Annals of Diagnostic Pathology 18 (2014) 326–328
Contents lists available at ScienceDirect
Annals of Diagnostic Pathology
8-Hydroxy-2′-deoxyguanosine expression predicts outcome of esophageal cancer Hongtao He, MD a, Yufei Zhao, MD a, Na Wang, MD a, Lan Zhang, MD a, Cuiju Wang, MD, PhD b,⁎ a b
Department of Gastroenterology and Hepatology, Shijiazhuang, PR China Department of Gynecology Ultrasound, Shijiazhuang, PR China
a r t i c l e
i n f o
Keywords: 8-OHdG Esophageal cancer Outcome Survival
a b s t r a c t Esophageal cancer is characterized by increased oxidative stress and the production of 8-hydroxy-2'deoxyguanosine (8-OHdG), which is one of the main mutagenic modifications of DNA. We analyzed the predictive value of 8-OHdG expression on postoperative survival of patients with esophageal cancer with univariate and multivariate analysis. The high levels of 8-OHdG are associated with significantly shorter survival time by log-rank test using Kaplan-Meier methods. Moreover, the level of 8-OHdG expression was identified as an independent predictor for esophageal cancer outcome using Cox proportional hazards model analysis (relative risk, 0.294; 95% confidence interval, 0.178-0.487; P = .000). These results suggest that oxidative damage marker of 8-OHdG is a useful prognostic marker in esophageal cancer. The analysis of 8-OHdG levels can help in the identification of patient subgroups that are at high risk for poor disease outcomes © 2014 Elsevier Inc. All rights reserved.
1. Introduction Reactive oxygen species is formed as byproducts of energy production in aerobic animals causing undesirable cellular damage to macromolecules such as DNA, proteins, and lipids and is thus involved in a variety of biological processes, such as aging, cancer, and degenerative diseases [1,2]. Oxidative damage to the base 2′-deoxyguanosine produces 8-hydroxy-2′-deoxyguanosine (8-OHdG), which could induce G-C to T-A transversion in daughter DNA strands. 8-OHdG is a useful marker for assessing oxidative DNA damage and has been a feature of carcinogenesis in several experiments [3,4]. Analysis of 8-OHdG levels in animal organ DNA is useful for assessment of their carcinogenesis risk [5]. Esophageal cancer is one of the most common cancers in the population of northern central China, with an age-standardized annual incidence rate more than 125 per 100 000 [6]. Cumulative mortality attributed to esophageal cancer is approximately 20% for women and 25% for men [7]. The prognosis for esophageal cancer remains poor, despite improved diagnosis and therapeutic strategies, mostly because of its aggressive nature. Performance status, TNM stage, and lymph node metastases seem to be predictive factors for esophageal cancer outcome [8]. Oxidative stress is also well recognized in the pathogenesis of esophageal cancer, and increase in 8-OHdG levels appears to be closely related to esophageal carcinogenesis [9,10]. In this study, we examined the correlation between 8-OHdG expression and the clinical ⁎ Corresponding author. Department of Gynecology Ultrasound, The Fourth Hospital of Hebei Medical University, 12 Jiankang Road, Shijiazhuang 050011, PR China. Tel.: +86 311 86095342; fax: +86 311 8383 2916. E-mail address: [email protected] (C. Wang). http://dx.doi.org/10.1016/j.anndiagpath.2014.08.008 1092-9134/© 2014 Elsevier Inc. All rights reserved.
outcome of esophageal cancer. Postoperative patients with esophageal cancer enrolled in this study were followed up with regular visits to our hospital, and overall survival rates were recorded. 2. Materials and methods 2.1. Tissue specimen collection We obtained histologically confirmed esophageal cancer tissues, according to the guidelines of the human tissue research committee of the hospital, from 150 patients with esophageal cancer who received esophageal cancer resection operations between January 2002 and June 2006 at the Thoracic Surgery Department of the Fourth Hospital of Hebei Medical University. Tissues were fixed in formalin (10%) immediately after resection, dehydrated in absolute ethanol, embedded in paraffin and serial sections (5 μm thick), and prepared for immunohistochemical analysis. All patients provided written informed consent for the collection of samples and subsequent analysis. 2.2. Measurements of 8-OHdG levels in esophageal cancerous tissues The 8-OHdG levels were measured by esophageal immunohistochemical staining. Briefly, tissue sections were incubated for 1 day at 4°C with anti–8-OHdG antibody (dilution 1:100; Abcam, Cambridge, UK) and then reacted with biotinylated secondary anti-mouse immunoglobulin G antibody for 1 hour at room temperature. Streptavidin was added, and the color was developed with 3,3'-diaminobenzidine. Levels of 8-OHdG were measured as described previously [11,12]; the 8-OHdG–stained esophageal epithelial cells in 10 different random fields at a magnification of ×400 were counted independently by 2
H. He et al. / Annals of Diagnostic Pathology 18 (2014) 326–328
327
statistically significant, and all statistical calculations were performed with SPSS18.0 software package (SPSS Inc, Chicago, IL, USA).
