worldwide , cervical cancer is the second most common malignant tumor in women and is one of the leading causes of cancer - related death in developing countries . despite the general downward trend in incidence of cervical cancer ,
it is still a serious public health problem worldwide , especially in developing countries ; in addition , cervical cancer is occurring in younger patients . during the past decades
, many distinct advances have been made in the prevention , surgical resection , radiotherapy , and chemotherapy of cervical cancer , but the prognosis of cervical cancer patients remains poor .
therefore , it is important that we better understand the molecular events involved in the invasion and metastasis of cervical cancer , and develop novel prognostic markers and therapeutic strategies for cervical cancer .
the infection of human papilloma virus ( hpv ) is considered the major cause of cervical cancer , but viral infection alone is not sufficient for the development of cervical cancer .
emerging evidence indicates that the chronic inflammatory response is significantly correlated with various types of cancers .
interleukin 1(il-1 ) , as a pleiotropic pro - inflammatory cytokine , plays an important role in tumor growth and metastasis .
two of them are active , il-1 and il-1 , and the third one is an antagonist , il-1ra . il-1 and il-1 are coded by different genes , but their biological functions are identical .
interleukin 6 ( il-6 ) , a 23- to 30-kda pleiotropic cytokine , is produced by many types of cells like fibroblasts , immune cells , and some tumor cells .
increasing evidence reveals that il-1 and il-6 play crucial roles in cancer initiation , development , and metastasis .
however , to the best of our knowledge , the final role of il-1 and il-6 in cervical cancer remains unclear . in the present study , we used tissue microarray and immunohistochemistry methods to detect the expression of il-1 and il-6 in 105 case - samples of human cervical cancer tissues and their paired adjacent tissues .
the purpose of this study was to investigate the association of il-1 and il-6 expression with clinicopathological parameters of cervical cancer and evaluate the prognostic value of il-1 and il-6 expression in cervical cancer patients .
this study included a total of 105 formalin - fixed , paraffin - embedded cervical cancer tissues and their adjacent non - tumor tissues taken from patients at the time of resection surgery , from january 2002 to november 2013 at east hospital , tongji university .
all cancers were confirmed as cervical cancer by the medical examination of hematoxylin and eosin after surgical resection .
none of patients had received adjuvant chemotherapy , radiation therapy , or other anti - tumor therapies .
important related clinicopathological parameters of the patients , such as age , tumor size , figo stage , lymphatic metastasis , stromal invasion , differentiation , and survival time were obtained from each patient s medical records and are shown in tables 1 and 2 .
the survival time was calculated from the date of surgery to the date of death , or the last known follow - up .
all cervical cancer tissue samples include in this investigation were obtained with patients written informed consent .
a standard immunohistochemistry method was used to inspect il-1 and il-6 expression in cervical cancer tissues and their adjacent tissues . briefly , the tumor tissues and adjacent tissues were fixed in 10% formaldehyde and embedded in paraffin ; then cut into 4-m sections .
all 4-m tissue sections are dewaxed and rehydrated with xylene and graded alcohol , respectively .
we washed the sections with buffer solution for 5 minutes and then added the primary antibody at 4c overnight .
afterwards , we washed the sections with phosphate buffered saline ( pbs ) and stained them with 3 , 3-diaminobenzidine ( dab ) for 5 minutes , then counterstained with hematoxylin .
finally , all the sections were assessed under an optical microscope by two independent investigators ; any discrepancy in immunohistochemistry was resolved by consensus .
the expression of il-1 and il-6 in cervical cancer and adjacent tissues was compared with paired wilcoxon test . chi - square test and fisher s exact test were used to assess the association between clinical characteristics of cervical cancer patients and il-1 and il-6 expression .
the prognostic indicators of cervical cancer and il-1 and il-6 expression were determined using kaplan - meier survival analysis and log - rank test for univariate analysis ; the significant variables resulting from univariate tests were included in the cox multivariate regression analysis . a p value of < 0.05 was considered statistically significant for all tests .
