In vitro assessment of the effects of temperature on phagocytosis, reactive oxygen species production and apoptosis in bovine polymorphonuclear cells

Abstract

Heat stress exerts a direct negative effect on farm animal health, triggering physiological responses. Environmental high temperature induces immunosuppression in dairy cows, increasing the risk of mastitis and milk somatic cell counts. The influence of heat stress on leukocytes activities has not been fully elucidated. The present in vitro study was aimed at assessing whether the exposure to temperature simulating conditions of severe whole body hyperthermia affects defensive functions of bovine blood polymorphonuclear cells.Blood was collected from seven clinically healthy, multiparous, late lactating Holstein cows. After isolation, PMN were incubated at either 39 or 41°C. Phagocytosis, respiratory burst and apoptosis were then investigated. The selected temperatures of 39°C or 41°C mimicked conditions of normothermia or severe heat stress, respectively. Phagocytosis assay was carried out by measuring the fluorescence of phagocyted fluorescein-labelled E. coli bioparticles. The modulation of oxidative burst activity was studied by the cytochrome C reduction method. Apoptosis was determined by measuring the activities of two enzymes that play an effector role in the process, namely Caspase-3 and Caspase-7. Statistical analyses were performed using SPSS 22.0. A Student t-test for paired samples and a Generalised Estimating Equation were used based on data distribution.The phagocytosis rate was reduced (−37%, P<0.01) when PMN were incubated for 2h at 41°C, when compared to phagocytosis rate measured at 39°C. The oxidative burst, as determined by extracellular production of reactive oxygen species (ROS), was also reduced by the exposure of cells to 41°C compared to 39°C. Such reduction ranged between −2 and −21% (P<0.05). Apoptosis rate was not affected by different temperatures.The results reported in this study suggest that phagocytosis and ROS production in PMN exposed to severe high temperature are impaired, partially explaining the higher occurrence of infections during periods of hot weather.