Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.26.9 (ribonuclease)
6,589 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Thrombin, via activation of vascular endothelial and smooth muscle cell thrombin receptors, modulates vascular wall healing. To understand the mechanisms that regulate human thrombin receptor (HTR) expression, we cloned and characterized the HTR gene. The HTR gene consists of Exon I, which contains the 5'-regulatory region and 85 nucleotides of coding sequence; a approximately 15-kb intron; and Exon II, which contains the remainder of the coding sequence and the entire 3'-untranslated region. Multiple transcription initiation sites were identified by S1 mapping and ribonuclease protection assay. DNA sequence analysis indicated the presence of two SP-1-AP-2 consensus binding sequences, near or within the transcription initiation sites, and consensus binding sequences for numerous regulatory proteins that potentially modulate HTR expression. Functional analysis of the HTR promoter was performed by transfecting human microvascular endothelial cells with HTR promoter region-luciferase constructs. The highest level of expression was obtained with a 0.7-kb promoter sequence and was progressively less with fragments of 0.54, 1.16, 1.6, and approximately3.2 kb. The data presented in this report provide a foundation for further characterization of the HTR gene and the mechanisms that regulate its expression within the blood vessel wall.
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PMID:Cloning and identification of regulatory sequences of the human thrombin receptor gene. 882 85

Thrombin is one of the first regulatory molecules present at sites of CNS trauma or injury. Exposure of neuronal and glial cells to thrombin produces potent morphological as well as cytoprotective and cytotoxic effects, but little is known about how this important modulator affects neurotransmitter signaling. In astrocyte cultures that have been morphologically differentiated by exposure to transforming growth factor-alpha, addition of thrombin induced a retraction of astrocytic processes and suppressed the stimulation of phosphoinositide hydrolysis by the selective metabotropic glutamate receptor (mGluR) agonist 1-aminocyclopentane-1S,3R-dicarboxylic acid. In addition to the suppression of phosphoinositide hydrolysis, thrombin treatment produced a corresponding reduction in level of mGluR5 mRNA as demonstrated with ribonuclease protection assay and reduced content of mGluR5 receptor protein as seen with western blotting. In contrast, thrombin exposure up-regulated astrocyte beta-actin mRNA levels. A synthetic hexapeptide with a sequence corresponding to the amino-terminus of the thrombin receptor's tethered ligand also mimicked the ability of thrombin to suppress mGluR5 levels and to increase beta-actin mRNA content, suggesting that these effects of thrombin are mediated by proteolytically activated cell surface thrombin receptors. Thrombin's suppressive effect on mGluR5 was resistant to pretreatment with pertussis toxin or various protein kinase and protein phosphatase inhibitors. However, the serine/threonine protein kinase inhibitor H-7 did prevent thrombin-induced reversal of astrocyte stellation and induction of beta-actin mRNA levels, indicating that these effects of thrombin involve a signaling pathway distinct from the one that mediates the suppressive effects of thrombin on mGluR5.
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PMID:Exposure of astrocytes to thrombin reduces levels of the metabotropic glutamate receptor mGluR5. 885 25

Calorie restriction (CR) and supplementation with fish oil (FO) are known to increase the life span and diminish histological evidence of glomerulonephritis in lupus prone (NZB x NZW)F1 (B/W) mice. Cellular proliferation is an important pathological element in the development of lupus nephritis, and we have examined the expression of thrombin receptor (TR) and the mitogenic agents PDGF-A and -B. Weanling B/W mice were fed either ad libitum or a calorie restricted (CR; 40% less calories than ad libitum) diet supplemented with either 5% (w/w) corn oil (CO) or FO. CR animals consumed 2.7-3.0 g of wet food per day versus 4.5-5.0 g for the ad libitum animals. Renal RNA was extracted from young (3.5-4.0 months of age) and old (8-10 months of age) mice. Densitometric analysis (reference gene GAPDH) of blots from Northern (PDGF-A and -B) and ribonuclease protection assays (TR) produced the following data: (i) in young mice no signal was detected for PDGF-A, -B and TR in all four groups, while the signals were readily detectable in old mice; (ii) in old mice low and similar levels of PDGF-B were detected, and neither CR nor the source of lipid altered its expression; (iii) CR significantly inhibited PDGF-A and TR expression in both CO (ad libitum versus CR; PDGF-A, 3.25-fold, P < 0.025; TR, 3.7-fold, P < 0.01) and FO (ad libitum versus CR; PDGF-A, 4.56-fold, P < 0.01; TR, 3.6-fold, P < 0.025) groups; (iv) although FO (versus CO) produced a trend towards decreased expression, results were not statistically significant. We conclude that suppression of renal disease in lupus-prone mice by CR is accompanied by decreased expression of PDGF-A and the thrombin receptor.
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PMID:Calorie restriction decreases platelet-derived growth factor (PDGF)-A and thrombin receptor mRNA expression in autoimmune murine lupus nephritis. 909 12

