Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P00750 (PLA)
 16,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In the present study we examined the in vitro regulation of plasminogen activator inhibitor I (PAI-1) expression in peritubular cells recovered from 20-day-old rat testes. We tested two growth factors, basic fibroblast growth factor (bFGF) and transforming growth factor-alpha (TGF alpha). They are synthesized by Sertoli cells, and peritubular cells exhibit the corresponding high affinity receptors. After exposure to bFGF or TGF alpha (0.1-30 ng/ml), PAI-1 messenger RNA levels, as determined by Northern hybridization analysis, increased in a dose-dependent manner. The first significant effects were noted after 2-h exposure to bFGF or TGF alpha (10 ng/ml), and PAI-1 messenger RNA levels were maximally stimulated approximately 12-fold (bFGF) and 8-fold (TGF alpha) after 4 h. The two growth factors increased the amount of immunoreactive (Western blots) and biologically active (Stachrom) PAI-1 measured in the culture medium. Actinomycin D inhibited the effects of these factors, whereas cycloheximide augmented them. Phorbol myristate acetate, an activator of protein kinase C, mimicked the effects of bFGF and TGF alpha. Interestingly, long term (24-h) pretreatment with phorbol myristate acetate resulted in a severe loss of responsiveness to bFGF or TGF alpha. Staurosporine, an inhibitor of protein kinase C, also significantly reduced the effects of bFGF and TGF alpha. Given that PAI-1 inhibits Sertoli cell plasminogen activator activity and that bFGF and TGF alpha are synthesized by Sertoli cells, these factors are likely to interact to regulate protease activity in localized regions of the seminiferous tubule.
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PMID:Plasminogen activator inhibitor-1 regulation in cultured rat peritubular cells by basic fibroblast growth factor and transforming growth factor-alpha. 882 83

We analysed how interactions between protein kinase-dependent intracellular signalling pathways were implicated in the control of the production of tissue-type plasminogen activator (tPA) and the generation of neurite outgrowth by PC12 cells. To that aim, cells were treated with agents that interact with the trk receptor and with protein kinases A and C. Nerve growth factor induced only the formation of large neurites. The release of the protease and the production of short neurite outgrowth were found to be protein-kinase-A-dependent events that could be enhanced by simultaneous activation of protein kinase C with phorbol ester. At high concentration, staurosporine, a nonselective inhibitor of protein kinases, induced the production of short neurites and mimicked the protein-kinase-A-dependent effect on tPA release. Such a response was not observed with K-252a, an analogue of staurosporine devoid of neurite-outgrowth-promoting activity. The responses to protein kinase A stimulation and the addition of staurosporine, although similar, seemed to occur through an activation of distinct, yet interacting, signalling pathways. In conclusion, tPA release and large neurite outgrowth from PC12 cells are controlled by parallel, albeit interacting, pathways, suggesting that these two potentially antagonistic events in PC12 cell differentiation can be modulated in a concerted way or independently of each other, depending on the activity of several protein kinases.
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PMID:Protein kinase- and staurosporine-dependent induction of neurite outgrowth and plasminogen activator activity in PC12 cells. 893 50

