| Purpose:Monocytes can differentiate into classically activated macrophages (M1) and alternatively activated macrophages (M2) under different stimulation when recruited into peripheral tissues from the circulation. Tumor associated macrophages (TAM) resemble M2-polarized macrophages. High TAM density predicts poor prognosis in patients with primary hepatocellular carcinoma after resection. High intratumoral cytotoxic T lymphocytes (CTL) infiltration was an independent prognostic factor for improved overall survival and disease-free survival in primary liver cancer. To investigate the influence of TAM on CTL and the role of TAM in tumorigenesis and tumor development, we designed our experiment.Methods:Walker-256 Wistar rat hepatoma models were finished. Then rats were randomly divided into three groups:A group (clodronate-lipsomes by intraperitoneal injection), B group (PBS-lipsomes by intraperitoneal injection), and C group (normal saline by intraperitoneal injection). Animals were killed at the 12th day after treatment. The tumor volume, positive rate of peritoneal metastasis and positive rate of ascites were measured, and the number of positive cells of CD68, CD8, CD163, and Granzyme B in tumor tissue, peritumoral tissue and distal liver tissue were detected by immunohistochemical method, respectively.(the CD68, CD8 positive cell was macrophage; the CD8, GranzymeB positive cell was T cell). The apoptosis of CD8+ T cells in tumor tissues was tested by the method of double staining with immunohistochemistry and TUNEL.Results:1 For A, B and C group, the tumor volume was (0.13±0.09) cm3, (0.27±0.14) cm3, and (0.41±0.36) cm3; the tumor weight was (0.16±0.14)g, (0.31±0.13)g and (0.34±0.35)g; the positive rate of peritoneal metastasis was 0%,42.86% and 33.33%; the positive rate of ascites was 0%,42.86% and 33.33%; respectively. There was no significant difference among three groups (p>0.05).2 In tumor tissue, peritumoral tissue and distal liver tissue, the number of positive cells for CD68 was (134.29±35.45), (41.72±13.62) and (38.20±13.43); for CD163 was (47.84±1.61), (32.55±31.23) and (31.66±17.68); for CD8 was (104.95±35.77), (119.14±28.24) and (106.12±19.57); for Granzyme B was (11.48±6.60), (13.13±1.84) and (12.39±1.62); respectively. There was significant difference between tumor and peritumoral tissue, between tumor and distal liver tissue for CD68 positive cells (p<0.05). There was no significant difference between tumor, peritumoral and distal liver tissue for CD163, CD8 and GranzymeB positive cells(p>0.05).3 In A, B and C group, the positive cells number was (91.73±22.36), (153.88±47.66) and (134.29±35.45) for CD68; (34.43±9.32), (53.33±1.45) and (47.84±1.61) for CD163; (194.52±71.60), (183.36±59.96) and (104.95±35.77) for CD8; (26.88±1.14), (18.44±6.70) and (11.48±6.60) for Granzyme B;respectively. There was significant difference between A and B group for CD68 and CD163 (p<0.05); between A and C group for CD8 and Granzyme B (p<0.05); between B and C group for CD8 (p<0.05).4 There was a negative correlation not only between the positive cells of CD68 and CD8 (r=-0.221, p=0.030), also between CD163 and Granzyme B (r=-0.464, p=0.030). Furthermore, there was no correlation between the positive cells of CD68 and Granzyme B (r=0.017, p=0.939), as well as between the positive cells of CD163 and CD8 (r=0.224, p=0.316).5 The apoptotic rate of CD8 T cells was (3.63±1.49)%, (6.60±6.60)% and (9.42±4.40)% in A, B and C group, respectively. There was significant difference between A and C group(p<0.05).Conclusions:1 The number of TAM in A group was much less than B group and C group in rat liver cancer, it suggested that clodronate lipsome can deplete macrophages effectively.2 The number of CTL in A group was much more than B group and C group in rat liver cancer, it suggested that TAM can inhibit CTL infiltration. 3 The apoptotic rate of CTL in A group was much less than B group and C group in rat liver cancer, it suggested that TAM can promote CTL apoptosis. |
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