Induced pluripotent stem cells can utilize lactate as a metabolic substrate to support proliferation

Abstract

Human-induced pluripotent stem cells (iPSCs) hold the promise to improve cell-based therapies. Yet, to meet rising demands and become clinically impactful, sufficient high-quality iPSCs in quantity must be generated, a task that exceeds current capabilities. In this study, K3 iPSCs cultures were examined using parallel-labeling metabolic flux analysis ( 13 C-MFA) to quantify intracellular fluxes at relevant bioprocessing stages: glucose concentrations representative of initial media concentrations and high lactate concentrations representative of fed-batch culture conditions, prior to and after bolus glucose feeds. The glucose and lactate concentrations are also representative of concentrations that might be encountered at different locations within 3D cell aggregates. Furthermore, a novel method was developed to allow the isotopic tracer [U-13 C 3 ] lactate to be used in the 13 C-MFA model. The results indicated that high extracellular lactate concentrations decreased glucose consumption and lactate production, while glucose concentrations alone did not affect rates of aerobic glycolysis. Moreover, for the high lactate cultures, lactate was used as a metabolic substrate to support oxidative mitochondrial metabolism. These results demonstrate that iPSCs have metabolic flexibility and possess the capacity to metabolize lactate to support exponential growth, and that high lactate concentrations alone do not adversely impact iPSC proliferation.

1H NMR Metabolite Monitoring during the Differentiation of Human Induced Pluripotent Stem Cells Provides New Insights into the Molecular Events That Regulate Embryonic Chondrogenesis

“…The IPSC 1 H NMR metabolic profile was consistent with the established understanding of metabolic flexibility demonstrated by pluripotent stem cells. High levels of pyruvate and lactate were identified in both the intracellular and media components of pluripotent cultures, underpinning their glycolytic phenotype [ 22 , 24 ]. Lactate, conventionally regarded as a waste product of aerobic glycolysis, is now also proposed to be a metabolic substrate through the recycling of carbon into anabolic pathways that generate cellular biomass required for self-renewal [ 24 ].…”

“…High levels of pyruvate and lactate were identified in both the intracellular and media components of pluripotent cultures, underpinning their glycolytic phenotype [ 22 , 24 ]. Lactate, conventionally regarded as a waste product of aerobic glycolysis, is now also proposed to be a metabolic substrate through the recycling of carbon into anabolic pathways that generate cellular biomass required for self-renewal [ 24 ]. We hypothesise that the future interrogation of lactate flux by 1 H NMR metabolomics would elucidate a novel mechanism for regulating pluripotency.…”

“…Pluripotent stem cells preferentially source energy requirements from aerobic glycolysis, whereby pyruvate is diverted away from entry into the TCA cycle by reduction to the endpoint metabolite lactate [ 6 , 17 , 18 , 19 ]. Commonly referred to as the Warburg effect [ 17 , 20 , 21 ], aerobic glycolysis was first identified as a phenomenon of cancer cell metabolism and is now established as a core metabolic pathway in highly proliferative cell populations, including pluripotent stem cells and pre-implantation embryos [ 18 , 22 , 23 , 24 ]. While glycolysis is energetically less efficient than oxidative phosphorylation, it occurs more rapidly when glucose is present in excess and results in an overall greater production of ATP per unit of time [ 25 , 26 ].…”

Metabolic regulatory network kinetic modeling with multiple isotopic tracers for iPSCs

“…given its pivotal role in stem cell proliferation, biosynthesis, and overall functionality. To estimate model parameters, this model utilized experimentally obtained data from well-designed monolayer K3 iPSC cultures, as described in the study (Odenwelder et al, 2021) Briefly, the data used to estimate the model parameters were derived from an experimental study of K3 iPSCs, where monolayer cultures were conducted in 6-well plates and 60 mm cell culture dishes (Odenwelder et al, 2021). Furthermore, 2.75 mM glutamine was included in each condition.…”

“…L-glutamine, and [U-C 13 3 ] sodium L-lactate (when lactate was added to the culture media). The time-course measurements, as illustrated in Figure 1, included the transition to metabolic and isotopic steady states (Odenwelder et al, 2021). These measurements were used to develop the metabolic regulatory network kinetic model for iPSC cultures.…”