Kidney organoids: accurate models or fortunate accidents.

There are now many reports of human kidney organoids generated via the directed differentiation of human pluripotent stem cells (PSCs) based on an existing understanding of mammalian kidney organogenesis. Such kidney organoids potentially represent tractable tools for the study of normal human development and disease with improvements in scale, structure, and functional maturation potentially providing future options for renal regeneration. The utility of such organotypic models, however, will ultimately be determined by their developmental accuracy. While initially inferred from mouse models, recent transcriptional analyses of human fetal kidney have provided greater insight into nephrogenesis. In this review, we discuss how well human kidney organoids model the human fetal kidney and how the remaining differences challenge their utility.

The BCL-2 family member BID plays a role during embryonic development in addition to its BH3-only protein function by acting in parallel to BAX, BAK and BOK.

The intrinsic apoptosis pathway, regulated by the BCL-2 protein family, is essential for embryonic development. Using mice lacking all known apoptosis effectors, BAX, BAK and BOK, we have previously defined the processes during development that require apoptosis. Rare Bok-/- Bax-/- Bak-/- triple knockout (TKO) mice developed to adulthood and several tissues that were thought to require apoptosis during development appeared normal. This raises the question if all apoptosis had been abolished in the TKO mice or if other BCL-2 family members could act as effectors of apoptosis. Here, we investigated the role of BID, generally considered to link the extrinsic and intrinsic apoptosis pathways, acting as a BH3-only protein initiating apoptosis upstream of BAX and BAK. We found that Bok-/- Bax-/- Bak-/- Bid-/- quadruple knockout (QKO) mice have additional developmental anomalies compared to TKO mice, consistent with a role of BID, not only upstream but also in parallel to BAX, BAK and BOK. Mitochondrial experiments identified a small cytochrome c-releasing activity of full-length BID. Collectively, these findings suggest a new effector role for BID in the intrinsic apoptosis pathway.

Kidney organoids reveal redundancy in viral entry pathways during ACE2-dependent SARS-CoV-2 infection.

With a high incidence of acute kidney injury among hospitalized COVID-19 patients, considerable attention has been focussed on whether SARS-CoV-2 specifically targets kidney cells to directly impact renal function, or whether renal damage is primarily an indirect outcome. To date, several studies have utilized kidney organoids to understand the pathogenesis of COVID-19, revealing the ability for SARS-CoV-2 to predominantly infect cells of the proximal tubule (PT), with reduced infectivity following administration of soluble ACE2. However, the immaturity of standard human kidney organoids represents a significant hurdle, leaving the preferred SARS-CoV-2 processing pathway, existence of alternate viral receptors, and the effect of common hypertensive medications on the expression of ACE2 in the context of SARS-CoV-2 exposure incompletely understood. Utilizing a novel kidney organoid model with enhanced PT maturity, genetic- and drug-mediated inhibition of viral entry and processing factors confirmed the requirement for ACE2 for SARS-CoV-2 entry but showed that the virus can utilize dual viral spike protein processing pathways downstream of ACE2 receptor binding. These include TMPRSS- and CTSL/CTSB-mediated non-endosomal and endocytic pathways, with TMPRSS10 likely playing a more significant role in the non-endosomal pathway in renal cells than TMPRSS2. Finally, treatment with the antihypertensive ACE inhibitor, lisinopril, showed negligible impact on receptor expression or susceptibility of renal cells to infection. This study represents the first in-depth characterization of viral entry in stem cell-derived human kidney organoids with enhanced PTs, providing deeper insight into the renal implications of the ongoing COVID-19 pandemic. IMPORTANCE: Utilizing a human iPSC-derived kidney organoid model with improved proximal tubule (PT) maturity, we identified the mechanism of SARS-CoV-2 entry in renal cells, confirming ACE2 as the sole receptor and revealing redundancy in downstream cell surface TMPRSS- and endocytic Cathepsin-mediated pathways. In addition, these data address the implications of SARS-CoV-2 exposure in the setting of the commonly prescribed ACE-inhibitor, lisinopril, confirming its negligible impact on infection of kidney cells. Taken together, these results provide valuable insight into the mechanism of viral infection in the human kidney.

