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Abstract A 3D microenvironment is known to endorse pancreatic islet development from human induced pluripotent stem cells (iPSCs). However, oxygen supply becomes a limiting factor in a scaffold culture. In this study, oxygen‐releasing biomaterials are fabricated and an oxygenated scaffold culture platform is developed to offer a better oxygen supply during 3D iPSC pancreatic differentiation. It is found that the oxygenation does not alter the scaffold's mechanical properties. The in situ oxygenation improves oxygen tension within the scaffolds. The unique 3D differentiation system enables the generation of islet organoids with enhanced expression of islet signature genes and proteins. Additionally, it is discovered that the oxygenation at the early stage of differentiation has more profound impacts on islet development from iPSCs. More C‐peptide⁺/MAFA⁺β and glucagon⁺/MAFB⁺α cells formed in the iPSC‐derived islet organoids generated under oxygenated conditions, suggesting enhanced maturation of the organoids. Furthermore, the oxygenated 3D cultures improve islet organoids’ sensitivity to glucose for insulin secretion. It is herein demonstrated that the oxygenated scaffold culture empowers iPSC islet differentiation to generate clinically relevant tissues for diabetes research and treatment. 

Abstract In vitro differentiation of human induced pluripotent stem cells (iPSCs) into functional islets holds immense potential to create an unlimited source of islets for diabetes research and treatment. A continuous challenge in this field is to generate glucose-responsive mature islets. We herein report a previously undiscovered angiopoietin signal for in vitro islet development. We revealed, for the first time, that angiopoietins, including angiopoietin-1 (Ang1) and angiopoietin-2 (Ang2) permit the generation of islets from iPSCs with elevated glucose responsiveness, a hallmark of mature islets. Angiopoietin-stimulated islets exhibited glucose synchronized calcium ion influx in repetitive glucose challenges. Moreover, Ang2 augmented the expression of all islet hormones, including insulin, glucagon, somatostatin, and pancreatic polypeptide; and β cell transcription factors, including NKX6.1, MAFA, UCN3, and PDX1. Furthermore, we showed that the Ang2 stimulated islets were able to regulate insulin exocytosis through actin-filament polymerization and depolymerization upon glucose challenge, presumably through the CDC42-RAC1-gelsolin mediated insulin secretion signaling pathway. We also discovered the formation of endothelium within the islets under Ang2 stimulation. These results strongly suggest that angiopoietin acts as a signaling molecule to endorse in vitro islet development from iPSCs. 

It is an inside front cover to highlight the interaction between stem cells and oxygenated scaffold material. 

Pluripotent stem cells are endless sources for in vitro engineering human tissues for regenerative medicine. Extensive studies have demonstrated that transcription factors are the key to stem cell lineage commitment and differentiation efficacy. As the transcription factor profile varies depending on the cell type, global transcriptome analysis through RNA sequencing (RNAseq) has been a powerful tool for measuring and characterizing the success of stem cell differentiation. RNAseq has been utilized to comprehend how gene expression changes as cells differentiate and provide a guide to inducing cellular differentiation based on promoting the expression of specific genes. It has also been utilized to determine the specific cell type. This review highlights RNAseq techniques, tools for RNAseq data interpretation, RNAseq data analytic methods and their utilities, and transcriptomics-enabled human stem cell differentiation. In addition, the review outlines the potential benefits of the transcriptomics-aided discovery of intrinsic factors influencing stem cell lineage commitment, transcriptomics applied to disease physiology studies using patients’ induced pluripotent stem cell (iPSC)-derived cells for regenerative medicine, and the future outlook on the technology and its implementation. 

Induced pluripotent stem cells (iPSCs) have enormous potential in producing human tissues endlessly. We previously reported that type V collagen (COL5), a pancreatic extracellular matrix protein, promotes islet development and maturation from iPSCs. In this study, we identified a bioactive peptide domain of COL5, WWASKS, through bioinformatic analysis of decellularized pancreatic ECM (dpECM)-derived collagens. RNA-sequencing suggests that WWASKS induces the formation of pancreatic endocrine progenitors while suppressing the development of other types of organs. The expressions of hypoxic genes were significantly downregulated in the endocrine progenitors formed under peptide stimulation. Furthermore, we unveiled an enhancement of iPSC-derived islets’ (i-islets) glucose sensitivity under peptide stimulation. These i-islets secrete insulin in a glucose responsive manner. They were comprised of α, β, δ, and γ cells and were assembled into a tissue architecture similar to that of human islets. Mechanistically, the peptide is able to activate the canonical Wnt signaling pathway, permitting the translocation of β-catenin from the cytoplasm to the nucleus for pancreatic progenitor development. Collectively, for the first time, we demonstrated that an ECM-derived peptide dictates iPSC fate toward the generation of endocrine progenitors and subsequent islet organoids. 

Decellularization of natural tissues to produce extracellular matrix is a promising method for three-dimensional scaffolding and for understanding microenvironment of the tissue of interest. Due to the lack of a universal standard protocol for tissue decellularization, recent investigations seek to develop novel methods for whole or partial organ decellularization capable of supporting cell differentiation and implantation towards appropriate tissue regeneration. This review provides a comprehensive and updated perspective on the most recent advances in decellularization strategies for a variety of organs and tissues, highlighting techniques of chemical, physical, biological, enzymatic, or combinative-based methods to remove cellular contents from tissues. In addition, the review presents modernized approaches for improving standard decellularization protocols for numerous organ types. 

Tissue microenvironments are rich in signaling molecules. However, factors in the tissue matrix that can serve as tissue-specific cues for engineering pancreatic tissues have not been thoroughly identified. In this study, we performed a comprehensive proteomic analysis of porcine decellularized pancreatic extracellular matrix (dpECM). By profiling dpECM collected from subjects of different ages and genders, we showed that the detergent-free decellularization method developed in this study permits the preservation of approximately 62.4% more proteins than a detergent-based method. In addition, we demonstrated that dpECM prepared from young pigs contained approximately 68.5% more extracellular matrix proteins than those prepared from adult pigs. Furthermore, we categorized dpECM proteins by biological process, molecular function, and cellular component through gene ontology analysis. Our study results also suggested that the protein composition of dpECM is significantly different between male and female animals while a KEGG enrichment pathway analysis revealed that dpECM protein profiling varies significantly depending on age. This study provides the proteome of pancreatic decellularized ECM in different animal ages and genders, which will help identify the bioactive molecules that are pivotal in creating tissue-specific cues for engineering tissues in vitro. 

Diabetes is one of the leading causes of death globally. Currently, the donor pancreas is the only source of human islets, placing extreme constraints on supply. Hence, it is imperative to develop renewable islets for diabetes research and treatment. To date, extensive efforts have been made to derive insulin-secreting cells from human pluripotent stem cells with substantial success. However, the in vitro generation of functional islet organoids remains a challenge due in part to our poor understanding of the signaling molecules indispensable for controlling differentiation pathways towards the self-assembly of functional islets from stem cells. Since this process relies on a variety of signaling molecules to guide the differentiation pathways, as well as the culture microenvironments that mimic in vivo physiological conditions, this review highlights extracellular matrix proteins, growth factors, signaling molecules, and microenvironments facilitating the generation of biologically functional pancreatic endocrine cells from human pluripotent stem cells. Signaling pathways involved in stepwise differentiation that guide the progression of stem cells into the endocrine lineage are also discussed. The development of protocols enabling the generation of islet organoids with hormone release capacities equivalent to native adult islets for clinical applications, disease modeling, and diabetes research are anticipated.