Human peritoneal mesothelial cells (HMC) play a critical role in maintaining the intraperitoneal balance between fibrinolysis and coagulation by expressing the fibrinolytic enzyme tissue-type plasminogen activator (t-PA) as well as a specific plasminogen activator inhibitor, PAI-1, and the procoagulant protein tissue factor (TF). Of three compounds known to stimulate t-PA synthesis in cultured human endothelial cells, i.e., retinoic acid, the protein kinase C activator 4 beta-phorbol 12-myristate 13-acetate (PMA), and sodium butyrate, only butyrate (1 mM) caused about a threefold increase in t-PA synthesis and mRNA expression in HMC after 24 h of incubation, without markedly affecting PAI-1 synthesis. PMA (10 nM) induced a threefold increase in urokinase-type plasminogen activator (u-PA) mRNA, but u-PA antigen levels in the HMC conditioned media remained below the detection level (0.5 ng/ml), possibly as a result of rapid uptake and degradation by the u-PA receptor. The u-PA receptor mRNA levels were about fivefold enhanced above control levels after PMA treatment of the cells. An increase in intracellular adenosine 3',5'-cyclic monophosphate levels by forskolin (10 microM) diminished t-PA and PAI-1 levels 43 and 17%, respectively. Among the inflammatory mediators tested [tumor necrosis factor-alpha (TNF-alpha), interleukin-1 alpha, and bacterial lipopolysaccharide], TNF-alpha (10-1,000 U/ml) showed the strongest procoagulant effects. We found that the isoflavone compound genistein (25 micrograms/ml) prevented the TNF-alpha-induced expression of PAI-1 and TF while also slightly counteracting the decrease in t-PA synthesis. The protein kinase C inhibitor R0-318220 (3 microM) only moderately opposed the TNF-alpha-induced changes in t-PA and PAI-1 synthesis but completely prevented the induction of TF mRNA. In summary, our results demonstrate that t-PA synthesis in HMC is relatively insensitive to pharmacological stimulation. To restore the balance between fibrinolysis and coagulation under inflammatory conditions, attempts to interfere with the TNF-alpha-signaling pathway were more successful.
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PMID:Modulation of procoagulant and fibrinolytic system components of mesothelial cells by inflammatory mediators. 894 61

Tissue-type plasminogen activator (t-PA) gene expression in human endothelial cells and HeLa cells is stimulated by the protein kinase C activator phorbol 12-myristate 13-acetate (PMA) at the level of transcription. To study the mechanism of transcriptional regulation, we have characterized a segment of the t-PA gene extending from -135 to +100 by in vivo footprinting analysis [dimethyl sulphate (DMS) method] and gel mobility shift assay. In vivo footprinting analysis revealed changes in cleavage pattern in five distinct promoter elements in both endothelial cells and HeLa cells, including a PMA-responsive element (TRE), a CTF/NF-1 binding site and three GC-boxes, and an altered cleavage pattern of the TRE and CTF/NF-1 element after PMA treatment of HeLa cells. Although endothelial cells and HeLa cells differed in the exact G residues protected by nuclear proteins,in vitro bandshift analysis showed that nuclear protein binding to the t-PA promoter was qualitatively and quantitatively very similar in both cell types, except for the TRE. Protein binding to the TRE under non- stimulated conditions was much higher in human endothelial cells than in HeLa cells, and this TRE-bound protein showed a lower dissociation rate in the endothelial cells than in HeLa cells. In endothelial cells, the proteins bound to the TRE consisted mainly of the AP-1 family members JunD and Fra-2, while in HeLa cells predominantly JunD, FosB and Fra-2 were bound. The proteins bound to the other protected promoter elements were identified as SP-1 (GC-box II and III) and CTF/NF-1 (CTF/NF-1 binding site). After PMA treatment of the cells, AP-1 and SP-1 binding was increased two-fold in endothelial cell nuclear extracts and >20-fold in HeLa nuclear extracts. In the endothelial cells, all Jun and Fos forms (c-Jun, JunB, JunD, c-Fos, FosB, Fra-1 and Fra-2) were part of the AP-1 complex after PMA induction. In HeLa cells, the complex consisted predominantly of c-Jun and the Fos family members FosB and Fra-2. In the light of previous studies involving mutational analysis of the human and murine t-PA promoter our results underline an important role of the five identified promoter regions in basal and PMA-stimulated t-PA gene expression in intact human endothelial cells and HeLa cells. The small differences in DMS protection pattern and differences in the individual AP-1 components bound in endothelial cells and HeLa cells point to subtle cell-type specific differences in t-PA gene regulation.
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PMID:Cell-type specific DNA-protein interactions at the tissue-type plasminogen activator promoter in human endothelial and HeLa cells in vivo and in vitro. 901 59

