Long-term expandable mouse and human-induced nephron progenitor cells enable kidney organoid maturation and modeling of plasticity and disease.

Nephron progenitor cells (NPCs) self-renew and differentiate into nephrons, the functional units of the kidney. Here, manipulation of p38 and YAP activity allowed for long-term clonal expansion of primary mouse and human NPCs and induced NPCs (iNPCs) from human pluripotent stem cells (hPSCs). Molecular analyses demonstrated that cultured iNPCs closely resemble primary human NPCs. iNPCs generated nephron organoids with minimal off-target cell types and enhanced maturation of podocytes relative to published human kidney organoid protocols. Surprisingly, the NPC culture medium uncovered plasticity in human podocyte programs, enabling podocyte reprogramming to an NPC-like state. Scalability and ease of genome editing facilitated genome-wide CRISPR screening in NPC culture, uncovering genes associated with kidney development and disease. Further, NPC-directed modeling of autosomal-dominant polycystic kidney disease (ADPKD) identified a small-molecule inhibitor of cystogenesis. These findings highlight a broad application for the reported iNPC platform in the study of kidney development, disease, plasticity, and regeneration.

[The role of artificial intelligence in psychiatry].

In recent decades a global problem in mental health has been the increase in the relative proportion of patients who do not receive care, which is associated with loss of life years and deterioration in quality of life. The practical application of artificial intelligence (AI) can help in the fields of data analysis, diagnosis, therapy planning, among others in psychiatric care, thus reducing the human resource input. Today's artificial narrow intelligence (ANI), also known as weak AI, can recognise patterns and correlations in large data sets with the help of machine learning procedures and to make autonomous decisions while making its own refinements. The use of AI-based systems may be effective in the classification of mental health disorders, in disease prevention, in clinical diagnosis and treatment without human input, and finally, it can play a supporting role in many areas of data analysis (quality care assessment, research). A key area of diagnostics is the estimation of suicidal risk and the assessment of mood status using machine learning, which can be used to make predictions with high accuracy, by analysing written text or speech. By examining correlations within large data sets, advances in precision medicine could also be made, allowing more accurate prediction of medication. Psychotherapeutic programs using artificial intelligence are already available today, which can provide users with easily accessible help, mainly using cognitive therapy tools. In addition to its obvious benefits, the use of artificial intelligence also raises ethical and methodological questions, making its regulation a key issue for the future.

Canonical Wnt transcriptional complexes are essential for induction of nephrogenesis but not maintenance or proliferation of nephron progenitors.

Wnt regulated transcriptional programs are associated with both the maintenance of mammalian nephron progenitor cells (NPC) and their induction, initiating the process of nephrogenesis. How opposing transcriptional roles are regulated remain unclear. Using an in vitro model replicating in vivo events, we examined the requirement for canonical Wnt transcriptional complexes in NPC regulation. In canonical transcription, Lef/Tcf DNA binding proteins associate the transcriptional co-activator ß-catenin. Wnt signaling is readily substituted by CHIR99021, a small molecule antagonist of glycogen synthase kinase-3ß (GSK3ß). GSK3ß inhibition blocks Gskß-dependent turnover of ß-catenin, enabling formation of Lef/Tcf/ß-catenin transcriptional complexes, and enhancer-mediated transcriptional activation. Removal of ß-catenin activity from NPCs under cell expansion conditions (low CHIR) demonstrated a non-transcriptional role for ß-catenin in the CHIR-dependent proliferation of NPCs. In contrast, CHIR-mediated induction of nephrogenesis, on switching from low to high CHIR, was dependent on Lef/Tcf and ß-catenin transcriptional activity. These studies point to a non-transcriptional mechanism for ß-catenin in regulation of NPCs, and potentially other stem progenitor cell types. Further, analysis of the ß-catenin-directed transcriptional response provides new insight into induction of nephrogenesis. Summary Statement: The study provides a mechanistic understanding of Wnt/ ß-catenin activity in self-renewal and differentiation of mammalian nephron progenitors.

The C9ORF72 repeat expansion alters neurodevelopment.

Genetic mutations that cause adult-onset neurodegenerative diseases are often expressed during embryonic stages, but it is unclear whether they alter neurodevelopment and how this might influence disease onset. Here, we show that the most common cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS), a repeat expansion in C9ORF72, restricts neural stem cell proliferation and reduces cortical and thalamic size in utero. Surprisingly, a repeat expansion-derived dipeptide repeat protein (DPR) not known to reduce neuronal viability plays a key role in impairing neurodevelopment. Pharmacologically mimicking the effects of the repeat expansion on neurodevelopment increases susceptibility of C9ORF72 mice to motor defects. Thus, the C9ORF72 repeat expansion stunts development of the brain regions prominently affected in C9ORF72 FTD/ALS patients.

