The role of monocytes in the effects of β-endorphin and selective agonists of μ-and δ-opiate receptors on the proliferative activity of peripheral blood lymphocytes

The effects of β-endorphin under the conditions of naloxone hydrochloride blockade of opiate receptors, as well as the effects of the selective agonists of μ-and δ-receptors DAGO and DADLE and the effects of melanocyte-potentiating factor (MPF), on the in vitro proliferative response of lymphocytes were studied. The dose-effect dependence indicated stimulating effects of β-endorphin, DAGO, and DADLE on the proliferative response in the presence of phytohemagglutinin (PHA). The tetrapeptide MPF, which is the C-terminal sequence of β-endorphin, had almost no effect on the proliferative activity of lymphocytes. β-Endorphin, naloxone, and the μ-and δ-receptor selective agonists enhanced the proliferative response of lymphocytes in an unfractionated cell culture, whereas β-endorphin, naloxone, and DAGO suppressed the proliferative activity of lymphocytes in the mononuclear fraction purified of monocytes. In both cases, the naloxone blockade of opiate receptors enhanced rather than eliminated the β-endorphin effect.     

Characterization of immunomodulatory properties and accessory cell function of small intestinal epithelial cells.

We have studied the immunomodulatory properties of epithelial cells from the small intestine on T cell immune function in vitro. Proliferation of lymph node cells stimulated either with antigen or with mitogen was inhibited by epithelial cells in a dose-dependent fashion. The epithelial cell-mediated suppression of lymphocyte proliferation was blocked by indomethacin, a cyclooxygenase pathway inhibitor, demonstrating that the suppressive effect of epithelial cells was related to prostaglandin secretion. Furthermore, the action of epithelial cell-secreted prostaglandin on lymphocytes was related to its effect on IL-2 as the suppressive effect of epithelial cells was abrogated by the addition of exogenous IL-2. As previously reported, epithelial cells constitutively express MHC class II and we found them able to present antigen in a class II-restricted fashion when their suppressive effects were blocked by indomethacin. Furthermore, epithelial cells activated by LPS secrete an IL-1 like molecule in a fashion analogous to other antigen-presenting cells. These results demonstrate that epithelial cells can both enhance and suppress in vitro T cell immune responses and further characterize the mechanisms by which intestinal epithelial cells may function in gut-associated immune responses.     

A new restriction fragment length polymorphism in the haptoglobin gene region

Summary Direct gene analysis of the haptoglobin gene region was carried out by Southern blotting using an Hp cDNA as probe. Two types of polymorphism were observed: one due to intragenic duplication, is characterized by a constant fragment length difference of 1700bp observed with several enzymes and by complete correspondence with the protein molecular weight polymorphism; the second type, due to point mutation, was represented by two additional restriction sites for Eco RI and Pst I, with a frequency comparable to that of other genes. These two mutations segregated together in families, suggesting that the recently described Hp related gene is closely linked to the Hp gene. Moreover, they were completely associated with each other. The evolutionary significance of this finding is discussed.     

Synthesis of N-carboxyalkyl and N-aminoalkyl functionalized dipeptide building units for the assembly of backbone cyclic peptides.

To improve the assembly of backbone cyclic peptides, N-functionalized dipeptide building units were synthesized. The corresponding N-aminoalkyl or N-carboxyalkyl amino acids were formed by alkylation or reductive alkylation of amino acid benzyl or tert-butyl esters. In the case of N-aminoalkyl amino acid derivatives the aldehydes for reductive alkylation were obtained from N,O-dimethyl hydroxamates of N-protected amino acids by reduction with LiAlH4. N-carboxymethyl amino acids were synthesized by alkylation using bromoacetic acid ester and the N-carboxyethyl amino acids via reductive alkylation using aldehydes derived from formyl Meldrums acid. Removal of the carboxy protecting group leads to free N-alkyl amino acids of very low solubility in organic solvents, allowing efficient purification by extraction of the crude product. These N-alkyl amino acids were converted to their tetramethylsilane-esters by silylation with N,O-bis-(trimethylsilyl)acetamide and could thus be used for the coupling with Fmoc-protected amino acid chlorides or fluorides. To avoid racemization the tert-butyl esters of N-alkyl amino acids were coupled with the Fmoc-amino acid halides in the presence of the weak base collidine. Both the N-aminoalkyl and N-carboxyalkyl functionalized dipeptide building units could be obtained in good yield and purity. For peptide assembly on the solid support, the allyl type protection of the branching moiety turned out to be most suitable. The Fmoc-protected N-functionalized dipeptide units can be used like any amino acid derivative under the standard conditions for Fmoc-solid phase synthesis.