Low-dose SKA preparation of Progesteron and Interleukin-10 is effective in the modulation of the inflammatory response in an in vitro model of endometriosis
Francesca Mancini (IT), Giuseppe Grande (IT), Domenico Milardi (IT), Piero Carfagna (IT), Fiorenzo De Cicco Nardone (IT), Riccardo Marana (IT)
[Mancini] International Scientific Institute "Paul VI" - Catholic University, Rome (Italy), [Grande] International Scientific Institute "Paul VI" - Catholic University, Rome (Italy), [Milardi] International Scientific Institute "Paul VI" - Catholic University, Rome (Italy), [Carfagna] International Scientific Institute "Paul VI" - Catholic University, Rome (Italy), [De Cicco Nardone] Department of Obstetrics and Ginecology, Fondazione Policlinico "A. Gemelli", Rome (Italy), [Marana] International Scientific Institute "Paul VI" - Catholic University, Rome (Italy) AND Department of Obstetrics and Ginecology, Fondazione Policlinico "A. Gemelli", Rome (Italy)
Context: The oral use of low-dose SKA of Anti-IL-1 antibodies and Interleukin-10 is effective in the modulation of the inflammatory response in models of diseases presenting an important inflammatory component. Objective: The aim of this pre-clinical study was evaluate the efficacy of low-dose SKA Progesterone (GUNA) and low-dose SKA IL-10 (GUNA) in the modulation of the inflammatory response in endometriotic cell lines. Methods: Immortalized human endometriotic epithelial (Z-12) cells were derived from peritoneal lesions. For Western blot, cells were lysed in RIPA buffer supplemented with a cocktail of protease inhibitors (Boehringer). Nuclear and cytoplasmic fractions were prepared as follows: Cells were resuspended in hypotonic lysis buffer added with protease inhibitors (Boehringer). After resuspension, NP-40 was added to a final concentration of 0.6% and the nuclei was isolated by centrifugation. NfkB Transcription factor Assay was performed according to manufacturer’s instruction (Abcam). Interventions: Z12 cells were treated with low-dose SKA progesterone (10pg/ml) and with low dose SKA IL-10 (10 fg/ml). Results: Treatment with low-dose SKA progesterone significantly decreased estradiol 17β-dehydrogenase1 (HSD17B1) protein expression (about 20% of reduction). Importantly, was observed a significant reduction (about 15-20%) in the levels of nuclear translocation of NF-kB determined by nuclear-cytoplasmic fractionation and NFkB DNA binding assay. Treating the Z12 cell line with low dose SKA IL-10 resulted in a significant reduction (about 15%) in the levels of nuclear translocation of NF-kB. Conclusions: These results clearly demonstrate the role of treatment with low-dose SKA Progesterone and SKA IL-10 in modulating the NF-kB inflammatory pathway in endometriosis.