Comparison between High-Frequency Irreversible Electroporation and Irreversible Electroporation Ablation of Small Swine Liver: Follow-Up of DCE-MRI and Pathological Observations
Introduction
Irreversible electroporation (IRE), also known as nanoknife ablation, is a non-thermal ablation technology that uses high-voltage electrical pulses to create nanometer-scale pores in cell membranes, leading to cell death and tissue necrosis. Its non-thermal nature allows it to be applied in sensitive areas such as the porta hepatis and pancreas, sparing critical structures like blood vessels, bile ducts, pancreatic ducts, and nerves. However, IRE has limitations, including severe muscle contractions induced by electrical impulses, which necessitate the use of paralytic agents and general anesthesia. These factors complicate the procedure and limit its broader clinical application.
To address these issues, high-frequency irreversible electroporation (H-FIRE) was developed. H-FIRE uses high-frequency bipolar bursts to minimize muscle contractions by canceling the effects of alternating polarity pulses. This innovation eliminates the need for paralytic agents and general anesthesia, simplifying the procedure and making it accessible to more patients. While both IRE and H-FIRE induce cell death, their distinct waveforms and frequencies may lead to different cellular responses and repair mechanisms. However, the in vivo effects and mechanisms of H-FIRE remain poorly understood, and its efficacy compared to IRE has not been thoroughly evaluated.
Magnetic resonance imaging (MRI) is an ideal tool for assessing IRE treatment responses in liver tissues. Dynamic contrast-enhanced MRI (DCE-MRI) provides valuable information about tissue perfusion, vascular blood flow, blood vessel permeability, and contrast uptake. Quantitative parameters such as the volume transfer constant (Ktrans), rate constant (Kep), and extravascular extracellular volume fraction (Ve) can be derived from DCE-MRI, offering insights into tissue changes post-treatment. While DCE-MRI has been used to distinguish between IRE and reversible electroporation (RE) zones, its application in evaluating H-FIRE treatment remains limited.
This study aimed to compare the efficacy of IRE and H-FIRE in vivo by analyzing temperature changes, ablation zone size, DCE-MRI parameters, and histologic findings in a porcine liver model. The results provide insights into the potential of H-FIRE as a viable alternative to IRE and the utility of DCE-MRI in monitoring treatment outcomes.
Methods
Ethical approval for the study was obtained from the Institutional Animal Ethics Committee of the General Hospital of the People’s Liberation Army. Ten Bama miniature swine were used in the experiments, divided into two groups: a 1-day group and a 7-day group. Each animal underwent both IRE and H-FIRE procedures at two sites in the liver, separated by a distance of over 6 cm to avoid interference.
IRE and H-FIRE Ablation
The liver was exposed via laparotomy, and two parallel needle electrodes with a 1.5 cm spacing were inserted into the liver to a depth of 1.5 cm. IRE was performed using a generator delivering 2200 V square wave pulses, while H-FIRE was delivered using a custom-built generator providing 3000 V asymmetric square wave pulses. The temperature of the electrodes was monitored using a Bragg grating to measure real-time temperature changes during ablation.
DCE-MRI Examination
DCE-MRI images were acquired using a 3.0 T MRI unit on days 1, 4, and 7 post-treatment in the 7-day group. A dual-input two-compartment pharmacokinetic model was used to calculate Ktrans, Kep, and Ve from the DCE-MRI data. The size of the ablation zone was measured on DCE-MRI images, and histologic analysis was performed on treated and untreated liver tissues.
Histologic and Transmission Electron Microscope Analysis
Liver tissues were fixed in formalin and processed for hematoxylin and eosin (H&E) staining, heat shock protein 70 (HSP70) staining, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) assays. Transmission electron microscopy was used to observe ultrastructural changes in hepatocytes following IRE and H-FIRE treatment.
Statistical Analysis
Data were analyzed using GraphPad Prism software. Continuous variables were presented as mean ± standard deviation or median (Q1, Q3). Student’s t-test, Mann-Whitney U test, one-way ANOVA, and Pearson correlation coefficient tests were used for statistical comparisons.
