The function of metabolic state in stemness is poorly understood. Mouse embryonic stem cells (ESC) and epiblast stem cells (EpiSC) are at distinct pluripotent states representing the inner cell mass (ICM) and epiblast embryos. Human embryonic stem cells (hESC) are similar to EpiSC stage. We now show a dramatic metabolic difference between these two stages. EpiSC/hESC are highly glycolytic, while ESC are bivalent in their energy production, dynamically switching from glycolysis to mitochondrial respiration on demand. Despite having a more developed and expanding mitochondrial content, EpiSC/hESC have low mitochondrial respiratory capacity due to low cytochrome c oxidase (COX) expression. Similarly, in vivo epiblasts suppress COX levels. These data reveal EpiSC/ hESC functional similarity to the glycolytic phenotype in cancer (Warburg effect). We further show that hypoxiainducible factor 1a (HIF1a) is sufficient to drive ESC to a glycolytic Activin/Nodal-dependent EpiSC-like stage. This metabolic switch during early stem-cell development may be deterministic.

The naïve pluripotent state has been shown in mice to lead to broad and more robust developmental potential relative to primed mouse epiblast cells. The human naïve ES cell state has eluded derivation without the use of transgenes, and forced expression of OCT4, KLF4, and KLF2 allows maintenance of human cells in a naïve state [Hanna J, et al. (2010) Proc Natl Acad Sci USA 107 (20):9222-9227]. We describe two routes to generate nontransgenic naïve human ES cells (hESCs). The first is by reverse toggling of preexisting primed hESC lines by preculture in the histone deacetylase inhibitors butyrate and suberoylanilide hydroxamic acid, followed by culture in MEK/ERK and GSK3 inhibitors (2i) with FGF2. The second route is by direct derivation from a human embryo in 2i with FGF2. We show that human naïve cells meet mouse criteria for the naïve state by growth characteristics, antibody labeling profile, gene expression, X-inactivation profile, mitochondrial morphology, microRNA profile and development in the context of teratomas. hESCs can exist in a naïve state without the need for transgenes. Direct derivation is an elusive, but attainable, process, leading to cells at the earliest stage of in vitro pluripotency described for humans. Reverse toggling of primed cells to naïve is efficient and reproducible.

Low oxygen levels have shown to promote self-renewal in many stem cells. In tumors, hypoxia is associated with aggressive disease course and poor clinical outcomes. Furthermore, many aggressive tumors have shown to display gene expression signatures characteristic of human embryonic stem cells (hESC). We now tested whether hypoxia might be responsible for the hESC signature observed in aggressive tumors. We show that hypoxia, through hypoxia inducible factor (HIF), can induce a hESC-like transcriptional program, including the iPSC inducers, OCT4, NANOG, SOX2, KLF4, cMYC and miRNA-302 in eleven cancer cell lines (from prostate, brain, kidney, cervix, lung, colon, liver and breast tumors). Further, non-degradable forms of HIFα, combined with the traditional iPSC inducers are highly efficient in generating A549 iPSC-like colonies that have high tumorigenic capacity. To test potential correlation between iPSC inducers and HIF expression in primary tumors, we analyzed primary prostate tumors and found a significant correlation between NANOG-, OCT4- and HIF1α-positive regions. Further, NANOG and OCT4 expression positively correlated with increased prostate tumor Gleason score. In primary glioma-derived CD133 negative cells neurospheres and hESC markers were induced in hypoxia but not in normoxia. Together, these findings suggest that HIF targets may act as key inducers of a dynamic state of stemness in pathological conditions.

