Tuberin deficiency for an epigenetic modification in LAM cells from chylous of a TSC2 patient

Tuberous sclerosis complex (TSC) is an autosomal dominant syndrome caused by mutations in TSC1 or TSC2 genes which encode hamartin and tuberin, respectively. Lymphangioleiomyomatosis (LAM) can be sporadic or associated with TSC. From chylous of a patient affected by LAM associated to TSC, -smooth muscle-like cells were isolated. These cells bear a germline TSC2 mutation in exon 21 and an epigenetic alteration demonstrated by the expression of tuberin following incubation with chromatin-remodeling agents and the presence of a large amount of non-coding DNA with closed chromatin regions. Proliferation of LAM/TSC cells was EGF-dependent and blockade of EGFR caused cell death. Rapamycin caused a reduction in proliferation without leading to complete cell death. LAM/TSC cells have characteristics of stemness and the ability to grow independently from adhesion. About 80% of nonadherent cells remained viable and underwent an extremely low rate of proliferation. In adherent cells S6 and Erk phosphorylation were much higher than in nonadherent cells. Anti-EGFR antibodies and rapamycin reduced viability of adherent and nonadherent cells. Incubation with 5-azacytidine slightly increased the percentage of detaching cells. In conclusion, LAM/TSC cells are particularly important to study migration and invasiveness of pathological cells and better elucidate the pathogenesis of LAM and TSC.