Toll-like receptor 4 is needed to restrict the invasion of Escherichia coli P4 into mammary gland epithelial cells in a murine model of acute mastitis

Abstract

SummaryMastitis, an inflammatory response of the mammary tissue to invading pathogenic bacteria, is a common disease in breast-feeding women and dairy animals. Escherichia coli is a leading cause of mastitis in dairy animals. During the course of the disease the host mounts a strong inflammatory response, but specific bacterial virulence factors have not yet been identified. Here we report the use of a murine mastitis model to investigate the innate inflammatory reaction of the mammary gland. We show that lipopolysaccharide (LPS) infusion induces mastitis in wild-type mice (C3H/HeN), but not in mice expressing mutated Toll-like receptor 4 (TLR4) (C3H/HeJ). The wild-type phenotype was restored by adoptive transfer of TLR4-expressing macrophages into the alveolar milk space of C3H/HeJ mice. In contrast to the LPS treatment, infection with E. coli P4 (ECP4) resulted in inflammation even in the absence of LPS/TLR4 signalling, indicating that additional factors play a role in the pathogenesis of the intact bacteria. Furthermore, in the absence of functional TLR4 the infecting ECP4 invade the epithelial cells with high efficiency, forming intracellular microcolonies. However, adoptive transfer with TLR4-expressing macrophages drastically reduced the epithelial invasion. Taken together, these results indicate that ECP4 has an invasive potential, which is restricted by alveolar macrophages in response to the LPS/TLR4 signalling.

Molecular Studies on E.Coli Isolate from Milk of Mastatic Cattle with Special Reference to Associated Biochemical Changes in Kaliobea Governorate

“…The California Mastitis Test (CMT) provided a useful tool for farmers and veterinarians for measuring the level of inflammation in the udder [22]. In current study founded increase in inflamatory parameters IL6,TNF andCRP factors, In our opinion, this elevation may be created as a result of proinflamatory response to infection with E.coli and stmulation of immunity system, these aforesaid results came in agreement with other reports recoreded that LPS triggers formation of proinflammatory and inflammatory cytokines, produced predominantly by monocytes and macrophages [23,24]. Cytokines, such as tumor necrosis factor alpha (TNF-α), initiate the inflammatory response [25] which induces the acute phase response (APR) by activating the production of acute phase proteins (APP) and LPS-binding protein (LBP) [26][27][28][29].…”

IL-17A Is an Important Effector of the Immune Response of the Mammary Gland to Escherichia coli Infection

“…To gain a more direct evidence of the implication of IL-17A in the course of mastitis, we used a mouse model of E. coli mastitis with an E. coli strain that has been used extensively in experimentally induced MG infections in dairy cows (34)(35)(36) and mice (32,37,38). Our findings show that IL-17A is involved in the response of the MG to infection by E. coli, and that an increase in IL-17A production in the MG at the onset of infection dramatically improved the containment of infection.…”

“…Pregnant C57BL/6J mice were purchased from Janvier breeding laboratories (St. Berthevin, France). They were inoculated when they were 7-10 d in lactation (32). Clean food and water were given to mice ad libitum.…”

CD44 is highly expressed on milk neutrophils in bovine mastitis and plays a role in their adhesion to matrix and mammary epithelium

“…We have previously shown the importance of LPS-TLR4 signaling in alveolar macrophages in the pathogenesis of experimental LPS and E. coli murine mastitis [3]. In mice, CD44 expressed on hemopoietic cells is an important regulator of LPS-TLR signaling in macrophages, affecting pulmonary inflammatory response and PMN recruitment [9].…”

“…In the BME adhesion inhibition assay, PMN were enumerated using the myeloperoxidase (MPO) assay [3]. The linearity and validity of the microplate colorimetric tetramethyl benzidine (TMB, Vector Labs., Burlingame, CA, USA) assay for enumeration of bovine peripheral blood PMN were assessed in the range of 0 to 10 6 cells using the checkerboard method (data not shown).…”