Tissue-Specific Renin–Angiotensin System in Pulmonary Lymphangioleiomyomatosis

Julio Valencia

Gustavo Pacheco-Rodriguez

Adriana Carmona

Janina Xavier

Patrick Bruneval

William Riemenschneider

Pulmonary-Critical Care Medicine Branch and Pathology Core, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland; Biophysics Department, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, Brazil; and Laboratoire d'Anatomie Pathologique, Hôpital Européen Georges Pompidou, Paris, France

^Deceased.

Correspondence and requests for reprints should be addressed to Joel Moss, M.D, Ph.D., Chief, Pulmonary–Critical Care Medicine Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Building 10/Room 6D05 MSC-1590, Bethesda, MD 20892-1590. E-Mail: vog.hin.iblhn@jssom

Received 2005 Oct 14; Accepted 2006 Jan 19.

Abstract

Lymphangioleiomyomatosis (LAM), a multisystem disease found in middle-aged women, is characterized by cystic lung destruction and abdominal tumors (e.g., angiomyolipomas, lymphangioleimyomas), resulting from proliferation of abnormal-appearing, smooth muscle–like cells (LAM cells). The LAM cells, in combination with other cells, form nodular structures within the lung interstitium and in the walls of the cysts. LAM cells contain mutations in the tuberous sclerosis complex TSC1 and/or TSC2 genes, which lead to dysregulation of the mammalian target of rapamycin, affecting cell growth and proliferation. Proliferation and migration of vascular smooth muscle cells and production of angiogenic factors are regulated, in part, by angiotensin II. To determine whether a LAM-specific renin–angiotensin system might play a role in the pathogenesis of LAM, we investigated the expression of genes and gene products of this system in LAM nodules. mRNA for angiotensinogen was present in RNA isolated by laser-captured microdissection from LAM nodules. Angiotensin I–converting enzyme and chymase-producing mast cells were present within the LAM nodules. We detected renin in LAM cells, as determined by the presence of mRNA and immunohistochemistry. Angiotensin II type 1 and type II receptors were identified in LAM cells by immunohistochemistry and immunoblotting of microdissected LAM nodules. Angiotensin II is localized in cells containing α–smooth muscle actin (LAM cells). A LAM-specific renin–angiotensin system appears to function within the LAM nodule as an autocrine system that could promote LAM cell proliferation and migration, and could represent a pharmacologic target.

Keywords: angiotensin II, lymphangioleiomyomatosis, mammalian target of rapamycin, smooth muscle cells, tuberous sclerosis complex

Abstract

Lymphangioleiomyomatosis (LAM) is a rare multisystem disease found primarily in middle-aged women. Clinically, pulmonary LAM is characterized by recurrent pneumothoraces, pleural effusions, and progressive cystic destruction of the lung, leading, in some patients, to respiratory failure. These clinical and radiographic findings result from the proliferation of abnormal-appearing, smooth muscle–like cells (LAM cells) that form nodules in the pulmonary interstitium and in the walls of the cysts (1, 2). The presence of α–smooth muscle actin (SMA), smooth muscle myosin heavy chains I and II, desmin, vimentin, calponin, and h-caldesmon in LAM cells is consistent with the smooth muscle phenotype of the LAM cell. Some LAM cells, however, react with a monoclonal antibody, HMB-45, which recognizes a protein, gp100, found in melanosomal structures in melanocytes and related cells (3–5). Thus, LAM cells have a mixed smooth muscle and melanocytic phenotype.

