The kidney plays a vital role in regulating bodily fluids and maintaining their composition. It performs crucial tasks such as blood filtration, hormone regulation, and waste removal, all of which are essential for maintaining organism homeostasis. The kidney’s remarkable anatomy enables it to carry out these functions effectively: its highly vascularized structure allows for efficient filtration from the blood vessels, while its connection to the urinary tract facilitates the excretion of waste in the form of urine. These processes are carefully regulated by signalling hormones within the body, which are essential for maintaining blood pressure, electrolyte concentration and acid-base balance.

The kidneys contain approximately 1 million nephrons, which are the functional units (Bertram et al., 2011). Each nephron consists of two major components: the renal corpuscle, responsible for blood filtration, and tubular structures that facilitate the uptake and secretion of solutes. At the distal end of each nephron, waste and excess water are drained into a network of collecting ducts, which eventually converge into the ureter.

In humans, the formation of nephrons (nephrogenesis), is completed around 36 weeks of pregnancy, ensuring the kidney’s complexity is established by birth. Throughout this developmental time, various environmental cues and genetic factors guide the precise localization and formation of tissue components within the organ. While the genetic and biochemical cues involved in kidney development are well understood, the role of mechanical cues has remained largely unexplored.

Given the kidney’s intrinsic complexity and its vital role in maintaining overall organism homeostasis, it has gathered significant interest in the scientific community. Researchers have been particularly focused on unravelling the morphogenetic principles that govern kidney development from early embryonic stages until birth. Advancements in developmental studies, including the use of human pluripotent stem cells- (hPSCs) or adult stem cell-derived organoids, have pushed the boundaries of our understanding of organogenesis and disease progression in this complex organ. However, despite these major breakthroughs, the precise mechanisms underlying the (bio) mechanical machinery that contributes to renal fate specification and the overall tissue organization in vivo remain unknown.

As the recognition of mechanical cues in development grows, there is an increasing demand for sophisticated bioengineering solutions to understand the mechanical signals and responses involved in organogenesis. Mechanobiological techniques, as reviewed in (Gómez-González et al., 2020), can help uncover the role of mechanical cues in tissue morphogenesis. At the same time, the generation of a more representative developmental architecture using hPSCs-derived organoids, as discussed in (Garreta et al., 2021), holds tremendous promise for gaining a deeper understanding of the interplay between the genetic and (bio) mechanical machinery contributing to organogenesis.

This review aims to summarize current knowledge on mammalian kidney development and provide insights into the state-of-the-art methodologies for in vitro kidney organoid differentiation, which pose as valuable tools for studying development and disease. The review will also discuss what is known so far regarding the presence of mechanical cues during embryo and organ development and will highlight state-of-the-art techniques commonly employed in the field of tissue mechanics to probe cell-to-tissues responses. At the same time, here we aim to provide a comprehensive view on how these tools may be translated to understand kidney morphogenesis exploiting hPSCs-derived kidney organoid technology. Finally, the review will explore the rising need for more interdisciplinarity between the fields of classical developmental biology and mechanobiology, which can significantly enhance our understanding of organ development and disease progression in humans.

