Quantitative Intravital Calcium Imaging in Mouse Kidney.

Intracellular calcium is an important regulator of solute transport in renal epithelial cells, and disordered calcium signaling may underlie the pathogenesis of certain kidney diseases. Intravital multiphoton imaging of the kidney in transgenic mice expressing highly sensitive fluorescent reporters allows detailed study of calcium signals within different specialized segments of the renal tubule and how these are integrated with other cellular processes. Moreover, changes in activity can be observed in real time in response to physiological interventions or disease-causing insults. In this chapter, we will provide a detailed protocol for performing this powerful research technique.

Recurrent Fanconi Syndrome Induced by Lenalidomide: A Case Report.

Fanconi syndrome (FS) is a complex disorder characterized by a reabsorption defect in the proximal renal tubule (PT), leading to urinary loss of molecules such as glucose, phosphate, calcium, amino acids, bicarbonate, potassium, and low-molecular-weight proteins. Its etiology can be genetic or acquired, with drug toxicity being a significant cause of the acquired forms. The heterogeneous manifestations of FS, whether in its partial or complete form, can pose challenges in the emergency department; nevertheless, it should be considered in certain patients, as understanding its cause is crucial for initiating effective treatment. We present the case of a 59-year-old female patient with FS who was treated with lenalidomide in the context of stage III IgG kappa multiple myeloma according to the Salmon Durie classification. We highlight the recurrent nature of this syndrome in this patient.

HO-1 activation contributes to cadmium-induced ferroptosis in renal tubular epithelial cells via increasing the labile iron pool and promoting mitochondrial ROS generation.

Cadmium (Cd), a prevalent environmental contaminant, has attracted widespread attention due to its serious health hazards. Ferroptosis is a form of iron-dependent oxidative cell death that contributes to the development of various kidney diseases. However, the mechanisms underlying the occurrence of ferroptosis in Cd-induced renal tubular epithelial cells (TECs) have not been fully elucidated. Hereby, both in-vitro and in-vivo experiments were established to elucidate this issue. In this study, we found that Cd elicited accumulation of lipid peroxides due to intracellular ferrous ion (Fe2+) overload and glutathione depletion, contributing to ferroptosis. Inhibition of ferroptosis via chelation of Fe2+ or reduction of lipid peroxidation can significantly mitigate Cd-induced cytotoxicity. Renal transcriptome analysis revealed that the activation of heme oxygenase 1 (HO-1) was closely related to ferroptosis in Cd-induced TECs injury. Cd-induced ferroptosis and resultant TECs injury are significantly alleviated due to HO-1 inhibition, demonstrating the crucial role of HO-1 in Cd-triggered ferroptosis. Further studies showed that accumulation of lipid peroxides due to iron overload and mitochondrial ROS (mtROS) generation was responsible for HO-1-triggered ferroptosis in Cd-induced cytotoxicity. In conclusion, the current study demonstrates that excessively upregulating HO-1 promotes iron overload and mtROS overproduction to trigger ferroptosis in Cd-induced TECs injury, highlighting that targeting HO-1-mediated ferroptosis may provide new ideas for preventing Cd-induced nephrotoxicity.

Revolutionizing renal research: The future of kidney-on-a-chip in biotechnology.

In vitro models of kidneys have limited effectiveness owing to the complex structure and functions of the kidney when compared with other organs. Therefore many renal function evaluations are currently being carried out through animal experiments. In contrast, efforts are being made to apply biomimetic systems, such as organ-on-a-chip, which is based on microfluidic device technology, to serve as an in vitro model for the kidney. These systems aimed to recreate a physiological cultivation environment. This review has provided an overview of organ-on-a-chip research focused on glomeruli and tubules as in vitro models for the kidney and discusses future prospects.

WWC1 upregulation accelerates hyperuricemia by reduction in renal uric acid excretion through Hippo signaling pathway.

Hyperuricemia (HUA) is a metabolic disorder characterized by elevated serum uric acid (UA), primarily attributed to the hepatic overproduction and renal underexcretion of UA. Despite the elucidation of molecular pathways associated with this underexcretion, the etiology of HUA remains largely unknown. In our study, using by Uox knockout rats, HUA mouse, and cell line models, we discovered that the increased WWC1 levels were associated with decreased renal UA excretion. Additionally, using knockdown and overexpression approaches, we found that WWC1 inhibited UA excretion in renal tubular epithelial cells. Mechanistically, WWC1 activated the Hippo pathway, leading to phosphorylation and subsequent degradation of the downstream transcription factor YAP1, thereby impairing the ABCG2 and OAT3 expression through transcriptional regulation. Consequently, this reduction led to a decrease in UA excretion in renal tubular epithelial cells. In conclusion, our study has elucidated the role of upregulated WWC1 in renal tubular epithelial cells inhibiting the excretion of UA in the kidneys and causing HUA.

