BACKGROUND:Chronic kidney disease (CKD) can be asymptomatic for many years and is often diagnosed late. Given the availability of new treatments, the early identification of relevant findings from screening of the kidney markers estimated glomerular filtration rate (eGFR) and albuminuria in the general population is becoming increasingly important. METHODS:In the NAKO study, self-reported medical diagnoses of kidney disease in 195 182 participants were compared with relevant findings from screening biomarkers (eGFR < 60 mL/min/1.73 m² and albuminuria). For the purpose of comparison, various equations for assessing kidney function were evaluated as well. RESULTS:2% of the participants reported having received a medical diagnosis of kidney disease, and 2% had an eGFR below 60 mL/min/1.73 m². There was, however, little overlap between these two groups: more than 80% of participants with an eGFR between 30 and 59 mL/min/1,73 m²) did not report any diagnosis of kidney disease. The additional inclusion of data on albuminuria did not materially affect this discrepancy: 6213 persons (17.5% of the cohort) with an abnormal eGFR or urinary albumin-to-creatinine ratio (UACR) did not report any diagnosis of kidney disease. Even among participants whose eGFR was in the range of 30-59 mL/min/1.73 m² and whose UACR was above 300 mg/g, less than half reported having a medically diagnosed kidney disease. CONCLUSION:These findings indicate a low level of awareness regarding the possible presence of CKD in the general population. Many people with abnormal screening findings needing further investigation due to their potential clinical relevance are unaware that they might be suffering from a kidney disease. As more effective treatments for kidney disease are now available, these findings indicate a need for structured screening and evaluation strategies to promote kidney health.

Focal segmental glomerulosclerosis (FSGS) is a major cause of nephrotic syndrome and progression to end-stage renal disease, yet its molecular pathogenesis remains still incompletely defined. While transcriptional alterations in podocytes have been extensively characterized, the contribution of post-transcriptional regulatory mechanisms is poorly understood. Here, we combined a zebrafish podocyte-specific injury model with glomerulus-resolved transcriptomic profiling to dissect RNA regulatory alterations during FSGS progression. Integrated analyses of bulk RNA sequencing, small RNA profiling, and alternative splicing revealed pronounced, time-dependent remodeling of the glomerular transcriptome. We demonstrate that podocyte injury is associated with loss of key podocyte-specific proteins, activation of inflammatory pathways, remodeling of the extracellular matrix, and altered microRNA expression, such as miR-21 and miR-193. Moreover, we found that alternative splicing influences key podocyte gene expression, affecting genes critical for slit diaphragm integrity, actin cytoskeleton organization, and glomerular basement membrane stability. Isoform analyses identified FSGS-associated isoform switches in SRSF3 and EPB41L5. Importantly, these changes were also evident in glomeruli from FSGS patients, demonstrating that the zebrafish model recapitulates key molecular features of human disease and highlighting alternative splicing as a central regulatory mechanism in FSGS. Post-transcriptional regulations, such as alternative splicing and microRNA dysregulation, are identified as central and underappreciated processes in injured podocytes in focal segmental glomerulosclerosis (FSGS), with disease-associated isoform switches in SRSF3 and EPB41L5. Post-transcriptional regulations, such as alternative splicing and microRNA dysregulation, are identified as central and underappreciated processes in injured podocytes in focal segmental glomerulosclerosis (FSGS), with disease-associated isoform switches in SRSF3 and EPB41L5.