Proliferation of the neoplastic LAM cells appears to result from mutations in the tuberous sclerosis complex (TSC) TSC1 and/or TSC2 genes (4), leading to dysregulation of the mammalian target of rapamycin (5). The mechanism(s) involved in LAM metastasis (6, 7) and formation of lymphatics (8) within the LAM nodule are less-well characterized. The renin–angiotensin system has been described as a regulator of smooth muscle cell proliferation (9, 10) and migration (11, 12), as well as a central player in vascular remodeling and angiogenesis (13). Most of these actions have been attributed to production of the peptide angiotensin II, which activates G protein–coupled, seven-transmembrane helix angiotensin II type 1 or type II receptors (AT₁R and AT₂R) (14). Whereas AT₁R activates the Janus kinase (JAK)/signal transducers and activators of transcription (STAT) pathway, AT₂R inhibits STAT activation via tyrosine and serine phosphorylation through JAK and extracellular signal–regulated kinase pathways, respectively (15).

Angiotensin II is generated through a series of enzyme-regulated events, which follows the initial cleavage of angiotensinogen by renin to form angiotensin I (16), which is converted subsequently to angiotensin II by angiotensin I–converting enzyme (ACE). Other enzymes, such as chymase, a chymostatin-inhibited serine protease, can also covert angiotensin I to angiotensin II (16, 17). In human lung, chymase is found in mast cells, which have been implicated in smooth muscle cell proliferation (18).

Given the smooth muscle phenotype of LAM cells, we questioned whether a local renin–angiotensin system is present in LAM cells. We describe here the presence of angiotensinogen, angiotensin II, ACE, renin, and receptors for angiotensin II in LAM cells, as well as the presence of degranulated mast cells within LAM nodules. These findings are consistent with participation of a renin–angiotensin system in the neoplastic phenotype of LAM cells and in LAM pathogenesis.

Acknowledgments

This work was initiated in the laboratory of Dr. Victor J. Ferrans. The authors are deeply indebted to him and dedicate this article to his memory. They thank Dr. Martha Vaughan for helpful discussions and critical review of the manuscript, and Michael Spencer for his help with the artwork. They thank the LAM foundation and the Tuberous Sclerosis Alliance for their assistance in recruiting patients for our studies.

Acknowledgments

Notes

This work was supported by the Intramural Research Program of the National Institutes of Health, National Heart, Lung, and Blood Institute.

Originally Published in Press as DOI: 10.1165/rcmb.2005-0387OC on February 10, 2006

Conflict of Interest Statement: J.C.V. does not have a financial relationship with a commercial entity that has an interest in the subject of this manuscript. G.P.-R. does not have a financial relationship with a commercial entity that has an interest in the subject of this manuscript. A.K.C. does not have a financial relationship with a commercial entity that has an interest in the subject of this manuscript. J.X. does not have a financial relationship with a commercial entity that has an interest in the subject of this manuscript. P.B. receives an annual fee of €10,000 from Guerbet group (radiology contrast products company in France) for assessment of new products for IRM in experimental preclinical models of atherosclerosis. W.K.R. does not have a financial relationship with a commercial entity that has an interest in the subject of this manuscript. Y.I. is a participant in the Senior Fellow Program from the Oak Ridge Institute for Science and Education. Z.-X.Y. does not have a financial relationship with a commercial entity that has an interest in the subject of this manuscript. V.J.F. is deceased. J.M. does not have a financial relationship with a commercial entity that has an interest in the subject of this manuscript.

Notes

This work was supported by the Intramural Research Program of the National Institutes of Health, National Heart, Lung, and Blood Institute.

Originally Published in Press as DOI: 10.1165/rcmb.2005-0387OC on February 10, 2006

