Introduction: Mastitis is a mammary gland inflammation with a bacterial aetiology. Invading pathogens activate the immune response engaging leukocytes and epithelial cells of the udder. To better understand the innate immune response in goat mammary gland, a model of S. aureus intramammary infection (IMI) has been established (1). We investigated the expression of TIR8 in blood and milk cells and in mammary tissue. TIR8 is an important molecule of the innate immunity involved in the early response to invading pathogens. TIR8 is a regulatory receptor, able to inhibit ILRs and TLRs signaling, preventing damages caused by inflammation (2). Aim: The aim of this study was to elucidate the role of TIR8 in the immune response of mammary gland. Given that no data are available on TIR8 in goat, we first studied its pattern of expression in a panel of goat healthy organs and tissues. Then we investigated the role of TIR8 during IMI, comparing its expression in healthy and infected blood, milk and tissues. Materials and Methods: Six goats were infused with PBS in the right udder and with S. aureus in the left udder. Mammary biopsies from both udders were collected 30 hours post infection, formalin fixed and routinely processed for microscopic evaluation or immediately stored in RNAlater (1). As controls, tissue samples from various non-lesional organs were collected at the slaughterhouse from healthy goats and immediately stored in RNAlater. Blood and milk were collected from healthy and infected goats; blood and milk cells were isolated and processed for RNA extraction or for cytospin; milk fat globules were obtained through milk centrifugation and immediately processed for RNA extraction. Total RNA from different organs, blood and milk cells, milk fat globules and mammary infected tissues was extracted and used as template in qPCR for TIR8. TIR8 protein expression pattern was investigated by immunohistochemistry on formalin fixed, paraffin embedded tissue samples from normal and infected mammary glands and on cytospin of isolated goat blood and milk cells. Results: TIR8 messenger was ubiquitously expressed with very high levels in pancreas, mammary gland, spleen and lymph nodes. High levels were also present in kidney, salivary gland, thymus, gastro intestinal tract, liver and lung. Given the high expression in the mammary gland, we investigated which cell population expressed TIR8. TIR8 was mainly expressed by ductal epithelial cells. During S. aureus infection the receptor was down-modulated in ductal cells, but up-regulated in antigen presenting cells (APCs). PMNs didn't show any modulation. Moreover, the secretum increased its TIR8 positivity during infection. We demonstrated that after infection TIR8 was down-modulated in lymphocytes and PMNs, but up-regulated in monocytes and in mammary secretum. Conclusions: TIR8 is ubiquitously expressed in goat tissues, with higher levels in organs with a predominant epithelial component and in lymphoid tissues. This pattern of expression is very similar to other mammals (3). TIR8 down-modulation after S. aureus infection by ductal cells could lead to an exaggerated inflammatory response, whereas the TIR8 up-regulation by APCs could impair pathogen clearance by infiltrating immune cells. Further experiments are needed to elucidate the role of TIR8 in the pathogenesis of S. aureus infection. References: 1) Brenaut et al. Vet Res (2014) 2) Riva et al. Frot Immunol (2012) 3) Riva et al. Vet Immnol Immunopath (2009)

S. Aures modulates tir8 expression during intramammary infection in goat / J. Filipe, G. Curone, P. Moroni, L. Turin, M.F. Addis, N. Rota, M. Caniatti, P. Roccabianca, V. Bronzo, F. Riva - In: Atti del LXIX convegno SISVET[s.l] : Società Italiana delle Scienze Veterinarie, 2015 Jun. - ISBN 978-88-909002-0-7. - pp. 414 (( Intervento presentato al 69. convegno Convegno Nazionale della Società Italiana Scienze Veterinarie tenutosi a Perugia nel 2015.

S. Aures modulates tir8 expression during intramammary infection in goat

J. Filipe
Primo
;
G. Curone
Secondo
;
P. Moroni;L. Turin;M.F. Addis;N. Rota;M. Caniatti;P. Roccabianca;V. Bronzo
Penultimo
;
F. Riva
Ultimo
2015

Abstract

Introduction: Mastitis is a mammary gland inflammation with a bacterial aetiology. Invading pathogens activate the immune response engaging leukocytes and epithelial cells of the udder. To better understand the innate immune response in goat mammary gland, a model of S. aureus intramammary infection (IMI) has been established (1). We investigated the expression of TIR8 in blood and milk cells and in mammary tissue. TIR8 is an important molecule of the innate immunity involved in the early response to invading pathogens. TIR8 is a regulatory receptor, able to inhibit ILRs and TLRs signaling, preventing damages caused by inflammation (2). Aim: The aim of this study was to elucidate the role of TIR8 in the immune response of mammary gland. Given that no data are available on TIR8 in goat, we first studied its pattern of expression in a panel of goat healthy organs and tissues. Then we investigated the role of TIR8 during IMI, comparing its expression in healthy and infected blood, milk and tissues. Materials and Methods: Six goats were infused with PBS in the right udder and with S. aureus in the left udder. Mammary biopsies from both udders were collected 30 hours post infection, formalin fixed and routinely processed for microscopic evaluation or immediately stored in RNAlater (1). As controls, tissue samples from various non-lesional organs were collected at the slaughterhouse from healthy goats and immediately stored in RNAlater. Blood and milk were collected from healthy and infected goats; blood and milk cells were isolated and processed for RNA extraction or for cytospin; milk fat globules were obtained through milk centrifugation and immediately processed for RNA extraction. Total RNA from different organs, blood and milk cells, milk fat globules and mammary infected tissues was extracted and used as template in qPCR for TIR8. TIR8 protein expression pattern was investigated by immunohistochemistry on formalin fixed, paraffin embedded tissue samples from normal and infected mammary glands and on cytospin of isolated goat blood and milk cells. Results: TIR8 messenger was ubiquitously expressed with very high levels in pancreas, mammary gland, spleen and lymph nodes. High levels were also present in kidney, salivary gland, thymus, gastro intestinal tract, liver and lung. Given the high expression in the mammary gland, we investigated which cell population expressed TIR8. TIR8 was mainly expressed by ductal epithelial cells. During S. aureus infection the receptor was down-modulated in ductal cells, but up-regulated in antigen presenting cells (APCs). PMNs didn't show any modulation. Moreover, the secretum increased its TIR8 positivity during infection. We demonstrated that after infection TIR8 was down-modulated in lymphocytes and PMNs, but up-regulated in monocytes and in mammary secretum. Conclusions: TIR8 is ubiquitously expressed in goat tissues, with higher levels in organs with a predominant epithelial component and in lymphoid tissues. This pattern of expression is very similar to other mammals (3). TIR8 down-modulation after S. aureus infection by ductal cells could lead to an exaggerated inflammatory response, whereas the TIR8 up-regulation by APCs could impair pathogen clearance by infiltrating immune cells. Further experiments are needed to elucidate the role of TIR8 in the pathogenesis of S. aureus infection. References: 1) Brenaut et al. Vet Res (2014) 2) Riva et al. Frot Immunol (2012) 3) Riva et al. Vet Immnol Immunopath (2009)
Settore VET/05 - Malattie Infettive degli Animali Domestici
giu-2015
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/349636
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