3. Results 3.1. Clinical characteristics of esophageal cancer patients A total of 150 patients with esophageal cancer were enrolled in our study. A review of the patients was performed every 3 months for 5 years. Six patients with squamous cell carcinoma were lost during follow-up: 2 in the second year, 1 in the third year, and 3 in the fourth year. The remaining 144 patients, including 130 with squamous cell carcinoma, 6 with adenocarcinoma, 7 with small cell carcinoma, and 1 with undifferentiated carcinoma, were assessed. Adjuvant chemotherapy and radiotherapy were not administered before or after esophageal cancer resection. The relationships between the clinical characteristics and postoperative survival data were analyzed by log-rank test using the Kaplan-Meier method. Sex, age, tumor size, and tumor histology were not associated with overall survival of esophageal cancer patients, whereas clinical stage showed an association with a 5-year survival rate at significantly statistical levels (Table 1).
3.2. 8-OHdG predicts prognosis of patients with esophageal cancer
Fig. 1. 8-OHdG immunostaining in esophageal tissues with low expression (A) and high expression (B). Cells with a brown-stained nucleus are regarded as positive. Original magnification, ×400.
pathologists. The percentage of positive cells versus total cells that we defined as 8-OHdG label index (LI) in each field was calculated and graded with semiquantitative methods as follows: grade “low” is defined as the percentage of positive cells staining no more than 50%, and grade “high” as the percentage greater than 50% (Fig. 1). 2.3. Statistical analysis
The immunoreactivity of 8-OHdG is observed in esophageal cancer tissue mostly deposited in nuclei (Fig. 1). We graded 8-OHdG with low and high based on percentage of 8-OHdG deposited esophageal epitelial cells. The different grades of LI are shown in Fig. 1, and the 8OHdG levels are measured with this semiquantitative method for further analysis with the prognosis for patients with esophageal cancer. We divided these patients to high 8-OHdG and low 8-OHdG groups based on their LI value for 5-year survival analysis, and cumulative survival was plotted as Kaplan-Meier curve, as shown in Fig. 2. Dramatically significant difference appeared between the low and high groups, with low 8-OHdG patients associating to long survival time by log-rank test (P = .000). These data demonstrate strong power for this oxidative DNA damage marker to predict prognosis of patients with esophageal cancer.
All the survival data were calculated by the Kaplan-Meier method, the difference was determined by the log-rank test, and the prognostic significance of clinical characteristic was assessed by the multivariate Cox proportional hazard model. P values less than .05 were considered Table 1 Analysis of clinical characteristics associated with overall survival of esophageal cancer Characteristics Sex Male Female Age, y ≤55 N55 Size of the tumor (diameter in cm) ≤5 N5 Clinical stages 0 I II III Pathologic classification Squamous cell carcinoma Adenocarcinoma Small cell carcinoma Undifferentiated carcinoma
No. of cases
5-y survival rate (%)
P .242
101 43
45.5 53.5
60 84
50.0 46.4
92 52
51.1 42.3
1 10 86 47
100.0 60.0 58.1 25.5
130 6 7 1
50.0 16.7 42.9 0.0
.709
.339
.000
.410
Fig. 2. Survival curve according to low and high 8-OHdG groups in patients with esophageal cancer.
