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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We tested the hypothesis that adrenomedullin (ADM), a recently isolated hypotensive peptide, is an autocrine transmitter in renal arterial smooth muscle cells (RASMCs). Fura-2 fluorometry revealed that ADM decreases both the cytosolic Ca2+ concentration ([Ca2+]i) and tension induced by 0.3 microM phenylephrine (PE) in porcine RASMCs. The expression of ADM mRNA in RASMCs was assessed by the reverse transcription polymerase chain reaction (RT-PCR), using the total RNA from the RASMCs and the specific primers for the porcine ADM mRNA. A band with an expected size could be detected only when the reverse transcriptase was added, thus indicating that the porcine RASMCs express ADM mRNA. These results therefore suggested that ADM plays an important role in the regulation of the renal vascular tone, not only as a circulating hormone, but also as an autocrine transmitter.
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PMID:Autocrine regulation of the renal arterial tone by adrenomedullin. 748

Although adrenomedullin (AM) previously has been identified in human tumors, its role has remained elusive. Analysis by reverse transcriptase-polymerase chain reaction (RT-PCR) revealed AM mRNA in 18 of 20 human normal tissues representing major organs, and 55 of 58 (95%) malignant cell lines. Western blot and high performance liquid chromatography analysis showed immunoreactive AM species of 18, 14, and 6 kDa that are consistent with the precursor, intermediate product, and active peptide, respectively. Immunohistochemistry and in situ RT-PCR performed on paraffin-embedded tumor cell lines of various tissue origins exhibited AM cytoplasmic staining. Neutralizing monoclonal antibody to AM inhibits tumor cell growth in a concentration-dependent manner, an effect that was reversed with the addition of exogenous AM. Responding tumor cells were shown to have approximately 50,000 AM receptors per cell by Scatchard analysis with 125I-AM and expressed AM receptor mRNA by RT-PCR. Our data showed 36 of 48 (75%) tumor cell lines expressed AM receptor mRNA by RT-PCR assessment, all of them also expressed AM. In the presence of AM, cAMP levels were shown to increase in tumor cells. Our collective data demonstrate that AM and AM receptor are expressed in numerous human cancer cell lines of diverse origin and constitute a potential autocrine growth mechanism that could drive neoplastic proliferation.
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PMID:Adrenomedullin expression in human tumor cell lines. Its potential role as an autocrine growth factor. 879 36

Recently a new type of proteins modulating the pharmacological profile of the calcitonin receptor-like receptor (CRLR) were identified. The receptor-activity-modifying proteins (RAMPs) were shown to be essential for the expression of a functional CRLR and furthermore the RAMPs seemed to modify ligand selectivity of CRLR: coexpression of CRLR and RAMP1 resulted in a CGRP1 type of receptor while an adrenomedullin receptor resulted when CRLR and RAMP2 were coexpressed. In the present study significant molecular expression of CRLR concomitant with RAMP1, 2 and 3 were demonstrated in human meningeal, cerebral and temporal arteries by use of reverse transcriptase polymerase chain reactions (RT-PCR). These findings support previous studies demonstrating functional CGRP1 receptors in human cranial arteries. Furthermore the present study suggests the potential for functional adrenomedullin receptors in human cranial arteries.
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PMID:Expression of calcitonin receptor-like receptor and receptor-activity-modifying proteins in human cranial arteries. 987 47