References

1. Pacheco-Rodriguez G, Kristof AS, Stevens LA, Zhang Y, Crooks D, Moss J. Giles F. Filley Lecture: genetics and gene expression in lymphangioleiomyomatosis. Chest 2002;121:56S–60S. [[PubMed]
2. Ferrans VJ, Yu ZX, Nelson WK, Valencia JC, Tatsuguchi A, Avila NA, Riemenschn W, Matsui K, Travis WD, Moss JLymphangioleiomyomatosis (LAM): a review of clinical and morphological features. J Nippon Med Sch 2000;67:311–329. [[PubMed][Google Scholar]
3. Dell'Angelica ECMelanosome biogenesis: shedding light on the origin of an obscure organelle. Trends Cell Biol 2003;13:503–506. [[PubMed][Google Scholar]
4. Carsillo T, Astrinidis A, Henske EPMutations in the tuberous sclerosis complex gene TSC2 are a cause of sporadic pulmonary lymphangioleiomyomatosis. Proc Natl Acad Sci USA 2000;97:6085–6090. [Google Scholar]
5. Goncharova EA, Goncharov DA, Eszterhas A, Hunter DS, Glassberg MK, Yeung RS, Walker CL, Noonan D, Kwiatkowski DJ, Chou MM, et al. Tuberin regulates p70 S6 kinase activation and ribosomal protein S6 phosphorylation: a role for the TSC2 tumor suppressor gene in pulmonary lymphangioleiomyomatosis (LAM). J Biol Chem 2002;277:30958–30967. [[PubMed]
6. Karbowniczek M, Astrinidis A, Balsara BR, Testa JR, Lium JH, Colby TV, McCormack FX, Henske EPRecurrent lymphangiomyomatosis after transplantation: genetic analyses reveal a metastatic mechanism. Am J Respir Crit Care Med 2003;167:976–982. [[PubMed][Google Scholar]
7. Crooks DM, Pacheco-Rodriguez G, DeCastro RM, McCoy JP Jr, Wang JA, Kumaki F, Darling T, Moss JMolecular and genetic analysis of disseminated neoplastic cells in lymphangioleiomyomatosis. Proc Natl Acad Sci USA 2004;101:17462–17467. [Google Scholar]
8. Kumasaka T, Seyama K, Mitani K, Sato T, Souma S, Kondo T, Hayashi S, Minami M, Uekusa T, Fukuchi Y, et al. Lymphangiogenesis in lymphangioleiomyomatosis: its implication in the progression of lymphangioleiomyomatosis. Am J Surg Pathol 2004;28:1007–1016. [[PubMed]
9. Berk BCVascular smooth muscle growth: autocrine growth mechanisms. Physiol Rev 2001;81:999–1030. [[PubMed][Google Scholar]
10. Meloche S, Pelletier S, Servant MJFunctional cross-talk between the cyclic AMP and Jak/STAT signaling pathways in vascular smooth muscle cells. Mol Cell Biochem 2000;212:99–109. [[PubMed][Google Scholar]
11. Kraemer RRegulation of cell migration in atherosclerosis. Curr Atheroscler Rep 2000;2:445–452. [[PubMed][Google Scholar]
12. Mifune M, Ohtsu H, Suzuki H, Nakashima H, Brailoiu E, Dun NJ, Frank GD, Inagami T, Higashiyama S, Thomas WG, et al. G protein coupling and second messenger generation are indispensable for metalloprotease-dependent, heparin-binding epidermal growth factor shedding through angiotensin II type-1 receptor. J Biol Chem 2005;280:26592–26599. [[PubMed]
13. Escobar E, Rodriguez-Reyna TS, Arrieta O, Sotelo JAngiotensin II, cell proliferation and angiogenesis regulator: biologic and therapeutic implications in cancer. Curr Vasc Pharmacol 2004;2:385–399. [[PubMed][Google Scholar]
14. Thomas WG, Qian HArresting angiotensin type 1 receptors. Trends Endocrinol Metab 2003;14:130–136. [[PubMed][Google Scholar]
15. de Gasparo M, Catt KJ, Inagami T, Wright JW, Unger T. International union of pharmacology. XXIII: the angiotensin II receptors. Pharmacol Rev 2000;52:415–472. [[PubMed]