328
H. He et al. / Annals of Diagnostic Pathology 18 (2014) 326–328
Table 2 Multivariate analysis of prognostic factors associated with overall survival of esophageal cancer patients with Cox proportional hazards model Factors
Relative risk
95% CI
p
8-OHdG expression status Clinical stages
0.294 2.145
0.178-0.487 1.412-3.258
0.000 0.000
analyze their relationship with survival rate. In conclusion, we report here that 8-OHdG is a novel strong prognosis factor in esophageal cancer. Further studies are also required to clarify whether extensive oxidative stress in highly malignant tumors is a prerequisite for carcinoma progression or secondary changes during the carcinogenesis. Conflict of interest
3.3. Multivariate analysis of prognosis factors for esophageal cancer patients
None declared. Acknowledgments
To evaluate the affect of this novel oxidative DNA marker on the predictive power of traditional clinical parameter, we performed multivariate analysis including predictors above with Cox proportional hazard method. The analysis results showed that 8-OHdG and clinical stage were independent factors for the postoperative survival of esophageal cancer patients (Table 2). 8-OHdG shows the strongest link to esophageal cancer survival with the hazard ratio of 0.294 (95% confidence interval [CI], 0.178-0.487; P = .000). Univariate and multivariate analyses suggested that 8-OHdG level in esophageal cancer tissue was one of the best predictor for postoperative survival of patients with esophageal cancer. 4. Discussion In this study, we used immunohistochemical approaches using a polyclonal antibody against 8-OHdG in formalin-fixed, paraffinembedded esophageal sections for assessment of oxidatively damaged DNA. 8-OHdG generated as a result of oxidative DNA damage could induce mutations in genes thereby being closely involved in carcinogenesis of esophageal cancer [9,10]. In this study, we also found that 8-OHdG expressional levels of cancerous tissues were associated significantly with postoperative survival for patients with esophageal cancer. 8-OHdG level is not only a risk factor for carcinogenesis of esophageal cancer but also a good marker for predicting the outcome of postoperative patients with esophageal cancer. Previous studies have suggested that oxidative DNA damage was associated with cancer progression including cellular proliferation, apoptosis, genetic instability, and chemoresistant phenotype [13–16], whereas the functional significance of persistent oxidative stress in esophageal cancerous tissue remains unknown. Miyake et al [17] have proven the association between 8-OHdG expression and tumor size, clinical stage, and venous invasion in renal cell carcinoma progress, and Li et al have also proven the association between 8-OHdG expression and postoperative hepatocellular carcinoma survival [12]. Consistent with previous studies, we found the prognosis value of 8-OHdG for esophageal cancer. Whether oxidative damage is the cause for these clinical characteristics to influence the tumor progress indirectly or just as cooperator of these characteristics that modify the tumor progress directly needs to be further analyzed. We measured 8-OHdG levels using an immunochemical method defined as LI to semiquantify 8-OHdG amount and to subsequently
This work was supported by Key basic research program of Hebei (14967713D). References [1] Ames BN, Shigenaga MK, Hagen TM. Oxidants, antioxidants, and the degenerative diseases of aging. Proc Natl Acad Sci U S A 1993;90:7915–22. [2] Shigenaga M, Gimeno CJ, Ames BN. Urinary 8-hydroxy-2′-deoxyguanosine as a biomarker of in vivo oxidative DNA damage. Proc Natl Acad Sci U S A 1989;86: 9697–701. [3] Halliwell B. Why and how should we measure oxidative damage in nutritional studies? How far could you come? Am J Clin Nutr 2000;72:1082–7. [4] Nakae D, Kobayashi Y, Akai H, Andoh N, Satoh H, Ohashi K, et al. Involvement of 8hydroxyguanine formation in the initiation of rat liver carcinogenesis by low dose levels of N-nitrosodiethylamine. Cancer