A genetically inducible endothelial niche enables vascularization of human kidney organoids with multilineage maturation and emergence of renin expressing cells.

Vascularization plays a critical role in organ maturation and cell-type development. Drug discovery, organ mimicry, and ultimately transplantation hinge on achieving robust vascularization of in vitro engineered organs. Here, focusing on human kidney organoids, we overcame this hurdle by combining a human induced pluripotent stem cell (iPSC) line containing an inducible ETS translocation variant 2 (ETV2) (a transcription factor playing a role in endothelial cell development) that directs endothelial differentiation in vitro, with a non-transgenic iPSC line in suspension organoid culture. The resulting human kidney organoids show extensive endothelialization with a cellular identity most closely related to human kidney endothelia. Endothelialized kidney organoids also show increased maturation of nephron structures, an associated fenestrated endothelium with de novo formation of glomerular and venous subtypes, and the emergence of drug-responsive renin expressing cells. The creation of an engineered vascular niche capable of improving kidney organoid maturation and cell type complexity is a significant step forward in the path to clinical translation. Thus, incorporation of an engineered endothelial niche into a previously published kidney organoid protocol allowed the orthogonal differentiation of endothelial and parenchymal cell types, demonstrating the potential for applicability to other basic and translational organoid studies.

The origin and role of the renal stroma.

The postnatal kidney is predominantly composed of nephron epithelia with the interstitial components representing a small proportion of the final organ, except in the diseased state. This is in stark contrast to the developing organ, which arises from the mesoderm and comprises an expansive stromal population with distinct regional gene expression. In many organs, the identity and ultimate function of an epithelium is tightly regulated by the surrounding stroma during development. However, although the presence of a renal stromal stem cell population has been demonstrated, the focus has been on understanding the process of nephrogenesis whereas the role of distinct stromal components during kidney morphogenesis is less clear. In this Review, we consider what is known about the role of the stroma of the developing kidney in nephrogenesis, where these cells come from as well as their heterogeneity, and reflect on how this information may improve human kidney organoid models.

Comparison of Tobacco Product Prices at Fort Liberty Army Installation and Surrounding Community Areas, 2021.

INTRODUCTION: High rates of tobacco use persist in the U.S. military, with 18.4% of service members smoking cigarettes in 2018. The Department of Defense's (DoD) 2017 policy required that tobacco retailers on military installations set tobacco product prices equal to the most common community price, including tax, but there is limited evidence confirming whether local retailers are adhering to this policy. We examined tobacco product pricing in tobacco retailers on- and off-post at the largest U.S. Army installation, Fort Liberty, and Cumberland County, North Carolina. METHODS: Between June-August 2021, we collected data on tobacco product availability, price, and promotions from retailers on Fort Liberty (n=14) and a random sample of off-post retailers within 10-miles of installation gates (n=52). We calculated the mode, mean, and median price of each product, plus the difference in these prices at on- and off-post retailers. We used Welch's t-test to test differences in mean prices between on- vs. off-post retailers. RESULTS: The mode, mean, and median prices of cigarette packs and cartons were lower on-post than off-post (e.g., $0.51-$0.55 cheaper for Marlboro cigarette packs on-post). However, the mode, mean, and median prices of smokeless tobacco products and little cigars were higher on-post than off-post (e.g., $0.82-$0.89 more costly for Swisher Sweets 2-packs on-post). CONCLUSION: Results highlight the need for continued enforcement to ensure compliance with the 2017 DoD policy. Comprehensive policy action to reduce tobacco price disparities on- and off-post is critical to reducing high rates of tobacco use among service members. IMPLICATIONS: Despite the implementation of the 2017 DoD pricing policy, some tobacco products remain cheaper at tobacco retailers on-post compared to off-post retailers. Our results highlight the need for greater routine surveillance to increase implementation of the policy-particularly for cigarettes-to reduce high rates of tobacco use among service members.

