Immunocytochemical detection of interleukin 1 within stimulated human monocytes.

Abstract

The term IL-1 has been used to describe a family of monocyte derived polypeptides that have immunomodulatory and proinflammatory biological properties (1, 2). While most studies have focused on IL-1 activity that is found in monocyte/macrophage culture supernatants, early studies showed that high amounts of activity could also be recovered from cell lysates. These studies suggested that IL-1 may accumulate within mononuclear phagocytes in a precursor form before release into the surrounding medium (3-5).Recently it has been shown that there are at least two distinct human IL-1 molecules termed IL-la and IL-13. Separate mRNAs for these IL-1 species each code for a precursor of ~31 kD, which is subsequently processed by still undefined mechanisms to a "mature" molecule of ~17.5 kD (6). The absence of a typical hydrophobic leader sequence in either of the IL-1 molecules identified to date suggests that they may not be typical secretory proteins. A 33-kD precursor for murine IL-1, which is ~62% homologous to human IL-la (6), has been documented at the protein level in cell lysates of peritoneal macrophages, as well as in lysates of the murine macrophage line P388D~ (7). Comparable studies have not yet been reported for human monocytes and macrophages.The successful development of highly specific heterologous antisera (8) for the species of human IL-1 having a pI of 6.8 (IL-18) (6, 9) has allowed us to identify its precursor in the cytoplasm of activated human monocytes. In the present report, the intracellular accumulation of IL-1 is documented using indirect immunofluorescence performed on fixed and permeabilized cells, and immunoblot analysis of cell lysates. These studies show that it is possible to identify IL-1-producing cells at the light microscopic level and suggest that anti-IL-1 antibodies may be useful for the in situ localization of IL-1 production in inflammed tissue. Materials and MethodsAntisera. A detailed description of the peptides used, as well as the methodology for raising and characterizing antisera, is presented elsewhere (8). Briefly, peptides Ala ~7-Asp l 2s (p 117 -128) and of the amino acid sequence derived from I L-1 cDNA (6, 10) were synthesized using standard solid phase methodology (11), coupled to KLH with the heterobifunctional reagent m-maleimidobenzyl-N-hydroxysuccinimide

Effect of local hyperthermia on lymph immune cells and lymphokines of normal human skin

“…Also, no IL-1 was found on the. membrane or in the cytoplasm of Langerhans cells, although membrane-associated IL-l -like activity was detected by others on rat dendritic cells [19] and human monocytes [20]. This rules out the lymph cells at the concentrations present in lymph as contributors of ETAF/IL-1 activity.…”

Cytokine regulation of human monocyte interleukin-1 (IL-1) productionin vitro. Enhancement of IL-1 production by interferon (IFN) gamma, tumour necrosis factor-alpha, IL-2 and IL-1, and inhibition by IFN-alpha

“…High concentrations of cytokines were required to demonstrate direct IL-1 induction, usually by unfractionated mononuclear cells. In addition, cells were cultured on plastic in the presence of serum, conditions which are now thought to alter monocyte responses (Bayne et al, 1986; Van der Meer et al, 1988).…”

C‐reactive protein and platelet activating factor complexes induce production and release of interleukin‐1 by human monocytes

“…The method described is essentially that of Bayne et al (33) with modifications. The assay for intracellular IL-1 was carried out as follows: 0.3 ml of PBM cells with the concentration of 3 X 106/ml in DMEM with 10% FCS was pipetted into each sterile tissue culture chamber on a 1 X 3-in glass slide (Miles Scientific, Naperville, IL).…”