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Silver (Ag) NPs are among the most commercialized NPs due to their antimicrobial potential. They are highly at- tractive for possible applications in manufacture of medical device. However there is a serious lack of information concerning their impact on the human health and the environment. Moreover studies on the effects of NPs on ejaculated sperm are rather limited. For these reasons our study explored the in vitro effects of Ag NPs on human ejaculated spermatozoa. Ag NPs have been produced, characterized, and furnished by Colorobbia Industry, Sovigliana (Vinci, Florence, Italy). Aliquots of total semen were incubated at 37°C for 60 minutes (min) and 120 min at the concentration of 125 μM, 250 μM, and 500 μM of engineered Ag NPs. The control was represented by specimens of semen samples treated with the same procedure without NPs. After the incubations, sperm motility was evaluated following WHO guidelines and sperm viability was eval- uated by Eosin Y test. At the end of incubation with Ag NPs the samples were processed by a Field Emission Gun-based Scanning Transmition Electron Microscope/ Energy Dispertion Spectrometry (STEM/EDS). We observed that sperm motility percentage was significantly reduced in semen samples treated with 125 μM, 250 μM and 500 μM of Ag NPs after 60 min and 120 min of incubation respect to controls (P<0.001; P<0.01, 125 μM at 60 min). Sperm viability percentage significantly decreased in a progressive manner after 125 μM (P<0.05), 250 μM (P<0.05) and 500 μM (P<0.001) Ag NPs incubation at 60 min and 120 min. We did not find any significant difference between the values assessed after 60 min of NPs incubation and those estimated after 120 min of incubation. In the control samples, the sperm motility and the sperm viability percentages significantly decreased after 120 min of incubation (P<0.001) respect to the basal values. Ag NPs were undetectable in all treated samples by STEM/EDS. These in vitro results show a decline in sperm motility and viability in even at the lowest concentration used and the cytotoxic effect occurs in a dose dependent manner. It is noteworthy that in each experiment, for each concentration of NPs used, the percentage of sperm viability was always higher than the percentage of sperm motility; it means that spermatozoa were viable but immotile. Moreover Ag NPs was undetectable in all the treated samples by STEM/EDS analysis. We may hypothesize that Ag NPs, under aqueous conditions, release Ag+ that could damage sperm membrane and/or penetrate inside the cells and interfere with disulphide bonds of proteins of the periassonemal structures of the sperm tail.
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