rabbit polyclonal antibody against pdcd4  (Proteintech)

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: Expression and distribution of PDCD4 in spinal cord of CCI mice. (a–b) Behavioral tests of CCI and sham mice, PWL and PWT were lower in CCI mice after operation. (* p < 0.05, ** p < 0.01, *** p < 0.001 compared with the sham group, n = 8 per group). (c) WB showed time-course of PDCD4 in the spinal cord of CCI mice. (*** p < 0.001; * p < 0.05 compared with the sham group, n = 6 per group). (d) Distribution of PDCD4 in the spinal cord dorsal horn following CCI. PDCD4 was colocalized mostly with neurons (NeuN), and a minority with microglia (IBA1), but not astrocytes (GFAP) in the spinal cord dorsal horn. Tissues were collected at day 14 after CCI ( n = 3 per group, scale bar = 100 μm).

Article Snippet: The membranes were blocked with 5% non-fat milk or BSA at room temperature for 2 h followed by overnight incubation with the following primary antibodies at 4°C: rabbit polyclonal anti-PDCD4 antibody (1:500, 9535S, CST, USA), Beclin1 (1:200, HA721216, Huabio, China), p62 (1:500, HA72117, Huabio, China), and glyceraldehyde-3-phosphate dehydrogenase (1:10000, GAPDH, G9545, Sigma, USA), respectively.

Techniques: Expressing

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: Effects of intrathecal injection of AAV encoded shPDCD4 (AAV-shPDCD4) on the expression of PDCD4 protein. (a) Schematics of AAV constructs shPDCD4 or control. CMV, cytomegalovirus promoter; EGFP, enhanced GFP; WPRE, woodchuck hepatitis virus posttranscriptional regulatory element; ITR, inverted terminal repeats. (b) EGFP immunoreactivity was detected in the superficial laminae of the dorsal horn at 4 weeks after AAV-shPDCD4 (5.0 × 10 12 v.g/mL) injection (scale bar = 100 μm). (c) Representative bands for the expression of PDCD4 in the spinal cord of mice at 4 weeks after intrathecal injection of AAV-shPDCD4 or AAV-control, CCI models were performed 2 weeks after AAV injection (** p < 0.01 compared with the indicated group, n = 4 per group).

Article Snippet: The membranes were blocked with 5% non-fat milk or BSA at room temperature for 2 h followed by overnight incubation with the following primary antibodies at 4°C: rabbit polyclonal anti-PDCD4 antibody (1:500, 9535S, CST, USA), Beclin1 (1:200, HA721216, Huabio, China), p62 (1:500, HA72117, Huabio, China), and glyceraldehyde-3-phosphate dehydrogenase (1:10000, GAPDH, G9545, Sigma, USA), respectively.

Techniques: Injection, Expressing, Construct, Control, Virus

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: Effects of PDCD4 inhibition on existing pain hyperalgesia after CCI. (a) Schematic timeline for drug administration, behavioral detection, and tissue extraction. (b–c) Analgesic effects of i.t. AAV-shPDCD4 on thermal hyperalgesia (PWL) and mechanical allodynia (PWT) in ipsilateral to CCI. (*** p < 0.001, ** p < 0.01 vs sham+AAV-control group. # p < 0.05, ### p < 0.001 vs CCI+AAV-control, n = 8 per group).

Article Snippet: The membranes were blocked with 5% non-fat milk or BSA at room temperature for 2 h followed by overnight incubation with the following primary antibodies at 4°C: rabbit polyclonal anti-PDCD4 antibody (1:500, 9535S, CST, USA), Beclin1 (1:200, HA721216, Huabio, China), p62 (1:500, HA72117, Huabio, China), and glyceraldehyde-3-phosphate dehydrogenase (1:10000, GAPDH, G9545, Sigma, USA), respectively.

