endothelin-1 and Kidney-Neoplasms

This review highlights new developments in miRNA as diagnostic and surveillance tools in diseases damaging the renal proximal tubule mediated by endothelin in the field of renal carcinoma, proteinuric kidney disease and tubulotoxicity. A new mechanism in the miRNA regulation of proteins leads to the binding of the miRNA directly to the DNA with premature transcriptional termination and hence the formation of truncated protein isoforms (Mxi2, Vim3). These isoforms are mediated through miRNA15a or miRNA 498, respectively. ET-1 can activate a cytoplasmic complex consisting of NF-κB p65, MAPK p38α, and PKCα. Consequently, PKCα does not transmigrate into the nucleus, which leads to the loss of suppression of a primiRNA15a, maturation of this miRNA in the cytoplasm, tubular secretion and detectability in the urine. This mechanism has been shown in renal cell carcinoma and in proteinuric disease as a biomarker for the activation of the signalling pathway. Similarly, ET-1 induced miRNA 498 transmigrates into the nucleus to form the truncated protein Vim3, which is a biomarker for the benign renal cell tumour, oncocytoma. In tubulotoxicity, ET-1 induced miRNa133a down-regulating multiple-drug-resistant related protein-2, relevant for proteinuric and cisplatin/cyclosporine A toxicity. Current advantages and limitations of miRNAs as urinary biomarkers are discussed.

Topics: Animals; Biomarkers; Carcinoma, Renal Cell; Endothelin-1; Gene Expression Profiling; Humans; Kidney Neoplasms; Kidney Tubules, Proximal; Mice; MicroRNAs; Proteinuria; Rats; Signal Transduction

The endothelin system comprises the three peptide hormones endothelin (ET)-1, -2, -3, their G protein-coupled receptors, endothelin-A-receptor (ET(A)R) and endothelin-B-receptor (ET(B)R), and the enzymes of endothelin biosynthesis and degradation. In the past two decades, an impressive amount of data has been accumulated investigating the role of the endothelin system in a variety of malignancies. In many cancers, ET-1/ET(A)R interaction induces proliferation, angiogenesis, antiapoptosis and resistance to chemotherapy. Data indicate a pivotal role of the endothelin system in tumorigenesis, local progression and metastasis. Subsequently, novel drugs have been designed inhibiting ET-1 biosynthesis or ET(A)R interaction. A wide range of preclinical data is available on the role of ET(A)R antagonists in gynecological, urological and breast cancers providing evidence for their antiangiogenic, proapoptotic and growth inhibitory effects. Of particular interest is the anti-invasive and antimetastatic efficacy of ET(A)R antagonists and synergism when co-administered with established cancer therapies. Data indicate a future role of ET(A)R antagonists in oncologic therapies.

Topics: Animals; Antineoplastic Agents; Aspartic Acid Endopeptidases; Breast Neoplasms; Endometrial Neoplasms; Endothelin Receptor Antagonists; Endothelin-1; Endothelin-Converting Enzymes; Female; Humans; Kidney Neoplasms; Male; Metalloendopeptidases; Ovarian Neoplasms; Prostatic Neoplasms; Receptors, Endothelin; Urinary Bladder Neoplasms; Urogenital Neoplasms; Uterine Cervical Neoplasms

