Expression Of Bcl-2 And NF-κB In Human Brain Glioma And Their Dependability

PrefaceBrain gliomas are malignant tumors with highest morbidity in central nervous system. There has been no obvious improvement in prognosis of gliomas in recent years though there are many advancements in operation, radiotherapy and chems. Progression in the study of molecular biology of gliomas including apop-tosis impulses the clinical application of gene therapy and biotherapy overwhelmingly and it has developed a new path to the cure of brain gliomas. Many studies regarded apoptosis as their targets.As an elemental vital phenomena, apoptosis is very important in maintaining homeostasis of organisms. Apoptosis exists not only in normal tissue but also in tumors. Growing downward in apoptosis can result in the genesis of tumors and escaping from apoptosis can promote malignant transformation of tumor cells.Bcl-2 gene has significant role in genesis and development of tumors by suppressing apoptosis of tumor cells and prolonging their live time. It is a hot spot in neurosurgery that expression of Bcl-2 in human brain gliomas and its relationship with the degrees in tissue and cell differentiation of gliomas.NF-kB involves in the course of generation and anti-apoptosis of tumor cells. Studies in the recent years have confirmed that abnormal expression of NF-kB has close relationship with the degrees on malignancy of gliomas and its activation has important regulated role in the genesis of gliomas.The mechanism of anti-apoptosis of NF-kB in gliomas is related to Bcl-2 closely. Our research mean to reveal the correlation between Bcl-2 and NF-kB in genesis, progression and apoptosis of gliomas by studying their expression inglioma cells then provide new theoretical basis for gene therapy and biotherapy of gliomas.Materials and methodsSpecimen selectionThere were totally 45 paraffin imbedded specimens of human brain gliomasobtained from Department of Neurosurgery of the Second AfHliated Hospital ofChina Medical University between February, 1999 and March, 2004 . All tumorscorresponding to each specimen were graded according to WHO criteria {GradeI :n=0, Grade II:n=13, Grade III = 12, Grade IV: n = 11).ImmunohistochemistryParaffin embedded specimens were cut into 5 micrometer-thick sections and deparaffinized with xylene and rehydrated. After endogenous peroxidase activity was blocked with 0.3% H2O2 for 10 minutes at room temperature, these sections were treated with 0. lmol/liter citrate (PH 6.0) in a 500-W microwave oven for 10 minutes for antigen retrieval. After blocking nonspecific reactivity with normal nonimmune serum for 30 minutes at room temperature, these sections were incubated with anti-Bcl-2 and anti-NF-KB monoclonal antibody for one hour at room temperature and then with biotin-conjugated second antibody for an additional 30 minutes at 37℃. Positive signals were detected by the S-P method with Ultrasensitive S-P kit. The sections were also counterstained with hematoxylin. Buffy in cytoplasm was considered positive reaction for Bcl-2 and Buffy in cell nucleus was considered positive for NF-kB.Data processingAll of the analysis were performed by SPSS 11.5 software. For the statistical analysis ,x~2 test and Sperman correlation analysis were performed according to requirement of the experimental data. P value < 0. 05 was considered to be significant.ResultsThe positive expression rate of Bcl-2 in 45 glioma specimens was 71. 1% and there was no positive expression of Bcl-2 in non-tumor tissue. The positive expression rate of Bcl-2 in lower potential malignant gliomas was lower than that of higher potential malignant gliomas significantly ( P < 0. 05 ). Sperman rank correlation indicated that the grades of gliomas were positively related with the expression intensity of Bcl-2(r = 0.429, P = 0.014).The positive expression rate of NF-kB in 45 glioma specimens was 48.9% and tliere was no positive expression of NF-kB in non-tumor tissue. The positive expression rate of NF-kB in lower potential malignant gliomas was lower than that of higher potential malignant gliomas significantly (P <0. 05).There were 22 NF-kB-positive specimens among 32 Bcl-2positive specimens and there were 1 NF-KB-positive specimens among 13 Bcl-2-negative specimens. There was a positive correlation between the expression rate of Bcl-2 and NF-KB(r =0.525, P<0.01).DiscussionIt is generally considered that the role of suppression of apoptosis is more important than that of excessive generation in the course of genesis and progression of malignant tumors. Furthermore chems, radiotherapy and biotherapy treat tumors by inducing apoptosis of tumor cells.With uninterrupted improvement of the study on apoptosis, people discovered that Bcl-2 gene family was a kind of protein that has important role in apoptosis. Some studies indicated that there was enchanced expression of Bcl-2 in many kind of tumors including gliomas. Other studies indicated that there was dependability between expression of Bcl-2 and grading and prognosis of gliomas , moreover, Bcl-2 involved in the sensibility of gliomas to radiotherapy and chems.Our study found that the positive rate of Bcl-2 in higher malignant gliomaswas higher than that of lower malignant gliomas significantly and the expression rate of Bcl-2 rised with the enhancement of the malignant degree of gliomas. The fact that positive rates of Bcl-2 in lower malignant gliomas (Grade I and Grade II ) were high(44.4% and 46.2% ) indicated that the expression of Bcl-2 was a kind of frequent events in gliomas and there was intimate correlation between the expression and genesis and progression of gliomas. Simultaneously, the intensity of the expression of Bcl-2 was enhanced with the rising of the ranks in pathology of gliomas(r = 0.441, P = 0. 015). According to these facts, we concluded that the expression level of Bcl-2 in gliomas can reflex the malignant degree of gliomas appropriately and it can be regarded as an effective index in the judgement of biological behaviour of gliomas.NF-kB is a kind of multi-dominance nuclear factor existing in cells wide-spresd and it belongs to Rel gene family. NF-kB has the role of anti-apoptosis in both tissues and cells of living organism and various models of cell culture. NF-kB existing in glioma cells can be activated by many kinds of ecto-stimulus and educe the ability of regulating transcription and express of many kinds of genes.In our study, the positive rate of NF-kB P65 in higher malignant degree gliomas was higher than that of lower malignant degree gliomas significantly and there was no positive expression of NF-kB P65 in non-tumor brain tissue. The fact that the positive rate of NF-kB P65 enhanced with the rising of malignant degrees of gliomas indicated that NF-kB P65 involved in the growth and malignant transformation of gliomas.As a kind of multi-dominance nuclear factor, the function of NF-kB depends on its activation, intranuclear congestion, DNA-binding ability and transcription and expression of corresponding genes causing by it. Its activation is the critical stage in showing activities. According to these facts we can attempt to achieve the objective that suppressing metastasis and recurrence of gliomas by molecular biological means.In our study, there was positive correlation between the expression of Bcl-2 and that of NF-kB in gliomas ( r =0.525, P <0. 01) and both two positive rates are high. This fact indicated that as a kind of nuclear factor can suppress apop-tosis, NF-kB can suppress the programmed death of tumor cells by regulating...