Suppressing Wnt Signaling by the Hedgehog Pathway through sFRP-1

The hedgehog (Hh) signaling pathway is essential for embryonic development and carcinogenesis. Activation of Hh signaling has been identified in several types of gastrointestinal cancers, including esophageal, gastric, pancreatic, and liver cancers. Several recent studies suggest that Hh signaling activation can inhibit Wnt signaling. However, the molecular basis underlying this inhibition remains unclear. As transcription factors in the Hh signaling pathway, Gli molecules transform cells in culture, and their expression are associated with cancer development. Here we report that expression of a secreted frizzled-related protein-sFRP-1 in mouse embryonic fibroblasts is dependent on Gli1 and Gli2. In human gastric cancer cells, inhibition of Hh signaling reduces the level of sFRP-1 transcript, whereas ectopic expression of Gli1 increases the level of sFRP-1 transcript. Results from chromatin immunoprecipitation indicate that Gli1 is involved in transcriptional regulation of sFRP-1. In 293 cells with Gli1 expression, Wnt-1-mediated β-catenin accumulation in the cytosol and DKK1 expression are all abrogated, which can be reversed by inhibiting sFRP-1 expression. Furthermore, while SIIA cells do not respond to Wnt-1-conditioned medium, inhibition of Hh signaling by smoothened (SMO) antagonist KAAD-cyclopamine (keto-N-aminoethylaminocaproyldihydrocinnamoylcyclopamine) leads to Wnt1-mediated β-catenin accumulation in the cytosol. These data indicate that sFRP-1, a target gene of the hedgehog pathway, is involved in cross-talk between the hedgehog pathway and the Wnt pathway. The hedgehog (Hh) signaling pathway is essential for embryonic development and carcinogenesis. Activation of Hh signaling has been identified in several types of gastrointestinal cancers, including esophageal, gastric, pancreatic, and liver cancers. Several recent studies suggest that Hh signaling activation can inhibit Wnt signaling. However, the molecular basis underlying this inhibition remains unclear. As transcription factors in the Hh signaling pathway, Gli molecules transform cells in culture, and their expression are associated with cancer development. Here we report that expression of a secreted frizzled-related protein-sFRP-1 in mouse embryonic fibroblasts is dependent on Gli1 and Gli2. In human gastric cancer cells, inhibition of Hh signaling reduces the level of sFRP-1 transcript, whereas ectopic expression of Gli1 increases the level of sFRP-1 transcript. Results from chromatin immunoprecipitation indicate that Gli1 is involved in transcriptional regulation of sFRP-1. In 293 cells with Gli1 expression, Wnt-1-mediated β-catenin accumulation in the cytosol and DKK1 expression are all abrogated, which can be reversed by inhibiting sFRP-1 expression. Furthermore, while SIIA cells do not respond to Wnt-1-conditioned medium, inhibition of Hh signaling by smoothened (SMO) antagonist KAAD-cyclopamine (keto-N-aminoethylaminocaproyldihydrocinnamoylcyclopamine) leads to Wnt1-mediated β-catenin accumulation in the cytosol. These data indicate that sFRP-1, a target gene of the hedgehog pathway, is involved in cross-talk between the hedgehog pathway and the Wnt pathway. Hedgehog (Hh) 2The abbreviations used are: Hh, hedgehog; SMO, smoothened; KAAD-cyclopamine, keto-N-aminoethylaminocaproyldihydrocinnamoylcyclopamine; MEF, mouse embryonic fibroblast; siRNA, small interfering RNA; RT, reverse transcription; ChIP, chromatin immunoprecipitation; PIPES, 1,4-piperazinediethanesulfonic acid. proteins are a group of secreted proteins whose active forms are derived from a unique protein cleavage process and at least two post-translational modifications (1Taipale J. Beachy P.A. Nature. 