Key elements of the BMP/SMAD pathway co-localize with CDX2 in intestinal metaplasia and regulate CDX2 expression in human gastric cell lines.
Helicobacter pylori infection induces intestinal metaplasia of the stomach, a preneoplastic lesion associated with an increased risk for gastric cancer development. Intestinal metaplasia is induced by the intestine-specific transcription factor CDX2 but the mechanisms responsible for this ectopic expression have never been described. We hypothesized that the BMP/SMAD pathway has a role in CDX2 regulation, in this context, for the following reasons: (1) the BMP pathway is crucial for normal intestinal differentiation and (2) there is an influx of BMP2 and BMP4-producing cells to the stomach upon Helicobacter pylori infection. We evaluated the expression of key elements of the BMP pathway in human stomach specimens with IM. Growth factor treatments, with BMP2 and BMP4, were performed in cultured cells and a knock-down experiment of SMAD4 was done using RNAi. We showed overexpression in IM of BMP2/4, BMPR1A, and SMAD4 in 56% of IM foci, and pSMAD1/5/8 in 100% of IM foci as compared to adjacent mucosa. In vitro, treatment of AGS cells with BMP2 and BMP4 increased endogenous CDX2 expression as well as the intestinal differentiation markers MUC2 and LI-cadherin. On the other hand, SMAD4 knock-down led to decreased endogenous CDX2, MUC2, and LI-cadherin in AGS. Treatment of the SMAD4 knock-down cells had no influence on CDX2 expression as opposed to wild-type cells. A 9.3 kb CDX2 promoter could be transactivated by SMAD4 and SMAD1 in a cell-dependent manner. In conclusion, we identified for the first time that the BMP pathway is active in intestinal metaplasia and that BMP2 and BMP4 regulate CDX2 expression and promote intestinal differentiation through the canonical signal transducers.
Barros
R
R
Institute of Molecular Pathology and Immunology of the University of Porto, Porto, Portugal.
Pereira
B
B
Duluc
I
I
Azevedo
M
M
Mendes
N
N
Camilo
V
V
Jacobs
R J
RJ
Paulo
P
P
Santos-Silva
F
F
van Seuningen
I
I
van den Brink
G R
GR
David
L
L
Freund
J-N
JN
Almeida
R
R
D015153:Blotting, Western / Q000379:methods; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D000071616:CDX2 Transcription Factor; D002277:Carcinoma / Q000235:genetics* / Q000378:metabolism; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D015972:Gene Expression Regulation, Neoplastic; D018398:Homeodomain Proteins / Q000032:analysis / Q000235:genetics*; D006801:Humans; D007150:Immunohistochemistry; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D051898:Smad1 Protein / Q000378:metabolism; D051901:Smad4 Protein / Q000235:genetics / Q000378:metabolism; D013274:Stomach Neoplasms / Q000235:genetics* / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology
C525718:BMP2 protein, human; C525809:BMP4 protein, human; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D000071616:CDX2 Transcription Factor; C491270:CDX2 protein, human; D018398:Homeodomain Proteins; D034741:RNA, Small Interfering; D051898:Smad1 Protein; D051901:Smad4 Protein; D016212:Transforming Growth Factor beta