Kidney Organoids Generated Using an Allelic Series of NPHS2 Point Variants Reveal Distinct Intracellular Podocin Mistrafficking.

BACKGROUND: NPHS2 variants are the most common cause of steroid-resistant nephrotic syndrome in children >1 month old. Missense NPHS2 variants were reported to cause mistrafficking of the encoded protein, PODOCIN, but this conclusion was on the basis of overexpression in some nonpodocyte cell lines. METHODS: We generated a series of human induced pluripotent stem cell (iPSC) lines bearing pathogenic missense variants of NPHS2 , encoding the protein changes p.G92C, p.P118L, p.R138Q, p.R168H, and p.R291W, and control lines. iPSC lines were also generated from a patient with steroid-resistant nephrotic syndrome (p.R168H homozygote) and a healthy heterozygous parent. All lines were differentiated into kidney organoids. Immunofluorescence assessed PODOCIN expression and subcellular localization. Podocytes were transcriptionally profiled and PODOCIN-NEPHRIN interaction interrogated. RESULTS: All variant lines revealed reduced levels of PODOCIN protein in the absence of reduced transcription. Although wild-type PODOCIN localized to the membrane, distinct variant proteins displayed unique patterns of subcellular protein trafficking, some unreported. P118L and R138Q were preferentially retained in the endoplasmic reticulum (ER); R168H and R291W accumulated in the Golgi. Podocyte profiling demonstrated minimal disease-associated transcriptional change. All variants displayed podocyte-specific apoptosis, which was not linked to ER stress. NEPHRIN-PODOCIN colocalization elucidated the variant-specific effect on NEPHRIN association and hence NEPHRIN trafficking. CONCLUSIONS: Specific variants of endogenous NPHS2 result in distinct subcellular PODOCIN localization within organoid podocytes. Understanding the effect of each variant on protein levels and localization and the effect on NEPHRIN provides additional insight into the pathobiology of NPHS2 variants. PODCAST: This article contains a podcast at https://dts.podtrac.com/redirect.mp3/www.asn-online.org/media/podcast/JASN/2023_01_05_JASN2022060707.mp3.

The potential feasibility of tobacco-focused medication therapy management in pharmacies affiliated with Federally Qualified Health Centers: Perspectives of pharmacists.

BACKGROUND: Tobacco-focused medication therapy management (MTM) interventions executed in pharmacies located in Federally Qualified Health Centers (FQHC) may provide an innovative means to reach smokers with low incomes and reduce health disparities. However, greater understanding of the intervention's potential feasibility in this setting is needed. OBJECTIVE: To inform the feasibility of implementing an MTM program to address tobacco and nicotine dependence in the FQHC setting by assessing the experience and perceptions of pharmacists working in pharmacies associated with FQHCs. METHODS: A convergent mixed methods approach was used to assess indicators associated with the domains of the Consolidated Framework for Implementation Research (CFIR). Pharmacists from FQHC-based pharmacies in the Southeast United States completed surveys (n=24) and interviews (n=15). Quantitative data were summarized descriptively. Qualitative data were content coded. RESULTS: Quantitative and qualitative data were mapped across all five CFIR domains. Pharmacists report high rates of tobacco and nicotine use among their patients and that addressing their use is important. 62.5% of pharmacists had some or a great deal of experience with tobacco and nicotine dependence. Quantitative and qualitative data demonstrate that the pharmacists and their FQHCs would support MTM efforts focused on tobacco and nicotine dependence. Qualitative findings highlight that pharmacists view an MTM intervention as aligning with their current workflow. Quantitative and qualitative data highlight how factors related to pharmacists' engagement in introducing tobacco and nicotine dependence treatment programs to patients, the electronic medical record, time, staffing, and patient-level barriers could impact the feasibility of an MTM intervention focused on tobacco and nicotine dependence. CONCLUSION: Findings suggest an MTM intervention focused on tobacco and nicotine dependence has the potential to be feasible within FQHC-based pharmacies. Considerations related to training, staffing, time, identifying participants, and supporting participant engagement must be taken into account to support its implementation.