Rat-2 fibroblasts demonstrate specific binding of 125I-labelled rat adrenomedullin (KD=0.43 nM; Bmax=50 fmol/mg of protein) in the absence of 125I-labelled calcitonin-gene-related peptide (CGRP) binding. Therefore Rat-2 cells were used to examine the pharmacology and signal transduction pathways of adrenomedullin receptors. We examined the effects of adrenomedullin, the CGRP receptor antagonist CGRP-(8-37) and the amylin antagonists AC187 and AC253 on receptor binding and cAMP production. AC253, AC187 and CGRP-(8-37) inhibited 125I-adrenomedullin binding, with respective IC50 values of 25+/-8, 129+/-39 and 214+/-56 nM. Adrenomedullin dose-dependently increased intracellular cAMP (approximate EC50=1.0 nM). CGRP-(8-37), AC253 and AC187 antagonized adrenomedullin-stimulated cAMP production at micromolar concentrations. Using kinase-substrate assays, Mono Q FPLC and 'phospho-specific' Western blotting, we found that adrenomedullin alone abolished basal mitogen-activated protein kinase (MAPK) activity and dose-dependently inhibited platelet-derived-growth-factor-stimulated MAPK activity. Radioimmunoassay for adrenomedullin of media from Rat-2 cells showed a linear release of adrenomedullin-like immunoreactivity of 3.1 fmol/h per 2x10(6) cells. Gel-filtration chromatography showed that this adrenomedullin-like immunoreactivity co-eluted with synthetic rat adrenomedullin. Northern blotting with a rat adrenomedullin cDNA probe was used to confirm the presence of adrenomedullin mRNA. However, neither Northern blotting nor reverse transcriptase-PCR showed the presence of the cloned adrenomedullin receptor (L1). We conclude that the Rat-2 cell line expresses a specific adrenomedullin receptor (coupled to cAMP production and regulation of MAPK) and secretes adrenomedullin, which may participate in a regulatory control loop.
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PMID:Rat-2 fibroblasts express specific adrenomedullin receptors, but not calcitonin-gene-related-peptide receptors, which mediate increased intracellular cAMP and inhibit mitogen-activated protein kinase activity. 993 Dec 92

In this study we characterized calcitonin (CT) receptors in human neuroblastoma IMR 32 cells. Saturation binding assays indicated that [125I]-human CT bound with high affinity to IMR 32 cell membranes (K(d) = 253.6 pM; Bmax = 3.84 fmol/ mg protein). In competition binding studies, human adrenomedullin displayed high affinity for these sites (IC50 = 30 nM) whereas human alpha calcitonin-gene related peptide (alphaCGRP; IC50 = 145 nM) and human amylin (IC50 = 415 nM) showed lower affinity. These peptides increased cAMP levels in viable cells; the relative potencies were: human CT > human adrenomedullin > human cCGRP > or = human amylin. The expression of mRNA coding for the published sequences of the human calcitonin receptor and of the human calcitonin receptor-like receptorwas evaluated by reverse transcriptase-polymerase chain reaction. Electrophoretic analysis did not confirm the occurrence of mRNA coding for the above mentioned receptors in these cells. This study suggests the presence of a novel, putative CT receptor in IMR 32 cells.
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PMID:Characterization of a putative calcitonin receptor in IMR 32 human neuroblastoma cells. 1051 85

Isometric contractile force were studied on isolated human myocardial trabeculae that were paced at 1.0 Hz in tissue baths. Alpha calcitonin gene-related peptide (alpha-CGRP) had a potent positive inotropic effect in most trabeculae from both the right atrium and left ventricle, and this effect was partially antagonized by the CGRP(1) receptor antagonist alpha-CGRP-(8-37) (10(-6) M). Amylin and the CGRP(2) receptor agonist [Cys(acetylmethoxy)(2, 7)]CGRP had a positive inotropic effect in some trabeculae, whereas adrenomedullin had no inotropic effect. Using reverse transcriptase-polymerase chain reaction (PCR) mRNAs encoding the human calcitonin receptor-like receptor and the receptor associated modifying proteins (RAMPs) RAMP1, RAMP2, and RAMP3 were detected in human myocardial trabeculae from both the right atrium and left ventricle. In conclusion, functional CGRP(1) and CGRP(2) receptors may mediate a positive inotropic effect at both the atrial and ventricular level of the human heart. mRNAs for calcitonin receptor-like receptor and specific RAMPs further support the presence of CGRP receptors.
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PMID:Positive inotropy mediated via CGRP receptors in isolated human myocardial trabeculae. 1084 37

