title: Effects of chitooligosaccharides on human red blood cell morphology and membrane protein structure.
abstract: Recent studies of chitosan have increased the interest in its conversion to chitooligosaccharides (COSs) because these compounds are water-soluble and have potential use in several biomedical applications. Furthermore, such oligomers may be more advantageous than chitosans because of their much higher absorption profiles at the intestinal level, which permit their facilitated access to systemic circulation and potential distribution throughout the entire human body. In that perspective, it is important to clarify their effect on blood further, namely, on human red blood cells (RBCs). The aim of this work was thus to study the effect of two COS mixtures with different molecular weight (MW) ranges, <3 and <5 kDa, at various concentrations (5.0-0.005 mg/mL) on human RBCs. The interactions of these two mixtures with RBC membrane proteins and with hemoglobin were assessed, and the RBC morphology and surface structure were analyzed by optical microscopy (OM) and atomic force microscopy (AFM). In the presence of either COS mixture, no significant hemolysis was observed; however, at COS concentrations >0.1 mg/mL, changes in membrane binding hemoglobin were observed. Membrane protein changes were also observed with increasing COS concentration, including a reduction in both alpha- and beta-spectrin and in band 3 protein, and the development of three new protein bands: peroxiredoxin 2, calmodulin, and hemoglobin chains. Morphologic evaluation by OM showed that at high concentrations COSs interact with RBCs, leading to RBC adhesion, aggregation, or both. An increase in the roughness of the RBC surface with increasing COS concentration was observed by AFM. Overall, these findings suggest that COS damage to RBCs was dependent on the COS MW and concentration, and significant damage resulted from either a higher MW or a greater concentration (>0.1 mg/mL).
authors.affiliation: Escola Superior de Biotecnologia, Universidade Católica Portuguesa, Rua Dr. António Bernardino de Almeida, P-4200-072 Porto, Portugal.
authors.forename: João C
authors.initials: JC
authors.lastname: Fernandes
authors.forename: Peter
authors.initials: P
authors.lastname: Eaton
authors.forename: Henrique
authors.initials: H
authors.lastname: Nascimento
authors.forename: Luís
authors.initials: L
authors.lastname: Belo
authors.forename: Susana
authors.initials: S
authors.lastname: Rocha
authors.forename: Rui
authors.initials: R
authors.lastname: Vitorino
authors.forename: Francisco
authors.initials: F
authors.lastname: Amado
authors.forename: Joana
authors.initials: J
authors.lastname: Gomes
authors.forename: Alice
authors.initials: A
authors.lastname: Santos-Silva
authors.forename: Manuela E
authors.initials: ME
authors.lastname: Pintado
authors.forename: F Xavier
authors.initials: FX
authors.lastname: Malcata
mesh_terms: D002448:Cell Adhesion / Q000187:drug effects; D002449:Cell Aggregation / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000000981:diagnostic imaging / Q000187:drug effects* / Q000378:metabolism; D006454:Hemoglobins / Q000378:metabolism; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D018625:Microscopy, Atomic Force; D008970:Molecular Weight; D009844:Oligosaccharides / Q000009:adverse effects / Q000737:chemistry / Q000494:pharmacology*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013499:Surface Properties; D014463:Ultrasonography
chemical_list: D006454:Hemoglobins; D008565:Membrane Proteins; D009844:Oligosaccharides