Influence of transarterial chemoembolization on serum opsonic activity in hepatocellular carcinoma measured by chemiluminescence

Abstract

This study aimed to evaluate the physical stress associated with transarterial chemoembolization (TACE), a catheter-based treatment for hepatocellular carcinoma, by examining changes in serum opsonic activity (SOA). SOA was examined by measuring reactive oxygen species (ROS) produced by neutrophils using lucigenin-dependent chemiluminescence (LgCL) and luminol-dependent chemiluminescence (LmCL). Sixty-four patients were enrolled, and SOA was measured at admission, the following day, and 3 days after TACE. The area under the curve (AUC) for LgCL did not change significantly from baseline to the day after TACE but increased significantly from the following day to 3 days post-TACE. In contrast, no changes were observed in the AUC of LmCL. Subgroup analyses revealed a significant increase in LgCL from day 1 to day 3 post-TACE among patients aged >75 years, males, body mass index (BMI) <25 ‍kg/m², those with a FIB-4 index of ≥2.67, cisplatin use, Hepatitis B virus/Hepatitis C virus-related liver disease, or a procedure time ≥120 ‍min. Multivariate analyses identified BMI <25 ‍kg/m² and cisplatin use as significant risk factors for increased LgCL. Although TACE is considered a minimally invasive procedure, low BMI and cisplatin use have been identified as notable sources of significant physical stresses.

Introduction

Hepatocellular carcinoma (HCC) is widespread, particularly in East Asia and Africa, and has one of the highest mortality rates among all cancer types.⁽¹⁾ HCC has various treatment methods, including surgical resection, systemic drug therapy, and local treatment. Transarterial chemoembolization (TACE), in which chemotherapeutic agents are administered directly to the arteries supplying the tumor using digital subtraction angiography, is recognized as a safe, minimally invasive, and effective treatment for HCC.⁽²⁾ Although some previous studies have investigated perioperative distress and fatigue during TACE, most have focused on psychological aspects of TACE, mainly through questionnaires.^(3–5) In contrast, few studies have investigated the physical stress experienced during the perioperative period of TACE.

Production of reactive oxygen species (ROS) by neutrophils is also associated with physical stress.^(6–9) Phagocytosis by neutrophils and subsequent ROS production play important roles in nonspecific immunity. The adhesion of immunoglobulins and complement components to foreign substances, termed opsonization, activates phagocytosis by neutrophils and subsequent ROS production. In sports medicine, chemiluminescence (CL) is used to measure the serum opsonic activity (SOA) and assess the stress of exercise.^(6,9,10) CL is widely used as a sensitive and accurate method to assess the ability of neutrophils to produce ROS.^(9,11) The species and amount of ROS can be measured using both lucigenin-dependent chemiluminescence (LgCL) and luminol-dependent chemiluminescence (LmCL).^(6,7,9,10) While lucigenin is closely associated with the detection of superoxide anions (O₂⁻), luminol has a higher oxidation potential and more broadly reflects the total production of ROS, including hypochlorous acid (HOCl) produced by myeloperoxidase. The effects of SOA on endoscopic therapy for early gastric and colorectal cancers have been reported.^(12,13) However, the effect of TACE on SOA for HCC has not been studied.

This study aimed to determine whether changes in SOA measured by LgCL and LmCL are useful for estimating physical stress during the perioperative period of TACE for HCC.

Materials and Methods

Patients

Sixty-four patients who provided consent for this study between August 2021 and August 2023 were included. Of these, 61 patients were included in the analysis after excluding two patients who were unable to complete the procedure due to arrhythmic attacks or difficulty maintaining rest during treatment because of back pain, and one patient with missing values in the acquired data (Fig. 1).

Fig. 1.

Flowchart of the study.

HCC was diagnosed using dynamic contrast-enhanced computed tomography or gadolinium-ethoxybenzyl diethylenetriamine penta-acetic acid-enhanced magnetic resonance imaging. The most appropriate treatment was selected based on the size and number of HCC, hepatic functional reserve, and patient preference.

TACE was performed using the Seldinger technique. A vascular catheter was inserted from the femoral artery into the hepatic artery, and computed tomography was performed during hepatic arteriography to confirm tumor localization and the nutrient artery. The selected artery was embolized with anticancer drugs or microspheres, and lipiodol was injected into the artery using a gel part. The procedure was completed after confirming that the drug had accumulated in the tumor using cone-beam computed tomography. With regard to drugs administered into the nutrient artery in TACE, cisplatin was not used in patients with renal impairment or pancytopenia, and epirubicin was not used if there was cardiac impairment.

The following clinical parameters were recorded on the day of admission: sex, age, height, body weight, body mass index (BMI; calculated by dividing body weight in kilograms by the square of height in meters), background liver, hepatic functional reserve, and the number and size of HCC.

Blood samples were collected from the intermediate antebrachial vein using a vacuum blood collection tube on admission, before TACE, on the day after TACE, and 3 days after TACE during bed rest. Blood samples were allowed to clot for 30 ‍min at room temperature and then separated by centrifugation at 3,000 ‍rpm for 10 ‍min. Serum samples were stored at –80°C until analysis. White blood cell counts and fractions, as well as platelet counts, were determined using the XN-9100 (Sysmex, Kobe, Japan). The serum levels of aspartate aminotransferase, alanine aminotransferase, γ-glutamyl transpeptidase, total bilirubin, albumin, C-reactive protein (CRP), immunoglobulins (lgG, IgM, and IgA), and complements (C3 and C4) were measured using the JCA-BM6070 (JEOL Ltd, Akishima, Japan).

The fibrosis (FIB)-4 index was used as an indicator of liver fibrosis. The scoring formula is as follows:

$$\text{FIB-4 index}=[\text{age (years)}\times \text{aspartate aminotransferase (U/L)}]/\left[\text{platelet count}\right(\mathrm{\times }{10}^9/\mathrm{L})\times \text{alanine aminotransferase (U/L)}].$$

Chemiluminescence method

Based on previous studies, SOA in peripheral blood was examined by measuring ROS production by neutrophils using LgCL and LmCL.^(12–17) Neutrophils were isolated from the peripheral blood of a healthy adult man using a Mono-Poly Resolving Medium (Sumitomo Pharma, Osaka, Japan) immediately before the CL assay. Neutrophil suspensions were prepared by adjusting neutrophil counts to 5.6 × 10³ cells/μl through dilution with Hanks’ Balanced Salt Solution (HBSS).^(14,15,18) Zymosan from Saccharomyces cerevisiae, an activator of the alternative pathway of the complement system, was used for the opsonized particles. In this study, opsonized zymosan was prepared as follows: zymosan A (Sigma-Aldrich, St. Louis, MO) was suspended in HBSS at a concentration of 5 ‍mg/ml and sonicated. Patient serum samples (250 ‍μl) were added to the suspended zymosan (5 ‍mg/ml) and incubated at 37°C for 30 ‍min. Two chemiluminescent probes, lucigenin and luminol, were used to detect ROS. Lucigenin was prepared by dissolving bis-N-methylacridinium nitrate (Sigma-Aldrich) in HBSS to a final concentration of 0.5 ‍mM (pH ‍7.4). Luminol was prepared by dissolving 5-amino-2,3-dihydro-1,4-phatalazinedione (Sigma-Aldrich) in 0.1 ‍M NaOH to make a clear solution, and the pH was adjusted to 7.4 using HCl and HBSS to achieve a final concentration of 0.5 ‍mM. The CL reaction of each sample was examined simultaneously in 96-well microplates (well capacity: 400 ‍μl; Greiner Japan, Tokyo, Japan). To each well, 50 ‍μl of neutrophil suspension, 50 ‍μl of opsonized zymosan, 50 ‍μl of lucigenin or luminol solution, and 100 ‍μl of HBSS were added. CL was measured continuously at 37°C using an auto-luminescence analyzer equipped with a microplate reader (FlexStation 3; Molecular Devices, Tokyo, Japan). The CL assay results were evaluated using area under the curve (AUC). For each CL assay, ROS production was also measured in stored serum samples from three healthy male volunteers as standard values, and the CL assay results for the patients were expressed as percentages of the mean values obtained from standard serum samples.

Ethics

This study was approved by the Hirosaki University Ethics Committee (approval number: 2021-063; approval date: May 11, 2021). Prior to admission, the study procedures and objectives were explained to all participants, and written informed consent was obtained from those who agreed to participate.

Statistical analysis

Categorical variables are presented as frequencies, and continuous variables as medians and interquartile ranges. Changes in clinical parameters before TACE, the day after TACE, and three days after TACE were analyzed using the Friedman test, with comparisons between the two groups performed using Bonferroni’s multiple comparison test. The relationships between increased or decreased SOA levels and related parameters were examined using univariate and multivariate analyses. Statistical analyses were performed using the Statistical Package for the Social Sciences ver. 28.0 (SPSS Inc., Chicago, IL) and EZR software (Saitama Medical Center, Jichi Medical University).⁽¹⁹⁾

Results

Patient characteristics

The participants’ characteristics are summarized in Table 1. The median age was 74.0 years, and 45 patients were male. The median BMI was 25.0 ‍kg/m². The median FIB-4 index was 3.17. The underlying liver diseases included hepatitis B in three patients, hepatitis C in 32, alcohol-related liver disease in 18, nonalcoholic fatty liver disease in six, and an unknown cause in 2 patients. According to the Child–Pugh classification, which is an index of hepatic functional reserve, 54 patients were classified as having good hepatic functional reserve (A), seven patients were classified as having intermediate hepatic functional reserve (B), and zero patients were classified as having severe liver cirrhosis (C). The number of HCC treated was one in 39 patients, two in 13 patients, and three or more in nine patients. The maximum tumor diameter was <3 ‍cm in 39 patients and ≥3 ‍cm in 22 patients. The median duration of the TACE procedure was 120 ‍min. The drugs administered to the nutrient vessels included cisplatin in 38 patients, epirubicin in 15, miriplatin in seven, and microspheres in one patient.

Table 1.

Patient characteristics

Parameter
Total patients	61
Age (years)	74.0 (69.0–80.0)
Sex
Male:Female	45:16
Body mass index	25.0 (21.1–26.6)
FIB-4 index	3.17 (2.31–4.76)
Underlying liver diseases
Hepatitis B	3 (4.9%)
Hepatitis C	32 (52.5%)
Alcohol	18 (29.5%)
Non-alcoholic fatty liver	6 (9.8%)
Uncertain	2 (3.3%)
Child Pugh
A	54 (88.5%)
B	7 (11.5%)
C	0 (0.0%)
Number of HCC
1	39 (63.9%)
2	13 (21.3%)
3 or more	9 (14.8%)
Maximum tumor diameter
<3 ‍cm	39 (63.9%)
≥3 ‍cm	22 (36.1%)
Time for TACE	120.0 (92.0–147.0)
Drugs administered into the nutrient vessels
Cisplatin	38 (62.3%)
Epirubicin	15 (24.6%)
Miriplatin	7 (11.5%)
Microspheres	1 (1.6%)

Data are presented as numbers (%) or median (range). HCC, hepatocellular carcinoma; TACE, transarterial chemoembolization.

Serial changes in peripheral blood and biochemical laboratory values

The changes in peripheral blood and biochemical parameters before and after TACE are shown in Table 2. White blood cell and neutrophil counts increased significantly from before TACE to the day after, with only slight changes observed between the day after and three days after TACE. In contrast, CRP showed only a slight increase from before TACE to the day after, but a significant increase was observed between the day after and 3 days post-TACE. The hepatic enzymes aspartate aminotransferase, alanine aminotransferase, γ-glutamyl transpeptidase, and total bilirubin showed a significant increasing trend from before TACE to the day after, and further on day 3 after TACE.

Table 2.