Modeling kidney development, disease, and plasticity with clonal expandable nephron progenitor cells and nephron organoids.

Nephron progenitor cells (NPCs) self-renew and differentiate into nephrons, the functional units of the kidney. Here we report manipulation of p38 and YAP activity creates a synthetic niche that allows the long-term clonal expansion of primary mouse and human NPCs, and induced NPCs (iNPCs) from human pluripotent stem cells. Cultured iNPCs resemble closely primary human NPCs, generating nephron organoids with abundant distal convoluted tubule cells, which are not observed in published kidney organoids. The synthetic niche reprograms differentiated nephron cells into NPC state, recapitulating the plasticity of developing nephron in vivo. Scalability and ease of genome-editing in the cultured NPCs allow for genome-wide CRISPR screening, identifying novel genes associated with kidney development and disease. A rapid, efficient, and scalable organoid model for polycystic kidney disease was derived directly from genome-edited NPCs, and validated in drug screen. These technological platforms have broad applications to kidney development, disease, plasticity, and regeneration.

[Current therapeutic and familial implications of the genetic background of prostate cancer]. / A prosztatarák genetikai hátterének aktuális terápiás és familiáris vonatkozásai.

Genetic testing for prostate cancer (PC) is becoming more widely used in the clinical routine, primarily due to the introduction of PARP inhibitors targeting genetically affected patients in their BRCA1/2 and other homologous recombination repair (HRR) genes. Simultaneously, the number of available therapies that are specifically targeting genetically defined PC subgroups is steadily increasing. As a result, the selection of treatment for PC patients is likely to require testing of multiple genes to enable more specific treatment sequences that consider the genetic characteristics of the tumor. Some of the mutations discovered by genetic testing may be hereditary, necessitating the use of germline testing from normal tissue, which is only permitted within the framework of clinical counseling. This change in PC care requires the collaboration by multiple specialists, including experts in molecular pathology, bioinformatics, biology, and genetic counseling. In this review, we aim to provide an overview on the currently relevant genetic alterations in PC for therapeutic purposes and their implications for familial testing.

[Current diagnostic and treatment strategies for urachal cancer]. / Az urachuscarcinoma aktuális diagnosztikai és kezelési lehetoségei.

Urachal carcinoma is a rare malignancy that is uniquely associated with the field of urology. Urachal carcinoma is mostly diagnosed in urological care centers due to its most frequently presenting symptom that is hematuria. Currently available diagnostic and therapeutic knowledge is solely based on case reports and single center series, while no prospective clinical studies are carried out due to the modest number of patients. These circumstances have made creating professional guidelines challenging, hence the treatment of urachal carcinoma is commonly based on individual clinical decisions. In this review, we summarize the epidemiology, diagnostic modalities, prognosis as well as local and systemic therapeutic approaches of urachal carcinoma. Furthermore, we aim to draw conclusions from this knowledge base that may guide clinical decision-making. Finally, we highlight some of the novel therapeutic strategies that hold the potential to improve urachal carcinoma patients' prognosis and quality of life. Orv Hetil. 2023; 164(16): 602-609.

Cadherin Adhesion Complexes Direct Cell Aggregation in the Epithelial Transition of Wnt-Induced Nephron Progenitor Cells.

In the developing mammalian kidney, nephron formation is initiated by a subset of nephron progenitor cells (NPCs). Wnt input activates a ß-catenin ( Ctnnb1 )-driven, transcriptional nephrogenic program. In conjunction, induced mesenchymal NPCs transition through a pre-tubular aggregate to an epithelial renal vesicle, the precursor for each nephron. How this critical mesenchymal-to-epithelial transition (MET) is regulated is unclear. In an in vitro mouse NPC culture model, activation of the Wnt pathway results in the aggregation of induced NPCs into closely-packed, cell clusters. Genetic removal of ß-catenin resulted in a failure of both Wnt pathway-directed transcriptional activation and the formation of aggregated cell clusters. Modulating extracellular Ca 2+ levels showed cell-cell contacts were Ca 2+ -dependent, suggesting a role for cadherin (Cdh)-directed cell adhesion. Molecular analysis identified Cdh2 , Cdh4 and Cdh11 in uninduced NPCs and the up-regulation of Cdh3 and Cdh4 accompanying the Wnt pathway-induced MET. Genetic removal of all four cadherins, and independent removal of α-catenin, which couples Cdh-ß-catenin membrane complexes to the actin cytoskeleton, abolished cell aggregation in response to Wnt pathway activation. However, the ß-catenin driven inductive transcriptional program was unaltered. Together with the accompanying paper (Bugacov et al ., submitted), these data demonstrate that distinct cellular activities of ß-catenin - transcriptional regulation and cell adhesion - combine in the mammalian kidney programs generating differentiated epithelial nephron precursors from mesenchymal nephron progenitors. Summary statement: Our study highlights the role of Wnt-ß-catenin pathway regulation of cadherin-mediated cell adhesion in the mesenchymal to epithelial transition of induced nephron progenitor cells.

