Sexual dimorphism in immunoneutralization of bioactivity of rat and ovine inhibin

In late-prepubertal female rats passive immunoneutralization of endogenous inhibin was achieved by injection of inhibin antiserum. Effects on follicle population, timing of sexual maturation, ovulation rate at first and second oestrus and serum FSH levels were studied.Rats were injected with antiserum, (non-immune) control serum from castrated sheep (castrated serum) or their IgG fractions, or with saline on day 33 or 3 or 2 days (days \ m=-\ 3/ \ m=-\2) before the expected day of first ovulation, day 38\m=.\5\ m=+-\ 0\m=.\2 (n = 70). Blood was collected from different subgroups at 8, 24 and 48 h, and at first and second oestrus after injection. At necropsy, ovaries were histologically prepared for differential counting of follicles (48 h and first oestrus) and counting of corpora lutea (CL; first and second oestrus) as an index of ovulation rate.Results from rats injected with either serum or its IgG fraction were not different, as was the case when rats were injected with either castrated serum or saline.Thus, results from groups treated with antiserum and antiserum IgG were combined and labelled 'antiserum', and the castrated serum, castrated serum IgG and saline-treated groups were combined and labelled 'control'. The activity of inhibin-neutralizing antibodies in the circulation ofantiserum-treated rats was reduced by 43% between 8 h and second oestrus after injection, as determined by the binding of purified bioactive radioiodinated 31 kDa bovine inhibin.After antiserum injection on day 33, more healthy antral follicles (vol. > 100 \ m=x\1 0 5 \ g=m\ m3, diameter > 260 \g=m\m) were present in the ovaries at 48 h (70\m=.\6 vs 54\m=.\4;P < 0\m=.\05) and at first oestrus (73\m=.\1 vs 50\m=.\8; P < 0\m=.\05) if first oestrus was reached within 5 days, but numbers were not different iffirst oestrus was more than 5 days after injection (52\m=.\6vs 50\m=.\8).The number of CL after injection of antiserum on day 33 was increased at first oestrus compared with control (13\m=.\4\ m=+-\ 0\m=.\5, n = 30, vs 10\m=.\0\ m=+-\ 0\m=.\2, n = 40; P<0\m=.\001), an effect that was even more clearly present in antiserum-injected rats ovulating within 5 days (14\m=.\4 \ m=+-\ 0\m=.\7, n = 20; P < 0\m=.\001).Rats injected with antiserum at days \m=-\ 3/ \m=-\2 showed a doubling of ovulation rate at first oestrus when compared with control animals (21\m=.\5 \ m=+-\ 0\m=.\8, n=12, vs 10\m=.\5\ m=+-\ 0\m=.\2, n = 15; P < 0\ m=. \ 001). No differences in the number of CL was seen at second oestrus. Age and body weight on the day of first ovulation were not influenced by antiserum treatment.Serum FSH was significantly (P < 0\m=.\01) increased at 8 h after antiserum injection on either day 33 or on days \m=-\3/\m=-\2 to a level of 250 and 800% of control levels respectively.Thus, injection with inhibin \p=n-\neutralizing antiserum into prepubertal female rats resulted, through an increase in serum FSH concentration 8 h after injection, in the growth of additional numbers of healthy antral follicles. Supranormal ovulation rate occurred if...

Section: Antiserummentioning

confidence: 73%

Section: Discussionmentioning

confidence: 99%

Section: Antiserummentioning

confidence: 99%

See 1 more Smart Citation

Initial ovulation rate and follicle population after injection of inhibin-neutralizing antiserum in the late-prepubertal rat

Sander

Meijs-Roelofs²,

Leeuwen³

et al. 1991

“…Antisera raised to inhibin partially purified from bovine follicular fluid have been shown to neutralize the FSH-suppressive activity of inhibin in follicular fluid in vitro (van Dijk, Steenbergen, Gielen & de Jong, 1986), but while active immunization of ewes against inhibin from various sources always results in an increase in ovulation rate, such immunizations have not produced consistent increases in the concen¬ tration of FSH (Henderson, Franchimont, LecomteYerna et Findlay, Doughton, Robertson & Forage, 1989;Wrathall, McLeod, Glenross et al 1989). Active immunization against oestradiol-17ß has been shown to produce more consistent rises in FSH concentration, as well as increases in the concentration of luteinizing hor¬ mone (LH) (Plant, Dobson & Ward, 1978;Rawlings, Kennedy & Henricks, 1978;Martensz, Scaramuzzi & Van Look, 1979;Scaramuzzi, Martensz & Van Look, 1980), but such immunizations prevent ovulation by blocking the oestradiol-induced LH surge.…”

