Development and Validation of a Radioimmunoassay for Peptides Related to β-Melanocyte-Stimulating Hormone in Human Plasma: The Lipotropins

Abstract

A radioimmunoassay is described for the measurement of human "beta-melanocyte-stimulating hormone" ("betah-MSH"). Two antisera have been used, one of which cross-reacts with synthetic betah-MSH as well as with the two larger pituitary peptides betah- and gammah-lipotropin (betah- and gammah-LPH) and the other mainly with betah-MSH and gammah-LPH. The sensitivity and reliability of the assay have been improved by employing a simple plasma extraction procedure, and the shelf-life of the iodinated betah-MSH tracer has been increased more than five-fold by storage in a concentrated human serum albumin solution. Using a 5 ml plasma sample the detection limit is 6 pg/ml. The mean resting "betah-MSH" level in normal subjects is 21 pg/ml (range 13-38 pg/ml) at 9 AM and 12 pg/ml (range 6-20 pg/ml) at 9 PM. Levels are considerably elevated (51-12,000 pg/ml) in patients with Addison's disease. Nelson's syndrome, Cushing's disease and the "ectopic" ACTH syndrome. After administration of insulin or pyrogen, the concentration of plasma "betah-MSH" increases in parallel with that of ACTH and they are approximately equivalent on a molar basis. The stability of purified betah- and gammah-LPH and endogenous "betah-MSH" when incubated in vitro in fresh blood or plasma are similar, in contrast to the less stable peptide synthetic betah-MSH. It is suggested that "betah-MSH" immunoreactivity in human plasma is due to betah- and gammah-LPH rather than betah-MSH.

Simultaneous Assay of Immunoreactive β-Lipotropin, γ-Lipotropin, and β-Endorphin in Plasma of Normal Human Subjects, Patients with ACTH/Lipotropin Hypersecretory Syndromes, and Patients undergoing Chronic Hemodialysis

“…2), and it is probable that their mean concentrations are, therefore, similar. Gilkes et al (8), using antiserum NZ, which crossreacts equally with (37-58)h-yLPH and h'yLPH, but only 3% as well with h,8LPH, reported plasma IR-(37-58)hyLPH in normal controls in the range of 25 pg/ml. Since the NZ antiserum presumably was measuring only hyLPH, there being no (37-58)hyLPH ("h,BMSH") in normal human plasma, this would correspond to about 10 fmol/ml of IR-hyLPH, a value compatil)le with our findings.…”

“…The human f8-melanocyte-stimulating hormone (hI3MSH)l immunoreactivity of human tissues and plasma is now generally accepted to be due to two larger molecules, called lipotropins (LPH), both of which contain the sequence of "hf3MSH" in their structures (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13). The LPH have been isolated from putuitary extracts of several species (14)(15)(16)(17)(18) including (7)(8)(9)(10)(11)(12)(13) and/or possibly hyLPH (11) on the basis of their apparent molecular weights.…”

“…The LPH have been isolated from putuitary extracts of several species (14)(15)(16)(17)(18) including (7)(8)(9)(10)(11)(12)(13) and/or possibly hyLPH (11) on the basis of their apparent molecular weights. Studies of extracts of human tissues, plasma, and media in which ACTH/LPH-producing human pituitary tumor cells were cultured, using gel exclusion chromatography and denaturing conditions, indicated the presence of hyLPH alone or both hyLPH and h,3LPH (11,13).…”

Beta endorphin selectively stimulates aldosterone secretion in hypophysectomized, nephrectomized dogs.

“…The authors concluded that #lMSH was the aldosterone-stimulating core of #lLPH. It should be noted, however, that f#MSH does not normally exist in human plasma, but that it may be an artefact produced by degradation of #LPH during extraction (29,30).…”

“…The peptides were dissolved in normal saline and equimolar doses (3 pmol/min) were infused into the arterial pouch for 50 min. Beginning 30 min after completion of the preparation of the animal, duplicate blood samples for measurement of plasma aldosterone and cortisol were collected from the venous pouch at 0, 20 min (control), 30 (23,24).…”