Techniques: Inhibition, Extraction, Control

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: The effects of PDCD4 inhibition proinflammatory factors. (a–c) ELISA data showed the expression of proinflammatory cytokines in the L4-6 spinal cord were downregulated by intrathecal injection of AAV-shPDCD4 after CCI (** p < 0.01, *** p < 0.001 compared with the sham group, ## p < 0.01 compared with CCI+AAV-control group, n = 4 per group). (d–f) qRT-PCR results of proinflammatory cytokines in the L4-6 spinal cord were downregulated by intrathecal injection of AAV-shPDCD4 after CCI (*** p < 0.001 compared with the sham group, ## p < 0.01, ### p < 0.001 compared with CCI+AAV-control group, n = 4 per group).

Article Snippet: The membranes were blocked with 5% non-fat milk or BSA at room temperature for 2 h followed by overnight incubation with the following primary antibodies at 4°C: rabbit polyclonal anti-PDCD4 antibody (1:500, 9535S, CST, USA), Beclin1 (1:200, HA721216, Huabio, China), p62 (1:500, HA72117, Huabio, China), and glyceraldehyde-3-phosphate dehydrogenase (1:10000, GAPDH, G9545, Sigma, USA), respectively.

Techniques: Inhibition, Enzyme-linked Immunosorbent Assay, Expressing, Injection, Control, Quantitative RT-PCR

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: The effects of PDCD4 inhibition on autophagy. (a–c) WB results showed expression of Beclin1, LC3B and p62 were reversed after AAV-shPDCD4 intrathecal injection in CCI mice. (* p < 0.05, ** p < 0.01, *** p < 0.001, *** p < 0.0001 compared with CCI+AAV-control; ## p < 0.01 compared with sham+AAV-control, n = 4 per group).

Article Snippet: The membranes were blocked with 5% non-fat milk or BSA at room temperature for 2 h followed by overnight incubation with the following primary antibodies at 4°C: rabbit polyclonal anti-PDCD4 antibody (1:500, 9535S, CST, USA), Beclin1 (1:200, HA721216, Huabio, China), p62 (1:500, HA72117, Huabio, China), and glyceraldehyde-3-phosphate dehydrogenase (1:10000, GAPDH, G9545, Sigma, USA), respectively.

Techniques: Inhibition, Expressing, Injection, Control

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: The effects of PDCD4 inhibition on MAPK activation. (a): Immunofluorescence showed the expression of p-ERK, p-JNK, and p-p38MAPK were inhibited by intrathecal injection of AAV-shPDCD4 in the L4-6 spinal cord ( n = 4 per group, scale bar = 100 μm). (b–e): WB showing the expression of p-ERK, p-JNK, and p-p38MAPK were inhibited by intrathecal injection of AAV-shPDCD4 in the L4-6 spinal cord (* p < 0.05, ** p < 0.01, compared with CCI+AAV-control; ### p < 0.001 compared with sham+AAV-control, n = 4 per group).

Article Snippet: The membranes were blocked with 5% non-fat milk or BSA at room temperature for 2 h followed by overnight incubation with the following primary antibodies at 4°C: rabbit polyclonal anti-PDCD4 antibody (1:500, 9535S, CST, USA), Beclin1 (1:200, HA721216, Huabio, China), p62 (1:500, HA72117, Huabio, China), and glyceraldehyde-3-phosphate dehydrogenase (1:10000, GAPDH, G9545, Sigma, USA), respectively.

Techniques: Inhibition, Activation Assay, Immunofluorescence, Expressing, Injection, Control

