hepatocellular carcinoma ( hcc ) is the second most common cause of cancer - related death worldwide . with the exception of japan , hcc in asian countries
most often occurs in patients with underlying chronic hepatitis b virus ( hbv ) infection through a multistep process of hepatocarcinogenesis . with the development of diagnostic and operative techniques , curative hepatic resection for this fatal disease
nevertheless , the high recurrence rate following curative resection hampers the long - term benefits of surgical treatment .
therefore , identifying factors that contribute to hbv - associated hcc recurrence could provide potential targets for novel therapeutic strategies .
recently , the expression profiling of fixed tissue hcc and nontumor tissue has been used to gain insight into hcc recurrence .
the results indicated that most cases of hcc recurrence after curative therapy are not metastasis from the primary tumor , but rather de novo cancers arising within the cirrhotic liver .
these findings prompted our investigation into the role of inflammation cytokines that are closed related to tumorigenesis in hcc recurrence .
interleukin-6 ( il-6 ) , produced by b - cells , t - cells , macrophages , and fibroblasts , regulates chronic inflammation and , thus , creates a cellular microenvironment beneficial to cancer growth .
the potential association of il-6 with the risk of hcc has been explored in previous studies .
however , it is not clear whether this inflammatory cytokine is associated with hcc recurrence after curative resection in patients with chronic hbv infection . despite recommendations regarding periodic follow - up with imaging and tumor markers such as alpha - fetoprotein ( afp ) and fibrosis markers , these strategies have not proven efficacious in the early prevention of hcc recurrence .
the currently study analyzed the relationship between preoperative serum il-6 and hbv - associated hcc recurrence after curative resection .
in addition , the correlation between serum il-6 levels and the original tumor characteristics was also explored to shed potential light on the underlying etiology of hbv - associated hcc recurrence .
this study was performed in accordance with national legislation and with the approval of the ethical committee of our hospital .
we retrospectively collected patients data from original medical records and the cancer patients follow - up database at our hospital .
serum il-6 was measured using eclia , modular - system ( roche , mannheim , germany ) . in the current study laboratory test results obtained immediately prior to the date of surgery
were selected to exclude interference from other preoperative invasive examinations . during 2008 to 2009 at our hospital , 183 consecutive patients with chronic hbv infection underwent liver resection for primary hcc .
out of the 183 patients , 37 patients were excluded from this study for the following reasons : 12 patients received noncurative resection , 3 patients died in the perioperative period , 15 patients received preoperative transarterial chemoembolization , and 7 patients had malignancy other than hcc .
all patients received laboratory assessment of liver function , preoperative viral serologic testing , and diagnostic imaging [ including computed tomography ( ct ) , magnetic resonance imaging ( mri ) , or positron emission tomography - computed tomography ( pet - ct ) ] to evaluate tumor location and tumor characteristics .
patients who had neither hilar nodal involvement nor extrahepatic metastases were treated with liver resection as the initial treatment for their primary hcc .
the selection criteria and type of operative procedures were determined according to preoperative indocyanine green retention rate at 15 minutes ( icgr15 ) , as previous described .
in general , patients with an icgr15 less than 35% were selected for anatomic resection .
after the operation , patients received routine follow - up with physical examination , serum afp level , and ultrasonography at 3-month intervals for the first year and then every 6 months .
suspected hcc recurrence was confirmed using mri , hepatic angiography , and percutaneous biopsy in some cases .
additional examinations , such as chest ct , bone scan , or pet - ct , were also performed if there was any sign of extrahepatic recurrence .
the end of follow - up was determined as either the time of last follow - up ( january 2013 ) or death .
case - controlled differences were assessed using student t - test , pearson chi - square test , and the fisher exact test , where appropriate ( table 1 ) .
the association between serum il-6 level and greatest tumor dimension and hbsag amount were analyzed by the pearson correlation test ( figures 1 and 2 ) .
exploratory stratified analysis was performed and p for interaction was calculated from the log likelihood ratio test comparing 2 nested models ( table 2 ) .
the relationship between serum il-6 and the risk of hcc recurrence was explored using a smoothing plot ( figure 3 ) .
a 2-piecewise linear regression model was used to examine the saturation effect of serum il-6 on the risk of hcc recurrence , according to the smoothing plot ( table 3 ) .
an inflection of serum il-6 level , at which the relationship between serum il-6 levels and the risk of hcc recurrence began to diminish was determined using a trial method .
the latter involved moving the trial inflection point along a predefined interval and detecting the inflection point that gave the maximum likelihood .
receiver operating characteristics ( roc ) curve was used to define the optimal cutoff value , sensitivity , and specificity ( figure 4 ) . the overall survival
( os ) and recurrence - free survival ( rfs ) rates were calculated using the kaplan - meier method .
the difference between the 2 groups was determined using the log rank test ( figure 5 ) .
baseline characteristics of 146 patients with chronic hbv infection who underwent curative hepatectomy for hepatocellular carcinoma characteristics significantly associated with serum il-6 levels .
( a ) serum il-6 is significantly higher in patients with multiple hepatic tumors compared with patients with only solitary tumor recurrence ( 3.5 vs 3.0  pg / ml , p  =  0.04 ) .
( b ) serum il-6 level has some relevance with hbsag amount ( r  =  0.504 , p  <  0.001 ) .
there is no significant difference in serum il-6 levels ( a ) between portal vein invasion versus no portal vein invasion ( 3.6 vs 3.2  pg / ml , p  =  0.20 ) , and ( b ) between microvascular invasion versus no microvascular invasion ( 3.5 vs 3.2  pg / ml , p  =  0.25 ) .
( c ) serum il-6 levels are not significantly associated with the maximal tumor dimension ( r  =  0.0003 , p  =  0.84 ) .
stratified analysis of factors affecting the correlation between preoperative serum il-6 and hcc recurrence the adjusted relationship between serum il-6 level and the risk of hcc recurrence . a nonlinear relationship between serum il-6 level and risk of hcc recurrence
is observed after adjusting for age , tumor number , portal vein invasion , and microvascular invasion .
threshold effect analysis of serum il-6 level on hepatocellular carcinoma recurrence using piecewise linear regression receiver operating characteristics ( roc ) curve analysis using serum il-6 level for predicting hcc recurrence .
the serum il-6 level predicts hcc recurrence using the area under the curve ( auc ) of 0.70 ( 95% ci : 0.620.78 ) . the optimal cutoff point for preoperative serum il-6 level in predicting
hcc recurrence is 3.1  pg / ml , with a sensitivity of 66.2% and a specificity of 74.2% .
the recurrence - free survival curves and the overall survival curves in patients with low versus high serum il-6 level .
( a ) the recurrence - free survival rates in the high serum il-6 group are significantly higher than those in the low serum il-6 group ( p  <  0.01 ) .
( b ) the overall survival rates in the high serum il-6 group are significantly higher than those in the low serum il-6 group ( p  <  0.01 ) .
this study was performed in accordance with national legislation and with the approval of the ethical committee of our hospital .
we retrospectively collected patients data from original medical records and the cancer patients follow - up database at our hospital .
serum il-6 was measured using eclia , modular - system ( roche , mannheim , germany ) . in the current study laboratory test results obtained immediately prior to the date of surgery
were selected to exclude interference from other preoperative invasive examinations . during 2008 to 2009 at our hospital , 183 consecutive patients with chronic hbv infection underwent liver resection for primary hcc .
out of the 183 patients , 37 patients were excluded from this study for the following reasons : 12 patients received noncurative resection , 3 patients died in the perioperative period , 15 patients received preoperative transarterial chemoembolization , and 7 patients had malignancy other than hcc .
all patients received laboratory assessment of liver function , preoperative viral serologic testing , and diagnostic imaging [ including computed tomography ( ct ) , magnetic resonance imaging ( mri ) , or positron emission tomography - computed tomography ( pet - ct ) ] to evaluate tumor location and tumor characteristics .
patients who had neither hilar nodal involvement nor extrahepatic metastases were treated with liver resection as the initial treatment for their primary hcc .
the selection criteria and type of operative procedures were determined according to preoperative indocyanine green retention rate at 15 minutes ( icgr15 ) , as previous described . in general ,
after the operation , patients received routine follow - up with physical examination , serum afp level , and ultrasonography at 3-month intervals for the first year and then every 6 months .
suspected hcc recurrence was confirmed using mri , hepatic angiography , and percutaneous biopsy in some cases .
additional examinations , such as chest ct , bone scan , or pet - ct , were also performed if there was any sign of extrahepatic recurrence .
the end of follow - up was determined as either the time of last follow - up ( january 2013 ) or death .
