Mapping the cord blood transcriptome of pregnancies affected by early maternal anemia to identify signatures of fetal programming

Objective Anemia during early pregnancy (EP) is common in developing countries and is associated with adverse health consequences for both mother and children. Offspring of women with EP anemia often have low birth-weight, the latter being a risk factor for cardiometabolic diseases including type 2 diabetes (T2D) later in life. Mechanisms underlying developmental programming of adult cardiometabolic disease include epigenetic and transcriptional alterations potentially detectable in umbilical cord blood (UCB) at time of birth. Methods We leveraged global transcriptome- and accompanying epigenome-wide changes in 48 UCB from newborns of EP-anemic Tanzanian mothers and 50 controls to identify differentially expressed genes (DEG) in UCB exposed to maternal EP-anemia. DEGs were assessed for association with neonatal anthropometry and cord insulin levels. These genes were further studied in expression data from human fetal pancreas and adult islets to understand their role in beta-cell development and/or function. Results The expression of 137 genes was altered in UCB of newborns exposed to maternal EP anemia. These putative signatures of fetal programming which included the birth-weight locus LCORL, were potentially mediated by epigenetic changes in 27 genes and associated with neonatal anthropometry. Among the DEGs were P2RX7, PIK3C2B, and NUMBL which potentially influence beta-cell development. Insulin levels were lower in EP anemia exposed UCB, supporting the notion of developmental programming of pancreatic beta-cell dysfunction and subsequently increased risk of T2D in offspring of EP anemic mothers. Conclusions Our data provide proof-of-concept on distinct transcriptional and epigenetic changes detectable in UCB from newborns exposed to maternal EP anemia.

A brief history of diabetes genetics: insights for pancreatic beta-cell development and function

Since the discovery of insulin 100 years ago, our knowledge and understanding of diabetes has grown exponentially. Specifically, with regards to the genetics underlying diabetes risk, our discoveries have paralleled developments in our understanding of the human genome and our ability to study genomics at scale; these advancements in genetics have both accompanied and led to those in diabetes treatment. This review will explore the timeline and history of gene discovery and how this has coincided with progress in the fields of genomics. Examples of genetic causes of monogenic diabetes are presented and the continuing expansion of allelic series in these genes and the challenges these now cause for diagnostic interpretation along with opportunities for patient stratification are discussed.

Loss of MANF Causes Childhood Onset Syndromic Diabetes due to Increased Endoplasmic Reticulum Stress

MANF is an endoplasmic reticulum resident protein that plays a crucial role in attenuating ER stress responses. Although MANF is indispensable for the survival and function of mouse beta cells, its precise role in human beta cell development and function is unknown. Herein, we show that lack of MANF in humans results in diabetes due to increased ER stress leading to impaired beta cell function. We identified two patients from different families with childhood diabetes and a neurodevelopmental disorder associated with homozygous loss-of-function mutations in the <i>MANF</i> gene. To study the role of MANF in human beta cell development and function, we knocked out the <i>MANF </i>gene in human embryonic stem cells and differentiated them into pancreatic endocrine cells. Loss of <i>MANF</i> induced mild ER stress and impaired insulin processing capacity of beta cells <i>in vitro</i>. Upon implantation to immunocompromised mice, the MANF knockout grafts presented elevated ER stress and functional failure, particularly in diabetic recipients. By describing a new form of monogenic neurodevelopmental diabetes syndrome caused by disturbed ER function, we highlight the importance of adequate ER stress regulation for proper human beta cell function and demonstrate the crucial role of MANF in this process.

Loss of MANF Causes Childhood Onset Syndromic Diabetes due to Increased Endoplasmic Reticulum Stress

MANF is an endoplasmic reticulum resident protein that plays a crucial role in attenuating ER stress responses. Although MANF is indispensable for the survival and function of mouse beta cells, its precise role in human beta cell development and function is unknown. Herein, we show that lack of MANF in humans results in diabetes due to increased ER stress leading to impaired beta cell function. We identified two patients from different families with childhood diabetes and a neurodevelopmental disorder associated with homozygous loss-of-function mutations in the <i>MANF</i> gene. To study the role of MANF in human beta cell development and function, we knocked out the <i>MANF </i>gene in human embryonic stem cells and differentiated them into pancreatic endocrine cells. Loss of <i>MANF</i> induced mild ER stress and impaired insulin processing capacity of beta cells <i>in vitro</i>. Upon implantation to immunocompromised mice, the MANF knockout grafts presented elevated ER stress and functional failure, particularly in diabetic recipients. By describing a new form of monogenic neurodevelopmental diabetes syndrome caused by disturbed ER function, we highlight the importance of adequate ER stress regulation for proper human beta cell function and demonstrate the crucial role of MANF in this process.

