Date Published: July 2, 2008
Publisher: BioMed Central
Author(s): Karin Östensson, Shichun Lun.
The first hours after antigen stimulation, interactions occur influencing the outcome of the immunological reaction. Immunoglobulins originate in blood and/or are locally synthesized. The transfer of Ig isotypes (Igs) in the udder has been studied previously but without the possibility to distinguish between the endothelium and the epithelium. The purpose of this study was to map the Ig transfer through each barrier, separately, and Ig transfer in the local lymph nodes of the bovine udder during the initial innate immune response.
The content of IgG1, IgG2, IgM, IgA and albumin (BSA) was examined in peripheral/afferent mammary lymph and lymph leaving the supramammary lymph nodes, and in blood and milk before (0 h) and during 4 hours after intramammary challenge with Esherichia coli endotoxin in 5 cows.
Igs increased most rapidly in afferent lymph resulting in higher concentrations than in efferent lymph at postinfusion hour (PIH) 2, contrary to before challenge. Ig concentrations in milk were lower than in lymph; except for IgA at 0 h; and they increased more slowly. Afferent lymph:serum and efferent lymph:serum concentration ratios (CR) of Igs were similar to those of BSA but slightly lower. Milk:afferent lymph (M:A) CRs of each Ig, except for IgG2, showed strikingly different pattern than those of BSA. The M:A CR of IgG1, IgM and IgA were higher than that of BSA before challenge and the CR of IgA and IgG1 remained higher also thereafter. At PIH 2 there was a drop in Ig CRs, except for IgG2, in contrast to the BSA CR which gradually increased. The M:A CR of IgM and Ig A decreased from 0 h to PIH 4, in spite of increasing permeability.
The transfer of Igs through the endothelium appeared to be merely a result of diffusion although their large molecular size may hamper the diffusion. The transfer through the epithelium and the Ig concentrations in milk seemed more influenced by selective mechanisms and local sources, respectively. Our observations indicate a selective mechanism in the transfer of IgG1 through the epithelium also in lactating glands, not previously shown; a local synthesis of IgA and possibly of IgM, released primarily into milk, not into tissue fluid; that IgG2 transfer through both barriers is a result of passive diffusion only and that the content of efferent lymph is strongly influenced by IgG1, IgM and IgA in the mammary tissue, brought to the lymph node by afferent lymph.
Bovine mastitis has been extensively studied but mainly as reflected in milk and circulating blood. Investigations of the reaction as it appears in the tissue, usually performed in tissue specimens, have added important information and improved the understanding of immunological reactions of the mammary gland. For studies of tissue reactions over time when repeated sampling is desirable, it appears more suitable to examine interstitial fluid that can be sampled frequently after it has entered the collecting vessels of the peripheral (afferent) lymphatic system in the tissue, through application of a semi-permanent catheter. This method was used in the present investigation parallel to examination of efferent lymph, leaving the local supramammary lymph nodes and analysis of milk. It enabled us to follow the inflammatory response along the entire pathway from the mammary milk compartment, through the interstitial space in the tissue, the afferent lymphatics and the local lymph node; a route where the immune events are initiated and of significant immunological interest. It further made it possible to separately study the transfer of various components through, on one hand the mammary endothelium and on the other hand the mammary epithelium.
The most rapid increase of Igs was observed in afferent lymph, resulting in significantly higher concentration of each Ig isotype, except for IgG2, in afferent than in efferent lymph at PIH 2, contrary to before challenge. Ig concentrations in milk were in general lower than in lymph and they increased later. The transfer of Igs through the endothelium appeared to be merely a result of diffusion while the transfer through the epithelium and the Ig concentrations in milk seemed to be more influenced by selective mechanisms and local sources, respectively. In addition, the molecular size of the Igs appeared to negatively affect their transfer through the endothelium, particularly under normal permeability conditions when the CR of each Ig isotype, except for IgG2, was lower than that of BSA. However, at the mammary epithelium the opposite was observed; the CR of each Ig isotype, except for IgG2, was higher than that of BSA, before challenge. Additionally, the alterations in the epithelial CR of the Igs (IgG1, IgM and IgA) during inflammation were strikingly different from those of BSA. Our observations indicate a selective mechanism being present in the transfer of IgG1 through the epithelium, also in lactating glands which has not been previously shown. The results also indicate a local synthesis in the tissue of IgA and possibly also of IgM, released primarily into milk, not into tissue fluid suggesting that the synthesis occurs close to the epithelium. The IgG2 transfer through endothelium as well as epithelium appeared to be a result of passive diffusion only. In the lymph node, the content of efferent lymph was strongly influenced by IgG1, IgM and IgA brought to the node by the afferent lymph, from the mammary tissue and less dependent on transfer from blood.
The authors declare that they have no competing interests.
KÖ designed and planned the study and methods used, performed the surgery, oversaw and participated in the practical work and prepared the major part of the final manuscript, SL performed the sample collection and laboratory analyses, prepared a first draft of the manuscript and participated in preparing the final manuscript. Both authors read and approved the final manuscript.