16. Belova LAAngiotensin II–generating enzymes. Biochemistry (Mosc) 2000;65:1337–1345. [[PubMed][Google Scholar]
17. Takai S, Jin D, Sakaguchi M, Miyazaki MChymase-dependent angiotensin II formation in human vascular tissue. Circulation 1999;100:654–658. [[PubMed][Google Scholar]
18. Page S, Ammit AJ, Black JL, Armour CLHuman mast cell and airway smooth muscle cell interactions: implications for asthma. Am J Physiol Lung Cell Mol Physiol 2001;281:L1313–L1323. [[PubMed][Google Scholar]
19. Matsui K, Takeda K, Yu ZX, Valencia J, Travis WD, Moss J, Ferrans VJDownregulation of estrogen and progesterone receptors in the abnormal smooth muscle cells in pulmonary lymphangioleiomyomatosis following therapy: an immunohistochemical study. Am J Respir Crit Care Med 2000;161:1002–1009. [[PubMed][Google Scholar]
20. Valencia JC, Matsui K, Bondy C, Zhou J, Rasmussen A, Cullen K, Yu ZX, Moss J, Ferrans VJDistribution and mRNA expression of insulin-like growth factor system in pulmonary lymphangioleiomyomatosis. J Investig Med 2001;49:421–433. [[PubMed][Google Scholar]
21. Rakugi H, Okamura A, Kamide K, Ohishi M, Sasamura H, Morishita R, Higaki J, Ogihara TRecognition of tissue- and subtype-specific modulation of angiotensin II receptors using antibodies against AT1 and AT2 receptors. Hypertens Res 1997;20:51–55. [[PubMed][Google Scholar]
22. Netzel-Arnett S, Mallya SK, Nagase H, Birkedal-Hansen H, Van Wart HEContinuously recording fluorescent assays optimized for five human matrix metalloproteinases. Anal Biochem 1991;195:86–92. [[PubMed][Google Scholar]
23. Araujo MC, Melo RL, Cesari MH, Juliano MA, Juliano L, Carmona AKPeptidase specificity characterization of C- and N-terminal catalytic sites of angiotensin I–converting enzyme. Biochemistry 2000;39:8519–8525. [[PubMed][Google Scholar]
24. Friedland J, Silverstein EA sensitive fluorimetric assay for serum angiotensin-converting enzyme. Am J Clin Pathol 1976;66:416–424. [[PubMed][Google Scholar]
25. Hardman JA, Hort YJ, Catanzaro DF, Tellam JT, Baxter JD, Morris BJ, Shine JPrimary structure of the human renin gene. DNA 1984;3:457–468. [[PubMed][Google Scholar]
26. Matsumoto Y, Horiba K, Usuki J, Chu SC, Ferrans VJ, Moss JMarkers of cell proliferation and expression of melanosomal antigen in lymphangioleiomyomatosis. Am J Respir Cell Mol Biol 1999;21:327–336. [[PubMed][Google Scholar]
27. Furuta H, Guo DF, Inagami TMolecular cloning and sequencing of the gene encoding human angiotensin II type 1 receptor. Biochem Biophys Res Commun 1992;183:8–13. [[PubMed][Google Scholar]
28. Tsuzuki S, Ichiki T, Nakakubo H, Kitami Y, Guo DF, Shirai H, Inagami TMolecular cloning and expression of the gene encoding human angiotensin II type 2 receptor. Biochem Biophys Res Commun 1994;200:1449–1454. [[PubMed][Google Scholar]
29. Servant G, Dudley DT, Escher E, Guillemette GThe marked disparity between the sizes of angiotensin type 2 receptors from different tissues is related to different degrees of N-glycosylation. Mol Pharmacol 1994;45:1112–1118. [[PubMed][Google Scholar]
30. Paxton WG, Runge M, Horaist C, Cohen C, Alexander RW, Bernstein KEImmunohistochemical localization of rat angiotensin II AT1 receptor. Am J Physiol 1993;264:F989–F995. [[PubMed][Google Scholar]