The development of the adult mammalian kidney begins during embryogenesis with the formation of three precursor structures: pronephros, mesonephros and metanephros. The prior two are transient, yet still necessary structures, the absence of which have been associated with renal agenesis. The metanephros on the other hand is the permanent structure that gives rise to the functional adult kidney (Reidy and Rosenblum, 2009). Primitive urinary system commitment occurs at the blastula-stage of embryonic development, where the blastocyst undergoes an invagination, referred to as the primitive streak (PS). At those early stages of development, antero-posterior (A-P) patterning information is conveyed by the movement of cells through the PS at different time points that consequently affects the expression of kidney genes. Importantly, other kidney-inductive signals are present along the whole axis, including anterior non-kidney-generating regions. Then, the posterior end of the PS, as largely demonstrated by making use of embryonic mouse experiments [reviewed in (Dressler, 2009; Halt and Vainio, 2014)] further evolves to the Intermediate Mesoderm (IM), from where the urogenital system -the kidneys, the gonads, and their respective duct systems-will be derived. Upon polarization, the IM will be segmented into the anterior IM (AIM) and posterior IM (PIM). The AIM eventually epithelializes into the Wolffian duct, also known as the mesonephric duct which are tubular structures leading to the cloaca (future bladder). In humans, at the 5th week of gestation, a small protrusion of the mesonephric duct is observed, named the ureteric bud (UB), indicating the initiation of the metanephros formation. This structure is rapidly enclosed by cells from the metanephric mesenchyme (MM) lineage which further aggregate surrounding the bud’s ends, forming cap-like structures, namely, the cap mesenchyme. From this stage onwards, reciprocal interactions between cells from the UB and MM lineage are established, guiding their co-development. MM cells produce GDNF, ultimately stimulating UB branching morphogenesis to form a urine draining tubular network. In parallel, canonical Wnt signals from UB cells to the MM population induces the mesenchymal-to-epithelial transition (MET) of nephron progenitor cells (NPCs). Upon activation of the Wnt pathway, differentiated NPCs further cluster together forming lumen-presenting spherical aggregates termed renal vesicles (RVs). These are then polarized, elongated and undergo a series of structural changes to eventually segment into the different sections of the mature nephron (Dressler, 2006; Lindström and Tran, 2018b; Khoshdel Rad et al., 2020).

These RV structures elongate into polarized comma-shaped bodies, then S-shaped bodies, readily connected to the UB-derived collecting duct. S-shaped bodies reportedly evolve into mature nephrons, exhibiting glomerulus and tubular structures, including the proximal tubule, loop of Henle and distal tubule. All these structural changes are known to be guided by the genetic blueprint. In humans nephrogenesis repeats until shortly before birth.

Like any other developmental process, kidney organogenesis exhibits a remarkable sensitivity. Indeed, disruptions or impairment at any stage of kidney morphogenesis can precipitate a cascade of severe inborn conditions, ultimately known as congenital anomalies of the kidneys and urinary tract (CAKUT). As such, CAKUT encompasses a family of urinary system malformation at birth, including kidney defects such as renal agenesis or hypoplasia, as well as ureteric malfunctions (Nicolaou et al., 2015). These anomalies, although generally associated with mutations in specific genes involved in the urinary system formation, may also be due to improper physical interactions between UB and MM cells (Jain and Chen, 2019). For instance, supernumerary kidneys, which present multiple ureters, arise upon impaired UB and MM interaction, where UB bifurcation occurs prior MM invasion. In the case of kidney hypoplasia, which manifests as small kidneys with low nephron number, reduced branching morphogenesis events of the UB have been recorded.

Several CAKUT manifestations have been reported to be clinically associated with hypertension, ultimately leading to, in certain occasions, end stage renal disease (ESRD). There is therefore an urge to understand the mechanisms underlying CAKUT anomalies in the human context. Indeed, several works have been developed in knock-out animal models, however, the phenotypical manifestation of the pathological mutations may vary from species to species (Khan et al., 2022).

Mammalian kidney development is a fascinating process that exhibits remarkable evolutionary conservation across species. The study of human-specific kidney development throughout gestation has been challenged by limited access to human embryonic kidney samples and the ethical concerns surrounding their origin. However, this limitation has been overcome by utilizing animal models of lower complexity, such as rats and mice, to investigate the intricate cellular and molecular processes involved in kidney organogenesis. By leveraging the remarkable conservation of embryogenesis across mammalian species, findings from these animal models have provided valuable insights into human kidney development (Cullen-McEwen et al., 2015). In fact, these studies have contributed to our understanding of the different stages of kidney morphogenesis and have identified fundamental signalling pathways, including the canonical Wnt pathway, that play a crucial role in this process (Carroll et al., 2005; Lindström and Tran, 2018b).

Comparison of embryonic kidney development across diverse mammalian species has revealed high levels of similarity. Notably, studies have documented shared morphological features between mouse and human embryonic and adult tissues, demonstrating clear parallels in tissue architecture at species-specific gestational timepoints (Lindström and Tran, 2018b). The blueprint of nephron morphology and positioning relative to other cell lineages within the kidney is conserved between both species. Nephron-patterning events also exhibit strong conservation, as observed through immunocytochemistry studies: the polarization of renal vesicles, primitive nephron-oriented structures, segmentation into proximal and distal sections can all be observed in mammalian species.