this study included a total of 105 formalin - fixed , paraffin - embedded cervical cancer tissues and their adjacent non - tumor tissues taken from patients at the time of resection surgery , from january 2002 to november 2013 at east hospital , tongji university .
all cancers were confirmed as cervical cancer by the medical examination of hematoxylin and eosin after surgical resection .
none of patients had received adjuvant chemotherapy , radiation therapy , or other anti - tumor therapies .
important related clinicopathological parameters of the patients , such as age , tumor size , figo stage , lymphatic metastasis , stromal invasion , differentiation , and survival time were obtained from each patient s medical records and are shown in tables 1 and 2 .
the survival time was calculated from the date of surgery to the date of death , or the last known follow - up .
all cervical cancer tissue samples include in this investigation were obtained with patients written informed consent .
a standard immunohistochemistry method was used to inspect il-1 and il-6 expression in cervical cancer tissues and their adjacent tissues .
briefly , the tumor tissues and adjacent tissues were fixed in 10% formaldehyde and embedded in paraffin ; then cut into 4-m sections .
all 4-m tissue sections are dewaxed and rehydrated with xylene and graded alcohol , respectively .
we washed the sections with buffer solution for 5 minutes and then added the primary antibody at 4c overnight .
afterwards , we washed the sections with phosphate buffered saline ( pbs ) and stained them with 3 , 3-diaminobenzidine ( dab ) for 5 minutes , then counterstained with hematoxylin .
finally , all the sections were assessed under an optical microscope by two independent investigators ; any discrepancy in immunohistochemistry was resolved by consensus .
the expression of il-1 and il-6 in cervical cancer and adjacent tissues was compared with paired wilcoxon test .
chi - square test and fisher s exact test were used to assess the association between clinical characteristics of cervical cancer patients and il-1 and il-6 expression .
the prognostic indicators of cervical cancer and il-1 and il-6 expression were determined using kaplan - meier survival analysis and log - rank test for univariate analysis ; the significant variables resulting from univariate tests were included in the cox multivariate regression analysis
typical immunohistochemistry images of il-1 and il-6 in cervical cancer tissues and paired adjacent non - tumor tissues are shown in figures 1 and 2 .
the positive percentages of il-1 expression in cervical cancer and adjacent non - tumor tissues were 63.8% ( 67/105 ) and 8.6% ( 9/105 ) , respectively . the positive percentages of il-6 expression in cervical cancer and adjacent non - tumor tissues were 60.0% ( 63/105 ) and 10.5% ( 11/105 ) , respectively .
the chi - square test was used to confirm that the difference in the expression level of il-1 and il-6 between cervical cancer tissues and paired adjacent non - tumor tissues was statistically significant ( p<0.05 ) .
the relationship between a patient s clinical parameters and the expressions of il-1 and il-6 are shown in tables 3 and 4 .
these results showed that the expression of il-1 in cervical cancer tissues was significantly correlated with tumor size ( =6.024 , p=0.014 ) , figo histology grade ( =19.661 , p<0.0001 ) , lymph node metastasis ( =5.135 , p=0.023 ) , stromal invasion ( =5.399 , p=0.020 ) , and tumor differentiation ( =7.513 , p=0.006 ) .
however , no statistical correlation was found between il-1 expression and patient age ( p>0.05 ) .
similarly , the expression of il-6 in cervical cancer tissues was significantly correlated with tumor size ( =5.695 , p=0.017 ) , figo histology grade ( =10.239 , p=0.001 ) , and tumor differentiation ( =5.210 , p=0.022 ) .
however , there was no statistical correlation found between il-6 expression and patient age , lymph node metastasis , or stromal invasion ( p>0.05 ) .
therefore , these results demonstrated that higher il-1 and il-6 expression in cervical cancer tissues was positively correlated with tumor metastasis and cancer progression , suggesting that il-1 and il-6 play important roles in tumor progression . in order to further evaluate the relationship between il-1 and il-6 expression and prognosis of cervical cancer , we performed log - rank survival analysis according to il-1 and il-6 expression level and patient survival data .