2001; 411: 349-354Crossref PubMed Scopus (1189) Google Scholar). Secreted Hh molecules bind to the receptor patched, thereby alleviating patched-mediated suppression of smoothened (SMO) (2Pasca di Magliano M. Hebrok M. Nat. Rev. Cancer. 2003; 3: 903-911Crossref PubMed Scopus (730) Google Scholar, 3Hooper J.E. Scott M.P. Nat. Rev. Mol. Cell. Biol. 2005; 6: 306-317Crossref PubMed Scopus (688) Google Scholar). Expression of sonic hedgehog appears to stabilize SMO protein possibly through post-translational modification of SMO. In Drosophila, SMO stabilization triggers complex formation with Costal-2, Fused, and Gli homologue cubitus interruptus, which prevents cubitus interruptus degradation and formation of a transcriptional repressor (4Lum L. Zhang C. Oh S. Mann R.K. von Kessler D.P. Taipale J. Weis-Garcia F. Gong R. Wang B. Beachy P.A. Mol. Cell. 2003; 12: 1261-1274Abstract Full Text Full Text PDF PubMed Scopus (193) Google Scholar, 5Jia J. Tong C. Jiang J. Genes Dev. 2003; 17: 2709-2720Crossref PubMed Scopus (136) Google Scholar, 6Ogden S.K. Ascano M. Stegman Jr., M.A. Suber L.M. Hooper J.E. Robbins D.J. Curr. Biol. 2003; 13: 1998-2003Abstract Full Text Full Text PDF PubMed Scopus (95) Google Scholar, 7Ruel L. Rodriguez R. Gallet A. Lavenant-Staccini L. Therond P.P. Nat. Cell. Biol. 2003; 5: 907-913Crossref PubMed Scopus (164) Google Scholar). SMO ultimately activates transcription factors of the Gli family. As transcriptional factors, Gli molecules can regulate target gene expression by direct association with a consensus binding site (5′-tgggtggtc-3′) located in the promoter region of the target genes (8Kinzler K.W. Vogelstein B. Mol. Cell. Biol. 1990; 10: 634-642Crossref PubMed Scopus (418) Google Scholar, 9Sasaki H. Hui C. Nakafuku M. Kondoh H. Development (Camb.). 1997; 124: 1313-1322Crossref PubMed Google Scholar). Constitutive activation of Hh signaling is detected in a variety of human cancers. The link between the Hh signaling pathway and human cancer has come from genetic analysis of Gorlin syndrome patients, who are predisposed to early onset of multiple basal cell carcinomas (10Hahn H. Wicking C. Zaphiropoulous P.G. Gailani M.R. Shanley S. Chidambaram A. Vorechovsky I. Holmberg E. Unden A.B. Gillies S. Negus K. Smyth I. Pressman C. Leffell D.J. Gerrard B. Goldstein A.M. Dean M. Toftgard R. Chenevix-Trench G. Wainwright B. Bale A.E. Cell. 1996; 85: 841-851Abstract Full Text Full Text PDF PubMed Scopus (1688) Google Scholar, 11Johnson R.L. Rothman A.L. Xie J. Goodrich L.V. Bare J.W. Bonifas J.M. Quinn A.G. Myers R.M. Cox D.R. Epstein Jr., E.H. Scott M.P. Science. 1996; 272: 1668-1671Crossref PubMed Scopus (1641) Google Scholar). In addition to basal cell carcinomas and medulloblastomas, Hh signaling activation occurs frequently in advanced prostate cancer, small cell lung cancer, and several gastrointestinal cancers, including gastric, esophageal, pancreatic, and liver cancers (12Berman D.M. Karhadkar S.S. Maitra A. Montes De Oca R. Gerstenblith M.R. Briggs K. Parker A.R. Shimada Y. Eshleman J.R. Watkins D.N. Beachy P.A. Nature. 2003; 425: 846-851Crossref PubMed Scopus (1129) Google Scholar, 13Thayer S.P. di Magliano M.P. Heiser P.W. Nielsen C.M. Roberts D.J. Lauwers G.Y. Qi Y.P. Gysin S. Fernandez-del Castillo C. Yajnik V. Antoniu B. McMahon M. Warshaw A.L. Hebrok M. Nature. 2003; 425: 851-856Crossref PubMed Scopus (1317) Google Scholar, 14Ma X. Chen K. Huang S. Zhang X. Adegboyega P.A. Evers B.M. Zhang H. Xie J. Carcinogenesis. 2005; 26: 1698-1705Crossref PubMed Scopus (184) Google Scholar, 15Sicklick J.K. Li Y.X. Jayaraman A. Kannangai R. Qi Y. Vivekanandan P. Ludlow J.W. Owzar K. Chen W. Torbenson M.S. Diehl A.M. Carcinogenesis. 2006; 27: 748-757Crossref PubMed Scopus (239) Google Scholar, 16Huang S. He J. Zhang X. Bian Y. Yang L. Xie G. Zhang K. Tang W. Stelter A.A. Wang Q. Zhang H. Xie J. Carcinogenesis. 