Section: Introductionmentioning

confidence: 99%

Effects of passively immunizing ewes against inhibin and oestradiol during the follicular phase of the oestrous cycle

Mann¹,

Campbell²,

McNeilly³

et al. 1990

Passive immunization was used to investigate the importance of inhibin and oestradiol in the control of FSH production during the follicular phase of the oestrous cycle in the sheep. Four groups of five mature Scottish Blackface ewes were injected with normal sheep plasma (control), antiserum to the 1-26 alpha peptide fragment of porcine inhibin, antiserum to oestradiol-17 beta, or a combination of the two antisera, 24 h following cloprostenol-induced luteal regression. There was no difference in the concentration of LH in jugular venous plasma between the control and inhibin-immunized groups following the injection of normal sheep plasma or inhibin antiserum, with both groups exhibiting normal LH surges. In both the groups immunized against oestradiol, the basal concentration of LH rose by 25-30% (P less than 0.05) during the 96-h period following injection, while the LH surge and consequent formation of a corpus luteum was inhibited. In all three immunized groups there was a significant (P less than 0.001) rise in the concentration of FSH starting 3.8-4.8 h after the injection of antiserum. The duration of the rise was similar in the groups injected with oestradiol antiserum alone (43.6 +/- 12.8 h) or in combination with inhibin antiserum (40.6 +/- 11.7 h), but was significantly (P less than 0.05) shorter in the group immunized against inhibin alone (17.0 +/- 0.5 h).(ABSTRACT TRUNCATED AT 250 WORDS)

“…Furthermore, Robertson et al (1988) reported different bioactivity/ immunoactivity ratios for inhibin in sera from men and from women, while van Dijk, Steenbergen, Gielen & de Jong (1986) found that inhibin from ovarian fol¬ licular fluid was immunoneutralized more effectively than testicular inhibin.…”

Section: Introductionmentioning

confidence: 99%

Inhibin and activin-like activity in fluids from male and female gonads: different molecular weight forms and bioactivity/immunoactivity ratios

Grootenhuis

Steenbergen²,

Timmerman³

et al. 1989

Self Cite

The existence of various molecular weight forms of inhibin in ovarian follicular fluid has been reported earlier, while there is no information on the form of inhibin in testicular tissue. Inhibin bioactivity was therefore estimated in eluates of slices, obtained after SDS-PAGE of rat testicular and ovarian homogenates, rat Sertoli cell-conditioned medium (rSCCM) and bovine ovarian follicular fluid (bFF). The only form of inhibin detected in testes from 22-day-old rats and in rSCCM was a 30 kDa protein. In rat ovarian extracts, larger forms of inhibin were also found as well as the predominant 30 kDa form. An activin-like activity was found in the 25 kDa SDS-PAGE eluates of both rSCCM and ovarian homogenates, which caused a dose-dependent increase of FSH release from cultured pituitary cells. Activin-like activity and several forms of inhibin were found in bFF after SDS-PAGE. After purification of inhibin from bFF using dye affinity, anion-exchange, lentil lectin affinity chromatography and a subsequent reversed phase chromatography step, two pools of inhibin activity were obtained. These were separated by SDS-PAGE revealing 30 and 58 kDa inhibin forms. The immunoactivity of these forms of inhibin was then estimated using antibodies against the 22 N-terminal amino acids of the alpha subunit of 30 kDa bovine inhibin. It appeared that the two molecular weight forms of inhibin had bioactivity/immunoactivity ratios which differed more than five-fold. This indicates that results of radioimmunoassays of inhibin of ovarian origin, using peptide antisera, should be interpreted with caution.