Background:Recent studies have indicated that body composition may affect kidney function. The aim of this study was to examine the sex- and age-related association between body composition markers and the estimated glomerular filtration rate (eGFR). Methods:Analyses were based on data from the population-based 'Study of Health in Pomerania' (SHIP), including data from 4211 individuals enrolled in the SHIP Trend-0 cohort. Body composition markers included measurements from classic anthropometry (body mass index, waist-to-hip ratio), body impedance analysis and magnetic resonance imaging (subcutaneous, hepatic and visceral fat mass). The eGFR was estimated using creatinine- and Cystatin C-based formulas. Cross-sectional analysis was performed using linear regression models adjusted for confounding variables. Results:Generally, higher values of body composition markers were associated with lower eGFR levels for all formulas, while higher relative fat-free mass was associated with higher eGFR. The effects were more pronounced in women compared to men. Regarding age dependency, increased values of body or fat mass were associated with higher eGFR in younger age groups, but with lower eGFR in older age groups. In contrast, higher relative fat-free mass was associated with increased eGFR in older age groups. Conclusions:Increased body and fat mass, regardless of distribution, and a lower percentage of muscle mass are associated with lower eGFR especially in women and older individuals.

Background Focal Segmental Glomerulosclerosis (FSGS) is a severe kidney disorder with complex and not yet fully understood pathogenesis. Alternative splicing (AS) – the generation of distinct protein isoforms from the same gene – might play a critical role by the regulation of gene functions and disease development. Methods To investigate the role of AS in FSGS, we used a zebrafish model, which mimics key human FSGS features, including foot process effacement, matrix accumulation, podocyte detachment and parietal epithelial cell activation. We performed total RNA sequencing of isolated zebrafish glomeruli and whole larvae, followed by integrative bioinformatic analysis to identify AS events and regulatory miRNAs. Results Our data revealed a downregulation of essential podocyte genes ( nphs1, nphs2, podxl, wt1 ) and an inhibition of pathways associated with nephron development and cytoskeletal organization. We also observed increased expression of the transcription factor stat3 and disease-associated miRNAs such as miR-21 and miR-193. AS analysis identified approximately ∼7,000 splicing events, primarily exon skipping (∼80%), affecting genes such as nphs1 , magi2 , and ptpro . A total of 136 and 612 alternatively spliced genes were found at 5 and 6 days post-fertilization (dpf), respectively. Isoform switch analysis uncovered 70 genes affected by AS in FSGS, including epb41l5 (linked to podocyte adhesion), fgfr1a (fibroblast growth signaling), and members of the SRSF splicing factor family (e.g., srsf3a ). Conclusions These findings emphasize the importance of transcriptional and post-transcriptional regulation, including AS, in FSGS pathogenesis. Furthermore, they support the zebrafish model as a valuable system for identifying novel mechanisms and potential therapeutic targets for kidney diseases. ### Competing Interest Statement The authors have declared no competing interest.

Key PointsMechanical stretch induced over 3000 alternative splicing events in podocytes, affecting gene expression and protein abundance.Seventeen genes showed consistent splicing events across multiple analysis tools, with key isoform changes.Shroom3 and Myl6 underwent isoform switches under mechanical stretch, altering the C-terminal sequence and interaction properties of Myl6.BackgroundAlterations in pre-mRNA splicing are crucial to the pathophysiology of various diseases. However, the effects of alternative splicing of mRNA on podocytes in hypertensive nephropathy are still unknown. The Sys_CARE project aimed to identify alternative splicing events involved in the development and progression of glomerular hypertension.MethodsMurine podocytes were exposed to mechanical stretch, after which proteins and mRNA were analyzed by proteomics, RNA sequencing, and several bioinformatic alternative splicing tools.ResultsUsing transcriptomic and proteomic analysis, we identified significant changes in gene expression and protein abundance because of mechanical stretch. RNA-Seq identified over 3000 alternative spliced genes after mechanical stretch, including all types of alternative splicing events. Among these, 17 genes exhibited an alternative splicing event across four different splicing analysis tools. From this group, we focused on Myl6, a component of the myosin protein complex, and Shroom3, an actin-binding protein essential for podocyte function. We identified two Shroom3 isoforms with significant expression changes under mechanical stretch, which was validated by quantitative RT-PCR and in situ hybridization. In addition, we observed an expression switch of two Myl6 isoforms after mechanical stretch, accompanied by an alteration in the C-terminal amino acid sequence.ConclusionsA comprehensive RNA-Seq analysis of mechanically stretched podocytes identified novel potential podocyte-specific biomarkers and highlighted significant alternative splicing events, notably in the mRNA of Shroom3 and Myl6.

