mitogen activated protein kinase mapk pathway inhibitors  (TargetMol)

Journal: Stem Cell Research & Therapy

Article Title: Melatonin enhances osteogenic differentiation of dental pulp mesenchymal stem cells by regulating MAPK pathways and promotes the efficiency of bone regeneration in calvarial bone defects

doi: 10.1186/s13287-022-02744-z

Figure Lengend Snippet: Effects of melatonin (0, 100, 10 and 1 μM) on ALP, BMP-2, OCN, RUNX2, COX-2, NF-κB and MAPK pathway protein level expression in melatonin-untreated DPSCs and melatonin-treated DPSCs with osteogenic differentiation after 24 h incubation. a , b DPSCs with 100 μM melatonin treatment enhanced ALP, OCN and RUNX2 protein expression and downregulated COX-2 and NF-κB protein expression. c DPSCs with 100 μM melatonin treatment induced phosphorylation of p38 and ERK. The phosphorylation of JNK was not observed. d Quantitative analysis for the ratios of p-p38/p38, p-JNK/JNK and p-ERK/ERK. ( GM growth medium, OM osteogenic medium, Mel melatonin) (* p < 0.05, ** p < 0.01)

Article Snippet: Melatonin (M5250), SB203580 (p38 mitogen-activated protein kinase [MAPK] pathway inhibitor), and PD98059 (ERK/MAPK pathway inhibitor) were purchased from Sigma-Aldrich (St Louis, MO, USA) and dissolved in dimethylsulfoxide (DMSO) and then diluted in PBS immediately before use.

Techniques: Expressing, Incubation

Journal: Stem Cell Research & Therapy

Article Title: Melatonin enhances osteogenic differentiation of dental pulp mesenchymal stem cells by regulating MAPK pathways and promotes the efficiency of bone regeneration in calvarial bone defects

doi: 10.1186/s13287-022-02744-z

Figure Lengend Snippet: Effects of MAPK pathway inhibitors on osteogenic differentiation of DPSCs treated with 100 μM melatonin at 7 and 14 days after incubation with osteogenic differentiation medium. a , b SB203580 (p38 MAPK inhibitor) and PD98059 (ERK MAPK inhibitor) significantly reduced the mineral matrix depositions of 100 μM melatonin-treated DPSCs. c The gene expression of ALP, BMP-2, OCN, and RUNX2 in 100 μM melatonin-treated DPSCs was suppressed in the experimental groups with SB203580 and PD98059 inhibitors. Scale bar = 100 μm. (* p < 0.05, ** p < 0.01)

Article Snippet: Melatonin (M5250), SB203580 (p38 mitogen-activated protein kinase [MAPK] pathway inhibitor), and PD98059 (ERK/MAPK pathway inhibitor) were purchased from Sigma-Aldrich (St Louis, MO, USA) and dissolved in dimethylsulfoxide (DMSO) and then diluted in PBS immediately before use.

Techniques: Incubation, Expressing

Journal: Frontiers in Pharmacology

Article Title: Glycyrrhetinic Acid Protects Renal Tubular Cells against Oxidative Injury via Reciprocal Regulation of JNK-Connexin 43-Thioredoxin 1 Signaling

doi: 10.3389/fphar.2021.619567

Figure Lengend Snippet: c-Jun N-terminal kinase (JNK)-mediated oxidative stress induces connexin 43 (Cx43). (A) Induction of JNK phosphorylation and expression of Cx43 by Px-12. NRK-52E cells in 6-well plates were incubated with Px-12 for 0, 1, 3, 6, 9, and 12 h separately. Cellular lysates were then analyzed by western blotting for phosphorylated JNK and Cx43 expression. Densitometric analysis of p -JNK and Cx43 are shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in control). (B) Effects of 18α-glycyrrhetinic acid (Ga) and sp600125, an inhibitor of JNK, on Px12-induced cell injury. NRK-52E cells were pretreated with Ga (20 μM) and sp600125 (20 μM) for 1 h and then challenged with Px-12 for another 24 h. Then, cell viability was evaluated using a Cell Counting Kit-8 (CCK-8) assay. Data are expressed as the percentage of living cells vs. the untreated control (mean ± SD, n = 6). * p < 0.01 vs. the control. ## p < 0.01 vs. Px-12 in control. (C) Influence of sp600125 and Ga on NRK-52E cell apoptosis staining. NRK-52E cells in 96-well plate were pretreated with 18α-Ga and sp600125 for 1 h and then incubated with Px-12 for another 24 h. Then, apoptotic cells were stained via DT-mediated dUTP nick end labeling (TUNEL) and 4,6-diamidino-2-phenylindole (DAPI) (magnification, ×400). The ratio of positive cells is shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in the control).