Angiogenesis inhibition with sunitinib, a multitarget tyrosine kinase inhibitor of the vascular endothelial growth factor receptor, is associated with hypertension and cardiac toxicity, of which the underlying pathophysiological mechanism is unknown. We investigated the effects of sunitinib on blood pressure (BP), its circadian rhythm, and potential mechanisms involved, including the endothelin-1 system, in 15 patients with metastatic renal cell carcinoma or gastrointestinal stromal tumors. In addition, we investigated in rats the effect of sunitinib on BP, serum endothelin-1 levels, coronary microvascular function, cardiac structure, and cardiac mitochondrial function. In patients, BP increased by ≈15 mm Hg, whereas heart rate decreased after 4 weeks of treatment. Furthermore, the nocturnal dipping of BP diminished. Plasma endothelin-1 concentration increased 2-fold (P<0.05) and plasma renin decreased (P<0.05), whereas plasma catecholamines and renal function remained unchanged. In rats, 8 days of sunitinib administration induced an ≈30-mm Hg rise in BP, an attenuation of the circadian BP rhythm, and a 3-fold rise in serum endothelin-1 and creatinine, of which all but the rise in creatinine reversed after sunitinib withdrawal. Coronary microvascular function studies after 8 days of sunitinib administration showed decreased responses to bradykinin, angiotensin II, and sodium nitroprusside, all normalizing after sunitinib withdrawal. Cardiac structure and cardiac mitochondrial function did not change. In conclusion, sunitinib induces a reversible rise in BP in patients and in rats associated with activation of the endothelin-1 system, suppression of the renin-angiotensin system, and generalized microvascular dysfunction.

Topics: Aged; Angiotensin II; Animals; Blood Pressure; Bradykinin; Carcinoma, Renal Cell; Cells, Cultured; Coronary Circulation; Endothelial Cells; Endothelin-1; Female; Gastrointestinal Stromal Tumors; Heart; Heart Rate; Humans; Hypertension; In Vitro Techniques; Indoles; Kidney Neoplasms; Male; Middle Aged; Pyrroles; Rats; Rats, Inbred WKY; Receptor Protein-Tyrosine Kinases; Renin; Sunitinib; Vascular Endothelial Growth Factor A

Recent reports suggest that complex interactions exist between the neuroendocrine and immune systems. It has been shown for example that cytokines are able to stimulate the hypothalamo-pituitary-adrenal axis. In addition, some studies present evidence that endothelin is able to modulate the activity of several hypothalamic-pituitary axes, e.g. by inducing the ACTH production.. We investigated the effects of interleukin-2 on endothelin levels and the hypothalamo-pituitary-adrenal axis. We determined the interleukin-6, big-endothelin, endothelin-1, ACTH, cortisol and AVP responses to intravenously and subcutaneously administered interleukin-2 in 8 cancer patients in a randomized placebo controlled trial.. 8 Patients (2 female and 6 male), age 44 +/- 4.8 years, were enrolled. All patients had a World Health Organization performance status of 1 or less and a Karnofsky Index of at least 80%.. Blood-samples were taken before and 15, 30, 45, 60, 120, 180, 240, 300 and 360 min after interleukin-2 injection. Cytokine serum levels and the plasma levels of big-endothelin, endothelin, ACTH and AVP were analysed using radioimmuno-assays. Cortisol was assayed by an enzyme-linked immunosorbent assay.. Interleukin-2 treatment significantly increased plasma big-endothelin levels (P < 0.01 vs basal) and endothelin-1 levels (P < 0.05 vs basal) within two hours and this was followed by an increase in ACTH (P < 0.01 vs basal) and cortisol (P < 0.05 vs basal) within three hours. Interleukin-6 levels increased two hours after interleukin-2 administration (P < 0.01 vs basal). Interleukin-2 had no detectable effect on AVP, blood pressure or heart rate.. Our data demonstrate that cytokines are able to activate the human hypothalamo-pituitary-adrenal axis in vivo. On the basis of the observed time kinetics and in connection with previous findings from in vitro and animal models, we conclude that endothelin may be a link between cytokines and corticotrophin-releasing hormone, most probably functioning as a cytokine-induced neuromodulator controlling pituitary functions.

Topics: Adrenocorticotropic Hormone; Adult; Arginine Vasopressin; Endothelin-1; Endothelins; Female; Humans; Hydrocortisone; Hypothalamo-Hypophyseal System; Interleukin-2; Interleukin-6; Kidney Neoplasms; Male; Melanoma; Middle Aged; Neoplasms; Pituitary-Adrenal System; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Protein Precursors; Stimulation, Chemical

