Colocalisation of GH and FSH in the human fetal pituitary by confocal immunofluorescence microscopy
ARM Chirculescu1, C Badiu1, JF Morris2 & M Coculescu1
Colocalisation of hormones has been demonstrated for growth hormone (GH) and prolactin in the normal adult human pituitary, and for many other hormones in pituitary adenomas. We have investigated whether colocalisation of GH and other hormones occurs earlier, during onto-genesis. Pituitaries from therapeutically aborted male human fetuses of 1520 weeks gestational age (with ethical approval) were fixed by immersion in 4% buffered formaldehyde and divided in halves, one of which was embedded in LRGold, the other cryoprotected and embedded in Tissue Tec. One-μm-thick sections of the LRGold-embedded tissue and 12-μm-thick cryostat sections were incubated for immunocytochem-ical double labeling with anti-rat GH and anti-human FSH primary anti-bodies. Rabbit polyclonal or mouse monoclonal antibodies to FSH were used. Primary antibodies were located with either fluorescein- or Texas red-linked secondary antibodies. After TPro nuclear staining the sections were examined by confocal microscopy. The combination of monoclonal anti-FSH and polyclonal anti-GH was best for double labelling.
In all fetuses, there were more GH-positive than FSH-positive cells. In the 15-weeks-old fetus, green-red superposition showed colocalisation of GH / FSH in cells in small clusters of FSH-positive cells. This super-position was not seen in microscopic fields where GH-positive cells pre-dominated. In cryostat sections, colocalisation decreased from 15 to 20 weeks in both intensity and the number of cells showing colocalisation: from 2.87±1.96 (SEM) cells/field in the 15-weeks-old fetus to 1.23±0.95 at 16 weeks and 0.14±0.55 at 20 weeks, where cells colocalizing GH and FSH were only occasionally seen. High magnification of colocalizing cells revealed distinct yellow and red spots suggesting that subcellular colocalization was rarely complete.
These data show that colocalisation/coexistence of GH and FSH occurs in normal pituitary cells early during human ontogenesis but decreases markedly from 15 to 20 weeks. We suggest, however, that the potential for colocalisation is preserved and can be reactivated in adulthood during tumoral transformation.