The Infiltration Of Tumor-associated Macrophages And Its Impacts On Patients' Prognosis In Esophageal Cancer

Background and ObjectivesThere is a large quantity of tumor infiltration of macrophages in malignant tumors, known as tumor associated macrophages (TAMs). Previous studies suggest that TAMs have the important function of anti-tumor immune regulation and can kill tumor cells directly, or through the presentation of tumor-associated antigens to induce immune responses to clear tumor cells. The early studies have found that tumor infiltrated TAMs release a variety of cytokines, growth factors and chemokines to participate in substrates reconstruction, the generation of tumor microvessels and lymphatic vessels, and a variety of mechanisms such as immune suppression and escape to promote tumor development, invasion and metastasis, so they are closely related to survival. With further research, scholars have recognized that different TAMs show activation of different nature when stimulated under different tumor microenvironment, including 2 types:(1) classical activated macrophages, also known as Ml macrophages, which can inhibit and kill resistance of tumor cells; (2) alternative activated macrophages, also known as M2 macrophages, which mainly promote tumor development. Previous studies suggested that the macrophages marked as CD68 in tumor tissues are TAMs. However, in recent years, studies have shown that TAMs are more likely to be M2 macrophages marked as CD 163, which can promote the proliferation, survival, invasion and metastasis of tumor cells, therefore promoting tumor development and affecting the survival time and prognosis of patients.Firstly this study uses non-biotin immunohistochemical staining method to detect the infiltration density of CD68- and CD 163-positive macrophages in 117 cases of esophageal carcinoma tissue and 30 cases of adjacent normal tissue. Then with the clinicopathologic features and follow-up data, it explores the relationship between the infiltration density of CD68- and CD163-positive macrophages under esophageal carcinoma tissue microenvironment and the lymph node metastasis and the survival time of esophageal cancer patients, and finally analyzes the influence of the infiltration density under different micro-environment on the prognosis of patients.Materials and methodsThis study collects 117 esophageal carcinoma tissue samples surgically removed and pathologically diagnosed with complete clinical and follow-up data in Henan Tumor Hospital and the First Affiliated Hospital of Zhengzhou University from January 2004 to May 2005, and all the patients did not receive any treatment before surgery, and at the same time collected 30 adjacent normal tissue samples as comparison. Then it uses two-step non-biotin immunohistochemical staining method to detect the infiltration of CD68- and CD163-positive macrophages in carcinoma tissue and adjacent normal tissue. SPSS16.0 software package is used for statistical analysis and processing. Then it tests the progressiveness and homogeneity of variance of the infiltration density of CD68- and CD163-positive macrophages in esophageal carcinoma tissue and adjacent normal tissue and it carries out independent sample T tests further. Spearman rank correlation test is carried out for the relationship between the density of macrophage infiltration and lymph node metastases and survival time. Finally it uses Kaplan-Meier method to draw the survival curve, Cox regression model for multivariate survival analysis, andχ2 test to check the relationship between the pathologic parameters. The difference limited to P<0.05 is considered statistically significant.Results1. The comparison between the density of tissue infiltration of CD68- and CD163-positive macrophages in adjacent normal tissue and that in esophageal carcinoma tissue.The mean of infiltration density of CD68- and CD163- positive macrophages in adjacent normal tissue are respectively (3.9±2.6)/HP vs (2.3±1.6)/HP; those in Esophageal carcinoma tissue are (36.2±15.5)/HP vs (28.5±12.8)/HP. Compared with the density in adjacent normal tissue the density in esophageal carcinoma tissue increases significantly(P=0.000; P=0.000), the difference was statistically significant. Esophageal carcinoma tissue CD68-positive macrophage infiltration density is higher than that of CD163-positive macrophages, and the pair correlation test shows the two are closely related (P=0.000), the difference was statistically significant.2. The the relationship between the infiltration density of CD68-and CD163-positive macrophages under different microenvironment in esophageal carcinoma tissue, and the metastasis of lymph nodes and survival time, and that of univariate survival alanalysis.2.1 The the relationship between the infiltration density of CD68-and CD 163-positive macrophages in esophageal esophageal carcinoma (cancer nest+ cancer stroma) and the metastasis of lymph nodes and survival time, and that of univariate survival