Journal: Molecular Medicine

Article Title: Deficiency of mindin reduces renal injury after ischemia reperfusion

doi: 10.1186/s10020-022-00578-2

Figure Lengend Snippet: Mindin deficiency inhibited TLR4/JNK/NF-κB signaling activation in renal IR injury A TLR4, phosphorylated JNK, phosphorylated P65, and phosphorylated IκBα measured by western blot analysis in kidneys of WT and mindin KO mice induced by IR or not. B – E Quantitative analysis of TLR4, phosphorylated JNK, phosphorylated P65, and phosphorylated IκBα protein expression. *P < 0.05 versus WT Sham or KO Sham. # P < 0.05 versus WT IR

Article Snippet: Mindin protein (HY-P70142, MedChemExpress) and TLR4 protein (HY-P73586, MedChemExpress) were used to study the protein binding in CM5 electronic chips by biomolecular interaction instrument (Biacore T2000).

Techniques: Activation Assay, Western Blot, Expressing

Journal: Molecular Medicine

Article Title: Deficiency of mindin reduces renal injury after ischemia reperfusion

doi: 10.1186/s10020-022-00578-2

Figure Lengend Snippet: Mindin overexpression promoted production of inflammatory mediators and activation of TLR4/JNK/ NF-κB signaling after HR in vitro. A Representative photomicrographs showing the immunofluorescence of GFP in adGFP and adMindin cells. B Mindin protein expression in adGFP and adMindin cells. C – E Protein levels of TNF-α and MCP-1 in the media of cultured TECs by Western blot. *P < 0.05 versus adGFP Con or adMindn Con. # P < 0.05 versus adGFP HR. F TLR4, phosphorylated JNK, phosphorylated P65, and phosphorylated IκBα measured by western blot analysis in adGFP and adMindin cells induced by HR or not. G – J Quantitative analysis of TLR4, phosphorylated JNK, phosphorylated P65, and phosphorylated IκBα protein expression. *P < 0.05 versus adGFP Con or adMindn Con. # P < 0.05 versus adGFP HR. K Quantitative analysis of TLR4, JNK, P65, TNF-α and MCP-1 mRNA expression in mindin-overexpressed HK-2 cells. ***P < 0.001 versus adGFP HR group

Article Snippet: Mindin protein (HY-P70142, MedChemExpress) and TLR4 protein (HY-P73586, MedChemExpress) were used to study the protein binding in CM5 electronic chips by biomolecular interaction instrument (Biacore T2000).

Techniques: Over Expression, Activation Assay, In Vitro, Immunofluorescence, Expressing, Cell Culture, Western Blot

Journal: Molecular Medicine

Article Title: Deficiency of mindin reduces renal injury after ischemia reperfusion

doi: 10.1186/s10020-022-00578-2

Figure Lengend Snippet: Mindin knockdown inhibited the activation of TLR4/JNK/ NF-κB signaling after HR in vitro. A TLR4, Mindin, phosphorylated JNK, phosphorylated P65, and phosphorylated IκBα measured by western blot analysis in si-Mindin, si-TLR4 and si-JNK cells induced by HR respectively. B – F Quantitative analysis of TLR4, Mindin, phosphorylated JNK, phosphorylated P65, and phosphorylated IκBα protein expression. * P < 0.05, ** P < 0.01 versus HR. G Quantitative analysis of TLR4, JNK, P65, TNF-α and MCP-1 mRNA expression in si-Mindin, si-TLR4 and si-JNK cells induced by HR respectively. ***P < 0.001 versus control HR group

Article Snippet: Mindin protein (HY-P70142, MedChemExpress) and TLR4 protein (HY-P73586, MedChemExpress) were used to study the protein binding in CM5 electronic chips by biomolecular interaction instrument (Biacore T2000).

Techniques: Knockdown, Activation Assay, In Vitro, Western Blot, Expressing, Control

Journal: Molecular Medicine

Article Title: Deficiency of mindin reduces renal injury after ischemia reperfusion

doi: 10.1186/s10020-022-00578-2

Figure Lengend Snippet: Co-immunoprecipitation of TLR4 and mindin in HK-2 cells. Lysates of HK-2 cells were incubated with anti-TLR4 antibody and immune complexes were precipitated by protein A/G beads. The TLR4 complex was determined TLR4 and mindin proteins in the complex. The whole cell lysate was used to detect TLR4 and mindin by Western blot

Article Snippet: Mindin protein (HY-P70142, MedChemExpress) and TLR4 protein (HY-P73586, MedChemExpress) were used to study the protein binding in CM5 electronic chips by biomolecular interaction instrument (Biacore T2000).

