8 research outputs found
Single-cell transcriptome profiling reveals β cell maturation in stem cell-derived islets after transplantation
Human pluripotent stem cells differentiated to insulin-secreting β cells (SC-β cells) in islet organoids could provide an unlimited cell source for diabetes cell replacement therapy. However, current SC-β cells generated in vitro are transcriptionally and functionally immature compared to native adult β cells. Here, we use single-cell transcriptomic profiling to catalog changes that occur in transplanted SC-β cells. We find that transplanted SC-β cells exhibit drastic transcriptional changes and mature to more closely resemble adult β cells. Insulin and IAPP protein secretions increase upon transplantation, along with expression of maturation genes lacking with differentiation in vitro, including INS, MAFA, CHGB, and G6PC2. Other differentiated cell types, such as SC-α and SC-enterochromaffin (SC-EC) cells, also exhibit large transcriptional changes. This study provides a comprehensive resource for understanding human islet cell maturation and provides important insights into maturation of SC-β cells and other SC-islet cell types to enable future differentiation strategy improvements
SIX2 regulates human β cell differentiation from stem cells and functional maturation in vitro
Generation of insulin-secreting β cells in vitro is a promising approach for diabetes cell therapy. Human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) are differentiated to β cells (SC-β cells) and mature to undergo glucose-stimulated insulin secretion, but molecular regulation of this defining β cell phenotype is unknown. Here, we show that maturation of SC-β cells is regulated by the transcription factor SIX2. Knockdown (KD) or knockout (KO) of SIX2 in SC-β cells drastically limits glucose-stimulated insulin secretion in both static and dynamic assays, along with the upstream processes of cytoplasmic calcium flux and mitochondrial respiration. Furthermore, SIX2 regulates the expression of genes associated with these key β cell processes, and its expression is restricted to endocrine cells. Our results demonstrate that expression of SIX2 influences the generation of human SC-β cells in vitro
Multidimensional analysis and therapeutic development using patient iPSC-derived disease models of Wolfram syndrome
Wolfram syndrome is a rare genetic disorder largely caused by pathogenic variants in the WFS1 gene and manifested by diabetes mellitus, optic nerve atrophy, and progressive neurodegeneration. Recent genetic and clinical findings have revealed Wolfram syndrome as a spectrum disorder. Therefore, a genotype-phenotype correlation analysis is needed for diagnosis and therapeutic development. Here, we focus on the WFS1 c.1672C\u3eT, p.R558C variant, which is highly prevalent in the Ashkenazi Jewish population. Clinical investigation indicated that patients carrying the homozygous WFS1 c.1672C\u3eT, p.R558C variant showed mild forms of Wolfram syndrome phenotypes. Expression of WFS1 p.R558C was more stable compared with the other known recessive pathogenic variants associated with Wolfram syndrome. Human induced pluripotent stem cell-derived (iPSC-derived) islets (SC-islets) homozygous for WFS1 c.1672C\u3eT variant recapitulated genotype-related Wolfram syndrome phenotypes. Enhancing residual WFS1 function through a combination treatment of chemical chaperones mitigated detrimental effects caused by the WFS1 c.1672C\u3eT, p.R558C variant and increased insulin secretion in SC-islets. Thus, the WFS1 c.1672C\u3eT, p.R558C variant causes a mild form of Wolfram syndrome phenotypes, which can be remitted with a combination treatment of chemical chaperones. We demonstrate that our patient iPSC-derived disease model provides a valuable platform for further genotype-phenotype analysis and therapeutic development for Wolfram syndrome
Single-nucleus multi-omics of human stem cell-derived islets identifies deficiencies in lineage specification
Insulin-producing β cells created from human pluripotent stem cells have potential as a therapy for insulin-dependent diabetes, but human pluripotent stem cell-derived islets (SC-islets) still differ from their in vivo counterparts. To better understand the state of cell types within SC-islets and identify lineage specification deficiencies, we used single-nucleus multi-omic sequencing to analyse chromatin accessibility and transcriptional profiles of SC-islets and primary human islets. Here we provide an analysis that enabled the derivation of gene lists and activity for identifying each SC-islet cell type compared with primary islets. Within SC-islets, we found that the difference between β cells and awry enterochromaffin-like cells is a gradient of cell states rather than a stark difference in identity. Furthermore, transplantation of SC-islets in vivo improved cellular identities overtime, while long-term in vitro culture did not. Collectively, our results highlight the importance of chromatin and transcriptional landscapes during islet cell specification and maturation
