Loss of the collagen IV modifier prolyl 3-hydroxylase 2 causes thin basement membrane nephropathy
Hande Aypek, … , Tobias B. Huber, Florian Grahammer
Hande Aypek, … , Tobias B. Huber, Florian Grahammer
Published May 2, 2022
Citation Information: J Clin Invest. 2022;132(9):e147253. https://doi.org/10.1172/JCI147253.
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Research Article Nephrology

Loss of the collagen IV modifier prolyl 3-hydroxylase 2 causes thin basement membrane nephropathy

Abstract

The glomerular filtration barrier (GFB) produces primary urine and is composed of a fenestrated endothelium, a glomerular basement membrane (GBM), podocytes, and a slit diaphragm. Impairment of the GFB leads to albuminuria and microhematuria. The GBM is generated via secreted proteins from both endothelial cells and podocytes and is supposed to majorly contribute to filtration selectivity. While genetic mutations or variations of GBM components have been recently proposed to be a common cause of glomerular diseases, pathways modifying and stabilizing the GBM remain incompletely understood. Here, we identified prolyl 3-hydroxylase 2 (P3H2) as a regulator of the GBM in an a cohort of patients with albuminuria. P3H2 hydroxylates the 3′ of prolines in collagen IV subchains in the endoplasmic reticulum. Characterization of a P3h2ΔPod mouse line revealed that the absence of P3H2 protein in podocytes induced a thin basement membrane nephropathy (TBMN) phenotype with a thinner GBM than that in WT mice and the development of microhematuria and microalbuminuria over time. Mechanistically, differential quantitative proteomics of the GBM identified a significant decrease in the abundance of collagen IV subchains and their interaction partners in P3h2ΔPod mice. To our knowledge, P3H2 protein is the first identified GBM modifier, and loss or mutation of P3H2 causes TBMN and focal segmental glomerulosclerosis in mice and humans.

Authors

Hande Aypek, Christoph Krisp, Shun Lu, Shuya Liu, Dominik Kylies, Oliver Kretz, Guochao Wu, Manuela Moritz, Kerstin Amann, Kerstin Benz, Ping Tong, Zheng-mao Hu, Sulaiman M. Alsulaiman, Arif O. Khan, Maik Grohmann, Timo Wagner, Janina Müller-Deile, Hartmut Schlüter, Victor G. Puelles, Carsten Bergmann, Tobias B. Huber, Florian Grahammer

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Figure 3

Ultrastructural analysis and histological phenotypes of P3h2ΔPod and P3h2fl/fl mice.

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Ultrastructural analysis and histological phenotypes of P3h2ΔPod and P3h...
(A) TEM micrographs of P3h2ΔPod and P3h2fl/fl mice at 6 weeks, 28 weeks, and 48 weeks. Red arrows indicate an abnormal GBM structure. Scale bars: 500 nm (top left and middle panels), 2 μm (top right panel), 1 μm (bottom left panel), and 2 μm (bottom middle and right panels). (B) Measurement of GBM thickness on TEM micrographs. At 6 weeks and 28 weeks, the KO mice had a thinner GBM, whereas at 48 weeks, the KO mice had a thicker GBM when compared with that of WT mice. (C) Foot process width measurement of WT and KO podocytes on TEM micrographs. Foot process widths were increased in 28- and 48-week-old KO mice when compared with WT foot process widths, indicating foot process effacement. (D) Bowman’s capsule thickness measurement on TEM micrographs of P3h2ΔPod and P3h2fl/fl mice at 48 weeks. The thickness of Bowman’s capsule was significantly increased in KO mice, indicating PEC activation. (E) PAS staining was performed at 6 weeks, 28 weeks, and 48 weeks. At 6 weeks and 28 weeks, the glomerular morphology was healthy in P3h2ΔPod mice. At 48 weeks, glomerulosclerosis and podocyte injury were observed in P3h2ΔPod kidney tissue (red arrow). Scale bar: 20 μm. (F) Quantification of glomerulosclerosis in P3h2ΔPod and P3h2fl/fl mice at 48 weeks. P3h2ΔPod mice had significantly more glomerulosclerosis than did P3h2fl/fl mice. n ≥4. Graphs show the mean ± SD. *P < 0.05, **P < 0.01, and ***P < 0.001, by unpaired, 2-tailed Student’s t test.