2006; 27: 1334-1340Crossref PubMed Scopus (200) Google Scholar, 17Patil M.A. Zhang J. Ho C. Cheung S.T. Fan S.T. Chen X. Cancer Biol. Ther. 2006; 5: 111-117Crossref PubMed Scopus (89) Google Scholar). Wnt signaling is also known to be involved in several types of gastrointestinal cancers (18Moon R.T. Sci. STKE. 2005; 2005: cm1PubMed Google Scholar). Two recent studies indicate that tumors with activated Hh signaling often do not have nuclear accumulation of β-catenin, a major indicator for the canonical Wnt signaling (16Huang S. He J. Zhang X. Bian Y. Yang L. Xie G. Zhang K. Tang W. Stelter A.A. Wang Q. Zhang H. Xie J. Carcinogenesis. 2006; 27: 1334-1340Crossref PubMed Scopus (200) Google Scholar, 19Akiyoshi T. Nakamura M. Koga K. Nakashima H. Yao T. Tsuneyoshi M. Tanaka M. Katano M. Gut. 2006; 55: 991-999Crossref PubMed Scopus (83) Google Scholar). One study indicates that Indian Hh negatively regulates Wnt signaling (20van den Brink G.R. Bleuming S.A. Hardwick J.C. Schepman B.L. Offerhaus G.J. Keller J.J. Nielsen C. Gaffield W. van Deventer S.J. Roberts D.J. Peppelenbosch M.P. Nat. Genet. 2004; 36: 277-282Crossref PubMed Scopus (318) Google Scholar); however, the molecular basis for such an inhibition remains elusive. SFRP-1 is a 30-kDa glycoprotein that was first identified as the antagonist of Wnt signaling and regulator of apoptosis (21Finch P.W. He X. Kelley M.J. Uren A. Schaudies R.P. Popescu N.C. Rudikoff S. Aaronson S.A. Varmus H.E. Rubin J.S. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 6770-6775Crossref PubMed Scopus (364) Google Scholar). The sFRP-1 gene is frequently up-regulated in Hh-activated tumors (22Romer J.T. Kimura H. Magdaleno S. Sasai K. Fuller C. Baines H. Connelly M. Stewart C.F. Gould S. Rubin L.L. Curran T. Cancer Cell. 2004; 6: 229-240Abstract Full Text Full Text PDF PubMed Scopus (452) Google Scholar). We have found that expression of sFRP-1 is dependent on Gli1 and Gli2 in MEF cells and regulated by the Hh pathway in gastric cancer cell lines. Gli1 protein appears to be involved in regulation of the endogenous sFRP-1 promoter. As a consequence of Hh signaling activation and consequent sFRP-1 expression, Wnt-1-mediated β-catenin accumulation in the cytosol and DKK1 expression are inhibited. This inhibition can be reversed when sFRP-1 expression is reduced. These data indicate that sFRP-1 serves as the molecular link for Hh signaling-mediated inhibition of Wnt signaling. Cell Culture and Plasmids—The SIIA cell line was established in the Department of Surgery in our institution (23Barranco S.C. Weintraub B. MacLean K.K. Beasley E.G. Jenkins V.K. Townsend Jr., C.M. Invest. New Drugs. 1991; 9: 29-36Crossref PubMed Scopus (19) Google Scholar). AGS is another cell line from gastric adenocarcinomas used in this study. Both cell lines were kindly provided by Dr. Mark Hellmich and cultured according to the suggested conditions (14Ma X. Chen K. Huang S. Zhang X. Adegboyega P.A. Evers B.M. Zhang H. Xie J. Carcinogenesis. 2005; 26: 1698-1705Crossref PubMed Scopus (184) Google Scholar). 293 cells were purchased from ATCC and cultured in Dulbecco's modified Eagle's medium with 10% fetal bovine serum. Ectopic expression of Gli1 in AGS cells was achieved by the stable retrovirus infection. For retrovirus infection, pLNCX-Gli1 (MYC-tagged) was transfected into Phoenix packaging cells using FuGENE 6 (Roche Applied Science). Supernatant was collected 24–48 h post-transfection (24Sheng T. Chi S. Zhang X. Xie J. J. Biol. Chem. 2006; 281: 9-12Abstract Full Text Full Text PDF PubMed Scopus (102) Google Scholar). Empty pLNCX vector was used to produce the control retrovirus. Infection for AGS cells was carried out overnight in the presence of 8 μg/ml polybrene. A day later, 250 μg/ml G418 was added to the culture medium for selection of a pool of Gli1 stable expressing clones. Western blot was used to detect the Gli1 protein. 