X-chromosomal genetic variants are understudied but can yield valuable insights into sexually dimorphic human traits and diseases. We performed a sex-stratified cross-ancestry X-chromosome-wide association meta-analysis of seven kidney-related traits ( n = 908,697), identifying 23 loci genome-wide significantly associated with two of the traits: 7 for uric acid and 16 for estimated glomerular filtration rate (eGFR), including four novel eGFR loci containing the functionally plausible prioritized genes ACSL4 , CLDN2 , TSPAN6 and the female-specific DRP2 . Further, we identified five novel sex-interactions, comprising male-specific effects at FAM9B and AR/EDA2R , and three sex-differential findings with larger genetic effect sizes in males at DCAF12L1 and MST4 and larger effect sizes in females at HPRT1 . All prioritized genes in loci showing significant sex-interactions were located next to androgen response elements (ARE). Five ARE genes showed sex-differential expressions. This study contributes new insights into sex-dimorphisms of kidney traits along with new prioritized gene targets for further molecular research.

Key Points Computational fluid dynamics were applied to estimate the shear stress challenge to the filtration barrier during glomerular filtration in rats.Shear forces were especially relevant in pathologic situations where they contribute to the loss of viable podocytes. Background The flow dynamic forces during glomerular filtration challenging the fixation of podocytes to the glomerular basement membrane (GBM) are insufficiently understood. Methods Numerical flow simulations were used to estimate these forces in the rat kidney. Simulations were run with a three-dimensional (3D) model of the slit diaphragm as a zipper structure according to Rodewald and Karnovsky. The GBM was modeled as a porous medium. Results Filtrate flow exerted a mean wall shear stress of 39 Pa with a maximum of 152 Pa on the plasma membrane of foot processes and up to 250 Pa on internal surfaces of the slit diaphragm. The slit diaphragm accounted for 25% of the hydrodynamic resistance of the glomerular filtration barrier. Based on the results of the 3D model, we developed a two-dimensional (2D) model that allowed us to perform extensive parameter variations. Reducing the filtration slit width from 40 to 30 nm almost doubled wall shear stress. Furthermore, increasing filtrate flow velocity by 50% increased wall shear stress by 47%. When increasing the viscous resistance of the slit diaphragm, the pressure drop across the slit diaphragm increased to intolerably high values. A lower viscous resistance of the slit diaphragm than that of the GBM accounted for a gradual pressure decline along the filtration barrier. The subpodocyte space tempered these challenges in circumscribed areas of filtration surface but had only a marginal impact on overall forces. Conclusions The filtration barrier experiences high levels of shear and pressure stress accounting for the detachment of injured but viable podocytes from the GBM—a hallmark in many glomerular diseases.

Alterations in pre-mRNA splicing play an important role in disease pathophysiology. However, the role of alternative splicing (AS) for podocytes in hypertensive nephropathy (HN) has not been investigated. The purpose of the Sys_CARE project was to identify AS events that play a role in the development and progression of HN. For this, murine podocytes were exposed to mechanical stretch, after which proteins and mRNA were analyzed by proteomics, RNA-Seq and several bioinformatic AS tools. Based on transcriptomics and proteomics analysis we could observe significant changes in gene expression and abundance of proteins under mechanical stretch compared to unstretched conditions. By RNA-Seq, we identified over 1000 different splicing events including all types of AS events. We identified 17 genes that showed an AS event in four different splicing analysis tools. We focused on Myl6 , a component of the myosin protein complex, and Shroom3 , an actin-binding protein crucial for podocyte function. We found two Shroom3 isoforms that showed significant changes in expression upon mechanical stretch, which was verified by qRT-PCR and in situ hybridization. Furthermore, we observed an expression switch of two Myl6 isoforms after mechanical stretch. This switch is accompanied by a change in a C-terminally located amino acid sequence. In summary, mechanical stretch of cultured podocytes is an excellent model to simulate hypertensive nephropathy. In depth RNA-Seq analysis disclosed alternative splicing events, such as in Shroom3 and Myl6 , which may play a crucial role in the pathophysiology of hypertension-induced nephropathy. ### Competing Interest Statement The authors have declared no competing interest.