The applicability of the Beck Depression Inventory and Hamilton Depression Scale in the automatic recognition of depression based on speech signal processing.

Depression is a growing problem worldwide, impacting on an increasing number of patients, and also affecting health systems and the global economy. The most common diagnostical rating scales of depression are self-reported or clinician-administered, which differ in the symptoms that they are sampling. Speech is a promising biomarker in the diagnostical assessment of depression, due to non-invasiveness and cost and time efficiency. In our study, we try to achieve a more accurate, sensitive model for determining depression based on speech processing. Regression and classification models were also developed using a machine learning method. During the research, we had access to a large speech database that includes speech samples from depressed and healthy subjects. The database contains the Beck Depression Inventory (BDI) score of each subject and the Hamilton Rating Scale for Depression (HAMD) score of 20% of the subjects. This fact provided an opportunity to compare the usefulness of BDI and HAMD for training models of automatic recognition of depression based on speech signal processing. We found that the estimated values of the acoustic model trained on BDI scores are closer to HAMD assessment than to the BDI scores, and the partial application of HAMD scores instead of BDI scores in training improves the accuracy of automatic recognition of depression.

A scalable organoid model of human autosomal dominant polycystic kidney disease for disease mechanism and drug discovery.

Human pluripotent stem-cell-derived organoids are models for human development and disease. We report a modified human kidney organoid system that generates thousands of similar organoids, each consisting of 1-2 nephron-like structures. Single-cell transcriptomic profiling and immunofluorescence validation highlighted patterned nephron-like structures utilizing similar pathways, with distinct morphogenesis, to human nephrogenesis. To examine this platform for therapeutic screening, the polycystic kidney disease genes PKD1 and PKD2 were inactivated by gene editing. PKD1 and PKD2 mutant models exhibited efficient and reproducible cyst formation. Cystic outgrowths could be propagated for months to centimeter-sized cysts. To shed new light on cystogenesis, 247 protein kinase inhibitors (PKIs) were screened in a live imaging assay identifying compounds blocking cyst formation but not overall organoid growth. Scaling and further development of the organoid platform will enable a broader capability for kidney disease modeling and high-throughput drug screens.

Reliability and validity of the Hungarian version of the Personality Inventory for DSM-5 (PID-5).

OBJECTIVES AND METHODS: In order to assess the internal consistency, fit indexes, test-retest reliability, and validity of the Personality Inventory for the DSM-5 (PID-5) and its associations with age, gender, and education, 471 non-clinical (69,6% female; mean age: 37,63) and 314 clinical participants (69,7% female, mean age: 37,41) were administered the Hungarian translation of the PID-5, as well as the SCL-90-R and the SCID-II Personality Questionnaire. RESULTS: We found that; (a) temporal consistency of the Hungarian PID-5 was confirmed by one-month test-retest reliability analysis, (b) validity of the PID-5 instrument is acceptable in the clinical and the non-clinical sample as well, based on significant correlations with SCID-II and SCL-90-R, (c) PID-5 facets' and domains' associations with gender, age, and level of education are in accordance with previous findings. CONCLUSION: These findings support that the Hungarian PID-5 is a reliable and valid instrument for both clinical and non-clinical populations.

Technical Skill Impacts the Success of Sequential Robotic Suturing Substeps.