31. Ciuffo GM, Johren O, Egidy G, Heemskerk FM, Saavedra JMHeterogeneity of rat angiotensin II AT2 receptor. Adv Exp Med Biol 1996;396:189–197. [[PubMed][Google Scholar]
32. Saridogan E, Djahanbakhch O, Puddefoot JR, Demetroulis C, Dawda R, Hall AJ, Vinson GPType 1 angiotensin II receptors in human endometrium. Mol Hum Reprod 1996;2:659–664. [[PubMed][Google Scholar]
33. AbdAlla S, Lother H, Quitterer UAT1-receptor heterodimers show enhanced G-protein activation and altered receptor sequestration. Nature 2000;407:94–98. [[PubMed][Google Scholar]
34. Marrero MB, Schieffer B, Li B, Sun J, Harp JB, Ling BNRole of Janus kinase/signal transducer and activator of transcription and mitogen-activated protein kinase cascades in angiotensin II– and platelet-derived growth factor–induced vascular smooth muscle cell proliferation. J Biol Chem 1997;272:24684–24690. [[PubMed][Google Scholar]
35. El-Hashemite N, Kwiatkowski DJInterferon-γ-Jak-Stat signaling in pulmonary lymphangioleiomyomatosis and renal angiomyolipoma: a potential therapeutic target. Am J Respir Cell Mol Biol 2005;33:227–230. [Google Scholar]
36. Gallagher PE, Li P, Lenhart JR, Chappell MC, Brosnihan KBEstrogen regulation of angiotensin-converting enzyme mRNA. Hypertension 1999;33:323–328. [[PubMed][Google Scholar]
37. Brosnihan KB, Senanayake PS, Li P, Ferrario CMBi-directional actions of estrogen on the renin–angiotensin system. Braz J Med Biol Res 1999;32:373–381. [[PubMed][Google Scholar]
38. Proudler AJ, Ahmed AI, Crook D, Fogelman I, Rymer JM, Stevenson JCHormone replacement therapy and serum angiotensin-converting-enzyme activity in postmenopausal women. Lancet 1995;346:89–90. [[PubMed][Google Scholar]
39. Oelkers WKEffects of estrogens and progestogens on the renin–aldosterone system and blood pressure. Steroids 1996;61:166–171. [[PubMed][Google Scholar]
40. Inoue Y, King TE Jr, Barker E, Daniloff E, Newman LSBasic fibroblast growth factor and its receptors in idiopathic pulmonary fibrosis and lymphangioleiomyomatosis. Am J Respir Crit Care Med 2002;166:765–773. [[PubMed][Google Scholar]
41. Silver RB, Reid AC, Mackins CJ, Askwith T, Schaefer U, Herzlinger D, Levi RMast cells: a unique source of renin. Proc Natl Acad Sci USA 2004;101:13607–13612. [Google Scholar]
42. Molteni A, Moulder JE, Cohen EF, Ward WF, Fish BL, Taylor JM, Wolfe LF, Brizio-Molteni L, Veno PControl of radiation-induced pneumopathy and lung fibrosis by angiotensin-converting enzyme inhibitors and an angiotensin II type 1 receptor blocker. Int J Radiat Biol 2000;76:523–532. [[PubMed][Google Scholar]
43. Patella V, Marino I, Lamparter B, Arbustini E, Adt M, Marone GHuman heart mast cells: isolation, purification, ultrastructure, and immunologic characterization. J Immunol 1995;154:2855–2865. [[PubMed][Google Scholar]
44. Beil WJ, Pammer J. In situ detection of the mast cell proteases chymase and tryptase in human lung tissue using light and electron microscopy. Histochem Cell Biol 2001;116:483–493. [[PubMed]
45. Deshayes F, Nahmias CAngiotensin receptors: a new role in cancer? Trends Endocrinol Metab 2005;16:293–299. [[PubMed][Google Scholar]
46. Inoki K, Corradetti MN, Guan KLDysregulation of the TSC–mTOR pathway in human disease. Nat Genet 2005;37:19–24. [[PubMed][Google Scholar]