Abstract
BackgroundGlioblastoma remains one of the most lethal brain cancers. T-cell immunoglobulin and mucin domain 1 (Tim-1) is associated with various immune diseases. The molecular mechanism of Tim-1 in regulating glioblastoma cell proliferation, invasion, and migration is still unknown. Moreover, it has shown that miR-133a plays an important role in glioblastoma. However, little is known about the interaction between Tim-1 and miR-133a in glioblastoma.MethodsTim-1 expression in glioblastoma and normal brain tissues was detected by qPCR, Western Blot and IHC. After Tim-1 knockdown in U251 and U87 cells, genes showing significantly differential expression, along with the significant differential miRNAs were analyzed using RNA-seq analysis. The binding sites were verified using dual-luciferase reporter gene assay. U251 and U87 cells were allocated into the small harpin-negative control (sh-NC), sh-Tim-1, sh-Tim-1 + inhibitor NC, and sh-Tim-1 + miR-133a inhibitor group. Cell proliferation, migration, and invasion were determined by CCK-8, flow cytometry, wound-healing and Transwell assays, respectively. Next, U251 and U87 cells were allocated into the mimic NC, miR-133a mimic, miR-133a mimic + pcDNA3.1, and miR-133a mimic + pcDNA3.1-TGFBR1 groups, followed by the detection of cell proliferation, migration, and invasion. Western blot was used to identify the expression of vital kinases in the Wnt/β-catenin pathway.ResultsTim-1 was highly expressed in glioblastoma tissues compared with that in normal brain tissues. RNA-seq analysis showed that Tim-1 knockdown could lead to the downregulation of TGFBR1 and the upregulation of miR-133a. The binding sites between TGFBR1 and miR-133a were confirmed. Tim-1 knockdown impaired the invasion, migration, proliferation of U251 and U87 cells, which could be reversed by miR-133a downregulation. miR-133a upregulation inhibited the proliferation, invasion, and migration of U251 and U87 cells, which could be reversed by TGFBR1 upregulation. Tim-1 knockdown and miR-133a upregulation could inhibit the activation of the Wnt/β-catenin pathway, while the elevation of TGFBR1 showed opposite effects.ConclusionTim-1 knockdown inhibited glioblastoma cell proliferation, invasion, and migration through the miR-133a/TGFBR1 axis and restrained the activation of the Wnt/β-catenin pathway.
Highlights
Glioblastoma remains one of the most lethal brain cancers
The cell density of glioblastoma tissues was elevated with the increase of T-cell immunoglobulin and mucin domain 1 (Tim-1) expression, which suggested that high Tim-1 expression may be negatively correlated with glioblastoma malignant proliferation
Tim‐1 regulated the miR‐133a/Transforming growth factor beta receptors type 1 (TGFBR1) axis To identify the downstream mechanism of Tim-1, the expressions of genes were analyzed through RNA-seq analysis after the downregulation of Tim-1 in U87 and U251 cell lines; the differential pathways were analyzed by bioinformatics (Fig. 2A, B)
Summary
Glioblastoma remains one of the most lethal brain cancers. The molecular mechanism of Tim-1 in regulating glioblastoma cell proliferation, invasion, and migration is still unknown. It has shown that miR-133a plays an important role in glioblastoma. Little is known about the interaction between Tim-1 and miR-133a in glioblastoma. Glioblastoma, as a common and severe neurologic tumor, ranks the highest level in World Health Organization’s brain tumor classification [1, 2]. Glioblastoma is characterized by rapid dissemination, invasive growth, and high-degree cell heterogeneity [3]. The common treatments (including surgery, radiotherapy, and chemotherapy) show no evident effect on the permanent therapy of glioblastoma [6]. A T cell immunity based-therapy has been reported as a promising method to enhance the prognosis of glioblastoma patients [7]
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