Changes in laboratory data

	The before TACE	The next day of TACE	The 3 days after TACE	The before vs next day	The before vs 3 days	The next day vs 3 days
White blood cell	4,800 (4,130–6,550)	7,520 (6,245–8,700)	8,110 (6,165–10,095)	<0.001	<0.001	<0.001
Neutrophils	3,098 (2,400–4,135)	5,913 (4,788–7,007)	6,167 (4,379–7,928)	<0.001	<0.001	<0.001
C-reactive protein	0.09 (0.04–0.33)	0.78 (0.47–2.02)	8.13 (5.14–13.88)	<0.001	<0.001	<0.001
Aspartate aminotransferase	29.0 (23.0–43.0)	114.0 (66.0–165.0)	140.0 (92.0–243.0)	<0.001	0.056	<0.001
Alanine aminotransferase	23.0 (14.0–39.0)	81.0 (48.0–129.0)	131.0 (198.0–318.0)	<0.001	<0.001	<0.001
γ-glutamyl transpeptidase	54.0 (26.0–84.0)	50.0 (24.0–77.0)	56.0 (32.0–88.0)	<0.001	0.2595	<0.001
Total bilirubin	0.8 (0.6–1.2)	1.3 (1.0–1.7)	1.4 (1.0–1.8)	<0.001	<0.001	0.49
IgG	1,311 (1,051–1,706)	1,219 (1,014–1,502)	1,212 (942–1,515)	<0.001	<0.001	0.029
IgA	303.0 (195.0–411.5)	283.0 (196.0–383.0)	283.5 (183.3–387.0)	<0.001	0.00088	0.999
IgM	62.0 (49.5–96.0)	61.0 (46.0–88.0)	61.0 (45.3–90.8)	<0.001	0.0024	0.999
C3	101.0 (91.5–113.5)	96.0 (88.0–109.5)	109.0 (92.3–120.8)	<0.001	0.0894	<0.001
C4	22.0 (17.0–26.0)	21.0 (17.0–25.5)	24.0 (21.0–29.0)	0.024	<0.001	<0.001

Data are presented as median (range). TACE, transarterial chemoembolization.

Immunoglobulin IgG, IgA, and IgM levels significantly decreased on the day after and 3 days after TACE compared with those before TACE. However, no significant changes were observed in IgA or IgM levels between the day after and 3 days after TACE. Complement components C3 and C4 decreased on the day after TACE but showed a re-increase by day 3.

Perioperative changes in SOA

Figure 2 shows the AUC of LgCL and LmCL. The AUC of LgCL before TACE was 79.1% (72.8–97.7%), and the day after TACE was 79.7% (74.1–83.9%), showing no significant change; however, at 3 days after TACE, the AUC increased to 82.1% (76.6–94.4%), indicating a significant increase compared with the day after TACE. In contrast, the AUC of LmCL was 105.6% (89.2–127.4%) before TACE, 107.5% (89.1–123.8%) on the next day, and 100.0% (89.5–114.8%) at 3 days after TACE, showing no significant change.

Fig. 2.

Changes in AUC of (A) LgCL and (B) LmCL. The results are expressed as percentages relative to the values obtained from the standard serum, which were defined as 100%. AUC, area under the curve; LgCL, lucigenin-dependent chemiluminescence; LmCL, luminol-dependent chemiluminescence.

The effects of age, sex, body size, background liver, liver fibrosis, number and size of liver cancers, drugs used in TACE, and treatment duration on LgCL were examined (Table 3). A significant increase in the AUC of LgCL from the day after TACE to 3 days after was observed in patients aged >75 years, males, BMI <25 ‍kg/m², those with a FIB-4 index ≥2.67, background liver disease due to hepatitis B or C, cisplatin use and those with treatment duration of ≥120 ‍min. Although no significant differences were observed in patients with three or more tumors or tumors >3 ‍cm treated with TACE, an increasing trend in the AUC of LgCL was noted from the day following to 3 days after TACE. Furthermore, patients treated with cisplatin and those undergoing procedures lasting ≥120 ‍min showed a significant increase in the AUC of LgCL from the next day after to 3 days following TACE.

Table 3.

