No abstract

Pluripotent stem cell-derived kidney organoids offer a promising solution to renal failure, yet current organoid protocols often lead to off-target cells and phenotypic alterations, preventing maturity. Here, various dynamic hydrogel architectures are created, conferring a controlled and biomimetic environment for organoid encapsulation. How hydrogel stiffness and stress relaxation affect renal phenotype and undesired fibrotic markers are investigated. The authors observe that stiff hydrogel encapsulation leads to an absence of certain renal cell types and signs of an epithelial-mesenchymal transition (EMT), whereas encapsulation in soft, stress-relaxing hydrogels leads to all major renal segments, fewer fibrosis or EMT associated proteins, apical proximal tubule polarization, and primary cilia formation, representing a significant improvement over current approaches to culture kidney organoids. The findings show that engineering hydrogel mechanics and dynamics have a decided benefit for organoid culture. These structure-property-function relationships can enable the rational design of materials, bringing us closer to functional engraftments and disease-modeling applications.

Electrospun fibrous materials have increasing applications in regenerative medicine due to the similarity of fibre constructs to the morphology of certain extracellular matrices. Although experimentally the electrospinning method is relatively simple, at the theoretical level the interactions between process parameters and their influence on the fibre morphology is not yet fully understood. Here, we hypothesised that a design of experiments (DoE) model could determine combinations of process parameters that result in significant effects on poly-D,L-lactic acid (PDLLA) fibre morphology. The process parameters used in this study were applied voltage, needle-to-collector distance, flow rate and polymer concentration. Data obtained for mean fibre diameter, standard deviation (SD) of the fibre diameter (measure of fibre morphology) and presence of 'beading' on the fibres (beads per μm) were evaluated as a measure of PDLLA fibre morphology. Uniform fibres occurred at SDs of ≤500 nm, 'beads-on-string' morphologies were apparent between ±500 and 1300 nm and large beads were observed at ±1300-1800 nm respectively. Mean fibre diameter was significantly influenced by the applied voltage and interaction between flow rate and polymer concentration. Fibre morphology was mainly influenced by the polymer concentration, while bead distribution was significantly influenced by the polymer concentration as well as the flow rate. The resultant DoE model regression equations were tested and considered suitable for the prediction of parameters combinations needed for desired PDLLA fibre diameter and additionally provided information regarding the expected fibre morphology.

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.