RNA-seq analysis of proximal tubule specific Fan1 knockout transgenic kidneys subjected to cisplatin injury
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE163862 | GEO | 2023/05/03
REPOSITORIES: GEO
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Project description:Acute kidney injury (AKI) remains a major global healthcare problem and there is a need to develop human-based models to study AKI in vitro. Towards this goal, we have characterized induced pluripotent stem cell-derived human kidney organoids and their response to cisplatin, a chemotherapeutic drug that induces AKI and preferentially damages the proximal tubule. We found that a single treatment with 50 µM cisplatin induces HAVCR1 and CXCL8 expression, DNA damage (γH2AX) and cell death in the organoids in a dose-dependent manner but greatly impairs organoid viability. DNA damage was not specific to the proximal tubule but also affected the distal tubule and interstitial cell populations. This lack of specificity correlated with low expression of the proximal tubule-specific SLC22A2/OCT2 transporter for cisplatin. To improve viability, we developed a repeated low-dose regimen of 4x 5 µM cisplatin over 7 days and found this causing less toxicity while still inducing a robust AKI response that included secretion of known AKI biomarkers and inflammatory cytokines. This work validates the use of human kidney organoids to model aspects of AKI, with the potential to identify new AKI biomarkers and develop better therapies.
2020-02-29 | GSE145085 | GEO
Project description:Genome-wide association studies (GWAS) have identified over 800 loci associated with kidney function, yet the specific genes, variants, and pathways involved remain elusive. By integrating kidney function GWAS, human kidney expression and methylation quantitative trait analyses, we identified Ten-Eleven Translocation (TET) DNA demethylase 2: TET2 as a novel kidney disease risk gene. Utilizing single-cell chromatin accessibility and CRISPR-based genome editing, we highlight GWAS variants that influence TET2 expression in kidney proximal tubule cells. Experiments using kidney/tubule-specific Tet2 knockout mice indicated its protective role in cisplatin-induced acute kidney injury, as well as chronic kidney disease and fibrosis, induced by unilateral ureteral obstruction or adenine diet. Single-cell gene profiling of kidneys from Tet2 knockout mice and TET2-knock-down tubule cells revealed the altered expression of DNA damage repair and chromosome segregation genes, notably including INO80, another kidney function GWAS target gene itself. Remarkably both TET2-null and INO80-null cells exhibited an increased accumulation of micronuclei after injury, leading to the activation of cytosolic nucleotide sensor cGAS-STING. Genetic deletion of cGAS or STING in kidney tubules or pharmacological inhibition of STING protected TET2 null mice from disease development. In conclusion, our findings highlight TET2 and INO80 as key genes in the pathogenesis of kidney diseases, indicating the importance of DNA damage repair mechanisms.
2024-09-13 | GSE273027 | GEO
Project description:After acute kidney injury (AKI), patients either recover or alternatively develop fibrosis and chronic kidney disease. Interactions between injured epithelia, stroma and inflammatory cells determine whether kidneys repair or undergo fibrosis, but the molecular events that drive these processes are poorly understood. Here, we use single nucleus RNA sequencing of a mouse model of AKI to characterize cell states during repair from acute injury. We identify a distinct proinflammatory and profibrotic proximal tubule cell state that fails to repair. Deconvolution of bulk RNA-seq datasets indicates that this “failed-repair proximal tubule cell” or FR-PTC, state can be detected in other models of kidney injury, increasing in the aging rat kidney and over time in human kidney allografts. We also describe dynamic intercellular communication networks and discern transcriptional pathways driving successful vs. failed repair. Our study provides a detailed description of cellular responses after injury and suggests that the FR-PTC state may represent a therapeutic target to improve repair.
2020-06-08 | GSE139107 | GEO
Project description:We combined lineage tracing of cycling (Ki67+) cells with single nuclear multiomics (single nucleus RNA-seq + single nucleus ATAC-seq) to characterize the long-term (4 weeks and 6 months) outcome of cells that initiate proliferation early after acute kidney injury (AKI). The data document a broad proliferative response to injury in epithelial and non-epithelial kidney cell types, identify novel transcription factors governing the adaptive and maladaptive proximal tubule cell state and highlight the importance of enhancer dynamics in determining cell states. Comparison of lineage traced with control proximal tubule cells reveals long-term effects of AKI on proximal tubule cells, even following adaptive repair.