A randomized controlled trial to reduce post-cessation weight gain.

BACKGROUND/OBJECTIVES: Weight gain is a barrier to smoking cessation. Previous interventions targeting weight gain while quitting smoking have largely been unsuccessful. The current study aimed to assess the efficacy of weight stability and weight loss interventions compared to a low-intensity, self-guided bibliotherapy weight management group. SUBJECTS/METHODS: A randomized controlled trial with 12-month follow-up from 2018 to 2022 was conducted with participants (N = 305) who reported smoking at least five cigarettes per day for the last year and interest in quitting initially recruited from the Memphis, TN, USA area. Recruitment was expanded nationally with the onset of the COVID-19 pandemic. Subsequently, 276 completed 12-month follow-up. INTERVENTIONS/METHODS: The Bibliotherapy group was provided a weight management book. Both the Stability and Loss groups met via telephone for eight weeks to learn strategies for maintaining/losing weight, respectively. All three groups then received the same six-week smoking cessation intervention, with six months of varenicline provided. RESULTS: Individuals in the Loss group lost more weight (-2.01 kg, SE = 1.58) than individuals in the Bibliotherapy group (+1.08 kg, SE = 1.49, p = 0.0004), while the Stability group (-0.30 kg, SE = 1.56) was not significantly different from the Bibliotherapy group (p = 0.17). Those in the Stability group did not gain a significant amount of weight. Participants in the Loss group did not gain back all weight lost after smoking cessation and ended the study approximately 2.01 kg lower than baseline. The Bibliotherapy group did not gain the amount of weight expected after cessation. There were no significant differences between groups related to self-reported smoking cessation at each time point except at eight-month follow-up (p = 0.005). CONCLUSIONS AND RELEVANCE: Results indicated the Stability and the Loss interventions were effective for preventing post-smoking cessation weight gain, with the Loss group having the benefit of sustained weight loss. These interventions may be helpful to implement to combat weight gain and potentially facilitate smoking cessation. TRIAL REGISTRATION: The trial is registered on clinicaltrials.gov (NCT03156660).

Examining Smoking Prevalence Disparities in Virginia Counties by Rurality, Appalachian Status, and Social Vulnerability, 2011-2019.

Objectives. To estimate county-level cigarette smoking prevalence in Virginia and examine cigarette use disparities by rurality, Appalachian status, and county-level social vulnerability. Methods. We used 2011-2019 Virginia Behavioral Risk Factor Surveillance System proprietary data with geographical information to estimate county-level cigarette smoking prevalence using small area estimation. We used the Centers for Disease Control and Prevention's social vulnerability index to quantify social vulnerability. We used the 2-sample statistical t test to determine the differences in cigarette smoking prevalence and social vulnerability between counties by rurality and Appalachian status. Results. The absolute difference in smoking prevalence was 6.16 percentage points higher in rural versus urban counties and 7.52 percentage points higher in Appalachian versus non-Appalachian counties in Virginia (P < .001). Adjusting for county characteristics, a higher social vulnerability index is associated with increased cigarette use. Rural Appalachian counties had 7.41% higher cigarette use rates than did urban non-Appalachian areas. Tobacco agriculture and a shortage of health care providers were significantly associated with higher cigarette use prevalence. Conclusions. Rural Appalachia and socially vulnerable counties in Virginia have alarmingly high rates of cigarette use. Implementation of targeted intervention strategies could reduce cigarette use, ultimately reducing tobacco-related health disparities. (Am J Public Health. 2023;113(7):811-814. https://doi.org/10.2105/AJPH.2023.307298).

A Postpartum Weight Loss-focused Stepped-care Intervention in a Military Population: A Randomized Controlled Trial.

