rabbit anti cleaved caspase3 antibody  (Cell Signaling Technology Inc)

Journal: Drug Design, Development and Therapy

Article Title: Obacunone Promotes Functional Recovery After Spinal Cord Injury by Attenuating Neuroinflammation by Targeting the TLR4/MyD88/p38 MAPK Pathway

doi: 10.2147/DDDT.S577707

Figure Lengend Snippet: Oba attenuated neuroinflammation and reduced cellular apoptosis following SCI. All experiments were performed using tissues from the Sham, SCI, and SCI + Oba (20 mg/kg) groups at 3 days post-SCI. ( A ) Representative Western blot image of iNOS, IL-1β and TNFα at 3 days post-SCI. ( B ) Quantitative analysis of iNOS, IL-1β and TNFα protein levels (n = 5). ( C ) Representative immunofluorescence images of iNOS (green) and DAPI (blue) in spinal cord tissue at 3 days post-SCI; Scale bar = 50 μm. ( D ) Quantitative analysis of iNOS fluorescence areas (n = 5). ( E ) Representative Western blot image of Bax and Cleaved-caspase3 at 3 days post-SCI. ( F ) Quantitative analysis of Bax and Cleaved-caspase3 protein levels (n = 5). ( G ) Representative immunofluorescence images of TUNEL (red) and DAPI (blue) in spinal cord tissue at 3 days post-SCI; Scale bar = 50 μm. ( H ) Quantitative analysis of TUNEL positive cells (n = 5). The values are presented as mean ± SD. *p<0.05, **p<0.01, ***p<0.001; NS, not significant.

Article Snippet: The primary antibodies included rabbit anti-iNOS (13120, Cell signaling technology, 1: 1000), rabbit anti-TNFα (IPB9396, Baijia, 1:1000), rabbit anti-IL-1β (IPB0002, Baijia, 1: 1000), rabbit anti-GAP43 (16971-1-AP, Proteintech, 1:1000), mouse anti- α-tubulin (11224-1-AP, Proteintech, 1:1000), mouse anti-GAPDH (ab307799, Abcam, 1:5000), rabbit anti-Cleaved-caspase3 (AF7022, Affinity, 1: 1000), rabbit anti-Bax (505992-Ig, Proteintech, 1: 1000), rabbit anti-TLR4 (A17436, ABclonal, 1: 1000), rabbit anti-p38 MAPK ( T55600 , ABmart, 1: 1000), rabbit anti-Phospho-p38 MAPK (Thr180/Tyr182) (TA4001, ABmart, 1: 1000), rabbit anti-MyD88 (YM8747, immunoway, 1: 1000).

Techniques: Western Blot, Immunofluorescence, Fluorescence, TUNEL Assay

Journal: Drug Design, Development and Therapy

Article Title: Obacunone Promotes Functional Recovery After Spinal Cord Injury by Attenuating Neuroinflammation by Targeting the TLR4/MyD88/p38 MAPK Pathway

doi: 10.2147/DDDT.S577707

Figure Lengend Snippet: Oba suppresses microglial inflammation and blocks the subsequent apoptosis of HT22 neurons. ( A ) Viability of BV-2 cells treated with different concentrations of Oba, as assessed by CCK-8 assay (n = 5). ( B ) Representative Calcein AM (green)/PI (red) staining images of BV-2 cells following treatment with Oba at the indicated concentrations (0, 25, and 50 μM). Scale bar = 100 μm. ( C ) Quantitative analysis of the ratio of viable cells (n = 5). ( D ) Representative Western blot images of iNOS, TNFα, and IL-1β expression in BV-2 cells. ( E ) Quantitative analysis of iNOS, TNFα and IL-1β protein levels (n = 5). ( F ) Representative immunofluorescence images of iNOS (green) and DAPI (blue) in BV-2 cells treated as follows: vehicle (Control), LPS, LPS+Oba 25 μM, and LPS+Oba 50 μM. Scale bar = 50 μm. ( G ) Quantitative analysis of fluorescence intensity of iNOS (n = 5). ( H ) mRNA expression levels of TNFα and IL-1β in BV-2 cells measured by qPCR (n = 6). ( I ) Representative Western blot image of Bax and Cleaved-caspase3 protein level in HT22 cells. ( J ) Quantitative analysis of Bax and Cleaved-caspase3 protein levels (n = 5). ( K ) Representative flow cytometry plots of HT22 cells stained with Annexin V-FITC (x-axis) and PI (y-axis). ( L ) The percentage of apoptotic HT22 cells, as determined by flow cytometry. (n = 5). The plus and minus signs (+, -) denote the presence or absence of the following reagents in culture: LPS, 25 μM Oba, and 50 μM Oba. The values are presented as mean ± SD. *p<0.05, **p<0.01, ***p<0.001; NS, not significant.

Article Snippet: The primary antibodies included rabbit anti-iNOS (13120, Cell signaling technology, 1: 1000), rabbit anti-TNFα (IPB9396, Baijia, 1:1000), rabbit anti-IL-1β (IPB0002, Baijia, 1: 1000), rabbit anti-GAP43 (16971-1-AP, Proteintech, 1:1000), mouse anti- α-tubulin (11224-1-AP, Proteintech, 1:1000), mouse anti-GAPDH (ab307799, Abcam, 1:5000), rabbit anti-Cleaved-caspase3 (AF7022, Affinity, 1: 1000), rabbit anti-Bax (505992-Ig, Proteintech, 1: 1000), rabbit anti-TLR4 (A17436, ABclonal, 1: 1000), rabbit anti-p38 MAPK ( T55600 , ABmart, 1: 1000), rabbit anti-Phospho-p38 MAPK (Thr180/Tyr182) (TA4001, ABmart, 1: 1000), rabbit anti-MyD88 (YM8747, immunoway, 1: 1000).

Techniques: CCK-8 Assay, Staining, Western Blot, Expressing, Immunofluorescence, Control, Fluorescence, Flow Cytometry