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Exposure of sensitized perfused rabbit lungs to human O-N type erythrocytes leads to pulmonary vasoconstriction and bronchoconstriction. To investigate whether platelet-activating factor (PAF) is a mediator of pulmonary vasoconstrictive and bronchoconstrictive responses after antigen challenge, we administered antigenic erythrocytes after the administration of PAF antagonist (.1 mg/kg; CV6209). Pulmonary arterial and airway pressures significantly increased after antigen challenge in the sensitized rabbit lungs, but not in the nonsensitized rabbit lungs. CV6209 significantly inhibited these pulmonary vasoconstrictive and bronchoconstrictive responses after antigen challenge. We concluded that PAF, at least in part, plays an important role in pulmonary vasoconstriction and bronchoconstriction after antigen challenge in rabbits.

To investigate whether thromboxane and/or platelet activating factor (PAF) mediate the pulmonary vasoconstrictive response to antigen in vivo, we intra-arterially injected human erythrocytes as antigen into sensitized rabbits after administration of putative inhibitors: a cyclooxygenase synthetase inhibitor (indomethacin, 5 mg.kg-1), a thromboxane synthetase inhibitor (OKY 046, 10 mg.kg-1 + 100 micrograms.kg-1.min-1), and a PAF blocker (CV6209, .1 mg.kg-1). Pulmonary artery and airway pressures significantly increased after the antigen challenge in sensitized rabbits, but did not in nonsensitized rabbits. Both indomethacin and OKY046 significantly inhibited the increase in pulmonary artery pressure after the antigen challenge, while CV6209 did not. CV6209 significantly attenuated the decrease in femoral artery pressure after the antigen challenge, while neither indomethacin nor OKY046 did. There were no significant differences in the increase in airway pressure among the groups. We conclude that thromboxane rather than PAF mediates the pulmonary vasoconstriction after the antigen challenge and that mediators other than thromboxane and PAF mediate bronchoconstriction after the antigen challenge in sensitized rabbits.

By adding antigen cells into perfusate circulation, a great increase in pulmonary arterial pressure was observed in isolated perfused lung from rabbits previously immunized with human O-N type erythrocytes. To investigate whether thromboxane A2 is the main mediator in pulmonary vasoconstrictive response, we injected antigen erythrocytes into the reservoir after administration of putative inhibition as follows: indomethacin (5 mg/kg, thromboxane A2 synthetase inhibitor), KT2-962 (.1 mg/kg, thromboxane receptor blocker), and pyrilamine (.1 mumol, H1 blocker). Pulmonary vasoconstrictive response after antigen challenge was significantly blocked by both indomethacin and KT2-962, but not by H1 blocker. Although the H1 blocker, pyrilamine, did not significantly block the pulmonary vasoconstrictive response, it did significantly block the bronchoconstrictive response after antigen challenge; however, the bronchoconstrictive response was not blocked by either indomethacin or KT2-962. We conclude that thromboxane is the main mediator in the pulmonary vasoconstrictive response, and histamine is the main mediator in the bronchoconstrictive response.

In the accompanying paper, we described the existence, molecular characterization, and ontogeny of a 30 kDa abnormal protein in chicken dystrophic muscles. In this study, we have purified chicken carbonic anhydrase III and the 30 kDa protein and directly compared them. In terms of its enzymological features, the 30 kDa protein is a typical carbonic anhydrase III. Like carbonic anhydrases, it contains one mole zinc per mole of protein. The protein selectively cross-reacted with a chicken carbonic anhydrase III antibody. Antibody to the 30 kDa protein cross-reacted with chicken skeletal muscle carbonic anhydrase III. Moreover, the distribution of the abnormal protein is exactly identical to that of carbonic anhydrase III; however, there is a possibility that the 30 kDa protein is a variant of carbonic anhydrase III. Slight differences were found in antigenicities and in the apparent molecular weights of the two proteins. We have compared the two proteins by 125I-labeled two-dimensional peptide mapping. Tryptic maps have shown that the two proteins are highly homologous. Combined, these results strongly indicate that the 30 kDa protein and carbonic anhydrase III are similar, if not identical.

An effective isolation of carbonic anhydrase C (CA-C) from red cell lysate was described. The lysate dialyzed against 0.0175 M phosphate buffer pH 6.3 was applied onto a CM-Sephadex column equilibrated with the same buffer. The elution was performed with the starting buffer and 0.1 M dibasic potassium phosphate containing 0.14 M NaCl. The hemoglobin fraction eluted with the second eluant was applied onto a DEAE-cellulose column and eluted with 0.2 M glycine containing 0.01 percent KCN, resulting in complete isolation of CA-C at high recovery rate. For the preparation in a large scale, the hemoglobin fraction prepared from CM-Sephadex semi-batch-type chromatography was treated with cold ethanol and chloroform. The purity of these preparations was confirmed by polyacrylamide gel disc electrophoresis and immunoelectrophoresis.

In the previous paper, the present authors reported on diminution in agglutinability of trypsin-treated M and MN red cells with anti-M IgG antibody. This paper deals with the time course of changes in agglutinability and quantitative differences of M agglutinogen of red cells. M agglutinogen of M red cells disappeared much faster than that of MN, and the lowest titer of anti-M IgG antibody to agglutinate M red cells was higher than that to agglutinate MN red cells. The significant difference of agglutination scores was observed in the frequency distribution in 100 cases of M and MN red cells each. Subgroups such as M1 and M2 in M agglutinogen of trypsin-treated red cells reacting with anti-M IgG antibody were unsettled.

The agglutinability of the proteolytic enzyme treated M, N and MN red cells with any one of the anti-M and -N antisera, eluted antibodies and class antibodies was studied. The enzyme-treated 141 and N red cells lost their group specificity in agglutination with the absorbed antisera. However, it was noted that the enzyme-treated M red cells acquired N agglutinogen instead of losing their own M agglutinogen in agglutination with the eluted antibodies. The enzyme-treated N red cells maintained the group specificity. Agglutination of the enzyme-treated red cells with the class antibodies showed clearly that the changes in agglutinability of the red cells were caused by each of the class antibodies.