OBJECTIVE: Postpartum weight retention is associated with adverse health among both civilian and military women. PURPOSE: The current study evaluated a stepped-care weight management intervention, Moms Fit 2 Fight, adapted for use in a pregnant and postpartum military population. METHODS: Active duty women and other TRICARE beneficiaries (N = 430) were randomized to one of three conditions: gestational weight gain only (GWG-only) intervention (n =144), postpartum weight loss only (PPWL-only) intervention (n =142), or a combined GWG + PPWL intervention (n = 144). Those participants who received the PPWL intervention (i.e., the PPWL-only and GWG+PPWL conditions) were combined consistently with the pre-registered protocol and compared to those participants who did not receive the PPWL intervention in the primary analyses. Primary outcome data (i.e., postpartum weight retention) were obtained at 6-months postpartum by unblinded data collectors, and intent-to-treat analyses were conducted. RESULTS: Retention at 6-months postpartum was 88.4%. Participants who received the PPWL intervention retained marginally less weight (1.31 kg) compared to participants that received the GWG-only intervention (2.39 kg), with a difference of 1.08 kg (p = .07). None of the measured covariates, including breastfeeding status, were significantly associated with postpartum weight retention. Of the participants who received the PPWL intervention, 48.1% participants returned to their pre-pregnancy weight at 6-months postpartum, with no significant differences compared to those who received the GWG-only intervention. CONCLUSIONS: A behavioral intervention targeting diet and physical activity during the postpartum period had a trend for reduced postpartum weight retention. CLINICAL TRIAL INFORMATION: The trial is registered on clinicaltrials.gov (NCT03057808).

Since postpartum weight retention is associated with negative health outcomes among women in the military and women in the general population, the Moms Fit 2 Fight study evaluated a stepped-care weight management intervention among active duty women and other military health insurance beneficiaries. Participants (N = 430) were recruited in their first trimester of pregnancy and randomized to one of three conditions: pregnancy weight gain-only intervention, postpartum weight loss (PPWL)-only intervention, or a combined pregnancy weight gain and PPWL intervention. Participants who received the PPWL intervention (i.e., the participants who received the PPWL-only intervention or the combined intervention) were compared to the participants who did not receive the PPWL intervention, based on weight retention at 6-months postpartum. Participants who received the PPWL intervention retained marginally less weight compared to participants that did not receive the PPWL intervention. Thus, this behavioral intervention targeting diet and physical activity during the postpartum period had a trend for reducing postpartum weight retention, which may be beneficial for achieving military fitness standards and avoiding escalating obesity over multiple pregnancies.

Reengagement for Long-Term Smoking-Cessation In Military Personnel, Retirees, Family Members (TRICARE): A Randomized Trial.

INTRODUCTION: We sought to determine what type of treatment reengagement after smoking relapse would increase long-term cessation. AIMS AND METHODS: Participants were military personnel, retirees, and family members (TRICARE beneficiaries) recruited across the United States from August 2015 through June 2020. At baseline, consented participants (n = 614) received a validated, four-session, telephonic tobacco-cessation intervention with free nicotine replacement therapy. At the 3-month follow-up, 264 participants who failed to quit or relapsed were offered the opportunity to reengage in cessation. Of these, 134 were randomized into three reengagement conditions: (1) repeat initial intervention ("recycle"), (2) Smoking reduction with eventual cessation goal ("rate reduction"), or (3) Choose #1 or #2 ("choice"). Prolonged abstinence and 7-day point prevalence abstinence were measured at 12 months. RESULTS: Despite being in a clinical trial advertised as having the opportunity for reengagement, only 51% (134 of the 264) of participants who still smoked at 3-month follow-up were willing to reengage. Overall, participants randomized to recycle had higher prolonged cessation rates at 12 months than rate reduction conditions (OR = 16.43, 95% CI: 2.52 to 107.09, Bonferroni adjusted p = .011). When participants who randomly received recycle or rate reduction were pooled, respectively, with participants who chose recycle or rate reduction in the Choice group, recycle had higher prolonged cessation rates at 12 months than rate reduction (OR = 6.50, 95% CI: 1.49 to 28.42, p = .013). CONCLUSIONS: Our findings suggest service members and their family members who fail to quit smoking but are willing to reengage in a cessation program are more likely to benefit from repeating the same treatment. IMPLICATIONS: Finding methods that are both successful and acceptable to reengage people who smoke who want to quit can have a significant impact on improving the health of the public by reducing the portion of the population who smoke. This study suggests that repeating established cessation programs will result in more people ready to quit successfully achieving their goal.

