ABSTRACT
Excessive accumulation of neutrophils and their uncontrolled death by necrosis at the site of inflammation exacerbates inflammatory responses and leads to self-amplifying tissue injury and loss of organ function, as exemplified in a variety of respiratory diseases. In homeostasis, neutrophils are inactivated by apoptosis, and non phlogistically removed by neighboring macrophages in a process known as efferocytosis, which promotes the resolution of inflammation. The present study assessed the potential anti-inflammatory and pro-resolution benefits of tylvalosin, a recently developed broad-spectrum veterinary macrolide derived from tylosin. Recent findings indicate that tylvalosin may modulate inflammation by suppressing NF-κB activation. Neutrophils and monocyte-derived macrophages were isolated from fresh blood samples obtained from 12- to 22-week-old pigs. Leukocytes exposed to vehicle or to tylvalosin (0.1, 1.0, or 10 µg/mL; 0.096-9.6 µM) were assessed at various time points for apoptosis, necrosis, efferocytosis, and changes in the production of cytokines and lipid mediators. The findings indicate that tylvalosin increases porcine neutrophil and macrophage apoptosis in a concentration- and time-dependent manner, without altering levels of necrosis or reactive oxygen species production. Importantly, tylvalosin increased the release of pro-resolving Lipoxin A4 (LXA4) and Resolvin D1 (RvD 1 ) while inhibiting the production of pro-inflammatory Leukotriene B4 (LTB4) in Ca2+ ionophore-stimulated porcine neutrophils. Tylvalosin increased neutrophil phospholipase C activity, an enzyme involved in releasing arachidonic acid from membrane stores. Tylvalosin also inhibited pro-inflammatory chemokine (C-X-C motif) ligand 8 (CXCL-8, also known as Interleukin-8) and interleukin-1 alpha (IL-1α) protein secretion in bacterial lipopolysaccharide-stimulated macrophages. Together, these data illustrate that tylvalosin has potent immunomodulatory effects in porcine leukocytes in addition to its antimicrobial properties.
ABSTRACT
The accumulation of neutrophils and proinflammatory mediators, such as leukotriene B4 (LTB4), is a classic marker of inflammatory disease. The clearance of apoptotic neutrophils, inhibition of proinflammatory signaling, and production of proresolving lipids (including lipoxins, such as lipoxin A4 [LXA4]) are imperative for resolving inflammation. Tulathromycin (TUL), a macrolide used to treat bovine respiratory disease, confers immunomodulatory benefits via mechanisms that remain unclear. We recently reported the anti-inflammatory properties of TUL in bovine phagocytes in vitro and in Mannheimia haemolytica-challenged calves. The findings demonstrated that this system offers a powerful model for investigating novel mechanisms of pharmacological immunomodulation. In the present study, we examined the effects of TUL in a nonbacterial model of pulmonary inflammation in vivo and characterized its effects on lipid signaling. In bronchoalveolar lavage (BAL) fluid samples from calves challenged with zymosan particles (50 mg), treatment with TUL (2.5 mg/kg of body weight) significantly reduced pulmonary levels of LTB4 and prostaglandin E2 (PGE2). In calcium ionophore (A23187)-stimulated bovine neutrophils, TUL inhibited phospholipase D (PLD), cytosolic phospholipase A2 (PLA2) activity, and the release of LTB4. In contrast, TUL promoted the secretion of LXA4 in resting and A23187-stimulated neutrophils, while levels of its precursor, 15(S)-hydroxyeicosatetraenoic acid [15(S)-HETE], were significantly lower. These findings indicate that TUL directly modulates lipid signaling by inhibiting the production of proinflammatory eicosanoids and promoting the production of proresolving lipoxins.
