ABSTRACT
Doctorate School in Biochemical, Nutritional and Metabolic Sciences
Dept. of Medicine, Surgery and Dentistry
Università degli Studi di Milano, Italy
LAM is a rare and progressive disease characterized by widespread proliferation of abnormal smooth muscle-like cells (LAM cells)(1). LAM cells cause cystic destruction of lung parenchyma, abdominal tumours (angiomyolipoma, AML) and infiltration of axial lymphatics in torax and abdomen (adenopathy and lymphangioleiomyoma)(1). LAM occurs sporadically or in association with tuberous sclerosis complex (TSC), an inherited disorder with variable penetrance, which results from mutations in TSC1 or TSC2 genes. TSC1 or TSC2 genes, encoding hamartin an tuberin respectively, regulate mammalian target of rapamycin (mTOR)(1). LAM affects primarly women of child-bearing age and the mechanisms causing the disease are not yet clarified. To explain the multisystemic clinical manifestations of LAM a metastatic model has been proposed where lymphangiogenesis has a key role in the development and progression of the disease(2). An experimental model of LAM is needed to study the pathological mechanism causing LAM and it may help to explain how LAM cells migrate from tissue to tissue and to develop a pharmacological approach. We recently isolated and characterized α-actin positive smooth muscle cells from chylous of a patient affected by LAM/TSC (LAM/TSC cells). These circulating cells showed reactivity to HMB45 and CD44v6 antibodies, markers of TSC and LAM, and bear a germline TSC2 mutation in exon 21. Like TSC2 smooth muscle cells previously isolated (TSC2-/- and TSC2-/meth ASM cells)(3-4), LAM/TSC cells from chylous required epidermal growth factor (EGF) to proliferate and the blockade of EGF receptor (EGFR) caused progressive cell death. To better study LAM pathogenesis we developed a procedure for a quick invasion of the respiratory system by endonasally administrating LAM/TSC cells. LAM/TSC cells were administrated in immunodeficient female nude mice (nu/nu Hsd: athymic nude mice, 3 weeks old) and following 26 weeks anti-EGFR antibody and rapamycin were intraperitoneally injected 2 times a week for 4 weeks. 30 weeks after endonasal administration LAM/TSC cells were detected in lungs, lymph nodes and uterus. In lung parenchyma LAM/TSC cells caused cystic destruction with emphysematous-like picture such as in LAM patient lungs. This lesion was reverted by anti-EGFR antibody, while rapamycin was less effective and caused hemoptysis. In lungs blood vessel number was increased and a strong S6 phosphorylation were detected. Using LYVE-1 staining, a significant increase of lymphatic vessel density (LVD) was observed in lungs of animals that received LAM/TSC cells administration suggesting a possible correlation between LAM/TSC cells and lymphangiogenesis. LVD decreased following anti-EGFR antibody and rapamycin treatments. In lymph nodes, LAM/TSC cells promoted a lymphatic vessel invasion, as showed by PROX-1-reactivity. Anti-EGFR antibody and rapamycin treatments decreased lymphatic vessels. Unlike to lungs, blood vessels in lymph nodes were not affected by LAM/TSC cells administration, as shown by CD31 immunoreactivity. Finally, LAM/TSC cells were detected in uteri where they promoted a significant increase of cells with high levels of estrogen (ER) and progesterone receptors (PR). Pharmacological treatments decreased ER and PR expressions. Our data show that endonasal administration of cells isolated from chylous of LAM/TSC patient developed a mouse LAM model. LAM/TSC cells invade lungs, lymph nodes and uterus causing LAM-like lesions. Anti-EGFR antibody is more effective than rapamycin in promoting lung restauration and reducing lymphangiogenesis suggesting that it may represent a useful pharmacological approch for LAM therapy.
- Taveira-daSilva AM et al. (2006) – Cancer Control vol 13, 276-85
- Seyama K et al (2010) – Lymphat Res Biol vol 8(1), 21-31
- Lesma E et al (2005) – Am J Pathol vol 167(4), 1093-103
- Lesma E et al (2009) –Am J Pathol vol 174(6), 2150-9