Article Snippet: The JNK-mitogen-activated protein kinase (MAPK) signaling pathway inhibitor (sp600125, A4604) was obtained from ApexBio Technology China.

Techniques: Expressing, Incubation, Western Blot, Cell Counting, CCK-8 Assay, Staining, End Labeling, TUNEL Assay

Journal: Frontiers in Pharmacology

Article Title: Glycyrrhetinic Acid Protects Renal Tubular Cells against Oxidative Injury via Reciprocal Regulation of JNK-Connexin 43-Thioredoxin 1 Signaling

doi: 10.3389/fphar.2021.619567

Figure Lengend Snippet: Reciprocal modulation of connexin 43 (Cx43) and c-Jun N-terminal kinase (JNK). (A) Effect of 18α-glycyrrhetinic acid (Ga) and sp600125 on JNK phosphorylation and expression of Cx43. NRK-52E cells in six-well plates were pretreated with Ga and sp600125 for 1 h and then challenged with Px-12 for another 1 h. Cellular lysates were then analyzed by western blotting for phosphorylated JNK and Cx43 expression. Densitometric analysis of p -JNK and Cx43 are shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in the control). (B) NRK-52E cells were transfected with either Cx43 siRNA or siRNA control for 24 h. Cellular lysates were analyzed by western blotting for Cx43 expression. Densitometric analysis of Cx43 is shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the siRNA control group). (C) Effect of Cx43 siRNA on JNK phosphorylation and Cx43 expression. The transfected or normal cells were incubated with Px-12 for 1 h separately. Then, cellular lysates were analyzed by western blotting for phosphorylated JNK and Cx43 expression. Densitometric analysis of p -JNK and Cx43 are shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in the siRNA control group). (D) Effect of Cx43 siRNA on cell viability. The transfected or normal cells were incubated with Px-12 for 24 h. Then, cell viability was evaluated using a Cell Counting Kit-8 (CCK-8) assay. Data are expressed as the percentage of living cells vs. the untreated control (mean ± SD, n = 6). * p < 0.01 vs. Px-12 in normal cells. ## p < 0.01 vs. Px12 in the siRNA control group. (E) Effect of Cx43 siRNA on caspase-3 activation. The transfected or normal cells were incubated with Px-12 for 24 h. Then, cellular lysates were analyzed by western blotting for expression of cleaved caspase-3. Densitometric analysis of cleaved caspase-3 and caspase-3 are shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. Px-12 in the siRNA control group).

Article Snippet: The JNK-mitogen-activated protein kinase (MAPK) signaling pathway inhibitor (sp600125, A4604) was obtained from ApexBio Technology China.

Techniques: Expressing, Western Blot, Transfection, Incubation, Cell Counting, CCK-8 Assay, Activation Assay

Journal: Frontiers in Pharmacology

Article Title: Glycyrrhetinic Acid Protects Renal Tubular Cells against Oxidative Injury via Reciprocal Regulation of JNK-Connexin 43-Thioredoxin 1 Signaling

doi: 10.3389/fphar.2021.619567

Figure Lengend Snippet: c-Jun N-terminal kinase (JNK)-mediated oxidative stress induces connexin 43 (Cx43). (A) Induction of JNK phosphorylation and expression of Cx43 by Px-12. NRK-52E cells in 6-well plates were incubated with Px-12 for 0, 1, 3, 6, 9, and 12 h separately. Cellular lysates were then analyzed by western blotting for phosphorylated JNK and Cx43 expression. Densitometric analysis of p -JNK and Cx43 are shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in control). (B) Effects of 18α-glycyrrhetinic acid (Ga) and sp600125, an inhibitor of JNK, on Px12-induced cell injury. NRK-52E cells were pretreated with Ga (20 μM) and sp600125 (20 μM) for 1 h and then challenged with Px-12 for another 24 h. Then, cell viability was evaluated using a Cell Counting Kit-8 (CCK-8) assay. Data are expressed as the percentage of living cells vs. the untreated control (mean ± SD, n = 6). * p < 0.01 vs. the control. ## p < 0.01 vs. Px-12 in control. (C) Influence of sp600125 and Ga on NRK-52E cell apoptosis staining. NRK-52E cells in 96-well plate were pretreated with 18α-Ga and sp600125 for 1 h and then incubated with Px-12 for another 24 h. Then, apoptotic cells were stained via DT-mediated dUTP nick end labeling (TUNEL) and 4,6-diamidino-2-phenylindole (DAPI) (magnification, ×400). The ratio of positive cells is shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in the control).

