Transforming growth factor beta (TGF-beta) expression in isolated and cultured rat hepatocytes.
It is still a subject of debate whether hepatocytes have the ability to express TGF-beta. Therefore, we investigated in freshly isolated and in monolayer cultures of rat hepatocytes the expression of TGF-beta isoform s at the RNA and protein level applying RT-PCR, immunocytochemistry, immunoblotting, and functional assays of TGF-beta. TFG-beta 1, -beta 2, and -beta 3 transcripts were detected in cultured cells, and the level of m RNA increased up to 48/72 h, but TGF-beta 1 transcripts were absent in freshly isolated cells. Using APAAP stainings the proteins of all three TGF-beta isoforms were observed in hepatocyte cultures from 5-96 h, but in hepatocytes in the liver in situ and in freshly isolated cell suspensions TGF-beta staining was negative. SDS-PAGE under reducing conditions followed by Western blotting detected in cell lysates the subunit of mature TGF-beta at about 13 kd. Analysis of TGF-beta bioactivity with the mink cell (Mv1Lu) proliferation inhibition assay and competitive radioligand assay confirmed in activated (i.e., acidified and subsequently neutralized) hepatocyte-conditioned media the presence of TGF-beta, which, however, is almost entirely in the latent form. It is concluded that TGF-beta can be expressed in cultured hepatocytes and that the level of expression is quickly upregulated under abnormal, not yet known, microenvironmental conditions.
Gao
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C
Department of Clinical Chemistry, Philipps University, Marburg, Germany.
Gressner
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G
Zoremba
M
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Gressner
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AM
D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002478:Cells, Cultured; D003470:Culture Media; D004587:Electrophoresis, Agar Gel; D005786:Gene Expression Regulation / Q000235:genetics*; D015151:Immunoblotting; D007150:Immunohistochemistry; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008297:Male; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011863:Radioimmunoassay; D051381:Rats; D017207:Rats, Sprague-Dawley; D016212:Transforming Growth Factor beta / Q000096:biosynthesis / Q000235:genetics*
D003470:Culture Media; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta