The study protocol was approved by the institutional review board, and written informed consent was obtained from all participating patients.

This prospective, single-center, non-randomized controlled study was conducted at Department of Surgery, The University of Hong Kong, from September 2021 to January 2023. Patients who had liver tumors indicating liver resection were recruited for the study.

Patients were given ICG (0.25 mg/kg) 1 day before liver resection. A fluorescent imaging system (NIR/ICG system FDA 510 (k), KARL STORZ, Germany) was used to examine resected specimens. In each case, two tissue biopsies were taken, one from the tumor and another from normal tissue. The color signal obtained from the ICG fluorescence imaging was compared with the biopsy samples for further analysis.

Descriptive data regarding tumor characteristics, operative details, postoperative outcomes, oncological outcomes and survival were recorded and listed as part of the study analysis.

Hypothesis

The use of intraoperative NIR/ICG fluorescence imaging would be a promising guiding tool for removing HCC with a better resection margin.

Research plan and methodology

This study was initiated by the investigators and conducted at Department of Surgery, The University of Hong Kong. Recruitment of patients was done by the principal investigator and co-investigators. Patients with newly diagnosed or recurrent liver tumors were recruited according to the following inclusion and exclusion criteria.

Inclusion criteria a.

Hepatectomy was indicated

b.

Laparoscopic or open liver resection was considered technically feasible

c.

Adequate liver reserve

d.

Absence of extrahepatic metastasis

e.

Absence of radiological evidence of venous invasion of the major portal vein or hepatic vein branches

f.

Liver function of Child–Pugh class A or B

g.

Karnofsky performance status ≥ 70%

h.

Total serum bilirubin ≤ 3 × upper normal limit

i.

Platelet count > 60 × 109/L

j.

Serum creatinine ≤ 2 × upper normal limit

Exclusion criteria a.

Presence of extrahepatic metastasis

b.

Unsuitability for general anesthesia

c.

Refusal to consent

d.

Presence of previously documented allergy to ICG

Resectability was determined based on the anatomical location of the tumor in relation to major hepatic vasculatures, regardless of tumor size. The Couinaud nomenclature was used to define major and minor resections. The criteria for selecting patients for major resection have been previously reported [9]. In cases where there was insufficient liver volume, portal vein embolization or ALPPS was considered. To assess liver anatomy and identify any systemic involvement, computed tomography or magnetic resonance imaging was conducted, sometimes supplemented with dual-tracer positron emission tomography in the later period of the study. Postoperative complications were graded according to the Clavien–Dindo classification [12].

Resections were carried out using the open or laparoscopic approach, with laparoscopy being the preferred method whenever feasible. The specific techniques for open and laparoscopic resections have been detailed in previous descriptions [13]. In cases where tumors involved the bile duct, en bloc resection of the tumor and the bile duct was performed. This was followed by biliary reconstruction using hepaticojejunostomy.

Administration of ICG

For patients who underwent liver resection more than 2 weeks after the initial ICG retention test, an additional intravenous injection of ICG (0.25 mg/kg) was administered 24 h prior to the operation.

Fluorescent imaging system

The endoscopic fluorescent imaging system (NIR/ICG System using the OPAL1® technology with the modular IMAGE1 S™ system, FDA 510(k), KARL STORZ, Germany) includes a 10-mm laparoscope, camera head, xenon light guide cable, video processor/illuminator, and 4 K-3D monitor. The charge-coupled device camera and xenon light source in the system have the ability to filter out light wavelengths below 810 nm and above 800 mm. This allows for optimal visualization of NIR light and has the capability to switch images from full floor to fluorescence images, enabling the detection and visualization of fluorescence during the surgical procedure. (Fig. 1).

Fig. 1

The ICG system used in the study

Harvesting tissue samples from resected specimens for histopathological evaluation

Two tissue biopsies, one from the tumor lesion and another form the normal tissue adjacent to the tumor, were collected from the resected specimen and sent for histological assessment. (Fig. 2).

Fig. 2

Example of resected tumor under ICG fluorescence (the hepatic specimen incised, confirming resection of the HCC)

The pathological tissue samples, which were obtained from paraffin-embedded liver sections, were examined by at least two experienced pathologists who were blinded to the results of the radiological examinations. The presence of venous permeation was recorded if tumor emboli were identified within the liver’s venous system, including the portal vein, its lobar and segmental branches, the hepatic vein, capsular vein, and inferior vena cava. The tumor was histologically graded according to the Edmondson criteria.

Follow-up

All patients were subjected to regular follow-up examinations, with monthly follow-ups conducted in the first year and quarterly follow-ups thereafter if no recurrence was detected. The first computed tomography or magnetic resonance imaging was performed approximately 1 month after the resection and subsequent scans were repeated every three to 4 months during the first year and every 6 months in the following years. The diagnosis of recurrence was primarily based on the identification of typical imaging findings on computed tomography or magnetic resonance imaging. If necessary, percutaneous biopsy was performed to confirm the recurrence. This approach allowed for timely detection and management of recurrent tumors.

Statistical analysis

Descriptive data of tumor characteristics, operative details, postoperative and oncological outcomes, and survival were recorded. Continuous variables were expressed as median with interquartile range. Pearson’s chi-squared test or Fisher’s exact test, where appropriate, was used to compare categorical variables. Receiver operating characteristic (ROC) curves were constructed to evaluate the adequacy of resection margins by visualization of the tumors using ICG fluorescence. The area under the curve was calculated to determine the sensitivity and specificity of ICG fluorescence in assessing the resection margin. The computer software SPSS, version 24.0, was used for statistical analyses. A p-value of less than 0.05 was considered statistically significant.

Consent

Prior to operation, every patient was given full explanation of the trial before they gave their voluntary written consent.

Conflicts of interests

None of the investigators had any financial interests in the materials or equipment used in the trial. The trial did not receive any financial support from any organization that had any relationship with the materials or equipment used in the trial.