In vivo evaluation of electron mediators for the reduction of methemoglobin encapsulated in liposomes by using electron energies produced by red blood cell glycolysis
(2017) KBK820 20162Pure and Applied Biochemistry
- Abstract
- Previous studies clarified that the electron enriched NADH and NADPH by RBC glycolysis could be used in an extracellular chemical reaction, where electron energies are extracted by an electron mediator methylene blue (MB). The reduced form of MB permeates the phospholipid bilayer of liposomes and reduce encapsulated ferric methemoglobin (metHb) to ferrous hemoglobin. Moreover, in vitro screening study clarified some other potential chemicals with comparable capacity to reduce metHb. With this background, 8 of these compounds: MB, 1,9-dimethyl methylene blue (DMB), azure A (AA), azure B (AB), azure (AC), toluidine blue (TDB), brilliant cresyl blue (BCB), toluylene blue (TLB), were evaluated by both in vitro and in vivo studies in the... (More)
- Previous studies clarified that the electron enriched NADH and NADPH by RBC glycolysis could be used in an extracellular chemical reaction, where electron energies are extracted by an electron mediator methylene blue (MB). The reduced form of MB permeates the phospholipid bilayer of liposomes and reduce encapsulated ferric methemoglobin (metHb) to ferrous hemoglobin. Moreover, in vitro screening study clarified some other potential chemicals with comparable capacity to reduce metHb. With this background, 8 of these compounds: MB, 1,9-dimethyl methylene blue (DMB), azure A (AA), azure B (AB), azure (AC), toluidine blue (TDB), brilliant cresyl blue (BCB), toluylene blue (TLB), were evaluated by both in vitro and in vivo studies in the present work. As compared with MB as a guideline, in vitro experiments showed that most compounds resulted in effective metHb reduction of HbV in the presence of RBC. However, in vivo experiment using Wistar rats for 6 h observation (injection of 10 mL HbV/kg to rats) showed some discrepancies and the effective reduction was found for the combination of AB (1.53 mL/kg body weight, 2.6 mM). To further evaluate the optimization with AB, a hemorrhagic shock and resuscitation model was used, where hemorrhagic shock was induced by delaying fluid resuscitation with 15 min after withdrawal of 50% blood (28 mL/kg body weight) from the femoral arteries and the rats were resuscitated with HbV. Again, AB (1.0-2.4 mL/kg body weight) resulted in sufficient reduction of metHb. The study clarified through in vivo experiments that AB is a suitable electron mediator to prolong the functional life-time of HbV. The study also implies the potential of the dyes for the treatment of methemoglobinemia with better outcomes than conventional MB therapy. (Less)
- Popular Abstract
- In many countries, worldwide, the present blood transfusion system is not sufficient on supplying the healthcare with sufficient and safe blood transfusion. The short shelf-life of approximately 42 days, and the various complicated, time consuming and expensive blood tests contribute to the shortage of blood supplies. The necessity of alternative blood supplies was recognized in the 1980’s when it was reported that HIV could be transmitted through transfusion. Subsequently, extensive research in development of artificial blood, such as hemoglobin-based oxygen carriers, has been made, since it has capacity to be feasible to ease the shortage of blood supplies for the future, and alleviate blood transfusion related safety issues.... (More)
- In many countries, worldwide, the present blood transfusion system is not sufficient on supplying the healthcare with sufficient and safe blood transfusion. The short shelf-life of approximately 42 days, and the various complicated, time consuming and expensive blood tests contribute to the shortage of blood supplies. The necessity of alternative blood supplies was recognized in the 1980’s when it was reported that HIV could be transmitted through transfusion. Subsequently, extensive research in development of artificial blood, such as hemoglobin-based oxygen carriers, has been made, since it has capacity to be feasible to ease the shortage of blood supplies for the future, and alleviate blood transfusion related safety issues. Furthermore, the artificial blood can be modified to be ubiquitously applicable for all blood types and attributes such as longer shelf-life can be achieved.