The DNA-damaging agent camptothecin (CPT) and its analogs demonstrate clinical utility for the treatment of advanced solid tumors, and CPT-based nanopharmaceuticals are currently in clinical trials for advanced kidney cancer; however, little is known regarding the effects of CPT on hypoxia-inducible factor-2α (HIF-2α) accumulation and activity in clear cell renal cell carcinoma (ccRCC). Here we assessed the effects of CPT on the HIF/p53 pathway. CPT demonstrated striking inhibition of both HIF-1α and HIF-2α accumulation in von Hippel-Lindau (VHL)-defective ccRCC cells, but surprisingly failed to inhibit protein levels of HIF-2α-dependent target genes (VEGF, PAI-1, ET-1, cyclin D1). Instead, CPT induced DNA damage-dependent apoptosis that was augmented in the presence of pVHL. Further analysis revealed CPT regulated endothelin-1 (ET-1) in a p53-dependent manner: CPT increased ET-1 mRNA abundance in VHL-defective ccRCC cell lines that was significantly augmented in their VHL-expressing counterparts that displayed increased phosphorylation and accumulation of p53; p53 siRNA suppressed CPT-induced increase in ET-1 mRNA, as did an inhibitor of ataxia telangiectasia mutated (ATM) signaling, suggesting a role for ATM-dependent phosphorylation of p53 in the induction of ET-1. Finally, we demonstrate that p53 phosphorylation and accumulation is partially dependent on mTOR activity in ccRCC. Consistent with this result, pharmacological inhibition of mTORC1/2 kinase inhibited CPT-mediated ET-1 upregulation, and p53-dependent responses in ccRCC. Collectively, these data provide mechanistic insight into the action of CPT in ccRCC, identify ET-1 as a p53-regulated gene and demonstrate a requirement of mTOR for p53-mediated responses in this tumor type.

Topics: Apigenin; Apoptosis; Basic Helix-Loop-Helix Transcription Factors; Camptothecin; Carcinoma, Renal Cell; Cell Hypoxia; Cell Line, Tumor; DNA Damage; Endothelin-1; Gene Expression Regulation, Neoplastic; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Kidney Neoplasms; Mechanistic Target of Rapamycin Complex 1; Mechanistic Target of Rapamycin Complex 2; Multiprotein Complexes; Protein Biosynthesis; RNA, Messenger; Signal Transduction; TOR Serine-Threonine Kinases; Tumor Suppressor Protein p53

Cellular senescence, leading to cell death through prevention of regular cell renewal, is associated with the upregulation of the tumor suppressor gene p16(INK4a). While this mechanism has been described as leading to progressive nephron loss, p16(INK4a) upregulation in renal cell carcinoma has been linked to a disease-specific improved patient survival rate. While in both conditions endothelin-1 is also upregulated, the signaling pathway connecting ET-1 to p16(INK4a) has not been characterized until this study.. Cell culture, qRT-PCR, Western Blot, immunoprecipitation (IP), proximity ligation assay (PLA), and non-radioactive electrophoretic mobility shift assay (EMSA).. In malignant renal proximal tumor cells (Caki-1), an activation of p16(INK4a) and p21(waf1/cip1) was observed. An increased expression of E-26 transformation-specific (ETS) transcription factors was detectable. Using specific antibodies, a complex formation between ETS1 and extracellular signal-regulated kinase-2 (ERK2) was shown. A further complex partner was Mxi2. EMSA with supershift analysis for ETS1 and Mxi2 indicated the involvement of both factors in the protein-DNA interaction. After specifically blocking the endothelin receptors, ETS1 expression was significantly downregulated. However, the endothelin B receptor dependent downregulation was stronger than that of the A receptor. In contrast, primary proximal tubule cells showed a nuclear decrease after ET-1 stimulation. This indicates that other ETS members may be involved in the observed p16(INK4a) upregulation (as described in the literature).. ETS1, ERK2 and Mxi2 are important complex partners initiating increased p16(INK4a) and p21w(af1/cip1) activation in renal tumor cells.

