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Heme-Induced Glycolytic Imbalance Contributes Towards Endothelial Cell Proliferation

Ruslan Rafikov

Anup Srivastava

Cody Eccles

Olga Rafikova


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Conference: 2018 PHA International PH Conference & Scientific Sessions

Release Date: 07.28.2018

Presentation Type: Abstracts

File Download: Conference 2018_1006

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2018 International PH Conference and Scientific SeAbstract presented at the 2018 International PH Conference and Scientific Sessions in Orlando, Fla., June 28-July 1, 2018.


An increasing number of studies have implicated glycolytic switch in the pathology of pulmonary arterial hypertension (PAH). Although, this “Warburg-like” effect is well reported for PAH patients and is well characterized in pre-clinical studies, there are no mechanistic insights on why this metabolic switch occurs. Decreased mitochondrial respiration and activation of HIF signaling have been shown to be associated with the metabolic switching in PAH. We have found elevated free hemoglobin (Hb) levels in PAH patients and animal models of PAH. Interestingly; our data indicates that free heme, a degradation product of free Hb, can directly affect the glycolytic metabolism in endothelial cells (EC).


Human lung microvascular endothelial cells (HLMVECs) were utilized to study heme-mediated effects on glycolytic pathway. We analysed the glycolytic function using Seahorse Extracellular Flux analyzer and the glycolytic enzyme activities were measured using commercially available kits. Cell index measured by the xCELLigence system (Acea) was used for proliferation assays.


Heme induced strong inhibition of glycolysis in HLMVEC after 1h of treatment (Figure 1A). Our data showed that on one hand activation of phosphofructokinase (PFK) (Figure 1C) and on the other inhibition of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (Figure 1D) after 1h of heme treatment. PFK catalyzed formation of fructose 1,6-bisphosphate is considered the rate-limiting step of glycolysis. Therefore, under the influence of heme, the glycolytic rate limiting step in endothelial cells is reassigned to GAPDH (Figure 1B). Further, our data indicates an increased activity of GAPDH after 24h of heme incubation. Thus, we believe that this could be the possible mechanism by which heme causes this shift in cellular metabolism toward glycolysis. Moreover, the rate of endothelial cell proliferation exhibited two-phase effect of the heme treatment, with growth inhibition in the acute phase (0-12h) and increased proliferation after 48h.


Our results indicate involvement of free heme in the endothelial glycolytic pathway. Further studies on identifying the role of heme signaling on the glycolytic shift could potentially lead to identification of new targets for vascular remodeling in PAH.

Heme induced glycolytic imbalance in EC

Figure 1: Heme induced glycolytic imbalance in EC.

A) heme induced glycolysis inhibition B) Bottle neck of glycolysis C) PFK activation D) GAPDH inhibition