Differential effects of the charge variants of human follicle-stimulating hormone.
FSH is synthesized and secreted by the anterior pituitary gland in multiple molecular forms; the release of these isoforms depends on the endocrine status of the donor at the time of sample collection. In the present study, we analysed the possibility that the FSH charge isoforms may exert differential effects at the target cell. Seven FSH isoform mixes were isolated from pooled anterior pituitary glycoprotein extracts by high resolution chromatofocusing, followed by affinity chromatography, which removed nearly 90% of the LH that co-eluted with the FSH isoforms during chromatofocusing. The isoforms (isoform I, pH >7.10; II, pH range 6.60-6.20; III, pH 5. 47-5.10; IV, pH 5.03-4.60; V, pH 4.76-4.12; VI, pH 4.05-3.82 and VII, pH <3.80) were then tested for their capacity to stimulate cAMP release, androgen aromatization and tissue-type plasminogen activator (tPA) enzyme activity and cytochrome P450 aromatase, tPA and inhibin alpha-subunit mRNA production by rat granulosa cells in culture. cAMP and oestradiol production were determined by RIA, tPA enzyme activity by SDS-PAGE and zymography and all mRNAs by northern blot hybridization analysis and semiquantitative RT-PCR. All isoforms, with the exception of isoform I, stimulated synthesis and release of cAMP, oestrogen and tPA enzyme activity in a dose-dependent manner; the potency of the less acidic isoforms (pH 6. 60-4.60) was greater than that exhibited by the more acidic/sialylated analogs (pH 4.76 to <3.80; potencies II>III>IV>V>VII>VI). A similar trend was observed in terms of cytochrome P450 aromatase and tPA mRNA production. In contrast, when FSH-stimulated production of alpha-inhibin mRNA was analysed, isoforms V-VII were significantly more potent (two- to threefold) than the less acidic/sialylated counterparts (II-IV). In contrast to isoforms II-VII (which behaved as FSH agonists), isoform I (elution pH >7.10) completely blocked P450 aromatase and tPA mRNA expression, without altering that of a constitutively expressed gene (glyceraldehyde-3-phosphate dehydrogenase). These results show for the first time that the naturally occurring human FSH isoforms may exhibit differential or even unique effects at the target cell level.
Timossi
C M
CM
Department of Pharmacology, Faculty of Medicine, Universidad Nacional Autónoma de México.
Barrios-de-Tomasi
J
J
González-Suárez
R
R
Arranz
M C
MC
Padmanabhan
V
V
Conn
P M
PM
Ulloa-Aguirre
A
A
D000704:Analysis of Variance; D000818:Animals; D001141:Aromatase / Q000032:analysis; D015152:Blotting, Northern / Q000379:methods; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000032:analysis; D004958:Estradiol / Q000032:analysis / Q000096:biosynthesis*; D005260:Female; D005640:Follicle Stimulating Hormone / Q000302:isolation & purification / Q000494:pharmacology*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000032:analysis; D006107:Granulosa Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D007265:Inhibins; D007986:Luteinizing Hormone / Q000032:analysis; D010455:Peptides / Q000032:analysis; D010903:Pituitary Gland, Anterior / Q000737:chemistry; D020033:Protein Isoforms / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D010959:Tissue Plasminogen Activator / Q000032:analysis / Q000096:biosynthesis*
D010455:Peptides; D020033:Protein Isoforms; D012333:RNA, Messenger; C094377:inhibin-alpha subunit; D004958:Estradiol; D007265:Inhibins; D007986:Luteinizing Hormone; D005640:Follicle Stimulating Hormone; D000242:Cyclic AMP; D001141:Aromatase; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010959:Tissue Plasminogen Activator