We have demonstrated previously that pemphigus vulgaris (PV)-IgG induces activation of phospholipase C (PLC), production of inositol 1,4,5-trisphosphate, and a rapid transient increase in [Ca2+]i in cultured human keratinocytes, leading to secretion of plasminogen activator and cell-cell detachment in cell culture. In the current study, to examine the involvement of protein kinase C (PKC) in the mechanism of blister formation in PV, we studied the PV-IgG-induced translocation of PKC isozymes from the cytosol to the particulate/cytoskeleton (p/c) fractions and the activation of PKC in human keratinocytes. Cells cultured in Eagle's minimum essential medium were incubated with PV-IgGs for 30 s, 1 min, 5 min, or 30 min. PV-IgG binding to the cell surface antigen (desmoglein III) induced translocation of PKC-alpha from the cytosol to the p/c fractions within 30 s, with a peak at 1 min that lasted at least 30 min. PKC-delta also was translocated within 1 min and reached a peak at 5 min but was reduced to basal levels at 30 min. Alternatively, PKC-eta translocation to the p/c fraction was induced slowly, taking more than 5 min, and was reduced to approximately half-maximum at 30 min, whereas PKC-zeta translocation reached a maximum at 30 s, rapidly returning to baseline by 5 min after PV-IgG stimulation. The total PKC activity in the p/c fraction also was increased after PV-IgG exposure, peaked at 1 min, and was sustained for at least 30 min. These findings suggest that a unique activation profile of PKC isomers may be involved in mediating the intracellular signaling events induced by PV-IgG binding to desmoglein III in cultured human keratinocytes.
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PMID:Pemphigus IgG activates and translocates protein kinase C from the cytosol to the particulate/cytoskeleton fractions in human keratinocytes. 907 78

Activation of protein kinase C- (PKC) and Fos/Jun-dependent signal transduction pathways are thought to be major effects of oncogene action in different tumor systems including human non-small-cell lung carcinoma (NSCLC). We have previously shown that the phorbol ester analogue phorbol-myristate-acetate (PMA), which is a potent activator of PKC, can induce squamous-type cellular differentiation and the expression of proteinases, such as plasminogen activators and pro-cathepsin L, in several NSCLC cell lines. To investigate the PMA-dependent effect on proteinase secretion in more detail, we have now analysed the role of a downstream transmitter of PKC activity in this process, namely Fos, which is part of the AP-1 transcription factor in the nucleus. We transfected a cell line derived from an undifferentiated squamous-cell lung carcinoma with different chimeric fos-estrogen receptor constructs (fos-ER) which makes selective activation of this transcription factor possible. The resulting clones were treated either with PMA as activator of PKC, or with diethylstilbestrol (DES), an estrogen analogue binding to and thereby activating preformed Fos-ER molecules. We show that cells treated with either substance undergo similar phenotypic changes (change from cuboidal to spindle-cell type) and decrease their doubling rates and cloning efficiencies. This is paralleled by the induction of several proteinase genes such as t-PA, urokinase, and pro-cathepsins B and L. Contrary to activated PKC, Fos in this system seems to be unable to initiate terminal squamous-cell differentiation, as assessed by the production of cornified envelopes. It is, however, efficient in the stimulation of neutral or lysosomal proteinase secretion as determined by Western-blot analysis and zymography. This Fos-ER expressing system thus seems to be a valuable tool in the molecular dissection of pathways that lead to the activation and secretion of proteinases in NSCLC cells.
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PMID:Control of proteinase expression by phorbol-ester- and Fos-dependent pathways in human non-small-cell lung-cancer cells. 913 54

