Variety of Angiotensin Receptors in 3T3-L1 Preadipose Cells and Differentiated Adipocytes

A local paracrine acting angiotensin (ANG) system of preadipocytes and mature adipocytes is involved in metabolic effects and tissue differentiation. The present study reports on the investigation of binding affinities for various angiotensin receptors including their relevance in 3T3-L1 adipocytes and preadipocytes and 3T3-442A preadipocytes. Competitive binding studies using both I-125-ANG II and its more stable analogue I-125-SARILE for investigating AT(1)/AT(2) binding sites in 3T3-L1 preadipocytes reveal a biphasic competition curve with KDS at a low and high nanomolar range. By using the AT(2) receptor selective ligand I-125-CGP4112A the presence of high affinity AT(2) binding sites in preadipocytes was observed. High nonspecific binding and a low receptor number is characteristic for all these experiments. An AT(4) binding site (binding site for ANG IV) exists in 3T3-L1 and F442A preadipocytes and adipocytes with a high nanomolar K-D. This low binding affinity was confirmed by a biological assay, the IRAP assay (= insulin regulated aminopeptidase assay). IRAP is associated with the AT(4) receptor, which is a binding site at the luminal part of membrane bound IRAP. The curves for competition binding and for inhibition of IRAP activity are superimposable with respect to angiotensin IV. In conclusion, AT(1) and AT(2) binding sites are present in preadipocytes. AT(2) receptor binding affinities are shown in preadipocytes for the first time. The description of a non-AT(1)/AT(2) binding site with low affinity remains speculative albeit of high interest because antidiabetic and obesity related effects of angiotensin peptides and sartanes as antagonists are observed at these high concentrations. Local concentrations of ANG II and their degradation products may be extremely high. The low amounts of AT(1) and AT(2) binding sites emphasize the relevance of other binding sites in adipose tissue development and metabolic effects. The AT(4) binding site seems to be one of the predominant receptors in adipose cells. Other degraded, but still bioactive peptides like ANG III, IV and ANG(1-7), activating receptors not influenced by ANG II, Could be of importance.