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: Effect of the autophagy inhibitor 3-MA on pain-related behaviors, autophagy and inflammatory cytokines following AAV-shPDCD4 injection. (a) A schematic timeline for drug administration, behavioral detection, and tissue extraction. (b–c) Intrathecal injection of autophagy inhibitor 3-MA markedly reversed the analgesic effect of PDCD4 inhibition (** p < 0.01, *** p < 0.001 compared with the CCI+AAV-shPDCD4 group, n = 8 per group). (d–f) Intrathecal injection of autophagy inhibitor 3-MA suppressed the autophagy activation induced by PDCD4 inhibition (** p < 0.01, *** p < 0.001 compared with the CCI+AAV-shPDCD4 group, n = 4 per group). (g–i) ELISA data showed intrathecal injection of autophagy inhibitor 3-MA markedly reversed the suppression of pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) by PDCD4 inhibition. (** p < 0.01, *** p < 0.001 compared with the CCI+AAV-shPDCD4 group, n = 4 per group). (j–l) qRT-PCR results showed intrathecal injection of autophagy inhibitor 3-MA markedly reversed the suppression of pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) by PDCD4 inhibition. (** p < 0.01, *** p < 0.001 compared with the CCI+AAV-shPDCD4 group, n = 4 per group).

Article Snippet: The membranes were blocked with 5% non-fat milk or BSA at room temperature for 2 h followed by overnight incubation with the following primary antibodies at 4°C: rabbit polyclonal anti-PDCD4 antibody (1:500, 9535S, CST, USA), Beclin1 (1:200, HA721216, Huabio, China), p62 (1:500, HA72117, Huabio, China), and glyceraldehyde-3-phosphate dehydrogenase (1:10000, GAPDH, G9545, Sigma, USA), respectively.

Techniques: Injection, Extraction, Inhibition, Activation Assay, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: Expression and distribution of PDCD4 in spinal cord of CCI mice. (a–b) Behavioral tests of CCI and sham mice, PWL and PWT were lower in CCI mice after operation. (* p < 0.05, ** p < 0.01, *** p < 0.001 compared with the sham group, n = 8 per group). (c) WB showed time-course of PDCD4 in the spinal cord of CCI mice. (*** p < 0.001; * p < 0.05 compared with the sham group, n = 6 per group). (d) Distribution of PDCD4 in the spinal cord dorsal horn following CCI. PDCD4 was colocalized mostly with neurons (NeuN), and a minority with microglia (IBA1), but not astrocytes (GFAP) in the spinal cord dorsal horn. Tissues were collected at day 14 after CCI ( n = 3 per group, scale bar = 100 μm).

Article Snippet: The sections were first blocked with 5% donkey serum in 0.3% Triton X-100 for 1 h at room temperature and then incubated overnight at 4°C with the following primary antibodies: rabbit anti-PDCD4 polyclonal antibody (1:500, 9535S, CST, USA), c-Fos rabbit mAb (1:500, #2250, CST, USA), rabbit anti-p-ERK polyclonal antibody (1:100, 28733-1-AP, Huabio), rabbit anti-p-JNK polyclonal antibody (1:400, 80024-1-RR, Huabio), rabbit anti-p-p38MAPK polyclonal antibody (1:200, 28796-1-AP, Huabio).

Techniques: Expressing

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: Effects of intrathecal injection of AAV encoded shPDCD4 (AAV-shPDCD4) on the expression of PDCD4 protein. (a) Schematics of AAV constructs shPDCD4 or control. CMV, cytomegalovirus promoter; EGFP, enhanced GFP; WPRE, woodchuck hepatitis virus posttranscriptional regulatory element; ITR, inverted terminal repeats. (b) EGFP immunoreactivity was detected in the superficial laminae of the dorsal horn at 4 weeks after AAV-shPDCD4 (5.0 × 10 12 v.g/mL) injection (scale bar = 100 μm). (c) Representative bands for the expression of PDCD4 in the spinal cord of mice at 4 weeks after intrathecal injection of AAV-shPDCD4 or AAV-control, CCI models were performed 2 weeks after AAV injection (** p < 0.01 compared with the indicated group, n = 4 per group).

Article Snippet: The sections were first blocked with 5% donkey serum in 0.3% Triton X-100 for 1 h at room temperature and then incubated overnight at 4°C with the following primary antibodies: rabbit anti-PDCD4 polyclonal antibody (1:500, 9535S, CST, USA), c-Fos rabbit mAb (1:500, #2250, CST, USA), rabbit anti-p-ERK polyclonal antibody (1:100, 28733-1-AP, Huabio), rabbit anti-p-JNK polyclonal antibody (1:400, 80024-1-RR, Huabio), rabbit anti-p-p38MAPK polyclonal antibody (1:200, 28796-1-AP, Huabio).

