Cardiopulmonary bypass (CPB) is essential for modern cardiac surgery, enabling complex procedures on the heart and major vessels. However, exposure of blood to artificial surfaces, mechanical shear stress, hypothermia, and hemodilution can negatively affect red blood cells (RBCs). In this 2026 pilot study published in the Journal of Clinical Medicine, Sergunova et al. investigated how CPB alone versus CPB combined with hypothermic circulatory arrest (CPB+HCA) alters RBC structure and mechanical properties at the nanoscale using atomic force microscopy (AFM) .
Study Design and Patient Groups
Fourteen patients undergoing cardiac surgery were divided into two groups. Group 1 (n=7) underwent heart valve surgery with normothermic CPB (35°C). Group 2 (n=7) underwent aortic arch surgery requiring CPB combined with moderate hypothermia (28°C) and hypothermic circulatory arrest. Blood samples were collected before anesthesia induction and immediately after CPB.
Unlike conventional hemorheological studies that evaluate whole-blood viscosity, this research focused on individual erythrocytes. AFM was used to assess:
- RBC morphology (discocytes, echinocytes, stomatocytes, planocytes)
- Membrane nanostructure via surface roughness (Rtm)
- Mechanical stiffness via Young’s modulus (E)
This approach allowed detection of subtle membrane and cytoskeletal alterations not visible with routine laboratory testing.
Morphological Changes in Red Blood Cells
The most striking findings involved RBC morphology. In the CPB-only group, the proportion of healthy discocytes decreased postoperatively, but some normal morphology remained. In contrast, in the CPB+HCA group, discocytes were completely absent after surgery in all patients. In one case, discocytes fell from 74% preoperatively to 0% postoperatively, with 100% of cells classified as echinocytes (page 5, Figure 2).
This dramatic shift suggests that combining hypothermia and circulatory arrest produces more severe membrane-level injury than normothermic CPB alone. The transformation into echinocytes and stomatocytes reflects membrane instability, altered lipid bilayer structure, and potential oxidative stress.
Nanostructural Membrane Damage
Membrane surface roughness (Rtm) was used as a quantitative marker of nanoscale membrane injury. Both groups demonstrated significant increases in Rtm after surgery (page 6–7, Figure 4).
- Group 1 (CPB): Mean Rtm increased 1.4-fold (p < 0.0001)
- Group 2 (CPB+HCA): Mean Rtm increased 1.6-fold (p < 0.0001)
These findings indicate that CPB causes measurable nanostructural membrane disruption, and that hypothermic circulatory arrest amplifies this effect. AFM images showed pronounced postoperative changes in surface topography, consistent with membrane micro-irregularities and structural destabilization.
Importantly, inter-patient variability was observed, emphasizing that RBC damage during CPB may depend on individual susceptibility, perfusion duration, and procedural complexity.
Mechanical Properties and Cytoskeletal Stiffness
Mechanical stiffness, measured via Young’s modulus, revealed a key mechanistic difference between groups (page 7–8, Figure 5).
In the CPB group, RBC stiffness increased significantly—on average 1.55-fold (p < 0.0001). This suggests cytoskeletal rigidification due to mechanical shear stress, turbulent flow, and interaction with extracorporeal circuit surfaces. Increased stiffness reduces RBC deformability, impairing microcirculatory flow and oxygen delivery.
Conversely, in the CPB+HCA group, Young’s modulus increased only modestly (1.14-fold), and in several patients the change was not statistically significant. Despite more severe morphological damage and membrane roughness, cytoskeletal elasticity was relatively preserved.
This dissociation between membrane topography and mechanical stiffness is a central contribution of the study. It suggests that:
- Normothermic CPB primarily induces mechanical stress leading to cytoskeletal stiffening.
- CPB+HCA causes more surface-level membrane injury driven by hypothermia, hemodilution, and reperfusion stress, without proportionally increasing cytoskeletal rigidity.
Clinical Implications
Although traditional laboratory parameters such as hematocrit and hemoglobin were comparable between groups (page 8, Table 1), AFM revealed significant cellular-level injury invisible to routine testing.
These nanoscale changes may impair:
- RBC deformability
- Microvascular perfusion
- Oxygen transport efficiency
- Cellular lifespan
Stiffer or morphologically altered RBCs are more rapidly cleared by the spleen and may contribute to postoperative organ dysfunction. Understanding how perfusion strategy affects RBC biomechanics may inform improvements in extracorporeal circulation design, temperature management, and protective interventions.
Mechanistic Model
The authors summarize these mechanisms in a schematic diagram (page 10, Figure 6).
Under CPB (normothermia):
- Shear stress and turbulence → cytoskeletal injury
- Increased Young’s modulus
- Moderate membrane changes
Under CPB+HCA (hypothermia):
- Increased blood viscosity and altered flow
- Greater membrane topography disruption
- Preservation of cytoskeletal elasticity
This conceptual distinction may influence perfusion protocols in complex aortic surgery.
Limitations
As a pilot study with only 14 patients, statistical power is limited. Clinical outcomes, transfusion requirements, and long-term correlations with organ dysfunction were not assessed. The study also did not quantify effects of blood–air contact or ultrafiltration. Larger prospective studies are needed to validate AFM-derived biomarkers as predictors of postoperative complications.
Conclusion
This study demonstrates that cardiopulmonary bypass alters red blood cell structure and biomechanics at the nanoscale. Normothermic CPB primarily increases RBC stiffness, while CPB combined with hypothermic circulatory arrest produces more profound morphological and membrane surface damage. Atomic force microscopy may serve as a powerful tool to detect subtle erythrocyte injury and guide optimization of cardiac surgery perfusion strategies .
