Isolation of Lymphocyte Activating Factors from Human Milk
The protective effect of breast-feeding against infections has mainly been attributed to the presence in human milk of factors such as secretory immunoglobulins, lactoferrin and lysozyme. However, milk is also a rich source of regulatory molecules with putative immunomodulating functions in the mammary gland, and in the recipient infant. Such factors include many hormones, peptide growth factors and interferon, but others still remain to be characterized. The present study explores human milk in regard to the presence of lymphocyte activating factors. Ammonium sulphate precipitated protein from cell-free and defatted mature human milk was tested for interleukin-1 (IL-1), interleukin-2 (IL-2), thymocyte growth peptide (TGP) as well as T and B lymphocyte stimulating activity in various lymphocyte culture assays. The following results were obtained. Employing the murine thymocyte proliferation assay, human milk protein displayed an IL-1-like activity, which eluted in three distinct peaks after Sephadex G 150 gel chromatography, with apparent Mr’s of 14K, 30K and 60K, respectively. Chromatofocusing revealed charge heterogeneity, with an isoelectric point (pI) of 5 of the major peak. No pI 7 activity was detected, indicating that milk-derived IL-1 is similar to macrophage-derived IL-1α, and that no IL-1β-like material is present. Studies in progress on the cellular origin of milk IL-1 indicate that it is produced in high amounts by milk macrophages. Also T and B lymphocyte stimulating factors were detected. Thus, milk protein was comitogenic in combination with dextran sulphate for rat splenic B lymphocytes and in combination with phytohemagglutinin for guinea pig lymph node T cells. Gel filtration showed four peaks of B cell stimulating activity with apparent Mr’s of less than 10K, 14K, 30K, and 70K, whereas the T cell stimulating activity was even more heterogeneous. IL-1 could be responsible for some, but not all, of this activity. Human milk protein displayed no IL-2 or TGP activity. Pasteurization at 72°C for 15 sec. abolished all activity, except for a slight B cell stimulating effect.