Abstract
Interactions between inflammatory cytokines and plasminogen (Pg) activation system on immune cells are yet to be established. In previous studies we reported a somatic cell-associated elevation of proteolytic activity in mammary secretion of drying-off goats and cows. The purposes of the present study were to examine the role of TNF-alpha in polymorphonuclear neutrophil (PMN)-associated Pg activation, and the significance of this activation pathway for overall plasmin (Pm) activity in mammary secretion of drying-off cows. Results of experiments in vitro showed that the spontaneous Pg activation observed on fresh preparations of bovine blood PMN was completely blocked by anti bovine TNF-alpha antibody, and was further up-regulated by exogenous bovine TNF-alpha. Monitoring the parameters of mammary secretion of drying-off cows revealed that both somatic cell counts and differential PMN ratio was significantly elevated at weeks 1, 2 and 3 of milk stasis. Nevertheless, specific activity of soluble Pm in mammary secretion increased and the level of 17-kDa TNF-alpha decreased immediately following milk stasis. Iimmunoblotting revealed that although both 26-kDa pro-TNF-alpha and 17-kDa TNF-alpha were consistently present in somatic cells of mammary secretion collected at weeks 0, 1, 2 and 3 of milk stasis, only 26-kDa pro-TNF-alpha was present in somatic cells of milk during lactation. In-vitro assay indicated that cell-free mammary secretion of drying-off cows exerted no Pg activation bioactivity towards bovine blood PMN. Altogether, the current study suggests the existence of an active TNF-alpha-Pg-Pm autocrine/paracrine loop on the massively infiltrated PMN inside udders of drying-off cows, which involves extensive binding and internalization of 17-kDa TNF-alpha on PMN and consequently activation of Pg, resulting in high Pm activity and low 17-kDa TNF-alpha level in mammary secretion. These coordinated mechanisms may play a role in the defence of drying-off mammary gland.
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