Res 1997;57:1281–7. [5] Kasai H. Analysis of a form of oxidative DNA damage 8-hydroxy-2′-deoxyguanosine, as a marker of cellular oxidative stress during carcinogenesis. Mutat Res 1997;387: 147–63. [6] Blot WJ, Li JY. Some considerations in the design of a nutrition intervention trial in Linxian, People's Republic of China. Natl Cancer Inst Monogr 1985;69:29–34. [7] Abnet CC, Huppi K, Carrera A. Control region mutations and the ‘common deletion’ are frequent in the mitochondrial DNA of patients with esophageal squamous cell carcinoma. BMC Cancer 2004;4:30. [8] Blanchard P, Quero L, Hennequin C. Prognostic and predictive factor of oesophageal carcinoma. Bull Cancer 2009;96:379–89. [9] Kubo N, Morita M, Nakashima Y, Kitao H, Egashira A, Saeki H, et al. Oxidative DNA damage in human esophageal cancer: clinicopathological analysis of 8hydroxydeoxyguanosine and its repair enzyme. Dis Esophagus 2014;3:285–93. [10] Sehitogullari A, Aslan M, Sayir F, Kahraman A, Demir H. Serum paraoxonase-1 enzyme activities and oxidative stress levels in patients with esophageal squamous cell carcinoma. Redox Rep 2014;19:199–205. [11] Maki A, Kono H, Gupta M, Asakawa M, Suzuki T, Matsuda M, et al. Predictive power of biomarkers of oxidative stress and inflammation in patients with hepatitis C virus-associated hepatocellular carcinoma. Ann Surg Oncol 2007;14:1182–90. [12] Li S, Wang X, Wu Y, Zhang H, Zhang L, Wang C, et al. 8-Hydroxy-2′-deoxyguanosine expression predicts hepatocellular carcinoma outcome. Oncol Lett 2012;3:338–42. [13] Tanaka T, Kondo S, Iwasa Y, Hiai H, Toyokuni S. Expression of stressresponseand cell proliferation genes in renal cell carcinoma inducedby oxidative stress. Am J Pathol 2000;156:2149–57. [14] Inoue S, Kawanishi S. Oxidative DNA damage induced by simultaneous generation of nitric oxide and superoxide. FEBS Lett 1995;371:86–8. [15] Erhola M, Toyokuni S, Okada K, Tanaka T, Hiai H, Ochi H, et al. Biomarker evidenceof DNA oxidation in lung cancer patients: association of urinary 8-hydroxy-2′deoxyguanosine excretion with radiotherapy, chemotherapy, and response to treatment. FEBS Lett 1997;409:287–91. [16] Miyake H, Hara I, Gleave ME, Eto H. Protection of androgen-dependent human prostate cancer cells from oxidative stress-induced DNA damage by overexpression of clusterin and its modulation by androgen. Prostate 2004;61:318–23. [17] Miyake H, Hara I, Kamidono S, Eto H. Prognostic significance of oxidative DNA damage evaluated by 8-hydroxy-2′-deoxyguanosine in patients undergoing radical nephrectomy for renal cell carcinoma. Urology 2004;64:1057–61.
Contents lists available at ScienceDirect
Annals of Diagnostic Pathology
8-Hydroxy-2′-deoxyguanosine expression predicts outcome of esophageal cancer Hongtao He, MD a, Yufei Zhao, MD a, Na Wang, MD a, Lan Zhang, MD a, Cuiju Wang, MD, PhD b,⁎ a b
Department of Gastroenterology and Hepatology, Shijiazhuang, PR China Department of Gynecology Ultrasound, Shijiazhuang, PR China
a r t i c l e
i n f o
Keywords: 8-OHdG Esophageal cancer Outcome Survival
a b s t r a c t Esophageal cancer is characterized by increased oxidative stress and the production of 8-hydroxy-2'deoxyguanosine (8-OHdG), which is one of the main mutagenic modifications of DNA. We analyzed the predictive value of 8-OHdG expression on postoperative survival of patients with esophageal cancer with univariate and multivariate analysis. The high levels of 8-OHdG are associated with significantly shorter survival time by log-rank test using Kaplan-Meier methods. Moreover, the level of 8-OHdG expression was identified as an independent predictor for esophageal cancer outcome using Cox proportional hazards model analysis (relative risk, 0.294; 95% confidence interval, 0.178-0.487; P = .000). These results suggest that oxidative damage marker of 8-OHdG is a useful prognostic marker in esophageal cancer. The analysis of 8-OHdG levels can help in the identification of patient subgroups that are at high risk for poor disease outcomes © 2014 Elsevier Inc. All rights reserved.
