Bhavanandan V P, Puch S, Guo X, Jiang W
Department of Biochemistry & Molecular Biology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA.
Adv Exp Med Biol. 2001;491:95-108. doi: 10.1007/978-1-4615-1267-7_7.
Defects in the glycocalyx of the bladder epithelium may be related to the development of bladder diseases including interstitial cystitis which is a chronic bladder disease of unknown etiology. Indirect evidence has implicated alterations in the bladder epithelial glycoconjugates in interstitial cystitis and vesicaler instillation of glycosaminoglycans is promoted as treatments. However, information on the nature of the glycoconjugates of the bladder epithelium and lectins that may interact with the exogenous instilled glycoconjugates is very limited. We have examined the endogenous lectin associated with bladder epithelium by immunohistochemistry using biotinylated neoglycoconjugates. The strong calcium-independent binding of beta-D-galactose probe suggested the presence of galectins in rabbit and human bladder. Extracts of rabbit bladder organ cultures metabolically labeled with [14C]-amino acids were subjected to affinity chromatography on immobilized lactose and the specifically bound material eluted with 0.2 M lactose. SDS-PAGE of the recovered proteins revealed a major band of approximately 30 kDa and a minor band of 21 kDa. Polymerase chain reaction and northern blot analysis showed that both galectin-3 and galectin-4 are expressed in rabbit bladder. Since galectin-3 from rabbit had been previously cloned, we cloned and sequenced galectin-4 from rabbit bladder. The deduced full length sequence of 328 amino acids revealed four distinct regions: a N-terminal peptide of 19 residues, two carbohydrate recognition domains of 130 residues each, and a linker region of 49 residues. Comparison of the rabbit galectin-4 sequence with those of human, pig, rat, and mouse revealed two invariant peptide motifs that are proposed as signature sequences for identifying related galectins.
膀胱上皮糖萼的缺陷可能与膀胱疾病的发生发展有关，包括间质性膀胱炎，这是一种病因不明的慢性膀胱疾病。间接证据表明间质性膀胱炎中膀胱上皮糖缀合物发生了改变，因此促进膀胱内灌注糖胺聚糖作为治疗方法。然而，关于膀胱上皮糖缀合物的性质以及可能与外源性灌注糖缀合物相互作用的凝集素的信息非常有限。我们通过使用生物素化新糖缀合物的免疫组织化学方法检测了与膀胱上皮相关的内源性凝集素。β-D-半乳糖探针的强非钙依赖性结合表明兔和人膀胱中存在半乳糖凝集素。用[14C]-氨基酸进行代谢标记的兔膀胱器官培养物提取物在固定化乳糖上进行亲和层析，并用0.2M乳糖洗脱特异性结合的物质。回收蛋白质的SDS-PAGE显示一条约30kDa的主要条带和一条21kDa的次要条带。聚合酶链反应和Northern印迹分析表明半乳糖凝集素-3和半乳糖凝集素-4在兔膀胱中均有表达。由于兔半乳糖凝集素-3先前已被克隆，我们克隆并测序了兔膀胱中的半乳糖凝集素-4。推导的328个氨基酸的全长序列显示有四个不同区域：一个19个残基的N端肽段、两个各130个残基的碳水化合物识别结构域以及一个49个残基的连接区。将兔半乳糖凝集素-4序列与人类、猪、大鼠和小鼠的序列进行比较，发现了两个不变的肽基序，被提议作为鉴定相关半乳糖凝集素的特征序列。