Effects of age, sex, body size, background liver, liver fibrosis, number and size of tumors, drugs used in TACE, and treatment duration on LgCL

	n	AUC measured by LgCL
		The before TACE	The next day of TACE	The 3 days after TACE
Age
< 75 years old	33	78.5 (70.5–90.8)	78.1 (73.5–81.9)	79.7 (75.4–90.8)
≥ 75 years old	28	82.1 (73.0–101.0)	80.0 (74.8–84.6)	84.5 (78.5–96.4)*
Sex
Male	45	81.4 (72.8–98.5)	79.8 (74.6–85.8)	82.9 (76.7–95.0)*
Female	16	78.6 (72.8–86.3)	79.0 (73.1–79.9)	80.4 (75.9–90.6)
Body mass index
< 25 ‍kg/m2	30	78.6 (72.8–98.9)	76.0 (73.0–80.0)	80.8 (76.2–94.0)**
≥ 25 ‍kg/m2	31	80.5 (72.3–91.0)	81.8 (78.5–85.5)	84.5 (76.7–94.0)
Background liver
Hepatitis B or C	35	81.2 (73.2–99.5)	78.1 (73.6–85.2)	84.5 (76.4–95.6)**
Others	26	78.2 (70.6–91.7)	79.9 (77.0–83.7)	80.8 (76.7–93.4)
FIB-4 index
<2.67	21	83.8 (73.6–99.3)	80.1 (74.8–85.8)	80.8 (76.7–93.6)
≥2.67	40	77.6 (70.1–91.2)	78.9 (73.6–83.7)	83.4 (76.5–94.5)*
Administered drug
Cisplatin	38	77.6 (69.6–97.0)	78.3 (73.6–83.4)	83.1 (76.7–94.0)**
Others	23	81.2 (73.4–94.2)	79.7 (77.9–84.8)	81.1 (76.1–94.3)
Number and size of tumors
Less than 3 tumors or less than 3 ‍cm	30	78.5 (71.0–89.1)	79.6 (74.4–83.4)	81.8 (77.0–93.5)
More than 3 tumors or more than 3 ‍cm	31	81.4 (72.9–99.5)	79.7 (73.9–85.0)	82.1 (76.4–94.7) †
Treatment time
<120 ‍min	29	78.7 (73.2–100.6)	80.1 (75.7–86.8)	81.1 (79.1–93.6)
≥120 ‍min	32	79.8 (70.9–93.8)	78.2 (73.3–82.6)	82.5 (75.4–94.5)*

Data are presented as median (range). TACE, transarterial chemoembolization; LgCL, lucigenin-dependent chemiluminescence; AUC, area under the curve. ^†p<0.1 vs the day after TACE, *p<0.05 vs the day after TACE, **p<0.01 vs the day after TACE.

Univariate and multivariate analyses of factors contributing to the increase in LgCL from the day after TACE to 3 days post-TACE are presented in Table 4. Both univariate and multivariate analyses found that BMI <25 ‍kg/m² and cisplatin use were significant risk factors for increased LgCL.

Table 4.

Factors contributing to the increase in LgCL from the day after to 3 days after TACE

	Univariable					Multivariable
	OR	95% CI		p value		OR	95% CI		p value
Male	2	0.56	7.19	0.288
Over 75 years old	1.03	0.36	2.94	0.958
Body mass index less than 25 ‍kg/m2	6.07	1.84	20	0.003		6.98	1.95	24.9	0.003
FIB-4 index 2.67 or higher	1.56	0.55	4.63	0.425
Hepatitis B or C	2.14	0.74	6.21	0.165
Tumor diameter greater than 3 ‍cm	2.63	0.81	8.56	0.109
Number of tumors 3 or more	0.72	0.25	2.13	0.555
Treatment time 120 ‍min or more	1.55	0.54	4.44	0.412
Cisplatin administration	3.05	1.03	9.1	0.049		3.73	1.08	12.9	0.037

OR, odds ratio; CI, confidence interval; LgCL, lucigenin-dependent chemiluminescence; TACE, transarterial chemoembolization.

