Nuclear Factor-kappa B And Its Tissue Inhibitor In Colorectal Carcinoma

Objective: Colorectal carcinoma is one of the most common malignant tumors in our country. The tendency of incidence is increasing today.Although therapy methods have been improved, survival rate of five years is still very low. Tumor invasion and metastasis are the most important cause of death. So the study for mechanism of invasion and metastasis of colorectal carcinoma has become a focus at present. The biological mechanisms of Colorectal carcinoma is very complicate. Many factors are associated the poor prognosis of colorectal carcinoma. NF-κB plays important roles in malignant transformation, development and metastasis of colorectal cancer by anti-apoptosis. It maybe affect the occurrence and progression of tumor via promoting the cell proliferation, inhibiting, apoptosis. NF-κB is released from I-κB complexes (Inhibitor kappa B), and translocates to the nucleus, then promotes gene transcription. The high expression and activity of NF-κB have been found in many tumors so far. On the other hand, there is a kind of tissue inhibitor of NF-κB (I-κB) in vivo, which can inhibit the activities of NF-κB and hold back the invasion and metastasis of carcinoma. I-κB is a kind of tissue inhibitor of NF-κB. I-κB masking nuclear localization signal (NLS) of NF-κB to prevent nuclear translocation of NF-κB. Once receive motivation from signaling molecule,NF-κB separate from I-κB, and translocate to the nucleus. I-κB can inhibits the activity of NF-κB,effects the process of invasion and metastasis of colorectal carcinoma. The present study aimed at finding the expressive patterns of NF-κB and I-κB and the relationship with progress of colorectal carcinoma using flow cytometry, immunohistochemistry staining and Western blotting.Methods: 1 Specimens: 48 cases of colorectal carcinoma specimens and paired normal tissues were collected from patients operated in the Surgery Department of the Fourth Hospital of Hebei Medical University during March 2006 to July 2005. All specimens were determined by pathology.2 Flow cytometry of NF-κB : Use flow cytometer to quantitative analysis the expression of NF-κB.3 Immunohistochemistry staining was used to detect the expression and distribution of NF-κB and I-κB in normal and colorectal carcinoma tissues. The experiment procedures were according to the Test Kit introduction.4 Western blotting of NF-κB and I-κB: The extracts from normal and colorectal carcinoma tissues were separated on 8%SDS-PAGE, and then blotted onto PVDF membrane. The membrane was immunologically stained with anti- NF-κB and anti- I-κB antibodies. The results were analyzed by digital imaging system. Results: 1 Flow cytofluorimetry suggested that the level of NF-κB in colorectal carcinoma tissues was obviously higher than that in the normal tissues. There was a significant difference (P<0.01). With the progression of Dukes stage, the contents of NF-κB were gradually increased.2 Western blotting suggested that the level of NF-κB in colorectal carcinoma tissues was obviously higher than that in the normal tissues. There was a significant difference (P<0.01).The expression levels of NF-κB in colorectal carcinoma tissues with serous membrane layer or surrounding soft tissue invasion was higher than those in the carcinoma tissues with muscular layer invasion. There was a significant difference (P<0.01) between them. The expression level of NF-κB in colorectal carcinoma tissues with lymph node metastasis was much higher than those in the colorectal carcinoma tissues without lymph node metastasis. There was a significant difference (P<0.05) between them. The expression level of NF-κB in lower differentiated colorectal carcinoma was higher than those in the higher differentiated colorectal carcinoma. There was a significant difference (P<0.01). With the progression of Dukes stage, the contents of NF-κB were gradually increased.Western blotting suggested that the level of I-κB in colorectal carcinoma tissues was obviously lower than that in the normal tissues ; There was a significant difference (P<0.01).The expression levels of I-κB in colorectal carcinoma tissues with serous membrane layer or surrounding soft tissue invasion was lower than those of the carcinoma tissues with muscular layer invasion. But nonsignificant difference; The expression level of I-κB in colorectal carcinoma tissues with lymph node metastasis was lower than those in the colorectal carcinoma tissues without lymph node metastasis. But nonsignificant difference. The expression level of I-κB in lower differentiated colorectal carcinoma was depressed compared with higher differentiated colorectal carcinoma. There was a significant difference (P<0.05). With the progression of Dukes stage, the contents of I-κB was gradually declined.(2)Western blotting suggested that the level of phosphorylated I-κB in colorectal carcinoma tissues was obviously higher than that in the normal tissues ; There was a significant difference (P<0.01).The levels of phosphorylated I-κB in colorectal carcinoma tissues with serous membrane layer or surrounding soft tissue invasion was higher than those of the carcinoma tissues with muscular layer invasion. But nonsignificant difference. The expression level of phosphorylated I-κB in colorectal carcinoma tissues with lymph node metastasis was higher than those in the colorectal carcinoma tissues without lymph node metastasis,But nonsignificant difference. With the progression of Dukes stage, the contents of phosphorylated I-κB were gradually increased.3 The results of immunohistochemistry staining suggested that there were more NF-κB positive particles in colorectal carcinoma tissues than that in normal tissues. The positive particles of I-κB in normal tissues were much more than those in colorectal carcinoma tissues. The positive particles of NF-κB were mainly distributed in the cytoplasm of carcinoma cells. The distribution of I-κB was the similar to NF-κB carcinoma. These results were consistent with the results of flow cytometry and Western blotting.Conclusions: 1 The level of NF-κB in colorectal carcinoma was significantly increased compared with normal tissues and was related to the invasion depth, lymph node metastasis and the Dukes stage. The results suggested that NF-κB plays an important role in the process of invasion and metastasis of colorectal carcinoma.2 The level of I-κB in colorectal carcinoma was significantly declined compared with normal tissues. The level of phosphorylated I-κB in colorectal carcinoma was significantly increased compared with normal tissues. The results show that the phosphorylation and degradation of I-κB play an important role in the process of the occurrence and progression of colorectal carcinoma. The level of I-κB in lower differentiated colorectal carcinoma was decreased compared with higher differentiated colorectal carcinoma. The results show that I-κB affects the prognosis of colorectal carcinoma.3 The level of NF-κB in lower differentiated colorectal carcinoma was increased compared with higher differentiated colorectal carcinoma. The results show that NF-κB affects the prognosis of colorectal carcinoma.