We thank the authors and reviewers for their contributions to this collection. We also acknowledge funding from the NIH (DK106284 and DK123542) and the Oxalosis &#38; Hyperoxaluria Foundation &#8211; American Society of Nephrology KidneyCure Transition to Independence Grant.

<ol>
	<li>Kovesdy, C. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epidemiology+of+chronic+kidney+disease:+An+update+2022.">Epidemiology of chronic kidney disease: An update 2022.</a> Kidney International Supplements. 12 (1), 7-11 (2022).</li><li>Bellomo, R., Kellum, J. A., Ronco, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acute+kidney+injury.">Acute kidney injury.</a> Lancet. 380 (9843), 756-766 (2012).</li><li>Malek, M., Nematbakhsh, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Renal+ischemia/reperfusion+injury;+From+pathophysiology+to+treatment.">Renal ischemia/reperfusion injury; From pathophysiology to treatment.</a> Journal of Renal Injury Prevention. 4 (2), 20-27 (2015).</li><li>Saba, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Manganese+porphyrin+reduces+renal+injury+and+mitochondrial+damage+during+ischemia/reperfusion.">Manganese porphyrin reduces renal injury and mitochondrial damage during ischemia/reperfusion.</a> Free Radical Biology and Medicine. 42 (10), 1571-1578 (2007).</li><li>Mitchell, T., Saba, H., Laakman, J., Parajuli, N., MacMillan-Crow, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+mitochondrial-derived+oxidants+in+renal+tubular+cell+cold-storage+injury.">Role of mitochondrial-derived oxidants in renal tubular cell cold-storage injury.</a> Free Radical Biology and Medicine. 49 (8), 1273-1282 (2010).</li><li>Liu, K. D., Humphreys, B. D., Endre, Z. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+ten+barriers+for+translation+of+animal+data+on+AKI+to+the+clinical+setting.">The ten barriers for translation of animal data on AKI to the clinical setting.</a> Intensive Care Medicine. 43 (6), 898-900 (2017).</li><li>Doulamis, I. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+large+animal+model+for+acute+kidney+injury+by+temporary+bilateral+renal+artery+occlusion.">A large animal model for acute kidney injury by temporary bilateral renal artery occlusion.</a> Journal of Visualized Experiments. (168), e62230 (2021).</li><li>Cantero-Navarro, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+macrophages+and+related+cytokines+in+kidney+disease.">Role of macrophages and related cytokines in kidney disease.</a> Frontiers in Medicine. 8, 688060 (2021).</li><li>Taguchi, K., Sugahara, S., Elias, B. C., Brooks, C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+the+source+of+secreted+proteins+in+the+kidney+by+brefeldin+A+injection.">Identification of the source of secreted proteins in the kidney by brefeldin A injection.</a> Journal of Visualized Experiments. (177), e63178 (2021).</li><li>Wei, Q., Mi, Q. S., Dong, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+regulation+and+function+of+microRNAs+in+kidney+diseases.">The regulation and function of microRNAs in kidney diseases.