Despite the diversity of mammalian species, the fundamental principles and mechanisms governing kidney development remain remarkably similar. Indeed, our current understanding of kidney organogenesis has been derived from mouse and rat embryonic kidney samples. Shared anatomy, patterning and signalling pathways have emphasized the utility of mouse embryonic models in recapitulating kidney development when human samples are unavailable. And while the overall process of embryonic kidney development remains evolutionarily conserved among mammalian species, inter-species molecular and anatomical variations still exist (O’Brien et al., 2016; Lindström and Tran, 2018b). Indeed, some divergent features have been observed when comparing mouse to human embryonic nephrogenesis (O’Brien et al., 2016). For example, only a minority of previously established mouse anchor genes, which are expected to distinctly encode kidney structures, display a conserved expression pattern in humans (Lindström and Guo, 2018a). Discrepancies in nephron number have been reported, with mouse kidneys containing approximately 12–16,000 nephrons, while human kidneys include an average of 1 million nephrons. This difference in nephron quantity is due to variations in time of gestation (9 months in humans, approximately 20 days in mice) and the overall kinetics of nephrogenesis (O’Brien and McMahon, 2014).

With regards to inborn conditions affecting the kidney, the mechanisms underlying CAKUT, for instance, have been more challenging to address in non-human models. Importantly, heterozygous or homozygous mutations in mice models have caused either embryonic or postnatal lethality, making it difficult to study the underlying pathophysiology (Stone et al., 2016).

The establishment of human embryonic and induced pluripotent stem cell lines (hESCs and hiPSCs, referred as hPSCs) has significantly impacted how scientists study human development and diseases outside of the human body (Thomson et al., 1998; Takahashi and Yamanaka, 2006; Takahashi et al., 2007). The capability of hPSCs to expand indefinitely while preserving pluripotent differentiation features (that is to give rise to cells of the three-germ layer of the embryo in vivo and in vitro) allows for the interrogation of early stages of cell lineage specification and differentiation. Thanks to this, specific cell culture conditions sustaining and promoting cell differentiation have been identified and are now consistently used as a tool in developmental research. For more than 2 decades researchers world-wide have started to define external conditions aiming to guide hPSCs differentiation while exploiting inherent characteristics of these cells, that are capable of self-organization and symmetry breaking (Armstrong, 1989). While initial studies of hPSCs differentiation promoted early stages of cell lineage specification and commitment in two dimensional culture conditions (Laflamme et al., 2007), seminal studies started to address these questions making use of embryoid bodies (EBs) as surrogates of differentiation and cell specification along the three germ lineages of the human embryo (Yang et al., 2008; Chambers et al., 2009).

A technical evolution of EB culture systems led to the overall explosion of the organoid field from hPSCs. This was based on the seminal work from Yoshiki Sasai on the establishment of the first self-patterned stratified cortical tissues generated by plating EBs in serum-free medium on a coated surface (Eiraku et al., 2008). At the present time, the term “organoid” refers to a three-dimensional (3D) collection of cells that resembles an organ [reviewed in detail in (Lancaster and Knoblich, 2014)]. To achieve resemblance, this 3D structure comprises of several different cell types that are characteristically present in the native organ. In vivo, different cell types arise from stem cells through a process of lineage commitment and cell sorting, that allow a spatial organization like an organ during development. In vitro, self-organized collection of hPSCs-derived cell types can recapitulate, until certain extent, specific functions of an organ.

Cutting-edge research from laboratories around the world have led to the generation of kidney organoids from hPSCs by mimicking in vitro renal inductive signals that occur during kidney organogenesis (Xia et al., 2014; Freedman et al., 2015; Morizane et al., 2015; Takasato and ErPei, 2015; Taguchi and Nishinakamura, 2017; Garreta et al., 2019). These studies have revealed specific morphogens and cytokines that can drive the differentiation of PSCs towards a renal specification ex vivo (Takasato et al., 2014; 2016; Xia et al., 2014; Morizane et al., 2015; Takasato and ErPei, 2015; Taguchi and Nishinakamura, 2017; Uchimura et al., 2020). Collectively, procedures allowing the generation of organoids resembling separate components of the mammalian kidney have been established (Figure 1), including: nephron progenitor (NP), ureteric bud (UB) and even stromal progenitor (SP) organoids (Zeng et al., 2021; Palakkan et al., 2022; Tanigawa et al., 2022; Vanslambrouck et al., 2022).