Association of high vibration perception threshold with reduced renal function in patients with type 2 diabetes.

Objective: To investigate the correlation between vibration sensory threshold (VPT) and renal function, including glomerulus and renal tubule, in patients with type 2 diabetes mellitus (T2DM). Methods: A total of 1274 patients with T2DM who were enrolled in the Department of Endocrinology of the First Affiliated Hospital of Fujian Medical University between January 2017 and June 2020 were included. Patients were grouped according to VPT levels and divided into three groups, including the normal VPT group (VPT<15V), the mild-moderate elevated VPT group (VPT15~25V), and the severely elevated VPT group (VPT≥25 V). Linear correlation analysis was used to analyze the correlation between VPT and renal functions, including glomerulus markers urine microalbumin (MA) and urinary immunoglobulin G (U-IgG), and renal tubule marker α1-microglobulin (α1-MG). Chronic kidney disease (CKD) was defined according to Kidney Disease Improving Global Outcomes (KDIGO) criteria. The binary logistic regression of the relation between VPT and CKD, eGFR<60 ml/min, and UACR >30 mg/g were expressed. Results: In the mild-moderate and severely elevated VPT group, injury biomarkers of glomerulus (MA and U-IgG), renal tubule (α1-MG), and the incidence of CKD, eGFR<60 ml/min, and UACR > 30 mg/g were gradually increased compared with the normal VPT group. Furthermore, patients with diabetes and severely elevated VPT had significantly higher levels of MA (ß=197.54, p=0.042) and α1-MG (ß=11.69, p=0.023) compared to those with normal VPT. Also, patients with mild-moderate elevated VPT demonstrate significantly higher levels of MA (ß=229.02, p=0.005). Patients in mild-moderate elevated VPT group (OR=1.463, 95% CI 1.005-2.127; OR=1.816, 95% CI 1.212-2.721) and severely elevated VPT group (OR=1.704, 95% CI 1.113-2.611; OR=2.027, 95% CI 1.248-3.294) are at a higher incidence of CKD and elevated levels of UACR>30mg/g compared to those in the VPT normal group. Moreover, the incidence of positive Upro was notably higher in the severely elevated VPT group (OR=1.738, 95% CI 1.182-2.556). However, this phenomenon was not observed in the incidence of eGFR <60 ml/min. Conclusion: A higher VPT is positively associated with the incidence of CKD in patients with T2DM, particularly with elevated UACR. VPT may serve as a marker for glomerulus and renal tubule injury.

Chronic Kidney Disease Diets for Kidney Failure Prevention: Insights from the IL-11 Paradigm.

Nearly every fifth adult in the United States and many older adults worldwide are affected by chronic kidney disease (CKD), which can progress to kidney failure requiring invasive kidney replacement therapy. In this review, we briefly examine the pathophysiology of CKD and discuss emerging mechanisms involving the physiological resolution of kidney injury by transforming growth factor beta 1 (TGFß1) and interleukin-11 (IL-11), as well as the pathological consequences of IL-11 overproduction, which misguides repair processes, ultimately culminating in CKD. Taking these mechanisms into account, we offer an overview of the efficacy of plant-dominant dietary patterns in preventing and managing CKD, while also addressing their limitations in terms of restoring kidney function or preventing kidney failure. In conclusion, this paper outlines novel regeneration strategies aimed at developing a reno-regenerative diet to inhibit IL-11 and promote repair mechanisms in kidneys affected by CKD.

Polarized HLA Class I Expression on Renal Tubules Hinders the Detection of Donor-Specific Urinary Extracellular Vesicles.

