ABSTRACT

Peripheral blood mononuclear cells were collected from a sheep immunized against progesterone-11α-hemisuccinate-ovalbumin. Following fusion with NS1 mouse myeloma or heteromyeloma cells, a large number of hybrid colonies was established. These were screened for the production of sheep antibodies to progesterone. Twenty-four cell lines were cloned and one was stabilized. This cell line, O/MP.1A9.D7B2, produced a high-affinity ovine immunoglobulin G1 (dissociation constant 4·8 pmol/l) with a high degree of specificity for progesterone. The antibody was substituted into a competitive enzyme-linked immunosorbant assay for the measurement of progesterone in bovine milk, originally established using an ovine polyclonal antibody, and the results were compared. The monoclonal antibody produced an assay with a lower limit of detection and a greater degree of discrimination than the polyclonal antiserum.

Journal of Endocrinology (1990) 126, 217–222

ABSTRACT

Evidence in the literature suggests that the trace element chromium may have a role in glucose homeostasis through the regulation of insulin action. We have previously reported a significant reduction in plasma chromium levels in healthy individuals, following a 75 g oral glucose load, and after meals and glucose-dependent uptake of chromium in insulindependent tissues in vitro. However, in vivo it is unclear whether the changes in plasma chromium relate to changes in plasma glucose or insulin.

The present study describes a series of euglycaemic hyperinsulinaemic clamps designed to attempt to define the initiator of changes in plasma chromium levels in ten healthy individuals.

The data showed a significant (P<0·01) reduction in fasting plasma chromium levels following glucose infusion and an initial bolus of insulin. Significant (P<0·02) increases in post-clamp urinary chromium excretion were insufficient to explain the decrease in plasma levels. During the recovery phase of an extended two-phase clamp protocol we found plasma insulin levels decreased by 70% within 10 min, associated with an increase in plasma chromium levels of 30% and no significant change in plasma glucose level.

These data indicate that alterations in plasma glucose are unlikely to be directly related to changes in plasma chromium, whilst supporting the hypothesis that plasma insulin may influence plasma levels of this trace element. In contrast, plasma zinc was unaffected throughout these clamp studies.

Journal of Endocrinology (1993) 139, 339–345

ABSTRACT

The putative endocrine role of inhibin in the control of FSH secretion during the luteal phase in the primate was investigated by immunoneutralization. Antisera against the 1–23 amino acid sequence of the N-terminus of the human inhibin α subunit were raised in a ewe and three macaques. Antisera (10–20 ml) were administered to macaques on day 8/9 of the luteal phase and serum samples collected during the treatment cycle and post-treatment cycle for determination of FSH, oestradiol and progesterone. In addition, localization of inhibin within the macaque ovary at this stage of the luteal phase was investigated using the ovine antiserum. Intense immunostaining was localized within the granulosa-lutein cells of the corpus luteum with absence of staining in the thecalutein cells or other ovarian compartments. Administration of antisera was without significant effect on serum concentrations of FSH when compared with control animals, either during the first 24 h of detailed observation or for the following 10-day period of the late luteal phase and subsequent early follicular phase. These results provide further evidence that the corpus luteum is the major source of inhibin immunoreactivity during the primate menstrual cycle, but fail to support an endocrine role for inhibin in the suppression of FSH secretion.

Journal of Endocrinology (1992) 133, 341–347

Macrophage migration inhibitory factor (MIF) is an essential regulator of the macrophage responses to endotoxin. MIF also has the ability to override the anti-inflammatory actions of glucocorticoids during an immune response, and is thus an important pro-inflammatory factor. The presence of MIF in cells of the anterior pituitary has been described, and high levels of MIF in other rapidly proliferating tIssues have also been demonstrated. It has been hypothesised that MIF release from these cells is influenced by the hypothalamo-pituitary-adrenal axis, and that ACTH and MIF are released simultaneously to exert counter-regulatory effects on cortisol. However, another intracellular role for MIF has also been suggested as it has been shown that MIF exerts an effect on the inhibitory cell cycle control protein p27 through an interaction with Jab1, a protein implicated in p27 degradation. We studied MIF expression in different normal and adenomatous human pituitary samples using immunohistochemistry and RT-PCR. There was evidence of co-immunoprecipitation of MIF with Jab1, suggesting an interaction of the two proteins. Our results showed that there is increased expression of MIF protein in the nuclei of all pituitary adenomas compared with normal tIssue (P=0.0067), but there was no statistically significant difference in nuclear MIF expression between the different adenoma types. Nuclear MIF expression correlated positively with p27 and its phosphorylated form in normal tIssue (P=0.0028 and P<0.0001); however, this relationship was not seen in the adenoma samples. Cytoplasmic expression of MIF was found to be variable both in normal and adenomatous samples, with no consistent pattern. MIF mRNA was demonstrated to be present in all tumour and normal samples studied. Somatotroph tumours showed higher MIF mRNA expression compared with normal pituitary or other types of adenomas. In conclusion, MIF is expressed in cell nuclei in pituitary adenomas to a greater extent than in normal pituitary tIssue. We speculate that it may play a role in the control of the cell cycle, but whether its higher level in adenomas is a cause or a consequence of the tumorigenic process remains to be clarified.