Todorov P T, McDevitt T M, Cariuk P, Coles B, Deacon M, Tisdale M J
Cancer Research Campaign Nutritional Biochemistry Research Group, Pharmaceutical Sciences Institute, Aston University, Birmingham, United Kingdom.
Cancer Res. 1996 Mar 15;56(6):1256-61.
Splenocytes from mice bearing a cachexia-inducing tumor (MAC16) have been fused with mouse myeloma cells to produce hybridomas, which have been cloned to produce antibody reactive to a material which copurified with a lipid-mobilizing factor isolated from the same tumor. The monoclonal antibody has been used to investigate factors potentially involved in the development of cachexia. The major protein detectable by immunoprecipitation of a partially purified lipid-mobilizing factor was M(r) 69,000, whereas Western blotting showed two bands of M(r) 69,000 and M(r) 24,000. Although the monoclonal antibody did not neutralize lipid-mobilizing activity in an in vitro assay, it did neutralize a serum factor capable of protein degradation in isolated gastrocnemius muscle. Affinity purification of MAC16 tumor homogenates using the monoclonal antibody yielded two immunoreactive bands of M(r) 69,000 and M(r) 24,000, which were further fractionated on a hydrophobic column (C8). This material was capable of inducing tyrosine release from isolated gastrocnemius muscle, and the effect could be blocked with the monoclonal antibody. The two immunoreactive bands from the hydrophobic column were capable of inducing weight loss in mice, whereas nonimmunoreactive fractions had no effect on body weight. The M(r) 24,000 species had a unique amino acid sequence, whereas the M(r) 69,000 species gave the same sequence as the M(r) 24,000 material, together with that for albumin. The M(r) 24,000 species contained carbohydrate, and lectin blotting showed a strong reaction with wheat germ and Erythrina crystagalli agglutinins. This suggests that the material is a glycoprotein or proteoglycan that shows strong binding affinity for albumin, possibly through the carbohydrate residues.
将携带可诱发恶病质肿瘤（MAC16）的小鼠的脾细胞与小鼠骨髓瘤细胞融合，以产生杂交瘤，这些杂交瘤已被克隆，以产生与一种与从同一肿瘤中分离出的脂质动员因子共纯化的物质发生反应的抗体。该单克隆抗体已被用于研究可能参与恶病质发展的因素。通过免疫沉淀部分纯化的脂质动员因子可检测到的主要蛋白质的分子量为69,000，而蛋白质印迹显示分子量为69,000和24,000的两条带。尽管该单克隆抗体在体外试验中不能中和脂质动员活性，但它确实能中和一种能够在离体腓肠肌中引起蛋白质降解的血清因子。使用该单克隆抗体对MAC16肿瘤匀浆进行亲和纯化，得到分子量为69,000和24,000的两条免疫反应带，将其在疏水柱（C8）上进一步分离。这种物质能够诱导离体腓肠肌释放酪氨酸，并且该作用可被单克隆抗体阻断。来自疏水柱的两条免疫反应带能够诱导小鼠体重减轻，而非免疫反应部分对体重没有影响。分子量为24,000的物质具有独特的氨基酸序列，而分子量为69,000的物质给出的序列与分子量为24,000的物质相同，以及与白蛋白的序列相同。分子量为24,000的物质含有碳水化合物，凝集素印迹显示与麦胚凝集素和刺桐凝集素发生强烈反应。这表明该物质是一种糖蛋白或蛋白聚糖，可能通过碳水化合物残基对白蛋白表现出强烈的结合亲和力。
