Thiol-ene cross-linked alginate hydrogel encapsulation modulates the extracellular matrix of kidney organoids by reducing abnormal type 1a1 collagen deposition

Abstract

Differentiated kidney organoids from induced pluripotent stem cells hold promise as a treatment for patients with kidney diseases. Before these organoids can be translated to the clinic, shortcomings regarding their cellular, extracellular compositions and developmental plateau needs to be overcome. We performed a proteomic analysis on kidney organoids cultured for a prolonged culture time and we found a specific change in the extracellular matrix composition with increased expression of types 1a1, 2 and 6a1 collagen. Such an excessive accumulation of specific collagen types is a hallmark of renal fibrosis that causes a life-threatening pathological condition by compromising key functions of the human kidney. Here we hypothesized the need for a three-dimensional environment to grow the kidney organoids, which could better mimic the in vivo surroundings of the developing kidney than standard culture on a transwell filter. Encapsulating organoids for four days in a soft, thiol-ene cross-linked alginate hydrogel resulted in decreased type 1a1 collagen expression. Furthermore, the encapsulation did not result in any changes of organoid structural morphology. Using a biomaterial to modulate collagen expression allows for a prolonged kidney organoid culture in vitro and a reduction of abnormal type 1a1 collagen expression bringing kidney organoids closer to clinical application.HIGHLIGHTSProlonging kidney organoid culture results in a developmental plateau instead of improved in vitro maturation.Proteomic analyses point to an increased expression of specific collagen subtypes associated with renal fibrosis.Encapsulating kidney organoids using a soft thiol-ene cross-linked alginate hydrogel reduces collagen type 1a1 and αSMA deposition.

Human pluripotent stem cell-derived kidney organoids for personalized congenital and idiopathic nephrotic syndrome modeling

“…The 3D configurations and the role of mechanical as well as extracellular matrix (ECM) cues that influence cellular (de)differentiation to develop accurate organoid models need to be addressed in order to establish improved glomerular models in vitro . The effect of ECM cues and material stiffness on organoid differentiation was nicely shown by Garetta et al and also by Geuens and co-workers (Garreta et al, 2019; Geuens et al, 2021). Organoids encapsulated in soft biomaterials showed accelerated differentiation, less fibrosis upon aging, and reduced off-target cell populations, all aspects that will aid in signaling to enhance the maturation of podocytes but also other kidney segments like the proximal tubule and the collecting duct.…”

Win, Lose, or Tie: Mathematical Modeling of Ligand Competition at the Cell–Extracellular Matrix Interface

“…Next, we simulated a 9-fold change [which was the maximum fold-change observed in kidney organoid ECM mass spectrometry experiments by Geuens et al (2020a) ] between days 18 and 25 ( Figure 2 condition 4) for L 2. This simulation was done to reflect the effect of a higher fold of the ligand with the lower integrin binding rate on the system.…”

“…Using the ODE system described above, we performed a set of in silico experiments, aimed at characterizing the effects of increased ECM ligand concentration on integrin binding in a cell culture system. Initial molar concentrations for fibronectin (0.18 nM) and vWA (0.33 nM) were taken from Hudson et al while we used the fold changes reported by Geuens et al (2020a) between days 18 and 25 for these two ligands (2.5-fold and 1.5-fold for fibronectin and vWA) in kidney organoid culture. This way we obtained the test condition 1 “Different Initial Conditions, Different Fold Change” in Table 2 and Figure 2 .…”

“…Maintaining the appropriate ECM composition is critical for kidney organoid development. For example, Geuens et al (2020a) reported an unwanted increase in specific ECM proteins when cell culture times were prolonged in an attempt to increase kidney organoid maturation. They performed a tandem mass spectrometry analysis to compare the ECM composition of kidney organoids that were cultured for 18 and 25 days.…”

“…The analysis also showed an increase in αSMA — a myofibroblast marker — in older kidney organoids, that further indicated tissue fibrosis. Following this analysis, they encapsulated the kidney organoids in a soft hydrogel system, which prevented the unwanted ECM deposition, perhaps by better mimicking the natural environment in kidney development ( Geuens et al, 2020a ).…”