</a> IUBMB Life. 65 (7), 602-614 (2013).</li><li>Cui, C., Cui, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+relationship+of+human+tissue+microRNAs+with+those+from+body+fluids.">The relationship of human tissue microRNAs with those from body fluids.</a> Scientific Reports. 10, 5644 (2020).</li><li>Yanai, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+real-time+polymerase+chain+reaction+evaluation+of+microRNA+expression+in+kidney+and+serum+of+mice+with+age-dependent+renal+impairment.">Quantitative real-time polymerase chain reaction evaluation of microRNA expression in kidney and serum of mice with age-dependent renal impairment.</a> Journal of Visualized Experiments. (182), e63258 (2022).</li><li>Holmes, H. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+3D+robotic+ultrasound+for+in+vivo+analysis+of+mouse+kidneys.">Use of 3D robotic ultrasound for in vivo analysis of mouse kidneys.</a> Journal of Visualized Experiments. (174), e62682 (2021).</li><li>Wagner, M. C., Sandoval, R. M., Campos-Bilderback, S. B., Molitoris, B. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Using+2-photon+microscopy+to+quantify+the+effects+of+chronic+unilateral+ureteral+obstruction+on+glomerular+processes.">Using 2-photon microscopy to quantify the effects of chronic unilateral ureteral obstruction on glomerular processes.</a> Journal of Visualized Experiments. (181), e63329 (2022).</li><li>Daudon, M., Hennequin, C., Boujelben, G., Lacour, B., Jungers, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Serial+crystalluria+determination+and+the+risk+of+recurrence+in+calcium+stone+formers.">Serial crystalluria determination and the risk of recurrence in calcium stone formers.</a> Kidney International. 67 (5), 1934-1943 (2005).</li><li>Kumar, P., Bell, A., Mitchell, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Estimation+of+urinary+nanocrystals+in+humans+using+calcium+fluorophore+labeling+and+nanoparticle+tracking+analysis.">Estimation of urinary nanocrystals in humans using calcium fluorophore labeling and nanoparticle tracking analysis.</a> Journal of Visualized Experiments. (168), e62192 (2021).</li><li>Kumar, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dietary+oxalate+loading+impacts+monocyte+metabolism+and+inflammatory+signaling+in+humans.">Dietary oxalate loading impacts monocyte metabolism and inflammatory signaling in humans.</a> Frontiers in Immunology. 12, 617508 (2021).</li><li>Yanai, K., Kaneko, S., Ishii, H., Aomatsu, A., Morishita, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Delivery+of+exogenous+artificially+synthesized+miRNA+mimic+to+the+kidney+using+polyethylenimine+nanoparticles+in+several+kidney+disease+mouse+models.">Delivery of exogenous artificially synthesized miRNA mimic to the kidney using polyethylenimine nanoparticles in several kidney disease mouse models.</a> Journal of Visualized Experiments. (183), e63302 (2022).</li></ol>