293-Wnt1 cells were made from the pLNCX-Wnt1 plasmid (25Chen R.H. Ding W.V. McCormick F. J. Biol. Chem. 2000; 275: 17894-17899Abstract Full Text Full Text PDF PubMed Scopus (205) Google Scholar, 26Papkoff J. Rubinfeld B. Schryver B. Polakis P. Mol. Cell. Biol. 1996; 16: 2128-2134Crossref PubMed Scopus (312) Google Scholar). SIIA Cells were treated with 2.5 μm keto-N-aminoethylaminocaproyldihydrocinnamoylcyclopamine (KAAD-cyclopamine, catalog number K171000 from Toronto Research Chemicals Inc., Toronto, Canada) for 10, 12, and 24 h (14Ma X. Chen K. Huang S. Zhang X. Adegboyega P.A. Evers B.M. Zhang H. Xie J. Carcinogenesis. 2005; 26: 1698-1705Crossref PubMed Scopus (184) Google Scholar). siRNA, RNA Isolation, RT-PCR, and Real-time PCR—Expression of sFRP-1 in 293 cells was knocked down using Ambion's pre-designed siRNA for sFRP-1 (siRNA ID number 121421) following transfection with oligofectAmine according to manufacturer's instruction. Total RNA of cells was extracted using a RNA extraction kit from Promega according to the manufacturer (Promega, Madison, WI), and quantitative PCR analyses were performed according to a previously published procedure using primers and probes from Applied Biosystems (16Huang S. He J. Zhang X. Bian Y. Yang L. Xie G. Zhang K. Tang W. Stelter A.A. Wang Q. Zhang H. Xie J. Carcinogenesis. 2006; 27: 1334-1340Crossref PubMed Scopus (200) Google Scholar). Triplicate CT values were analyzed in Microsoft Excel using the comparative CT(ΔΔCT) method as described by the manufacturer (Applied Biosystems, Foster City, CA). The amount of target (2–ΔΔCT) was obtained by normalization to an endogenous reference (18 S RNA) and relative to a calibrator. We performed RT-PCR of Gli1, sFRP-1, and HIP with 32 cycles of 96 °C for 30 s, 55 °C for 45 s, and 72 °C for 45 s with the following primers: Gli1, 5′-GGAATTCTGTTTCCCCAGGT-3′ (forward primer) and 5′-ACCCCCTGGACTCTCTTGAT-3′ (reverse primer); sFRP-1, 5′-CCCTCGGGGAACTTGTCACA-3′ (forward primer) and 5′-GCTCAACAAGAACTGCCACA-3′ (reverse primer); HIP, 5′-TGCTAAGCCTCGCATTCCA-3′ (forward primer) and 5′-ACAACCCTAAGAATGTGGTCATGA-3′ (reverse primer). Western Blotting—Cells were lysed in protein loading buffer, and proteins were separated by 10% SDS-PAGE. After electrophoresis, protein was electrotransferred on nitrocellulose membrane, blocked with 5% nonfat dry milk in TBST (50 mm Tris-HCl, pH 7.5, 150 mm NaCl, 0.1% Tween 20) for 1 h at room temperature and blotted against appropriate primary antibodies (sFRP-1 antibodies were from R&D System, Inc., catalog number AF 1384; Myc-tag (9B-11) antibodies were from Cell Signaling Inc., catalog number 2276; β-catenin antibodies were from BD Transduction Laboratory, catalog number 610154; β-actin antibodies were purchased from Sigma) overnight at 4 °C, which was followed by incubation with horseradish peroxidase-conjugated secondary antibody (1:5,000) for 1 h at room temperature. Different dilutions were used for different primary antibodies: anti-sFRP-1, 1:1,000; anti-β-catenin, 1:1,000; anti-MYC 9B11, 1:5,000; and anti-β actin, 1:5,000. The protein bands were visualized by enhanced chemiluminescence (24Sheng T. Chi S. Zhang X. Xie J. J. Biol. Chem. 2006; 281: 9-12Abstract Full Text Full Text PDF PubMed Scopus (102) Google Scholar). Chromatin Immunoprecipitation (ChIP) Assay—Log-phase AGS cells and the AGS cells with ectopic Gli1 expression (2 × cells for immunoprecipitation were cultured and by the addition of for After addition of mm cells were with in of mm PIPES, pH mm and for at 4 was by to an of and by at × for at 4 were with the mm mm Tris-HCl, pH mm was used as the and the of was with 1 of the control antibody at 4 °C