Comparative Study In Vivo and In Vitro of Uniformly ¹⁴C‐Labelled and ¹²⁵I‐Labelled Recombinant Fibroblast Growth Factor 2

Abstract

Recombinant bovine fibroblast growth factor (FGF2), uniformly labelled with 14C ([14C]FGF2), was purified and showed to be highly stable and to retain full biological activity. Organ distribution of [14C]FGF2 after intravenous injection of young rats was assessed by autoradiography of whole body sections and compared with those obtained with [125I]iodinated FGF2 (125I‐FGF2). Thyroid, stomach, intestine, bladder and skin were radioactively labelled only in the case of 125I‐FGF2. This tissue‐labelling is artefactual, probably due to free iodide binding not observed when using [14C]FGF2. High‐resolution autoradiography showed a complex tissue distribution of [14C]FGF2 in kidney and adrenal organs. Incubation of frozen eye sections with [14C]FGF2 showed a specific and high‐resolution labelling pattern of ocular tissues. After cellular internalization, [14C]FGF2 was processed into five distinct polypeptides of 16, 14, 8, 7, and 5.5 kDa. The 14‐kDa and 7‐kDa polypeptides are novel catabolic fragments not detected with radioiodinated FGF2. In terms of stability, tissue distribution specificity, and autoradiographic resolution, [14C]FGF2 proved to have more advantages than 125I‐FGF2 for pharmacokinetic and catabolism studies.

In Vivo Involvement of Heparan Sulfate Proteoglycan in the Bioavailability, Internalization, and Catabolism of Exogenous Basic Fibroblast Growth Factor

“…In a recent study, we used the uniformly 14 C-labeled FGF2 and 125 I-FGF2 to study the in vivo distribution of FGF2 (Colin et al, 1997). Our results have shown that the use of 125 I-FGF2 leads to artifactual labeling of many tissues and organs, probably due to the nature of the labeling agent ( 125 I atoms).…”

“…The 14 C-PPIase was then purified as described by Liu and Walsh (1990 C-FGF2 as a bolus through the saphenous vein. After injection, animals were anesthetized at different times with the same mixture and sacrificed, and 20-m-thick sagittal sections were performed as described previously (Colin et al, 1997). The FGF2 distribution was observed by autoradiography of tissue sections using Hyperfilm-␤ Max and quantified using a PhosphorImager (Molecular Dynamics, Sunnyvale, CA).…”

“…The E. coli strain DH5a was transformed with the plasmid pJLEC-2B encoding E. coli periplasmic PPIase (Liu and Walsh, 1990). The uniform 14 C labeling of PPIase was obtained by the method described by Colin et al (1997). Briefly, E. coli strain overproducing PPIase was fed with 14 C-glucose as the sole source of carbon.…”

“…The uniform labeling of the growth factor leads to a specific activity of 2.4 Ci/g. The growth-promoting activity was controlled in adult bovine aortic endothelial (ABAE) and retinal pigmented epithelial (RPE) cells (Colin et al, 1997). …”

“…14 C-Uniform Labeling of Recombinant FGF2. The recombinant FGF2 was produced, biosynthetically labeled with 14 C, and purified as described previously (Colin et al, 1997). The uniform labeling of the growth factor leads to a specific activity of 2.4 Ci/g.…”