Type 2 diabetes mellitus (T2D) is a growing health problem, but little is known about its early disease stages, its effects on biological processes or the transition to clinical T2D. To understand the earliest stages of T2D better, we obtained samples from 106 healthy individuals and individuals with prediabetes over approximately four years and performed deep profiling of transcriptomes, metabolomes, cytokines, and proteomes, as well as changes in the microbiome. This rich longitudinal data set revealed many insights: first, healthy profiles are distinct among individuals while displaying diverse patterns of intra- and/or inter-personal variability. Second, extensive host and microbial changes occur during respiratory viral infections and immunization, and immunization triggers potentially protective responses that are distinct from responses to respiratory viral infections. Moreover, during respiratory viral infections, insulin-resistant participants respond differently than insulin-sensitive participants. Third, global co-association analyses among the thousands of profiled molecules reveal specific host–microbe interactions that differ between insulin-resistant and insulin-sensitive individuals. Last, we identified early personal molecular signatures in one individual that preceded the onset of T2D, including the inflammation markers interleukin-1 receptor agonist (IL-1RA) and high-sensitivity C-reactive protein (CRP) paired with xenobiotic-induced immune signalling. Our study reveals insights into pathways and responses that differ between glucose-dysregulated and healthy individuals during health and disease and provides an open-access data resource to enable further research into healthy, prediabetic and T2D states.

A new wave of portable biosensors allows frequent measurement of health-related physiology. We investigated the use of these devices to monitor human physiological changes during various activities and their role in managing health and diagnosing and analyzing disease. By recording over 250,000 daily measurements for up to 43 individuals, we found personalized circadian differences in physiological parameters, replicating previous physiological findings. Interestingly, we found striking changes in particular environments, such as airline flights (decreased peripheral capillary oxygen saturation [SpO2] and increased radiation exposure). These events are associated with physiological macro-phenotypes such as fatigue, providing a strong association between reduced pressure/oxygen and fatigue on high-altitude flights. Importantly, we combined biosensor information with frequent medical measurements and made two important observations: First, wearable devices were useful in identification of early signs of Lyme disease and inflammatory responses; we used this information to develop a personalized, activity-based normalization framework to identify abnormal physiological signals from longitudinal data for facile disease detection. Second, wearables distinguish physiological differences between insulin-sensitive and -resistant individuals. Overall, these results indicate that portable biosensors provide useful information for monitoring personal activities and physiology and are likely to play an important role in managing health and enabling affordable health care access to groups traditionally limited by socioeconomic class or remote geography.

Precision health relies on the ability to assess disease risk at an individual level, detect early preclinical conditions and initiate preventive strategies. Recent technological advances in omics and wearable monitoring enable deep molecular and physiological profiling and may provide important tools for precision health. We explored the ability of deep longitudinal profiling to make health-related discoveries, identify clinically relevant molecular pathways, and impact behavior in a prospective longitudinal cohort ( n = 109) enriched for risk of type 2 diabetes mellitus (DM). The cohort underwent integrative Personalized Omics Profiling (iPOP) from samples collected quarterly for up to 8 years (median 2.8 years) using clinical measures and emerging technologies including genome, immunome, transcriptome, proteome, metabolome, microbiome, and wearable monitoring. We discovered over 67 clinically actionable health discoveries and identified multiple molecular pathways associated with metabolic, cardiovascular and oncologic pathophysiology. We developed prediction models for insulin resistance using omics measurements illustrating their potential to replace burdensome tests. Finally, study participation lead the majority of participants to implement diet and exercise changes. Altogether, we conclude that deep longitudinal profiling can lead to actionable health discoveries and provide relevant information for precision health.

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SUMMARY Pluripotent stem cells have distinct metabolic requirements, and reprogramming cells to pluripotency requires a shift from oxidative to glycolytic metabolism. Here, we show that this shift occurs early during reprogramming of human cells and requires Hypoxia Inducible Factors in a stage-specific manner. HIF1α and HIF2α are both necessary to initiate this metabolic switch and for acquisition of pluripotency, and stabilization of either protein during early phases of reprogramming is sufficient to induce the switch to glycolytic metabolism. In contrast, stabilization of HIF2α during later stages represses reprogramming, due at least in part to up-regulation of TNF-related apoptosis-inducing ligand (TRAIL). TRAIL inhibits iPSC generation by repressing apoptotic caspase 3 activity specifically in cells undergoing reprogramming, but not hESCs, and inhibiting TRAIL activity enhances hiPSC generation. These results shed light on the mechanisms underlying the metabolic shifts associated with acquisition of a pluripotent identity during reprogramming.