IRE has been gaining increasing interest because the non-thermal properties of this method permit the ability to ablate tumours adjacent to vital structures. Previous studies have demonstrated that US findings after IRE application in the liver evolve sequentially over time, from seconds to at least hours[3, 11, 12]. Our results show the evolution of post-IRE ablation patterns over the clinically relevant time period.
As noted in an animal-based experiment, a strong association exists between conventional US and histopathology. In our study, on conventional US, IRE ablation zones of most patients appear as a developing hypo-echoic area that demonstrates an increasingly hyperechogenic ablation zone starting one day after the procedure.
Our US findings after IRE ablation evolved sequentially over time. The findings showed good correlation with those in previous studies[5, 13]. In the study, specimens obtained immediately after ablation showed that the widened oedematous sinusoidal spaces were filled mainly by fluid, and very little haemorrhagic infiltrate was observed. As time passed, the haemorrhagic infiltrate became more dominant, which might be due to the widened fluid-filled sinusoids at the beginning of the procedure. The authors attribute this hyper-echogenicity to red blood cell accumulation over time. The degree of red blood cell infiltrate was qualitatively associated with echogenicity. Lee et al also reported the US findings of 55 ablation zones immediately and 1 day after ablation and showed that the immediate hypo-echogenicity transformed to total hyper-echogenicity in the treatment zone 1 day after the procedure. Appelbaum et al speculated that red blood cells progressively infiltrated into deep regions of the ablated zone so that the hyper-echoic rim on US at 90–120 minutes transitioned to complete hyper-echogenicity within 24 hours. There is concordance between our findings and those of previous studies that described the characteristics of images obtained after ablation.
In CEUS, most of the ablated zones showed hypo-enhancement immediately after IRE. Chung et al found the following zones with different enhancement patterns on CT perfusion images in normal porcine liver: an inner non-enhanced zone; a middle well-defined progressive internal enhancement zone, and an outer ill-defined arterial enhancement zone. On histopathology, the inner and middle zones accounted for the extent of cell death. The histological examination suggested that the apoptotic process was involved with complete cell death in the pathophysiology of cell death caused by IRE.
The CEUS images of the IRE ablation zones in normal liver tissue differed from the images of liver tumours treated with IRE. Our study also found that 10 ablated zones appeared with hypo-enhancement immediately after IRE but became non-enhanced one day after IRE, which is due to diverse reasons. Lee et al attributed the focal hyper-attenuation to the release of contrast medium into an ablation defect caused by an IRE-induced microvasculature leakage within the defect zone. Because the zones became non-enhanced on the follow-up CEUS images, the hyper-enhancement was probably caused by extraluminal contrast material. Another opinion is about the differences in contrast agent concentrations. Guo et al suggested that nanometre-scale pores in the tissue cell membrane caused contrast agent to accumulate in the IRE zone and allowed for the contrast agent to be internalized into the intracellular environment rather than remaining extracellularly, which makes evaluating whether the viable tissue is a residual tumour difficult after ablation. Therefore, follow-up CEUS is necessary to assess the viable portion. However, unlike normal regeneration activity, the residual tumour can continue growing and results in a newly enhanced region.
Most of the non-enhancement pattern on CEUS 1 month after ablation is a sign of effective IRE treatment, besides there are still false negatives. Such non-enhancement patterns are different to those of the thermal ablation zone after complete ablation. Previous histologic examinations of the ablation zones showed cell death caused by apoptosis. Cell death is seen with full preservation of the peri-ablative zone structures, such as blood vessels and bile ducts.
In the present study, only one patient continuously showed an enhanced ablation zone immediately and on follow-up CEUS images. The ablation zone became hyper-enhanced during the early arterial phase, slightly wash out in the late arterial phase and presented as hypo-enhanced in the late phase. To elucidate the reason for this pattern, the patient underwent a liver biopsy one month after IRE. The physiology results showed small patchy necrosis, inflammatory fibrosis tissue hyperplasia and foam cell aggregation. The result ruled out the possibility of recurrence. A reasonable explanation for this finding is hyperplasia of the inflammatory tissue. This result can cause confusion in clinical practice. Therefore, sequential follow-up is essential. Further investigation is needed to study the histological and cytological mechanisms underlying this process.
In our study, only 2 ablation zones still appeared as hypo-echoic areas, and both zones were in the same patient. After 1 month of follow-up, recurrence occurred near one of the zones treated with IRE ablation. In a previous multi-institutional review from 2009 through 2012, 31% of the patients had recurrence during a median follow-up of 18 months. Of the total 31% of patients with recurrence, 10.7% had local recurrences at the ablated site. Previous work attributed these recurrences to electric field sinks that resulted from the heterogeneous structure and conductivity of the liver. Later work from the authors indicated that the IRE-treated extracellular matrix (ECM) provides an environment for the activation and differentiation of progenitor cells, but the mechanism is not completely understood; in contrast, some studies have suggested that abnormal ECM affects cancer progression by directly promoting cellular transformation and metastasis and facilitating tumour-associated angiogenesis and inflammation, leading to the generation of a tumorigenic microenvironment. The role of the IRE-spared tumour matrix in follow-up recurrences requires further research. Another theory is about the size of the treated liver tumours. Niessen et al found that large tumour volumes (> 5 cm3) portended early local recurrence.
In our study, the intra-hepatic blood vessels and bile duct remained almost completely intact after IRE. The hepatohilar bile duct of only one patient showed an unclear contrast agent pattern immediately after IRE. After 15 minutes, the phenomenon disappeared. However, a consensus on the mechanism of IRE has not yet been realized. This effect could be due to the high proportion of collagenous connective tissue. A further hypothesis is that the gap junctions, which are present in large numbers in the muscularis propria of the blood vessels and the bile duct walls, could act as a conductive structure for the electrical currents; thus, the current can be passed from cell to cell without leading to destruction of the cell membrane. Whether the effects of IRE are caused by a thermal or non-thermal mechanism remains unclear.
This study has several limitations. First, our study had a small sample size for the span of more than 2 years. Another limitation was that almost all of the patients we included in our study group previously underwent right or left liver resection and thermal ablation or catheter chemical ablation. The origin of the ablation zones was heterogeneous. However, this circumstance mirrors the status of our clinical treatment strategies. IRE as a novel technique and was not the first choice in our hospital. Only patients who experienced tumour recurrence after liver resection and thermal ablation were considered for IRE. Because of this situation, assessing the images of the ablated zones during subsequent follow-up imaging examinations is important. We conclude that CEUS may be a useful tool for assessing the characteristics of post-IRE ablation changes. And it is an effective way to evaluate the therapeutic efficacy 1 month after ablation. Further studies are needed to evaluate more patients to precisely depict the appearance of hepatic zones on CEUS.