The authors have nothing to disclose.

Introduction 
Research within the field of nephrology has significantly improved the well-being of individuals impacted by kidney disease. However, kidney disease is a still major healthcare problem that impacts more than 10% of the population and is responsible for increased healthcare costs and mortality worldwide1. Unfortunately, the precise mechanisms leading to kidney disease have not been fully elucidated. This JoVE Methods Collection highlights recent strategies to define kidney pathophysiology. Specifically, the research groups present a novel animal model and methods to evaluate or treat kidney disease using experimental models or clinical samples. The work presented in this collection is another series of excellent contributions to the field of nephrology and may provide new avenues to prevent, diagnose, or treat kidney disease in the future.

A translational animal model of acute kidney injury 
Acute kidney injury (AKI) is a common form of kidney disease2. Several experimental models have been used to study renal ischemia/reperfusion (I/R) injury3,4,5. However, translating these findings to improve the clinical setting has posed many challenges since there are significant differences amongst species6. Doulamis et al. developed a swine renal I/R model using a bilateral balloon catheter to address this issue7. Their model was highly reproducible, caused reduced urinary output and estimated glomerular filtration rate, and increased plasma creatinine and blood urea nitrogen levels in animals. Further, gross tissue and histological examination revealed infarction and hemorrhaging within the kidneys. This new model is clinically relevant and may aid in further understanding AKI in humans.
Strategies to examine cellular mechanisms involved in kidney disease 
Cytokines can contribute to kidney injury and repair8. Thus, it is important to identify the source and role of cytokines within the kidney. Taguchi et al. developed a technique to understand protein secretion using brefeldin A (BFA), a protein secretion inhibitor, in AKI and chronic kidney disease mouse models9. They injected BFA into the tail vein of mice and subsequently evaluated their kidneys using immunofluorescence staining. They determined that BFA inhibited protein secretion and caused cytokine buildup in certain cell types using cell-specific markers. This innovative protocol could be used to study protein secretion in several renal pathologies.
Another proposed mechanism thought to contribute to kidney disease is microRNAs (miRNA)10, which are known to degrade and inhibit mRNA transcription. It has been reported that there is a correlation between miRNA expression in tissue and serum in humans11. However, there is no method to purify and quantify both mouse kidneys and serum miRNA. Yanai et al. optimized a quantitative real-time polymerase chain reaction method to evaluate miRNA expression in the kidney and serum of mice with age-dependent renal impairment12. They also established a correlation between both miRNA levels. This protocol is high-throughput and could be applicable to many pathological conditions.
New imaging techniques to evaluate kidney disease 
Noninvasive imaging of the kidney can assess structural and functional defects. However, there are some limitations with the current methods. To circumvent this, Holmes et al. developed a simple, cost-effective 3D imaging technique based on robotic ultrasound (US) technology to quantify kidney, liver, and cardiac function13. Their protocol provided consistently high-quality images and could be used for studies where gross tissue, tumor sizes, or the benefits of therapeutics could be monitored. The greatest strength of this method is the high output of data. For example, three rats can be imaged at once, or 20&#8211;30 mice can be imaged per hour. This protocol is a quick alternative to traditional noninvasive imaging modalities.
Another research group in the collection used imaging to study unilateral ureteral obstruction (UUO), an injury model that induces surface glomeruli. Wagner et al. used intravital two-photon microscopy to monitor UUO in rats that develop surface glomeruli and those that do not develop surface glomeruli14. Two-photon microscopy provides an increased depth of penetration and examination of different regions of the kidney. The authors found that UUO induced inflammation and fibrosis, and also decreased red blood cell flow in rat kidneys. Quantifying blood flow, vasoconstriction, and dilatation in response to drugs and inflammation are a few benefits this method provides. Collectively, these studies provide new imaging strategies to study various tissues.
Estimating nanocrystalluria as a predictor of kidney disease 
It has been proposed that assessing urinary crystals could be useful to predict kidney stone risk15. We developed a protocol to quantify urinary nanocrystals (&#8804;1 &#181;m) from healthy adults before and after they consumed a dietary oxalate load known to induce urinary oxalate levels16. We isolated and analyzed calcium-containing urinary nanocrystals using a calcium fluorophore and nanoparticle tracking analysis (NTA). We determined that NTA can specifically detect calcium-containing nanocrystals in urine, and the results were consistent with our previous findings17. This protocol can be used to study urinary nanocrystals in stone formers or various fields of research where crystalopathies occur.
Therapeutic approaches to treat kidney disease 
Artificially synthesized miRNA mimics have been suggested as a possible treatment for kidney disease. However, serum RNAase can degrade exogenous miRNA mimics18. Yanai et al. produced non-viral vectors based on linear polymer, polyethylenimine nanoparticles (PEI-NPs), to safely deliver a miRNA mimic to the kidneys of mice18. They proved that the mimic was effective in overexpressing the targeted miRNA. The limitations of this method include the substantial amount of PEI-NPs required for larger animal models and the lack of specificity to the kidneys. However, based on the success of RNA vaccines, miRNA mimics could be an ideal therapeutic option for kidney disease.
Concluding remarks 
The comprehensive research presented here&#160;introduces a new animal model and techniques to study prevalent issues surrounding nephrology. It is important to note that some of these methods could apply to other areas outside of nephrology. The next logical steps will be to investigate how these methods could be tested in larger animal models and clinical trials. We hope that the sharing of these protocols will expand our knowledge about renal pathophysiology and birth new ideas to mitigate kidney disease in individuals.

