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DIFFERENTIATION POTENTIAL OF INTESTINAL MESENCHYME AND ITS INTERACTION WITH EPITHELIAL CELLS: A STUDY USING β‐GALACTOSIDASE‐EXPRESSING FIBROBLAST LINES
Author(s) -
Subramanian V.,
Sneddon S. F.,
Martin L.,
Evans G. S.
Publication year - 2001
Publication title -
cell biology international
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.932
H-Index - 77
eISSN - 1095-8355
pISSN - 1065-6995
DOI - 10.1006/cbir.2000.0705
Subject(s) - transfection , electroporation , green fluorescent protein , mesenchyme , cell culture , biology , microbiology and biotechnology , plasmid , fibroblast , reporter gene , beta galactosidase , gene expression , gene , embryo , genetics
Rat intestinal fibroblast lines (F1:G9 and A1:F1) differing in their potential to support intestinal mucosal development were marked with reporter genes to investigate their differentiation potential. The fibroblasts were transfected with plasmids expressing either β‐galactosidase (with or without a nuclear localisation signal) or green fluorescent protein (GFP). Transfection using Tfx50 or Fugene™ was more efficient than electroporation. The expression of β‐galactosidase was more stable and stronger than GFP. Cells were optimally labelled using the plasmid pL27B‐GAL, and sub‐clones with a strong and uniform nuclear expression of β‐galactosidase were isolated. These clones expressed β‐galactosidase even after prolonged passage in the absence of selection. The β‐galactosidase tagged lines (F1:G9 gal and A1:F1 gal ) retained the morphological characteristics, viability and differentiation properties of the parental non‐transfected lines. In co‐culture with a colorectal tumour cell line Caco‐2, the F1:G9 gal and A1:F1 gal cells differed in their morphological organisation but this did not change their expression of smooth muscle α‐actin.