IDENTIFICATION OF GENES REGULATED BY ADVANCED GLYCATION END PRODUCTS IN CULTURED GLOMERULAR MESANGIAL CELLS

非酶糖基化被认为是糖尿病肾病的病因之一。虽然在肾小球系膜细胞中发现了晚期糖基化终产物（AGEs）的受体，但AGEs诱导的细胞功能变化尚未明确。我们进行了mRNA差异显示分析，以确定ages诱导的大鼠系膜细胞基因表达的改变。将融合系膜细胞与AGE-BSA （200mug/ml）或对照BSA （200mug/ml）孵育48小时，提取总RNA。用T12MN引物反转录后，用T12MN引物和随机10粒引物进行聚合酶链反应扩增cDNA，并在序列凝胶上分析产物。在筛选的6000个mrna中，有78个候选PCR产物在序列凝胶上被检测到。northern blot分析证实有6个克隆被AGEs改变。其中3个增加了AGEs， 3个减少了AGEs。经AGEs增加的3个无性系中，2个分别与线粒体延伸因子Tu和间质叉头-1同源。AGEs减少的三个克隆中有一个与肌动蛋白解聚因子具有同源性。据报道，这三个基因参与蛋白质合成或细胞增殖。对这些基因的进一步分析可能为糖尿病肾病的发病机制提供新的认识。综上所述，通过mRNA差异显示分析，确定了系膜细胞中三个受AGEs调控的基因。

Nonenzymatic glycation is proposed to be one of the etiologic factors of diabetic nephropathy. Although receptors for advanced glycation end products (AGEs) were found in glomerular mesangial cells, AGEs-induced changes in cellular functions have not been clarified yet. We performed mRNA differential display analysis, to identify AGEs-induced alterations of gene expression in cultured rat mesangial cells. Confluent mesangial cells were incubated with AGE-BSA (200mug/ml) or control BSA (200mug/ml) for 48 hours and total RNA was extracted. After reverse transcription with T12MN primers, cDNA was amplified by polymerase chain reaction using T12MN primers and random 10 mer, and the products were analyzed on a sequence gel. Of 6,000 mRNAs screened, 78 candidate PCR products were detected on a sequence gel. Northen blot analysis confirmed 6 clones changed by AGEs. Three of them were increased by AGEs and three clones were decreased. Two of three clones increased by AGEs showed homology to mitochondrial elongation factor Tu and mesenchyme fork head-1, respectively. One of three clones decreased by AGEs showed homology to actin depolymerizing factor. These three genes have been reported to be involved in protein synthesis or cell proliferation. Further analysis of these genes may provide new insight on pathogenesis of diabetic nephropathy. In conclusion, three genes regulated by AGEs in mesangial cells were identified by mRNA differential display analysis.