Abstract Background and Aims High blood pressure is considered one of the most important risk factors leading to chronic kidney disease. The pathogenesis of hypertension-induced damage to podocytes remains still elusive. Since recent reports have shown that alternative splicing (AS) has a significant impact on disease progression, we investigated alternative splicing of mechanically stretched podocytes, a well-established model for glomerular hypertension, in the Sys_CARE (Systems Medicine Investigation of AS in Cardiac and Renal Diseases) project. Our results provide new insights into the molecular mechanisms that might drive the progression of hypertensive nephropathy. Method Differentiated murine podocytes were exposed to mechanical stretch for 3 days under low and high stretch conditions. Subsequently, mRNA and proteins were analyzed by RNA-Seq and LC-MS/MS, respectively. Enrichment analysis identified transcripts which were classified according to the related biological processes, molecular function, and cellular components. Splicing and transcript expression were evaluated with bioinformatical tools such as MAJIQ, Multivariate Analysis of Transcript Splicing (rMATS), leafcutter, Whippet and IsoformSwitchAnalyzeR. Results Transcriptome analyses of mechanically stretched podocytes by RNA sequencing showed that mechanical stress leads to a high number of differentially expressed genes (1323 with log2 fc ± 1) compared to unstretched podocytes. Detailed gene set enrichment analysis for GO-Terms showed an enrichment among up-regulated transcripts related to mRNA processing and RNA splicing. In contrast, most transcripts with decreased expression upon stretch are associated with cytoskeleton function. To identify AS events, different AS tools were used. We found a wide variety of splice events. The most frequent event was exon skipping (between 63-82% of all AS events), followed by intron retention and alternative 5′ or 3′ splice site. 290 alternatively spliced genes were detected after mechanical stretch by three different splice analysis tools (rMATS, leafcutter, Whippet). Out of these candidates, the IsoformSwitchAnalyzeR identified 17 genes exhibiting significant isoform switches. To prioritize the candidates, we performed a screening that included only those genes that were identified by proteomic analysis and that are expressed in podocytes in vivo or showed an altered expression pattern in glomerular disease. This screening led to the two candidates Shroom3 and Myl6. Shroom3, an actin-binding protein, is crucial for podocyte morphology and function. Genetic variants of Shroom3 have been linked with chronic kidney disease by GWAS. Here we found that two of four Shroom3 isoforms showed significant expression changes due to mechanical stretch. Isoform 2 which showed the highest expression in cultured podocytes, significantly decreased, but the shorter isoform X1 was significantly up-regulated after mechanical stretch. Both changes were verified by qRT-PCR as well as by in situ hybridization in mouse tissue. Myosin light chain 6 (Myl6), a component of the myosin motor protein complex, has two isoforms which are detected in cultured podocytes as well as in vivo. We found an expression switch from isoform 1 to 2 due to mechanical stretch. Our analysis showed that this is accompanied by a change of the amino acid sequence AFVRHILS to ELVRMVLN, sequences that are well conserved in mice, rat, and human. The analysis of the protein structure by FoldX revealed that this change in the amino acid sequence could lead to an impairment of the binding of the motor protein complex to actin and might therefore influence the functionality of the actin cytoskeleton, which is highly important for proper podocyte foot process morphology. Conclusion Mechanical stretch of cultured mouse podocytes, which is an excellent model to simulate hypertensive nephropathy, leads to alterations in isoform expression due to alternative splicing. These changes in the expression of isoforms, such as Shroom 3 and Myl6, have the potential to play a key role in the pathophysiology of hypertension-induced nephropathy.

