Fig 1: MAP kinase‐interacting serine/threonine‐protein kinase 2/hypoxia‐inducible factor‐1α (MKNK2/HIF‐1α) axis is critical for M1 polarization in rat microglia exposed to oxygen‐glucose deprivation/reoxygenation (OGD/R). (A) mRNA expression of MKNK2 in microglia exposed to OGD/R or not was assessed by RT‐qPCR. (B) Plasmid transfection efficiency was observed using fluorescence microscopy, and mRNA expression of MKNK2 in microglia after knockdown of MKNK2 was assessed by RT‐qPCR. (C) The protein expression of MKNK2 and HIF‐1α in cells with different treatments was assessed by Western blot assays. (D) Detection of inflammatory factors IL‐1β and IL‐10 secreted by microglia by enzyme‐linked immunosorbent assay (ELISA). (E) Expression of genes related to M1 phenotype (CD16 and iNOS) in microglia after OGD/R and knockdown of MKNK2 detected by RT‐qPCR. (F) Expression of genes related to M2 phenotype (Arg‐1 and CD206) in microglia after OGD/R and knockdown of MKNK2 was detected by RT‐qPCR. (G) The relative fluorescence intensity of M1 phenotype (CD16) or M2 phenotype (Arg‐1) in rat microglia after OGD/R and knockdown of MKNK2 was detected by immunocytochemistry. (H) Protein expression of MKNK2 and HIF‐1α in OGD/R‐induced cells treated with knockdown of MKNK2 in combination with the HIF‐1α pathway activator, Fenbendazole‐d3, was analyzed by Western blot analysis. (I) Expression of CD16 and Arg‐1 in OGD/R‐induced cells treated with knockdown of MKNK2 in combination with the HIF‐1α pathway activator, Fenbendazole‐d3, detected by RT‐qPCR. Values are mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. n = 3. p‐values were calculated using an unpaired t‐test or ANOVA.
Fig 2: Downregulation of MAP kinase‐interacting serine/threonine‐protein kinase 2 (MKNK2) expression significantly alleviates brain and neuronal injury in rats induced with middle cerebral artery occlusion/reperfusion (MCAO/R). (A) The neurological score of rats was assessed in MCAO/R‐induced rats with MKNK2 knockdown or not. (B) The area of cerebral infarction between the sham‐operated group, the MCAO/R rats, and the lentivirus‐infected MCAO/R rats was analyzed by 2,3,5‐triphenyltetrazolium chloride (TTC) staining. (C) HE staining analysis of brain damage in rats with sh‐MKNK2 induced by MCAO/R. (D) Neuronal degeneration in MCAO/R rats with downregulated MKNK2 expression was analyzed using FJC staining. (E) Detection of apoptosis in sh‐MKNK2‐treated MCAO/R rats by TUNEL. (F) Detection of CD16 in brain tissues of sh‐MKNK2‐treated MCAO/R rats by immunohistochemistry. (G) The content of inflammatory markers (IL‐6 and IL‐1β) secreted by microglia in sh‐MKNK2‐treated MCAO/R rats was analyzed using enzyme‐linked immunosorbent assay (ELISA). (H) Dual‐labeling immunofluorescence analysis of MKNK2 expression (green) in rat brain microglia, iba1 (red). Values are mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. n = 5 per group. p‐values were calculated using an unpaired t‐test or ANOVA.
Fig 3: Krueppel‐like factor 7 (KLF7) mediates MAP kinase‐interacting serine/threonine‐protein kinase 2/hypoxia‐inducible factor‐1 (MKNK2/HIF‐1) signaling to promote microglia M1 polarization. (A) Plasmid transfection efficiency was observed using fluorescence microscopy, and mRNA expression of KLF7, MKNK2, and HIF‐1α in cells transfected with oe‐KLF7 alone or combined with oe‐MKNK2 plasmid before oxygen‐glucose deprivation/reoxygenation (OGD/R) induction was examined using RT‐qPCR. (B) The protein expression of KLF7, MKNK2, and HIF‐1α in cells transfected with oe‐KLF7 + oe‐MKNK2 or oe‐NC before OGD/R induction was examined using Western blot. (C) The expression of M1 microglia‐related genes (CD16 and iNOS) in the cells was examined using RT‐qPCR. (D) The expression of M2 microglia‐related genes (CD206 and Arg‐1) in the cells was examined using RT‐qPCR. (E) The relative fluorescence intensity of M1 phenotype (CD16) or M2 phenotype (Arg‐1) in rat microglia was detected by immunocytochemistry. (F) The contents of IL‐1β and IL‐6 in the cells were evaluated using enzyme‐linked immunosorbent assay (ELISA). Values are mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. n = 3. p‐values were calculated using an unpaired t‐test or ANOVA.
Fig 4: Krueppel‐like factor 7/MAP kinase‐interacting serine/threonine‐protein kinase 2/hypoxia‐inducible factor‐1α (KLF7/MKNK2/HIF‐1α) axis mediates middle cerebral artery occlusion/reperfusion (MCAO/R)‐induced neuronal injury. (A) Assessment of neurobehavioral scores in rats at 24 h postoperatively. (B) The mRNA expression of KLF7 and MKNK2 in brain tissues of oe‐KLF7 combined with oe‐MKNK22 lentivirus‐injected MCAO/R rats was examined using RT‐qPCR. (C) The expression of CD16 in the brain tissues of rats was examined using RT‐qPCR. (D) The concentrations of IL‐1β and IL‐6 in the brain tissues of rats were evaluated using enzyme‐linked immunosorbent assay (ELISA). (E) Detection of cerebral infarction in oe‐KLF7 combined with oe‐MKNK2 lentivirus‐infected MCAO/R rats using 2,3,5‐triphenyltetrazolium chloride (TTC) staining. (F) The degree of brain damage in MCAO/R rats was observed using HE staining. (G) The degeneration of neurons in the brains of the rats was observed using FJC staining. (H) Dual‐labeling immunofluorescence analysis of MKNK2 expression (green) in rat brain microglia, iba1 (red). Values are mean ± SEM, **p < 0.01, ***p < 0.001, and ****p < 0.0001. n = 5. p‐values were calculated using ANOVA.
Supplier Page from Novus Biologicals, a Bio-Techne Brand for Fc gamma RIII (CD16) Antibody (5B11) - BSA Free
Available conjugates: Available conjugates: UnconjugatedSpecificity: Recognizes CD16 antigenSizes Available: 0.1 mg