Techniques: Injection, Expressing, Construct, Control, Virus

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: Effects of PDCD4 inhibition on existing pain hyperalgesia after CCI. (a) Schematic timeline for drug administration, behavioral detection, and tissue extraction. (b–c) Analgesic effects of i.t. AAV-shPDCD4 on thermal hyperalgesia (PWL) and mechanical allodynia (PWT) in ipsilateral to CCI. (*** p < 0.001, ** p < 0.01 vs sham+AAV-control group. # p < 0.05, ### p < 0.001 vs CCI+AAV-control, n = 8 per group).

Article Snippet: The sections were first blocked with 5% donkey serum in 0.3% Triton X-100 for 1 h at room temperature and then incubated overnight at 4°C with the following primary antibodies: rabbit anti-PDCD4 polyclonal antibody (1:500, 9535S, CST, USA), c-Fos rabbit mAb (1:500, #2250, CST, USA), rabbit anti-p-ERK polyclonal antibody (1:100, 28733-1-AP, Huabio), rabbit anti-p-JNK polyclonal antibody (1:400, 80024-1-RR, Huabio), rabbit anti-p-p38MAPK polyclonal antibody (1:200, 28796-1-AP, Huabio).

Techniques: Inhibition, Extraction, Control

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: The effects of PDCD4 inhibition proinflammatory factors. (a–c) ELISA data showed the expression of proinflammatory cytokines in the L4-6 spinal cord were downregulated by intrathecal injection of AAV-shPDCD4 after CCI (** p < 0.01, *** p < 0.001 compared with the sham group, ## p < 0.01 compared with CCI+AAV-control group, n = 4 per group). (d–f) qRT-PCR results of proinflammatory cytokines in the L4-6 spinal cord were downregulated by intrathecal injection of AAV-shPDCD4 after CCI (*** p < 0.001 compared with the sham group, ## p < 0.01, ### p < 0.001 compared with CCI+AAV-control group, n = 4 per group).

Article Snippet: The sections were first blocked with 5% donkey serum in 0.3% Triton X-100 for 1 h at room temperature and then incubated overnight at 4°C with the following primary antibodies: rabbit anti-PDCD4 polyclonal antibody (1:500, 9535S, CST, USA), c-Fos rabbit mAb (1:500, #2250, CST, USA), rabbit anti-p-ERK polyclonal antibody (1:100, 28733-1-AP, Huabio), rabbit anti-p-JNK polyclonal antibody (1:400, 80024-1-RR, Huabio), rabbit anti-p-p38MAPK polyclonal antibody (1:200, 28796-1-AP, Huabio).

Techniques: Inhibition, Enzyme-linked Immunosorbent Assay, Expressing, Injection, Control, Quantitative RT-PCR

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: The effects of PDCD4 inhibition on autophagy. (a–c) WB results showed expression of Beclin1, LC3B and p62 were reversed after AAV-shPDCD4 intrathecal injection in CCI mice. (* p < 0.05, ** p < 0.01, *** p < 0.001, *** p < 0.0001 compared with CCI+AAV-control; ## p < 0.01 compared with sham+AAV-control, n = 4 per group).

Article Snippet: The sections were first blocked with 5% donkey serum in 0.3% Triton X-100 for 1 h at room temperature and then incubated overnight at 4°C with the following primary antibodies: rabbit anti-PDCD4 polyclonal antibody (1:500, 9535S, CST, USA), c-Fos rabbit mAb (1:500, #2250, CST, USA), rabbit anti-p-ERK polyclonal antibody (1:100, 28733-1-AP, Huabio), rabbit anti-p-JNK polyclonal antibody (1:400, 80024-1-RR, Huabio), rabbit anti-p-p38MAPK polyclonal antibody (1:200, 28796-1-AP, Huabio).