Rapid regeneration of the endothelium is a critical component of vascular wall repair because limitations of this process enhance early thrombotic and vasospastic complications, as well as late sequelae of recurrent lesion formation. We have postulated that direct activation of the thrombin receptor initiates both mitogenic and chemokinetic endothelial behavior which facilitates intimal repair. To characterize the role of the thrombin receptor in human endothelial cell (EC) proliferation and migration, we investigated the effects of both alpha-thrombin (0.5-10 U/ml) and its receptor-activating peptide (TRAP; 1-100 microM). Responses of human aortic (HAEC) and umbilical vein (HUVEC) were characterized using [3H]thymidine and 61Cr microcarrier bead assays of proliferation and migration, respectively. Expression of motility-related genes was evaluated using a ribonuclease protection assay. Thrombin exerts both of its chemokinetic and mitogenic effects differentially in human endothelial cells. Following 2 or 4 days in culture, HUVEC proliferation increased two- to threefold after exposure to thrombin, primarily in the low concentration range (P < 0.05). However, HACE proliferation was inhibited up to 50% after a 4-day incubation period (P < 0.005). These mitogenic effects, including the inhibition of aortic endothelial cell proliferation, were reproduced, in part, by thrombin receptor activation with TRAP. In contrast, thrombin stimulates migratory responses in HAEC, but not HUVEC. However, this behavior was not reproduced by TRAP. It is noteworthy that urokinase-plasminogen activator (u-PA) expression was much more strongly expressed in migrating HAEC than in the HUVEC population. Moreover, when stimulated with thrombin, u-PA gene expression was significantly augmented in HAEC. It has been speculated that an effective human thrombin receptor (HTR) antagonist may reduce the proliferation of vascular smooth muscle cells and the development of a restenotic lesion following arterial wall injury. Our data suggest that such an inhibitor will likely also accelerate intimal regeneration through a dominant effect on limiting the HTR inhibitory effect on endothelial proliferation.
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PMID:Endothelial cells exhibit differential chemokinetic and mitogenic responsiveness to alpha-thrombin. 918 72

Initial work has shown that clonal B cells from B-chronic lymphocytic leukemia (B-CLL) are able to synthesize pro-angiogenic molecules. In this study, our goal was to study the spectrum of angiogenic factors and receptors expressed in the CLL B cell. We used ELISA assays to determine the levels of basic fibroblast growth factors (bFGF), vascular endothelial growth factor (VEGF), endostatin, interferon-alpha (IFN-alpha) and thrombospondin-1 (TSP-1) secreted into culture medium by purified CLL B cells. These data demonstrated that CLL B cells spontaneously secrete a variety of pro- and anti-angiogenic factors, including bFGF (23.9 pg/ml +/- 7.9; mean +/- s.e.m.), VEGF (12.5 pg/ml +/- 2.3) and TSP-1 (1.9 ng/ml +/- 0.3). Out of these three factors, CLL B cells consistently secreted bFGF and TSP-1, while VEGF was expressed in approximately two-thirds of CLL patients. Of interest, hypoxic conditions dramatically upregulated VEGF expression at both the mRNA and protein levels. We also employed ribonuclease protection assays to assay CLL B cell expression of a variety of other angiogenesis-related molecules. These analyses revealed that CLL B cells consistently express mRNA for VEGF receptor 1 (VEGFR1), thrombin receptor, endoglin, and angiopoietin. Further analysis of VEGFR expression by RT-PCR revealed that CLL B cells expressed both VEGFR1 mRNA and VEGFR2 mRNA. In summary, these data collectively indicate that CLL B cells express both pro- and anti-angiogenic molecules and several vascular factor receptors. Because of the co-expression of angiogenic molecules and receptors for some of these molecules, these data suggest that the biology of the leukemic cells may also be directly impacted by angiogenic factors as a result of autocrine pathways of stimulation.
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PMID:B-CLL cells are capable of synthesis and secretion of both pro- and anti-angiogenic molecules. 1198 54

