| ObjectiveBlood-brain barrier ( BBB) is composed of capillary endothelial cells, intercellular tight junction, base membrane, astroglial processes. BBB limits adequate delivery of drugs to brain. Scientists have been looking for a safety method to open the BBB. Now the most noticeable method is opening BBB by vasoactive compounds. Vasoactive compounds can increase the permeability of blood vessel. Normal brain capillaries appear to resist the efforts of vasoactive compounds. In contrast, brain tumor capillaries or injured brain capillaries appear to lose the ability to resist the effects of increased permeability by these vasoactive compounds.Malignant brain tumors represent a serious medical challenge for they are the second leading neurological cause of death, with the majority of patients dying within a year of initial diagnosis. Brain tumor capillaries have a blood -tumor barrier ( BTB) that limits adequate delivery of antitumor agents to tumor tissue. Gliomas are the most prevalent type of brain tumor. Recently, biochemical opening of the BTB with vasoactive compounds have been used to increase drug delivery to gliomas. Studies have demonstrated that this approach do not increase drug delivery in normal brain, but result in a selective, significant increase in permeability in the tumor. Currently, the most effective compound to selectively open the BTB is bradykinin or its analog, receptor-mediated perme-abilizer-7 ( RMP-7). The selective increases in BTB permeability induced by bradykinin are mediated solely by B2 receptor. In clinical study there is very great difference of increase of BTB permeability induced by bradykinin in brain tumors with the same diagnose. The variability in response of tumors to bradykinin is likely related to the expression level of bradykinin B2 receptor of tumor. It is the urgent task to recognize the reason of the variability in response of braintumors to bradykinin.This study used fresh glioma samples to determine expression level of bradykinin B2 receptor of glioma with different pathological grade and analyzed the difference of bradykinin B2 receptor expression level among gliomas with different pathological grade. The result would supply experimental basis for clinical application of bradykinin.Materials1. Experimental specimens: fresh tissue samples were obtained from 28 patients with brain tumor who underwent surgery in the Department of Neurosurger-y, the Affiliated Hospital of China Medical University, Shenyang, China. After resection the samples were immediately frozen in liquid nitrogen then kept at -70 C. Two cases of normal brain tissues of the edge of tumor were selected as control.2. Experimental reagents: goat anti human bradykinin B2 receptor antibody and enhanced chemiluminescence kit ( Santa Cruz Biotechnology, U. S. A. ) ; streptavidin-peroxidase (SP) kit (Zhorigshan Company, Beijing).3. Experimental instruments: stir apparatus, low temperature centrifugal machine, electrophoresis apparatus, transmark apparatus, freeze microtome, Luzex-F image analysis system.MethodsThe clinicopathologic findings were diagnosed by reviewing all hematoxylin and eosin (H&E) stained tissue sections. The tumor grade was determined according to the 2000 World Health Organization (WHO) Central Nervous System Tumor Classification.According to the result of H&E screened 8 cases of grade 1, 8 cases of grade II, 8 cases of grade III. Devide the specimens into 3 groups. Every grade was as one group.To determine the bradykinin B2 receptor expression level of glioma, Immu-nohistochemistry and Western blot methods were used.Integrated Density Value (IDV) for the B2 receptor protein doublet band was calculated by Fluor Chen 2. 0 software from scanned immunoblot by Chemi Imager 5500 V2.03 software. The distribution of cells immunopositive for anti-B2 receptor antibody was analysed semiquantitatively by Luzex-F Real-Time Image Analysis System. Target area/election district area ratio was calculated.Data from Immunohistochemical and We...
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