alanalysis.The mean of infiltration density of CD68-positive macrophage in esophageal carcinoma tissue(cancer nest+cancer stroma) is (36.22±15.50)/HP. The infiltration density of CD68- positive macrophages is positively correlated with the lymph node metastasis (P=0.000); negative correlation with survival time (P=0.002),and the difference were statistically significant. The 5-year survival rate of the high density group is obviously lower than that of low density group, which are respectively 30.5% vs 56.9% (P=0.005), the difference was statistically significant. The mean of infiltration density of CD163-positive macrophages in esophageal carcinoma tissue (cancer nest+cancer stroma) is (28.50±12.80)/HP. The infiltration density of CD163- positive macrophages is positively correlated with the lymph node metastasis (P=0.000); it is negatively correlated with the survival time (P=0.000) and the difference were statistically significant. The 5-year survival rate of the high density group is obviously lower than that of low density group, which are respectively 27.1% vs 60.3% (P=0.000), and the difference is statistically significant.2.2 The the relationship between the infiltration density of CD68- and CD163-positive macrophages in esophageal cancer stroma and the metastasis of lymph nodes and survival time, and that of univariate survival analysis.The mean of infiltration density of CD68-positive macrophages in esophageal cancer stroma is (29.26±13.47)/HP. The infiltration density of CD68-positive macrophages in esophageal cancer stroma is positively correlated with the lymph node metastasis(P=0.000); it is negatively correlated with the survival time (P=0.000) and the difference were statistically significant. The 5-year survival rate of the high density group is obviously lower than that of low density group, which are respectively 27.1% vs 60.3% (P=0.000), and the difference was statistically significant.The mean of infiltration density of CD163-positive macrophages in esophageal cancer stroma is (23.09.±0.92)/HP. The infiltration density of CD163-positive macrophages is positively correlated with the lymph node metastasis (P=0.000); it is negatively correlated with the survival time (P=0.001) and the difference were statistically significant. The 5-year survival rate of the high density group is obviously lower than that of low density group, which are respectively 33.9% vs 53.4% (P=0.032), and the difference was statistically significant.2.3 The the relationship between the infiltration density of CD68-and CD163-positive macrophages in esophageal cancer nest and the metastasis of lymph nodes and survival time, and that of univariate survival analysis.The mean of infiltration density of CD68-positive macrophages in esophageal cancer nest is (6.85±4.42)/HP. The infiltration density of CD68-positive macrophages in esophageal cancer nest has no correlation with the lymph node metastasis (P=0.101); it has no correlation with the survival time (P=0.081) and the difference has no statistic significance. The 5-year survival rate of the high density group is obviously higher than that of low density group, which are respectively 64.4% vs 22.4% (P=0.000), and the difference was statistically significant.The mean of infiltration density of CD163-positive macrophages in esophageal cancer nest is (5.26±0.47)/HP. The infiltration density of CD163-positive macrophages is positively correlated with the lymph node metastasis (P=0.000); it is negatively correlated with the survival time (P=0.000) and the difference were statistically significant. The 5-year survival rate of the high density group is obviously lower than that of low density group, which are respectively 30.5% vs 56.9% (P=0.002), and the difference was statistically significant.3. Cox regression model for multivariate survival analysis in esophageal carcinoma prognosis.Cox regression model for multivariate survival analysis shows that the independent prognostic factors are the infiltration density of CD163-positive lymph node metastasis macrophages in esophageal cancer stroma and cancer nest (P=0.023; P=0.000), the infiltration density of CD68- positive macrophages in cancer nest (P=0.000) and the lymph node metastases(P=0.000), and the difference was statistically significant.Conclusion1.The infiltration density of CD68- and CD163-positive macrophages in esophageal cancer is obviously higher than that of adjacent normal tissues.2. The infiltration of CD68-positive macrophages under different microenvironment in esophageal cancer may have bidirectional influences on tumors. In the cancer nest it has anti-tumor effect, and in the cancer stroma it primarily facilitates tumor progression.3. The infiltration of CD163-positive macrophages under different microenvironment in esophageal cancer promotes lymph node metastasis and shortens the survival time, which is a bad prognostic factor.4. The infiltration of CD163-positive macrophages in esophageal cancer has good independent prognostic value.