Topics: Carcinoma, Renal Cell; Cell Line; Cell Line, Tumor; Cellular Senescence; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinase Inhibitor p21; Down-Regulation; Endothelin-1; Humans; Kidney Neoplasms; Kidney Tubules, Proximal; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 14; Proto-Oncogene Protein c-ets-1; Proto-Oncogene Proteins c-ets; Up-Regulation

We evaluated urinary endothelin (ET)-1-like Immunoreactivity (uET-1 L) excretion in Wilms tumor (WT) survivors and investigated its relationships with glomerular filtration rate (GFR) and effective renal plasma flow (ERPF). Glomerular hemodynamics were also assessed by Gomez formulae.. Seventeen WT survivors underwent renal sequential scintigraphy for residual kidney function determination including ERPF and GFR. Forty-five healthy individuals were selected as the control group. uET-1 L was measured by radioimmunoassay from the 24-h urine collection.. In WT survivors, uET-1 L excretion was significantly higher than in controls. Significant correlations were found between uET-1 L and ERPF and GFR. Cluster analysis, applied on uET-1 L, identified two different patient groups. Between them, GFR and ERPF were significantly different. No significant difference existed between the two clusters for age and sex, elapsed time from nephrectomy, treatment, or nephrectomy side. Applying Gomez formulae, significant difference was found for afferent and total renal resistance.. According to our results, uET-1 L seems to be a marker of glomerular injury in patients with renal mass loss revealing renal overload condition. The uET-1 L role in renal damage progression and hemodynamic glomerular worsening in nephrectomized patients should be proven by prospective long-term follow-up studies, even for potential ET-1 receptor antagonist therapeutic use.

Topics: Adolescent; Adult; Age of Onset; Biomarkers; Child; Child, Preschool; Endothelin-1; Female; Glomerular Filtration Rate; Humans; Infant; Kidney Neoplasms; Male; Renal Circulation; Survivors; Wilms Tumor; Young Adult

Topics: Blood Pressure; Carcinoma, Renal Cell; Endothelin-1; Gastrointestinal Stromal Tumors; Heart Rate; Humans; Hypertension; Indoles; Kidney Neoplasms; Models, Biological; Proteinuria; Pyrroles; Receptor Protein-Tyrosine Kinases; Renin; Sunitinib; Vascular Endothelial Growth Factor A

Endothelin-1 (ET-1) is a potent vasoconstrictor that has been shown to significantly impact many benign and malignant tissues by signaling through its two cognate receptors: ET(A) and ET(B). As ET-1 has a role in both normal and diseased kidney, we initiated studies to investigate endothelin axis expression and function in renal cell carcinoma (RCC). In this study, relatively high levels of ET-1 were detected in all six human RCC cell lines investigated. RT-PCR and Southern analyses revealed that all six RCC cell lines expressed ET(A) receptor mRNA, while 3/6 cell lines also expressed ET(B) mRNA. High affinity ET-1 binding occurred in all but one RCC cell line and quantitative RT-PCR demonstrated ET(A) mRNA expression in all six cell lines. Methylation of the ET(B) promoter (EDNRB) in 4/6 RCC cell lines was observed, suggesting a mechanism for repressed ET(B) expression. Moreover, methylation occurred in 32/48 of renal tumors and in 27/55 of histologically normal adjacent tissue samples studied, while no methylation was evident in any normal tissue isolated from nephrectomy or at autopsy. Functionally, ET-1 significantly inhibited paclitaxel-induced apoptosis in RCC cells through binding ET(A) with the ET-1 signaling mediated via the PI3-kinase/Akt pathway. Collectively, these data support the therapeutic targeting of the ET(A) receptor as a novel treatment strategy for RCC.

Topics: Androstadienes; Antineoplastic Agents, Phytogenic; Apoptosis; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Survival; Chromones; DNA Methylation; Endothelin A Receptor Antagonists; Endothelin-1; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation, Neoplastic; Humans; Immunoblotting; Kidney Neoplasms; Morpholines; Paclitaxel; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Promoter Regions, Genetic; Protein Binding; Proto-Oncogene Proteins c-akt; Receptor, Endothelin A; Receptor, Endothelin B; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Wortmannin