Transforming growth factor-beta (TGF-beta) mediates the production of extracellular matrix proteins, proteases and protease inhibitors in epithelial cells. Both TGF-beta and phorbol-12-myristate-13-acetate (PMA) exert both positive and negative effects on mitogenesis in these as well as other cell types. Phorbol esters act through stimulation of protein kinase C (PKC) and are among the most potent tumor promoters known. The present study was conducted to determine whether the effect of TGF-beta in human non-small cell lung cancer (NSCLC) and normal human bronchial epithelial (NHBE) cells parallels that of the phorbol esters and whether this effect of TGF-beta involves PKC. TGF-beta 1 and PMA increased expression of TGF-beta 1 mRNA 24 hr after their addition to both NSCLC and NHBE cells. The effects of these agents on expression of the mRNAs for TGF-beta 2 and TGF-beta 3 were more complex; while TGF-beta 2 and TGF-beta 3 mRNAs increased transiently in response to TGF-beta 1 in NHBE cells and TGF-beta 3 mRNA increased transiently in some NSCLC cells, expression of these mRNAs decreased in most of these cells in response to PMA with the exception of the carcinoid NCI-H727 where TGF-beta 2 mRNA increased dramatically, TGF-beta 1 and PMA both caused a persistent increase in expression of the mRNAs for both plasminogen activator inhibitor-1 (PAI-1) and plasminogen activator (PA) up to 24 hr in most NSCLC cells, with the increase in PAI-1 mRNA beginning several hours before that of PA mRNA. In contrast, while TGF-beta 1 also increased expression of PAI-1 mRNA in NHBE cells, the expression of PA mRNA decreased simultaneously. The effect of PMA on PAI-1 and PA mRNAs was opposite of TGF-beta 1 in these cells, with expression of PAI-1 mRNA decreasing and PA mRNA increasing after addition of PMA. These data show that there is parallel regulation of the genes for TGF-beta 1, PAI-1 and PA by TGF-beta 1 and PMA in NSCLC, but differential regulation of the genes for PAI-1 and PA by these agents in NHBE cells. The responses of the mRNAs and proteins of TGF-beta 1, PAI-1 and PA to TGF-beta 1 and PMA were inhibited by the serine/ threonine kinase inhibitor H7 in NSCLC cells. Treatment of NSCLC cells with TGF-beta 1 and PMA resulted in a persistent increase in the expression of fibronectin mRNA and protein. This response was blocked by the addition of H7. Inhibition of these effects by H7 in NSCLC cells suggests that H7 blocks TGF-beta responses by inhibiting a protein serine/threonine kinase(s). Because the effects of TGF-beta and PMA on the different TGF-beta isoforms, PA, PAI and fibronectin in NHBE and NSCLC cells are complex, our data suggest that there are distinct mechanisms for controlling the different TGF-beta isoforms, PA, PAI and extracellular matrix proteins in normal lung and lung cancer cells.
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PMID:Effects of transforming growth factor-beta 1 and phorbol ester on PAI-1 and PA genes in human lung cells. 925 8

Hexadecylphosphocholine (HePC) is the main representative of a new group of antineoplastic agents, the alkylphosphocholines, which were originally derived from cytotoxic etherlysophospholipids. HePC shows antiproliferative action against a whole variety of tumor cells and tumors in vitro and in vivo. Furthermore, it also induces differentiation in some hematologic cell lines and prevents invasive growth of neoplastic cells in vitro. To date, the precise molecular mechanisms mediating the biological effects of HePC have not been identified yet. As etherlysophospholipids seem to inhibit some pathways of lipid-dependent intracellular signalling, similar effects may be relevant for HePC. We therefore investigated the influence of HePC on phospholipase A2 (PLA2-EC 3.1.1) in the human leukemia cell line U 937. HePC seems to inhibit enzyme activity independently of protein kinase C (PKC) in differentiated U 937 cells stimulated by tumor necrosis factor alpha (TNFalpha). Inhibition of purified secretory PLA2 from snake venom (EC 3.1.1.4) in vitro shows characteristics of a non-competitive mode. In contrast, HePC leads to an enhancement of PLA2 activity in immature cells which cannot be explained by changes in membrane composition. Our data suggest that PLA, inhibition is most probably not the mechanism by which HePC mediates its antiproliferative effects.
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PMID:Differential regulation of phospholipase A2 in human leukemia cells by the etherphospholipid analogue hexadecylphosphocholine. 926 26

