HCC generally develops following an orderly progression from cirrhosis to dysplastic nodules to early cancer development, which can be reliably cured if discovered before the development of vascular invasion . Early detection of HCC in those patients provides the best chance for a curative treatment, but AFP levels are frequently normal in patients with small HCC and are not elevated in a significant proportion of patients with early-stage, potentially curable HCC.
Elevated concentrations of cytokines represent a characteristic feature of CLD, regardless of the underlying etiology, and may represent a consequence of liver dysfunction instead of an inflammatory disorder . Cytokines imbalance between T-helper 1 (Th1) and T-helper 2 (Th2) can prolong inflammation, leading to necrosis, fibrosis and CLD  in addition to the development and progression of HCC . Cytokine production is thought to play an important role in the recruitment of tumor associated inflammatory cells, induction of angiogenesis and direct modulation of tumor cell proliferation [38, 39]. The cytokines studied in this work were carefully chosen to include cytokines of the Th1 repertoire (IL-2R and sTNFR-II), in addition to one of the important pro-inflammatory cytokines (IL-8), and other factors as sFas.
In the present study, liver function tests were significantly elevated whereas log-HCV titer was significantly lower in HCC patients (p < 0.001) when compared to PNALT and CLD patients. In agreement with our findings, HCC group had the highest values (86.3%) for various concurrently-measured liver function tests, significant higher values of AST/ALT, ALT, AST (each, p < 0.001) than cirrhotic patients as previously reported . On the other hand, HCV levels were markedly higher in non-cancerous liver than in HCC (p = 0.001) . Moreover, comparing HCV titers of four HCC isolates and surrounding cirrhotic liver tissues in two anti-HCV positive patients; the copy numbers of HCV-RNA were 1 × 106 and 4 × 106/gm wet weight of HCC, and 8 × 107 and 3.2 × 108/gm wet weight of cirrhotic liver tissues from patient-1 and -2, respectively . The present study showed that men had higher log-HCV RNA titer than that detected in women; then, a strong evidence is provided in favour of a higher HCV clearance rate in women compared with that in men .
Fas (APO-1 or CD95) is a cell-surface receptor that transduces apoptotic signals from Fas ligand (Fas-L) . Apoptosis is tightly regulated throughout a variety of mechanisms, one of which is postulated to be the production of soluble forms of Fas (sFas) that normally binds to Fas-L, thus blocking the signaling of the membrane-bound form of Fas. Peripheral blood mononuclear cells in HCV infection exhibit decreased susceptibility to Fas-L induced cell death. This may signify a mean by which HCV escapes immune surveillance; however, it would be worth a further investigation on this phenomenon. The sFas appeared to increase in advanced stages of HCV-induced liver disease, as a result of host-related immunological factors . In the present series, the mean values of sFas were significantly higher in HCC patients compared to the other groups (p < 0.001). This could be explained by the role of sFas in the inhibition of apoptosis, progression to end stage liver damage, and subsequent development of HCC. Similarly, a significant elevation of serum levels of sFas in HCC patients compared with liver cirrhosis and healthy control was previously reported . Previous studies [47, 48] have reported mRNA encoding secreted sFas in a number of hepatitis and HCC cases indicating that sFas may function as an inhibitor of the Fas/Fas-L system and escape of tumor cells from immune surveillance may then occur. In chronic hepatitis, sFas was correlated with the severity of disease  and its expression can illustrate the mechanism of liver injury caused by death receptors throughout the multistep process of fibrosis/carcinogenesis. So, the increased incidence of HCC is correlated not only with the higher degree of hepatic fibrosis, but also with the lower expression of Fas protein .
The rate of progression to end-stage liver disease might be related to an up-regulation of the TNF-α/Fas pathways and an age-dependent host response . Pro-inflammatory TNF-α released by host and tumor cells is an important factor involved in initiation, proliferation, angiogenesis as well as metastasis of various cancer types . Activities of TNF-α are mediated through TNFR-I and TNFR-II . Our results showed that levels of sTNFR-II were elevated in patients with PNALT, CLD and HCC with a significant difference between HCC in relation to the other two groups (p < 0.001). These results are in agreement with previous published results [13, 29, 53], where it was found that sTNFR-IIα were closely correlated with disease progression in chronic HCV infection. Enhanced TNF-α and TNFRs in chronic HCV infection may reflect the histological activity of the disease and TNFRs up-regulation might modify host response and potentially contribute to liver damage .
