Collectively, our results demonstrated for the first time that in thyroid cancer cells hydroxytyrosol promoted apoptosis at higher doses with respect to other cancer cells lines. Therefore, further studies will reveal the mechanisms by which thyroid cancer cells are more resistant to the proapoptotic effect exerted by hydroxytyrosol as well as the potential application as novel target therapeutic in thyroid cancer.

Experimental evidence demonstrated that macroautophagy/autophagy exerts a crucial role in maintain renal cellular homeostasis and represents a protective mechanism against renal injuries. Interestingly, it has been demonstrated that in the human proximal tubular renal cell line, HK-2, the MTOR inhibitor rapamycin enhanced autophagy and mitigated the apoptosis damage induced by urinary protein overload. However, the underlying molecular mechanism has not yet been elucidated. In our study we demonstrated, for the first time, that in HK-2 cells, the exposure to low doses of rapamycin transactivated the NGFR promoter, leading to autophagic activation. Indeed, we observed that in HK-2 cells silenced for the NGFR gene, the rapamycin-induced autophagic process was prevented, as the upregulation of the proautophagic markers, BECN1, as well as LC3-II, and the autophagic vacuoles evaluated by transmission electron microscopy, were not found. Concomitantly, using a series of deletion constructs of the NGFR promoter we found that the EGR1 transcription factor was responsible for the rapamycin-mediated transactivation of the NGFR promoter. Finally, our results provided evidence that the cotreatment with rapamycin plus albumin further enhanced autophagy via NGFR activation, reducing the proapoptotic events promoted by albumin alone. This effect was prevented in HK-2 cells silenced for the NGFR gene or pretreated with the MTOR activator, MHY1485. Taken together, our results describe a novel molecular mechanism by which rapamycin-induced autophagy, mitigates the tubular renal damage caused by proteinuria, suggesting that the use of low doses of rapamycin could represent a new therapeutic strategy to counteract the tubule-interstitial injury observed in patients affected by proteinuric nephropathies, avoiding the side effects of high doses of rapamycin.

The mesothelial cells (MCs) play an important role in the morpho-functional alterations of the peritoneal membrane (PM) undergoing peritoneal dialysis (PD). MCs, through the epithelial-mesenchymal transition process (EMT), progressively acquire a myofibroblast-like phenotype, promoting peritoneal fibrosis (PF) and failure of peritoneal membrane function. Transforming growth factor β1 (TGFβ1), through canonical and non-canonical pathways, promotes the epithelial-mesenchymal transition (EMT) process leading to PF. To investigate the therapeutic potential of an olive leaf extract (OLE) on preserving peritoneal membrane function, we evaluated the effect of OLE on the TGFβ1-induced EMT in mesothelial cells, Met5A, and elucidated the underlying molecular mechanisms. As assessed by changes in the expression of epithelial, mesenchymal, and fibrotic cell markers (such as E-cadherin, N-cadherin, α-SMA, fibronectin, vimentin), levels of matrix metalloproteinases (MMP2 and MMP9), and cell migration, OLE inhibited the TGFβ1-induced EMT. Importantly, the beneficial effect of OLE was mediated by reduction of the TGFβ1-induced activation of Smad2/3 signaling and the mitigation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (MAPK) phosphorylation. Smad/non-Smad signaling pathways, activated by TGFβ1, both reduce expression of epithelial marker E-cadherin which has a crucial role in EMT initiation. Interestingly, we observed that in presence of OLE activity of the E-cadherin, promoter was increased and concomitantly OLE reduced the nuclear content of its co-repressor SNAIL. Our results suggest the potential therapeutic of OLE to counteract fibrotic process in peritoneal dialysis patients.

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Our results suggest that in TX recipients an appropriate sexual hormonal evaluation should be performed, as we found a high prevalence of TD. However, further studies are needed to clarify the association between TD and patient and graft survival.