Of note, the metanephric adult kidney as an organ comprises a highly complex tissue architecture. The current state-of-the-art of procedures in kidney organoid differentiation show a remarkable resemblance to the first/second trimester human embryonic kidney (Garreta et al., 2019). Yet, these methodologies still lack a higher order of anatomical resemblance, where the developing nephrons would connect with the collecting duct to allow for filtration and re-absorption of nutrients. To address these limitations, specialized tissue culture techniques are being widely explored to drive organoid maturation forward, particularly in terms of nephron development (bioreactors), collective duct development (patterned substrates), and vascularization (microfluidics) that better mimic physiologically-relevant cues (Czerniecki et al., 2018; Przepiorski et al., 2018; Homan et al., 2019; Glass et al., 2020; Shi et al., 2023).

One of the advantages of using hPSC-based organoid systems is that they are highly adaptable for genetic manipulation, making it possible to mimic genetic kidney diseases by introducing alterations in the corresponding genes. Patient-derived and/or gene-edited hPSCs have been an excellent source for studying adult kidney disorders such as chronic kidney disease (CKD) and polycystic kidney diseases (PKD), among others [reviewed in more detail in (Karp et al., 2022; Liu et al., 2022)]. While studies with hPSC- derived organoids capturing the PKD genetic background have been seminal in modelling cyst formation (Freedman et al., 2015) and response to drugs (Cruz et al., 2017; Tran et al., 2022), the immature nature of in vitro kidney organoids makes it difficult to model diseases exhibiting adult anomalies. In contrast to chronic-related manifestations, CAKUT maybe effectively studied exploiting hPSCs-derived kidney organoids.

In this regard, CAKUT is an umbrella term encompassing an extensive spectrum of embryonic kidney and urinary tract malformations that affect approximately 1 in 100–500 new-borns (Westland et al., 2021). The phenotypical profile of CAKUT anomalies is quite heterogeneous: the most typical observed phenotypes include hypoplasia, dysplasia, agenesis, ureteral abnormalities among others. More than twenty genes have been identified as liable for the onset of syndromic and non-syndromic CAKUT, potentially explaining the heterogeneous clinical presentation of these congenital anomalies (Nicolaou et al., 2015). Mutations in PAX2, HNF1B, BMP7, RET, GATA3, SALL1, SIX5, and EYA1 among others developmental genes have been previously reported.

Paired box gene 2 (PAX2) is an important developmental gene in kidney organogenesis, expressed throughout nephron differentiation: starting from progenitors to epithelial renal vesicles to distal fragments of the renal tubules. Particularly, in mice, Pax2 was shown to play an indispensable role in MET of NPs, and the positioning and outgrowth of the ureteric bud (Torres et al., 1995; Brophy et al., 2001). The latter phenomenon is likely responsible for the renal agenesis phenotype observed in mice with an absence of this gene. In humans, on the other hand, frameshift mutations have been associated with the development of renal coloboma (Sanyanusin et al., 1995; Bower et al., 2012; Chang et al., 2022). Interestingly, in vitro research performed by Kaku & Nishinakamura et al. has demonstrated that PAX2 is in fact dispensable for MET in NPs developed from hPSCs (Kaku et al., 2017). In their study, PAX2-null hPSCs could still develop into nephron progenitors, and even epithelialize into tubular and glomerular structures. However, while human PAX2 may not be necessary for MET to occur, it is needed for the proper differentiation of parietal epithelial cells of the glomeruli (Kaku et al., 2017).

Hepatocyte nuclear factor 1—β (HNF1B) has been shown to play an important role in nephrogenesis. Studies in zebrafish have identified the role of this transcription factor in nephron patterning and segmentation, where embryos lacking hnf1ba/b would not express markers characteristic for distal and proximal segments, yet still form an epithelial tubule (Naylor et al., 2013). Similar findings have been observed in murine and xenopus nephrogenesis, where a disruption or overexpression of this gene leads to a failure in SSB patterning and segmentation (Naylor et al., 2013). In humans, mutations in HNF1B, together with mutations in PAX2 account for approximately 15% of known CAKUT cases (Nicolaou et al., 2015). Human iPSC-derived kidney organoids have been used as a platform to mimic congenital kidney anomalies involving HNF1B by generating biallelic deletions in the gene. Przepiorski et al. have demonstrated that organoids with disrupted HNF1B fail to develop regions positive for markers of proximal and distal tubules, similar to phenotypes previously observed in Hnf1b conditionally deficient mice (Przepiorski et al., 2018).