IL-2 is a cytokine produced by T cells in response to inflammatory stimuli. It induces the surface expression of IL-2 receptor (IL-2R) and, consequently, the production of its soluble form, sIL-2R. The excess of sIL-2R is capable of binding IL-2 and causes the inhibition of an appropriate immune response. IL-2R is the protein that mediates the action of IL-2, which is normally not displayed at a significant number on T and B cell surfaces. Stimulation of the immune system causes two IL-2R changes: more molecules of "IL-2R" expressed on the cell plasma membrane and sIL-2Rα is released by the activated cells into the surrounding fluid . Our results showed that levels of IL-2Rα were elevated in all studied patients with a statistically significant difference in HCC patients when compared to those with PNALT (p = 0.001). This could be attributed to the binding of IL-2 due to excess of its receptor and thus inducing an inhibition of the appropriate immune response with subsequent progression of chronic liver disease and the development of HCC. Previous results [13, 17, 56] are in agreement with ours, where it is was shown that serum levels of sIL-2R are correlated with the histological severity of liver damage in HCV patients, which may be used as a marker in patients at high risk of getting HCC as the highest levels of soluble IL-2R occurred in those patients. The sIL-2R may be an important marker for assessing the phase of active chronic hepatitis and the degree of liver damage . High sIL-2R levels, found in patients with chronic HBV [58, 59], were related to the activity of the disease rather than to the virus replication; thus, those levels may be a useful marker of T-cells immune response. In contrast to our results, it was concluded that IL-2R was not detectable in HCC patients in comparison to patients with chronic hepatitis and liver cirrhosis . Regarding the levels of IL-2R in patients with HCC, and in agreement with our findings, there was no statistically significant difference (p = 0.62) between its values in men and women .
IL-8 is a chemoattractant cytokine which is produced after stimulation with numerous exogenous and endogenous agents. Viruses induce IL-8 production leading to enhanced viral RNA replication and cytopathic effects. Furthermore, evidence was provided that induction of that interleukin was able to attenuate the IFN-α mediated inhibition of viral replication . In the current study, levels of IL-8 were significantly lower in HCC patients than in the other groups (p < 0.001). On the contrary, other results found that serum IL-8 levels were markedly elevated in most HCC patients compared with healthy subjects  and was found to be over expressed in the HCC tumor cells compared with the non-tumorous livers . Furthermore, multivariate analyses revealed that the levels of the interleukin under consideration may play an important role in the progression and dissemination of HCC and is an independent predictor of long-term survival among those patients. High-serum level of that cytokine may reflect active angiogenesis and rapid tumor growth in HCC. Therefore, targeting IL-8 can represent a potential approach to control angiogenesis and invasion of HCC . In agreement with our results, there was no significant correlation between serum concentration of that cytokine and patient gender (p = 0.215) .
The present series showed that HCV viral load was significantly correlated with sTNFR-II and IL-8. The production of the latter was found to enhance viral RNA replication , thus the low levels of the interleukin in our HCC patients are in accordance with the low HCV viral load. Moreover, there is a good correlation between reduction in virus load and IL-8 level which may indicate that it is related to viral infection rather than to hepatocarcinogenesis.
In the current series, the studied cytokines were significantly correlated to each other. The sFAS was positively correlated with sTNFR-II and IL-2R; sTNFR-II positively correlated with IL-2R and negatively with IL-8; lastly IL-2R and IL-8 were negatively correlated.
Th1 cytokines, which include IL-2R and sTNFR-II, are in favor of an effective immune response against viral infection, whereas Th2 (represented by IL-8 in our study), is in favor of progressive inflammation, continuous cell injury and persistent HCV infection .
The depicted correlations could highlight the imbalance between pro- and anti-inflammatory cytokines among patients with CLD and HCC. Furthermore, the rate of progression of CHC to end-stage liver disease might be related to an up-regulation of the TNF-α/Fas pathways .
Analysis of sTNFR-II and IL-8 by ROC curves revealed satisfactory values regarding sensitivity and specificity at a cutoff value of ≥ 398 pg/ml and ≤ 290 pg/ml, respectively, when both markers were combined. Therefore, a simultaneous assessment of both sTNFR-II and IL-8 would be beneficial for the diagnosis of HCC; in fact, they were capable of differentiating between patients with PNALT and HCC -- hence, an early detection of HCC among apparently healthy patients with PNALT levels. Nonetheless, these values must be evaluated on a larger scale of patients with various stages of CLD and HCC, in order to be used as new markers for an early detection of HCC.