Increased mTOR Signaling Secondary to a Human Integrin-Linked Kinase (ILK) Missense Variant Inhibits Nephrogenesis with Decreased ATP Generation
Bibliographic record
Abstract
Background: Signaling pathways that control nephrogenesis are critical to our understanding of kidney health and disease. Previously, we demonstrated increased mTOR signaling in vitro by overexpression of a CAKUT-associated human missense variant (T173I) of Integrin-Linked Kinase (ILK), a key regulator of renal development. Here, we investigate the hypothesis that ILK-T173I disrupts renal development in vivo via dysregulation of mTOR signaling. Methods: mTOR signaling was analyzed in Ilk-T173I knock-in mice with Ilk-T173I replacing the WT Ilk allele. Morphogenic effects of Ilk-T173I on renal development were quantitated. Ureteric and non-ureteric cell populations were isolated by FAC sorting. Gene expression was analyzed by whole genome RNA microarray and qPCR. Cell proliferation and nephron maturation were evaluated by immunostaining and specific markers. Energy metabolism was characterized using the Seahorse assay. Results: Homozygous Ilk-T173I knock-in mice were characterized by a 13.6% decrease in nephron number (n=6, P=0.04), a 35.2% decrease in ureteric branching (n=5, P=0.006) and an 88% increase in mTOR signaling (n=3, P=0.01). Rapamycin treatment of Ilk-T173I-knock-in embryonic kidney explants rescued ureteric branching to normal levels. Genome-wide RNA expression analysis in Ilk-T173I-knock-in ureteric and nonureteric cell populations demonstrated elevated mTOR signaling in non-ureteric cells only. Ilk-T173I-knock-in non-ureteric populations expressed a 431% increase in Osr1 (n=3, P=0.002), a mesenchymal cell progenitor marker, and a 54% decrease in Lhx1 (n=3, P=0.04), a marker for maturing nephrons. Ilk-T173I knock-in embryonic kidneys demonstrated a 2075% increase in phospho-Histone H3-labeled cells (n=3, P=0.0005), an indicator of cell proliferation, in the ureteric, nephrogenic and stromal lineages, and a 16.1% decrease in the percentage of S-shaped bodies (n=4, P=0.01), the mature form of nephrogenic intermediates. Metabolic profile of Ilk-T173I-knock-in, non-ureteric cells exhibited a 17% decrease in both oxidative ATP production (n=3, P=0.04) and total ATP production (n=3, P=0.02). Conclusions: These data show that human Ilk-T173I variant impairs renal development in a mTOR-dependent manner, specifically acting within the non-ureteric cell population by increasing proliferation and disrupting maturation and cell metabolism. Funding: Government Support - Non-U.S.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.002 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".