case - controlled differences were assessed using student t - test , pearson chi - square test , and the fisher exact test , where appropriate ( table 1 ) .
the association between serum il-6 level and greatest tumor dimension and hbsag amount were analyzed by the pearson correlation test ( figures 1 and 2 ) .
exploratory stratified analysis was performed and p for interaction was calculated from the log likelihood ratio test comparing 2 nested models ( table 2 ) . the relationship between serum il-6 and the risk of hcc recurrence
a 2-piecewise linear regression model was used to examine the saturation effect of serum il-6 on the risk of hcc recurrence , according to the smoothing plot ( table 3 ) .
an inflection of serum il-6 level , at which the relationship between serum il-6 levels and the risk of hcc recurrence began to diminish was determined using a trial method .
the latter involved moving the trial inflection point along a predefined interval and detecting the inflection point that gave the maximum likelihood .
receiver operating characteristics ( roc ) curve was used to define the optimal cutoff value , sensitivity , and specificity ( figure 4 ) . the overall survival
( os ) and recurrence - free survival ( rfs ) rates were calculated using the kaplan - meier method .
the difference between the 2 groups was determined using the log rank test ( figure 5 ) .
baseline characteristics of 146 patients with chronic hbv infection who underwent curative hepatectomy for hepatocellular carcinoma characteristics significantly associated with serum il-6 levels .
( a ) serum il-6 is significantly higher in patients with multiple hepatic tumors compared with patients with only solitary tumor recurrence ( 3.5 vs 3.0  pg / ml , p  =  0.04 ) .
( b ) serum il-6 level has some relevance with hbsag amount ( r  =  0.504 , p  <  0.001 ) .
there is no significant difference in serum il-6 levels ( a ) between portal vein invasion versus no portal vein invasion ( 3.6 vs 3.2  pg / ml , p  =  0.20 ) , and ( b ) between microvascular invasion versus no microvascular invasion ( 3.5 vs 3.2  pg / ml , p  =  0.25 ) .
( c ) serum il-6 levels are not significantly associated with the maximal tumor dimension ( r  =  0.0003 , p  =  0.84 ) .
stratified analysis of factors affecting the correlation between preoperative serum il-6 and hcc recurrence the adjusted relationship between serum il-6 level and the risk of hcc recurrence . a nonlinear relationship between serum il-6 level and risk of hcc recurrence
is observed after adjusting for age , tumor number , portal vein invasion , and microvascular invasion .
threshold effect analysis of serum il-6 level on hepatocellular carcinoma recurrence using piecewise linear regression receiver operating characteristics ( roc ) curve analysis using serum il-6 level for predicting hcc recurrence .
the serum il-6 level predicts hcc recurrence using the area under the curve ( auc ) of 0.70 ( 95% ci : 0.620.78 ) .
the optimal cutoff point for preoperative serum il-6 level in predicting hcc recurrence is 3.1  pg / ml , with a sensitivity of 66.2% and a specificity of 74.2% .
the recurrence - free survival curves and the overall survival curves in patients with low versus high serum il-6 level .
( a ) the recurrence - free survival rates in the high serum il-6 group are significantly higher than those in the low serum il-6 group ( p  <  0.01 ) .
( b ) the overall survival rates in the high serum il-6 group are significantly higher than those in the low serum il-6 group ( p  <  0.01 ) .
patients clinicopathological characteristics are summarized in table 1 . out of 146 patients , 80 ( 54.8% )
patients were documented as having hcc recurrence during the follow - up period ( group 1 ) . for patients with hcc recurrence compared with those patients without tumor recurrence ( controls , group 2 ) ,
serum il-6 levels were significantly higher ( 3.7 vs 2.8  pg / ml , respectively , p  <  0.01 ) and the maximal tumor dimensions were significantly larger ( 6.2 vs 5.3  cm , respectively , p  <  0.01 ) . moreover , multiple tumors and portal vein invasion were more common in hcc recurrence patients .
there were no significant differences , however , regarding child - pugh score , alcohol consumption , afp concentration , liver function tests , intraoperative factors , hcc stages , or degree of tumor differentiation ( p  >  0.05 ) .
the relationship between serum il-6 levels and primary tumor characteristics is shown in figures 1 and 2 .
serum il-6 levels were significantly higher in patients with multiple tumors compared with patients with a solitary tumor ( 3.5 vs 3.0  pg / ml , respectively , p  =  0.04 , figure 1a ) .