2022-11-22 | GSE209610 | GEO
Project description:Proximal tubule has a remarkable capacity for repair after acute kidney injury. Whether dedifferentiation or a fixed progenitor population is responsible for this repair remains the subject of ongoing controversy. We have generated a novel genetic mouse model to address this question. We generated a Kim-1-GFPCreERt2 knockin mouse line (Kim1GCE) by homologous recombination. Kim-1 is expressed exclusively in dedifferentiated proximal tubule cells. We crossed this mouse line to the EGFP-L10a mouse line to perform Translating Ribosome Affinity Purification coupled with next generation sequencing (TRAP-seq) to identify the transcriptional signature of tubular epithelial cells during the course of injury and repair. TRAP-seq was performed in kidney samples at day 2, 7, and 14 after bilateral ischemia reperfusion injury and sham.
2019-11-29 | GSE121191 | GEO
Project description:<p><strong>BACKGROUND:</strong> Ischemia/reperfusion injury (IRI) is the leading cause of acute kidney injury (AKI). The current standard of care focuses on supporting kidney function, stating the need for more efficient and targeted therapies to enhance repair. Mesenchymal Stromal Cells (MSCs) and their secretome, either as conditioned medium (CM) or extracellular vesicles (EVs), have emerged as promising options for regenerative therapy, however, their full potential in treating AKI remains unknown.</p><p><strong>METHODS:</strong> In this study, we employed an in vitro model of chemically-induced ischemia using antimycin A combined with 2-deoxy-D-glucose to induce ischemic injury in proximal tubule epithelial cells. Afterwards, we evaluated the effects of MSC secretome, CM or EVs obtained from adipose tissue, bone marrow and umbilical cord, on ameliorating the detrimental effects of ischemia. To assess the damage and treatment outcomes, we analyzed cell morphology, mitochondrial health parameters (mitochondrial activity, ATP production, mass and membrane potential) and overall cell metabolism by metabolomics.</p><p><strong>RESULTS:</strong> Our findings show that ischemic injury caused cytoskeletal changes confirmed by disruption of the F-actin network, energetic imbalance as revealed by a 50% decrease in the oxygen consumption rate, increased oxidative stress, mitochondrial dysfunction and reduced cell metabolism. Upon treatment with MSC secretome, the morphological derangements were partly restored and ATP production increased by 40-50%, with umbilical cord-derived EVs being most effective. Furthermore, MSC treatment led to phenotype restoration as indicated by an increase in cell bioenergetics, including increased levels of glycolysis intermediates, as well as an accumulation of antioxidant metabolites.</p><p><strong>CONCLUSION:</strong> Our in vitro model effectively replicated the in vivo-like morphological and molecular changes observed during ischemic injury. Additionally, treatment with MSC secretome ameliorated proximal tubule damage, highlighting its potential as a viable therapeutic option for targeting AKI.</p>
2023-10-16 | MTBLS6279 | MetaboLights
Project description:Mammalian kidney has very limited ability to repair or regenerate after acute kidney injury (AKI). The maladaptive repair of AKI promotes the progression to chronic kidney disease (CKD). Therefore, it is extremely urgent to explore new strategies to promote the repair/regeneration of injured renal tubules after AKI. It has been shown that hypoxia induces heart regeneration in adult mice. However, it is unknown whether hypoxia can induce kidney regeneration after AKI. In this study, we used a prolyl hydroxylase domain inhibitor (PHDI), MK-8617, to mimic hypoxia condition and found that MK-8617 significantly ameliorates ischemia reperfusion injury (IRI) induced acute kidney injury. We then showed that MK-8617 dramatically facilitates renal regeneration via promoting the proliferation of injured renal proximal tubular cells (RPTCs) after IRI-induced AKI. We then performed bulk mRNA sequencing and discovered that multiple nephrogenesis- related genes were significantly upregulated with MK-8617 pretreatment. Furthermore, we showed that MK-8617 may alleviate proximal tubule injury via stabilizing HIF-1α protein specifically in renal proximal tubular cells. We also demonstrated that MK-8617 promotes the reprogramming of renal proximal tubular cells to Sox9+ renal progenitor cells, and the regeneration of renal proximal tubules. In summary, we discovered that inhibition of prolyl hydroxylase improves renal proximal tubule regeneration after IRI-induced AKI via promoting the reprogramming of renal proximal tubular cells to Sox9+ renal progenitor cells.
