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)

Changes in gene expression could play a central role in the phenotypic abnormalities of the retinal vascular cells observed in diabetic retinopathy and other retinal diseases. To measure gene expression in human retinal microvessels, a RNA-probe excess solution hybridization assay was used. Retinal microvessels were isolated from eyes obtained within 36 hr of death, and intact RNA was extracted by the guanidine method. Hybridization of poly(A)+ RNA northern blots revealed only the cytoskeletal beta-actin message; by using the more sensitive solution hybridization assay, the plasminogen activator-inhibitor 1 (PAI-1) and von Willebrand factor (vWF) mRNAs were quantified. The prevalence of these transcripts in the retinal microvessels was 0.04 x 10(6) copies/ng RNA for PAI-1 and 0.14 x 10(6) copies/ng RNA for vWF, much less than the prevalence in human umbilical vein endothelial cells (1.93 x 10(6) and 3.90 x 10(6), respectively). The PAI-1 mRNA levels in retinal microvessels isolated from five type II diabetic patients were significantly higher than those in vessels isolated from ten age-matched controls (0.06 x 10(6) versus 0.04 x 10(6) copies/ng RNA, P less than 0.05). The solution hybridization assay accurately measured low-abundance mRNAs in human retinal microvessels; determination of gene expression in these cells could aid in understanding the pathogenesis of important ophthalmologic diseases such as diabetic retinopathy.
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PMID:Measurement of gene expression in human retinal microvessels by solution hybridization. 170 59

The effects of recombinant tissue-type plasminogen activator (rt-PA) and of an inactive mutant of rt-PA, obtained by mutagenesis of the active site Ser478 to Ala (rt-PA-Ala478), on the synthesis and secretion of plasminogen activator inhibitor-1 (PAI-1) by human umbilical vein endothelial cells (HUVEC) in culture were studied. Under base-line conditions, PAI-1 antigen secretion was 4.3 +/- 1.0 micrograms (mean +/- S.D., n = 8) per 10(6) cells in 24 h. This PAI-1 had a low specific activity (6,000 +/- 1,600 units/mg) and Mr of 50,000, which was not altered by addition of rt-PA. In HUVEC cultured with 2 micrograms/ml rt-PA-Ala478, PAI-1 antigen secretion was 2.1 +/- 0.8 micrograms (n = 5) per 10(6) cells in 24 h with a specific activity of 120,000 +/- 42,000 units/mg and Mr of 50,000. Addition of rt-PA to this conditioned medium resulted in generation of three main components: 16% migrated as an Mr 106,000 rt-PA.PAI-1 complex, 16% as an Mr 81,000 degraded rt-PA.PAI-1 complex and the remainder as an Mr 45,000 degradation product of PAI-1. HUVEC cultured with 2 micrograms/ml rt-PA secreted 3.9 +/- 0.6 micrograms (n = 8) PAI-1 antigen per 10(6) cells within 24 h, of which 20-50% occurred as intact or degraded complexes with t-PA (Mr 106,000 and 81,000) and the rest as an inactive Mr 45,000 degradation product of PAI-1. PAI-1 mRNA levels, determined by Northern blot analysis and expressed relative to beta-actin mRNA levels, were very similar for HUVEC cultured in the absence or the presence of rt-PA or rt-PA-Ala478. It is concluded that PAI-1 is secreted by HUVEC in culture in fully active form which spontaneously inactivates. PAI-1 can be stabilized by addition of rt-PA-Ala478 to the culture medium, resulting in a 20-fold increase in specific activity. Interaction of rt-PA with active PAI-1 produces both t-PA.PAI-1 complex and an inactive degradation product of PAI-1.
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PMID:Synthesis and secretion of plasminogen activator inhibitor 1 by human endothelial cells in vitro. Effect of active site mutagenized tissue-type plasminogen activator. 189 37

A high-level and stable expression system of human tissue-type plasminogen activator (t-PA) was accomplished in human cells by selecting a promoter and a host cell line. First, we have constructed two types of t-PA expression plasmids containing 3 kb of the human beta-actin promoter region or 0.3 kb of SV40 early promoter region and these plasmids were transfected into HeLa cells, respectively, and the resulting transfectants were found to secrete various amounts of t-PA derived from the plasmids to the culture media. Southern blot analysis revealed that the beta-actin promoter was more efficient than the SV40 early promoter with regard to the expression level per single copy of the t-PA gene in the transfected HeLa cells. Next, the t-PA expression plasmid containing the beta-actin promoter was also transfected into WI-38 VA13 cells, a human fibroblastic cell line, and KMS-5 cells, a human lymphoid cell line, in order to compare the expression ability of the promoter among these three cell lines. Some of the transfectants from both cell lines were also found to produce t-PA. It was also found that the expression levels in HeLa and WI-38 VA13 seemed to be more efficient than that in KMS-5.
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PMID:Stable expression of human tissue-type plasminogen activator regulated by beta-actin promoter in three human cell lines: HeLa, WI-38 VA13 and KMS-5. 193 13

