D'Andrea G, Di Giulio A, Salucci M L, Oratore A
Dipartimento di Scienze e Tecnologie Biomediche e di Biometria, Università dell'Aquila, Italy.
Cell Signal. 1995 Jan;7(1):67-74. doi: 10.1016/0898-6568(94)00063-h.
Binding and iron delivering of ovotransferrin (OTf) were evaluated using 14-day old chick-embryo red blood cells (CERBC) and cholesterol-depleted by treatment with chicken egg phosphatidyl choline (E-PC) liposomes. Liposome-treated CERBC assayed for their cholesterol content showed a cholesterol depletion depending on the incubation time, being 25% (w/w) of the maximum cellular removal of cholesterol seen after 22 h incubation at 37 degrees C. Total phosphorus content did not change either for the various samples or during the different incubation times, indicating that specific cholesterol removal occurred, as confirmed also by the increased membrane fluidity revealed through fluorescence anisotropy measurements. The apparent dissociation constant (Kd) of control and treated CERBC was almost of the same value at the same incubation time, ranging from 0.30 microM after 0.25 h incubation to 0.19 microM after 14 or 22 h incubation. In all experiments, the maximum value of bound OTf molecules per cell (Bmax) notably decreased as incubation time increased. But, in cholesterol partly depleted CERBC, the decrease of the Bmax values was less pronounced as the incubation time increased. As far as binding experiments were concerned, iron uptake studies showed that uptaking capacities decreased as incubation time increased. Considering both binding and iron uptake, at the same incubation time, liposome-treated CERBC were slightly more efficient with respect to untreated samples. In any case a passive iron delivering could be evidenced after 22 h incubation. It is suggested that cholesterol may tune binding and iron uptake by either regulating or affecting the expression or mobility of the OTf receptor.
使用14日龄鸡胚红细胞（CERBC）以及经鸡蛋卵磷脂（E-PC）脂质体处理而胆固醇耗竭的细胞，评估了卵转铁蛋白（OTf）的结合和铁传递情况。对经脂质体处理的CERBC进行胆固醇含量测定，结果显示胆固醇耗竭程度取决于孵育时间，在37℃孵育22小时后，胆固醇最大细胞去除量的25%（w/w）出现胆固醇耗竭。不同样品或不同孵育时间下，总磷含量均未改变，这表明发生了特异性胆固醇去除，通过荧光各向异性测量显示的膜流动性增加也证实了这一点。在相同孵育时间下，对照和处理后的CERBC的表观解离常数（Kd）几乎相同，孵育0.25小时后为0.30微摩尔，孵育14或22小时后为0.19微摩尔。在所有实验中，随着孵育时间增加，每个细胞结合的OTf分子最大值（Bmax）显著降低。但是，在胆固醇部分耗竭的CERBC中，随着孵育时间增加，Bmax值的降低不太明显。就结合实验而言，铁摄取研究表明，随着孵育时间增加，摄取能力降低。综合考虑结合和铁摄取情况，在相同孵育时间下，经脂质体处理的CERBC相对于未处理样品效率略高。无论如何，孵育22小时后可证明存在被动铁传递。提示胆固醇可能通过调节或影响OTf受体的表达或流动性来调节结合和铁摄取。
