Lipp Sarah N, Jacobson Kathryn R, Hains David S, Schwarderer Andrew L, Calve Sarah
Weldon School of Biomedical Engineering, Purdue University, West Lafayette, Indiana.
Medical Scientist/Engineer Training Program, Indiana University, Indianapolis, Indiana.
J Am Soc Nephrol. 2021 Jul;32(7):1649-1665. doi: 10.1681/ASN.2020081204. Epub 2021 Apr 19.
The extracellular matrix (ECM) is a network of proteins and glycosaminoglycans that provides structural and biochemical cues to cells. In the kidney, the ECM is critical for nephrogenesis; however, the dynamics of ECM composition and how it relates to 3D structure during development is unknown.
Using embryonic day 14.5 (E14.5), E18.5, postnatal day 3 (P3), and adult kidneys, we fractionated proteins based on differential solubilities, performed liquid chromatography-tandem mass spectrometry, and identified changes in ECM protein content (matrisome). Decellularized kidneys were stained for ECM proteins and imaged in 3D using confocal microscopy.
We observed an increase in interstitial ECM that connects the stromal mesenchyme to the basement membrane (TNXB, COL6A1, COL6A2, COL6A3) between the embryo and adult, and a transient elevation of interstitial matrix proteins (COL5A2, COL12A1, COL26A1, ELN, EMID1, FBN1, LTBP4, THSD4) at perinatal time points. Basement membrane proteins critical for metanephric induction (FRAS1, FREM2) were highest in abundance in the embryo, whereas proteins necessary for integrity of the glomerular basement membrane (COL4A3, COL4A4, COL4A5, LAMB2) were more abundant in the adult. 3D visualization revealed a complex interstitial matrix that dramatically changed over development, including the perinatal formation of fibrillar structures that appear to support the medullary rays.
By correlating 3D ECM spatiotemporal organization with global protein abundance, we revealed novel changes in the interstitial matrix during kidney development. This new information regarding the ECM in developing kidneys offers the potential to inform the design of regenerative scaffolds that can guide nephrogenesis .
细胞外基质（ECM）是由蛋白质和糖胺聚糖组成的网络，为细胞提供结构和生化信号。在肾脏中，ECM对肾发生至关重要；然而，在发育过程中ECM组成的动态变化及其与三维结构的关系尚不清楚。
我们使用胚胎第14.5天（E14.5）、E18.5、出生后第3天（P3）和成年肾脏，根据不同溶解度对蛋白质进行分级分离，进行液相色谱-串联质谱分析，并鉴定ECM蛋白质含量（基质体）的变化。对脱细胞肾脏进行ECM蛋白质染色，并使用共聚焦显微镜进行三维成像。
我们观察到，在胚胎期和成年期之间，连接基质间充质与基底膜的间质ECM增加（TNXB、COL6A1、COL6A2、COL6A3），并且在围产期时间点间质基质蛋白（COL5A2、COL12A1、COL26A1、ELN、EMID1、FBN1、LTBP4、THSD4）有短暂升高。对后肾诱导至关重要的基底膜蛋白（FRAS1、FREM2）在胚胎中丰度最高，而肾小球基底膜完整性所需的蛋白（COL4A3、COL4A4、COL4A5、LAMB2）在成年期更为丰富。三维可视化显示了一个复杂的间质基质，其在发育过程中发生了显著变化，包括围产期出现的纤维状结构，这些结构似乎支持髓放线。
通过将三维ECM时空组织与整体蛋白质丰度相关联，我们揭示了肾脏发育过程中间质基质的新变化。这些关于发育中肾脏ECM的新信息为设计能够指导肾发生的再生支架提供了可能性。