Purpose: Kidney transplantation is the optimal treatment for patients with end-stage kidney disease. Donor-specific urinary extracellular vesicles (uEVs) hold potential as biomarkers for assessing allograft status. We aimed to develop a method for identifying donor-specific uEVs based on human leukocyte antigen (HLA) mismatching with the kidney transplant recipients (KTRs). Patients and Methods: Urine and plasma were obtained from HLA-A2+ donors and HLA-A2- KTRs pre-transplant. CD9 (tetraspanin, EV marker) and HLA-A2 double-positive (CD9+ HLA-A2+) EVs were quantified using isolation-free imaging flow cytometry (IFCM). Healthy individuals' urine was used to investigate CD9+ HLA-class-I+ uEV quantification using IFCM, time-resolved fluoroimmunoassay (TR-FIA), and immunogold staining cryo-electron microscopy (cryo-EM). Culture-derived CD9+ HLA-class-I+ EVs were spiked into the urine to investigate urine matrix effects on uEV HLA detection. Deceased donor kidneys and peritumoral kidney tissue were used for HLA class I detection with histochemistry. Results: The concentrations of CD9+ HLA-A2+ EVs in both donor and recipient urine approached the negative (detergent-treated) control levels for IFCM and were significantly lower than those observed in donor plasma. In parallel, universal HLA class I+ uEVs were similarly undetectable in the urine and uEV isolates compared with plasma, as verified by IFCM, TR-FIA, and cryogenic electron microscopy. Culture supernatant containing HLA class I+ vesicles from B, T, and human proximal tubule cells were spiked into the urine, and these EVs remained stable at 37°C for 8 hours. Immunohistochemistry revealed that HLA class I was predominantly expressed on the basolateral side of renal tubules, with limited expression on their urine/apical side. Conclusion: The detection of donor-specific uEVs is hindered by the limited release of HLA class I+ EVs from the kidney into the urine, primarily due to the polarized HLA class I expression on renal tubules. Identifying donor-specific uEVs requires further advancements in recognizing transplant-specific uEVs and urine-associated markers.

[Evaluation of chemical constituents of Draconis Sanguis and its protective effect on renal tubular injury in diabetic kidney disease].

The chemical constituents of Draconis Sanguis were preliminarily studied by macroporous resin, silica gel, dextran gel, and high-performance liquid chromatography. One retro-dihydrochalcone, four flavonoids, and one stilbene were isolated. Their chemical structures were identified as 4-hydroxy-2,6-dimethoxy-3-methyldihydrochalcone(1), 4'-hydroxy-5,7-dimethoxy-8-methylflavan(2), 7-hydroxy-4',5-dimethoxyflavan(3),(2S)-7-hydroxy-5-methoxy-6-methylflavan(4),(2S)-7-hydroxy-5-methoxyflavan(5), and pterostilbene(6) by modern spectroscopy, physicochemical properties, and literature comparison. Compound 1 was a new compound. Compounds 2 and 6 were first found in the Arecaceae family. Compound 5 had the potential to prevent and treat diabetic kidney disease.

Ferroptosis-related genes, a novel therapeutic target for focal segmental glomerulosclerosis.

Recent studies have suggested that ferroptosis participates in various renal diseases. However, its effect on focal segmental glomerulosclerosis remains unclear. This study analyzed the GSE125779 and GSE121211 datasets to identify the differentially expressed genes (DEGs) in renal tubular samples with and without FSGS. The Cytoscape was used to construct the protein-protein interaction network. Moreover, the ferroptosis-related genes (FRGs) were obtained from the ferroptosis database, while ferroptosis-related DEGs were obtained by intersection with DEGs. The target genes were analyzed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. The GSE108112 dataset was used to verify the expression of target FRGs. Besides, we built the mRNA-miRNA network regarding FRGs using the NetworkAnalyst database, and circRNAs corresponding to key miRNAs were predicted in the ENCORI database. In this study, 16 ferroptosis-related DEGs were identified between FSGS and healthy subjects, while five co-expressed genes were obtained by three topological algorithms in Cytoscape. These included the most concerned Hub genes JUN, HIF1A, ALB, DUSP1 and ATF3. The KEGG enrichment analysis indicated that FRGs were associated with mitophagy, renal cell carcinoma, and metabolic pathways. Simultaneously, the co-expressed hub genes were analyzed to construct the mRNA-miRNA interaction network and important miRNAs such as hsa-mir-155-5p, hsa-mir-1-3p, and hsa-mir-124-3p were obtained. Finally, 75 drugs targeting 54 important circRNAs and FRGs were predicted. This study identified the Hub FRGs and transcriptomic molecules from FSGS in renal tubules, thus providing novel diagnostic and therapeutic targets for FSGS.

An aldose reductase inhibitor, WJ-39, ameliorates renal tubular injury in diabetic nephropathy by activating PINK1/Parkin signaling.

