Effect of TGF-beta on differentiated organoids of the colon carcinoma cell line LIM 1863.
The LIM 1863 colon carcinoma cell line grows in suspension as morphologically and functionally organized organoids in serum-containing medium. Addition of TGF-beta caused the organoids to adhere and inhibited DNA synthesis. A 20 min incubation with TGF-beta was sufficient to induce adherence and this could be inhibited by cycloheximide. The adhesion and DNA synthesis inhibition were demonstrated to be separate events. We were not able to detect any changes in matrix or cell membrane antigens. Similarly there were no changes in synthesized proteins (by two-dimensional gel electrophoresis), and no upregulation of proteoglycan. When adhered organoids were lysed from the tissue culture plastic surface, untreated organoids adhered to this surface. This 'conditioned' surface was destroyed by trypsin but not collagenase or medium from normal LIM 1863 cultures. However, the adherent phenotype was prevented when organoids were treated with transforming growth factor-beta (TGF-beta) in the presence of medium conditioned by normal LIM 1863 cultures rather than in fresh medium. The adhesion process was inhibited by an antibody (QE2E5) against beta 1 integrin although no quantitative changes in integrins were observed (by immunoprecipitation or RNA analysis). A second anti-beta 1 integrin antibody (61.2C4) inhibited LIM 1863 adhesion to collagen but not TGF-beta induced adhesion, implying that TGF-beta induced a specific conformational change or interaction of a beta 1 integrin. In this morphologically structured system TGF-beta induced a number of subtle effects including formation of new extracellular matrix and conformational change of a beta 1 integrin, rather than the major quantitative changes in cell/matrix molecules reported previously.
Hayward
I P
IP
Melbourne Tumour Biology Branch, Ludwig Institute for Cancer Research, Parkville, Australia.
Whitehead
R H
RH
Ward
L
L
Gianello
R
R
Dempsey
P
P
Bates
R
R
Burns
G F
GF
D002277:Carcinoma / Q000188:drug therapy / Q000473:pathology*; D002448:Cell Adhesion / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D003110:Colonic Neoplasms / Q000188:drug therapy / Q000473:pathology*; D004247:DNA / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D006801:Humans; D016023:Integrins / Q000032:analysis; D009924:Organ Culture Techniques; D010641:Phenotype; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D016023:Integrins; D016212:Transforming Growth Factor beta; D004247:DNA