Overexpression of sense or antisense human gastrin mRNA does not affect proliferation of normal rat kidney fibroblasts.
Progastrin-derived peptides have been reported to stimulate mitogenesis in Swiss 3T3 fibroblasts [P. Singh, A. Owlia, R. Espeijo, B. Dai, Novel gastrin receptors mediate mitogenic effects of gastrin and processing intermediates of gastrin on Swiss 3T3 fibroblasts: Absence of detectable cholecystokinin (CCK)-A and CCK-B receptors. J. Biol. Chem. 270 (1995) 8429-8438]. The aim of the present study was to determine the generality of these findings, by investigating the effect of endogenous and exogenous progastrin-derived peptides on the proliferation of the normal rat kidney fibroblast cell line NRK. Levels of endogenous progastrin-derived peptides were modified by stable transfection of NRK cells with tetracycline-repressible plasmids containing sequences encoding human gastrin in either the sense or antisense orientation. Expression of sense and antisense gastrin mRNA was demonstrated by reverse transcriptase PCR and by radioimmunoassay, and cell proliferation rates were determined by the colorimetric MTT assay. Sense clones produced full length human progastrin, but significant quantities of glycine-extended or amidated gastrin17 were not detected. Concentrations of endogenous rat progastrin in antisense clones were significantly lower than concentrations in clones transfected with vector only. However no difference in proliferation rate was observed between sense, antisense and vector-transfected clones. No stimulation of proliferation was observed in synchronised untransfected NRK cells after supplementation of media with gastrin17 or gastrin17gly in the concentration range 0.3 to 100 nM. Our results do not provide evidence in support of the hypothesis that endogenous or exogenous progastrin-derived peptides act as growth factors in NRK fibroblasts.
Yang
Z
Z
University Department of Surgery, Austin & Repatriation Medical Centre, Melbourne, Vic., Australia.
Paterson
A
A
Karelina
Y
Y
Zhang
Q X
QX
Shulkes
A
A
Baldwin
G S
GS
D000818:Animals; D002455:Cell Division; D002845:Chromatography; D016373:DNA, Antisense / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism*; D005755:Gastrins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000378:metabolism*; D010957:Plasmids; D011498:Protein Precursors / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism*; D051381:Rats; D013997:Time Factors; D014162:Transfection
D016373:DNA, Antisense; D005755:Gastrins; D011498:Protein Precursors; D012333:RNA, Messenger; C013706:big gastrin