The aim of the present study was to determine functional and molecular characteristics of receptors for calcitonin gene-related peptide (CGRP) and adrenomedullin in three different diameter groups of lenticulostriate arteries. Furthermore, the presence of perivascular neuronal sources of CGRP was evaluated in these arteries. In the functional studies, in vitro pharmacological experiments demonstrated that both CGRP and adrenomedullin induce alpha-CGRP-(8-37) sensitive vasodilation in artery segments of various diameters. The maximal amounts of vasodilation induced by CGRP and adrenomedullin were not different, whereas the potency of CGRP exceeded that of adrenomedullin by 2 orders of magnitude. Significant negative correlations between artery diameters and maximal responses were demonstrated for CGRP and adrenomedullin. In addition, the potency of both peptides tended to increase in decreasing artery diameter. In the molecular experiments, levels of mRNAs encoding CGRP receptors and receptor subunits were compared using reverse transcriptase polymerase chain reactions (RT-PCR). The larger the artery, the more mRNA encoding receptor activity-modifying proteins 1 and 2 (RAMP1 and RAMP2) was detected relative to the amount of mRNA encoding the calcitonin receptor-like receptor. By immunohistochemistry, perivascular CGRP containing nerve fibres were demonstrated in all the investigated artery sizes. In conclusion, both CGRP and adrenomedullin induced vasodilation via CGRP receptors in human lenticulostriate artery of various diameter. The artery responsiveness to the CGRP receptor agonists increased with smaller artery diameter, whereas the receptor-phenotype determining mRNA ratios tended to decrease. No evidence for CGRP and adrenomedullin receptor heterogeneity was present in lenticulostriate arteries of different diameters.
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PMID:CGRP and adrenomedullin receptor populations in human cerebral arteries: in vitro pharmacological and molecular investigations in different artery sizes. 1108 May 25

The aim of this study was to assess human intracranial tumours for their gene expression pattern of the vasoactive peptide adrenomedullin (AM), its receptor (AM-R) and leptin, which exerts multiple biological effects including proliferation and angiogenesis via the leptin receptor (OB-Rb). Gene activity of neuropeptide Y (NPY) was monitored additionally. We investigated whether there was a characteristic gene expression pattern of AM and leptin in different intracranial tumours, depending on their proliferation activity and biological behaviour. We investigated 35 non-functioning pituitary adenomas (including eight null cell, four silent plurihormonal, 23 silent gonadotroph adenomas), seven somatotropinomas, seven prolactinomas, eight meningiomas, five astrocytomas, two glioblastoma multiformes and unaffected temporal lobe (n = 8). Quantitative reverse transcriptase-polymerase chain reaction (TaqMan RT-PCR) was performed. AM mRNA was detectable in all tumour specimens. AM/GAPDH (glyceraldehyde-3-phosphate dehydrogenase) ratio was significantly higher in somatotropinomas, as was AM/CD31 ratio in prolactinomas, compared with inactive adenomas (P < 0.05). AM-R mRNA was found in all tumour subgroups in small quantities but, in general, higher in tumours than in temporal lobe tissue, respectively. AM-R/CD31 ratio was significantly higher in prolactinomas than in inactive adenomas (P < 0.05). Leptin was detectable in very low quantities in each subgroup. OB-Rb gene expression was found in all tumour subgroups, OB-Rb/GAPDH ratio was highest for meningiomas (P < 0.0001, compared with temporal lobe). NPY mRNA was detectable in temporal lobe in higher quantities than in tumours (P < 0.0001), and almost undetectable in prolactinomas and astrocytomas. Our data demonstrate that AM and AM-R, NPY, as well as leptin and OB-Rb, are expressed in various intracranial tumours in humans but their particular function has to be elucidated further. At present, there is no evidence for a cross-talk on transcriptional level between the peptidergic vasodilative system AM and the putative angiogenic and proliferation affecting factor leptin.
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PMID:Gene expression of adrenomedullin, leptin, their receptors and neuropeptide Y in hormone-secreting and non-functioning pituitary adenomas, meningiomas and malignant intracranial tumours in humans. 1148 41

