HSULF-1 inhibits ERK and AKT signaling and decreases cell viability in vitro in human lung epithelial cells

BACKGROUND: Heparan sulfate proteoglycans (HSPGs) modulate the binding and activation of signaling pathways of specific growth factors, such as fibroblast growth factor-2 (FGF-2). Human endosulfatase 1 (HSULF-1) is an enzyme that selectively removes 6-O sulfate groups from HS side chains and alter their level and pattern of sulfation and thus biological activity. It is known that HSULF-1 is expressed at low levels in some cancer cell lines and its enhanced expression can inhibit cancer cell growth or induce apoptosis, but the mechanism(s) involved has not been identified. METHODS: HSULF-1 mRNA expression was assessed in five normal cells (primary human lung alveolar type 2 (hAT2) cells, adult lung fibroblasts (16Lu), fetal lung fibroblasts (HFL), human bronchial epithelial cells (HBE), and primary human lung fibroblasts (HLF)) and five lung cancer cell lines (A549, H292, H1975, H661, and H1703) using quantitative real time polymerase chain reaction (qRT-PCR). H292 and hAT2 cells over-expressing HSULF-1 were analyzed for cell viability, apoptosis, and ERK/Akt signaling, by MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, TUNEL (Terminal deoxynucleotidyl transferase dUTP nick end labeling) assay, and Western Blot, respectively. Apoptosis pathway activation was confirmed by PCR array in hAT2, H292, and A549 cells. RESULTS: HSULF-1 was expressed at a significantly lower level in epithelial cancer cell lines compared to normal cells. Infection with recombinant adenovirus for HSULF-1 over-expression resulted in decreased cell viability in H292 cells, but not in normal hAT2 cells. HSULF-1 over-expression induced apoptosis in H292 cells, but not in hAT2 cells. In addition, apoptosis pathways were activated in both H292 and A549 cells, but not in hAT2 cells. HSULF-1 over-expression reduced ERK and Akt signaling activation in H292 cells, which further demonstrated its inhibitory effects on signaling related to proliferation. CONCLUSIONS: These results indicate that HSULF-1 is expressed at lower levels in H292 lung cancer cells than in normal human alveolar cells and that its over-expression reduced cell viability in H292 cells by inducing apoptotic pathways, at least in part by inhibiting ERK/Akt signaling. We hypothesize that HSULF-1 plays important roles in cancer cells and functions to modify cell signaling, inhibit cancer proliferation, and promote cancer cell death.