Gestational Weight Gain During the COVID-19 Pandemic.

INTRODUCTION: Healthy gestational weight gain (GWG) is associated with improved pregnancy and delivery outcomes. The COVID-19 pandemic changed eating behaviours and physical activity, and thus may have impacted GWG. This study examines the impact of the COVID-19 pandemic on GWG. METHODS: Participants (N = 371, 86% of the larger study) were part of a study focused on GWG among TRICARE beneficiaries (i.e., active-duty military personnel and other beneficiaries). Participants were randomized to two treatment groups (GWG intervention (n = 149 pre-COVID and n = 98 during COVID), and usual care condition (n = 76 pre-COVID and n = 48 during COVID). GWG was calculated as the difference between screening weight and at 36 weeks gestation. Participants who delivered prior to the COVID-19 pandemic (March 1, 2020, N = 225) were compared to participants whose pregnancies occurred during the pandemic (N = 146). RESULTS: We found no significant difference in GWG between those who delivered prior to the pandemic (11.2 ± 4.3 kg) and those whose pregnancies occurred during COVID-19 (10.6 ± 5.4 kg), with no effect of intervention arm. While excessive GWG was higher pre-COVID (62.8%) than during the pandemic (53.7%), this difference was not significant overall or by intervention arm. In addition, we found lower attrition during the pandemic (8.9%) than in the pre-COVID period (18.7%). DISCUSSION: In contrast to prior research that indicated challenges with engaging in health behaviors during the COVID-19 pandemic, we found that women did not have increased GWG or higher odds of excessive GWG. This research contributes to our understanding of how the pandemic impacted pregnancy weight gain and engagement in research.

Regrow or Repair: An Update on Potential Regenerative Therapies for the Kidney.

Fifteen years ago, this journal published a review outlining future options for regenerating the kidney. At that time, stem cell populations were being identified in multiple tissues, the concept of stem cell recruitment to a site of injury was of great interest, and the possibility of postnatal renal stem cells was growing in momentum. Since that time, we have seen the advent of human induced pluripotent stem cells, substantial advances in our capacity to both sequence and edit the genome, global and spatial transcriptional analysis down to the single-cell level, and a pandemic that has challenged our delivery of health care to all. This article will look back over this period of time to see how our view of kidney development, disease, repair, and regeneration has changed and envision a future for kidney regeneration and repair over the next 15 years.

Examination of Tobacco-Related Messaging and Tobacco Use over Time among U.S. Military Young Adults.

Background: People from minoritized populations have historically been targeted by tobacco companies. Little is known about exposure to tobacco-related messages among military personnel from disadvantaged backgrounds. Objectives: The current study aimed to examine exposure to tobacco-related messaging across many nicotine products and through a variety of mediums (i.e., family, friends, advertisements, event promotions, social media) among diverse military populations and use one year later in a sample of young adults who recently enlisted in the U.S. Air Force. Methods: In this study, 8,901 U.S. Air Force trainees reported on demographics, tobacco use, and exposure to positive tobacco messages from social sources (i.e., friends, family, social media) and environmental sources (i.e., advertisements and promotions). Tobacco use was reported one-year later. Results: Compared to others of the same reported racial/ethnic background, Latino/a/x (Relative Risk Ratio [RRR] = 1.354, 95% CI: [1.145, 1.563]) and multiracial (RRR = 1.594, 95% CI: [1.173, 2.016]) participants who were exposed to positive tobacco messages from social sources were significantly more likely to report tobacco product use at one-year follow-up than those who were not exposed to social messages. Exposure to positive tobacco messages from environmental sources were not significantly associated with tobacco use one year later. Conclusions: Social messages may play an important role in increasing risk of tobacco use among some minoritized populations. Cultural as well as systemic factors could be addressed in future tobacco prevention programs to decrease the potency of positive tobacco-related social messages among Latino/a/x and multiracial communities.

