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Extended longevity geometrically-inverted proximal tubule organoids

ABSTRACT: This study reports the cellular self-organization of primary human renal proximal tubule epithelial cells (RPTECs) around a minimal Matrigel scaffold to produce basal-in and apical-out proximal tubule organoids (tubuloids). These tubuloids are produced and maintained in hanging drop cultures for 90+ days, the longest such culture of any kind reported to date. The tubuloids upregulate maturity markers, such as aquaporin-1 (AQP1) and megalin (LRP2), and exhibit less mesenchymal and proliferation markers, such as vimentin and Ki67, compared to 2D cultures. They also experience changes over time as revealed by a comparison of gene expression patterns of cells in 2D culture and in day 31 and day 67 tubuloids. Gene expression analysis and immunohistochemistry reveal an increase in the expression of megalin, an endocytic receptor that can directly bind and uptake protein or potentially assist protein uptake. The tubuloids, including day 90 tubuloids, uptake fluorescent albumin and reveal punctate fluorescent patterns, suggesting functional endocytic uptake through these receptors. Furthermore, the tubuloids release kidney injury molecule-1 (KIM-1), a common biomarker for kidney injury, when exposed to albumin in both dose- and time-dependent manners. While this study focuses on potential applications for modeling proteinuric kidney disease, the tubuloids may have broad utility for studies where apical proximal tubule cell access is required.

ORGANISM(S): Homo sapiens

PROVIDER: GSE198857 | GEO | 2022/10/07

REPOSITORIES: GEO

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Project description:Chronic kidney disease (CKD) affects more than 20 million Americans and approximately 10% of the population worldwide, and thus represents a major public health concern. Genome-wide association studies (GWAS) of kidney functional decline have identified genes associated with CKD, but the precise roles and mechanisms by which they influence kidney function remained largely unexplored. Here, we examine the role of one GWAS identified gene by creating mice deficient for Galnt11, which encodes a member of the enzyme family that initiates protein Oglycosylation, an essential post-translational modification known to influence protein function and stability. We find that Galnt11-deficient mice display low molecular weight proteinuria and have specific defects in proximal tubule-mediated resorption of vitamin D binding protein, a1-microglobulin and retinol binding protein. Moreover, we identify the endocytic receptor megalin (LRP2) as a direct target of Galnt11 in vivo. Megalin in Galnt11-deficient mice displays reduced ligand binding and undergoes age-related loss within the kidney. Differential mass spectrometry revealed specific sites of Galnt11-mediated glycosylation within mouse kidney megalin/LRP2 that are known to be involved in ligand binding, suggesting that Oglycosylation directly influences the ability to bind ligands. In support of this, recombinant megalin containing these sites displayed reduced albumin binding in cells deficient for Galnt11. Our results provide insight into the association between GALNT11 and CKD, and identify a role for Galnt11 in proper kidney function.

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TMAO Suppresses Megalin Expression and Albumin Uptake in Human Proximal Tubular Cells Via PI3K and ERK Signaling

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The soluble intracellular domain of megalin does not affect renal proximal tubular function in vivo

2011-01-01 | GSE19778 | GEO

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2021-06-16 | PXD020046 | Pride

Impact of Diabetic Stress Conditions on Renal Cell Metabolome

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