in addition , serum il-6 level has some relevance with hbsag amount ( r  =  0.504 p  <  0.001 , figure 1b ) .
however , no significant difference in serum il-6 levels were found between portal vein invasion versus no portal vein invasion ( 3.6 vs 3.2  pg / ml , p  =  0.20 , figure 2a ) , and between microvascular invasion versus no microvascular invasion ( 3.5 vs 3.2  pg / ml , p  =  0.25 , figure 2b ) . furthermore , there was no significant correlation between preoperative serum il-6 levels and maximal tumor dimension ( r  =  0.0003 , p  =  0.84 , figure 2c ) .
the effect of different clinicopathological features on the association between preoperative serum il-6 levels and hcc recurrence is shown in table 2 .
the predictive capability of preoperative serum il-6 levels was not significantly influenced by the number of tumors .
the preoperative serum il-6 was not associated with hcc recurrence in either the solitary or multiple tumor groups ( p for interaction  >  0.05 ) .
in addition , there was no evidence for an interaction between preoperative serum il-6 and maximal tumor dimension , microvascular invasion , or portal vein invasion ( all p  >  0.05 ) .
a multivariate regression analysis was performed to estimate the independent relationship between serum il-6 level and hcc recurrence , while adjusting for potential confounders . adjusted smoothed plots
suggested a nonlinear relationship between serum il-6 levels and risk of hcc recurrence ( figure 3 ) .
the risk of hcc recurrence increased with increasing serum il-6 level up to the turning point of 3.7  pg / ml ( or  =  3.8 , 95% ci 2.16.8 , p  <  0.001 ) . a serum il-6 level over 3.7  pg / ml was not associated with a higher risk of hcc recurrence ( or  =  0.89 , 95% ci 0.471.7 , p  =  0.71 ) ( table 3 ) .
receiver operating characteristic ( roc ) analysis was used to further verify the discriminating capacity of serum il-6 level to predict hcc recurrence ( figure 4 ) .
the results demonstrated that the preoperative serum il-6 level was able to distinguish hcc recurrence patients from controls with an area under the curve of 0.70 ( 95% ci 0.620.78 ) .
the optimal cutoff point for preoperative serum il-6 level in predicting hcc recurrence was 3.1  pg / ml , with a sensitivity of 66.2% and a specificity of 74.2% . according to the optimal cutoff value , patients with preoperative serum il-6 levels 3.1  pg / ml were defined as low il-6 ( n  =  76 ) , while patients with preoperative serum il-6 levels > 3.1  pg / ml were defined as high il-6 ( n  =  70 ) .
the rfs rate in the high il-6 group was significantly lower compared with the rate in the low il-6 group ( figure 5a ) ( p  <  0.01 ) .
the 1 , 3 , and 5-year rfs rates were 87.1% , 31.4% , and 21.4% , respectively , in the high il-6 group and 86.8% , 71.1% , and 61.8% , respectively , in the low il-6 group .
the os rate in the high il-6 group was also significantly lower compared with the os rate in the low il-6 group ( figure 5b ) ( p  <  0.01 ) .
the 1 , 3 , and 5-year os rates were 98.6% , 34.3% , and 22.9% , respectively , in the high il-6 group and 98.7% , 73.7% , and 65.8% , respectively , in the low il-6 group .
in patients with hbv - associated hcc , we determined that the preoperative serum il-6 level may serve as an independent and significant prognostic indicator of hcc recurrence after curative hepatic resection .
both rfs and os in the high il-6 group were significantly lower compared with the rates in the low il-6 group .
different from previous study , we also found a saturation effect when using serum il-6 to predict hcc recurrence .
inflammation is recognized as an important factor in the development and progression of malignancy . as an inflammatory cytokine ,
il-6 could activate the nuclear factor-b and the signal transducer and activator of transcription 3 , thereby contributing to hcc development .
our current data demonstrated that serum il-6 levels significantly correlated with tumor recurrence in patients with hbv - associated hcc , even after they received curative resection .
previous studies have demonstrated a significant correlation between hcc recurrence and unfavorable tumor factors , such as tumor size , multiple tumors , and the presence of microvascular invasion .
some researchers have explained the elevation in inflammatory cytokines as 1 aspect of a paraneoplastic syndrome , and as such , may predict hcc recurrence . in the current study , we found that tumor size , multiple tumors , and portal vein invasion were associated with hcc recurrence .
