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

Respiratory Research

Philip L Sannes¹  Donna R Newman¹  Huiying Zhang² 

[1] Department of Molecular Biomedical Sciences, Center for Comparative Molecular Translational Research, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606, USA;Department of Environmental and Molecular Toxicology, College of Agriculture and Life Sciences, North Carolina State University, Raleigh, NC, 27606, USA

关键词: Heparan sulfate proteoglycans;    Human endosulfatase 1;   

Others  :  796688
DOI  :  10.1186/1465-9921-13-69

 received in 2012-01-27, accepted in 2012-07-31,  发布年份 2012

【 摘 要 】

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.

【 授权许可】

   
2012 Zhang et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Clayton A, Thomas J, Thomas GJ, Davies M, Steadman R: Cell surface heparan sulfate proteoglycans control the response of renal interstitial fibroblasts to fibroblast growth factor-2. Kidney Int 2001, 59(6):2084-2094.
• [2]Fujiwara Y, Yamamoto C, Kaji T: Proteoglycans synthesized by cultured bovine aortic smooth muscle cells after exposure to lead, Lead selectively inhibits the synthesis of versican, a large chondroitin sulfate proteoglycan. Toxicology 2000, 154(1–3):9-19.
• [3]Iozzo RV: Heparan sulfate proteoglycans, Intricate molecules with intriguing functions. J Clin Invest 2001, 108(2):165-167.
• [4]Iozzo RV: Matrix proteoglycans, From molecular design to cellular function. Annu Rev Biochem 1998, 67:609-652.
• [5]Selva EM, Perrimon N: Role of heparan sulfate proteoglycans in cell signaling and cancer. Adv Cancer Res 2001, 83:67-80.
• [6]Ai X, Do AT, Lozynska O, Kusche-Gullberg M, Lindahl U, Emerson CP Jr: QSulf1 remodels the 6-O sulfation states of cell surface heparan sulfate proteoglycans to promote wnt signaling. J Cell Biol 2003, 162(2):341-351. PMCID, PMC2172803
• [7]Guimond S, Maccarana M, Olwin BB, Lindahl U, Rapraeger AC: Activating and inhibitory heparin sequences for FGF-2 (basic FGF). distinct requirements for FGF-1, FGF-2, and FGF-4. J Biol Chem 1993, 268(32):23906-23914.
• [8]Lai JP, Chien J, Strome SE, Staub J, Montoya DP, Greene EL, Smith DI, Roberts LR, Shridhar V: HSULF-1 modulates HGF-mediated tumor cell invasion and signaling in head and neck squamous carcinoma. Oncogene 2004, 23(7):1439-1447.
• [9]Lai JP, Chien JR, Moser DR, Staub JK, Aderca I, Montoya DP, Matthews TA, Nagorney DM, Cunningham JM, Smith DI, Greene EL, Shridhar V, Roberts LR: HSULF1 sulfatase promotes apoptosis of hepatocellular cancer cells by decreasing heparin-binding growth factor signaling. Gastroenterology 2004, 126(1):231-248.
• [10]Narita K, Staub J, Chien J, Meyer K, Bauer M, Friedl A, Ramakrishnan S, Shridhar V: HSULF-1 inhibits angiogenesis and tumorigenesis in vivo. Cancer Res 2006, 66(12):6025-6032.
• [11]Kimberley FC, van Bostelen L, Cameron K, Hardenberg G, Marquart JA, Hahne M, Medema JP: The proteoglycan (heparan sulfate proteoglycan) binding domain of APRIL serves as a platform for ligand multimerization and cross-linking. FASEB J 2009, 23(5):1584-1595.