Discussion

In this study, LgCL, which is used to assess SOA, did not change from before TACE to the day after TACE but significantly increased from the following day to 3 days after TACE. Furthermore, various factors were found to contribute to the increase in LgCL levels, including age, sex, hepatic functional reserve, drugs used, and treatment duration. Among these, low BMI and the use of cisplatin during TACE were identified as particularly significant factors.

Neutrophils produce O₂⁻, through the activation of NADPH oxidase during phagocytosis. O₂⁻ is rapidly converted, either spontaneously or via superoxide dismutase, into hydrogen peroxide (H₂O₂). Neutrophil azurophilic granules contain high concentrations of myeloperoxidase, which is released during degranulation and reacts with H₂O₂ and Cl⁻ to generate HOCl. The ROS produced by neutrophils exhibit varying levels of toxicity; HOCl has a significantly higher oxidation potential than its precursors, O₂⁻ and H₂O₂.^(6,11) It is understood that LgCL reflects O₂⁻ production, whereas LmCL reflects total ROS production, including HOCl. In the LgCL measurements of the participants in this study, the ratio of LgCL to that of serum from healthy participants was approximately 80% before TACE, the next day, and 3 days after TACE. The healthy participants used as controls in this study were young adults with a mean age of approximately 30 years. In contrast, the median age of patients with HCC who underwent TACE was 74.0 years. Neutrophil function is known to declines with age.⁽²⁰⁾ In a previous report, we also demonstrated that the ROS-producing capacity of neutrophils differs between young and elderly patients.⁽²¹⁾ These findings suggest that superoxide production, an initial response of neutrophils to opsonizing activity, may be reduced in elderly individuals compared with that in younger adults.

SOA measured by LgCL has been reported to increase with intense exercise and training, including 7-man rugby matches, prolonged judo training, and intense sumo training.^(18,22,23) No significant changes in SOA measured by LmCL were observed during the perioperative period of endoscopic treatment for early gastric cancer, colorectal cancer, or spinal surgery, which was consistent with the results observed in this study.^(12,13,24) In contrast, SOA measured using LgCL was reported to increase after endoscopic treatment of early gastric and colorectal cancers, similar to the findings of this study.^(12,13) Based on the results obtained using LgCL and LmCL, it is possible that changes in SOA during the perioperative period of TACE may reflect an increase in the production of substances with lower oxidative capacity, such as O₂⁻, rather than those with higher oxidative capacity.

In this study, LgCL increased significantly from the day after to 3 days after TACE, whereas no difference was observed between the day after TACE and the pre-treatment level. This finding differs from a previous study, which demonstrated an increase in LgCL after endoscopic treatment for early gastric and colorectal cancers compared with levels before treatment.^(12,13) In addition, although neutrophil counts were approximately twice as high the day after TACE as before the procedure, the increase in LgCL was delayed. This may be attributed to differences in treatment methods compared with the previous study. In a previous study, the tumor located on the mucosal surface was endoscopically resected, whereas TACE did not involve tumor resection but rather embolization of the tumor’s nutrient artery, resulting in tumor necrosis. Since ischemia and necrosis require a relatively longer period to develop following arterial embolization, it is possible that oxidative stress became apparent on the third day, rather than on the day after TACE.

Factors associated with increased LgCL from the day after to 3 days after TACE were examined, and a BMI <25 ‍kg/m² and cisplatin use were identified as significant factors in both univariate and multivariate analyses. Obesity is generally associated with increased oxidative stress.^(25–27) In this study, the median age of the participants was 74.0 years, with a predominance of elderly individuals. The median BMI of the patients in this study was 25.0 ‍kg/m², indicating that only a small proportion were classified as highly obese. In elderly individuals, BMI tends to be lower than in younger populations because of reduced muscle strength. The BMI associated with the highest life expectancy in the elderly has been reported to range from 27.0–27.9 ‍kg/m².⁽²⁸⁾ A low BMI in older adults also suggests the presence of sarcopenia.⁽²⁹⁾ Oxidative stress plays a major role in the development and progression of sarcopenia.^(30–32) This study suggests that individuals with low BMI may be more vulnerable to oxidative stress during TACE.