for were by at in an for 1 and were with 1 of (50 mm pH mm and with 1 of immunoprecipitation mm pH mm for with After of (50 mm pH 1 mm mm NaCl, μg/ml was added to at 55 °C for followed by 6 h at The were extracted with following by with at °C in the presence of of The were in of mm mm pH PCR was performed with cycles of 96 °C for 30 s, 55 °C for 45 s, and 72 °C for 45 s using the following primers the in the human sFRP-1 Cell with KAAD-cyclopamine for 24 h SIIA medium from 293-Wnt1 cells for h 293 the cells were with in 1 of for and collected by at × for The cell were with of Tris-HCl, pH 1 After on the were at × for at 4 The as the and were at × for 30 at 4 The the were collected (25Chen R.H. Ding W.V. McCormick F. J. Biol. Chem. 2000; 275: 17894-17899Abstract Full Text Full Text PDF PubMed Scopus (205) Google Scholar, 26Papkoff J. Rubinfeld B. Schryver B. Polakis P. Mol. Cell. Biol. 1996; 16: 2128-2134Crossref PubMed Scopus (312) Google Scholar). were for protein by protein and separated by 10% for to β-catenin Transduction were used to detect the level of β-catenin in different Hh target we gene expression between and MEF We found that expression of sFRP-1 was dependent on Gli1 and Gli2. As in the sFRP-1 level in was that in MEF This was by the blot analysis not the in cancer cells, we detected the sFRP-1 expression in the human cancer cell lines with activated Hh pathway. As in the Hh activation level was in AGS cells that in SIIA with Hh signaling a level of sFRP-1 expression was detected in SIIA cells analysis of primary human cancers that sFRP-1 was in gastric cancer with activated Hh signaling not ectopic expression of Gli1 gene in AGS cells, we that sFRP-1 expression was of SIIA cells for h with SMO KAAD-cyclopamine, the sFRP-1 level by in SIIA cells the expression of sFRP-1 at the protein we performed Western analysis using medium of SIIA cells sFRP-1 is a secreted As in we found that sFRP-1 protein expression was KAAD-cyclopamine in SIIA These data indicate that expression of sFRP-1 is regulated by Hh signaling. Gli transcription factors are involved in regulation of the sFRP-1 we analyzed the promoter of sFRP-1 and found two We performed to Gli1 is involved in binding the endogenous sFRP-1 promoter. As in we found that immunoprecipitation of Gli1 can down the endogenous sFRP-1 promoter binding that Gli1 is involved in regulation of the sFRP-1 promoter. is known that sFRP-1 is an for Wnt signaling (21Finch P.W. He X. Kelley M.J. Uren A. Schaudies R.P. Popescu N.C. Rudikoff S. Aaronson S.A. Varmus H.E. Rubin J.S. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 6770-6775Crossref PubMed Scopus (364) Google Scholar). sFRP-1 expression Wnt we β-catenin accumulation in the cytosol of 293 cells in different with published (25Chen R.H. Ding W.V. McCormick F. J. Biol. Chem. 2000; 275: 17894-17899Abstract Full Text Full Text PDF PubMed Scopus (205) Google Scholar, 26Papkoff J. Rubinfeld B. Schryver B. Polakis P. Mol. Cell. Biol. 1996; 16: 2128-2134Crossref PubMed Scopus (312) Google medium β-catenin accumulation in the cytosol However, following expression of Gli1, Wnt-1-mediated β-catenin accumulation was In of β-catenin were this we the expression of a Wnt target gene D.R. A. Varmus H.E. J. 2005; PubMed Scopus Google in As in while Wnt-1-conditioned medium expression of DKK1 by ectopic expression of Gli1 Wnt-1-mediated of As Gli1 expression was by level of sFRP-1 in the medium A and This was of protein addition of not the level of sFRP-1 sFRP-1 is for inhibition of we knocked down sFRP-1 in 293 We found that following sFRP-1 Wnt-1-conditioned were to accumulation of β-catenin as as DKK1 expression These that Hh signaling Wnt signaling in 293 cells through regulation of sFRP-1 expression. We that sFRP-1 is in cancer cells with activated