• [12]Rapraeger AC, Krufka A, Olwin BB: Requirement of heparan sulfate for bFGF-mediated fibroblast growth and myoblast differentiation. Science 1991, 252(5013):1705-1708.
• [13]Lamanna WC, Frese MA, Balleininger M, Dierks T: Sulf loss influences N-, 2-O-, and 6-O-sulfation of multiple heparan sulfate proteoglycans and modulates fibroblast growth factor signaling. J Biol Chem 2008, 283(41):27724-27735.
• [14]Pye DA, Vives RR, Hyde P, Gallagher JT: Regulation of FGF-1 mitogenic activity by heparan sulfate oligosaccharides is dependent on specific structural features, Differential requirements for the modulation of FGF-1 and FGF-2. Glycobiology 2000, 10(11):1183-1192.
• [15]Lundin L, Larsson H, Kreuger J, Kanda S, Lindahl U, Salmivirta M, Claesson-Welsh L: Selectively desulfated heparin inhibits fibroblast growth factor-induced mitogenicity and angiogenesis. J Biol Chem 2000, 275(32):24653-24660.
• [16]Verhagen AP, Ramaekers FC, Aalders TW, Schaafsma HE, Debruyne FM, Schalken JA: Colocalization of basal and luminal cell-type cytokeratins in human prostate cancer. Cancer Res 1992, 52(22):6182-6187.
• [17]Dhoot GK, Gustafsson MK, Ai X, Sun W, Standiford DM, Emerson CP Jr: Regulation of wnt signaling and embryo patterning by an extracellular sulfatase. Science 2001, 293(5535):1663-1666.
• [18]Lai J, Chien J, Staub J, Avula R, Greene EL, Matthews TA, Smith DI, Kaufmann SH, Roberts LR, Shridhar V: Loss of HSULF-1 up-regulates heparin-binding growth factor signaling in cancer. J Biol Chem 2003, 278(25):23107-23117.
• [19]Lai JP, Sandhu DS, Shire AM, Roberts LR: The tumor suppressor function of human sulfatase 1 (SULF1) in carcinogenesis. J Gastrointest Cancer 2008, 39(1-4):149-158. PMCID, PMC2925118
• [20]Li J, Mo ML, Chen Z, Yang J, Li QS, Wang DJ, Zhang H, Ye YJ, Xu JP, Li HL, Zhang F, Zhou HM: HSULF-1 inhibits cell proliferation and invasion in human gastric cancer. Cancer Sci 2011, 102(10):1815-21.
• [21]Dobbs LG: Isolation and culture of alveolar type II cells. Am J Physiol 1990, 258(4 Pt 1):L134-L147.
• [22]Dai Y, Yang Y, MacLeod V, Yue X, Rapraeger AC, Shriver Z, Venkataraman G, Sasisekharan R, Sanderson RD: HSULF-1 and HSULF-2 are potent inhibitors of myeloma tumor growth in vivo. J Biol Chem 2005, 280(48):40066-40073.
• [23]Lai JP, Yu C, Moser CD, Aderca I, Han T, Garvey TD, Murphy LM, Garrity-Park MM, Shridhar V, Adjei AA, Roberts LR: SULF1 inhibits tumor growth and potentiates the effects of histone deacetylase inhibitors in hepatocellular carcinoma. Gastroenterology 2006, 130(7):2130-2144.
• [24]Chen Z, Fan JQ, Li J, Li QS, Yan Z, Jia XK, Liu WD, Wei LJ, Zhang FZ, Gao H, Xu JP, Dong XM, Dai J, Zhou HM: Promoter hypermethylation correlates with the HSULF-1 silencing in human breast and gastric cancer. Int J Cancer 2009, 124(3):739-744.
• [25]Presta M, Dell’Era P, Mitola S, Moroni E, Ronca R, Rusnati M: Fibroblast growth factor/fibroblast growth factor receptor system in angiogenesis. Cytokine Growth Factor Rev 2005, 16(2):159-178.
• [26]Li J, Kleeff J, Abiatari I, Kayed H, Giese NA, Felix K, Giese T, Buchler MW, Friess H: Enhanced levels of HSULF-1 interfere with heparin-binding growth factor signaling in pancreatic cancer. Mol Cancer 2005, 4(1):14. PMCID, PMC1087876 BioMed Central Full Text
