UCL Discovery
UCL home » Library Services » Electronic resources » UCL Discovery

Some studies on the effect of α-chymotrypsin on mast cells

Khiav, Behzad Emadi; (1993) Some studies on the effect of α-chymotrypsin on mast cells. Doctoral thesis (Ph.D), UCL (University College London). Green open access

[thumbnail of Some_studies_on_the_effect_of_.pdf]

Download (9MB) | Preview


It has been suggested that activation of one or more proteolytic enzymes might constitute the earliest biochemical change in the sequence of events that ultimately leads to mediator secretion from the mast cell. This effect may be mimicked by the addition of exogenous serine esterases and, with this in mind, we first characterized the effect of α-chymotrypsin (α-CT) on isolated mast cells from different sources. α-CT (10-500 μg/ml) induced a dose-dependent secretion of histamine (≤ 80%) from purified and non-purified populations of rat peritoneal mast cells. The release was non-cytotoxic and was inhibited by metabolic blockers and extremes of temperature. The process was relatively slow, being essentially complete within 20 min, and was unaffected by phosphatidylserine. A substantial component of the secretion persisted in the absence of extracellular calcium. The release was suppressed by extremes of pH and a variety of antiallergic compound and serine esterase inhibitors. α-CT (10-300 μg/m l), in addition to the secretion of preformed mediators, also induced the metabolism of arachidonic acid, resulting in the release of prostaglandin D2 (PGD2) in a dose-related manner from purified rat peritoneal mast cells. α-CT exhibited a marked tissue and species selectivity in its action. The protease was a much weaker releaser of histamine from tissue mast cells of the rat. It was effective against human cells from lung, intestine and skin only at cytotoxic concentrations and ineffective against mouse peritoneal mast cells. The effect of inhibitors of, and substrates for, α-CT in normal and permeabilized rat mast cells were investigated. Rat peritoneal mast cells were recovered by direct lavage and purified by density gradient centrifugation over Percoll. Where appropriate, agents were introduced into the cells after reversible permeabilization with ATP. The seryl enzyme inhibitor phenyl methyl sulphonyl fluoride (PMSF), the suicide inactivator isatoic anhydride, and a number of chymotryptic substrates all effectively inhibited histamine release from rat mast cells stimulated with anti-IgE but not with compound 48/80. Their potency was strikingly increased in permeabilized cells, indicating their effective incorporation into the cytosol. Activation of a chymotrypic enzyme as evidenced by hydrolysis of the fluorescent substrate, was directly demonstrated following immunologic stimulation of permeabilized mast cells containing N-Succinyl-Leu-Leu-Val-Tyr-7-amino-4-methylcoumarin. No such activation was observed with compound 48/80. Immunological stimulation also led to a significant increase in total chymotryptic activity recoverable from rat mast cells. In total, this study provides the most direct evidence to date for the involvement of a serine esterase in immunologic but not polyamine-induced mast cell activation.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Some studies on the effect of α-chymotrypsin on mast cells
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
URI: https://discovery.ucl.ac.uk/id/eprint/10098045
Downloads since deposit
Download activity - last month
Download activity - last 12 months
Downloads by country - last 12 months

Archive Staff Only

View Item View Item