Renal tubular injury is a critical factor during the early stages of diabetic nephropathy (DN). Proximal tubular epithelial cells, which contain abundant mitochondria essential for intracellular homeostasis, are susceptible to disruptions in the intracellular environment, making them especially vulnerable to diabetic state disorders, which may be attributed to their elevated energy requirements and reliance on aerobic metabolism. It is widely thought that overactivation of the polyol pathway is implicated in DN pathogenesis, and inhibition of aldose reductase (AR), the rate-limiting enzyme in this pathway, represents a promising therapeutic avenue. WJ-39, a novel aldose reductase inhibitor, was investigated in this study for its protective effects on renal tubules in DN and the underlying mechanisms. Our findings revealed that WJ-39 significantly ameliorated the renal tubular morphology in high-fat diet (HFD)/streptozotocin (STZ)-induced DN rats, concurrently inhibiting fibrosis. Notably, WJ-39 safeguarded the structure and function of renal tubular mitochondria by enhancing mitochondrial dynamics. This involved the regulation of mitochondrial fission and fusion proteins and the promotion of PTEN-induced putative kinase 1 (PINK1)/Parkin-mediated mitophagy. Furthermore, WJ-39 demonstrated the inhibition of endogenous apoptosis by mitigating the production of mitochondrial reactive oxygen species (ROS). The protective effects of WJ-39 on mitochondria and apoptosis were countered in high glucose-treated HK-2 cells upon transfection with PINK1 siRNA. Overall, our findings suggest that WJ-39 protects the structural and functional integrity of renal tubules in DN, which may be attributed to its capacity to inhibit aldose reductase activity, activate the PINK1/Parkin signaling pathway, promote mitophagy, and alleviate apoptosis.

When Two Syndromes Collide: Managing Fanconi and Refeeding Syndrome in a Single Patient.

Refeeding syndrome is the potentially fatal shift in fluids and electrolytes that may occur in malnourished patients after receiving artificial refeeding. Its hallmark feature is hypophosphatemia, although other electrolytes might also be affected. Fanconi syndrome is a generalized dysfunction of the proximal tubule characterized by proximal renal tubular acidosis (RTA), phosphaturia, glycosuria, aminoaciduria, and proteinuria. The etiology of Fanconi syndrome can be either acquired or inherited, and drugs, among them tenofovir, are a common acquired cause of this disease. We present the case of a patient with AIDS and polysubstance abuse who was admitted due to pneumonia, completed treatment, was then started on antiretroviral medication (ART) that included tenofovir alafenamide (TAF) and began presenting severe episodes of hypophosphatemia along with other electrolyte imbalances, leading the workup denoted in the case, severe complications and finally to the patient's demise. Most cases of tenofovir-related Fanconi syndrome are related to tenofovir disoproxil fumarate, but very few cases have been reported with TAF. Our case highlights this rare complication of therapy with TAF and how artificial feeding can contribute to severe electrolyte abnormalities and worsen outcomes.

Renal proximal tubule-on-a-chip in PDMS: fabrication, functionalization, and RPTEC:HUVEC co-culture evaluation.

'On-a-chip' technology advances the development of physiologically relevant organ-mimicking architecture by integrating human cells into three-dimensional microfluidic devices. This method also establishes discrete functional units, faciliting focused research on specific organ components. In this study, we detail the development and assessment of a convoluted renal proximal tubule-on-a-chip (PT-on-a-chip). This platform involves co-culturing Renal Proximal Tubule Epithelial Cells (RPTEC) and Human Umbilical Vein Endothelial Cells (HUVEC) within a polydimethylsiloxane microfluidic device, crafted through a combination of 3D printing and molding techniques. Our PT-on-a-chip significantly reduced high glucose level, exhibited albumin uptake, and simulated tubulopathy induced by amphotericin B. Remarkably, the RPTEC:HUVEC co-culture exhibited efficient cell adhesion within 30 min on microchannels functionalized with plasma, 3-aminopropyltriethoxysilane, and type-I collagen. This approach significantly reduced the required incubation time for medium perfusion. In comparison, alternative methods such as plasma and plasma plus polyvinyl alcohol were only effective in promoting cell attachment to flat surfaces. The PT-on-a-chip holds great promise as a valuable tool for assessing the nephrotoxic potential of new drug candidates, enhancing our understanding of drug interactions with co-cultured renal cells, and reducing the need for animal experimentation, promoting the safe and ethical development of new pharmaceuticals.