1. Calcitonin gene-related peptide (CGRP), amylin and adrenomedullin (AM) belong to the same family of peptides. Accumulating evidence indicate that the calcitonin (CT) receptor, the CT receptor-like receptor (CRLR) and receptor-activity-modifying proteins (RAMPs) form the basis of all the receptors in this family of peptides. 2. Using reverse transcriptase - polymerase chain reaction the presence of mRNA sequences encoding the CRLR, RAMP1 and RAMP2 were demonstrated in porcine left anterior descending (LAD) coronary arteries, whereas porcine calcitonin (CT) receptor mRNA was not present. The partial porcine mRNA sequences shared 82 - 92% nucleotide identity with human sequences. 3. The human peptides alphaCGRP, betaCGRP, AM and amylin induced relaxation with pEC(50) values of 8.1, 8.1, 6.7 and 6.1 M respectively. 4. The antagonistic properties of a novel non-peptide CGRP antagonist 'Compound 1' (WO98/11128), betaCGRP(8 - 37) and the proposed AM receptor antagonist AM(22 - 52) were compared to the well-known CGRP(1) receptor antagonist alphaCGRP(8 - 37). 5. The alphaCGRP(8 - 37) and betaCGRP(8 - 37) induced concentration-dependent (10(-7) - 10(-5) M) rightward shift of both the alphaCGRP and betaCGRP concentration-response curves. betaCGRP(8 - 37) (10(-6) M) had the same effect as alphaCGRP(8 - 37) (10(-6) M), but with less potent rightward shift of the concentration-response curves for alphaCGRP, AM and amylin. 6. Preincubation with 'Compound 1' (10(-7) - 10(-5) M) and AM(22 - 52) (10(-6) M) had no significant antagonistic effect. 7. In conclusion, the building blocks forming CGRP and AM receptors were present in the porcine LAD, whereas those of the amylin receptor were not. alphaCGRP, betaCGRP, AM and amylin mediated vasorelaxation via the CGRP receptors. No functional response was detected to adrenomedullin via the adrenomedullin receptor.
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PMID:CGRP receptors mediating CGRP-, adrenomedullin- and amylin-induced relaxation in porcine coronary arteries. Characterization with 'Compound 1' (WO98/11128), a non-peptide antagonist. 1149 28

Presently, there is no effective treatment for glioblastoma, the most malignant and common brain tumor. Growth factors are potential targets for therapeutic strategies because they are essential for tumor growth and progression. Peptidylglycine alpha-amidating monooxygenase is the enzyme producing alpha-amidated bioactive peptides from their inactive glycine-extended precursors. The high expression of peptidylglycine alpha-amidating monooxygenase mRNA in glioblastoma and glioma cell lines points to the involvement of alpha-amidated peptides in tumorigenic growth processes in the brain. After screening of amidated peptides, it was found that human glioblastoma cell lines express high levels of adrenomedullin (AM) mRNA, and that immunoreactive AM is released into the culture medium. AM is a multifunctional regulatory peptide with mitogenic and angiogenic capabilities among others. Real-time quantitative reverse transcriptase-polymerase chain reaction analysis showed that AM mRNA was correlated to the tumor type and grade, with high expression in all glioblastomas analyzed, whereas a low expression was found in anaplastic astrocytomas and barely detectable levels in low-grade astrocytomas and oligodendrogliomas. In the present study we also demonstrate the presence of mRNA encoding the putative AM receptors, calcitonin receptor-like receptor/receptor activity-modifying protein-2 and -3 (CRLR/RAMP2; CRLR/RAMP3) in both glioma tissues and glioblastoma cell lines and further show that exogenously added AM can stimulate the growth of these glioblastoma cells in vitro. These findings suggest that AM may function as an autocrine growth factor for glioblastoma cells. One way to test the autocrine hypothesis is to interrupt the function of the endogenously produced AM. Herein, we demonstrate that a polyclonal antibody specific to AM, blocks the binding of the hormone to its cellular receptors and decreases by 33% (P < 0.001) the growth of U87 glioblastoma cells in vitro. Intratumoral administration of the anti-AM antibody resulted in a 70% (P < 0.001) reduction in subcutaneous U87 xenograft weight 21 days after treatment. Furthermore, the density of vessels was decreased in the antibody-treated tumors. These findings support that AM may function as a potent autocrine/paracrine growth factor for human glioblastomas and demonstrate that inhibition of the action of AM (produced by tumor cells) may suppress tumor growth in vivo.
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PMID:Neutralization of adrenomedullin inhibits the growth of human glioblastoma cell lines in vitro and suppresses tumor xenograft growth in vivo. 1194 13


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