Introduction: Robotic surgical performance, in particular suturing, has been associated with postoperative clinical outcomes. Suturing can be deconstructed into substep components (needle positioning, needle entry angle, needle driving, and needle withdrawal) allowing for the provision of more specific feedback while teaching suturing and more precision when evaluating suturing technical skill and prediction of clinical outcomes. This study evaluates if the technical skill required for particular substeps of the suturing process is associated with the execution of subsequent substeps in terms of technical skill, accuracy, and efficiency. Materials and Methods: Training and expert surgeons completed standardized sutures on the Mimic™ Flex virtual reality robotic simulator. Video recordings were deidentified, time annotated, and provided technical skill scores for each of the four suturing substeps. Hierarchical Poisson regression with generalized estimating equation was used to examine the association of technical skill rating categories between substeps. Results: Twenty-two surgeons completed 428 suturing attempts with 1669 individual technical skill assessments made. Technical skill scores between substeps of the suturing process were found to be significantly associated. When needle positioning was ideal, needle entry angle was associated with a significantly greater chance of being ideal (risk ratio [RR] = 1.12, p = 0.05). In addition, ideal needle entry angle and needle driving technical skill scores were each significantly associated with ideal needle withdrawal technical skill scores (RR = 1.27, p = 0.03; RR = 1.3, p = 0.03, respectively). Our study determined that ideal technical skill was associated with increased accuracy and efficiency of select substeps. Conclusions: Our study found significant associations in the technical skill required for completing substeps of suturing, demonstrating inter-relationships within the suturing process. Together with the known association between technical skill and clinical outcomes, training surgeons should focus on mastering not just the overall suturing process, but also each substep involved. Future machine learning efforts can better evaluate suturing, knowing that these inter-relationships exist.

Serial intravital imaging captures dynamic and functional endothelial remodeling with single-cell resolution.

Endothelial cells are important in the maintenance of healthy blood vessels and in the development of vascular diseases. However, the origin and dynamics of endothelial precursors and remodeling at the single-cell level have been difficult to study in vivo owing to technical limitations. Therefore, we aimed to develop a direct visual approach to track the fate and function of single endothelial cells over several days and weeks in the same vascular bed in vivo using multiphoton microscopy (MPM) of transgenic Cdh5-Confetti mice and the kidney glomerulus as a model. Individual cells of the vascular endothelial lineage were identified and tracked owing to their unique color combination, based on the random expression of cyan/green/yellow/red fluorescent proteins. Experimental hypertension, hyperglycemia, and laser-induced endothelial cell ablation rapidly increased the number of new glomerular endothelial cells that appeared in clusters of the same color, suggesting clonal cell remodeling by local precursors at the vascular pole. Furthermore, intravital MPM allowed the detection of distinct structural and functional alterations of proliferating endothelial cells. No circulating Cdh5-Confetti+ cells were found in the renal cortex. Moreover, the heart, lung, and kidneys showed more significant clonal endothelial cell expansion compared with the brain, pancreas, liver, and spleen. In summary, we have demonstrated that serial MPM of Cdh5-Confetti mice in vivo is a powerful technical advance to study endothelial remodeling and repair in the kidney and other organs under physiological and disease conditions.

Isoprostanes evoke contraction of the murine and human detrusor muscle via activation of the thromboxane prostanoid TP receptor and Rho kinase.

Local or systemic inflammation can severely impair urinary bladder functions and contribute to the development of voiding disorders in millions of people worldwide. Isoprostanes are inflammatory lipid mediators that are upregulated in the blood and urine by oxidative stress and may potentially induce detrusor overactivity. The aim of the present study was to investigate the effects and signal transduction of isoprostanes in human and murine urinary bladders in order to provide potential pharmacological targets in detrusor overactivity. Contraction force was measured with a myograph in murine and human urinary bladder smooth muscle (UBSM) ex vivo. Isoprostane 8-iso-PGE2 and 8-iso-PGF2α evoked dose-dependent contraction in the murine UBSM, which was abolished in mice deficient in the thromboxane prostanoid (TP) receptor. The responses remained unaltered after removal of the mucosa or incubation with tetrodotoxin. Smooth muscle-specific deletion of Gα12/13 protein or inhibition of Rho kinase by Y-27632 decreased the contractions. In Gαq/11-knockout mice, responses were reduced and in the presence of Y-27632 abolished completely. In human UBSM, the TP agonist U-46619 evoked dose-dependent contractions. Neither atropine nor the purinergic receptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid decreased the effect, indicating that TP receptors directly mediate detrusor muscle contraction. 8-iso-PGE2 and 8-iso-PGF2α evoked dose-dependent contraction in the human UBSM, and these responses were abolished by the TP antagonist SQ-29548 and were decreased by Y-27632. Our results indicate that isoprostanes evoke contraction in murine and human urinary bladders, an effect mediated by the TP receptor. The G12/13-Rho-Rho kinase pathway plays a significant role in mediating the contraction and therefore may be a potential therapeutic target in detrusor overactivity.NEW & NOTEWORTHY Voiding disorders affect millions of people worldwide. Inflammation can impair urinary bladder functions and contribute to the development of detrusor overactivity. The effects and signal transduction of inflammatory lipid mediator isoprostanes were studied in human and murine urinary bladders ex vivo. We found that isoprostanes evoke contraction, an effect mediated by thromboxane prostanoid receptors. The G12/13-Rho-Rho kinase signaling pathway plays a significant role in mediating the contraction and therefore may be a potential therapeutic target.

