Abstract
Background/Aim: Anti- and proinflammatory cytokines and plasma high-sensitivity C-reactive protein (hs-CRP) are used to assess inflammatory stress response (ISR) following surgery. However, the serum IL-18 (interleukin-18) cytokine values versus numeric rating scale (NRS) pain score and number of analgesic doses (NAD) postoperatively are unknown. Patients and Methods: Blood levels of six interleukins (IL-18, IL-1ra, IL-6, IL-10, IL-1β, and IL-8) and hs-CRP were measured at three time points; before operation (PRE), immediately after operation (POP1), and six hours after operation (POP2) in 114 patients with cholelithiasis. Results: Following surgery, the blood levels of hs-CRP and IL-1ra, IL-6, IL-10, and IL-1β cytokines had a trend for increase (p<0.001 and p=0.014, respectively). The serum IL-18 concentrations inversely correlated to NRS and NAD during the first 24 h postoperatively. Conclusion: The correlation of IL-18 levels to NRS and NAD values supports the hypothesis that ISR and pain are related.
Cytokines (CYTs) are cell-signaling molecules that aid cell-to-cell communication and enhance the movement of human cells towards sites of trauma, and inflammation (1). About 200 different CYTs has been reported, with multiple functions in regulating acute phase response (APR), steering cell growth/differentiation, activating antimicrobial defense and regulating homeostasis. CYTs especially mediate innate immunity and assist in the development of adaptive immunity regulating white cell growth and differentiation. Interleukin-18 (IL-18) is a member of the IL-1 family of CYTs (1-3). IL-18 is also known as interferon-gamma inducing factor (IGFN-γ), which in humans is encoded by the IL18-gene (2, 3). IL-18 can modulate both innate and adaptive immunity and its dysregulation can cause autoimmune or inflammatory diseases (2, 3). IL-18 has also been found to increase amyloid-beta production in human neurons in patients with Alzheimer’s disease (4). Zhang et al. (5) suggested IL-18 as a biomarker of early diabetic nephropathy in patients with type 2 diabetes associated with enhanced urine protein excretion. APR proteins and CYTs are thought to be early measures of inflammatory stress response (ISR) induced by trauma. There is also evidence suggesting that ISR and pain are related (6). Aspinen et al. (7) reported in 2016 that the ISR in patients with minilaparotomy cholecystectomy (MC) versus laparoscopic cholecystectomy (LC) was equal based on the interleukin (IL)-8, IL-10 and IL-1β values. Interestingly, Purdy et al. (6) found a significant correlation in the NRS pain scores versus the values of anti-inflammatory CYT IL-10 and pro-inflammatory CYT IL-1β suggesting that ISR and pain are related in midline laparotomy patients. However, the correlation of IL-18 and the ISR in patients with LC versus MC is unknown. The aim of this study was to assess the association between the ISR biomarkers and patients’ pain experience and number of analgesic doses (NAD) following surgery.
Patients and Methods
The study was approved by the Ethics Committee of Kuopio University Hospital District, Kuopio, Finland (DNRO 27/02/2013), it was registered in the ClinicalTrials.gov database (ClinicalTrials.gov Identifier: NCT01723540, Consort diagram, Figure 1), and was conducted in accordance with the Declaration of Helsinki. The study protocol and inclusion/exclusion criteria of the study patients are detailed in previous reports by Saimanen et al. (8, 9).
The blood samples were taken before surgery (PRE), immediately after surgery (POP1) and 6 h post-operatively (POP2) and centrifuged at 1,000 × g (2,900 rpm) for 15 min. The interleukin assays were performed using ELISA methods from R&D Systems (Minneapolis, MN, USA). The manufacturer’s intra-assay and the inter-assay coefficient of variations for IL-18 were 2.5-3.1 % and 7.9-8.7%, respectively. The sensitivity of the other CYT assays were detailed in previous reports by Aspinen et al. (7). Plasma high-sensitivity C-reactive protein (hs-CRP) was analyzed with a Cobas 6000-analyzer (Roche Diagnostics, Penzberg, Germany).
Data are presented as means and standard deviations or frequencies and percentages, in Table I. In Table II and Table III, the results of the laboratory measurements are presented as median concentrations with interquartile range as distributions were right-skewed. Differences in baseline characteristics between groups were tested by the Fisher’s exact test and in the case of continuous data, the analysis was performed using t-test. The blood CYT concentrations and differences in MC and LC groups of patients were tested by the Mann–Whitney U-test. The alterations between time points were tested by the Wilcoxon signed rank test. In the linear mixed effect (LME) analysis interleukin and hs-CRP blood concentration were log transformed. The Pearson’s method was used to test for correlation of IL-18 concentrations to NRS and NDA values (Figure 2 and Figure 3). Data were analyzed using the IBM SPSS statistical software (IBM SPSS Statistics for Windows, version 26.0, IBM Corporation Armonk, NY, USA).
Results
The basic patient data of the study. In the MC group there were 60 patients (49 females and 11 males) versus 54 patients in the LC group (36 females and 18 males, Figure 1), with the mean (SD) age of 50.7 (13.2) years versus 53.2 (13.1) years. There were no significant differences between the MC and LC study groups in mean age, mean height (167.4 and 168.9), mean weight (77.3 and 83.0), mean body mass index (BMI, 27.6 and 29.1), operative time (70 and 70.1 min), time in the operative room (120.4 and 127.1 min), perioperative bleed (41 and 31 ml) or conversion rate (3 and 3). Interestingly, the surgical incision was significantly shorter in the MC than in the LC group (49.7 mm and 78.3 mm, p<0.001, Table I).