Renal tubule-specific Atgl deletion links kidney lipid metabolism to glucagon-like peptide 1 and insulin secretion independent of renal inflammation or lipotoxicity.

OBJECTIVE: Lipotoxic injury from renal lipid accumulation in obesity and type 2 diabetes (T2D) is implicated in associated kidney damage. However, models examining effects of renal ectopic lipid accumulation independent of obesity or T2D are lacking. We generated renal tubule-specific adipose triglyceride lipase knockout (RT-SAKO) mice to determine if this targeted triacylglycerol (TAG) over-storage affects glycemic control and kidney health. METHODS: Male and female RT-SAKO mice and their control littermates were tested for changes in glycemic control at 10-12 and 16-18 weeks of age. Markers of kidney health and blood lipid and hormone concentrations were analyzed. Kidney and blood lysophosphatidic acid (LPA) levels were measured, and a role for LPA in mediating impaired glycemic control was evaluated using the LPA receptor 1/3 inhibitor Ki-16425. RESULTS: All groups remained insulin sensitive, but 16- to 18-week-old male RT-SAKO mice became glucose intolerant, without developing kidney inflammation or fibrosis. Rather, these mice displayed lower circulating insulin and glucagon-like peptide 1 (GLP-1) levels. Impaired first-phase glucose-stimulated insulin secretion was detected and restored by Exendin-4. Kidney and blood LPA levels were elevated in older male but not female RT-SAKO mice, associated with increased kidney diacylglycerol kinase epsilon. Inhibition of LPA-mediated signaling restored serum GLP-1 levels, first-phase insulin secretion, and glucose tolerance. CONCLUSIONS: TAG over-storage alone is insufficient to cause renal tubule lipotoxicity. This work is the first to show that endogenously derived LPA modulates GLP-1 levels in vivo, demonstrating a new mechanism of kidney-gut-pancreas crosstalk to regulate insulin secretion and glucose homeostasis.

Phosphate: An underrated component of primary hyperparathyroidism.

Primary hyperparathyroidism (PHPT) is a systemic disease that affects all the systems of the body, specifically the bones and the kidneys. Its main action is on calcium homeostasis. It tries to preserve the body's calcium level at the cost of phosphate. The criteria for surgery in asymptomatic PHPT patients revolve around raised serum calcium levels, renal dysfunction or nephrolithiasis, and bone health. It does not take into account the serum phosphate levels. Depending on the serum level, Hypophosphatemia is divided into mild, moderate, and severe categories. In PHPT, several studies have suggested that asymptomatic PHPT patients with moderate hypophosphatemia may warrant surgical intervention. Treatment of hypophosphatemia in PHPT is based upon the degree of hypophosphatemia, and treatment is given according to that oral or intravenous route; after surgical and medical treatment of PHPT, phosphate levels gradually normalized. But even after these considerations, phosphate levels in PHPT are not given much importance.

The Versatile Role of Uromodulin in Renal Homeostasis and Its Relevance in Chronic Kidney Disease.

Uromodulin, also known as the Tamm-Horsfall protein, is predominantly expressed in epithelial cells of the kidney. It is secreted mainly in the urine, although small amounts are also found in serum. Uromodulin plays an important role in maintaining renal homeostasis, particularly in salt/water transport mechanisms and is associated with salt-sensitive hypertension. It also regulates urinary tract infections, kidney stones, and the immune response in the kidneys or extrarenal organs. Uromodulin has been shown to be associated with the renal function, age, nephron volume, and metabolic abnormalities and has been proposed as a novel biomarker for the tubular function or injury. These findings suggest that uromodulin is a key molecule underlying the mechanisms or therapeutic approaches of chronic kidney disease, particularly nephrosclerosis and diabetic nephropathy, which are causes of end-stage renal disease. This review focuses on the current understanding of the role of uromodulin from a biological, physiological, and pathological standpoint.

Analysis of potential biomarkers for diabetic kidney disease based on single-cell RNA-sequencing integrated with a single-cell sequencing assay for transposase-accessible chromatin.