• [27]Fannon M, Forsten KE, Nugent MA: Potentiation and inhibition of bFGF binding by heparin, A model for regulation of cellular response. Biochemistry 2000, 39(6):1434-1445.
• [28]Newman DR, Li CM, Simmons R, Khosla J, Sannes PL: Heparin affects signaling pathways stimulated by fibroblast growth factor-1 and -2 in type II cells. Am J Physiol Lung Cell Mol Physiol 2004, 287(1):L191-L200.
• [29]Miao HQ, Ishai-Michaeli R, Atzmon R, Peretz T, Vlodavsky I: Sulfate moieties in the subendothelial extracellular matrix are involved in basic fibroblast growth factor sequestration, dimerization, and stimulation of cell proliferation. J Biol Chem 1996, 271(9):4879-4886.
• [30]Yue X, Li X, Nguyen HT, Chin DR, Sullivan DE, Lasky JA: Transforming growth factor-beta1 induces heparan sulfate 6-O-endosulfatase 1 expression in vitro and in vivo. J Biol Chem 2008, 283(29):20397-20407. PMCID, PMC2459296
• [31]Ji W, Yang J, Wang D, Cao L, Tan W, Qian H, Sun B, Qian Q, Yin Z, Wu M, Su C: HSULF-1 gene exhibits anticancer efficacy through negatively regulating VEGFR-2 signaling in human cancers. PLoS One 2011, 6(8):e23274.
• [32]Balsara BR, Pei J, Mitsuuchi Y, Page R, Klein-Szanto A, Wang H, Unger M, Testa JR: Frequent activation of AKT in non-small cell lung carcinomas and preneoplastic bronchial lesions. Carcinogenesis 2004, 25(11):2053-2059.
• [33]Vicent S, Lopez-Picazo JM, Toledo G, Lozano MD, Torre W, Garcia-Corchon C, Quero C, Soria JC, Martin-Algarra S, Manzano RG, Montuenga LM: ERK1/2 is activated in non-small-cell lung cancer and associated with advanced tumours. Br J Cancer 2004, 90(5):1047-1052. PMCID, PMC2409626
• [34]Lu ZJ, Zhou Y, Song Q, Qin Z, Zhang H, Zhou YJ, Gou LT, Yang JL, Luo F: Periplocin inhibits growth of lung cancer in vitro and in vivo by blocking AKT/ERK signaling pathways. Cell Physiol Biochem 2010, 26(4–5):609-618.
• [35]Iacobuzio-Donahue CA, Maitra A, Olsen M, Lowe AW, van Heek NT, Rosty C, Walter K, Sato N, Parker A, Ashfaq R, Jaffee E, Ryu B, Jones J, Eshleman JR, Yeo CJ, Cameron JL, Kern SE, Hruban RH, Brown PO, Goggins M: Exploration of global gene expression patterns in pancreatic adenocarcinoma using cDNA microarrays. Am J Pathol 2003, 162(4):1151-1162.
• [36]Nawroth R, van Zante A, Cervantes S, McManus M, Hebrok M, Rosen SD: Extracellular sulfatases, elements of the wnt signaling pathway, positively regulate growth and tumorigenicity of human pancreatic cancer cells. PLoS One 2007, 2(4):e392. PMCID, PMC1849966
• [37]Bret C, Moreaux J, Schved JF, Hose D, Klein B: SULFs in human neoplasia, Implication as progression and prognosis factors. J Transl Med 2011, 9(1):72. BioMed Central Full Text
• [38]Sahota AP, Dhoot GK: A novel SULF1 splice variant inhibits wnt signalling but enhances angiogenesis by opposing SULF1 activity. Exp Cell Res 2009, 315(16):2752-2764.
• [39]Han CH, Huang YJ, Lu KH, Liu Z, Mills GB, Wei Q, Wang LE: Polymorphisms in the SULF1 gene are associated with early age of onset and survival of ovarian cancer. J Exp Clin Cancer Res 2011, 30:5. PMCID, PMC3025876 BioMed Central Full Text