Mitochondrial superoxide disrupts the metabolic and epigenetic landscape of CD4+ and CD8+ T-lymphocytes.

While the role of mitochondrial metabolism in controlling T-lymphocyte activation and function is becoming more clear, the specifics of how mitochondrial redox signaling contributes to T-lymphocyte regulation remains elusive. Here, we examined the global effects of elevated mitochondrial superoxide (O2-) on T-lymphocyte activation using a novel model of inducible manganese superoxide dismutase (MnSOD) knock-out. Loss of MnSOD led to specific increases in mitochondrial O2- with no evident changes in hydrogen peroxide (H2O2), peroxynitrite (ONOO-), or copper/zinc superoxide dismutase (CuZnSOD) levels. Unexpectedly, both mitochondrial and glycolytic metabolism showed significant reductions in baseline, maximal capacities, and ATP production with increased mitochondrial O2- levels. MnSOD knock-out T-lymphocytes demonstrated aberrant activation including widespread dysregulation in cytokine production and increased cellular apoptosis. Interestingly, an elevated proliferative signature defined by significant upregulation of cell cycle regulatory genes was also evident in MnSOD knock-out T-lymphocytes, but these cells did not show accelerated proliferative rates. Global disruption in T-lymphocyte DNA methylation and hydroxymethylation was also observed with increased mitochondrial O2-, which was correlated to alterations in intracellular metabolite pools linked to the methionine cycle. Together, these results demonstrate a mitochondrial redox and metabolic couple that when disrupted may alter cellular processes necessary for proper T-lymphocyte activation.

Measurement of bovine body and scrotal temperature using implanted temperature sensitive radio transmitters, data loggers and infrared thermography.

Synchronous and continuous measurement of body (BT) and scrotal temperature (ST) without adverse welfare or behavioural interference is essential for understanding thermoregulation of the bull testis. This study compared three technologies for their efficacy for long-term measurement of the relationship between BT and ST by means of (1) temperature sensitive radio transmitters (RT), (2) data loggers (DL) and (3) infrared imaging (IRI). After an initial pilot study on two bulls to establish a surgical protocol, RTs and DLs were implanted into the flank and mid-scrotum of six Wagyu bulls for between 29 and 49 days. RT frequencies were scanned every 15 min, whilst DLs logged every 30 min. Infrared imaging of the body (flank) and scrotum of each bull was recorded hourly for one 24-h period and compared to RT and DL data. After a series of subsequent heat stress studies, bulls were castrated and testicular tissue samples processed for evidence of histopathology. Radio transmitters were less reliable than DLs; RTs lost >11 % of data, whilst 11 of the 12 DLs had 0 % data loss. IRI was only interpretable in 35.8 % of images recorded. Pearson correlations between DL and RT were strong for both BT (r > 0.94, P < 0.001) and ST (r > 0.80, P < 0.001). Surgery produced temporary minor inflammation and scrotal hematoma in two animals post-surgery. Whilst scar tissue was observed at all surgical sutured sites when bulls were castrated, there was no evidence of testicular adhesion and normal active spermatogenesis was observed in six of the eight implanted testicles. There was no significant correlation of IRI with either DL or RT. We conclude that DLs provided to be a reliable continuous source of data for synchronous measurement of BT and ST.

Thermoregulation of the bovine scrotum 1: measurements of free-range animals in a paddock and pen.

The bull's scrotum and scrotal cord vasculature has traditionally been regarded as a thermoregulatory device for maintaining optimal testicular temperature for normal spermatogenesis. This assumption has mostly been derived from discrete measurements using thermocouples with limited data correlating continuous scrotal temperature (ST) to body temperature (BT). From mid-summer to early autumn, four Wagyu bulls (9-18 months) were surgically implanted with two data loggers (DL) logging at 30 min intervals: one on the right hand side flank and the other was attached to the visceral vaginal tunic of the mid-testis. Bulls were firstly housed in a paddock (PK) for 13 days and then moved to individual pens (IP), again for 13 days. Repeated measures analysis modelled the long-term and diurnal trends in BT and ST. While both day and time of day (TOD) were significant effects for ST at both housing locations (P < 0.005), only TOD showed significance for BT at both locations (P < 0.0001). Significant effects were seen between bulls with ST (F = 167.2, P < 0.001) but not BT (F = 0.03, P = 0.863), suggestive of variation in individual bull thermoregulatory capacity. Dual peaks were observed in ST at 0500 and 2130 h when housed in PK but not IP, suggesting ST may be influenced by external stimuli such as postural or behavioural changes. Reporting concurrent and continuous BT and ST will allow further investigation into factors influencing bovine ST and should be useful in selecting bulls with high degrees of thermoregulation capacity.

Artificial insemination in South American camelids and wild equids.

An overview of the present status of the use of artificial insemination (AI) in South American camelids and wild equids is offered. Technical aspects of semen collection, dilution and cryopreservation have limited the development and use of AI in camelid and equid species. To-date, efficiency is low but progress has been made and viable offspring have been produced through the use of AI in domestic South American camelids using both fresh and frozen semen. The origin, composition, and function of the viscous component of camelid seminal plasma remain a mystery and an obvious area for future research. A better understanding of the normal constituents of seminal plasma will enable the rational design of semen extenders suitable for camelids. Post-thaw sperm viability is very low, and studies are needed to address questions of optimal freezing and thawing procedures as well as the insemination dose. The basis for differences in reported pregnancy rates with sexed and frozen semen in domestic equids, and the ultimate success of AI in wild equids will require continued research into the "stallion effect", extenders and cryoprotectants, optimal volume and number of spermatozoa, temperatures during handling, processing an transport, and insemination techniques. In both camelids and equids, research on domestic species under controlled conditions provides and excellent opportunity to develop effective semen handling techniques for application in wild and endangered species of the respective families.