1. Introduction Reactive oxygen species is formed as byproducts of energy production in aerobic animals causing undesirable cellular damage to macromolecules such as DNA, proteins, and lipids and is thus involved in a variety of biological processes, such as aging, cancer, and degenerative diseases [1,2]. Oxidative damage to the base 2′-deoxyguanosine produces 8-hydroxy-2′-deoxyguanosine (8-OHdG), which could induce G-C to T-A transversion in daughter DNA strands. 8-OHdG is a useful marker for assessing oxidative DNA damage and has been a feature of carcinogenesis in several experiments [3,4]. Analysis of 8-OHdG levels in animal organ DNA is useful for assessment of their carcinogenesis risk [5]. Esophageal cancer is one of the most common cancers in the population of northern central China, with an age-standardized annual incidence rate more than 125 per 100 000 [6]. Cumulative mortality attributed to esophageal cancer is approximately 20% for women and 25% for men [7]. The prognosis for esophageal cancer remains poor, despite improved diagnosis and therapeutic strategies, mostly because of its aggressive nature. Performance status, TNM stage, and lymph node metastases seem to be predictive factors for esophageal cancer outcome [8]. Oxidative stress is also well recognized in the pathogenesis of esophageal cancer, and increase in 8-OHdG levels appears to be closely related to esophageal carcinogenesis [9,10]. In this study, we examined the correlation between 8-OHdG expression and the clinical ⁎ Corresponding author. Department of Gynecology Ultrasound, The Fourth Hospital of Hebei Medical University, 12 Jiankang Road, Shijiazhuang 050011, PR China. Tel.: +86 311 86095342; fax: +86 311 8383 2916. E-mail address: [email protected] (C. Wang). http://dx.doi.org/10.1016/j.anndiagpath.2014.08.008 1092-9134/© 2014 Elsevier Inc. All rights reserved.
outcome of esophageal cancer. Postoperative patients with esophageal cancer enrolled in this study were followed up with regular visits to our hospital, and overall survival rates were recorded. 2. Materials and methods 2.1. Tissue specimen collection We obtained histologically confirmed esophageal cancer tissues, according to the guidelines of the human tissue research committee of the hospital, from 150 patients with esophageal cancer who received esophageal cancer resection operations between January 2002 and June 2006 at the Thoracic Surgery Department of the Fourth Hospital of Hebei Medical University. Tissues were fixed in formalin (10%) immediately after resection, dehydrated in absolute ethanol, embedded in paraffin and serial sections (5 μm thick), and prepared for immunohistochemical analysis. All patients provided written informed consent for the collection of samples and subsequent analysis. 2.2. Measurements of 8-OHdG levels in esophageal cancerous tissues The 8-OHdG levels were measured by esophageal immunohistochemical staining. Briefly, tissue sections were incubated for 1 day at 4°C with anti–8-OHdG antibody (dilution 1:100; Abcam, Cambridge, UK) and then reacted with biotinylated secondary anti-mouse immunoglobulin G antibody for 1 hour at room temperature. Streptavidin was added, and the color was developed with 3,3'-diaminobenzidine. Levels of 8-OHdG were measured as described previously [11,12]; the 8-OHdG–stained esophageal epithelial cells in 10 different random fields at a magnification of ×400 were counted independently by 2
H. He et al. / Annals of Diagnostic Pathology 18 (2014) 326–328
327
statistically significant, and all statistical calculations were performed with SPSS18.0 software package (SPSS Inc, Chicago, IL, USA).
3. Results 3.1. Clinical characteristics of esophageal cancer patients A total of 150 patients with esophageal cancer were enrolled in our study. A review of the patients was performed every 3 months for 5 years. Six patients with squamous cell carcinoma were lost during follow-up: 2 in the second year, 1 in the third year, and 3 in the fourth year. The remaining 144 patients, including 130 with squamous cell carcinoma, 6 with adenocarcinoma, 7 with small cell carcinoma, and 1 with undifferentiated carcinoma, were assessed. Adjuvant chemotherapy and radiotherapy were not administered before or after esophageal cancer resection. The relationships between the clinical characteristics and postoperative survival data were analyzed by log-rank test using the Kaplan-Meier method. Sex, age, tumor size, and tumor histology were not associated with overall survival of esophageal cancer patients, whereas clinical stage showed an association with a 5-year survival rate at significantly statistical levels (Table 1).
