1. Separate binding sites for intact PTH 1-84 and synthetic PTH 1-34 in canine kidney
J C Garcia, C L McConkey, K J Martin Calcif Tissue Int. 1989 Mar;44(3):214-9. doi: 10.1007/BF02556567.
The present studies examine the characteristics of parathyroid hormone (PTH) receptor binding in canine basolateral renal cortical membranes using iodinated preparations of intact bovine PTH 1-84 and [Nle8,Nle18,Tyr34] bPTH 1-34 amide. A solid phase lactoperoxidase technique was used to iodinate the bPTH 1-84. The PTH 1-34 analog was iodinated using chloramine T. Both radioligands were purified by reverse phase high pressure liquid chromatography (HPLC). Specific binding of 125I PTH 1-84 reached equilibrium at 3 hours whereas binding of the 125I PTH 1-84 analog reached equilibrium at 45 minutes. Excess bPTH 1-84 resulted in complete inhibition of binding of 125I bPTH 1-84, whereas 22 +/- 1.6% of the bound radioligand remained bound in the presence of excess synthetic bPTH 1-34. These data suggested the possibility of a binding site for the carboxy-terminal region of intact PTH, or binding sites selective for intact hormone. Therefore, additional studies were performed with PTH fragments, PTH 28-53, PTH 35-84, and PTH 53-84. In contrast to previous studies in other systems, these fragments did not result in significant displacement of 125I PTH 1-84. Analysis of binding of 125I PTH 1-84 and 125I [Nle8,Nle18,Tyr34] PTH 1-34 amide, using LIGAND, both indicated a single site model with similar affinities. Thus, the data are consistent either with multiple receptors with similar affinities or a second binding site for bPTH 1-84 on the same receptor.(ABSTRACT TRUNCATED AT 250 WORDS)
2. Dynamics of secretion and metabolism of PTH during hypo- and hypercalcaemia in the dog as determined by the 'intact' and 'whole' PTH assays
Jose C Estepa, Ignacio Lopez, Arnold J Felsenfeld, Ping Gao, Tom Cantor, Mariano Rodríguez, Escolastico Aguilera-Tejero Nephrol Dial Transplant. 2003 Jun;18(6):1101-7. doi: 10.1093/ndt/gfg104.
Background: Recent evidence has shown that the assay for 'intact' parathyroid hormone (I-PTH) not only reacts with 1-84 PTH but also with large non-1-84 PTH fragments, most of which is probably 7-84 PTH. As a result, an assay specific for 1-84 PTH named 'whole' PTH (W-PTH) has been developed. The present study was designed: (i) to determine whether the W-PTH assay reliably measures PTH values in the dog; (ii) to evaluate differences between the W-PTH and I-PTH assays during hypo- and hypercalcaemia; and (iii) to assess the peripheral metabolism of W-PTH and I-PTH. Methods: In normal dogs, hypocalcaemia was induced by EDTA infusion and was followed with a 90 min hypocalcaemic clamp. Hypercalcaemia was induced with a calcium infusion. Results: I-PTH and W-PTH values increased from 36+/-8 and 13+/-3 pg/ml (P=0.01) at baseline to a maximum of 158+/-40 and 62+/-15 pg/ml (P=0.02 vs I-PTH) during hypocalcaemia. The W-PTH/I-PTH ratio, 38+/-4% at baseline, did not change during the induction of hypocalcaemia, but sustained hypocalcaemia increased (P<0.05) this ratio. During hypercalcaemia, maximal suppression for I-PTH was 2.0+/-0.5 and only 5.7+/-0.6 pg/ml for W-PTH, due to a decreased sensitivity of the W-PTH assay at values <5 pg/ml. The disappearance rate of PTH was determined in five additional dogs which underwent a parathyroidectomy (PTX). At 2.5 min after PTX, W-PTH was metabolized more rapidly, with a value of 25+/-2% of the pre-PTX value vs 30+/-3% for I-PTH (P<0.05). Conclusions: (i) The W-PTH/I-PTH ratio is less in the normal dog than in the normal human, suggesting that the percentage of non-1-84 PTH measured with the I-PTH assay is greater in normal dogs than in normal humans; (ii) the lack of change in the W-PTH/I-PTH ratio during acute hypocalcaemia is different from the situation observed in humans; and (iii) the dog appears to be a good model to study I-PTH and W-PTH assays during hypocalcaemia.
3. Difference between 1-84 parathyroid hormone and the 1-34 fragment on renal tubular calcium transport in the dog
J B Puschett, J McGowan, J Fragola, T C Chen, H Keutmann, M Rosenblatt Miner Electrolyte Metab. 1984;10(4):271-4.
Acute clearance studies were performed in chronically thyroparathyroidectomized dogs to determine similarities and differences between the effects of 1-34 and 1-84 parathyroid hormone (PTH) on urinary calcium excretion. In a small (physiological) dose, 1-84 PTH caused no mean change in percentage calcium excretion, while the 1-34 fragment was frankly calciuric. At this dose, the two hormone preparations were equally phosphaturic. Fragments of PTH in the range 25-84 did not alter calcium or phosphate transport. We conclude that the entire molecule is required in order for the distal nephron effect of the hormone to enhance net calcium transport in the kidney to become manifest.