NKX6.1 Transcription Factor: A Crucial Regulator of Pancreatic β-Cell Development, Identity, and Proliferation

Understanding the biology underlying the mechanisms and pathways regulating pancreatic &beta;-cell development is necessary to understand the pathology of diabetes mellitus (DM), which is characterized by the progressive reduction in insulin producing &beta;-cell mass. Pluripotent stem cells (PSCs) can potentially offer an unlimited supply of functional &beta;-cells for cellular therapy and disease modeling of DM. Homeobox protein NKX6.1 is a transcription factor (TF) that plays a critical role in pancreatic &beta;-cell function and proliferation. In human pancreatic islet, NKX6.1 expression is exclusive to&beta;-cells and is undetectable in other islet cells. Several reports showed that activation of NKX6.1 in PSC-derived pancreatic progenitors (MPCs), expressing PDX1 (PDX1+/NKX6.1+), warrants their future commitment to monohormonal &beta;-cells. However, further differentiation of MPCs lacking NKX6.1 expression (PDX1+/NKX6.1-) results in an undesirable generation of non-functional polyhormonal &beta;-cells. The importance of NKX6.1 as a crucial regulator in MPC specification into functional &beta;-cells directs attentions to further investigating its mechanism and enhancing NKX6.1 expression as a mean to increase &beta;-cell function and mass. Here, we shed light on the role of NKX6.1 during pancreatic &beta;-cell development and in directing the MPCs to functional monohormonal lineage. Furthermore, we address the transcriptional mechanisms and targets of NKX6.1 as well as its association with diabetes.

Activation of pancreatic β-cell genes by multiplex epigenetic CRISPR-editing

ABSTRACTCRISPR-based systems for epigenetic editing are promising molecular tools that could be harnessed for directed differentiation of pluripotent stem cells. We used the CRISPR/dCas9-VP160, CRISPR/dCas9-TET1 and CRISPR/dCas9-P300 systems for multiplex epigenetic editing and activation of human beta pancreatic genes (PDX1, NEUROG3, PAX4 and INS). The CRISPR/dCas9-P300 system was the most effective at activating genes with reduced number of sgRNA. Using small number of sgRNA per gene was important to induce multiplex gene activation. Combined activation of transcription factors (TFs) involved in beta cell development resulted in INS gene expression; in which sequential TFs activation was more effective than simultaneous activation. Full CRISPR RNA-based delivery system was able to activate all targeted genes. Overall, this study shows the utility of CRISPR tools for epigenetic editing and directed cellular differentiation.

NKX6.1 Transcription Factor: A Crucial Regulator of Pancreatic β-Cell Development, Identity, and Proliferation

Understanding the biology underlying the mechanisms and pathways regulating pancreatic &beta;-cell development is necessary to understand the pathology of diabetes mellitus (DM), which is characterized by the progressive reduction in insulin producing &beta;-cell mass. Pluripotent stem cells (PSCs) can potentially offer an unlimited supply of functional &beta;-cells for cellular therapy and disease modeling of DM. Homeobox protein NKX6.1 is a transcription factor (TF) that plays a critical role in pancreatic &beta;-cell function and proliferation. In human pancreatic islet, NKX6.1 expression is exclusive to&beta;-cells and is undetectable in other islet cells. Several reports showed that activation of NKX6.1 in PSC-derived pancreatic progenitors (MPCs), expressing PDX1 (PDX1+/NKX6.1+), warrants their future commitment to monohormonal &beta;-cells. However, further differentiation of MPCs lacking NKX6.1 expression (PDX1+/NKX6.1-) results in an undesirable generation of non-functional polyhormonal &beta;-cells. The importance of NKX6.1 as a crucial regulator in MPC specification into functional &beta;-cells directs attentions to further investigating its mechanism and enhancing NKX6.1 expression as a mean to increase &beta;-cell function and mass. Here, we shed light on the role of NKX6.1 during pancreatic &beta;-cell development and in directing the MPCs to functional monohormonal lineage. Furthermore, we address the transcriptional mechanisms and targets of NKX6.1 as well as its association with diabetes.