RBC storage lesions:

What they are, and how we can

minimize them

Tatsuro Yoshida PhD

New Health Sciences Inc. Bethesda MD

Biomedical Engineering, Boston University
Boston MA
Storage lesions
 What are they?
 How are they related?
 What are the consequences?
 What can we do about them?
 Prevention
 Reversal
 Targets for ideal refrigerated RBC
storage.

Recent review literatures [1-3]

 Refrigerated RBC storage:
altered parameters
Biochemical Biomechanical Oxidative
Metabolic

SNO-Hb ↓
 Refrigerated RBC storage:
altered parameters
Biochemical Biomechanical Oxidative
Metabolic

SNO-Hb ↓
 Refrigerated RBC storage:
altered parameters
Biochemical Biomechanical Oxidative
Metabolic

SNO-Hb ↓
 Refrigerated RBC storage:
altered parameters
Biochemical Biomechanical Oxidative
Metabolic

SNO-Hb ↓
 Storage lesions of metabolic origin
Biochemical Biomechanical Oxidative
Metabolic

SNO-Hb ↓
Storage lesions of metabolic origin

References [4,5]
Consequences of ATP depletion

References [4,5]
Consequences of ATP depletion (2)

References [4,5]
Storage lesion: loss of
vasoregulation
 Hemolysis [gradual increase]
 NO scavenger--vasoconstriction
 ATP release [gradual decrease]
 Loss of ATP mediated vasodilation

 Loss of NO delivery by RBC [fast]

 Impaired vasoregulation
Regulation of microvascular perfusion
mediated by ATP

References [4,5] From: Ellsworth ML et al., Am J Physiol

1995;269(6 Pt 2):H2155-61.
 Consequence of 2,3-DPG depletion

Release DPG from Hb

References [4,5]
Reduced oxygen delivery capacity
Consequences of depleted 2,3-DPG
—reduction in tissue oxygen delivery

From Bunn & Forget 1986 [7]

 2,3-DPG and hemoglobin

Release DPG from Hb

Modified figure from Perutz, Nature 228:734,1970

References [4,5]
Reduced oxygen delivery capacity
Loss of NO-linked vasoregulation by RBC
 Studies: Stored blood lacks nitric oxide: Associated Press
October 8, 2007 08:06:20 PM PST

 Vasodiatory activity of RBC is lost 3 hrs

after blood collection

Bennett-Guerrero E, Veldman TH, Doctor A, et al. Evolution of

adverse changes in stored RBCs. Proceedings of the National
Academy of Sciences 2007;104(43):17063-68.
 RBC is a carrier of NO for vasoregulation
Regulation of microvascular perfusion
mediated by Hemoglobin + NO

O
SN

SNO---biologically active S-nirtrosothiol

References [27,28]
 Hemoglobin and nitric oxide
NO
(Hb)4 (Hb)3Hb-NO T-State

RSNO O2

(Hb)4-SNO (HbO2)4-SNO R-State

O2
LUNG
Loss of NO-linked RBC
mediated vasoregulation
 Immediately after transfusion, RBCs
may not only fail to increase peripheral
blood flow, but may also function as NO
sink
Loss of NO-linked RBC
mediated vasoregulation
 Immediately after transfusion, RBCs may
not only fail to increase peripheral blood
flow, but may also function as NO sink

 How serious is this effect in relation to other

storage lesions?
 Is it reversible in vivo---most likely
 How fast can it recover in vivo??
 Storage lesions linked to oxidative
damage
Biochemical Biomechanical Oxidative
Metabolic

SNO-Hb ↓
Oxidative damage pathway

References [8-11]
 Storage lesions linked to oxidative damage

SNO-Hb ↓
References [8-11]
Consequences: oxidative damage

References [12,13]
 Damage pathways
Biochemical Oxidative
/ Metabolic Damage
alterations

Bio-mechanical
changes

Hemolysis
Post-transfusion removal
TRALI
Reduction or delaying
development of storage lesions

Metabolically linked
RBC metabolic pathways

[26]
 Prevention of metabolic storage lesions:
Embden-Meyerhof pathway— ATP and 23DPG

Metabolic modulation [14-15]

Prevention of metabolic storage lesions:
manipulations with additive solution

Review: Hess 2006 [16]

Prevention of metabolic storage lesions:
storage under anaerobic condition

References [17-19]
Reduction or delaying
development of storage lesions

Oxidative damages
Reduction of oxidative damage:
RBC redox system
Reduction of oxidative damage:
metabolic manipulations

•GSH precursors [20]

•L-carnitine (protection of RBC

membrane [21])

•Anti-oxidant / free radical

scavenger in storage solution
Mannitol, N-acetylcysteine
tirilazad mesylate, etc
[11,22,23]
 Reduction of oxidative damage:
storage under anaerobic condition

Store RBC without oxygen

• Stop hydroxyl radical-
mediated peroxidation cycles
• Prevent hemoglobin
denaturation