Reduced glomerular filtration rate (GFR) can progress to kidney failure. Risk factors include genetics and diabetes mellitus (DM), but little is known about their interaction. We conducted genome-wide association meta-analyses for estimated GFR based on serum creatinine (eGFR), separately for individuals with or without DM (n DM = 178,691, n noDM = 1,296,113). Our genome-wide searches identified (i) seven eGFR loci with significant DM/noDM-difference, (ii) four additional novel loci with suggestive difference and (iii) 28 further novel loci (including CUBN ) by allowing for potential difference. GWAS on eGFR among DM individuals identified 2 known and 27 potentially responsible loci for diabetic kidney disease. Gene prioritization highlighted 18 genes that may inform reno-protective drug development. We highlight the existence of DM-only and noDM-only effects, which can inform about the target group, if respective genes are advanced as drug targets. Largely shared effects suggest that most drug interventions to alter eGFR should be effective in DM and noDM.

Chronic kidney disease (CKD) is a major public health burden affecting more than 500 million people worldwide. Podocytopathies are the main cause for the majority of CKD cases due to pathogenic morphological as well as molecular biological alterations of postmitotic podocytes. Podocyte de-differentiation is associated with foot process effacement subsequently leading to proteinuria. Since currently no curative drugs are available, high throughput screening methods using a small number of animals are a promising and essential tool to identify potential drugs against CKD in the near future. Our study presents the implementation of the already established mouse GlomAssay as a semi-automated high-throughput screening method-shGlomAssay-allowing the analysis of several hundreds of FDA-verified compounds in combination with downstream pathway analysis like transcriptomic and proteomic analyses from the same samples, using a small number of animals. In an initial prescreening we have identified vitamin D3 and its analog calcipotriol to be protective on podocytes. Furthermore, by using RT-qPCR, Western blot, and RNA sequencing, we found that mRNA and protein expression of nephrin, the vitamin D receptor and specific podocyte markers were significantly up-regulated due to vitamin D3- and calcipotriol-treatment. In contrast, kidney injury markers were significantly down-regulated. Additionally, we found that vitamin D3 and calcipotriol have had neither influence on the expression of the miR-21 and miR-30a nor on miR-125a/b, a miRNA described to regulate the vitamin D receptor. In summary, we advanced the established mouse GlomAssay to a semi-automated high-throughput assay and combined it with downstream analysis techniques by using only a minimum number of animals. Hereby, we identified the vitamin D signaling pathway as podocyte protective and to be counteracting their de-differentiation.

Given the increasing prevalence of chronic kidney disease (CKD) and its impact on health care, it is important to better understand the multiple factors influencing health-related quality of life (HRQOL), particularly since they have been shown to affect CKD outcomes. Determinants of HRQOL as measured by the validated Kidney Disease Quality of Life questionnaire (KDQOL) and the Patient Health Questionnaire depression screener (PHQ-9) were assessed in a routine CKD patient sample, the Greifswald Approach to Individualized Medicine (GANI_MED) renal cohort (N = 160), including a wide range of self-reported data, sociodemographic and laboratory measures. Compared to the general population, CKD patients had lower HRQOL indices. Dialysis was associated with (1) low levels of physical functioning, (2) increased impairments by symptoms and problems, and (3) more effects and burden of kidney disease. HRQOL is seriously affected in CKD patients. However, impairments were found irrespective of eGFR decline and albuminuria. Rather, the comorbid conditions of depression and diabetes predicted a lower HRQOL (physical component score). Further studies should address whether recognizing and treating depression may not only improve HRQOL but also promote survival and lower hospitalization rates of CKD patients.

