Impact of cell-free hemoglobin on skeletal muscle vascular and metabolic control
Research Summary: Sickle cell disease (SCD) is a complex clinical syndrome that, in a 1-year period in the United States alone, results in ~25,000 hospitalizations costing ~$160 million (2). Decreased exercise capacity and quality of life in these patients is due to both cardiopulmonary and peripheral vascular factors that conspire to reduce maximal oxygen uptake (VO2max) and instill premature fatigue during exercise (1). While central cardiopulmonary abnormalities contribute to this decrement, understanding the effects of hemolysis and the resulting contribution of hemoglobin (Hb)-mediated peripheral vascular abnormalities on exercise performance is of paramount importance for restoring the quality of life in SCD patients.
Furthermore, despite the well-established benefits of exercise training on cardiovascular health, there is considerable debate amongst clinicians whether patients with SCD should participate in regular physical activity as prolonged exercise may induce metabolic changes that can precipitate vaso-occlusive crises.
Consequently, exercise prescriptions in SCD are often imprecise (when given at all) due to the lack of substantial evidence regarding the mechanisms of exercise intolerance and effects of exercise training on SCD morbidities (5). The inability to participate in regular exercise and activities of daily living places an enormous psychosocial burden on this patient population (8), therefore understanding the mechanistic underpinnings of SCD associated cardiopulmonary abnormalities as they relate to exercise tolerance is crucial to resolving the current discrepancies in clinical recommendations.
Nitric oxide (NO) scavenging caused by Hb is a primary mechanism of vasculopathy and organ damage in SCD due to exacerbated platelet activation and cell adhesion (4, 6) and these effects likely disrupt skeletal muscle capillary hemodynamics and metabolic control. Conversely, a plethora of investigations have demonstrated the robust efficacy of dietary nitrate (NO3–) supplementation in the treatment of diseases associated with reduced NO bioavailability (3, 7); accordingly, it has been hailed as an “unrecognized nutrient.” During my post-doctoral training, I established a detrimental role of free Hb on muscle function and relied on in vivo techniques to help facilitate the bench-to-bedside research transition of our work. In this capacity, our laboratory has generated substantial evidence suggesting that cell-free hemoglobin plays a deterministic role in skeletal muscle vascular and metabolic control during exercise and that dietary NO3– supplementation improves exercise tolerance in mice with SCD.
These results strongly support the premise that dietary NO3– will attenuate Hb-mediated NO depletion in patients with SCD, and therefore reduce vascular crises, improve vascular and metabolic function, and evoke improved quality of life. Despite this intriguing potential, there are few investigations into the effects of SCD on skeletal muscle vascular and metabolic control during contractions or the therapeutic potential of a NO3– therapy to restore functionality in a preclinical or SCD patient population. In addition to other contributors of vascular dysfunction in SCD, we hypothesize that NO scavenging of cell-free Hb is a key mechanistic driver in the disruption of skeletal muscle vascular and microvascular control during exercise and that a NO3– based therapy, alone and in combination with moderate-intensity exercise training, will improve SCD sequelae. Aim 1 and 2 will be addressed during the INBRE pilot funding period while Aim 3 will be included in future NIH R-level (or equivalent) funding proposals.
Aim 1: Determine the impacts of cell-free hemoglobin (Hb) on the skeletal muscle microvascular O2 delivery-utilization (O2) balance (PO2mv, the determinant of blood-myocyte O2 flux) during skeletal muscle contractions and examine if the peripheral (e.g., skeletal muscle) effects of Hb are due to impaired nitric oxide (NO) bioavailability.
Using a rat model of hemolysis, we will determine if acute and chronic exposure to Hb impacts muscle NO biology and lowers muscle PO2mv at rest and during contractions in fast vs. slow-twitch skeletal muscle.
Aim 2: Determine the impacts of dietary nitrate supplementation via beetroot juice on the skeletal muscle PO2mv following exposure to acute hemolysis.
We will determine if dietary nitrate supplementation can ameliorate the cell-free hemoglobin induced reductions in the skeletal muscle PO2mv during muscle contractions in vivo. We will also determine if the beneficial impacts of dietary nitrate supplementation are muscle fiber-type specific, which has been the case in humans.
References
1. Anthi A, Machado RF, Jison ML, Taveira-Dasilva AM, Rubin LJ, Hunter L, Hunter CJ, Coles W, Nichols J, Avila NA, Sachdev V, Chen CC, and Gladwin MT. Hemodynamic and functional assessment of patients with sickle cell disease and pulmonary hypertension. American journal of respiratory and critical care medicine 175: 1272-1279, 2007.
2. Ashley-Koch A, Yang Q, and Olney RS. Sickle hemoglobin (HbS) allele and sickle cell disease: a HuGE review. Am J Epidemiol 151: 839-845, 2000.
3. Ashworth A, Bailey SJ, Hayward GM, DiMenna F, Vanhatalo A, and Jones AM. Dietary nitrate – An unrecognized nutrient? Clin Nutr ESPEN 10: e201, 2015.
4. Azuma H, Ishikawa M, and Sekizaki S. Endothelium‐dependent inhibition of platelet aggregation. British journal of pharmacology 88: 411-415, 1986.
5. Connes P, Machado R, Hue O, and Reid H. Exercise limitation, exercise testing and exercise recommendations in sickle cell anemia. Clinical hemorheology and microcirculation 49: 151-163, 2011.
6. Loscalzo J. Nitric oxide insufficiency, platelet activation, and arterial thrombosis. Circulation research 88: 756-762, 2001.
7. Lundberg JO, Weitzberg E, and Gladwin MT. The nitrate-nitrite-nitric oxide pathway in physiology and therapeutics. Nat Rev Drug Discov 7: 156-167, 2008.
8. Thomas V, and Taylor L. The psychosocial experience of people with sickle cell disease and its impact on quality of life: Qualitative findings from focus groups. British journal of health psychology 7: 345-363, 2002.
