in chick chondrocyte primary cultures at different differentiation times, i.e. at 48, 55, 62 and 69 days after cell collection from tibiae of 6-day old chick embryos. We would also detect cell differentiation stages towards the osteoblast-like cell phenotype by observing the immunocytochemical expression of the specific osteoblast marker, type I collagen. At the considered culture times, cells exhibited immunocytochemical positivity for type I collagen, thus showing their differentiation towards the osteoblast-like phenotype. PKC-zeta was the isoenzyme that exhibited the most relevant immunocytochemical expression in all considered culture times, whereas PKC-epsilon always less expressed in comparison to the other PKC-isoforms. No relevant differences were observed for the immunocytochemical expressions of PKC-alpha and PKC-delta. On the basis of the immunocytochemical data obtained from the present investigation, we could affirm that PKC-alpha, -delta, -epsilon, and -zeta may play peculiar roles in the differentiation process of chick chondrocytes towards the osteoblast-like cell phenotype.
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PMID:Expression of protein kinase C (PKC) alpha, delta, epsilon, zeta in primary chick chondrocyte cultures: immunocytochemical study. 1514 76

The role of endothelin-1 (ET) in tissue remodeling/fibrogenesis has been demonstrated in various in vitro and in vivo models. Our previous studies have revealed ET-induced expression of type I collagen in cardiac myofibroblasts (myoFb). Here we report that protein kinase Cdelta (PKCdelta) and mitogen-activated protein kinase/extracellular signal-regulated kinase-1/2 (MAPK/ERK1/2) play a role in ET-induced type I collagen expression using specific pharmacological inhibitors. The present study also reveals the expression of various isoforms of PKC including PKCalpha, PKCbetaI, PKCbetaII, PKCgamma, PKCdelta, PKCepsilon, PKCeta, and PKCzeta in cardiac myoFb. Our results from mRNA and protein studies demonstrate that calphostin-C, a PKC inhibitor, decreased the ET-induced type I collagen expression suggesting a role for the PKC pathway. Further treatment with rottlerin, a PKCdelta isoform-specific inhibitor, demonstrated attenuation of 80 to 90% of type I collagen expression induced by ET. However, Go6976 [12-(2-cyanoethyl)-6,7,12,13-tetrahydro-13-methyl-5-oxo-5H-indolo[2,3-a]pyrrolo [3,4-c]carbazole]], an inhibitor of Ca(2+)-dependent PKC isoforms (PKCalpha and PKCbetaI), showed little to no effect on ET-stimulated type I collagen expression. Furthermore, the MAPK inhibitor PD98059 (2'-amino-3'-methoxyflavone) attenuated ET-dependent activation of p44/42 MAPK (pERK1/2) and also down-regulated type I collagen expression. Similarly, rottlerin inhibited the activation of p44/42 MAPK (pERK) implicating the involvement of PKC and MAPK/ERK1/2 in ET-induced type I collagen expression. Our protein/DNA array and reverse transcription-polymerase chain reaction results from ET-treated samples showed a significant increase in Sp1 expression. PD98059 and rottlerin decreased ET-induced Sp1 expression, suggesting a possible interaction of Sp1 with PKCdelta and MAPK in ET-induced type I collagen expression in cardiac myoFb.
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PMID:Role of protein kinase Cdelta in endothelin-induced type I collagen expression in cardiac myofibroblasts isolated from the site of myocardial infarction. 1524 Aug 25

Interactions of integrin cellular adhesion molecules with matrix proteins play important roles in complex bidirectional signaling pathways. To investigate these interactions, a novel flow-cytometry-based cellular adhesion assay has been developed. Based on the concept of microcarrier cell culture, derivatized polystyrene microspheres (9.6 microm) are used as a substrate for the immobilization of type I collagen to which cells then adhere. Using cytometric detection, the extent of cellular adhesion can be precisely determined by comparison of adhered and nonadhered populations based on the side scatter properties of the microspheres. In combination with immunostaining, the novel format of this assay enables the correlation of adhesive function to other cellular characteristics such as surface expression. In this work, the protein kinase C activator 12-O-tetradecanoylphorbol-13-acetate (TPA) was used to stimulate increased adhesion in Chinese hamster ovary cells stably transfected with the collagen receptor integrin alpha2beta1. Multiple clones of varying expression distributions were analyzed, and correlations of adherent populations versus receptor distributions show a threefold increase in functional cellular adhesion to collagen upon treatment with TPA. Probability binning analysis of duplexed data revealed subtle changes in adhesion versus receptor distribution mediated by TPA which otherwise would not have been detectable.
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PMID:A duplexed microsphere-based cellular adhesion assay. 1569 4