Mutations in the receptor tyrosine kinase RET gene are also more commonly found in the spectrum of congenital urinary anomalies like renal agenesis and aplasia. A cohort study found that mutations along the GDNF-RET signalling pathway are present in about 5% of patients with CAKUT and that the nature of the mutations affects the penetrance of CAKUT clinical presentation (Chatterjee et al., 2012). Studies on mutant mice carrying genetic alterations along the GDNF-Gfra1-Ret axis have shed light on the expression patterns of Ret throughout the kidney and lower urinary tract [reviewed in detail in (Jain, 2009)]. Briefly, Ret plays a crucial role in early UB induction and branching, among other roles, without which extreme phenotypes such as renal agenesis or prenatal fatality can be observed. Human PSC-derived UB organoids have been successfully used to mimic congenital anomalies resulting from mutations in RET, proving the feasibility of modelling such diseases by in vitro genetic editing and 3D culture (Zeng et al., 2021). In their study, the authors have demonstrated how RET-null organoids fail to undergo branching and exhibit a phenotype like renal agenesis.

Similarly, hPSC-derived kidney organoid systems have been successfully utilized to model certain podocytopathies by either genetically introducing mutations into wild type PSCs, or by inducing them from patient-specific cells (Kim et al., 2017; Hale et al., 2018). Together, all these studies provide a promising outlook on the use of hPSC-derived kidney organoids as a platform to study early stages of kidney development and diseases, where cases of CAKUT could be of particular interest.

A major advantage of using in vitro kidney organoid systems to model organ development and CAKUT is that they can be interrogated via a variety of functional, physiological, and molecular techniques. One of the simplest and most accessible techniques is morphological analysis by brightfield microscopy, where structures like RVs, complex nephron like structures and cysts (in case of disease modelling) can be roughly identified during the differentiation protocol (Taguchi and Nishinakamura, 2017; Przepiorski et al., 2018; Garreta et al., 2019; Schutgens et al., 2019). Moreover, these structures can be subjected to more detailed histological or immunofluorescence analysis by confocal microscopy, where either whole organoids or tissue sections can be stained for markers such as PAX2, WT1, LHX1, JAG1, E-CADHERIN, and NEPHRIN, corresponding to different segments of the developing nephron (Figure 2A).

Single cell transcriptional profiling of human foetal tissue has been also a key tool in understanding the many intermediate transcriptional states of kidney development. Applying this technique to analyse kidney organoids at different stages of differentiation has also helped map the different cell types present more accurately, as well as provide a transcriptional comparison to human embryonic kidney development (Lindström et al., 2021; Wilson et al., 2022).

Developmental biology has largely focused on discovering the function of morphogens and the molecular processes that drive embryogenesis and tissue formation. However, it has become more evident that tissue formation and morphogenesis do not solely rely on biochemical and genetic cues. In fact, these processes are also influenced by intrinsic and extrinsic biomechanical forces [reviewed in detail in (Heisenberg and Bellaïche, 2013; Goodwin and Nelson, 2021)]. On a subcellular level, cells can experience phenomena such as actomyosin contractility, membrane stretching and tension during cell division or upon exposure to very stiff microenvironments (Colombo et al., 2003; Krieg et al., 2008; Daley et al., 2011). Extracellularly, cells can undergo compression and extrusion during events like apoptosis and tissue remodelling (Yang et al., 2000; Eisenhoffer et al., 2012). On a much grander tissue scale, contractions, for example, by muscle fibres can be observed (Harrington, 1979). Extrinsic forces can even be observed on an organismal level, an example of which is the effect of gravity.

Nowadays, in developmental biology there has been a shift towards investigating how mechanical forces impact the development of tissues and understanding the physical mechanisms that facilitate morphogenesis. The idea that tissue morphogenesis is in fact a biomechanical process has been reviewed by pioneers in the 1980–1990 s (Wolpert, 1981; Beloussov et al., 1994). In fact, they brought forward the ideas that biochemical and mechanical morphogenesis within the developing tissue are not necessarily distinct, as the physical properties of local regions can affect the shape of morphogen fields and the movement of cells can alter how tissues respond to molecular gradients.

