Rosmarinic acid improves boar sperm quality, antioxidant capacity and energy metabolism at 17°C via AMPK activation.

Boar sperm are susceptible to oxidative damage caused by reactive oxygen species (ROS) during storage. Adenosine monophosphate (AMP)-activated protein kinase (AMPK) is an important therapeutic target, because it is a cellular metabolism energy sensor and key signalling kinase in spermatozoa. We evaluated the effects of rosmarinic acid (RA), an antioxidant, on boar sperm during liquid storage to determine whether it protects boar sperm via AMPK activation. Boar ejaculates were diluted with Modena extender with different concentrations of RA and stored at 17°C for 9 days. Sperm quality parameters, antioxidant capacity, energy metabolism, AMPK phosphorylation and fertility were analysed. Compared with the control, 40 µmol/L significantly improved sperm motility, plasma membrane integrity and acrosome integrity (p < .05). The effective storage time of boar sperm was up to 9 days. On the third and seventh days, the sperm with RA exhibited increased total antioxidant capacity (T-AOC), superoxide dismutase (SOD) activity, adenosine triphosphate (ATP) content, mitochondrial membrane potential (ΔΨm) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activity, whereas malondialdehyde (MDA) content was significantly decreased (p < .05). Western blot showed that RA, as well as AICAR (AMPK activator), promoted AMPK phosphorylation, whereas Compound C (AMPK inhibitor) inhibited this effect. The sperm-zona pellucida binding experiment showed that 40 µmol/L RA increased the number of sperm attached to the zona pellucida (p < .05). These findings suggest meaningful methods for improved preservation of boar sperm in vitro and provide new insights into the mechanism by which RA protects sperm cells from oxidative damage via AMPK activation.

Melatonin Protects Goat Spermatogonial Stem Cells against Oxidative Damage during Cryopreservation by Improving Antioxidant Capacity and Inhibiting Mitochondrial Apoptosis Pathway.

Spermatogonial stem cells (SSCs) are the only adult stem cells that pass genes to the next generation and can be used in assisted reproductive technology and stem cell therapy. SSC cryopreservation is an important method for the preservation of immature male fertility. However, freezing increases the production of intracellular reactive oxygen species (ROS) and causes oxidative damage to SSCs. The aim of this study was to investigate the effect of melatonin on goat SSCs during cryopreservation and to explore its protective mechanism. We obtained SSCs from dairy goat testes by two-step enzymatic digestion and differential plating. The SSCs were cryopreserved with freezing media containing different melatonin concentrations. The results showed that 10-6 M of melatonin increased significantly the viability, total antioxidant capacity (T-AOC), and mitochondrial membrane potential of frozen-thawed SSCs, while it reduced significantly the ROS level and malondialdehyde (MDA) content (P < 0.05). Further analysis was performed by western blotting, flow cytometry, and transmission electron microscopy (TEM). Melatonin improved significantly the enzyme activity and protein expression of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) (P < 0.05), thereby activating the antioxidant defense system of SSCs. Furthermore, melatonin inhibited significantly the expression of proapoptotic protein (Bax) and increased the expression of antiapoptotic proteins (Bcl-2 and Bcl-XL) (P < 0.05). The mitochondrial apoptosis pathway analysis showed that the addition of melatonin reduced significantly the mitochondrial swelling and vacuolation, and inhibited the release of cytochrome C from mitochondria into the cytoplasm, thereby preventing the activation of caspase-3 (P < 0.05) and inhibiting SSC apoptosis. In addition, melatonin reduced significantly the autophagosome formation and regulated the expression of autophagy-related proteins (LC3-I, LC3-II, P62, Beclin1, and ATG7) (P < 0.05), thereby reversing the freeze-induced excessive autophagy. In summary, melatonin protected goat SSCs during cryopreservation via antioxidant, antiapoptotic, and autophagic regulation.

Location of the supraorbital and infraorbital foramen with references to the soft tissue landmarks in a Chinese population.

The purpose of the current study was to determine the supraorbital foramen (SOF) and infraorbital foramen (IOF) based on soft tissue landmarks, to facilitate prediction of the location of this structure during facial surgery. Forty-two hemispheres of 21 adult cadavers (16 men and 5 women; aged 30-75 years) were dissected to expose the SOF and IOF. The locations of the SOF and IOF were evaluated with direct and photographic measurements. The data gained were analyzed by statistical method. The SOF localized 23.11 ± 2.35 mm superior and 9.48 ± 3.06 mm lateral to the angulus oculi medialis (AOM). The vertical angle from AOM to SOF was 68.3 (SD, 6.44) degrees. The SOF localized 24.81 (SD, 3.39) mm inferior and 10.89 (SD, 2.78) mm lateral to the AOM on the front view. The vertical angle from AOM to IOF was 66.5 (SD, 5.18) degrees. The SOF localized 11.22 (SD, 2.01) mm inferior and 6.09 (SD, 2.32) mm lateral to the ala of the nose (AL) on the front view. The vertical angle from AL to IOF was 61.7 (SD, 7.61) degrees. These results were a little different from the results of some other populations. We found the IOFs located on the point of one-fifth proportion distant to the ALs along the vertical direction distance from AL to SOF, whereas the AOMs located on the point of three-fifths proportion distant from the AL. Our results may provide more detailed information to predict the location of the SOFs and IOFs and help to prevent nerve or vessel damage.

Location of mental foramen based on soft- and hard-tissue landmarks in a chinese population.

The purpose of the present study was to determine the location of the mental foramen (MF) based on soft- and hard-tissue landmarks, to facilitate prediction of the location of this structure during facial and dental surgery. Forty-two hemispheres of 21 adult cadavers (16 men and 5 women; aged 30-75 years) were dissected to expose the MF. The locations of the MFs were evaluated with direct and photographic measurements. Most of the MFs presented a single foramen (95%), except for only 2 cases with double foramina (5%). The MFs localized 23.38 +/- 2.00 mm inferior and 3.55 +/- 1.70 mm medial to the cheilion in the front view while 23.59 +/- 2.11 mm inferior and 7.19 +/- 3.03 mm posterior to the cheilion in the lateral view. Based on the hard-tissue landmarks, we found that most of the MFs localized inferior the second premolar in most of the cases (73.8%), and the MFs localized 23.34 +/- 2.39 mm below the cusp tip of the second premolar, 16.56 +/- 2.53 mm below the inferior alveoli, and 15.56 +/- 1.74 mm superior the bottom of the mandible. The position of the MF varied from 8.7 degrees medial to 15.5 degrees posterior in the vertical angle with the change of surgical body position from supine to lay-side position. Our results may provide a more detailed information to predict the location of the MFs.