Objective To investigate the expressions of the putative brain tumor stem cell (BTSC) marker CD133, nestin and proliferating cell nuclear antigen (PCNA) in formalin-fixed and paraffin-embedded gliomas, and discuss the BTSC microenvironment: the composition, morphology and distribution of the niche. Methods The samples of 74 patients performed resection of the glioma in our hospital from January 2007 and October 2008 were chosen. According to WHO 2008 classification of nervous system tumors, they were assigned into low-grade glioma group (grade Ⅱ, 22) and high-grade glioma group(grade Ⅲ, 27 and grade Ⅳ,25). Immunohistochemistry was used to detect the expression of CD 133 in 74 cases of brain gliomas. Double immunofluorescence staining was employed to detect the co-expressions of CD133/nestin or CD133/PCNA. The percentages of the CD133+cells, CD133+ blood vessels and CD 133+ niches were calculated, and correlation analysis was also performed on their percentage and the pathological grading of the tumor. Results Accumulated CD 133+ cells grew in the niches. CD 133+ niches could be observed in all grade gliomas; low-grade group showed lower level expression of the CD 133+ niches, fewer proliferating cells, clearer boundary between the 2 neighboring niches and fewer surrounding blood vessels as compared with high-grade group. Besides CD113+/nestin+ BTSCs, some subsets as CD133+/nestin-cells and CD133-/PCNA+ cells were noted in the niches in the high-grade group. The percentages of CD 133+ cells, CD 133+ blood vessels and CD 133+ niches were different in different grades of gliomas and the higher the grade of gliomas was, the higher their expressions were. Positive correlation was observed between the percentage of CD 133+ niches and the percentage of CD133+ blood vessels (r=0.425, P=0.000). The expression of CD133+ niches in high-grade glioma were higher than that in low-grade tumors (F=5.324, P=0.002). Conclusion Glioma tissues have niche structures, which composed of CD133+/nestin+ BTSCs and some subsets. CD133+ blood vessels were playing a key role in maintaining the niche structure, and the expression of CD133+ niches is statistically different in different pathological grading tumors.