Biology subjects

Isaacson, A.

Publications and source records attributed to Isaacson, A..

4 recordsLinked to original sources

bioRxiv2024

Generation of human iPSC-derived pancreatic organoids to study pancreas development and disease

The pancreas has vital endocrine and exocrine functions that can be affected by life-threatening diseases such as diabetes and pancreatic cancer. While animal models have been essential for understanding pancreatic development and disease, they are limited by their low-throughput and major species-specific molecular and physiological differences. Generating 3D in vitro models, such as organoids, that are physiologically relevant is thus primordial for investigating pancreatic development and disease in the human context. However, producing human stem cell-derived pancreatic organoids with proper branched architecture and correct patterning of cell domains has remained challenging. Here, we successfully developed a protocol that efficiently and reproducibly generates such organoids from human induced pluripotent stem cells (hiPSCs) by optimising organoid culture format and media. Our differentiation protocol promotes acinar cell differentiation and generates organoids with branches patterned into central trunk and peripheral tip domains, without relying on animal-derived matrices for organoid culture. This platform opens the door to high-throughput investigations of human pancreas development in a system recapitulating the most important aspects of pancreatic tissue architecture. Lastly, we anticipate that this system will contribute to the partial replacement of animal models used to investigate diseases such as pancreatic cancer.

developmental biology↗

bioRxiv2023

Heterozygous missense variant in GLI2 impairs human endocrine pancreas development

Missense variants are the most common type of coding genetic variants. Their functional assessment is fundamental for defining any implication in human diseases and may also uncover genes that are essential for human organ development. Here, we applied CRISPR-Cas9 gene editing on human iPSCs to study a heterozygous missense variant in GLI2 identified in a paediatric hyperglycaemic patient and family members. GLI2 is a primary mediator of the Hedgehog pathway, which regulates pancreatic {beta}-cell development in mice. However, neither mutations in GLI2 nor Hedgehog dysregulation have been reported as cause or predisposition to diabetes. A set of isogenic iPSC lines harbouring the missense variant were studied for their ability to differentiate into pancreatic {beta}-like cells. Interestingly, iPSCs carrying the missense variant showed altered GLI2 transcriptional activity and impaired endocrine differentiation. Collectively, our findings underscore an essential role for GLI2 in human endocrine development and identify a novel variant that predisposes to diabetes.

developmental biology↗

bioRxiv2022

A cell circuit approach to dissect fibroblast-macrophage interactions in the tumor microenvironment

The tumor microenvironment (TME) is composed of various nonmalignant cell types that interact with each other and with cancer cells, impacting all aspects of cancer biology. The TME is complex and heterogeneous, and thus simplifying systems and concepts are needed. Here we provide a tractable experimental system and powerful mathematical circuit concepts to identify the main molecular interactions that govern the composition of the TME. We focus on two major components of the TME - cancer associated fibroblasts (CAFs) and tumor associated macrophages (TAMs), define their interactions and verify our predictions in mouse and human breast cancer. We measure the population dynamics starting from many initial conditions of co-cultures of macrophages and organ-derived fibroblasts from mammary, lung, and fat, and explore the effects of cancer-conditioned medium on the circuits. We define the circuits and their inferred parameters from the data using a mathematical approach, and quantitatively compare the cell circuits in each condition. We find that while the homeostatic steady-states are similar between the organs, the cancer-conditioned medium profoundly changes the circuit. Fibroblasts in all contexts depend on autocrine secretion of growth factors whereas macrophages are more dependent on external cues, including paracrine growth factors secreted from fibroblasts and cancer cells. Transcriptional profiling reveals the molecular underpinnings of the cell circuit interactions and the primacy of the fibroblast autocrine loop. The same fibroblast growth factors are shared by the co-cultures and mouse and human breast cancer. The cell circuit approach thus provides a quantitative account of cell interactions in the cancer microenvironment.

cancer biology↗

bioRxiv2019

Single-cell RNA-seq of differentiating iPS cells reveals dynamic genetic effects on gene expression

Recent developments in stem cell biology have enabled the study of cell fate decisions in early human development that are impossible to study in vivo. However, understanding how development varies across individuals and, in particular, the influence of common genetic variants during this process has not been characterised. Here, we exploit human iPS cell lines from 125 donors, a pooled experimental design, and single-cell RNA-sequencing to study population variation of endoderm differentiation. We identify molecular markers that are predictive of differentiation efficiency, and utilise heterogeneity in the genetic background across individuals to map hundreds of expression quantitative trait loci that influence expression dynamically during differentiation and across cellular contexts.

genomics↗