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Small noncoding RNAs and sperm nuclear basic proteins reflect the environmental impact on germ cells.

BACKGROUND: Molecular techniques can complement conventional spermiogram analyses to provide new information on the fertilizing potential of spermatozoa and to identify early alterations due to environmental pollution. METHODS: Here, we present a multilevel molecular profiling by small RNA sequencing and sperm nuclear basic protein analysis of male germ cells from 33 healthy young subjects residing in low and high-polluted areas. RESULTS: Although sperm motility and sperm concentration were comparable between samples from the two sites, those from the high-pollution area had a higher concentration of immature/immune cells, a lower protamine/histone ratio, a reduced ability of sperm nuclear basic proteins to protect DNA from oxidative damage, and an altered copper/zinc ratio in sperm. Sperm levels of 32 microRNAs involved in intraflagellar transport, oxidative stress response, and spermatogenesis were different between the two areas. In parallel, a decrease of Piwi-interacting RNA levels was observed in samples from the high-polluted area. CONCLUSIONS: This comprehensive analysis provides new insights into pollution-driven epigenetic alterations in sperm not detectable by spermiogram.

Male

On the cytochemical demonstration of basic proteins in the cell nucleus, including the nucleolus.

A combined Feulgen-alkaline fast green method is described for simultaneous demonstration of DNA and basic proteins in the cell nucleus. The method is based on preserving both types of substances in the tissue section and releasing in them reactive groups for the 2 kinds of staining. These conditions are best provided, as proved by staining tests on tissue hydrolysates, if formalin-containing mixtures (SERRA's or LILLIE's fluids) are employed for fixation, and cold 5 N HCl is used for FEULGEN hydrolysis. In this way, a good cytological picture is also achieved. Nuclear euchromatin stains with this method red, while heterochromatin, pycnotic nuclei and sperm heads exhibit a deep violet to blue-violet colour. Prominent nucleoli of metabolically active cells display a distinct blue-green staining thus manifesting their high content of basic proteins. Acetylation test reveals that these proteins are of lysine-rich type. The known negative reaction of the nucleoli with the routine alkaline fast green method according to ALFERT and GESCHWIND must be attributed to an extraction of the nucleolar basic proteins with the hot TCA used in this method. Certain analogy in the cytochemical behaviour between the nucleolous and the chromatin under various conditions of hydrolysis leads to the suggestion that the nucleolar basic proteins demonstrated should be in the form of a ribonucleoprotein complex, probably of the pre-ribosomal material of the nucleolus.

Animals