Article Snippet: The JNK-mitogen-activated protein kinase (MAPK) signaling pathway inhibitor (sp600125, A4604) was obtained from ApexBio Technology China.

Techniques: Expressing, Incubation, Western Blot, Cell Counting, CCK-8 Assay, Staining, End Labeling, TUNEL Assay

Journal: Frontiers in Pharmacology

Article Title: Glycyrrhetinic Acid Protects Renal Tubular Cells against Oxidative Injury via Reciprocal Regulation of JNK-Connexin 43-Thioredoxin 1 Signaling

doi: 10.3389/fphar.2021.619567

Figure Lengend Snippet: Reciprocal modulation of connexin 43 (Cx43) and c-Jun N-terminal kinase (JNK). (A) Effect of 18α-glycyrrhetinic acid (Ga) and sp600125 on JNK phosphorylation and expression of Cx43. NRK-52E cells in six-well plates were pretreated with Ga and sp600125 for 1 h and then challenged with Px-12 for another 1 h. Cellular lysates were then analyzed by western blotting for phosphorylated JNK and Cx43 expression. Densitometric analysis of p -JNK and Cx43 are shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in the control). (B) NRK-52E cells were transfected with either Cx43 siRNA or siRNA control for 24 h. Cellular lysates were analyzed by western blotting for Cx43 expression. Densitometric analysis of Cx43 is shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the siRNA control group). (C) Effect of Cx43 siRNA on JNK phosphorylation and Cx43 expression. The transfected or normal cells were incubated with Px-12 for 1 h separately. Then, cellular lysates were analyzed by western blotting for phosphorylated JNK and Cx43 expression. Densitometric analysis of p -JNK and Cx43 are shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in the siRNA control group). (D) Effect of Cx43 siRNA on cell viability. The transfected or normal cells were incubated with Px-12 for 24 h. Then, cell viability was evaluated using a Cell Counting Kit-8 (CCK-8) assay. Data are expressed as the percentage of living cells vs. the untreated control (mean ± SD, n = 6). * p < 0.01 vs. Px-12 in normal cells. ## p < 0.01 vs. Px12 in the siRNA control group. (E) Effect of Cx43 siRNA on caspase-3 activation. The transfected or normal cells were incubated with Px-12 for 24 h. Then, cellular lysates were analyzed by western blotting for expression of cleaved caspase-3. Densitometric analysis of cleaved caspase-3 and caspase-3 are shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. Px-12 in the siRNA control group).

Article Snippet: The JNK-mitogen-activated protein kinase (MAPK) signaling pathway inhibitor (sp600125, A4604) was obtained from ApexBio Technology China.

Techniques: Expressing, Western Blot, Transfection, Incubation, Cell Counting, CCK-8 Assay, Activation Assay

Journal: Frontiers in Pharmacology

Article Title: Glycyrrhetinic Acid Protects Renal Tubular Cells against Oxidative Injury via Reciprocal Regulation of JNK-Connexin 43-Thioredoxin 1 Signaling

doi: 10.3389/fphar.2021.619567

Figure Lengend Snippet: Connexin 43 (Cx43) inhibition suppresses oxidative stress by regulating thioredoxin 1. (A) Effect of 18α-glycyrrhetinic acid (Ga) and sp600125 on reactive oxygens species (ROS)/O 2 − production. NRK-52E cells in a 96-well plate were pretreated with Ga and sp600125 separately for 1 h, challenged with Px-12 for another 1 h, and then incubated with a ROS/O 2 − probe for 30 min. Images were captured using an inverted fluorescence microscope (×100). Mean fluorescence intensity is shown at the bottom (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in the control). (B) Effects of Ga on thioredoxin 1. NRK-52E cells in six-well plates were pretreated with Ga for 1 h and then challenged with Px-12 for another 1 h. Then, cellular lysates were analyzed by western blotting for the expression of thioredoxin 1. Densitometric analysis of thioredoxin 1, as determined by ImageJ, is shown on the right (mean ± SD, n = 3; ** p < 0.01 vs. the control, ## p < 0.01 vs. Px-12 in the siRNA control group).

Article Snippet: The JNK-mitogen-activated protein kinase (MAPK) signaling pathway inhibitor (sp600125, A4604) was obtained from ApexBio Technology China.

Techniques: Inhibition, Incubation, Fluorescence, Microscopy, Western Blot, Expressing