Endothelin-1 and its receptors ETAR and ETBR, commonly referred to as the Endothelin-axis, are emerging to play a role in cancer. The Endothelin-axis has been shown to be involved in proliferation, angiogenesis and metastasis in various human tumours. To assess the role of the Endothelin-axis in renal cell carcinoma, we analysed its expression in archival tumour tissue of 183 patients. Representative tumour blocks were selected for constructing a tissue microarray. Paraffin sections were assessed immunohistochemically using monoclonal and polyclonal antibodies for Endothelin-1, ETAR and ETBR. Staining intensities were analysed semiquantitatively and the results were correlated with various histopathologic factors. Overexpression of Endothelin-1, ETAR and ETBR was identified in 12.8%, 84.1% and 93.3% of cases, respectively. No association with pathological tumour stage and histologic grading was found. Papillary renal cell carcinomas expressed highly significantly more Endothelin-1 than clear cell renal cell carcinomas (34.5% vs. 6.7%, p<0.001), while there was no difference between ETAR- and ETBR-expression in these histologic subtypes. However, ETAR tended to be overexpressed in the subgroup of G3-tumours (p=0.044). Studies are underway assessing the role of the Endothelin-axis and its potential use as a molecular target in renal cell carcinoma.

Topics: Antibodies, Monoclonal; Apoptosis; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Proliferation; Endothelin-1; Endothelins; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; Kidney Neoplasms; Neovascularization, Pathologic; Oligonucleotide Array Sequence Analysis; Receptor, Endothelin A; Receptor, Endothelin B

The endothelin axis has been implicated in cancer growth, angiogenesis, and metastasis, but to the authors' knowledge the expression of endothelin genes has not been defined in renal cell carcinoma (RCC).. Tissue specimens were harvested from both normal and tumor-affected regions at the time of radical nephrectomy from 35 patients with RCC (22 with clear cell RCC [ccRCC] and 13 with papillary RCC [PRCC]). Real-time reverse transcriptase-polymerase chain reaction analysis determined the expression profile of the preproendothelins (PPET-1, PPET-2, and PPET-3), the endothelin receptors (ET(A) and ET(B)), and the endothelin-converting enzymes (ECE-1 and ECE-2).. PPET-1 was found to be up-regulated in ccRCC tumor specimens and down-regulated in PRCC tumor specimens. ET(A) was significantly down-regulated in PRCC tumor specimens. ECE-1 was expressed in all tissue specimens at comparable levels, with moderate but significant elevation in normal tissue specimens associated with PRCC. Of the other genes, PPET-2 and ET(B) were expressed in all tissue specimens and no differences were observed between tumor subtypes or tumor-affected and normal tissue specimens, whereas PPET-3 and ECE-2 were present in all tissue specimens but were barely detectable.. The endothelin axis was expressed differently in the two main subtypes of RCC and appeared to match macroscopic features commonly observed in these tumors (i.e., high expression of PPET-1 in hypervascular ccRCC contrasted against low PPET-1 and ET(A) expression in hypovascular PRCC). The presence of ECE-1 mRNA in these tissue specimens suggested that active endothelin ligands were present, indicating endothelin axis activity was elevated in ccRCC compared with normal kidney, but impaired in PRCC. The current study provided further evidence that it is not appropriate to consider ccRCC and PRCC indiscriminately in regard to treatment.

Topics: Aspartic Acid Endopeptidases; Biomarkers, Tumor; Carcinoma, Papillary; Carcinoma, Renal Cell; Endothelin-1; Endothelin-Converting Enzymes; Endothelins; Gene Expression Regulation, Neoplastic; Humans; Kidney Neoplasms; Metalloendopeptidases; Nephrectomy; Protein Precursors; Receptor, Endothelin A; Receptor, Endothelin B; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transcription, Genetic

