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Home » Archive » 2020

TDK conference 2020

Pharmacological and toxicological characterization of S1P as a treatment option in anemias
Paréj Zsuzsanna - year 5
University of Veterinary Medicine Budapest, Department of Pharmacology and Toxicology
Supervisors: Judit Mercédesz Pomothy, Dr. Eszter Ágnes Czimmermann

Abstract:

Sphingosine-1-phosphate (S1P) has been reported as a nonselective matriptase activator. Matriptase-1 (MT-1) and matriptase-2 (MT-2) belong to the type II transmembrane serine protease family. However MT-1 is widely expressed in epithelial tissues, MT-2 is mainly expressed in the liver and also in small amounts in the kidney. MT-2 can regulate the iron homeostasis via modulation of hepcidin level. The aim of our study was to reveal if S1P can influence hepcidin production on primary human and rat hepatocytes after 72 hours of incubation via modulating the MT-2. Investigations were done on how S1P can affect the viability and the redox status of primary hepatocytes of human and rat primary hepatocytes. Also the distribution of the occludin, as a tight junction protein on primary rat hepatocytes was examined. In the case of human cells, cytochrom P450 3A4 (CYP3A4) enzyme interaction was also measured. Primary rat hepatocytes were treated with S1P at 50, 200 és 1000 ng/ml concentrations, while human hepatocytes were incubated with 500, 1000, 200 ng/ml S1P. Cell-free supernatants were collected every 24 hours. Cell viability was tested by a colorimetric method on both cells, the hepcidin levels in the cell-free supernatants were examined by hepcidin sandwich ELISA to determine the effect of S1P on the hepcidin-modulating ability of MT-2. In order to estimate the extent of S1P generated oxidative stress, extracellular H2O2 measurements were performed by the use of fluorescent method. Based on the findings, S1P treatment did not cause cell death for 72 hours at the highest concentrations nor in rat neither in human primary hepatocytes at any applied concentrations within the investigated time period. S1P did not influence the extracellular H2O2 production for 72 hours in any of our examined species. The distribution of occludin was not altered after treating the primary rat hepatocytes with S1P. The hepcidin levels were significantly suppressed after 72 hours in hepatocytes exposed to S1P treatment in both species. In the case of human primary hepatocytes we observed inhibition of CYP3A4 enzyme activity only in the highest concentration. These findings might presume that S1P might be a drug candidate in treatment of iron deficient anaemias, as any of the used concentrations were safe to apply and the hepcidin level significantly altered after the treatment, however drug-drug interaction might appear at the highest applied concentration.



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