Histamine Reduces ZO-1 Tight-Junction Protein Expression in Human Immortalized Podocytes

E Veglia1, A Moggio1, A Pini2, C Grange3, P Chazot4, AC Rosa1. 1Dipartimento di Scienza e Tecnologia del Farmaco Università di Torino, Torino, Italy, 2Dipartimento di Medicina Sperimentale e Clinica Sezione di Anatomia e Istologia, Università degli Studi di Firenze, Firenze, Italy, 3Dipartimento di Scienze Mediche, Università di Torino, Torino, Italy, 4School of Biological and Biomedical Sciences and Wolfson Research Institute, Durham University, Durham, UK

Independent studies suggest that the biogenic amine histamine may be involved in renal pathophysiology (1,2), affecting and regulating the renal microcirculation, increasing salt and water excretion, as well as decreasing the ultrafiltration coefficient (kf) by reducing the total filtration surface area, and increasing renin release (3-6). In particular, histamine reduces kf in the rat and this effect is blocked by a histamine H1 receptor (H1R) antagonist (3). It is well recognized that among the different cellular components of glomeruli, podocytes, terminally differentiated epithelial cells constituents of the glomerular filtration barrier (GFB), play a crucial role in retaining plasma proteins, by controlling the capillary surface area and thus the kf (7).

Although among the four known histamine receptors only H1R and H2R were previously identified in the glomeruli (1,8), the full component of histamine receptor subtype(s) present on podocytes remains to be determined. Thus, the present study was designed to evaluate the comparative expression and distribution of the histamine receptors in human immortalized podocytes and whether histamine affects the expression of the zonula occludens-1, ZO-1. Indeed, this tight-junction protein is localized in the podocytes foot processes and its alterations in expression, localization and phosphorylation are associated with increased paracellular permeability (7).

For this purpose, in human immortalized podocytes the gene and protein expression of the four histamine receptors was evaluated by RT-PCR and immunofluorescence. Furthermore, the molecular analysis was confirmed pharmacologically by TR-FRET measurement of both IP3 and cAMP production evoked by histamine (3 pM-10 nM). Finally, the effect of histamine (range 0.1-1000 nM) on ZO-1 expression was assessed.

Our data revealed the presence of H1R, H2R and H4R. However, only H1R was predominantly localized on the membrane, with H2R and H4R expressed intracellularly. Consistently with the anatomical observations, histamine did not modulate cAMP production, while it affected IP3 levels. Indeed, the amine elicited a sigmoid dose-response curve for IP3 in the range 3 pM-10 nM which was shifted to the right by the competitive H1R selective antagonist chlorpheniramine maleate at 10 μM.

When the effect of histamine on ZO-1 expression was evaluated, a significant reduction was measured with a maximum effect (30%) after 6 h of histamine exposure.

In conclusion, the data reported herein provide, for the first time, the topological and pharmacological identification of H1R in the human podocytes, demonstrating that histamine exert a detrimental direct effect on this cell type by affecting ZO-1 expression.

All together this new knowledge support the hypothesis for the role of histamine in renal pathophysiology.

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