Results
Temperature Monitoring
The temperature variations in the electrodes during IRE and H-FIRE ablation were similar, with no significant difference between the two treatments (18.00 ± 3.77°C for IRE vs. 16.20 ± 7.45°C for H-FIRE, t = 0.682, P = 0.504).
DCE-MRI Findings
No significant differences were observed in Ktrans or Kep between IRE and H-FIRE zones at any time point. However, the Ve value was significantly higher in the IRE zone than in the H-FIRE zone on day 4 (0.14 ± 0.02 vs. 0.08 ± 0.05, t = 2.408, P = 0.043). In the IRE zone, the Ktrans value was significantly higher on day 7 compared to day 1 (P = 0.033), indicating increased vascular permeability and plasma flow over time.
Ablation Zone Size
The ablation zone size was significantly larger in the H-FIRE zone than in the IRE zone on day 1 (4.74 ± 0.88 cm2 vs. 3.20 ± 0.77 cm2, t = 3.241, P = 0.009) and day 4 (2.22 ± 0.83 cm2 vs. 1.30 ± 0.50 cm2, t = 2.343, P = 0.041). Both IRE and H-FIRE zones showed a significant decrease in size over time, reflecting tissue regeneration.
Histologic and Electron Microscopy Findings
Electron microscopy revealed lysis of the plasma membrane, cytoplasmic swelling, and intracellular vacuoles in hepatocytes following both IRE and H-FIRE treatment. Histologic analysis showed extensive necrosis, hemorrhage, and overexpression of HSP70 in the ablation zones. The apoptotic index was significantly higher in the IRE and H-FIRE zones compared to untreated tissue (P < 0.001), but no significant difference was observed between the two treatments.
Correlation Between DCE-MRI Parameters and Hepatocyte Count
In the H-FIRE zone, the Ktrans and Kep values were positively correlated with the number of hepatocytes on day 7 (r = 0.940, P = 0.017 for Ktrans; r = 0.895, P = 0.040 for Kep). No significant correlations were found in the IRE zone.
Discussion
This study demonstrated that H-FIRE has a comparable ablation effect to IRE, with a larger ablation zone size and similar temperature changes during treatment. The absence of significant differences in Ktrans and Kep between the two treatments suggests that H-FIRE achieves similar levels of tissue damage and vascular permeability. The higher Ve value in the IRE zone on day 4 may reflect differences in extracellular fluid dynamics, but this did not translate into a significant difference in overall efficacy.
The larger ablation zone size observed with H-FIRE could be attributed to its unique waveform and frequency, which may enhance tissue penetration and cell death. However, the difference in ablation zone size decreased over time, indicating that both treatments induce similar regenerative responses in the liver.
Histologic and electron microscopy findings confirmed that both IRE and H-FIRE induce apoptosis and necrosis in hepatocytes, with no significant differences in apoptotic index or HSP70 expression between the two treatments. The preservation of blood vessels and bile ducts in the ablation zones further highlights the non-thermal nature of both techniques.
DCE-MRI proved to be a valuable tool for monitoring the pathophysiological changes in the ablation zones. The positive correlation between Ktrans, Kep, and hepatocyte count in the H-FIRE zone suggests that DCE-MRI can provide insights into tissue regeneration and vascular permeability following treatment.
Limitations of the study include the small sample size and the use of normal liver tissue, which may not fully replicate the conditions of tumor ablation. Future studies should investigate the effects of IRE and H-FIRE in tumor models and explore the potential of DCE-MRI in guiding treatment strategies.
Conclusion
H-FIRE is a promising alternative to IRE, offering comparable ablation efficacy with reduced procedural complexity. The larger ablation zone size and similar safety profile make H-FIRE a viable option for clinical applications. DCE-MRI provides valuable insights into tissue changes post-treatment, highlighting its potential as a monitoring tool for H-FIRE ablation.
doi.org/10.1097/CM9.0000000000001663
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