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  • Author or Editor: Cleverson D. Souza x
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Abstract

Objective—To evaluate the role of the mitogen-activated protein kinase extracellular signal-regulated kinase (MAPKERK) pathway in the interaction between Mycobacterium avium subsp paratuberculosis (MAP) organisms and bovine monocytes.

Sample Population—Monocytes obtained from healthy adult Holstein dairy cows that were not infected with MAP organisms.

Procedures—Monocytes and MAP organisms were incubated together with or without a specific inhibitor of the MAPKERK pathway (PD98059), and the capacity of monocytes to express tumor necrosis factor alpha (TNF)-α and interleukin (IL)-10 and -12, produce nitric oxide, acidify phagosomes, kill MAP organisms, and undergo apoptosis was evaluated.

Results—The MAPKERK pathway was activated within 10 minutes after addition of MAP organisms to monocytes. Addition of PD98059 to monocyte-MAP mixtures decreased monocyte TNF-α and IL-12 mRNA expression but had no effect on IL-10 mRNA expression. Treatment with PD98059 failed to induce significant alterations in phagosome acidification, organism killing, nitric oxide production, or apoptosis of MAP-exposed monocytes.

Conclusions and Clinical Relevance—Results indicated that the MAPKERK pathway was activated during the interaction of MAP organisms with monocytes, which initiated TNF-α and IL-12 mRNA expression but failed to initiate antimicrobial activity. The MAPKERK pathway may be involved in initiating proinflammatory and proimmune responses in MAP infection in cattle.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To determine cell membrane receptors involved in phagocytosis of Mycobacterium avium subsp paratuberculosis (MAP) organisms.

Sample Population—Monocytes were obtained from healthy adult Holstein dairy cows that were test negative for MAP infection on the basis of bacteriologic culture of feces and serologic test results.

Procedures—Monocytes or bovine macrophage cell line (BoMac) cells were incubated with MAP organisms for 30, 60, or 120 minutes with or without inhibitors of integrins, CD14, or mannose receptors. Phagocytosis was evaluated by light microscopy or by flow cytometry. CD11a/CD18, CD11b, and CD14 expression on monocytes and BoMac cells was evaluated by use of flow cytometry.

Results—Monocytes and BoMac cells rapidly phagocytized MAP organisms. However, compared with BoMac cells, monocytes had a greater total capacity to phagocytize MAP organisms. Addition of neutralizing anti-integrin antibodies (anti-CD11a/CD18 and anti-CD11b) substantially inhibited phagocytosis by monocytes during the first 60 minutes of incubation with MAP organisms, but were less effective at 120 minutes of incubation. Anti-CD11a/CD18 and anti-CD11b antibodies were less effective in inhibiting phagocytosis by BoMac cells. Addition of inhibitors of CD14 or mannose receptors also inhibited phagocytosis of MAP by monocytes. Addition of a combination of integrin and mannose inhibitors had an additive effect in reducing phagocytosis, but addition of integrin and CD14 inhibitors did not have an additive effect.

Conclusions and Clinical Relevance—Multiple receptors are involved in phagocytosis of MAP organisms. Although CD11/CD18 receptors appear to be the major receptors used by MAP at early time points, mannose receptors and CD14 also contribute substantially to phagocytosis.

Full access
in American Journal of Veterinary Research