novel translational approaches to study kidney disease

ARTICLES DISCUSSED:
Kumar, P., Bell, A., Mitchell, T., Estimation of urinary nanocrystals in humans using calcium fluorophore labeling and nanoparticle tracking analysis. Journal of Visualized Experiments. (168), e62192 (2021).
Doulamis, I. P. et al. A large animal model for acute kidney injury by temporary bilateral renal artery occlusion. Journal of Visualized Experiments. (168), e62230 (2021).
Taguchi, K., Sugahara, S., Elias, B. C., Brooks, C. R. Identification of the source of secreted proteins in the kidney by brefeldin A injection. Journal of Visualized Experiments. (177), e63178 (2021).
Yanai, K. et al. Quantitative real-time polymerase chain reaction evaluation of microRNA expression in kidney and serum of mice with age-dependent renal impairment. Journal of Visualized Experiments. (182), e63258 (2022).
Holmes, H. L. et al. Use of 3D robotic ultrasound for in vivo analysis of mouse kidneys. Journal of Visualized Experiments. (174), e62682 (2021).
Yanai, K., Kaneko, S., Ishii, H., Aomatsu, A., Morishita, Y. Delivery of exogenous artificially synthesized miRNA mimic to the kidney using polyethylenimine nanoparticles in several kidney disease mouse models. Journal of Visualized Experiments. (183), e63302 (2022).
Wagner, M. C., Sandoval, R. M., Campos-Bilderback, S. B., Molitoris, B. A. Using 2-photon microscopy to quantify the effects of chronic unilateral ureteral obstruction on glomerular processes. Journal of Visualized Experiments. (181), e63329 (2022).

Read this Scientific Article Protocol about Novel Translational Approaches To Study Kidney Disease at JoVE.com

Novel Translational Approaches To Study Kidney Disease

ARTICLES DISCUSSED:
Kumar, P., Bell, A., Mitchell, T., Estimation of urinary nanocrystals in humans using calcium fluorophore labeling and nanoparticle ...

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481 ビュー.  University of Alabama at Birmingham. Novel Translational Approaches To Study Kidney Disease

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spectral karyotyping to study chromosome abnormalities in humans and mice with polycystic kidney disease

Spectral Karyotyping to Study Chromosome Abnormalities in Humans and Mice with Polycystic Kidney Disease

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chemical cartography approaches to study trypanosomatid infection

Chemical Cartography Approaches to Study Trypanosomatid Infection

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phage phenomics: physiological approaches to characterize novel viral proteins

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exploring caspase mutations and post-translational modification by molecular modeling approaches

Exploring Caspase Mutations and Post-Translational Modification by Molecular Modeling Approaches

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delivery of exogenous artificially synthesized mirna mimic to the kidney using polyethylenimine nanoparticles in several kidney disease mouse models

Delivery of Exogenous Artificially Synthesized miRNA Mimic to the Kidney Using Polyethylenimine Nanoparticles in Several Kidney Disease Mouse Models

Here, we deliver exogenous artificially synthesized miRNA mimics to the kidney via tail vein injection of a nonviral vector and polyethylenimine nanoparticles in several kidney disease mouse models. This led to significant overexpression of target miRNA in the kidney, resulting in inhibited progression of kidney disease in several mouse models.

assessment of kidney function in mouse models of glomerular disease

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a novel clinical grade isolation method for human kidney perivascular stromal cells

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intravenous microinjections of zebrafish larvae to study acute kidney injury

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orthotopic rat kidney transplantation: a novel and simplified surgical approach

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isolation of intermediate filament proteins from multiple mouse tissues to study aging-associated post-translational modifications

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mass spectrometric approaches to study protein structure and interactions in lyophilized powders

Mass Spectrometric Approaches to Study Protein Structure and Interactions in Lyophilized Powders

Here, we present detailed protocols for solid-state amide hydrogen/deuterium exchange mass spectrometry (ssHDX-MS) and solid-state photolytic labeling mass spectrometry (ssPL-MS) for proteins in solid powders. The methods provide high-resolution information on protein conformation and interactions in the amorphous solid-state, which may be useful in formulation design.

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assessment of vascular function in patients with chronic kidney disease

Assessment of Vascular Function in Patients With Chronic Kidney Disease

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experimental approaches to study mitochondrial localization and function of a nuclear cell cycle kinase, cdk1

Experimental Approaches to Study Mitochondrial Localization and Function of a Nuclear Cell Cycle Kinase, Cdk1

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bilateral renal ischemia-reperfusion model for acute kidney injury in mice

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Developing Murine Acute Kidney Injury Model Using Bilateral Ischemia-Reperfusion

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