NMR spectroscopy of live human asthenozoospermic and normozoospermic sperm metabolism

Sperm motility varies between ejaculates from different men and from individual men. We studied normozoospermic and asthenozoospermic ejaculates after density gradient centrifugation washing (DCG, 80/40%) and compared high (80%) and low (40%) motility sperm populations within the same sample. Our objective was to identify differences in endogenous metabolomes and energy metabolism in relation to sperm motility. 1H-Nuclear Magnetic Resonance Spectroscopy (NMR) measured the endogenous metabolome of live human sperm. Incubating sperm with 13C-labelled substrates detected energy metabolism by 13C-NMR. The studied examined 850 ejaculates and diagnosed asthenozoospermia in 6.1%. DGC was used to wash 160 normozoospermic (N) and 52 asthenozoospermic (A) ejaculates to recover high motility sperm from the pellet (80N/80A) and low motility from the interface (40N/40A). 1H-NMR spectra, 45(N), 15(A), were binned and the integrals normalised by sperm concentration. Sperm from 126(N) and 36(A) ejaculates were incubated with either 13C-glucose, 13C-fructose or 13C-pyruvate. 13C-NMR lactate and bicarbonate integrals were normalised by motile or vital sperm concentrations. 1H-NMR spectra choline integrals from the 80A population were significantly lower than the 80N, p<0.0001. 13C-substrate conversion to lactate was significantly higher for 40A sperm than 80A sperm when normalised by motile sperm concentration. Bicarbonate integrals were sporadically observed. Sperm from asthenozoospermic ejaculates had similar glycolytic requirements to normozoospermic ones, with larger differences observed between 40% and 80% sperm populations. Higher lactate levels produced by 40% sperm may indicate that impaired sperm motility is due to dysregulated energy metabolism. The alteration in choline metabolism provides opportunities to understand the aetiology of asthenozoospermia.