Cytokine hormones have a short plasma half-life and require frequent administration. For example, growth hormone replacement involves daily injections. In common with other cytokines, the extracellular domain of the growth hormone receptor circulates as a binding protein, which naturally prolongs the biological half-life of growth hormone. Here we have studied the biological actions of a ligand-receptor fusion of growth hormone and the extracellular domain of its receptor. The genetically engineered ligand-receptor fusion protein was purified from mammalian cell culture. In rats, the ligand-receptor fusion had a 300-times reduced clearance as compared to native growth hormone, and a single injection promoted growth for 10 d, far exceeding the growth seen after administration of native growth hormone. The ligand-receptor fusion forms a reciprocal, head-to-tail dimer that provides a reservoir of inactive hormone similar to the natural reservoir of growth hormone and its binding protein. In conclusion, a ligand-receptor fusion of cytokine to its extracellular receptor generates a potent, long-acting agonist with exceptionally slow absorption and elimination. This approach could be easily applied to other cytokines.
In anesthetized and ventilated guinea pigs intravenous injection of ET-1, ET-2, or ET-3 (1 or 2 nmol/kg) induced similar dose-dependent increases in pulmonary inflation pressure (PIP) associated with increases in mean arterial blood pressure (MBP). Pretreatment of the guinea pigs with 1 mg/kg intravenous indomethacin significantly inhibited the increase in PIP evoked by 2 nmol/kg of ET-1, ET-2, or ET-3. In contrast, the increase in MBP following injection of the various ET isotypes was not significantly affected by indomethacin. Injection of ET-1, ET-2, or ET-3 (40, 120, and 400 pmol) via the pulmonary artery of isolated and perfused guinea pig lungs induced dose-dependent increases in PIP and pulmonary perfusion pressure (PPP), thromboxane B2 (TXB2) release, and formation of lung edema. In keeping with the in vivo results, no marked differences were observed between the activities of ET-1, ET-2, and ET-3 on isolated and perfused guinea pig lungs. Indomethacin (5 microM) added to the perfusion medium significantly inhibited the alterations of PIP and PPP, TXB2 release, and edema formation evoked by 400 pmol ET-1, ET-2, or ET-3. High-affinity binding sites for ET-1, ET-2, and ET-3 exhibiting similar characteristics were identified on guinea pig lung membrane. Therefore ET-1, ET-2, and ET-3 exert comparable bronchopulmonary and pressor activities in the guinea pig and probably act via interaction with the same binding site. In addition, the ET-induced increase in PIP and pulmonary vasoconstriction are primarily mediated via the production of cyclooxygenase metabolites.
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