An enzyme-linked immunosorbent assay to study human relaxin in human pregnancy and in pregnant rhesus monkeys

A sensitive and specific double-antibody enzyme-linked immunoassay, using a synthetic analogue of human relaxin for standard and immunogen, was developed for the measurement of human relaxin (hRLX) in serum and plasma. No cross-reactivity was observed for human insulin, human insulin-like growth factor-I, hGH, human chorionic gonadotropin, hFSH, hLH or human prolactin. The assay was used to monitor RLX concentrations in samples from men, non-pregnant and pregnant women, and in pregnant rhesus monkeys infused with hRLX. RLX was not detected in serum from men nor from non-pregnant women, while a concentration of 600 ng/l was measured in pooled sera from two pregnant women (pregnancies achieved by in-vitro fertilization). Immunoreactive RLX (1.1 micrograms/g) was found in human corpora lutea taken from ectopic pregnancies at 7 weeks. In an experiment with a pregnant rhesus monkey infused with human RLX analogue, less than 1.5% of the maternal concentration was measured in the fetal circulation. Even though preliminary, these data suggest a low level of transfer of human analogue relaxin across the placenta in a rhesus monkey. Further studies of the physiology of RLX in human pregnancy will be facilitated by the availability of this immunoassay.     

Pharmacokinetics and Tissue Distribution of Recombinant Human Transforming Growth Factor Beta1 After Topical and Intravenous Administration in Male Rats

Recombinant human transforming growth factor beta (rhTGF-₁) enhances the healing process after topical application to various animal wound models. A detailed pharmacokinetic and tissue distribution study was performed to support the clinical development of rhTGF-₁ for wound healing indications. Rats received radioiodinated or unlabeled rhTGF-₁ as an intravenous (iv) bolus or as a topical formulation applied to a full thickness wound. Plasma concentrations of TGF-₁ were estimated from TCA-precipitable radioactivity or were measured by ELISA. Following iv administration, the initial half-life was rapid (<11 min), regardless of whether radi-olabeled or unlabeled rhTGF-₁ was used. The terminal half-life was long (163 min) when the test material was radioiodinated and administered as a trace dose and relatively short (61 min) when given at high doses and assayed by ELISA. Analysis of plasma radioactivity by SDS-PAGE revealed a time-dependent clearance of the 25-kDa parent molecule without a significant appearance of lower molecular weight radiolabeled metabolites. The majority of the radioactivity was associated with highly perfused organs, known iodide elimination pathways, and the thyroid at 1 and 8 hr after iv injection. After topical administration of a high dose (0.8 mg/kg), no immunoreactive TGF-₁ was detectable in plasma samples taken over a 48-hr period. However, trace amounts (0.05 ng/mL) of acid-precipitable radioactivity were detected in plasma after topical application of [¹²⁵I]rhTGF-₁ (1 µg/kg, 126 µCi/kg). A significant portion (35%) of the [¹²⁵I]rhTGF-₁ persisted intact (25 kDa) at the wound site 24 hr after application. In conclusion, rhTGF-₁ was rapidly cleared after iv bolus administration and distributed primarily to the liver, lungs, kidney, and spleen. Little systemic exposure was observed after applying a single topical dose of rhTGF-₁ to a wound, and the intact molecule persisted at the wound site.