We have used subtractive hybridization to isolate cDNAs overexpressed in SK-PC-1 pancreas cancer cells. Forty-five independent clones corresponding to 11 genes were identified. Their expression in cultured pancreas cancer cells, normal pancreas tissue, and normal exocrine pancreas cultures was examined by Northern blotting. cDNA clones can be grouped into two broad categories: (1) those corresponding to genes expressed at high levels both in tumor cell lines and in primary cultures of normal pancreas, but not in normal tissue (i.e. thymosin beta4(3), cytokeratin 18, beta-actin, pyruvate kinase and mitochondrial genes); and (2) those corresponding to genes expressed at high levels in pancreas cancer cultures but not in normal pancreas tissue or cultured cells (i.e. tissue-type plasminogen activator and cathepsin H). The overexpression of these proteases in pancreas cancers suggests that they play a role in the aggressive biological behavior of this tumor.
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PMID:Isolation of tissue-type plasminogen activator, cathepsin H, and non-specific cross-reacting antigen from SK-PC-1 pancreas cancer cells using subtractive hybridization. 864 71

Effects of nicotine treatment (4.5 mg/kg of nicotine-free base/day administered s.c. by osmotic minipumps for 14 days) on focal ischemic stroke and expression of tissue plasminogen activator (t-PA) and plasminogen activator inhibitor-1 (PAI-1) in cerebral microvessels were studied in rats in vivo using a reversible (1 h) middle cerebral artery occlusion model. Plasma levels of nicotine and its major metabolite cotinine after 14 days of treatment were 88 and 364 ng/ml, respectively. Nicotine treatment resulted in 35-40% (p < 0.001) decrease in the blood flow in the periphery of the ischemic core during reperfusion, an increase in the neurologic score of 2.6-fold (p < 0.01), and 36% (p < 0.05) and 121% (p < 0.01) increases in the injury and edema volume in the pallium, respectively. A free pool of brain microvascular t-PA antigen was completely depleted by nicotine, while the expression of the PAI-1 antigen and/or PAI-1-t-PA complexes remained unchanged. The relative abundance of cerebromicrovascular t-PA mRNA transcript versus beta-actin mRNA transcript did not change with nicotine. It is concluded that chronic nicotine treatment impairs the restoration of blood flow, worsens the neurologic outcome, and enhances brain injury following an ischemic insult. These nicotine effects are associated with depletion of brain microvascular t-PA antigen.
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PMID:Chronic nicotine treatment enhances focal ischemic brain injury and depletes free pool of brain microvascular tissue plasminogen activator in rats. 904 Apr 92

We have generated a mouse strain carrying a transgene driven by a strong and ubiquitous promoter (human cytomegalovirus hCMV/beta-actin) and containing an enhanced green fluorescent protein (eGFP) coding sequence upstream of the 3' untranslated region (3'UTR) of tissue-type plasminogen activator (t-PA) mRNA. The 3'UTR of t-PA mRNA is known to be involved in the reversible deadenylation and translational repression of transcripts in mouse oocytes. hCMV/beta-actin-eGFP-3'UTR t-PA transgenic mice express eGFP mRNA in all brain structures analyzed but lack eGFP fluorescence, with the exception of blood vessels, choroid plexus, and Purkinje cells. Taking advantage of these features, we tested whether certain pathological conditions, in particular injuries of the nervous system, might trigger eGFP fluorescence in traumatized cells or neurons. From this perspective, we analyzed eGFP mRNA expression and eGFP fluorescence in experimental models of nervous system lesions, such as motoneuron axotomy and cerebral stroke induced by middle cerebral artery occlusion. We found an increase in eGFP fluorescence in specific brain areas in cells suffering or reacting to these injuries. This increased fluorescence is correlated with an increased transcription of eGFP in lesioned cells, presumably enhanced by a release of the translational silencing mediated by the 3'UTR region of the t-PA mRNA. This transgenic mouse model may prove useful to study the development of neurodegenerative lesions.
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PMID:Induction of enhanced green fluorescent protein expression in response to lesions in the nervous system. 1515 81

Phospholipase A(2) (PLA(2)) enzymes participate in a potent inflammatory pathway through the liberation of arachidonic acid upon hydrolysis of membrane glycerophospholipids. The presence of implanted polycarbonate-urethane (PCNU) materials, used in several medical applications, has the ability to influence inflammatory responses of human macrophages that are recruited to a tissue-material interface; however, the specific inflammatory pathways that are activated upon macrophage attachment to PCNU are largely unknown. Previous studies suggested the participation of PLA(2) pathways in material degradation with the use of chemical inhibitors, such as aristolochic acid (ARIST), however not accurately defining the specific PLA(2) enzymes involved. The current study aimed to establish specific groups of PLA(2) involved in the macrophage foreign body response to PCNU. ARIST was assessed for specific effects on secretory PLA(2) (sPLA(2)) protein expression and non-specific effects on key proteins, beta-actin and monocyte-specific esterase, implicated in the macrophage attack on PCNU materials. Macrophage attachment to PCNU materials induced increased intracellular expression of cytosolic PLA(2) (cPLA(2)), but not sPLA(2), relative to tissue culture polystyrene (TCPS) as detected by immunoblot analysis, demonstrating an early and delayed stimulation during the time course of increased cPLA(2) protein expression. Laser scanning confocal microscopy images indicated a change in location of cPLA(2) in macrophages adherent to PCNU surfaces compared to TCPS. This study has illustrated changes in macrophage cPLA(2) expression in response to cell-attachment to PCNU surfaces, demonstrating that the macrophage foreign body response to biomaterials induces a potent inflammatory pathway, which may lead to tissue damage near the site of material implantation.
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PMID:Intracellular phospholipase A2 expression and location in human macrophages: influence of synthetic material surface chemistry. 1756 22