2024-10-10 | GSE247560 | GEO
Project description:Acute kidney injury (AKI) is a major health care concern. There are no therapies for the treatment of AKI. The primary site of damage during AKI is the proximal tubule, which are highly metabolically active cells that rely upon fatty acids for energy. Proximal tubules are notably mitochondria- and peroxisomes-rich cell type that mediate fatty acid oxidation (FAO). Sirtuins reverse post-translational lysine acylation and control many biological processes including FAO. Sirtuin1 and sirtuin3 are protective against AKI. However, the role of the mitochondrial Sirtuin5 (Sirt5) during AKI has yet to be determined. We found Sirt5 to be highly expressed in the proximal tubule. At baseline Sirtuin5 knockout (Sirt5-/-) animals had modestly decreased mitochondrial function but significantly increased FAO, which was localized to the peroxisome. While no overt kidney phenotype was observed in SIRT5-/- mice, following ischemia reperfusion injury Sirt5-/- animals had significantly improved kidney function and less tissue damage. This coincided with increased peroxisomal activity in the Sirt5-/- proximal tubules in preference to the mitochondria. Here we have identified a novel mechanism driving protection of kidneys from ischemic injury. If this can be harnessed it may prove to be an effective therapeutic.
2019-02-13 | MSV000083439 | MassIVE
Project description:The kidney plays a critical role in fluid homeostasis, glucose control, and drug excretion. Loss of kidney function due to drug-induced nephrotoxicity affects over 20% of the adult population. The kidney proximal tubule is a complex vascularized structure that particularly vulnerable to drug-induced nephrotoxicity. Here we introduce a model of vascularized human kidney organoids with integrated tissue embedded micro-sensors for oxygen, glucose, lactate and glutamine, providing real time assessment of cellular metabolism. Our model shows that both the immunosuppressive drug cyclosporin (Neoral®) and the anti-cancer drug cisplatin (Platinol®) disrupt proximal tubule polarity at sub-toxic concentrations, leading to glucose accumulation and lipotoxicity. Impeding glucose reabsorption using glucose transport inhibitors blocked cyclosporin and cisplatin toxicity by 1,000 to 10-folds, respectively. Retrospective study of 247 patients receiving cyclosporin or cisplatin in combination with the SGLT-2 inhibitor gliflozin showed significant reduction in creatinine and uric acid recognized markers of kidney damage. These results demonstrate the potential of sensor-integrated organoid-on-chip platforms to elucidate new mechanisms of action and rapidly reformulate effective therapeutic solutions, increasing drug safety and reducing the cost of clinical and commercial failures.
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Project description:The cellular mechanisms of kidney tubule repair are poorly characterized in human. Here, we applied single-cell RNA sequencing to analyze the kidney in the first days after acute injury in 5 patients with severe COVID19. We found that tubule repair follows two converging patterns involving the plasticity of mature tubule cells and the expansion and differentiation of progenitor-like cells. Tubule repair by cell plasticity displayed substantial similarities between mouse and man and determined the transient expansion of undifferentiated tubule cells with altered functional and metabolic properties. Progenitor-like cells marked by PROM1 proliferated in response to injury and followed a differentiation process characterized by the sequential activation of the WNT, NOTCH and HIPPO signaling pathways. Taken together, our analyses reveal cell state transitions and fundamental cellular hierarchies underlying kidney injury and repair in patients.
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