The isolation from spleen tissue of high molecular weight proteins which alter vascular permeability: Investigation of their chemical and pharmacological properties

Abstract

Lymphocytes have been implicated in the inflammatory response associated with delayed hypersensitivity reactions. It was considered that these cells might release substances which cause this alteration in vascular permeability. Maillard et al. (Int. Arch. Allergy 42, P. 50, 1972) have shown that different classes of mediators are released one which increases vascular permeability in the first 10 minutes, the second in 20 minutes to 4 hours after intradermal injection. It is the first group of mediators we wish to discuss. Material from lymph node cells (LNPF) which increases vascular permeability within 10 minutes after its intradermal injection was first described by Willoughby et al. [8]. They showed this material could be obtained from other tissues such as liver and spleen. It was an extract of spleen tissue studied here as large quantities of starting material were desired for best results in purification and isolation of activity. The protein extract from spleen tissue causes a marked increase in vascular permeability in various species at the site of the intradermal injection, the dye which leaks into the tissue at the injection site being quantitatively assessed by extraction and colorimetric assay. This material was shown to be a mixture of proteins between 50,000 and 100,000 in molecular weight and was termed spleen permeability factor (SPF). A number of purification methods have been used to isolate the active factor(s) -ion exchange chromatography, molecular filtration through gels and membranes, preparative isoeleetric focusing and precipitation methods. As the fractionation of SPF by the above methods yields protein fractions of different molecular size and/or charge type, many of these fractions possessing considerable permeability activity, permeability activity may be shared by a variety of different proteins and could depend upon a particular structural feature shared by these molecules, rather than complex features of tertiary structure. SPF does not appear to depend on that part of the secondary and tertiary protein structure which is affected by urea but modification of the e-NH~ group with three different reagents practically abolished activity. Conversely, block of aspartic and glutamic carboxylate (and C-terminal) groups increases the activity of SPF by factors ranging from 5 to 24 X. Samples of SPF contain no appreciable amounts of low molecular wight permeability factors and produce no contraction or relaxation of isolated smooth muscle. The material contains no esterolytic or kallikrein-like activity and its permeability effect is not abolished by DPF treatment. The permeability effect of SPF is exerted indirectly through the release of histamine and 5-HT as indicated by the following findings.(a) The blueing effect of SPF is almost completely abolished by the pretreatment of the animal with a combination of mepyramine and methysergide; (b) local depletion of rat skin of histamine and 5-HT reduces the permeability response to SPF by more than 80 %; (c) there is a dose-dependen...

Anaphylactoid-Inflammation-Promoting Factor

“…While LNPF [23] and spleen permeability factor [11] cannot accurately be lo calized by any kind of chromatographic methods [5,18], AIPF consis tently runs with BSA during molecular sieving on Sephadex G-100. Moreover, AIPF alone has no effect on vascular permeability when in jected intradermally to rats.…”

“…The factor responsible for this effect has been termed lymph node permeability factor (LNPF). Other reports have de scribed permeability factors present in the extract of spleen cells [11]. It was of interest, therefore, to examine the relationship between AIPF and various permeability factors obtained from lymph node cells.…”