the survival analysis demonstrated that the cervical cancer survival rate of patients with negative il-1 or il-6 expression was significantly better than that of patients with positive expression ( p<0.05 , figures 3 , 4 ) .
furthermore , a multivariate cox regression analysis demonstrated that il-1 expression and lymph node metastasis were independent predictors of overall survival in cervical cancer patients .
typical immunohistochemistry images of il-1 and il-6 in cervical cancer tissues and paired adjacent non - tumor tissues are shown in figures 1 and 2 .
the positive percentages of il-1 expression in cervical cancer and adjacent non - tumor tissues were 63.8% ( 67/105 ) and 8.6% ( 9/105 ) , respectively . the positive percentages of il-6 expression in cervical cancer and adjacent non - tumor tissues were 60.0% ( 63/105 ) and 10.5% ( 11/105 ) , respectively .
the chi - square test was used to confirm that the difference in the expression level of il-1 and il-6 between cervical cancer tissues and paired adjacent non - tumor tissues was statistically significant ( p<0.05 ) .
the relationship between a patient s clinical parameters and the expressions of il-1 and il-6 are shown in tables 3 and 4 .
these results showed that the expression of il-1 in cervical cancer tissues was significantly correlated with tumor size ( =6.024 , p=0.014 ) , figo histology grade ( =19.661 , p<0.0001 ) , lymph node metastasis ( =5.135 , p=0.023 ) , stromal invasion ( =5.399 , p=0.020 ) , and tumor differentiation ( =7.513 , p=0.006 ) .
however , no statistical correlation was found between il-1 expression and patient age ( p>0.05 ) .
similarly , the expression of il-6 in cervical cancer tissues was significantly correlated with tumor size ( =5.695 , p=0.017 ) , figo histology grade ( =10.239 , p=0.001 ) , and tumor differentiation ( =5.210 , p=0.022 ) .
however , there was no statistical correlation found between il-6 expression and patient age , lymph node metastasis , or stromal invasion ( p>0.05 ) .
therefore , these results demonstrated that higher il-1 and il-6 expression in cervical cancer tissues was positively correlated with tumor metastasis and cancer progression , suggesting that il-1 and il-6 play important roles in tumor progression .
in order to further evaluate the relationship between il-1 and il-6 expression and prognosis of cervical cancer , we performed log - rank survival analysis according to il-1 and il-6 expression level and patient survival data .
the survival analysis demonstrated that the cervical cancer survival rate of patients with negative il-1 or il-6 expression was significantly better than that of patients with positive expression ( p<0.05 , figures 3 , 4 ) .
furthermore , a multivariate cox regression analysis demonstrated that il-1 expression and lymph node metastasis were independent predictors of overall survival in cervical cancer patients .
in the past decade , various studies have provided substantial evidence to support the role of inflammation and inflammation - related pathways in the pathogenesis of numerous human cancers , including cervical cancer [ 810 ] .
it has been shown that the inflammatory microenvironment consists of many important components such as tumor cells , stromal cells , and immune and inflammatory cells .
all of these components interact intimately and produce chemokines , growth factors , and adhesion molecules , and further promote the initiation and progression of many cancers .
il-1 , a key inflammatory signaling cytokine , is secreted by various types of cells such as monocytes , macrophages , neutrophils , smooth muscle cells , fibroblasts , and cervical epithelium .
increasing evidence suggests that il-1 within the cancer microenvironment can induce the expression of growth factors , such as il-6 , il-8 , tumor necrosis factor- ( tnf- ) , vascular endothelial growth factor ( vegf ) ; metastatic genes such as the matrix metalloproteinases ( mmps ) ; transforming growth factor- ( tgf ) , and stimulate the production of angiogenic proteins [ 1416 ] .
il-1 plays an important role in the development , invasion , and metastasis of cancer .
previous studies have found elevated expression of il-1 in a variety of cancers such as breast cancer , pancreatic cancer , and head and neck cancer .
moreover , the enhanced expression of il-1 has been correlated with poorer prognosis of cancer .