NK2 receptor-mediated detrusor muscle contraction involves Gq/11-dependent activation of voltage-dependent Ca2+ channels and the RhoA-Rho kinase pathway.

Tachykinins (TKs) are involved in both the physiological regulation of urinary bladder functions and development of overactive bladder syndrome. The aim of the present study was to investigate the signal transduction pathways of TKs in the detrusor muscle to provide potential pharmacological targets for the treatment of bladder dysfunctions related to enhanced TK production. Contraction force, intracellular Ca2+ concentration, and RhoA activity were measured in the mouse urinary bladder smooth muscle (UBSM). TKs and the NK2 receptor (NK2R)-specific agonist [ß-Ala8]-NKA(4-10) evoked contraction, which was inhibited by the NKR2 antagonist MEN10376. In Gαq/11-deficient mice, [ß-Ala8]-NKA(4-10)-induced contraction and the intracellular Ca2+ concentration increase were abolished. Although Gq/11 proteins are linked principally to phospholipase Cß and inositol trisphosphate-mediated Ca2+ release from intracellular stores, we found that phospholipase Cß inhibition and sarcoplasmic reticulum Ca2+ depletion failed to have any effect on contraction induced by [ß-Ala8]-NKA(4-10). In contrast, lack of extracellular Ca2+ or blockade of voltage-dependent Ca2+ channels (VDCCs) suppressed contraction. Furthermore, [ß-Ala8]-NKA(4-10) increased RhoA activity in the UBSM in a Gq/11-dependent manner and inhibition of Rho kinase with Y-27632 decreased contraction force, whereas the combination of Y-27632 with either VDCC blockade or depletion of extracellular Ca2+ resulted in complete inhibition of [ß-Ala8]-NKA(4-10)-induced contractions. In summary, our results indicate that NK2Rs are linked exclusively to Gq/11 proteins in the UBSM and that the intracellular signaling involves the simultaneous activation of VDCC and the RhoA-Rho kinase pathway. These findings may help to identify potential therapeutic targets of bladder dysfunctions related to upregulation of TKs.

In Vivo Developmental Trajectories of Human Podocyte Inform In Vitro Differentiation of Pluripotent Stem Cell-Derived Podocytes.

The renal corpuscle of the kidney comprises a glomerular vasculature embraced by podocytes and supported by mesangial myofibroblasts, which ensure plasma filtration at the podocyte-generated slit diaphragm. With a spectrum of podocyte-expressed gene mutations causing chronic disease, an enhanced understanding of podocyte development and function to create relevant in vitro podocyte models is a clinical imperative. To characterize podocyte development, scRNA-seq was performed on human fetal kidneys, identifying distinct transcriptional signatures accompanying the differentiation of functional podocytes from progenitors. Interestingly, organoid-generated podocytes exhibited highly similar, progressive transcriptional profiles despite an absence of the vasculature, although abnormal gene expression was pinpointed in late podocytes. On transplantation into mice, organoid-derived podocytes recruited the host vasculature and partially corrected transcriptional profiles. Thus, human podocyte development is mostly intrinsically regulated and vascular interactions refine maturation. These studies support the application of organoid-derived podocytes to model disease and to restore or replace normal kidney functions.

Advances in Renal Cell Imaging.

A great variety of cell imaging technologies are used routinely every day for the investigation of kidney cell types in applications ranging from basic science research to drug development and pharmacology, clinical nephrology, and pathology. Quantitative visualization of the identity, density, and fate of both resident and nonresident cells in the kidney, and imaging-based analysis of their altered function, (patho)biology, metabolism, and signaling in disease conditions, can help to better define pathomechanism-based disease subgroups, identify critical cells and structures that play a role in the pathogenesis, critically needed biomarkers of disease progression, and cell and molecular pathways as targets for novel therapies. Overall, renal cell imaging has great potential for improving the precision of diagnostic and treatment paradigms for individual acute kidney injury or chronic kidney disease patients or patient populations. This review highlights and provides examples for some of the recently developed renal cell optical imaging approaches, mainly intravital multiphoton fluorescence microscopy, and the new knowledge they provide for our better understanding of renal pathologies.