Free hemoglobin solution is toxic to the body and can cause complications such as vasoconstriction and nephrotoxicity. Therefore, we encapsulate the hemoglobin solution in lipid membranes, so called hemoglobin vesicles, to inhibit the toxicity caused by free hemoglobin. Hemoglobin vesicles are made to mimic the properties and conditions of red blood cells. The safety and efficacy of hemoglobin vesicles as a transfusion alternative have been clarified by animal experiments. Nonetheless, gradual auto-oxidation, of the iron in hemoglobin, from ferrous state (Fe2+) to ferric (Fe3+) methemoglobin remains a problem since it impairs the ability of binding oxygen. In red blood cells, the level of methemoglobin is normally controlled by several enzymatic systems. However, in hemoglobin vesicles these enzymatic systems are eliminated due to the usage of purified and pasteurized hemoglobin solution. Therefore, we add an external substance, such as methylene blue that can mediate in the reduction of the oxidized and non-functional hemoglobin. Addition of methylene blue reduces the level by mediating electrons from the electron carriers NADH and NADPH, produced in the red blood cell glycolysis, to the oxidized hemoglobin inside the vesicle.
In this study, we evaluate the potential of eight different cationic organic dyes, including methylene blue, in the reduction of encapsulated methemoglobin. The results were evaluated using methylene blue as reference due to it is conventional application in the treatment of methemoglobinemia. It was observed that toluylene blue, toluidine O and brilliant cresyl blue show similar reductive effect as compared to methylene blue. Derivatives of methylene blue (azure A, azure B and azure C) showed improved effect, while di-methyl methylene blue did not show in vivo effectiveness. It is thought that the observed variation in reduction among the dyes is due to differences in their structure and metabolism. Among the dyes tested we believe that azure B deserves more attention as it is a suitable electron mediator which can be utilized to prolong the functional life-time of artificial blood and does also deserve further investigation as an alternative therapy for methemoglobinemia. (Less)
Please use this url to cite or link to this publication:
http://lup.lub.lu.se/student-papers/record/8920470
- author
- Ghirmai, Semhar LU
- supervisor
-
- Leif Bülow LU
- organization
- course
- KBK820 20162
- year
- 2017
- type
- H2 - Master's Degree (Two Years)
- subject
- keywords
- Blood substitutes, hemoglobin vesicles, hemoglobin-based oxygen carriers, hemorrhage, methemoglobin, methemoglobinemia, phenothiazine, red blood cells, applied biochemistry
- language
- English
- id
- 8920470
- date added to LUP
- 2023-05-08 11:05:39
- date last changed
- 2023-05-08 11:05:39
@misc{8920470,
abstract = {{Previous studies clarified that the electron enriched NADH and NADPH by RBC glycolysis could be used in an extracellular chemical reaction, where electron energies are extracted by an electron mediator methylene blue (MB). The reduced form of MB permeates the phospholipid bilayer of liposomes and reduce encapsulated ferric methemoglobin (metHb) to ferrous hemoglobin. Moreover, in vitro screening study clarified some other potential chemicals with comparable capacity to reduce metHb. With this background, 8 of these compounds: MB, 1,9-dimethyl methylene blue (DMB), azure A (AA), azure B (AB), azure (AC), toluidine blue (TDB), brilliant cresyl blue (BCB), toluylene blue (TLB), were evaluated by both in vitro and in vivo studies in the present work. As compared with MB as a guideline, in vitro experiments showed that most compounds resulted in effective metHb reduction of HbV in the presence of RBC. However, in vivo experiment using Wistar rats for 6 h observation (injection of 10 mL HbV/kg to rats) showed some discrepancies and the effective reduction was found for the combination of AB (1.53 mL/kg body weight, 2.6 mM). To further evaluate the optimization with AB, a hemorrhagic shock and resuscitation model was used, where hemorrhagic shock was induced by delaying fluid resuscitation with 15 min after withdrawal of 50% blood (28 mL/kg body weight) from the femoral arteries and the rats were resuscitated with HbV. Again, AB (1.0-2.4 mL/kg body weight) resulted in sufficient reduction of metHb. The study clarified through in vivo experiments that AB is a suitable electron mediator to prolong the functional life-time of HbV. The study also implies the potential of the dyes for the treatment of methemoglobinemia with better outcomes than conventional MB therapy.}},
author = {{Ghirmai, Semhar}},
language = {{eng}},
note = {{Student Paper}},
title = {{In vivo evaluation of electron mediators for the reduction of methemoglobin encapsulated in liposomes by using electron energies produced by red blood cell glycolysis}},
year = {{2017}},
}