Biology subjects

Melton, D. A.

Publications and source records attributed to Melton, D. A..

4 recordsLinked to original sources

bioRxiv2024

IAPP Marks Mono-hormonal Stem-cell Derived β Cells that Maintain Stable Insulin Production in vitro and in vivo

Islet transplantation for treatment of diabetes is limited by availability of donor islets and requirements for immunosuppression. Stem cell-derived islets might circumvent these issues. SC-islets effectively control glucose metabolism post transplantation, but do not yet achieve full function in vitro with current published differentiation protocols. We aimed to identify markers of mature subpopulations of SC-{beta} cells by studying transcriptional changes associated with in vivo maturation of SC-{beta} cells using RNA-seq and co-expression network analysis. The {beta} cell-specific hormone islet amyloid polypeptide (IAPP) emerged as the top candidate to be such a marker. IAPP+ cells had more mature {beta} cell gene expression and higher cellular insulin content than IAPP- cells in vitro. IAPP+ INS+ cells were more stable in long-term culture than IAPP- INS+ cells and retained insulin expression after transplantation into mice. Finally, we conducted a small molecule screen to identify compounds that enhance IAPP expression. Aconitine up-regulated IAPP and could help to optimize differentiation protocols. HighlightsO_LIIAPP expression in vitro marks a mono-hormonal subpopulation of SC-{beta} cells excluding endocrine hormones other than insulin C_LIO_LIOnly INS+ IAPP+ cells maintain stable INS expression in vitro up to 100 days after differentiation C_LIO_LIThe small molecule aconitine accelerates IAPP expression in SC-{beta} cells in vitro C_LI

developmental biology↗

bioRxiv2022

Toward Improving Immunotolerance for Stem Cell-Derived Islets

Transplanting human stem cell-derived islets (SC-islets) is a promising therapy for insulin-dependent diabetes. While functional SC-islets have been produced for clinical application, immune rejection by the host remains a challenge. Present attempts, including chronic immunosuppression and/or physical encapsulation, have some disadvantages. Here we explore a strategy to induce an immune-tolerant environment based on the immune privilege observed in the male gonad. Sperm appears after the maturation of the immune system and development of systemic self-tolerance and the testis protects these autoreactive germ cells by the physical structure of blood-testis-barrier (BTB) and active local immunosuppression. Human SC-islets transplanted into the mouse testis can be physically protected by the BTB and we find that the testis secretes cytokines that induce a population of regulatory T cells (Tregs) that express both CD4 and CD8. We identified cytokines secreted by testis and used a cocktail of IL-2, IL-10, and TGF-{beta} for in vitro co-culture and in vivo transplantation demonstrating improved survival of SC-islets and the induction of Tregs. One Sentence SummaryInducing local immunotolerance by suppressive cytokines for islet Transplantation.

developmental biology↗

bioRxiv2019

Dissecting mechanisms of human islet differentiation and maturation through epigenome profiling

Investigating pancreatic islet differentiation from human stem cells in vitro provides a unique opportunity to dissect mechanisms that operate during human development in utero. We developed methods to profile DNA methylation, chromatin accessibility, and histone modifications from pluripotent stem cells to mature pancreatic islet cells, uncovering widespread epigenome remodeling upon endocrine commitment. Key lineage-defining loci are epigenetically primed before activation, foreshadowing cell fate commitment, and we show that priming of -cell-specific enhancers steers polyhormonal cells toward an -cell fate. We further dissect pioneer factors and core regulatory circuits across islet cell differentiation and maturation stages, which identify LMX1B as a key regulator of in vitro-derived endocrine progenitors. Finally, by contrasting maturing stem cell-derived to natural {beta}-cells, we discover that circadian metabolic cycles trigger rhythmic control of insulin synthesis and release and promote mature insulin responsiveness via an increased glucose threshold. These findings form a basis for understanding mechanisms orchestrating human islet cell specification and maturation.

genomics↗

bioRxiv2018

Identifying Gene Expression Programs of Cell-type Identity and Cellular Activity with Single-Cell RNA-Seq

Identifying gene expression programs underlying both cell-type identity and cellular activities (e.g. life-cycle processes, responses to environmental cues) is crucial for understanding the organization of cells and tissues. Although single-cell RNA-Seq (scRNA-Seq) can quantify transcripts in individual cells, each cells expression profile may be a mixture of both types of programs, making them difficult to disentangle. Here we illustrate and enhance the use of matrix factorization as a solution to this problem. We show with simulations that a method that we call consensus non-negative matrix factorization (cNMF) accurately infers identity and activity programs, including the relative contribution of programs in each cell. Applied to published brain organoid and visual cortex scRNA-Seq datasets, cNMF refines the hierarchy of cell types and identifies both expected (e.g. cell cycle and hypoxia) and intriguing novel activity programs. We propose that one of the novel programs may reflect a neurosecretory phenotype and a second may underlie the formation of neuronal synapses. We make cNMF available to the community and illustrate how this approach can provide key insights into gene expression variation within and between cell types.

bioinformatics↗