Cisplatin is the most commonly used drug for TACE.^(33–35) It has demonstrated favorable antitumor effects in TACE. However, due to its water solubility, cisplatin formulations exhibit physical instability and impose a burden on the kidneys.^(36,37) In the present study, alternative drugs were administered to patients with impaired renal function; nevertheless, a significant increase in LgCL was observed among those treated with cisplatin. These findings suggests that cisplatin use in TACE may be associated with greater neutrophil-derived oxidative stress compared with other anticancer agents.

In the present study, in addition to low BMI and cisplatin use, a significant increase in LgCL from the day after to 3 days after TACE was observed in patients aged >75 years, males, those with a FIB-4 index ≥2.67, background liver disease due to hepatitis B or C, and treatment duration ≥120 ‍min. ROS, which exert various harmful effects via inflammatory cytokines such as tumor necrosis factor-α, increase with age.⁽²⁰⁾ Sex hormones, particularly female hormones, are involved in various aspects of the immune system, including oxidative stress.^(38,39) Women tend to have higher immune function and neutrophil function than men.^(40–42) Among the pathological factors associated with liver fibrosis, oxidative stress has been reported to be associated with metabolic syndrome, obesity, glucose intolerance, and lipid abnormalities.^(21,43–45) These factors are more prevalent among elderly individuals, contributing to the progression of liver fibrosis. Oxidative stress have also been reported to be higher in individuals infected with hepatitis B or C compared with healthy individuals.^(46,47) Additionally, increased oxidative stress has been observed in alcoholic liver injury and nonalcoholic fatty liver disease.^(48–51) Although the reason for the increased LgCL in hepatitis B or C compared to other background liver diseases could not be elucidated in this study, the findings suggest that neutrophil-derived oxidative stress may be greater in viral hepatitis. The results also suggest that, although TACE is a minimally invasive treatment performed under local anesthesia, prolonged treatment may be associated with high physical stress. In this study, although no significant differences were observed in the group with a large number of tumors and greater tumor size, a trend toward increased in LgCL from the day after to 3 days after TACE was observed. Treatment of tumors with large number and size requires extended procedure time, which is considered a contributing factor to the elevated LgCL observed in cases with longer treatment durations.

This study has several limitations. First, it was conducted at a single institution. However, as TACE was performed according to the standard method approved by the Japanese Health Insurance System, similar results could be obtained in a multicenter study. Second, a control group to assess physical stress associated with TACE was not included in this study. Comparative studies with other local treatments, such as radiofrequency ablation, may provide more meaningful findings. Third, the physical stress assessed in this study could not be entirely attributed to physical stress caused by TACE. Other factors such as fasting and pain during the perioperative period may have contributed to physical stress.

In conclusion, SOA measured using CL can be used to safely and conveniently estimate the physical stress associated with TACE. Thus, this method may be applicable in various clinical settings to evaluate physical stress in emerging, less invasive procedures. The observed differences between LgCL and LmCL suggests that SOA during the perioperative period of TACE were associated with the production of substances with lower oxidation potentials. These results supported the hypothesis that TACE is a minimally invasive procedure. However, low BMI and the use of cisplatin are relatively significant physical stressors, considering the high frequency of TACE in elderly individuals.

Author Contributions

SS and DC wrote the manuscript and interpreted data. SS, DC, MO, NA, and TA performed data management and analysis. DC conceived the study design. SS, DC, MO, NA, TA, KY, CI, SKakehata, FT, SKakeda, and HS reviewed and revised the manuscript. All authors have approved the final draft of the manuscript.

Abbreviations

AUC

area under the curve

CL

chemiluminescence

CRP

C-reactive protein

HBSS

Hanks’ Balanced Salt Solution

HCC

hepatocellular carcinoma

HOCl

hypochlorous acid

H₂O₂

hydrogen peroxide

LgCL

lucigenin-dependent chemiluminescence

LmCL

luminol-dependent chemiluminescence

O₂⁻

superoxide anion

ROS

reactive oxygen species

SOA

serum opsonic activity

TACE

transarterial chemoembolization

WBC

white blood cells

Conflict of Interest

No potential conflicts of interest were disclosed.