Serum Interleukin-22 as a Marker of Decompensated Liver Cirrhosis and Hepatocellular Carcinoma in Chronic Hepatitis C Virus Patients

INTRODUCTION

Hepatitis C virus infection is one of the most significant health concerns in human civilizations. Today, the virus infects 170 to 200 million individuals worldwide, with an additional 40,000 persons adding to the figure each year, (more than 3% of the global population).[1] Egypt had the highest HCV prevalence worldwide. Egypt developed a feasible paradigm of care for managing that severe illness which has the highest global disease prevalence. This paradigm could treat around 4 million patients.   Although Egypt has made great efforts in managing such diseases, several obstacles can delay the disease's total elimination [2].

Hepatitis C virus (HCV) is the most common reason for chronic liver disease, cirrhosis, hepatocellular carcinoma, and eventually liver transplantation in several countries, and it is also the most common reason for liver transplantation. Several factors affect the prognosis of chronic HCV infection such as age, race, gender, and the immune response of the virus. A chronic HCV infection will develop in about 55% to 85% of people infected with the virus. They also have a higher risk of developing extrahepatic symptoms,  compensated and decompensated cirrhosis, and ultimately HCC [3].

Liver cirrhosis is the last stage of a multistep process, initiated from a primary hepatotoxic effect and resulting in insufficient liver function due to excessive hepatocyte destruction, Despite having the highest functional reserve of any organ, the cirrhotic liver rapidly loses its ability of compensation for the chronic influence of a causal agent due to severe biochemical and physiological alterations [4].

Necrosis occurs after hepatic injury by HCV and is followed by hepatocyte proliferation [5]  This repetitive cycle of destruction and regeneration leads to chronic liver disease development and finally cirrhosis [6]. Cirrhosis is marked by abnormal liver nodule formation which is encircled by collagen deposition and liver scarring. Eventually, hyperplastic nodules emerge, progressing to dysplastic nodules, and ultimately resulting in the formation of HCC [7].

In Egypt, hepatocellular carcinoma (HCC) ranks as the fourth cause of cancer-related deaths [8] and the sixth most common cancer worldwide [9].  The incidence of this disease has increased due to the advancements in screening programs and diagnostic technologies, raising the rate of cirrhotic patient survival, which raises the rate of HCC development, as well as hepatitis C virus (HCV), which is considered the commonest cause of HCC and hepatic cancers in Egypt. It takes 20 to 40 years for HCC to develop as a result of HCV [10].

 Both host-induced immune responses and virus-induced factors influence HCV carcinogenesis. Although there is only evidence of HCV's direct carcinogenic effect on liver cells in experimental animals, studies have revealed that HCV core protein may induce oxidative stress metabolism and cause lipogenesis [11]. 

HCV viral proteins may promote the development of HCC by deactivating tumor suppressor genes, and cell cycle checkpoints, or by stimulating growth and division-promoting signaling pathways [12]. HCV's nonstructural protein genes activate hepatic stellate cells and trigger transforming growth factor-beta, which both lead to fibrosis and hepatocellular carcinoma (HCC) [13]. HCV-induced immune response is modulated through persistent inflammation, tumor necrosis factor, and interferons (IFNs) [14].

IL-22 was initially discovered as a gene that reacted to interleukin-9 (IL-9) in T lymphoma cells of mice  . It was originally known as IL-10-related T-cell-derived inducible factor (IL-TIF) due to its encoding of a glycosylated protein sharing 22% sequence identity with interleukin-10 (IL-10) [15].

 In human genome, the gene of IL-22 is found on chromosome 12q15, near to the interleukin-26 (IL-26) and interferon-c (IFN-c) genes, forming an interleukin gene cluster . The chromosomal structure of IL-22 contains six exons and five introns, showing comparable sizes and organizational patterns when compared to IL-10 [16].

It has been shown that Th17 cells are able to produce IL-22 and IL-17 together [17]. When it was found that IL-22 affects hepatocytes by interacting with the IL-22R1/IL-10R2 receptor complex, its dual function became evident. Investigations were done to determine IL-22R1 receptor expression to determine the functional targets of IL-22 [18].

HCV patients had considerably high levels of IL-22-producing T helper (Th17) cells in their livers [19].  In contrast to cholestatic liver disorders, HCV hepatitis showed higher hepatic IL-22 mRNA expression. However, antiviral proteins 2',5'-oligoadenylates, and MxA synthesis as well as  IFN-α/β expression were not significantly affected by IL-22, indicating that in vitro models of virus C infection and replication show a lack of antiviral activity [20].