Quite like biochemical signalling molecules that instigate an intracellular sequence of events leading to changes in gene expression, mechanical stimuli can trigger a similar cascade. Here, cell adhesion complexes localized on cell membranes can adhere to components of the extracellular matrix (ECM) and create connections like cell-to-ECM and cell-to-cell junctions that are necessary for transmitting the biomechanical signals towards the nucleus and ultimately affect cell function. This ability of cells to sense their mechanical microenvironment is termed mechanosensing, while the signalling cascade of events is known as mechnotransduction. Cell fate decisions that are ultimately influenced by the cell’s mechanical microenvironment include migration, differentiation, proliferation, and cell death, among others.

During embryo development and organogenesis, the composition, stiffness and viscoelasticity of the ECM represent key factors previously reported to influence stem cell fate and differentiation [reviewed in detail in (Guilak et al., 2009; Elosegui-Artola, 2021)]. Indeed, in utero, the embryonic microenvironment contains a plethora of ECM proteins (such as fibronectin, collagen, laminin, etc.,) and a highly viscoelastic matrix (ranging from 0.1 to 100 kPa in stiffness) which has been shown to influence the fate of stem cells within the forming tissue (Guilak et al., 2009; Rozario and DeSimone, 2010; Chaudhuri et al., 2020).

A seminal example of this phenomena is the effect of ECM elasticity on lineage specification of mesenchymal stem cells (MSCs). Studies have shown how upon exposure to soft matrices, these multipotent MSCs can be driven to the adipogenic lineage, while stiffer microenvironments can induce osteogenesis in the same naïve stem cells (Engler et al., 2006; Guilak et al., 2009; Huebsch et al., 2010). Vascular progenitors in vitro have been shown to differentiate into endothelial lineage when exposed to soft microenvironments, while stiff matrices guide them to the smooth muscle cell fate (Wong et al., 2019). Specific ECM conditions are also crucial for the maintenance of tissue homeostasis. For example, the maintenance of mammary gland tissue homeostasis requires a soft and laminin-rich microenvironment (Alcaraz et al., 2008).

At the same time, stiffer matrices with altered ECM protein composition/density are typically conditions that support tumour growth and metastasis (Wullkopf et al., 2018; Bauer et al., 2020). A fascinating study on obesity and cancer invasion has shown how changes in the adipose tissue microenvironment, such as stiffening of the ECM from enrichment of myofibroblasts in obese mice, can increase the malignant potential of breast cancer cells (Seo et al., 2015; Ling et al., 2020). This study demonstrates the effect of ECM stiffness on tumorigenesis and the importance of mechanical stimuli on cell fate decisions (Elosegui-Artola, 2021). Meanwhile, the viscosity vs. elasticity of the ECM is becoming increasingly recognized as an additional modulator of cell fate decision (Chaudhuri et al., 2020; Elosegui-Artola et al., 2022). Matrix viscoelasticity has been demonstrated to guide single cell behaviour, as well as collective behaviours of spheroid/organoid growth and differentiation (Elosegui-Artola et al., 2022). Apart from the biophysical properties of the ECM, fluid flow in the extraembryonic environment has been shown to influence cell behaviour too [reviewed in (Freund et al., 2012)]. Briefly, fluid flow has been previously shown to influence left-right symmetry breaking during organ development (Tanaka et al., 2005), regulation of vascularization and sprouting (Song and Munn, 2011), and normal renal tubule function (Nauli et al., 2003).

The effect of fluid flow has also been explored on kidney organoid differentiation in vitro (Homan et al., 2019). By applying a fluid flow, Homan et al. were able to enhance kidney organoid vascularization and stimulate the maturation of podocytes, a specialized cell type in the kidney. The stiffness of the extracellular environment has also been explored in embryonic kidney differentiation. Garreta et al. and others have shown how a soft synthetic hydrogel can mimic the in utero mechanical microenvironment and enhance the maturation of kidney organoids cultured in vitro (Garreta et al., 2019; Ruiter et al., 2022). These studies highlight the importance of mechanical stimuli in regulating the differentiation of hPSCs into renal lineages and the generation of mature kidney organoids.