The vasodilator hydralazine, used clinically in cardiovascular therapy, relaxes arterial smooth muscle by inhibiting accumulation of intracellular free Ca2+ via an unidentified primary target. Collagen prolyl hydroxylase is a known target of hydralazine. We therefore investigated whether inhibition of other members of this enzyme family, namely the hypoxia-inducible factor (HIF)-regulating O2-dependent prolyl hydroxylase domain (PHD) enzymes, could represent a novel mechanism of action. Hydralazine induced rapid and transient expression of HIF-1alpha and downstream targets of HIF (endothelin-1, adrenomedullin, haem oxygenase 1, and vascular endothelial growth factor [VEGF]) in endothelial and smooth muscle cells and induced endothelial cell-specific proliferation. Hydralazine dose-dependently inhibited PHD activity and induced nonhydroxylated HIF-1alpha, evidence for HIF stabilization specifically by inhibition of PHD enzyme activity. In vivo, hydralazine induced HIF-1alpha and VEGF protein in tissue extracts and elevated plasma VEGF levels. In sponge angiogenesis assays, hydralazine increased stromal cell infiltration and blood vessel density versus control animals. Thus, hydralazine activates the HIF pathway through inhibition of PHD activity and initiates a pro-angiogenic phenotype. This represents a novel mechanism of action for hydralazine and presents HIF as a potential target for treatment of ischemic disease.

Topics: Adrenomedullin; Angiogenesis Inducing Agents; Animals; Breast Neoplasms; Carcinoma; Carcinoma, Renal Cell; Cell Hypoxia; Cell Line, Tumor; Cells, Cultured; DNA-Binding Proteins; Dose-Response Relationship, Drug; Endothelial Cells; Endothelin-1; Enzyme Inhibitors; Gene Expression Regulation; Heme Oxygenase (Decyclizing); Heme Oxygenase-1; Humans; Hydralazine; Hypoxia-Inducible Factor 1; Hypoxia-Inducible Factor 1, alpha Subunit; Implants, Experimental; Kidney Neoplasms; Membrane Proteins; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Nude; Myocytes, Smooth Muscle; Neovascularization, Physiologic; Nuclear Proteins; Peptides; Procollagen-Proline Dioxygenase; Transcription Factors; Vascular Endothelial Growth Factor A; Vasodilator Agents

Evidence has accumulated showing that vasoactive peptides, such as endothelin-1, adrenomedullin and urotensin-II, are expressed in various kinds of tumour cells. In the present study, the expression of endothelin-1 and endothelin receptors was studied in eight human tumour cell lines: T98G (glioblastoma), IMR-32 and NB69 (neuroblastoma), BeWo (choriocarcinoma), SW-13 (adrenocortical carcinoma), DLD-1 (colonic carcinoma), HeLa (cervical carcinoma) and VMRC-RCW (renal carcinoma). Reverse transcriptase-PCR showed expression of endothelin-1 mRNA in seven out of the eight cell lines, the exception being BeWo cells. ET(A) receptor mRNA was expressed in T98G, IMR-32 and NB69 cells, but weakly in the other cells. ET(B) receptor mRNA was expressed in IMR-32, NB69 and BeWo cells, but only weakly in T98G and HeLa cells. Immunoreactive endothelin was detected in the culture media of six out of the eight cell lines, but not in that of IMR-32 or BeWo cells. Treatment of T98G cells with an anti-endothelin-1 antibody or an anti-adrenomedullin antibody for 24 h decreased cell numbers to approx. 84% and 90% of control respectively. Treatment with the ET(A) receptor antagonist BQ-610 (1 microM) significantly decreased cell number to about 90% of control, whereas the ET(B) receptor antagonist BQ-788 had no significant effect. On the other hand, exogenously added endothelin-1, adrenomedullin or urotensin-II (0.1 microM) had no significant effects on cell number. These results suggest that endothelin-1 acts as a paracrine or autocrine growth stimulator in tumours. The effect of endothelin-1 on tumour growth appears to be mediated by the ET(A) receptor.