SIP24 is an acute phase iron binding lipocalin physiologically expressed in vivo in developing cartilage by prehypertrophic/hypertrophic chondrocytes. Taking advantage of the chondrocytic cell line MC615 and using SIP24 as a marker we investigated the pathways active in cartilage differentiation and inflammation. MC615 cells were cultured as: (i) proliferating prechondrogenic cells expressing type I collagen (ii) differentiated hyperconfluent cells expressing Sox9 and type II collagen. In proliferating cells the pathway PKC/ERK1, ERK2 was activated and SIP24 was not expressed while in differentiated cells the pathway p38/NF-kappaB was activated and SIP24 was expressed. Proliferating cells treated with inflammatory agents expressed a large amount of SIP24 and showed activation of p38/NF-kappaB pathway and inhibition of PKC/ERK1, ERK2 pathway indicating that in inflammation and differentiation the same factors are activated (p38, NF-kappaB) or inactivated (PKC, ERKs). Treatment of proliferating cells with the p38 specific inhibitor SB203580 inhibited the inflammation induced activation of p38 and the synthesis of SIP24. PMA treatment induced activation of PKC, inactivation of p38 and suppression of SIP24 synthesis, suggesting that PKC activation inhibits p38 activation. In differentiated hyperconfluent cells the same factors (p38/NF-kappaB/SIP24) are constitutively activated: treatment with inflammatory agents does not increase synthesis of SIP24 while treatment with SB203580 and with PMA does not repress activation of p38 nor synthesis of SIP24. We propose that the SIP24 stress related protein is expressed via p38 activation/NF-kappaB recruitment both in chondrocyte differentiation and inflammation and that a signaling pathway active in the acute phase response is physiologically activated in differentiation.
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PMID:A common pathway in differentiation and inflammation: p38 mediates expression of the acute phase SIP24 iron binding lipocalin in chondrocytes. 1622 8

Retinal pigment epithelial cells (RPEs) are thought to be one of the main components of fibrous membrane observed in eyes with proliferative vitreo-retinopathy. We investigated the signalling mechanisms of TGF-beta2-dependent collagen gel contraction by RPEs. An in vitro type I collagen gel contraction assay was performed to evaluate the effect of TGF-beta2 on gel contraction. The expression of alpha-smooth muscle actin (alpha-SMA) and the phosphorylation state of myosin light chain (MLC) were analyzed by Western blotting. The involvement of protein kinases such as p44/42 mitogen-activated protein kinase (MAPK), protein kinase C (PKC), p38 MAPK and phosphatidylinositol-3 kinase was investigated. The contribution of Rho-kinase and/or MLC-kinase was also evaluated using respective kinase inhibitors (Y27632, hydroxyfasudil and ML7). Additionally, RPEs were immunostained to examine whether the expression of alpha-SMA detected in our western blotting correlated to the stress fiber formation within the cells. TGF-beta2 caused time (0-5 days)-and dose (0 10 ng ml(-1))-dependent gel contraction associated with overexpression of alpha-SMA and phosphorylation of MLC (p < 0.01, respectively). PKC inhibitor (GF109203X, 5 microM) and p38 MAPK inhibitor (SB203580, 10 microM) significantly attenuated TGF-beta2-elicited gel contraction via partial downregulation of both alpha-SMA expression and MLC phosphorylation (p < 0.01, respectively). The gel contraction was prominently inhibited in the presence of Y27632 (10 microM) or hydroxyfasudil (10 microM) with strong suppression of MLC phosphorylation but had no significant effect on alpha-SMA expression. Treatment with ML7, in contrast, resulted in a marginal inhibition of MLC phosphorylation and gel contraction. Finally, pretreatment of the cells with Y27632 or hydroxyfasudil prevented the formation of stress fiber within the cells. These results indicate that TGF-beta2-dependent myofibroblastic transdifferentiation and MLC phosphorylation by RPEs involve both PKC and p38 MAPK pathways at least in part. Myofibroblastic transdifferentiation of RPEs appears to be independent of the Rho-kinase pathway, and the presence of alpha-SMA does not necessarily reflect the contractile potential of a cell. While Rho-kinase inhibitors are incapable of preventing myofibroblastic transdifferentiation itself, this pathway could be one of the critical targets of cell-mediated contraction of the tissue containing fibrillar collagens by transdifferentiated RPEs.
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PMID:Critical role of the Rho-kinase pathway in TGF-beta2-dependent collagen gel contraction by retinal pigment epithelial cells. 1631 Jan 90

Hypoxia promotes keratinocyte migration on wound bed connective tissues and is a profound biological signal that transforms a basal keratinocyte, destined to differentiate, into a motile cell that is essential for re-epithelialization. In this study, we examined the effect of hypoxia on keratinocyte-derived collagenases associated with keratinocyte migration. Cells plated on various connective tissue matrices under normoxic and hypoxic conditions, demonstrated a two-fold increase in the 92 kDa, type IV collagenase (MMP-9) when examined by quantitative zymography and ELISA. Western blotting and ELISA demonstrated a two-fold increase in tissue inhibitor of metalloproteinase (TIMP-1), an enzyme that binds to MMP-9 and inhibits its activity. The hypoxia-induced increase in cell motility could be inhibited by a neutralizing antibody to MMP-9. Northern blotting demonstrated that MMP-9 and TIMP-1 mRNA increased 2.5- to 4-fold, 2-12 h after the cells were made hypoxic. The hypoxia-induced changes in MMP-9 and TIMP-1 were inhibited by staurosporine and bisindolylmaleimide, inhibitors of protein kinase C (PKC), but not by inhibitors of tyrosine phosphorylation and the mitogen-activated protein kinase pathway. Inhibition of PKC also inhibited hypoxia-induced keratinocyte migration on type I collagen. These data provide evidence that hypoxia-induced keratinocyte migration is mediated by increased cellular secretion of MMP-9 via the PKC pathway.
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PMID:Hypoxia induces epidermal keratinocyte matrix metalloproteinase-9 secretion via the protein kinase C pathway. 1755 70


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