背景:体积调节阴离子通道(VRAC)是LRRC8A与LRRC8B的异六聚体,-C,-D,或-E的各种组合。根据亚基组成,这些膨胀激活的通道传导氯化物,氨基酸,有机渗透压,和毒品。尽管VRACs在细胞体积调节中起作用,肾脏的渗透压变化很大,VRACs在肾脏中的定位和功能都未知。
方法:使用表达表位标记的LRRC8亚基的小鼠来确定所有VRAC亚基的肾定位。携带Lrrc8b-e组成型缺失的小鼠,或Lrrc8a的诱导型或细胞特异性消融,进行分析以评估VRACs的肾功能。分析包括组织学,不同利尿状态下的尿液和血清参数,和代谢组学。
结果:肾脏表达所有5个VRAC亚基,并具有明显不同的定位。而LRRC8C只在血管内皮中发现,所有其他亚单位都在肾单位中发现。LRRC8E对嵌入细胞具有特异性,而LRRC8A,LRRC8B,LRRC8D在近端小管的基底外侧膜中突出。LRRC8A在近端而非远端小管中的条件性缺失和LRRC8D的组成性缺失导致近端肾小管损伤,增加利尿,和轻微的范可尼样症状.
结论:VRAC/LRRC8通道对于近端小管的功能和完整性至关重要,但不适用于更远端的肾单位段,尽管它们对容量调节的需求更大。许多有机化合物的基底外侧出口可能需要LRRC8A/D通道,包括细胞代谢物,在近端小管中。近端肾小管损伤可能是由于在不存在VRAC通道的情况下几种转运分子的组合积累所致。
BACKGROUND: Volume-regulated anion channels (VRACs) are heterohexamers of LRRC8A with LRRC8B, -C, -D, or -E in various combinations. Depending on the subunit composition, these swelling-activated channels conduct chloride, amino acids, organic osmolytes, and drugs. Despite VRACs\' role in cell volume regulation, and large osmolarity changes in the kidney, neither the localization nor the function of VRACs in the kidney is known.
METHODS: Mice expressing epitope-tagged LRRC8 subunits were used to determine the renal localization of all VRAC subunits. Mice carrying constitutive deletions of Lrrc8b-e, or with inducible or cell-specific ablation of Lrrc8a, were analyzed to assess renal functions of VRACs. Analysis included histology, urine and serum parameters in different diuresis states, and metabolomics.
RESULTS: The kidney expresses all five VRAC subunits with strikingly distinct localization. Whereas LRRC8C is exclusively found in vascular endothelium, all other subunits are found in the nephron. LRRC8E is specific for intercalated cells, whereas LRRC8A, LRRC8B, and LRRC8D are prominent in basolateral membranes of proximal tubules. Conditional deletion of LRRC8A in proximal but not distal tubules and constitutive deletion of LRRC8D cause proximal tubular injury, increased diuresis, and mild Fanconi-like symptoms.
CONCLUSIONS: VRAC/LRRC8 channels are crucial for the function and integrity of proximal tubules, but not for more distal nephron segments despite their larger need for volume regulation. LRRC8A/D channels may be required for the basolateral exit of many organic compounds, including cellular metabolites, in proximal tubules. Proximal tubular injury likely results from combined accumulation of several transported molecules in the absence of VRAC channels.