The APR in MC versus LC patients. The median biomarker concentrations between MC and LC groups were equal. Changes in blood hs-CRP and interleukin concentrations preoperatively and following surgery in different time points between MC and LC patients are shown in Table II. IL-18, hs-CRP, IL-1ra, IL-6, and IL-10 had a trend for increase following surgery in MC and LC patients and the LME interaction time between MC and LC groups was statistically significantly different for hs-CRP and IL-1ra biomarkers (p=0.033 and p=0.042, respectively, Table II). In all patients, the LME time-effect between different time points was statistically significantly different for hs-CRP, IL-1ra, IL-6, IL-10 and IL-1β biomarkers (p<0.001, p<0.001, p<0.001, p<0.001 and p=0.014, respectively, Table III).
The IL-18 cytokine versus NRS and NAD. IL-18 concentrations correlated significantly to IL-1ra concentrations (r=0.153, p=0.03). However, there were no significant correlations in Pearson’s method in between IL-18 concentrations and hs-CRP, IL-ra, IL-6, IL-10, and IL-1β concentrations. There was a significant inverse correlation between IL-18 concentrations and NRS pain scores during the first 24 h following surgery (r=−0.173, p=0.020, Figure 2) and a significant inverse correlation between the median IL-18 concentrations and NAD in all patients (r=−0.255, p=0.002, Figure 3).
Discussion
The pain following surgery is the main negative effect associated with surgical procedure. Postoperative pain after cholecystectomy derives from irritation of multiple origins: the peritoneum, abdominal wall, and abdominal viscera. The modern treatment of pain following surgery includes regional pain blocks, paracetamol, nonsteroidal anti-inflammatory drugs (NSAIDs), and opioid analgesics (6). In addition, enhanced recovery after surgery protocols (ERAS) are increasingly used in surgical patients and these programs warrant effective pain treatment that promote early mobilization of the patient (10). It is evident that appropriate pain control is necessary as cholecystectomy decreases pulmonary function by as much as 30% even when effective pain management is administered (11). Future goals include the development of peripherally restricted opioid agonists, selective targeting of opioid-containing immune cells to sites of trauma, and the augmentation of peripheral ligand and receptor synthesis. The main aim is to avoid harmful side effects of available opioid analgesics such as respiratory depression, cognitive impairment, addiction, gastrointestinal bleeding, and thromboembolic complications (12).
Presently, there is evidence suggesting that ISR and pain are related. Purdy et al. (6) found a correlation between NRS pain scores and blood values of anti-inflammatory IL-10 and pro-inflammatory IL-1β, suggesting that ISR and pain are related in midline laparotomy patients. However, the correlation of IL-18 and ISR in patients with LC or MC is unknown. The aim of our study was to assess the correlation between the ISR and patients’ pain experience and NAD. The present study showed that the ISR in LC and MC groups was quite similar based on the blood levels of IL-1ra and IL-10, an anti-inflammatory CYT preventing tissue damage caused by ISR and IL-1β, a proinflammatory acute-phase protein. A new finding in the present work is the slightly higher relative elevation in the mean concentrations of IL-18 and IL-8 postoperatively in the LC group. Although, the median concentrations of hs-CRP did increase in the LC group, the plasma levels of hs-CRP were doubled in the MC group at six hours following surgery compared to baseline. In addition, the LME p-values for interaction time between the MC and LC study groups in hs-CRP and IL-1ra biomarkers were statistically significant. As the median values of IL-1ra are several folds higher than other CYTs, even quite small relative alterations in median values could be reliably detected. Plasma levels of Il-6, a proinflammatory CYT, were increased in both groups, but the increase was 6-fold in the MC group compared to 3-fold increase in the LC group at six hours after surgery. The blood levels of hs-CRP and six interleukins (IL-18, IL-1ra, IL-6, IL-8, IL-10, IL-1β) measured at three time points; before operation (PRE), immediately after operation (POP1) and six hours after operation (POP2) were log-transformed to avoid showing right skewed values (Table II).
The hypothesis that the ISR and pain could associate is based mostly on suggested interactions between opioid ligands and receptors. Inflammation activates the opioid receptors on peripheral sensory neurons and leads to local production of endogenous opioid substances (1-3, 12). These findings have interesting implications for the development of safer analgesics, where modern treatment can communicate with peripheral sensory neurons to modulate pain. Future strategies take in to account that medication should be particularly active in local pathology rather than producing a suppression of the central nervous system, as with classical opioid analgesics. It is possible that in the future, specific CYTs or antagonists will be developed to act as blockers of the hyperexcitability in the sensory neurons (12, 13). Interestingly, Palat et al. found (13), that blockade of IL-18 signaling diminished neuropathic pain and enhanced the efficacy of the opioid analgesics morphine and buprenorphine.
In conclusion, our results suggest that the ISR for cholecystectomy in MC and LC patients was quite equal based on the interleukin and hs-CRP values. A novel finding in the present study is that IL-18, a member of the IL-1 family of CYTs, correlates to NRS pain scores and NAD in cholecystectomy patients. The correlation in IL-18 levels to NRS and NAD values supports the hypothesis that the ISR and pain are related.
Acknowledgements
The study was funded by the Päivikki and Sakari Sohlberg Foundation, Helsinki, Finland.
Footnotes
Authors’ Contributions
All Authors contributed to the collection and analysis of data, drafting and revising the manuscript, read and approved the final article.
Conflicts of Interest
The Authors report no conflicts of interest or financial ties to disclose.
- Received August 12, 2022.
- Revision received August 30, 2022.
- Accepted September 5, 2022.
- Copyright © 2022 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.
This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY-NC-ND) 4.0 international license (https://creativecommons.org/licenses/by-nc-nd/4.0).