Topics: Adrenal Cortex Neoplasms; Adrenomedullin; Antibodies, Monoclonal; Cell Division; Choriocarcinoma; Colonic Neoplasms; Endothelin Receptor Antagonists; Endothelin-1; Glioblastoma; Growth Substances; HeLa Cells; Humans; Kidney Neoplasms; Neuroblastoma; Oligopeptides; Peptides; Piperidines; Receptor, Endothelin A; Receptor, Endothelin B; RNA, Messenger; Tumor Cells, Cultured; Urotensins; Vasodilator Agents

Endothelin-1 (ET-1) is a potent vasoconstrictor and blood pressure modulator. Renin secretion from juxtaglomerular (JG) cells is crucial for blood pressure and electrolyte homeostasis and has been shown to be modulated by ET-1; however, the cellular and molecular mechanism of this regulation is not clear. The purpose of this study was to gain a better understanding of the cellular and molecular pathways activated by ET-1 by using a renin-producing cell line As4.1. ET-1 caused an increase in As4.1 cell intracelluar Ca(2+) concentration ([Ca(2+)](i)) mediated by the ET(A) receptor as its antagonist, BQ-123, abolished the response. The nitric oxide donor nitroprusside, but not 8-bromo-cGMP, reduced the time necessary for successive ET-1 responses. Endothelin-3 had no effect on [Ca(2+)](i). ET-1 dose dependently increased total inositol phosphates with an EC(50) of 2.1 nM. ET-1 reduced renin mRNA by 68% independently of changes in message decay. With the use of a renin-luciferase reporter system in As4.1 cells, ET-1 reduced luciferase activity by 51%, suggesting that renin gene transcription is directly modified by ET-1.

Topics: Animals; Calcium; Cell Line; Clone Cells; Cyclic GMP; Dose-Response Relationship, Drug; Endothelin Receptor Antagonists; Endothelin-1; Endothelin-3; Gene Expression Regulation; Inositol Phosphates; Juxtaglomerular Apparatus; Kidney; Kidney Neoplasms; Mice; Mice, Transgenic; Nitric Oxide Donors; Peptides, Cyclic; Receptor, Endothelin A; Renin; RNA, Messenger; Second Messenger Systems; Signal Transduction

Ectopic hormone secretion in tumor cells is here described as an amplification of hormone production already present in normal, nonendocrine tumor-originated tissue. This idea is tested on the available data regarding endothelin-1 (ET-1) secreting tumors. The endothelins are ubiquitous regulatory peptides produced by various tissues. The precursor cells of many tumor types secrete endothelins. ET-1 protein expression was detected in situ in all tested prostate cancers as well as in normal prostate tissue. The majority of hepatocellular carcinomas produce ET-1, while ET-1 is secreted by the normal hepatic stellate cells. Human breast cancer cells produce immunoreactive ET-1. Similar data exist for pancreatic tissue, the thyroid and large bowel. We can conclude that tumor cells might sustain endothelin secretions already present in the normal tumor-originated tissue. The model that is presented of the pseudoectopic hormone secretion consists of relations between a few parameters. The proportion of hormone-secreting tumors (Th) among all tumors (T) of that organ depends on the amount of the hormone-secreting cells (Ch) among all cells (C) susceptible to malignant transformation. The corrective factor (k) was introduced in the expression Th/T=Ch/C*k, to represent specific conditions altering the malignant transformation probability for a certain normal hormone-secreting cell. In prostate, breast and colon, the kvalue is predicted to be approximately 1, suggesting that ET-1-secreting normal cells are not more prone to the malignant transformation than their neighbours. In liver and pancreas, the incidence of ET-1-secreting tumors outnumbers the proportions of normal ET-1-secreting cells (k values >1). In these organs, normal ET-1-secreting cells seem more likely to turn malignant in comparison to their neighbours, perhaps due to their function, position and exposition to oncogenic factors, or even due to their ET-1 secretion. There are similar data for thyroid and adrenal glands. No ET-1 secretion was reported in kidney neoplasms. Normal renal ET-1 secreting cells might be less prone to turn malignant than other renal cells. Unlse the normal lung tissue, small cell lung cancers often secrete adrenocorticotrophic hormone (ACTH). The pancreatic islet cells do not secrete gastrin, but their tumors often do. Constant k would exceed 1 in both cases. We speculate that these tumors might originate from a small subset of cells with the described feature. Tumor cells s

Topics: Animals; Breast Neoplasms; Endothelin-1; Female; Hormones; Humans; Kidney Neoplasms; Male; Models, Biological; Neoplasms

