Tài liệu Báo cáo khoa học: Cell surface heparan sulfate proteoglycans Target and partners of the

Cell surface heparan sulfate proteoglycans

Target and partners of the basic ®broblast growth factor in rat Sertoli cells

Sylvie Brucato, Jean Bocquet and Corinne Villers

Laboratoire de Biochimie, IRBA, Universite de Caen, France

Basic ®broblast growth factor (bFGF) regulates diversi®ed

biological functions in rat Sertoli cells. This report demon￾strates that bFGF inhibits steroidogenesis in developing rat

Sertoli cells. Follicle stimulating hormone (FSH)-stimulated

estradiol production was reduced by bFGF. Moreover, the

amount of cytochrome P450 aromatase, responsible for the

irreversible transformation of androgens into estrogens, is

decreased by bFGF at the transcriptional level. The bFGF

inhibitory e€ect was also observed in the presence of dibu￾tyryl-cAMP, cholera toxin or RO-20-1724, all inducing high

levels of cAMP, the second messenger of FSH.

Heparan sulfate proteoglycans (HSPGs) were shown to be

required as cofactors for bFGF signaling. Indeed, sodium

chlorate, described to drastically decrease proteoglycan sul￾fation, abolishes the bFGF downregulation of FSH-stimu￾lated estradiol synthesis previously observed. Glypican-1,

syndecan-1 and -4, potential bFGF coreceptors, are mainly

regulated at the transcriptional level. This report shows that

the bFGF regulation of their expression speci®cally depends

on the nature of HSPG and of the Sertoli cell developmental

stage.

In conclusion, HSPG are partners and the target of bFGF

in rat Sertoli cells.

Keywords: bFGF; aromatase;heparansulfateproteoglycans;

RT-PCR; Sertoli cells.

The basic ®broblast growth factor (bFGF or FGF-2)

belongs to a large FGF family of 21 structurally related

members [1]. This growth factor is produced by many cell

types and tissues, including testis [2]. Its biological activity is

pleiotropic [3] as it in¯uences aspects of both cellular

growth, differentiation but also angiogenesis, tissue repair

and cell migration. In rat testis, bFGF affects, for instance,

Leydig and Sertoli cell steroidogenesis [4,5], Sertoli cell

transferrin production [6] and plasminogen activator activity

[7] but also c-fos [8] and FGFR-1 [9] mRNA expression.

The biological activity of bFGF is mediated by interac￾tion with high af®nity cell surface bFGF receptors (FGFR-1

to FGFR-4) [10]. In addition, bFGF binds to heparan

sulfate proteoglycans (HSPG) on the cell surface [11].

Oligosaccharidic sequences of HS chains are de®ned for the

bFGF binding and for the recognition of the speci®c bFGF

receptor, leading to the formation of a ternary complex

comprising HSPG±bFGF±FGFR. These oligosaccharidic

motifs are differently sulfated related to the synthesis

pathway itself and depending on the cell type. The resulting

structural microheterogeneity modulates bFGF af®nity for

its coreceptor and, as a consequence, the growth factor

activity. Studies indicated that bFGF binding to HSPG

facilitates bFGF receptor binding and activation. bFGF

receptor binding to cells that do not express HSPG is

signi®cantly reduced when compared to cells expressing

HSPG [4,11±15].

Sertoli cells are the principal source of estradiol produc￾tion in the immature testis [16,17]. Signi®cant estrogen

synthesis is present in Sertoli cells of early postnatal rats,

with a sharp reduction during subsequent maturation

[18,19].

The present work ®rstly aims to evaluate the effect of

bFGF on follicle stimuling hormone (FSH)-estradiol syn￾thesis and cytochrome P450 aromatase mRNA expression

in 20 days old-rat Sertoli cells. The involvement of the

cAMP pathway was evaluated using three approaches, all

inducing differently high levels of cAMP: (a) dibutyryl cyclic

AMP (dbcAMP), a structural analogue of cAMP; (b)

cholera toxin, a protein Gs activator; and (c) RO-20±1724, a

speci®c phosphodiesterase inhibitor.

Then, we investigated bFGF effect on FSH-estradiol

synthesis in the absence of HSPG in 20-day-old-rat Sertoli

cells. These cells were treated with sodium chlorate to

completely inhibit sulfatation of proteoglycans and, in

consequence, abolish bFGF binding to HSPG.

Our previous studies indicated that in immature rat Sertoli

cells, cell surface proteoglycans are mainly represented by

HSPG [20,21] and among these, at least glypican-1, syn￾decan-1 and syndecan-4 mRNAs are expressed [22]. More￾over, syndecan-1 [23], syndecan-4 [24] and glypican-1 [25] are

potential coreceptors of bFGF, and are essentially regulated

at the transcriptional level [26]. Thus, using a semi-quanti￾tative RT-PCR, we had demonstrated in immature Sertoli

cells that glypican-1 and syndecan-1 mRNA expression was

Correspondence to S. Brucato, Laboratoire de Biochimie, IRBA,

Universite de Caen, Esplanade de la Paix, 14032 Caen cedex, France.

Fax: + 33 2 31 95 49 40, Tel.: + 33 2 31 56 65 76,

E-mail: [email protected]

Abbreviations: bFGF, basic ®broblast growth factor; FSH, follicle

stimulating hormone; HSPG, heparan sulfate proteoglycan; PAPS,

phosphoadenosine phosphosulfate; FiRE, FGF-inducible response

element; FIN-1, FGF-inducible nuclear protein-1; DMEM, Dul￾becco's modi®ed Eagle's medium; RhFGF, recombinant human basic

FGF; PAPS, phosphoadenosine phosphosulfate; AMV, avian myelo￾blastosis virus.

(Received 30 May 2001, revised 1 October 2001, accepted 14

November 2001)

Eur. J. Biochem. 269, 502±511 (2002) Ó FEBS 2002