Development and Maturation of Human Pluripotent Stem Cell Derived Pancreatic Islets
Cell replacement therapy for severe diabetes is a promising strategy to restore long term normoglycemia. With limited availability of cadaveric islets from compatible donors, cellular engineering via human pluripotent stem cells differentiation to insulin-secreting stem cell derived islets (SC-islets) provides a useful alternative as they can be generated in unlimited supply. Nonetheless, SC-islets generated with current protocols are not as well performing when compared to native human islets due to poor β cell yields and low mature β cell genes expression. Herein, I use single-cell technologies and bioinformatics to uncover mechanisms associated with pancreatic development to recapitulate β cell differentiations. Gene regulatory network modifications were done using soluble factors and gene editing tools to develop methodologies for generating mature islets from stem cells
Acquisition of Dynamic Function in Human Stem Cell-Derived β Cells
Summary: Recent advances in human pluripotent stem cell (hPSC) differentiation protocols have generated insulin-producing cells resembling pancreatic β cells. While these stem cell-derived β (SC-β) cells are capable of undergoing glucose-stimulated insulin secretion (GSIS), insulin secretion per cell remains low compared with islets and cells lack dynamic insulin release. Herein, we report a differentiation strategy focused on modulating transforming growth factor β (TGF-β) signaling, controlling cellular cluster size, and using an enriched serum-free media to generate SC-β cells that express β cell markers and undergo GSIS with first- and second-phase dynamic insulin secretion. Transplantation of these cells into mice greatly improves glucose tolerance. These results reveal that specific time frames for inhibiting and permitting TGF-β signaling are required during SC-β cell differentiation to achieve dynamic function. The capacity of these cells to undergo GSIS with dynamic insulin release makes them a promising cell source for diabetes cellular therapy. : In this study, Millman and colleagues report a differentiation strategy to generate β-like cells from human pluripotent stem cells with islet-like dynamic insulin release that rapidly reverses diabetes in mice. The authors elucidate that stage-specific control of TGF-β signaling during endocrine induction and maturation to be critical for robust function. Keywords: human embryonic stem cells, human induced pluripotent stem cells, diabetes, differentiation, glucose-stimulated insulin secretion, transplantation, cell therapy, β cells, pancreas, islet
Tailoring Nanostructure Morphology for Enhanced Targeting of Dendritic Cells in Atherosclerosis
Atherosclerosis,
a leading cause of heart disease, results from
chronic vascular inflammation that is driven by diverse immune cell
populations. Nanomaterials may function as powerful platforms for
diagnostic imaging and controlled delivery of therapeutics to inflammatory
cells in atherosclerosis, but efficacy is limited by nonspecific uptake
by cells of the mononuclear phagocytes system (MPS). MPS cells located
in the liver, spleen, blood, lymph nodes, and kidney remove from circulation
the vast majority of intravenously administered nanomaterials regardless
of surface functionalization or conjugation of targeting ligands.
Here, we report that nanostructure morphology alone can be engineered
for selective uptake by dendritic cells (DCs), which are critical
mediators of atherosclerotic inflammation. Employing near-infrared
fluorescence imaging and flow cytometry as a multimodal approach,
we compared organ and cellular level biodistributions of micelles,
vesicles (<i>i</i>.<i>e</i>., polymersomes), and
filomicelles, all assembled from poly(ethylene glycol)-<i>bl</i>-poly(propylene sulfide) (PEG-<i>bl</i>-PPS) block copolymers
with identical surface chemistries. While micelles and filomicelles
were respectively found to associate with liver macrophages and blood-resident
phagocytes, polymersomes were exceptionally efficient at targeting
splenic DCs (up to 85% of plasmacytoid DCs) and demonstrated significantly
lower uptake by other cells of the MPS. In a mouse model of atherosclerosis,
polymersomes demonstrated superior specificity for DCs (<i>p</i> < 0.005) in atherosclerotic lesions. Furthermore, significant
differences in polymersome cellular biodistributions were observed
in atherosclerotic compared to naïve mice, including impaired
targeting of phagocytes in lymph nodes. These results present avenues
for immunotherapies in cardiovascular disease and demonstrate that
nanostructure morphology can be tailored to enhance targeting specificity