Thrombin receptors, i.e. proteinase-activated receptors (PARs), are expressed in endothelial cells (ECs) and neutrophils and directly affect platelet function and thrombosis. Although endothelial dysfunction and neutrophil activation have been demonstrated in women with preeclampsia (PE), the expression and regulation of PARs have not been defined in PE. In this study, we measured the expression of PARs in ECs and in neutrophils derived from normal and preeclamptic pregnancies. We also examined the effects of placental factors on PAR expression in these cells in vitro. ECs were isolated from umbilical cords (human umbilical vein ECs) from normal and preeclamptic pregnancies. Neutrophils were isolated from blood obtained from nonpregnant, uncomplicated pregnant, and preeclamptic women. Total RNA was extracted from the first-passage (P1) ECs (normal and PE) and from normal P1 ECs incubated with conditioned media derived from normal and preeclamptic placental cultures. The mRNA expression of thrombin receptor (PAR1), PAR2, and PAR3 was measured by ribonuclease protective assay. The expression of glyceraldehyde 3-phosphate dehydrogenase was used as an internal control for each sample. We found that: 1) PAR1 expression was enhanced in ECs from PE, compared with ECs from normal pregnancies; 2) PAR2 expression was expressed in PE ECs but not in normal ECs; 3) neutrophils from nonpregnant women, normal, and preeclamptic pregnancies expressed PAR2, whereas only neutrophils from normal and preeclamptic pregnancies expressed PAR1; and 4) factors released from preeclamptic placenta up-regulated PAR1 and PAR2 expression in ECs but not in neutrophils. We conclude that mRNA expression of PAR1 and PAR2 is increased in ECs derived from preeclamptic pregnancies. Up-regulation of thrombin receptor expression in neutrophils may be a unique phenomenon during pregnancy but not apparently unique to PE. Factors released from the placenta are likely candidates in regulating PAR expression in ECs and may contribute to the platelet activation and vascular endothelial dysfunction in PE.
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PMID:Expression of thrombin receptors in endothelial cells and neutrophils from normal and preeclamptic pregnancies. 1216 2

Reticulated platelets (RPs) are larger, hyperreactive platelets that contain significantly more ribonucleic acid (RNA) compared with mature platelets (MPs). High levels of RPs in peripheral blood are predictors of an insufficient response to dual antiplatelet therapy in cardiovascular patients and of adverse cardiovascular events. However, the mechanisms underlying these correlations remain widely unknown and the biology of RPs has not been investigated yet. Here, we compared for the first time the transcriptomic profiles of RPs and MPs isolated from peripheral blood of healthy donors. Total RNA sequencing revealed 1,744 differentially expressed genes (670 downregulated, 1,074 upregulated) in RPs compared with MPs. In particular, transcripts for the collagen receptor GP6, thromboxane receptor A2 (TBXA2R), thrombin receptor PAR4 (F2RL3), and adenosine triphosphate receptors P2RX1, ORAI2, and STIM1 (both involved in calcium signaling) were significantly upregulated in RPs, whereas several RNA regulators as the ribonuclease PARN, the RISC-component TNRC6A, and the splicing factor LUC7L3 were downregulated in RPs. Gene ontology analysis revealed an enrichment of relevant biological categories in RPs including platelet activation and blood coagulation. Gene Set Enrichment Analysis showed an overrepresentation of several platelet activation pathways like thrombin, thromboxane, and glycoprotein IIb/IIIa signaling in RPs. Small-RNA sequencing reported 9 micro-RNAs significantly downregulated in RPs with targets involved in platelet reactivity. Our data show for the first time an enrichment of several prothrombotic transcripts in RPs providing a first biological explanation for their hyperreactive phenotype.
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PMID:Transcriptome Analysis of Reticulated Platelets Reveals a Prothrombotic Profile. 3147 89

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