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Production and properties of histamine-releasing factor of guinea pigs
R Alam1, D M Lewis, S A Olenchock
1Division of Respiratory Disease Studies, National Institute for Occupational Safety and Health, Morgantown, WV 26505.
Abstract:
The production of histamine-releasing factor (HRF) by human mononuclear cells has previously been reported. In this paper we describe the production of HRF by guinea pig spleen cells, thymocytes, and PBMC. Guinea pig lymphoid cells were cultured either alone or in the presence of mitogens (PHA and Con A) or specific Ag(OVA and keyhole limpet hemocyanin) and the dialyzed cell-free supernatant was tested for histamine-releasing activity on guinea pig lung mast cells and blood basophils. Lung mast cells were isolated by enzymatic digestion and partially purified by countercurrent elutriation and discontinuous Percoll gradient centrifugation. Guinea pig spleen cells, thymocytes, and PBMC spontaneously produced significant amounts of HRF. The production was enhanced upon stimulation with PHA or specific Ag in animals immunized with Ag in CFA. Two distinct species of HRF were identified with m.w. of 50,000 to 70,000 and 5000 to 8000 by gel chromatography. HRF is a trypsin- and chymotrypsin-sensitive heat-stable protein. It does not bind to Con A-Sepharose and its production is not inhibited by tunicamycin. HRF-induced histamine release from lung mast cells is a temperature-dependent process and is complete in 10 min at 37 degrees C. Intradermal injection of HRF caused an immediate ear-swelling reaction in guinea pigs. The most severe ear-swelling reactions did not resolve within 1 h, but instead evolved over a period of 12 to 24 h.
Insights
Guinea pig immune cells produce histamine-releasing factor (HRF), a protein that triggers histamine release from mast cells. This factor, identified in two sizes, causes immediate and delayed swelling reactions upon injection.
Area of Science:
- Immunology
- Cell Biology
Background:
- Histamine-releasing factor (HRF) production by human cells is known.
- This study investigates HRF production in guinea pig immune cells.
Purpose of the Study:
- To characterize HRF production by guinea pig spleen cells, thymocytes, and peripheral blood mononuclear cells (PBMCs).
- To determine the properties and biological activity of guinea pig HRF.
Main Methods:
- Culturing guinea pig lymphoid cells with or without mitogens/antigens.
- Testing cell-free supernatants for histamine-releasing activity on mast cells and basophils.
- Purifying and characterizing HRF using gel chromatography and enzymatic sensitivity assays.
Main Results:
- Guinea pig spleen cells, thymocytes, and PBMCs spontaneously produced HRF.
- HRF production increased with mitogen or antigen stimulation in immunized animals.
- Two HRF species (50-70 kDa and 5-8 kDa) were identified; HRF is heat-stable and sensitive to trypsin/chymotrypsin.
- HRF-induced histamine release is rapid at 37°C.
- Intradermal HRF induced immediate and prolonged ear swelling in guinea pigs.
Conclusions:
- Guinea pigs produce HRF from various lymphoid cells, with production modulated by immune stimulation.
- HRF is a heat-stable protein with distinct molecular weights and potent histamine-releasing capabilities.
- HRF elicits both acute and delayed inflammatory responses in vivo.