E. Nagyova et al., Secretion of paracrine factors enabling expansion of cumulus cells is developmentally regulated in pig oocytes, BIOL REPROD, 63(4), 2000, pp. 1149-1156
To demonstrate secretion of cumulus expansion-enabling factor (CEEF) by por
cine oocytes, we used an interspecies testing system. Porcine oocytes were
used to condition culture medium, and the presence of CEEF was tested using
mouse oocytectomized complexes (OOX), which require CEEF for expansion. Fo
llicle-stimulating hormone-stimulated expansion and synthesis of hyaluronic
acid (HA) by mouse OOX were assessed after 18 h of culture in media condit
ioned by porcine oocytes: 1) at different stages of maturation and 2) in wh
ich maturation was inhibited with a specific inhibitor of cdk-kinases, buty
rolactone I. Fully grown (GV-germinal vesicle), late-diakinesis (LD), metap
hase 1 (MI), and metaphase II (MII) oocytes were prepared by culture of ooc
yte-cumulus complexes (OCC) for 0, 22, 27, and 42 h, respectively. To block
CV breakdown, porcine oocytes were cultured for 27 h in medium supplemente
d with butyrolactone 1 (50 mu M). Medium conditioned by oocytes in GV, LD,
and after butyrolactone I block allowed full expansion of >90% of mouse OOX
, whereas oocytes in MI and MII caused disintegration of mouse OOX without
cumulus mucification. To measure synthesis of HA by cumulus cells, 25 mouse
OOX were cultured in the conditioned media in the presence of 2.5 mu Ci of
D-[6-H-3]glucosamine hydrochloride. After 18 h, incorporation of the [H-3]
glucosamine into HA was determined either in complexes (retained HA) or in
medium plus complexes (total HA). Total HA accumulation by mouse OOX was no
t different from that of intact OCC. However, oocytes in CV, LD, and after
butyrolactone I treatment enabled mouse OOX to retain significantly more HA
within the complex than oocytes in MI and MII. The results indicate that s
ecretion of factors that promote the retention of HA within the complex is
developmentally regulated during oocyte maturation.