Techniques: Immunoprecipitation, Incubation, Western Blot

Journal: Molecular Medicine

Article Title: Deficiency of mindin reduces renal injury after ischemia reperfusion

doi: 10.1186/s10020-022-00578-2

Figure Lengend Snippet: Binding assy of mindin to TLR4 proteins by SPR. Different concen-trations of TLR4 protein (3.125, 6.25, 12.5, 25.0, 50.0, 100.0 nM) were performed to analyse the binding and dissociation rate constants between mindin and TLR4 proteins. The binding constant of them was calculated to evaluate protein binding

Article Snippet: Mindin protein (HY-P70142, MedChemExpress) and TLR4 protein (HY-P73586, MedChemExpress) were used to study the protein binding in CM5 electronic chips by biomolecular interaction instrument (Biacore T2000).

Techniques: Binding Assay, Protein Binding

Journal: Neural Plasticity

Article Title: Toll-Like Receptor 2 Attenuates Traumatic Brain Injury-Induced Neural Stem Cell Proliferation in Dentate Gyrus of Rats

doi: 10.1155/2020/9814978

Figure Lengend Snippet: Immunofluorescence (IF) microphotographs in the dentate gyrus (DG) of the DG in sham and TBI groups at different time points. (a) BrdU-labelled NSCs (red fluor), expression of TLR2 (green fluor), cell nuclei (blue fluor), and BrdU + /TLR2 + /DAPI + cells indicated that TLR2 expression in labelled proliferating cells was possible NSCs; (b) BrdU + cells showed the proliferation of NSCs in the DG during different time points posttrauma. BrdU + cells were more in the TBI group than that in the sham group ( ∗ p < 0.05), and numbers of these cells were significantly different among different time points posttrauma ( # p < 0.05); (c) numbers of BrdU + /TLR2 + cells indicated that the expression of TLR2 was quite different in proliferating cells of the DG among different time points posttrauma. There were more BrdU + cells in the TBI group than that in the sham group ( ∗ p < 0.05), and the numbers of these cells are obviously different among different time points posttrauma ( # p < 0.05). Scale bar: 50 μ m; data is shown as mean ± SEM.

Article Snippet: Primary antibodies were used as follows: mouse anti-TLR2 antibody (1 : 500, Biorbyt, orb191498, San Francisco, CA, USA) and mouse anti- β -actin antibody (1 : 3000, TDY BIOTECH, TDY041, Beijing, China) overnight at 4°C.

Techniques: Immunofluorescence, Expressing

Journal: Neural Plasticity

Article Title: Toll-Like Receptor 2 Attenuates Traumatic Brain Injury-Induced Neural Stem Cell Proliferation in Dentate Gyrus of Rats

doi: 10.1155/2020/9814978

Figure Lengend Snippet: IF in the DG of the TBI group (mouse brain got from 3 days posttrauma). (a) BrdU (red), nestin (green), and DAPI (blue), respectively, exhibited proliferating cells, NSCs, and cell nuclei in the DG. Merged pictures of BrdU + /nestin + /DAPI + showed NSCs (the percentage of NSCs in proliferating cells was 84.30% ± 6.54%); scale bar: 50 μ m; data were expressed as mean ± SEM. (b) Nestin (green), TLR2 (red), and DAPI (blue), respectively, exhibited NSCs, TLR2 expression, and cell nuclei in the DG. Merged pictures of nestin + /TLR2 + /DAPI + showed the expression of TLR2 on NSCs. Scale bar: 50 μ m; data were expressed as mean ± SEM.

Article Snippet: Primary antibodies were used as follows: mouse anti-TLR2 antibody (1 : 500, Biorbyt, orb191498, San Francisco, CA, USA) and mouse anti- β -actin antibody (1 : 3000, TDY BIOTECH, TDY041, Beijing, China) overnight at 4°C.

Techniques: Expressing

Journal: Neural Plasticity

Article Title: Toll-Like Receptor 2 Attenuates Traumatic Brain Injury-Induced Neural Stem Cell Proliferation in Dentate Gyrus of Rats

doi: 10.1155/2020/9814978

Figure Lengend Snippet: Expression of TLR2 protein and mRNA in the DG (western blotting and PCR). (a) Western blotting: electrophoresis bands of TLR2 protein controlled with β -actin; (b) western blotting: the optical density of TLR2 electrophoresis; (c) real-time PCR: the expression of TLR2 mRNA and GAPDH was used as the endogenous reference gene. The TLR2 expression in the protein and mRNA level was significantly higher in the TBI group than that in the sham group ( ∗ p < 0.05), and the TLR2 expression was significantly different among various time points ( # p < 0.05). Data were expressed as mean ± SEM.

Article Snippet: Primary antibodies were used as follows: mouse anti-TLR2 antibody (1 : 500, Biorbyt, orb191498, San Francisco, CA, USA) and mouse anti- β -actin antibody (1 : 3000, TDY BIOTECH, TDY041, Beijing, China) overnight at 4°C.

