Recently, microRNA-133a (miR-133a) has been found to function in many diseases in previous studies, yet few studies have been focused on its role in glioma. This study aims to investigate the mechanism of miR-133a/CTGF on regulating the malignant phenotypes of glioma cells via the JAK/STAT signaling pathway. Sixty-five human glioma specimens were collected and 30 normal brain tissues were selected as controls. The expression of connective tissue growth factor (CTGF) and miR-133a in tissues was detected, and the relationship between their expression and the clinicopathological features as well as prognosis of glioma was analyzed. MiR-133a and CTGF expression in U87, A172, and HEB cell lines was determined. The expression of CTGF, signaling pathway-, proliferation-, migration-, invasion-, apoptosis- and epithelial-mesenchymal transition (EMT)-related factors was detected. A number of assays were used to detect cell proliferation, migration, invasion, cell cycle, apoptosis, glioma growth, and the targeting site between CTGF and miR-133a. MiR-133a was downregulated and CTGF was upregulated in human glioma tissues and cells. MiR-133a and CTGF expression was related to glioma's WHO staging and size. Downregulated miR-133a and upregulated CTGF caused unfavorable prognosis in glioma. Upregulated miR-133a suppressed CTGF expression and the activation of JAK/STAT signaling pathway, thereby constraining cell colony formation, proliferation, migration and invasion, and promoting apoptosis in glioma. Our study reveals that upregulated miR-133a and downregulated CTGF suppress cell proliferation, migration, and invasion in human glioma through the inhibition of the JAK/STAT signaling pathway.