To investigate the presence and the distribution of preproendothelin-1 (prepro-ET-1) mRNA in human kidney, eight human kidneys obtained at surgery from patients affected by localized renal tumors were studied. Northern blot analysis using a human prepro-ET-1 cDNA probe labeled with 32P showed the presence of a single band of approximately 2.3 kb that was present both in the renal cortex and medulla of all the kidneys studied. Densitometric analysis of hybridization signals demonstrated that prepro-ET-1 mRNA levels in the renal medulla were 2.2-fold higher than those in the renal cortex. The distribution of prepro-ET-1 mRNA in human kidney was investigated by in situ hybridization using a human prepro-ET-1 RNA probe labeled with 35S. The greatest density of prepro-ET-1 mRNA was observed in the renal medulla, where hybridization signal was demonstrated in vasa recta bundles and capillaries and in collecting ducts. By combining in situ hybridization with immunohistochemical detection of von Willebrand factor, we demonstrated that 93 +/- 2.5% of nontubular medullary cells containing prepro-ET-1 mRNA were endothelial cells. In the cortex, prepro-ET-1 mRNA was localized in the endothelial layer of arcuate and interlobular arteries and veins and in the endothelial cells of afferent arterioles. The results of the present study demonstrate that ET-1 gene expression is present in vascular and tubular structures of the human kidney. It is possible that ET-1 synthesized locally in the human kidney represents a local system affecting renal hemodynamics and functions through paracrine and/or autocrine actions on different renal structures.

Topics: Aged; Blotting, Northern; DNA Probes; Endothelin-1; Endothelins; Female; Gene Expression; Humans; In Situ Hybridization; Kidney; Kidney Cortex; Kidney Medulla; Kidney Neoplasms; Male; Middle Aged; Protein Precursors; RNA, Messenger; Sulfur Radioisotopes

The biosynthetic pathway of endothelin (ET)-2 was analyzed in cultured ACHN cells. In the supernatant, we detected three ET-2-related peptides, ET-2, big ET-2(1-38) and big ET-2(22-38). Phosphoramidon decreased the amount of ET-2 and increased that of big ET-2(1-38) dose-dependently. The amount of big ET-2(1-37) did not significantly change. These results suggest that big ET-2 is composed of 38 and not 37 amino acid residues, and that a putative ET-2-converting enzyme (ECE-2) should be classified as a phosphoramidon-sensitive neutral metalloprotease, bearing a resemblance to the putative ET-1-converting enzyme (ECE-1) in endothelial cells.

Topics: Adenocarcinoma; Aspartic Acid Endopeptidases; Chromatography, High Pressure Liquid; Cross Reactions; Endothelin-1; Endothelin-Converting Enzymes; Endothelins; Glycopeptides; Humans; Kidney Neoplasms; Metalloendopeptidases; Protein Precursors; Radioimmunoassay; Tumor Cells, Cultured

Analysis of culture medium of human renal adenocarcinoma cells ACHN by RP-HPLC suggested that the cells specifically secreted human endothelin-2 (ET-2). cDNAs encoding human ET-2 precursor were cloned from a cDNA library constructed with mRNA derived from the ACHN cells, and the nucleotide and deduced amino acid sequences were determined. The ET-2 cDNA was revealed to contain 1.3 kb and encode prepro-ET-2 protein consisting of 178 amino acid residues. The ET-like sequences found in the prepro-ET-1 and -ET-3 was conserved in this prepro-ET-2. The Northern blot analysis of mRNA suggested that the transcript of ET-2 gene was 1.4 kb. This is the first direct evidence that human ET-2 gene was expressed specifically in tumor cells.

Topics: Adenocarcinoma; Amino Acid Sequence; Base Sequence; Cell Line; Chromatography, High Pressure Liquid; DNA, Neoplasm; Endothelin-1; Endothelins; Gene Expression; Gene Library; Genes; Humans; Kidney Neoplasms; Molecular Sequence Data; Oligonucleotide Probes; Poly A; Protein Precursors; Protein Sorting Signals; Restriction Mapping; RNA; RNA, Messenger