Glioma　is　the　most　common　malignancy　in　the　brain,　with　poor　survival　and　often　highly　resistant　to　chemotherapy　and　radiotherapy.　Temozolomide　(TMZ)　is　an　alkylating　agent　widely　used　for　glioma　treatment.　However,　resistance　to　TMZ　results　in　treatment　failure,　while　the　underlying　mechanisms　remain　unclear.　Mounting　evidence　suggests　that　dysregulated　microRNA　(miRNA)　expression　plays　a　critical　function　in　glioma　development　and　resistance　to　TMZ　treatment.　In　this　study,　we　found　that　miR-23b-5p　was　downregulated　in　glioma　tissues　and　cell　lines.　Overexpression　of　miR-23b-5p　inhibited　cell　proliferation　and　promoted　cell　apoptosis　in　glioma　cells,　while　miR-23b-5p　enhanced　the　chemosensitivity　of　TMZ　in　glioma　cells.　Furthermore,　we　identified　that　Toll-like　receptor　4　(TLR4)　is　a　direct　target　of　miR-23b-5p　in　glioma　cells.　Knockdown　of　TLR4　suppressed　cell　proliferation　and　enhanced　cell　apoptosis　and　promoted　chemosensitivity　to　TMZ　treatment　in　glioma　cells.　In　addition,　we　demonstrated　that　overexpression　of　TLR4　abrogated　the　regulatory　function　of　miR-23b-5p　in　glioma　cells　on　cell　proliferation,　cell　apoptosis,　and　the　chemosensitivity　of　TMZ　treatment.　In　summary,　our　data　suggest　that　miR-23b-5p　promotes　the　chemosensitivity　of　TMZ　via　negatively　regulating　TLR4　in　glioma,　which　provides　a　new　therapeutic　strategy　for　TMZ-resistant　glioma　treatment.