il-6 , a 23- to 30-kda multifunctional cytokine , is produced by numerous kinds of immune and non - immune cells , including fibroblasts , immune cells , and some cancer cells .
emerging evidence revealed that il-6 is a master regulator of cell proliferation , immune defense , and hematopoiesis , and plays a crucial role in the development and progression of various cancers [ 2224 ] .
il-6 can regulate cancer cell survival , proliferation , and metastasis by activating jak - stat3 signaling pathway and the ras - mapk signaling pathway .
the activation of stat3 has been shown to maintain nf-b activity and enhance the expression of angiogenesis - related genes such as vegf . therefore , angiogenesis oncology signaling pathways within the inflammatory microenvironment maybe a crucial mechanism of il-6 induced cervical cancer .
furthermore , il-6 can promote b cell differentiation and proliferation and then induce antibody secretion in b cells .
moreover , il-6 can enhance , directly or indirectly , nk cell cytotoxicity and t cell tumoricidal activity .
the results of our study demonstrated that the expression of il-1 and il-6 was correlated with invasion and progression of cervical cancer .
meanwhile , the cervical cancer survival rate of patients with negative il-1 or il-6 expression was significantly better than that of patients with positive expression . il-1 and
il-6 are implicated as important to the cytokine network in the development of cervical cancer through several complicated mechanisms that involve cell proliferation , apoptosis , angiogenesis , inflammatory microenvironment , and numerous complex signaling pathways .
it is well known that cancer pathogenesis is a multi - factor , multi - step , complicated process that involves gene - gene and gene - environment interactions .
large and well - designed studies are still needed to elucidate the pathogenesis of cervical cancer .
our study showed that the expression of il-1 and il-6 was significantly elevated in cervical cancer tissues .
kaplan - meier and cox regression analysis demonstrated that il-1 and il-6 was associated with cervical cancer progression and metastasis and they might act as important molecular markers for prognosis , and be potential therapeutic targets for cervical cancer .
 backgroundil-1 and il-6 are associated with the prognosis of a wide range of cancers , but their value in cervical cancer remains controversial . 
 the aim of this study was to investigate the expression of il-1 and il-6 in cervical cancer and their significance in clinical prognosis.material/methodsthe expression of il-1 and il-6 in 105 formalin - fixed , paraffin - embedded cervical cancer tissues and adjacent non - tumor tissues was examined by immunohistochemistry . 
 the results were semi - quantitatively scored and analyzed by chi - square test . 
 patient overall survival ( os ) data was collected by follow - up and analyzed by kaplan - meier analysis.resultsthe expression level of both il-1 and il-6 in cervical cancer tissue was higher than in adjacent non - tumor tissues ( p<0.05 ) . 
 il-1 expression was shown to be correlated with tumor size , figo histology grade , lymph node metastasis , stromal invasion , and tumor differentiation ( p<0.05 ) . 
 il-6 expression was shown to be correlated with tumor size , figo histology grade , and tumor differentiation ( p<0.05 ) . 
 patients with positive expression of il-1 or il-6 tended to have much shorter survival times than patients with negative expression . 
 in addition , a multivariate cox regression analysis demonstrated that il-1 expression and lymph node metastasis were independent predictors of os in cervical cancer patients.conclusionsthe expression of il-1 was significantly associated with tumor size , figo histology grade , lymph node metastasis , stromal invasion , and tumor differentiation . 
 the expression of il-6 was significantly associated with tumor size , figo histology grade , and tumor differentiation . 
 positive il-1 and il-6 expression was significantly correlated with poor prognosis . 
 they may be considered valuable biomarkers for prognosis and potential therapeutic targets for cervical cancer . 
Background
Material and Methods
Patients and tissues specimens
Immunohistochemical staining
Statistical analysis
Results
Expression of IL-1 and IL-6 significantly increased in cervical cancer tissues
Increased expression of IL-1 and IL-6 correlates with clinicopathological parameters of patients with cervical cancer
Correlation between IL-1 and IL-6 and prognosis for cervical cancer patients
Discussion
Conclusions