Estimated glomerular filtration rate (eGFR) reflects kidney function. Progressive eGFR-decline can lead to kidney failure, necessitating dialysis or transplantation. Hundreds of loci from genome-wide association studies (GWAS) for eGFR help explain population cross section variability. Since the contribution of these or other loci to eGFR-decline remains largely unknown, we derived GWAS for annual eGFR-decline and meta-analyzed 62 longitudinal studies with eGFR assessed twice over time in all 343,339 individuals and in high-risk groups. We also explored different covariate adjustment. Twelve genome-wide significant independent variants for eGFR-decline unadjusted or adjusted for eGFR-baseline (11 novel, one known for this phenotype), including nine variants robustly associated across models were identified. All loci for eGFR-decline were known for cross-sectional eGFR and thus distinguished a subgroup of eGFR loci. Seven of the nine variants showed variant-by-age interaction on eGFR cross section (further about 350,000 individuals), which linked genetic associations for eGFR-decline with age-dependency of genetic cross-section associations. Clinically important were two to four-fold greater genetic effects on eGFR-decline in high-risk subgroups. Five variants associated also with chronic kidney disease progression mapped to genes with functional in-silico evidence (UMOD, SPATA7, GALNTL5, TPPP). An unfavorable versus favorable nine-variant genetic profile showed increased risk odds ratios of 1.35 for kidney failure (95% confidence intervals 1.03-1.77) and 1.27 for acute kidney injury (95% confidence intervals 1.08-1.50) in over 2000 cases each, with matched controls). Thus, we provide a large data resource, genetic loci, and prioritized genes for kidney function decline, which help inform drug development pipelines revealing important insights into the age-dependency of kidney function genetics.

The majority of kidney diseases arise from the loss of podocytes and from morphological changes of their highly complex foot process architecture, which inevitably leads to a reduced kidney filtration and total loss of kidney function. It could have been shown that microRNAs (miRs) play a pivotal role in the pathogenesis of podocyte-associated kidney diseases. Due to their fully functioning pronephric kidney, larval zebrafish have become a popular vertebrate model, to study kidney diseases in vivo . Unfortunately, there is no consensus about a proper normalization strategy of RT-qPCR-based miRNA expression data in zebrafish. In this study we analyzed 9 preselected candidates dre-miR-92a-3p, dre-miR-206-3p, dre-miR-99-1, dre-miR-92b-3p, dre-miR-363-3p, dre-let-7e, dre-miR-454a, dre-miR-30c-5p, dre-miR-126a-5p for their capability as endogenous reference genes in zebrafish experiments. Expression levels of potential candidates were measured in 3 different zebrafish strains, different developmental stages, and in different kidney disease models by RT-qPCR. Expression values were analyzed with NormFinder, BestKeeper, GeNorm, and DeltaCt and were tested for inter-group differences. All candidates show an abundant expression throughout all samples and relatively high stability. The most stable candidate without significant inter-group differences was dre-miR-92b-3p making it a suitable endogenous reference gene for RT-qPCR-based miR expression zebrafish studies.

Under healthy conditions, foot processes of neighbouring podocytes are interdigitating and connected by an electron-dense slit diaphragm. Besides slit diaphragm proteins, typical adherens junction proteins are also found to be expressed at this cell-cell junction. It is therefore considered as a highly specialized type of adherens junction. During podocyte injury, podocyte foot processes lose their characteristic 3D structure and the filtration slits typical meandering structure gets linearized. It is still under debate how this change of structure leads to the phenomenon of proteinuria. Using super-resolution 3D-structured illumination microscopy, we observed a spatially restricted up-regulation of the tight junction protein claudin-5 (CLDN5) in areas where podocyte processes of patients suffering from minimal change disease (MCD), focal and segmental glomerulosclerosis (FSGS) as well as in murine nephrotoxic serum (NTS) nephritis and uninephrectomy DOCA-salt hypertension models, were locally injured. CLDN5/nephrin ratios in human glomerulopathies and NTS-treated mice were significantly higher compared to controls. In patients, the CLDN5/nephrin ratio is significantly correlated with the filtration slit density as a foot process effacement marker, confirming a direct association of local CLDN5 up-regulation in injured foot processes. Moreover, CLDN5 up-regulation was observed in some areas of high filtration slit density, suggesting that CLND5 up-regulation preceded the changes of foot processes. Therefore, CLDN5 could serve as a biomarker predicting early foot process effacement.

