Leid R W, Coley S C, Blanchard D P, Perryman L E
Vet Immunol Immunopathol. 1985 May;9(1):71-85. doi: 10.1016/0165-2427(85)90131-x.
The equine alternative complement pathway has been partially characterized and compared to the equine classical activation pathway. A dose-dependent lysis of RbRBC was observed with peak lytic values noted within 10 minutes at 37 degrees C when rabbit red blood cells (RbRBC) were used as an alternative pathway activator. Sheep red blood cells (SRBC) sensitized with rabbit hemolysin or partially purified equine IgM antibodies were equally sensitive to lysis. Dilution of the commercial hemolysin by 1/5 reduced lysis from 90% to 38% in the presence of constant cell numbers. Hemolysis of SRBC peaked at 10 minutes and the majority of lysis occurred within 10 minutes. Dilution of equine sera by as little as 1/5 decreased hemolytic activity for SRBC to 21.5% from greater than 90% with undiluted sera. The alternative pathway protein, equine factor B, was tested using RbRBC and monitored by its differential susceptibility to heat treatment at 50 degrees C. This treatment led to almost complete inactivation after a 15-minute incubation. An apparent heat-dependent decay of certain classical pathway components was also observed after 50 degrees C treatment. This sensitivity was indicated by a reduction in the lytic activity for sensitized SRBC. Treatment for 15 minutes at 56 degrees C with either RbRBC or SRBC was sufficient to abolish hemolytic activity in all equine sera tested. Chelation of cations with 0.04 M EDTA blocked expression of alternative and classical pathway activation; however, chelation of Ca++ ions with 10 mM EGTA containing 1 mM Mg++ ions permitted lysis of the RbRBC but not the SRBC. A dose-related Mg++-ion dependence for RbRBC hemolytic activity was observed as the concentration of Mg++ was increased to 1.0 mM. In addition, our results obtained with pre-colostral foal serum strongly suggest that natural antibody to RbRBC was of little importance in the lysis observed with these cells. These results also show that the equine alternative pathway activation may require Ca++ ions. If Ca++ ions are required, the equine alternative pathway is quite different from any other mammalian complement system so far described. Our results suggest that the alternative pathway of activation is of major importance in the equine complement system. Confirmation of this hypothesis requires both purification of the components involved as well as further characterization.
马的替代补体途径已得到部分表征，并与马的经典激活途径进行了比较。当使用兔红细胞（RbRBC）作为替代途径激活剂时，在37℃下观察到RbRBC的剂量依赖性溶解，在10分钟内达到峰值溶解值。用兔溶血素或部分纯化的马IgM抗体致敏的绵羊红细胞（SRBC）对溶解同样敏感。在细胞数量恒定的情况下，将商业溶血素稀释1/5可使溶解率从90%降至38%。SRBC的溶血在10分钟时达到峰值，大部分溶解发生在10分钟内。将马血清稀释至仅1/5，SRBC的溶血活性就从未稀释血清时的大于90%降至21.5%。使用RbRBC对替代途径蛋白马因子B进行了测试，并通过其在50℃下对热处理的不同敏感性进行监测。这种处理在孵育15分钟后导致几乎完全失活。在50℃处理后，还观察到某些经典途径成分明显的热依赖性衰变。这种敏感性表现为致敏SRBC的溶解活性降低。用RbRBC或SRBC在56℃处理15分钟足以消除所有测试马血清中的溶血活性。用0.04 M EDTA螯合阳离子可阻断替代途径和经典途径激活的表达；然而，用含有1 mM Mg++离子的10 mM EGTA螯合Ca++离子可使RbRBC溶解，但不能使SRBC溶解。随着Mg++浓度增加到1.0 mM，观察到RbRBC溶血活性与剂量相关的Mg++离子依赖性。此外，我们用初乳前马驹血清获得的结果强烈表明，针对RbRBC的天然抗体在这些细胞的溶解中作用不大。这些结果还表明，马的替代途径激活可能需要Ca++离子。如果需要Ca++离子，马的替代途径与迄今为止描述的任何其他哺乳动物补体系统有很大不同。我们的结果表明，替代激活途径在马补体系统中至关重要。证实这一假设需要纯化相关成分并进一步表征。