In this study, we investigated the role of Ca2+ and G proteins in thrombin-induced acute release (regulated secretion) of tissue-type plasminogen activator (TPA) and von Willebrand factor (vWF), using a previously described system of primary human umbilical vein endothelial cells (HUVECs). The acute release of TPA and vWF, as induced by alpha-thrombin, was almost zero after chelation of Ca2+i, showing that an increase in [Ca2+]i was required. It did not matter whether the increase in [Ca2+]i came from an intracellular or extracellular Ca2+ source. Thrombin-induced release of TPA and vWF already started at low [Ca2+]i, around 100 nmol/L. Half-maximal release was found at a [Ca2+]i, of 261 nmol/L for TPA and at 222 nmol/L for vWF. The Ca2+ signal was transduced to calmodulin, as calmodulin inhibitors inhibited TPA and vWF release. The Ca2+ ionophore ionomycin dose dependently released vWF; half-maximal vWF release occurred at a [Ca2+]i of 311 nmol/L. In contrast, no TPA release was found at all below a [Ca2+]i of 500 nmol/L. Thus, below 500 nmol/L [Ca2+]i, an increase in [Ca2+]i alone was sufficient to induce vWF release but not sufficient to induce TPA release. Protein kinase C did not appear to be involved in TPA or vWF release, as neither an activator nor an inhibitor of protein kinase C significantly influenced release. Inhibition of phospholipase A2 also did not reduce thrombin-induced TPA and vWF release. The involvement of G proteins was studied by using both saponin-permeabilized and intact cells. GDP-beta-S, which inhibits heterotrimeric and small G proteins, significantly inhibited thrombin-induced vWF and TPA release from permeabilized cells. AlF-4, which activates heterotrimeric G proteins, induced TPA and vWF release in both intact and permeabilized HUVECs. Preincubation of HUVECs with pertussis toxin significantly inhibited thrombin-induced vWF release, due to inhibition of thrombin-induced Ca2+ influx. Pertussis toxin did not affect ionomycin-induced release. The inhibitory effect of pertussis toxin was less obvious in thrombin-induced TPA release, because it was counterbalanced by a positive effect of the toxin on TPA release. Thus, both inhibitory and stimulatory (pertussis toxin-sensitive) G proteins were involved in TPA release. Therefore, thrombin-induced acute release of TPA and vWF differed in two respects. First, below a [Ca2+]i of 500 nmol/L, an increase in Ca2+ was sufficient for vWF release but not for TPA release. Second, pertussis toxin-sensitive G proteins were differentially involved in acute TPA and vWF release.
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PMID:Involvement of calcium and G proteins in the acute release of tissue-type plasminogen activator and von Willebrand factor from cultured human endothelial cells. 935 87

Previous studies in our laboratory using a rat model of fetal alcohol exposure (FAE) suggest that FAE-induced behavioral deficits are, in part, linked to neurochemical and electrophysiological deficits in long-term potentiation (LTP) in the entorhinal cortical perforant path projection to the hippocampal formation. Several findings suggest that signal-activated phospholipase C (PLC) and phospholipase A2 (PLA2) are critical to the induction and maintenance of LTP. Thus, alterations in phospholipid metabolism may play a significant role in the LTP deficits observed in FAE offspring. To test this hypothesis, we measured PLC-beta 1 and PLA2 activities in the hippocampus and medial frontal cortex of adult rats prenatally exposed to ethanol. PLC-beta 1 activities were significantly decreased by 20 to 30% in both the hippocampus and medial frontal cortex of FAE rats, compared with ad libitum and pair-fed controls. Total Ca(2+)-dependent PLA2 activity was 25% lower in the medial frontal cortex of FAE rats, but did not significantly differ from controls in the hippocampal formation. Approximately 30% of the measured activity in both the medial frontal cortex and hippocampal formation of ad libitum and pair-fed animals was associated with an 85 kDa cytosolic PLA2 form. Cytosolic PLA2 activities were significantly reduced in both the medial frontal cortex and hippocampal formation of FAE rats, compared with controls. These changes in Ca(2+)-dependent PLA 2 and PLC-beta 1 activities, coupled with reports of FAE-induced deficits in protein kinase C activity, indicate that prenatal exposure to moderate quantities of ethanol causes profound and long-lasting deficits in the cellular signaling mechanisms associated with activity-dependent synaptic plasticity and memory formation.
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PMID:Effects of prenatal ethanol exposure on phospholipase C-beta 1 and phospholipase A2 in hippocampus and medial frontal cortex of adult rat offspring. 939 28


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