however , serum il-6 levels were not significantly associated with tumor size or portal vein invasion .
subsequent subgroup analysis suggested that serum il-6 levels could effectively predict hcc recurrence , whether in a solitary tumor or in multiple tumors .
in addition , although preoperative serum il-6 levels had some relevance with hbv amount , the association between serum il-6 levels and hcc recurrence was not significantly changed , after adjusting the hbv amount in the multivariate regression analysis .
these suggest that il-6 can serve as an independent factor for hcc recurrence and not just the reflection of unfavorable primary tumor factors or hbv infection status .
since our study population involved patients with chronic hbv infection , this difference may be explained as follows : serum il-6 levels are primarily affected by chronic hbv infection , liver fibrosis , and coexistent cirrhosis .
previous studies have shown that hbv x protein can regulate the expression of il-6 via protein phosphatase type 2 c. a portion of hcc recurrences after curative therapy might be de novo cancers arising in cirrhotic livers , as proposed by some researchers .
serum il-6 levels could be a comprehensive reflection of systemic metabolism , inflammation , and immune status .
the finding of a preoperative serum il-6 saturation effect when predicting hbv - related hcc recurrence after curative resection suggests a complex link between il-6 and tumor recurrence , that is , il-6 could have strong protumorigenic activity due to its multiple effects on tumor cell proliferation and survival , angiogenesis , tumor metabolism , and inflammation .
in addition , il-6 , alone or in combination with other cytokines , is an architect for shaping and generating immune responses which can exert a profound influence on hcc development .
in hepatoma cell lines and in rodents injected with hepatoma cells , il-6 actually inhibits instead of enhances tumor growth .
only after il-6 reaches a certain concentration can assume the multifaceted role needed to reach a balance and not increase the risk of tumor recurrence . to date , few studies have obtained such a quantitative finding , although the mechanisms underlying this result need to be further studied .
in addition , we also found that both rfs and os were significantly lower if preoperative serum il-6 levels exceeded 3.1  pg / ml .
since few studies have defined a cutoff value for il-6 when predicting hcc recurrence , measurements of preoperative il-6 level may help in selecting patients who would benefit from additional therapy following surgery , even after curative resection .
our study had several limitations including its retrospective nature , which limited our ability to detect il-6 in the tissues surrounding hcc .
, a complex interaction exists between tumor  host immunity and inflammatory response , but the key mechanisms underlying this response are not fully understood . however , elevated serum levels of il-6 were significantly associated with an increased risk of hbv - associated hcc recurrence , even after adjusting for unfavorable primary
. the causal role of il-6 in hcc recurrence can provide valuable information when selecting additional therapy following surgery .
targeting the il-6 system may also prove beneficial in the early prevention of this disease , and this area may prove fertile ground for future research .
 abstractthe aim of this study is to assess whether preoperative serum interleukin-6 ( il-6 ) can predict recurrence of hepatitis b virus ( hbv)-associated hepatocellular carcinoma ( hcc).the association between preoperative il-6 levels and hcc recurrence following curative hepatectomy in 146 patients with chronic hbv infection was determined . 
 patients were divided into groups based on the presence or absence of hcc recurrence . 
 serum il-6 levels were compared between groups , and the association between serum il-6 level and greatest tumor dimension was also analyzed . 
 receiver operating characteristics ( roc ) curve was used to define the optimal cutoff value for predicting recurrence - free survival ( rfs ) and overall survival ( os ) rates . 
 the os and rfs rates were calculated using the kaplan - meier method.out of 146 patients , 80 ( 54.8% ) patients were documented as having hcc recurrence during the follow - up period . after adjusting for potential confounders , serum il-6 levels were significantly associated with hcc recurrence , and a saturation effect existed with serum il-6 levels up to 3.7  pg / ml . in addition , patients with preoperative serum il-6 levels over 3.1  pg / ml had lower rfs and os rates ( p  <  0.01 ) . 
 there was no significant correlation between preoperative serum il-6 levels and maximal tumor dimension ( r  =  0.0003 , p  =  0.84).elevated serum levels of il-6 were significantly associated with an increased risk of hbv - associated hcc recurrence suggesting that preoperative il-6 serum level is potential biomarker for early prediction of hbv - associated hcc recurrence . 
INTRODUCTION
METHODS
Study Population
Preoperative Assessment and Surgical Procedures
Postoperative Follow-Up
Statistical Analysis
RESULTS
DISCUSSION