Reference [19]
Reversal of storage lesions
 Reversibility of storage lesions

SNO-Hb
Rejuvenation
 Post-storage metabolic manipulations
 Mixture of pyruvate, inosine, Pi,
adenine, PEP etc
 Rejuvesol (Cytosol Laboratory Inc)
 37°C incubation followed by cell washing [24]
 Experimental
 PEP [references in 25]
RBC metabolic pathways

[26]
Rejuvenation by PIPA (Rejuvesol)
Rejuvenation by PIPA (Rejuvesol)
Rejuvenation by PIPA (Rejuvesol)
Rejuvenation with PEP

Reference [25]
Result of rejuvenation
 Restored ATP
 Elevated 2,3-DPG
 Increased 24-hr recovery rate
 Rejuvesol is approved by FDA, but RBC must be
washed
Result of rejuvenation
 Restored ATP
 Elevated 2,3-DPG
 Increased 24-hr recovery rate
 Rejuvesol is approved by FDA, but RBC must be washed

 Rejuvesol can be added during storage

 High levels of ATP and 2,3DPG throughout extended
storage period
 Prevent PS accumulation on RBC surface
 Increased 24-hr recovery rate
Goals for improved RBC
storage
Reduced toxicity
 Reduce hemolysis
 Reduce dead-on-arrival RBC
 Iron toxicity
 Vasoconstriction via NO scavenging

 Maintain deformability / prevent aggregation

 Prevent capillary blockage
Increase 24-hr recovery rate and / or
extend shelf-life
 Maintenance of high ATP
 Minimize oxidative damage

 Maintain below threshold parameters causing

removal after transfusion [to provide sufficient
time for recovery in body after transfusion]
Targets for ideal refrigerated RBC storage
Reduced toxicity
 Reduce hemolysis
 Reduce dead-on-arrival RBC
 Iron toxicity
 Vasoconstriction via NO scavenging

 Maintain deformability / prevent aggregation

 Prevent capillary blockage
 Prevent release, and/or remove bio-active
substances
 Prevention of TRALI
 Leukoreduction, irradiation, pathogen-reduction, etc.
Higher functionality for efficient
tissue oxygenation
 Maintain high levels of:
 2,3-DPG [oxygen release]
 SNO-Hb [regulation of vascular perfusion]
 ATP [regulation of vascular
perfusion]
 Deformability [capillary perfusion]
 Selected recent reviews
Storage lesion
 Ho J, Sibbald WJ, Chin-Yee IH. Effects of storage on efficacy of red cell transfusion: when is it not safe? Crit Care Med
2003;31(12 Suppl):S687-97.
 Chin-Yee I, Arya N, d'Almeida MS. The red cell storage lesion and its implication for transfusion. Transfus Sci
1997;18(3):447-58.

Clinical consequences of transfusion (not covered in this talk)

 Tinmouth A, Fergusson D, Yee IC, Hebert PC. Clinical consequences of red cell storage in the critically ill. Transfusion
2006;46(11):2014-27.
 Solheim BG, Flesland O, Seghatchian J, Brosstad F. Clinical implications of red blood cell and platelet storage lesions: an
overview. Transfus Apher Sci 2004;31(3):185-9.
 Vincent JL, Baron JF, Reinhart K, et al. Anemia and blood transfusion in critically ill patients. Jama 2002;288(12):1499-507.
 Spiess BD. Blood transfusion: the silent epidemic. Ann Thorac Surg 2001;72(5):S1832-7.

Additive solutions
 Hess JR. An update on solutions for red cell storage. Vox Sang 2006;91(1):13-9.

Vasoregulatory function of stored RBC

 Bennett-Guerrero E, Veldman TH, Doctor A, et al. Evolution of adverse changes in stored. RBCs. Proceedings of the
National Academy of Sciences 2007;104(43):17063-68.
 References (1)

1. Tinmouth A, Fergusson D, Yee IC, Hebert PC. Clinical consequences of red cell storage in the critically ill. Transfusion
2006;46(11):2014-27.
2. Ho J, Sibbald WJ, Chin-Yee IH. Effects of storage on efficacy of red cell transfusion: when is it not safe? Crit Care Med
2003;31(12 Suppl):S687-97.
3. Chin-Yee I, Arya N, d'Almeida MS. The red cell storage lesion and its implication for transfusion. Transfus Sci
1997;18(3):447-58.
4. Ellsworth ML, Forrester T, Ellis CG, Dietrich HH. The erythrocyte as a regulator of vascular tone. Am J Physiol
1995;269(6 Pt 2):H2155-61.
5. Sprague RS, Ellsworth ML, Stephenson AH, Lonigro AJ. ATP: the red blood cell link to NO and local control of the
pulmonary circulation. Am J Physiol 1996;271(6 Pt 2):H2717-22.
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Vox Sang 2007;92(1):22-31.
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 References (3)

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RBCs. Proceedings of the National Academy of Sciences 2007;104(43):17063-68.