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Biology subjects

Shapiro, A. J.

Publications and source records attributed to Shapiro, A. J..

4 recordsLinked to original sources

Using the ancestral recombination graph to study the history of rare variants in founder populations

Gene genealogies represent the ancestry of a sample and are often encoded as ancestral recombination graphs (ARG). It has recently become possible to infer these gene genealogies from sequencing or genotyping data and use them for evolutionary and statistical genetics. Unfortunately, inferred gene genealogies can be noisy and subject to biases, making their applications more challenging. This project aims to study the application of ARG methods to systematically impute and trace the transmission of all disease variants in founder populations where long-shared haplotypes allow for accurate timing of relatedness. We applied these methods to the population of Quebec, where multiple founder events led to an uneven distribution of pathogenic variants across regions and where extensive population pedigrees are available. We validated our approach with nine founder mutations for the SLSJ region, demonstrating high accuracy for mutation age, imputation, and regional frequency estimation. Moreover, we showed that this subset of high-quality carriers is sufficient to capture previously described associations with pathogenic variants in the LPL gene. This method systematically characterizes rare variants in founder populations, establishing a fast and accurate approach to inform genetic screening programs.

genetics↗

Age-Related Cerebral Ventriculomegaly occurs in Patients with Primary Ciliary Dyskinesia

Primary ciliary dyskinesia (PCD) is a genetic disorder causing motile ciliary dysfunction primarily affecting the respiratory and reproductive systems. However, the impact of PCD on the central nervous system, through dysfunction of motile cilia in multiciliated ependymal cells, remains poorly understood. We hypothesized that patients with PCD exhibit sub-clinical ventriculomegaly due to ependymal ciliary dysfunction, which may influence neuropsychiatric diagnoses. We demonstrated highly specific expression levels of known PCD-related genes in human brain ependymal cells (p<0.0001), supporting their potential role in regulating ependymal ciliary function. Computed tomography sinus images from patients with PCD (n=33) and age/sex-matched controls (n=64) were analysed. Patients with PCD displayed significantly larger ventricular areas (p<0.0001) and Evans index (p<0.01), indicating ventriculomegaly that was consistent across all genetic subgroups. Ventricular enlargement correlated positively with increasing age in patients with PCD compared to controls (p<0.001). Chart review demonstrated a high prevalence (39%) of neuropsychiatric/neurological disorders in adult PCD patients that did not correlate with degree of ventriculomegaly. Our findings suggest that patients with PCD may have unrecognized, mild ventriculomegaly potentially due to ependymal ciliary dysfunction which correlates with ageing. Further study is required to determine if ventricular enlargement contributes to neuropsychiatric/neurological or other morbidity in PCD.

neuroscience↗

Complete Suspension Differentiation of Human Pluripotent Stem Cells into Pancreatic Islets Using Vertical-Wheel Bioreactors

Advanced protocols to produce human pluripotent stem cell (SC)-derived islets show promise in functional, metabolic, and transcriptional maturation of cell therapy product to treat diabetes. Available protocols are either developed as complete planar (2D) or, in later stages, combined with suspension cultures (3D). Despite marked progress, both approaches have clear limitations for scalability, cell loss and batch to batch heterogeneity during differentiation. Using a Vertical Wheel(R) bioreactor system, we present a highly efficient and scalable complete suspension protocol across all stages for directed differentiation of human pluripotent stem cells into functional pancreatic islets. Here, we generate homogeneous, metabolically functional, and transcriptionally enriched SC-islets and compared against adult donor islets. Generated SC-islets showed enriched endocrine cell composition ([~]63% CPEP+NKX6.1+ISL1+) and displayed functional maturity for glucose stimulated insulin secretion ([~]5-fold) during in vitro and post transplantation. Comprehensive stage-specific single-cell mass flow cytometry characterization with dimensional reduction analysis at stage-4 and -6 confirmed optimal maturation was achieved without heterogeneity. Notably, by 16-weeks transplantation follow-up, normal glycemic homeostasis was restored, and glucose responsive human c-peptide secretion response (2-fold) was achieved. Four months post engraftment, graft-harvested single cells displayed islet hormonal cell composition with flow cytometry, improved functional maturity by in vivo glucose-stimulated insulin secretion (GSIS) and enhanced transcriptional landscape with real-time expression that closely resembled patterns comparable to adult human islets. Our comprehensive evaluation of a complete suspension method applied across all stages using Vertical Wheel(R) bioreactors for SC-islets generation highlight progressive molecular and functional maturation of islets while reducing potential cell loss and cellular heterogeneity. Such a system could potentially be scaled to deliver clinical grade SC-islet products in a closed good manufacturing practice type environment. One Sentence SummaryThis study describes all-stages complete suspension protocol for SC-islets generation.

bioengineering↗

Comprehensive human cell-type methylation atlas reveals origins of circulating cell-free DNA in health and disease

Methylation patterns of circulating cell-free DNA (cfDNA) contain rich information about recent cell death events in the body. Here, we present an approach for unbiased determination of the tissue origins of cfDNA, using a reference methylation atlas of 25 human tissues and cell types. The method is validated using in silico simulations as well as in vitro mixes of DNA from different tissue sources at known proportions. We show that plasma cfDNA of healthy donors originates from white blood cells (55%), erythrocyte progenitors (30%), vascular endothelial cells (10%) and hepatocytes (1%). Deconvolution of cfDNA from patients reveals tissue contributions that agree with clinical findings in sepsis, islet transplantation, cancer of the colon, lung, breast and prostate, and cancer of unknown primary. We propose a procedure which can be easily adapted to study the cellular contributors to cfDNA in many settings, opening a broad window into healthy and pathologic human tissue dynamics.

molecular biology↗