Diabetic kidney disease (DKD) is a renal microvascular disease caused by hyperglycemia that involves metabolic remodeling, oxidative stress, inflammation, and other factors. The mechanism is complex and not fully unraveled. We performed an integrated single-cell sequencing assay for transposase-accessible chromatin (scATAC-seq) and single-cell RNA-sequencing (scRNA-seq) analyses of kidneys from db/db and db/m mice to identify differential open chromatin regions and gene expression, particularly in genes related to proximal tubular reabsorption and secretion. We identified 9,776 differentially expressed genes (DEGs) and 884 cell type-specific transcription factors (TFs) across 15 cell types. Glucose and lipid transporters, and TFs related to the circadian rhythm in the proximal tubules had significantly higher expression in db/db mice than in db/m mice (P<0.01). Crosstalk between podocytes and tubular cells in the proximal tubules was enhanced, and renal inflammation, oxidative stress, and fibrosis pathways were activated in db/db mice. Western blotting and immunohistochemical staining results showed that Wfdc2 expression in the urine and kidneys of DKD patients was higher than that in non-diabetic kidney disease (NDKD) controls. The revealed landscape of chromatin accessibility and transcriptional profiles in db/db mice provide insights into the pathological mechanism of DKD.

Signaling pathways upregulating glutathione­specific γ­glutamylcyclotransferase 1 by 3­(5'­hydroxymethyl­2'­furyl)­1­benzylindazole.

Glutathione­specific γ­glutamylcyclotransferase 1 (CHAC1), is an unfolded protein response­induced gene. Although it has been previously reported that CHAC1 transcription is regulated by activating transcription factor (ATF) 4, ATF3 and CCAAT/enhancer­binding protein ß (C/EBPß), the signaling pathways that regulate CHAC1 are largely unknown. It was revealed that 3­(5'­hydroxymethyl­2'­furyl)­1­benzylindazole (YC­1; PubChem ID: 5712), a nitric oxide­independent activator of soluble guanylyl cyclase (sGC), increases CHAC1 levels in cultured human kidney proximal tubular cells (HK­2). Therefore, in the present study, the signaling pathways that induce CHAC1 by YC­1 were investigated in HK­2 cells. YC­1 induced CHAC1 expression in a dose­ and time­dependent manner. KT5823, an inhibitor of cGMP­dependent protein kinase (PKG), partially inhibited CHAC1 upregulation, indicating that the sGC­cGMP­PKG pathway participates in CHAC1 regulation. These results also suggested that other signaling pathways are involved in the regulation of CHAC1. Since antibody array analysis showed the activation of p38, mTOR and Akt, the involvement of these factors was further investigated. Although LY294002 and KU0063794 (inhibitors of Akt and mTOR, respectively) inhibited YC­1­induced CHAC1 expression, SB203580 (an inhibitor of p38) did not. These results indicated that CHAC1 is regulated by the Akt­mTOR pathway. In addition, YC­1 induced endoplasmic reticulum (ER) stress, a regulator of CHAC1 induction. These findings suggested that CHAC1 is regulated by YC­1 through the sGC­cGMP­PKG, Akt­mTOR and ER stress pathways. The present study demonstrated that CHAC1 induction reduced the intracellular glutathione concentration, indicating that CHAC1 plays an important role in intracellular redox homeostasis in tubular cells.

Renal Proximal Tubular Epithelial Cells: From Harvesting to Use in Studies.

The kidneys are the body's main excretion organ with several additional functions, and the nephron represents their central structural unit. It is comprised of endothelial, mesangial, glomerular, and tubular epithelial cells, as well as podocytes. Treatment of acute kidney injury or chronic kidney disease (CKD) is complex due to broad etiopathogenic mechanisms and limited regeneration potential as kidney cells finish their differentiation after 34 weeks of gestation. Despite the ever-increasing prevalence of CKD, very limited treatment modalities are available. The medical community should therefore strive to improve existing treatments and develop new ones. Furthermore, polypharmacy is present in most CKD patients, while current pharmacologic study designs lack effectiveness in predicting potential drug-drug interactions and the resulting clinically relevant complications. An opportunity for addressing these issues lies in developing in vitro cell models based on patient-derived renal cells. Currently, several protocols have been described for isolating desired kidney cells, of which the most isolated are the proximal tubular epithelial cells. These play a significant role in water homeostasis, acid-base control, reabsorption of compounds, and secretion of xenobiotics and endogenous metabolites. When developing a protocol for the isolation and culture of such cells, one must focus on several steps. These include harvesting cells from biopsy specimens or after nephrectomies, using different digestion enzymes and culture mediums to facilitate the selective growth of only the desired cells. The literature reports several existing models, from simple 2D in vitro cultures to more complex ones created with bioengineering methods, such as kidney-on-a-chip models. While their creation and use depend on the target research, one should consider factors such as equipment, cost, and, even more importantly, source tissue quality and availability.