Techniques: Expressing, Western Blot, Electrophoresis, Real-time Polymerase Chain Reaction

Journal: Neural Plasticity

Article Title: Toll-Like Receptor 2 Attenuates Traumatic Brain Injury-Induced Neural Stem Cell Proliferation in Dentate Gyrus of Rats

doi: 10.1155/2020/9814978

Figure Lengend Snippet: Gene sequences for primer synthesis.

Article Snippet: Primary antibodies were used as follows: mouse anti-TLR2 antibody (1 : 500, Biorbyt, orb191498, San Francisco, CA, USA) and mouse anti- β -actin antibody (1 : 3000, TDY BIOTECH, TDY041, Beijing, China) overnight at 4°C.

Techniques: Sequencing

Journal: Antioxidants

Article Title: By-Product Extracts from Castanea sativa Counteract Hallmarks of Neuroinflammation in a Microglial Model

doi: 10.3390/antiox12040808

Figure Lengend Snippet: Primer used for Real-time qPCR.

Article Snippet: Rabbit anti-TLR4 polyclonal FITC-conjugated antibody was purchased from StressMarq.

Techniques: Sequencing

Journal: Antioxidants

Article Title: By-Product Extracts from Castanea sativa Counteract Hallmarks of Neuroinflammation in a Microglial Model

doi: 10.3390/antiox12040808

Figure Lengend Snippet: Cell surface level of TLR4 in BV-2 cells treated with chestnut leaf or spiny bur extracts and LPS. BV-2 cells were pre-treated (or not) with spiny bur (SB) or leaf (L) extracts (0.5 mg/mL) for 3 h, incubated with or without LPS (0.5 μg/mL) for a total of 24 h, then subjected to flow cytometric analysis of the immunostaining for TLR4 receptor. ( a ) Representative plots of untreated BV-2 cells ( b ) Representative plots of BV-2 cells not activated with LPS ( c ) Representative plots of LPS-stimulated BV-2 cells ( d ) Relative quantification is expressed as MFI, median fluorescence intensity. Results are expressed as means ± SEM of three independent experiments. Statistical analysis was performed by Fisher’s LSD test following one-way ANOVA. ° p < 0.05 significantly different from control cells; * p < 0.05 significantly different from LPS-treated cells.

Article Snippet: Rabbit anti-TLR4 polyclonal FITC-conjugated antibody was purchased from StressMarq.

Techniques: Incubation, Immunostaining, Fluorescence, Control

Journal: Antioxidants

Article Title: By-Product Extracts from Castanea sativa Counteract Hallmarks of Neuroinflammation in a Microglial Model

doi: 10.3390/antiox12040808

Figure Lengend Snippet: RT-PCR analysis of TLR4 gene expression in BV-2 cells treated with chestnut extracts. BV-2 cells were treated with spiny bur or leaf extracts (0.5 mg/mL) for 3 h, then stimulated (or not) with LPS (0.5 µg/mL) for a total of 24 h. At the end of incubation, RNA was extracted from cells and samples were subjected to RT-PCR analysis using a specific primer for TLR4, as explained in the Materials and Methods section. The relative mRNA content of TLR4 was normalised to ( a ) untreated control and ( b ) untreated LPS control. Relative quantification is obtained as reported in the Materials and Methods section; results are expressed as means ± SEM of three independent experiments. Statistical analysis was performed by Fisher’s LSD test following one-way ANOVA. ° p < 0.05 significantly different from control cells; * p < 0.05 significantly different from LPS-treated cells.

Article Snippet: Rabbit anti-TLR4 polyclonal FITC-conjugated antibody was purchased from StressMarq.

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Incubation, Control

Journal: Antioxidants

Article Title: By-Product Extracts from Castanea sativa Counteract Hallmarks of Neuroinflammation in a Microglial Model

doi: 10.3390/antiox12040808

Figure Lengend Snippet: RT-PCR analysis of TLR4 and CD14 gene expression in BV-2 cells treated with different chestnut leaf extract fractions. BV-2 cells were treated for 3 h with 50 μg/mL of fractions at different polarity (L Fr.), then stimulated with 0.5 μg/mL LPS for a total of 24 h. At the end of incubation, RNA was extracted from cells and samples were subjected to RT-PCR analysis using a specific primer for TLR4 ( a ) and CD14 ( b ), as reported in the Materials and Methods section. The relative mRNA contents of TLR4 and CD14 were normalised to LPS-treated cells (red bars). Relative quantification is obtained as described in the Materials and Methods section, and results are expressed as means ± SEM of three independent experiments. Statistical analysis was performed by Fisher’s LSD test following one-way ANOVA. * p < 0.05 significantly different from LPS-treated cells.

Article Snippet: Rabbit anti-TLR4 polyclonal FITC-conjugated antibody was purchased from StressMarq.

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Incubation