Abstract Background and Aims The zebrafish (Danio rerio) is a powerful animal model to study glomerular morphology and the function of the permselectivity of the glomerular filtration barrier. Since zebrafish larvae develop quickly and can be bred to become transparent, in vivo observation of these animals is possible. At 48 hours post fertilization, zebrafish larvae develop a single glomerulus which is attached to a pair of tubules. Like in mammals, the glomerular filtration barrier consists of a fenestrated endothelium, the glomerular basement membrane and interdigitating podocyte foot processes bridged by a slit diaphragm. By using genetically modified zebrafish strains with fluorescently labeled podocytes, it is possible to study alterations of the glomerulus during the development of renal disease like focal segmental glomerulosclerosis (FSGS) directly in vivo. FSGS is characterized by podocyte loss, the effacement of their foot processes as well as scarring of the glomerulus. To study FSGS in zebrafish larvae, we induced podocyte detachment by the use of a zebrafish strain expressing the enzyme nitroreductase converting metronidazole into a toxic substance specifically in podocytes. The aim of our study was to collect glomeruli for the identification of mRNAs as well as miRNAs by RNA_Seq that are up- and down-regulated in the glomeruli of this FSGS-like disease model. Method The transgenic zebrafish strain Cherry (Tg(nphs2:GAL4); Tg(UAS:Eco.nfsB-mCherry); mitfaw2/w2; mpv17a9/a9) which expresses the prokaryotic enzyme nitroreductase (NTR) fused to mCherry, a red fluorescent protein, under the control of the podocyte-specific podocin (nphs2) promoter in a transparent zebrafish strain, was utilized. After addition of metronidazole (MTZ) into the tank water, MTZ is converted into a cytotoxin by NTR leading to dose-dependent apoptosis exclusively in podocytes. Cherry larvae were treated at 4 days post fertilization (dpf) for 48 h with 80 µM MTZ. MTZ-treated and control larvae were homogenized at 6 dpf. The cell suspension was diluted, and red-fluorescent glomeruli were collected using a micropipette and a microscope. Total RNA was isolated, and integrity was checked by a Bioanalyzer. Libraries were generated with a MACE kit and True Quant small RNA seq kit by GenXPro. Constructs were amplified by PCR and sequenced on an Illumina Hiseq 2000. Normalization and statistical analysis for differential gene expression were done using DESeq2. Results Zebrafish larvae showed severe whole-body edema, proteinuria, loss of podocytes and an increased mortality rate after MTZ-treatment. The glomerular histology resembled mammalian FSGS. We found that only the RNA of manually collected glomeruli had an excellent quality. Using RNA_Seq, we identified a total of 16941 genes. DESeq2 analysis showed 494 up-regulated and 473 down-regulated genes. Gene ontology (GO) enrichment analysis of up-regulated genes revealed a total of 167 that are significantly enriched in GO terms (e.g. metabolic processes, immune response and ion transport). Down-regulated genes were enriched in 14 GO terms and most of them are linked to normal glomerular function and the slit diaphragm. DESeq2 analysis identified 200 miRNAs of 777 small RNAs. Some of these miRNA are already described to be regulated in different glomerular diseases like FSGS, lupus nephritis, IgA nephropathy and diabetic nephropathy. Conclusion We analyzed isolated glomeruli from transgenic zebrafish larvae that developed a FSGS-like disease. By sequencing, we have found mRNAs and miRNAs that were significantly regulated after the onset of disease. Detailed knowledge of these mRNAs and miRNA-based gene regulation will help to uncover the pathomechanism as well as to develop therapeutics for the treatment of FSGS.