Pereira Oliveira Paulo Ricardo, Rodrigues-Junior Valnes, Rech Virginia Cielo, Duval Wannmacher Clovis Milton
Departamento de Bioquímica, Instituto de Ciências Básicas de Saúde, UFRGS, Porto Alegre, RS, Brazil.
Arch Med Res. 2007 Feb;38(2):164-9. doi: 10.1016/j.arcmed.2006.09.013. Epub 2006 Dec 11.
Cystinosis is an autosomal recessive disorder associated with lysosomal cystine accumulation caused by defective cystine efflux. Visual deficit is a possible consequence of cystine accumulation in cornea and retina. Fibroblasts from cystinotic patients present ATP deficit with intact mitochondrial energy-generating capacity by an unknown mechanism. Considering that creatine kinase is a thiol enzyme crucial for energy homeostasis in retina, and disulfides like cystine may alter thiol enzymes, the main objective of the present study was to investigate the effect of cystine and cysteamine, the drug used for treatment of cystinotic patients, on creatine kinase activity in cytosolic and mitochondrial fractions of the retina from adult pigs.
Retina was isolated from 6-month-old Landrace pigs, homogenized and mitochondrial and cytosolic fractions separated by centrifugation. Cytosolic and mitochondrial creatine kinase activities were determined in the presence of different concentrations of cystine and/or cysteamine.
Cystine inhibited the enzyme activity in a dose- and time-dependent manner and cysteamine prevented and reversed the inhibition caused by cystine, suggesting that cystine inhibits creatine kinase activity by oxidation of the sulfhydryl groups of the enzyme.
Considering that creatine kinase is a crucial enzyme for retina energy homeostasis, in case cystine leaves lysosome these results provide a possible mechanism for cystine toxicity and also another beneficial effect for the use of cysteamine in patients with cystinosis.
胱氨酸病是一种常染色体隐性疾病，与由于胱氨酸外流缺陷导致的溶酶体胱氨酸蓄积相关。视力缺陷是角膜和视网膜中胱氨酸蓄积的一个可能后果。胱氨酸病患者的成纤维细胞存在ATP缺陷，但其线粒体能量生成能力保持完好，机制不明。鉴于肌酸激酶是视网膜能量稳态中一种关键的硫醇酶，且像胱氨酸这样的二硫化物可能会改变硫醇酶，本研究的主要目的是探究胱氨酸及用于治疗胱氨酸病患者的药物半胱胺对成年猪视网膜胞质和线粒体部分中肌酸激酶活性的影响。
从6月龄长白猪分离视网膜，匀浆后通过离心分离线粒体和胞质部分。在不同浓度的胱氨酸和/或半胱胺存在的情况下测定胞质和线粒体肌酸激酶活性。
胱氨酸以剂量和时间依赖的方式抑制该酶活性，半胱胺可预防并逆转由胱氨酸引起的抑制作用，这表明胱氨酸通过氧化该酶的巯基来抑制肌酸激酶活性。
鉴于肌酸激酶是视网膜能量稳态的关键酶，如果胱氨酸离开溶酶体，这些结果为胱氨酸毒性提供了一种可能的机制，也为半胱胺用于胱氨酸病患者提供了另一个有益作用。