In adult kidneys, Choudhury et al., have explored the role of mechanical forces that drive fluid transport across the tubule epithelial cells. They described differences in fluid flux and pressure gradients in human healthy kidney cells vs. autosomal dominant PKD cells, which shed light on the importance of mechanical forces in kidney function and their implications in pathophysiological conditions (Choudhury et al., 2022). In efforts to understand PKD pathophysiology better, Cruz et al. have looked into the role of the microenvironment on disease progression. PKD is associated with the formation of fluid filled cysts from kidney tubular epithelia. The authors have identified that the microenvironment of tubule organoids can substantially increase or decrease the probability of cyst formation (Cruz et al., 2017).

These findings have significant implications for the development of new models for studying kidney development and disease, as well as the potential use of human kidney models in regenerative medicine.

All these studies demonstrate how mechanical input from the surrounding environment and the cell’s ability to mechano-sense these signals contribute to cell fate decisions during healthy tissue development and disease evolution. Nowadays, insights on how these mechanical signals are sensed and internalized by the cells to elicit a genetic response are being slowly uncovered (Discher et al., 2005; Heisenberg and Bellaïche, 2013; De Belly et al., 2022).

To this day, mechanical stimuli like substrate stiffness, physical confinement and fluid flow have been clearly proven to affect cell fate decisions in 2D-in vitro environment (s) [see reviewed in (Mammoto et al., 2012; Wagh et al., 2021)]. In vivo, however, tissues are far more complex, where viscoelasticity plays a key role dictating cell behaviour (Cheng et al., 2008; Geerligs et al., 2008; Safshekan et al., 2017). Here, a multitude of cellular and extracellular components, including the cytoskeleton, ECM proteins, membrane adhesion complexes, cellular cortex, among others, can influence the biomechanical signalling pathway and lead to variable cellular responses in time and space. Moreover, the practical difficulty of accessing organs and tissues during development in vivo increases the complexity of studying their mechanics. All this makes the measurement and probing of the mechanical environment of living tissues in vivo far more complicated.

However, with the rise of organoid technologies it has become possible to mimic embryonic organ development in vitro more closely and to investigate the mechanical changes occurring during early steps of organogenesis in a dish. By combining organoid technology with experimental mechanobiology techniques, it becomes possible to study the mechanical properties of cells and tissues and how they influence organ development and disease in a more controlled and physiologically relevant environment. This can provide new insights into the role of mechanical forces in these processes and help develop new therapeutic strategies for a variety of diseases.

There are several ways to study the mechanobiology of organoid development: through bioengineering of their microenvironment (exposure to synthetic and natural hydrogels, micropatterning, microfluidics devices, etc.), application of mechanobiology techniques (micro-rheology, TFM, optical tweezers, laser ablation, etc.,) and genetic manipulation of key genes involved in mechanosensing and transduction pathways (YAP/TAZ—HIPPO pathway, WNT signalling pathway, SRF pathway).

Altogether, accumulated findings within the last decades have revealed the immense value of hPSC-derived models as a platform on which organogenesis could be recapitulated. By investigating the specification of stem cells towards the kidney lineage in vivo, researchers gained an in-depth knowledge regarding the principles governing nephrogenesis, an invaluable asset to ultimately recreate this process in vitro. While in vitro models of human kidney organogenesis can to some extent faithfully recapitulate the organ’s microanatomy and physiology, there are still major limitations in reproducibility, control of cell type, composition, and lack of vascularized and innervated structures. Here, by acknowledging the mechanical microenvironment of in vivo tissues during development, and by closely mimicking these external cues, it becomes possible to drive organoid maturation a step further and gain a better understanding of organogenesis and disease progression.

While current advances in bioengineering and mechanobiology are paving new strategies for studying and guiding tissue morphogenesis in vitro, there is still a long way to go. In order to guide cells in the direction of organ development, biochemical and mechanical signalling cascades within and between cells need to be perfectly coordinated to navigate through the complex dynamics of tissue patterning. Hence in kidney organogenesis, more consistent efforts are needed to truly understand the underlying biomechanical machinery and how it works in concert with the genetic program. Only then will it be possible to engineer functional kidney tissues.