POTENTIATION OF THE ANAPHYLATOXINS INVIVO USING AN INHIBITOR OF SERUM CARBOXYPEPTIDASE-N (SCPN) .1. LETHALITY AND PATHOLOGIC EFFECTS ON PULMONARY TISSUE

Abstract

Carboxypeptidase N (EC 3.4.12.7) (SCPN) is a plasma enzyme that efficiently inactivates the anaphylatoxins C3a [complement component 3a] and C4a and significantly reduces C5a spasmogenic activity by removing the C-terminal arginyl residue from each of these factors. The arginine analog DL-2-mercaptomethyl-3-guanidinoethylthiopropanoic acid (SCPN-INH) is a potent competitive inhibitor of SCPN with a Ki for this carboxypeptidase in serum of 2 .times. 10-9 M. The SCPN inhibitor was used to potentiate biologic activity of the anaphylatoxins in vivo. Infusion via the carotid artery of about 40 mg of SCPN-INH into each of 8 adult guinea pigs inactivated the SCPN for at least 3 h and caused no measurable toxic effects. When cobra venom factor (CVF) is infused into guinea pigs, it activates the alternative pathway of complement, thereby generating the anaphylatoxins C3a and C5a. Ordinarily, infusion of CVF is nonlethal, because the generated anaphylatoxins are rapidly converted to C3a des Arg and C5a des Arg by SCPN. CVF (200 .mu.g) plus SCPN-INH delivered i.v. in 5 animals induced a lethal reaction in less than 5 min. The lethal effect is due largely to the anaphylatoxins. Histologic sections of the lungs from treated animals show dramatic structural changes consistent with peripheral small airway constriction, bronchial constriction, and vasoconstriction of small muscular arteries. Cell aggregates are present in blood vessels. Other histologic changes include severe congestion, pulmonary edema, and an interstitial infiltrate of mononuclear cells. Large doses of chlorpheniramine prevent this lethal reaction. Lethality is apparently attributable to asphyxia and is dependent on the level of CVF administered: e.g., 100 .mu.g CVF was not lethal in 4 animals given SCPN inhibitor, although signs of respiratory distress were observed. On histologic examination of lungs from guinea pigs given CVF and SCPN-INH, the features are similar to those described when anaphylatoxins are instilled into guinea pig lungs. Application of purified C3a plus SCPN-INH i.v. also proved lethal in 3 of the 6 animals challenged. This is the 1st evidence that the C3a anaphylatoxin can elicit a lethal response.