3.2. 8-OHdG predicts prognosis of patients with esophageal cancer
Fig. 1. 8-OHdG immunostaining in esophageal tissues with low expression (A) and high expression (B). Cells with a brown-stained nucleus are regarded as positive. Original magnification, ×400.
pathologists. The percentage of positive cells versus total cells that we defined as 8-OHdG label index (LI) in each field was calculated and graded with semiquantitative methods as follows: grade “low” is defined as the percentage of positive cells staining no more than 50%, and grade “high” as the percentage greater than 50% (Fig. 1). 2.3. Statistical analysis
The immunoreactivity of 8-OHdG is observed in esophageal cancer tissue mostly deposited in nuclei (Fig. 1). We graded 8-OHdG with low and high based on percentage of 8-OHdG deposited esophageal epitelial cells. The different grades of LI are shown in Fig. 1, and the 8OHdG levels are measured with this semiquantitative method for further analysis with the prognosis for patients with esophageal cancer. We divided these patients to high 8-OHdG and low 8-OHdG groups based on their LI value for 5-year survival analysis, and cumulative survival was plotted as Kaplan-Meier curve, as shown in Fig. 2. Dramatically significant difference appeared between the low and high groups, with low 8-OHdG patients associating to long survival time by log-rank test (P = .000). These data demonstrate strong power for this oxidative DNA damage marker to predict prognosis of patients with esophageal cancer.
All the survival data were calculated by the Kaplan-Meier method, the difference was determined by the log-rank test, and the prognostic significance of clinical characteristic was assessed by the multivariate Cox proportional hazard model. P values less than .05 were considered Table 1 Analysis of clinical characteristics associated with overall survival of esophageal cancer Characteristics Sex Male Female Age, y ≤55 N55 Size of the tumor (diameter in cm) ≤5 N5 Clinical stages 0 I II III Pathologic classification Squamous cell carcinoma Adenocarcinoma Small cell carcinoma Undifferentiated carcinoma
No. of cases
5-y survival rate (%)
P .242
101 43
45.5 53.5
60 84
50.0 46.4
92 52
51.1 42.3
1 10 86 47
100.0 60.0 58.1 25.5
130 6 7 1
50.0 16.7 42.9 0.0
.709
.339
.000
.410
Fig. 2. Survival curve according to low and high 8-OHdG groups in patients with esophageal cancer.
328
H. He et al. / Annals of Diagnostic Pathology 18 (2014) 326–328
Table 2 Multivariate analysis of prognostic factors associated with overall survival of esophageal cancer patients with Cox proportional hazards model Factors
Relative risk
95% CI
p
8-OHdG expression status Clinical stages
0.294 2.145
0.178-0.487 1.412-3.258
0.000 0.000
analyze their relationship with survival rate. In conclusion, we report here that 8-OHdG is a novel strong prognosis factor in esophageal cancer. Further studies are also required to clarify whether extensive oxidative stress in highly malignant tumors is a prerequisite for carcinoma progression or secondary changes during the carcinogenesis. Conflict of interest
3.3. Multivariate analysis of prognosis factors for esophageal cancer patients
None declared. Acknowledgments
To evaluate the affect of this novel oxidative DNA marker on the predictive power of traditional clinical parameter, we performed multivariate analysis including predictors above with Cox proportional hazard method. The analysis results showed that 8-OHdG and clinical stage were independent factors for the postoperative survival of esophageal cancer patients (Table 2). 8-OHdG shows the strongest link to esophageal cancer survival with the hazard ratio of 0.294 (95% confidence interval [CI], 0.178-0.487; P = .000). Univariate and multivariate analyses suggested that 8-OHdG level in esophageal cancer tissue was one of the best predictor for postoperative survival of patients with esophageal cancer. 4. Discussion In this study, we used immunohistochemical approaches using a polyclonal antibody against 8-OHdG in formalin-fixed, paraffinembedded esophageal sections for assessment of oxidatively damaged DNA. 8-OHdG generated as a result of oxidative DNA damage could induce mutations in genes thereby being closely involved in carcinogenesis of esophageal cancer [9,10]. In this study, we also found that 8-OHdG expressional levels of cancerous tissues were associated significantly with postoperative survival for patients with esophageal cancer. 