Generating Kidney from Stem Cells.

Human kidney tissue can now be generated via the directed differentiation of human pluripotent stem cells. This advance is anticipated to facilitate the modeling of human kidney diseases, provide platforms for nephrotoxicity screening, enable cellular therapy, and potentially generate tissue for renal replacement. All such applications will rely upon the accuracy and reliability of the model and the capacity for stem cell-derived kidney tissue to recapitulate both normal and diseased states. In this review, we discuss the models available, how well they recapitulate the human kidney, and how far we are from application of these cells for use in cellular therapies.

Returning to kidney development to deliver synthetic kidneys.

There is no doubt that the development of transplantable synthetic kidneys could improve the outcome for the many millions of people worldwide suffering from chronic kidney disease. Substantial progress has been made in the last 6 years in the generation of kidney tissue from stem cells. However, the limited scale, incomplete cellular complexity and functional immaturity of such structures suggests we are some way from this goal. While developmental biology has successfully guided advances to date, these human kidney models are limited in their capacity for ongoing nephrogenesis and lack corticomedullary definition, a unified vasculature and a coordinated exit path for urinary filtrate. This review will reassess our developmental understanding of how the mammalian embryo manages to create kidneys, how this has informed our progress to date and how both engineering and developmental biology can continue to guide us towards a synthetic kidney.

What can we learn from kidney organoids?

The ability to generate 3-dimensional models of the developing human kidney via the directed differentiation of pluripotent stem cells-termed kidney organoids-has been hailed as a major advance in experimental nephrology. Although these provide an opportunity to interrogate human development, model-specific kidney diseases facilitate drug screening and even deliver bioengineered tissue; most of these prophetic end points remain to be realized. Indeed, at present we are still finding out what we can learn and what we cannot learn from this approach. In this review, we will summarize the approaches available to generate models of the human kidney from stem cells, the existing successful applications of kidney organoids, their limitations, and remaining challenges.

Single cell analysis of the developing mouse kidney provides deeper insight into marker gene expression and ligand-receptor crosstalk.

Recent advances in the generation of kidney organoids and the culture of primary nephron progenitors from mouse and human have been based on knowledge of the molecular basis of kidney development in mice. Although gene expression during kidney development has been intensely investigated, single cell profiling provides new opportunities to further subsect component cell types and the signalling networks at play. Here, we describe the generation and analysis of 6732 single cell transcriptomes from the fetal mouse kidney [embryonic day (E)18.5] and 7853 sorted nephron progenitor cells (E14.5). These datasets provide improved resolution of cell types and specific markers, including subdivision of the renal stroma and heterogeneity within the nephron progenitor population. Ligand-receptor interaction and pathway analysis reveals novel crosstalk between cellular compartments and associates new pathways with differentiation of nephron and ureteric epithelium cell types. We identify transcriptional congruence between the distal nephron and ureteric epithelium, showing that most markers previously used to identify ureteric epithelium are not specific. Together, this work improves our understanding of metanephric kidney development and provides a template to guide the regeneration of renal tissue.

Kidney micro-organoids in suspension culture as a scalable source of human pluripotent stem cell-derived kidney cells.

Kidney organoids have potential uses in disease modelling, drug screening and regenerative medicine. However, novel cost-effective techniques are needed to enable scaled-up production of kidney cell types in vitro We describe here a modified suspension culture method for the generation of kidney micro-organoids from human pluripotent stem cells. Optimisation of differentiation conditions allowed the formation of micro-organoids, each containing six to ten nephrons that were surrounded by endothelial and stromal populations. Single cell transcriptional profiling confirmed the presence and transcriptional equivalence of all anticipated renal cell types consistent with a previous organoid culture method. This suspension culture micro-organoid methodology resulted in a three- to fourfold increase in final cell yield compared with static culture, thereby representing an economical approach to the production of kidney cells for various biological applications.