Physiologic Determinants of Endothelin Concentrations in Human Saliva

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Abstract
Background: Salivary endothelin (ET) concentrations have been shown to correlate with disease severity in patients with chronic heart failure (CHF). We undertook the present study to evaluate the stability of salivary ET under different handling conditions to assess its suitability as a biochemical marker in screening, diagnosis, and management of CHF. Methods: Saliva samples were collected from healthy individuals and/or CHF patients, subjected to different handling conditions, and then stored at −80 °C until assayed by an ELISA for ET. Results: Salivary ET concentrations showed a time-dependent increase during storage at room temperature. After 72 h of incubation at room temperature, ET increased ∼2.8-fold (P = 0.03). Simultaneously, salivary big ET showed a time-dependent 11.2-fold decrease (P <0.0001). This activity was blocked by an ET-converting enzyme (ECE) inhibitor, suggesting that these changes were attributable to ECE-dependent cleavage of endogenous big ET in saliva. Ex vivo conversion was also observed when samples were stored at 4 °C, but the magnitude of these changes was markedly smaller (P <0.0001). Posture also affected salivary ET concentrations in CHF patients. With a change from supine to seated rest, salivary ET concentrations increased 1.5- and 1.8-fold after 20 and 40 min, respectively (P = 0.01). With a return to supine rest, salivary ET concentrations returned to baseline concentrations (P = 0.008). Conclusions: These data suggest that saliva sampling and handling conditions could markedly affect measurement of salivary ET. In particular, care should be taken to minimize ECE-dependent enzymatic conversion of endogenous big ET in saliva.