Whittle C, Gillespie K, Harrison R, Mathieson P W, Harper S J
Academic Renal Unit, University of Bristol, Southmead Hospital, Westbury-on-Trym, Bristol BS10 5NB, U.K.
Clin Sci (Lond). 1999 Sep;97(3):303-12.
Vascular endothelial growth factor (VEGF) mediates increased vascular permeability and endothelial mitogenesis, and may orchestrate normal glomerular permselectivity and proteinuria. Distinct isoforms result from differential gene splicing. VEGF binds to two cell surface tyrosine-kinase receptors, KDR (kinase domain region) and Flt-1 (fms-like tyrosine kinase-1). The latter also exists in a soluble form (sFlt), which is inhibitory. We have studied patterns of VEGF-isoform and VEGF-receptor expression in isolated single normal human glomeruli. mRNA from 190 glomeruli (from 20 individuals) was harvested on to magnetic beads, and nested reverse transcription-PCR was performed using primers for the VEGF isoforms and VEGF receptors. Simultaneous nested reverse transcription-PCR for CD45 was conducted in order to exclude leucocyte contamination. Unexpected products were isolated, cloned and sequenced. Multiple patterns of glomerular VEGF mRNA isoform expression were identified. Most frequently (58%), all three common forms were expressed. VEGF(189) (i.e. 189-amino-acid form of VEGF) was expressed in 63%, VEGF(165) in 85% and VEGF(121) in 84% of glomeruli. Two unexpected PCR products were also identified: 18% of glomeruli expressed VEGF(145), and 27% of glomeruli expressed a new truncated VEGF splice variant, VEGF(148), lacking exon 6, the terminal part of exon 7 and exon 8. Multiple patterns of VEGF-receptor expression were also identified, the most common being expression of all three isoforms (28%). Overall, KDR was seen in 59% of glomeruli, Flt-1 in 45% and sFlt in 57%. Thus the expression of VEGF within normal glomeruli is complex and variable, with inter- and intra-individual variation. Furthermore, sFlt appears to be the co-dominant form of VEGF receptor expressed within glomeruli, suggesting that, in healthy individuals, a degree of VEGF autoregulation is the norm. The physiological importance of VEGF(148) remains to be confirmed.
血管内皮生长因子（VEGF）介导血管通透性增加和内皮细胞有丝分裂，可能调控正常肾小球的电荷选择性和蛋白尿。不同的异构体由基因的差异剪接产生。VEGF与两种细胞表面酪氨酸激酶受体结合，即KDR（激酶结构域区域）和Flt-1（fms样酪氨酸激酶-1）。后者也以可溶性形式（sFlt）存在，具有抑制作用。我们研究了分离的单个正常人肾小球中VEGF异构体和VEGF受体的表达模式。从20名个体的190个肾小球中提取mRNA并吸附到磁珠上，使用针对VEGF异构体和VEGF受体的引物进行巢式逆转录PCR。同时进行CD45的巢式逆转录PCR以排除白细胞污染。对意外产物进行分离、克隆和测序。鉴定出多种肾小球VEGF mRNA异构体表达模式。最常见的情况（58%）是三种常见形式均有表达。VEGF(189)（即189个氨基酸形式的VEGF）在63%的肾小球中表达，VEGF(165)在85%的肾小球中表达，VEGF(121)在84%的肾小球中表达。还鉴定出两种意外的PCR产物：18%的肾小球表达VEGF(145)，27%的肾小球表达一种新的截短的VEGF剪接变体VEGF(148)，其缺少外显子6、外显子7的末端部分和外显子8。还鉴定出多种VEGF受体表达模式,最常见的是三种异构体均表达（28%）。总体而言，59%的肾小球中有KDR，45%中有Flt-1，57%中有sFlt。因此，正常肾小球内VEGF的表达复杂且多变，存在个体间和个体内差异。此外，sFlt似乎是肾小球内表达的VEGF受体的共同主导形式，这表明在健康个体中，一定程度的VEGF自调节是常态。VEGF(148)的生理重要性仍有待证实。