8-OHdG level is not only a risk factor for carcinogenesis of esophageal cancer but also a good marker for predicting the outcome of postoperative patients with esophageal cancer. Previous studies have suggested that oxidative DNA damage was associated with cancer progression including cellular proliferation, apoptosis, genetic instability, and chemoresistant phenotype [13–16], whereas the functional significance of persistent oxidative stress in esophageal cancerous tissue remains unknown. Miyake et al [17] have proven the association between 8-OHdG expression and tumor size, clinical stage, and venous invasion in renal cell carcinoma progress, and Li et al have also proven the association between 8-OHdG expression and postoperative hepatocellular carcinoma survival [12]. Consistent with previous studies, we found the prognosis value of 8-OHdG for esophageal cancer. Whether oxidative damage is the cause for these clinical characteristics to influence the tumor progress indirectly or just as cooperator of these characteristics that modify the tumor progress directly needs to be further analyzed. We measured 8-OHdG levels using an immunochemical method defined as LI to semiquantify 8-OHdG amount and to subsequently
This work was supported by Key basic research program of Hebei (14967713D). References [1] Ames BN, Shigenaga MK, Hagen TM. Oxidants, antioxidants, and the degenerative diseases of aging. Proc Natl Acad Sci U S A 1993;90:7915–22. [2] Shigenaga M, Gimeno CJ, Ames BN. Urinary 8-hydroxy-2′-deoxyguanosine as a biomarker of in vivo oxidative DNA damage. Proc Natl Acad Sci U S A 1989;86: 9697–701. [3] Halliwell B. Why and how should we measure oxidative damage in nutritional studies? How far could you come? Am J Clin Nutr 2000;72:1082–7. [4] Nakae D, Kobayashi Y, Akai H, Andoh N, Satoh H, Ohashi K, et al. Involvement of 8hydroxyguanine formation in the initiation of rat liver carcinogenesis by low dose levels of N-nitrosodiethylamine. Cancer Res 1997;57:1281–7. [5] Kasai H. Analysis of a form of oxidative DNA damage 8-hydroxy-2′-deoxyguanosine, as a marker of cellular oxidative stress during carcinogenesis. Mutat Res 1997;387: 147–63. [6] Blot WJ, Li JY. Some considerations in the design of a nutrition intervention trial in Linxian, People's Republic of China. Natl Cancer Inst Monogr 1985;69:29–34. [7] Abnet CC, Huppi K, Carrera A. Control region mutations and the ‘common deletion’ are frequent in the mitochondrial DNA of patients with esophageal squamous cell carcinoma. BMC Cancer 2004;4:30. [8] Blanchard P, Quero L, Hennequin C. Prognostic and predictive factor of oesophageal carcinoma. Bull Cancer 2009;96:379–89. [9] Kubo N, Morita M, Nakashima Y, Kitao H, Egashira A, Saeki H, et al. Oxidative DNA damage in human esophageal cancer: clinicopathological analysis of 8hydroxydeoxyguanosine and its repair enzyme. Dis Esophagus 2014;3:285–93. [10] Sehitogullari A, Aslan M, Sayir F, Kahraman A, Demir H. Serum paraoxonase-1 enzyme activities and oxidative stress levels in patients with esophageal squamous cell carcinoma. Redox Rep 2014;19:199–205. [11] Maki A, Kono H, Gupta M, Asakawa M, Suzuki T, Matsuda M, et al. Predictive power of biomarkers of oxidative stress and inflammation in patients with hepatitis C virus-associated hepatocellular carcinoma. Ann Surg Oncol 2007;14:1182–90. [12] Li S, Wang X, Wu Y, Zhang H, Zhang L, Wang C, et al. 8-Hydroxy-2′-deoxyguanosine expression predicts hepatocellular carcinoma outcome. Oncol Lett 2012;3:338–42. [13] Tanaka T, Kondo S, Iwasa Y, Hiai H, Toyokuni S. Expression of stressresponseand cell proliferation genes in renal cell carcinoma inducedby oxidative stress. Am J Pathol 2000;156:2149–57. [14] Inoue S, Kawanishi S. Oxidative DNA damage induced by simultaneous generation of nitric oxide and superoxide. FEBS Lett 1995;371:86–8. [15] Erhola M, Toyokuni S, Okada K, Tanaka T, Hiai H, Ochi H, et al. Biomarker evidenceof DNA oxidation in lung cancer patients: association of urinary 8-hydroxy-2′deoxyguanosine excretion with radiotherapy, chemotherapy, and response to treatment. FEBS Lett 1997;409:287–91. [16] Miyake H, Hara I, Gleave ME, Eto H. Protection of androgen-dependent human prostate cancer cells from oxidative stress-induced DNA damage by overexpression of clusterin and its modulation by androgen. Prostate 2004;61:318–23. [17] Miyake H, Hara I, Kamidono S, Eto H. Prognostic significance of oxidative DNA damage evaluated by 8-hydroxy-2′-deoxyguanosine in patients undergoing radical nephrectomy for renal cell carcinoma. Urology 2004;64:1057–61.