IL-22 is implicated in several liver illnesses, including cirrhosis, fibrosis, and inflammation.    After the hepatic injury, HSCs predominantly cause liver fibrosis, expressing elevated levels of IL-22R1 and IL-10R2. Both in vivo and in vitro studies show that IL-22's ability to suppress HSC apoptosis and improve their survival by stimulating anti-apoptotic genes. [21]

Notably, the recognized tumor-infiltrating leukocytes (TILs) of human HCC have markedly elevated IL-22 expression, with higher levels seen in advanced tumor grades [22].  Additionally, IL-22 enhanced the progression of HCC by activating STAT3 signaling, thereby inducing the expression of pro-tumorigenic genes like CyclinD1, Bcl-2, Bcl-XL, and the known vascular endothelial growth factor (VEGF), which facilitate growth and metastasis of the tumor   [23].

Our study aimed to identify the serum levels and diagnostic performance of Interleukin-22 in HCV decompensated cirrhotic patients as well as HCC patients. We also studied the relation between serum IL-22 and Fib-4, APRI, and Child-Pugh scores.

PATIENTS AND METHODS

The study involved eighty participants divided into four groups: 20 patients had decompensated liver cirrhosis caused by HCV (Child-Pugh B or C),20 patients had compensated liver cirrhosis caused by HCV (Child-Pugh A),20 patients with HCV-induced liver cirrhosis and HCC, and 20 healthy volunteers as a control group. Participants with other conditions like sepsis, spontaneous bacterial peritonitis, hepatorenal syndrome, or malignancies were excluded. All participants underwent a clinical examination and medical history, with a focus on identifying signs of chronic liver disease.

Laboratory tests were conducted to assess liver function, kidney function, blood glucose levels, and serum electrolytes. The APRI and FIB-4 scores, which are non-invasive measures to diagnose liver fibrosis, were also calculated. Serum Interleukin-22 level was measured using an ELISA test.

Imaging studies, including abdominal ultrasounds, were performed to assess the liver, spleen, and presence of ascites or focal lesions. If any focal lesions were detected on ultrasound, a triphasic CT scan of the liver was conducted for further evaluation. Informed consent was obtained from all the participants before their involvement in the study.

Statistical analysis

 Data were fed to the computer and analyzed using IBM SPSS software package version 20.0. (Armonk, NY: IBM Corp) Qualitative data were described using numbers and percentages. The Shapiro-Wilk test was used to verify the normality of distribution Quantitative data were described using range (minimum and maximum), mean, standard deviation, median, and interquartile range (IQR). The significance of the obtained results was judged at the 5% level. The used tests were the Chi-square test, Monte Carlo correction, F-test (ANOVA),  Kruskal Wallis test, Receiver operating characteristic curve (ROC), Sensitivity,  Specificity, Positive Predictive value (PPV), and Negative Predictive value (NPV).

RESULTS

Patients with decompensated cirrhosis (Mean = 49.8 ng/L) and HCC (Mean = 97.3 ng/L) had significantly higher serum interleukin-22 levels than patients with compensated cirrhosis (Mean = 28 ng/L) and healthy controls (Mean = 9 ng/L; P<0.001), according to the study. Moreover, IL22 levels were significantly higher in HCC patients than in decompensated patients (P<0.05).

Additionally, IL-22 levels showed a significant positive correlation with tumor size in HCC patients (r = 0.481, P = 0.032), and with serum AFP levels in decompensated cirrhosis (r = 0.459, P = 0.042) and HCC patients (0.581, P = 0.007). Also, IL-22 levels were positively correlated with non-invasive fibrosis markers (APRI and FIB-4 scores) in all patient groups (P<0.05). Additionally, in patients with HCC, IL-22 levels in patients with Child Class C (Mean = 58.4 ng/L) were higher than in patients with Child Class B (Mean = 45.1 ng/L; P = 0.016).

Importantly, a serum IL-22 level of >34 ng/ml was identified as a potential cutoff value for discriminating between compensated and decompensated cirrhosis with high sensitivity (95%) and specificity (80%). Moreover, a cutoff value of >81 ng/L showed good diagnostic performance in differentiating between HCC patients from decompensated cirrhosis (AUC = 0.976, sensitivity = 95%, specificity = 95%). High levels of IL22 were a good predictor of HCC in decompensated cirrhosis according to Univariate logistic regression analysis.

 Table I Comparison between the four studied groups considering Serum Interleukin 22-level

Serum Interleukin 22 level Units (ng/L= nanogram per liter ),

n= number of patients,

IQR=Interquartile rang

SD =Standard deviation.

         DISCUSSION