Hh cells be to we found that Wnt-1-conditioned medium not β-catenin accumulation in SIIA cells, which have active hedgehog signaling. However, following inhibition of Hh signaling by KAAD-cyclopamine which is by expression of sFRP-1 Wnt-1-conditioned medium was to accumulation of β-catenin These data that Hh signaling Wnt signaling through regulation of sFRP-1. The of our were by the that gastrointestinal cancer with Hh signaling activation not have accumulation of β-catenin (16Huang S. He J. Zhang X. Bian Y. Yang L. Xie G. Zhang K. Tang W. Stelter A.A. Wang Q. Zhang H. Xie J. Carcinogenesis. 2006; 27: 1334-1340Crossref PubMed Scopus (200) Google Scholar, 19Akiyoshi T. Nakamura M. Koga K. Nakashima H. Yao T. Tsuneyoshi M. Tanaka M. Katano M. Gut. 2006; 55: 991-999Crossref PubMed Scopus (83) Google Scholar). As we in liver and gastric cancers, out of tumors with activated Hh signaling nuclear of β-catenin (16Huang S. He J. Zhang X. Bian Y. Yang L. Xie G. Zhang K. Tang W. Stelter A.A. Wang Q. Zhang H. Xie J. Carcinogenesis. 2006; 27: 1334-1340Crossref PubMed Scopus (200) Google Scholar). In our data direct to that sFRP-1 is the target gene of Hh signaling that the on Wnt signaling. data from suggest that the Hh signaling pathway regulates sFRP-1 expression through the promoter. The activation of Hh and Wnt signaling occurs frequently in studies indicate that serves as an for Wnt signaling development and in cancer (20van den Brink G.R. Bleuming S.A. Hardwick J.C. Schepman B.L. Offerhaus G.J. Keller J.J. Nielsen C. Gaffield W. van Deventer S.J. Roberts D.J. Peppelenbosch M.P. Nat. Genet. 2004; 36: 277-282Crossref PubMed Scopus (318) Google Scholar). Furthermore, is that Gli1 an in the development of cancer Wnt signaling T. Nakamura M. Koga K. Nakashima H. Yao T. Tsuneyoshi M. Tanaka M. Katano M. Gut. 2006; 55: 991-999Crossref PubMed Scopus (83) Google Scholar). We have that activation of Hh signaling is associated with nuclear of β-catenin in liver cancers (16Huang S. He J. Zhang X. Bian Y. Yang L. Xie G. Zhang K. Tang W. Stelter A.A. Wang Q. Zhang H. Xie J. Carcinogenesis. 2006; 27: 1334-1340Crossref PubMed Scopus (200) Google Scholar). data indicate that expression of sFRP-1 following activation of the Hh pathway the molecular link for the on Wnt signaling. We have that sFRP-1 expression is dependent on Gli1 and Gli2. with activated Hh signaling have a level of sFRP-1 expression, and inhibition of Hh signaling also reduces the level of sFRP-1. We that Gli1 is involved in binding the endogenous sFRP-1 promoter. SIIA cells in which Hh signaling is activated do not respond to Wnt-1-mediated β-catenin accumulation in the whereas inhibition of Hh signaling We that this signaling is in cancers in which sFRP-1 is through promoter H. Watkins D.N. K.W. Chen Yang B. Y. M. M. T. Y. K. Nat. Genet. 2004; 36: PubMed Scopus Google which can be in of sFRP-1 not Wnt signaling to control also the from the Hh pathway. In addition to expression of sFRP-1, be also for this such as β-catenin accumulation T. Nakamura M. Koga K. Nakashima H. Yao T. Tsuneyoshi M. Tanaka M. Katano M. Gut. 2006; 55: 991-999Crossref PubMed Scopus (83) Google Scholar). the of has been to their inhibition of Wnt of was to cell and inhibit apoptosis through an association with a complex and of signaling L.L. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). A of using sFRP-1 identified several molecules involved in such as and a human homologue of receptor Y. Uren A. J. C. V. Rubin J.S. 2004; PubMed Scopus Google Scholar). be to are involved in signaling in human We and for and Dr. A. Aaronson for the sFRP-1 with

Fetched live from OpenAlex and de-inverted. Abstracts are not stored in this database: the inverted indexes are 8.6 GB of the frame’s 9.3 GB of text, and the host has 13 GB free.