Techniques: Inhibition, Expressing, Injection, Control

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: The effects of PDCD4 inhibition on MAPK activation. (a): Immunofluorescence showed the expression of p-ERK, p-JNK, and p-p38MAPK were inhibited by intrathecal injection of AAV-shPDCD4 in the L4-6 spinal cord ( n = 4 per group, scale bar = 100 μm). (b–e): WB showing the expression of p-ERK, p-JNK, and p-p38MAPK were inhibited by intrathecal injection of AAV-shPDCD4 in the L4-6 spinal cord (* p < 0.05, ** p < 0.01, compared with CCI+AAV-control; ### p < 0.001 compared with sham+AAV-control, n = 4 per group).

Article Snippet: The sections were first blocked with 5% donkey serum in 0.3% Triton X-100 for 1 h at room temperature and then incubated overnight at 4°C with the following primary antibodies: rabbit anti-PDCD4 polyclonal antibody (1:500, 9535S, CST, USA), c-Fos rabbit mAb (1:500, #2250, CST, USA), rabbit anti-p-ERK polyclonal antibody (1:100, 28733-1-AP, Huabio), rabbit anti-p-JNK polyclonal antibody (1:400, 80024-1-RR, Huabio), rabbit anti-p-p38MAPK polyclonal antibody (1:200, 28796-1-AP, Huabio).

Techniques: Inhibition, Activation Assay, Immunofluorescence, Expressing, Injection, Control

Journal: Molecular Pain

Article Title: PDCD4 inhibition alleviates neuropathic pain by regulating spinal autophagy and neuroinflammation

doi: 10.1177/17448069251333928

Figure Lengend Snippet: Effect of the autophagy inhibitor 3-MA on pain-related behaviors, autophagy and inflammatory cytokines following AAV-shPDCD4 injection. (a) A schematic timeline for drug administration, behavioral detection, and tissue extraction. (b–c) Intrathecal injection of autophagy inhibitor 3-MA markedly reversed the analgesic effect of PDCD4 inhibition (** p < 0.01, *** p < 0.001 compared with the CCI+AAV-shPDCD4 group, n = 8 per group). (d–f) Intrathecal injection of autophagy inhibitor 3-MA suppressed the autophagy activation induced by PDCD4 inhibition (** p < 0.01, *** p < 0.001 compared with the CCI+AAV-shPDCD4 group, n = 4 per group). (g–i) ELISA data showed intrathecal injection of autophagy inhibitor 3-MA markedly reversed the suppression of pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) by PDCD4 inhibition. (** p < 0.01, *** p < 0.001 compared with the CCI+AAV-shPDCD4 group, n = 4 per group). (j–l) qRT-PCR results showed intrathecal injection of autophagy inhibitor 3-MA markedly reversed the suppression of pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) by PDCD4 inhibition. (** p < 0.01, *** p < 0.001 compared with the CCI+AAV-shPDCD4 group, n = 4 per group).

Article Snippet: The sections were first blocked with 5% donkey serum in 0.3% Triton X-100 for 1 h at room temperature and then incubated overnight at 4°C with the following primary antibodies: rabbit anti-PDCD4 polyclonal antibody (1:500, 9535S, CST, USA), c-Fos rabbit mAb (1:500, #2250, CST, USA), rabbit anti-p-ERK polyclonal antibody (1:100, 28733-1-AP, Huabio), rabbit anti-p-JNK polyclonal antibody (1:400, 80024-1-RR, Huabio), rabbit anti-p-p38MAPK polyclonal antibody (1:200, 28796-1-AP, Huabio).

Techniques: Injection, Extraction, Inhibition, Activation Assay, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR

Journal: Journal of Clinical Laboratory Analysis

Article Title: Generation of MiRNA sponge constructs targeting multiple MiRNAs

doi: 10.1002/jcla.24527

Figure Lengend Snippet: Primers of quantitative real‐time PCR

Article Snippet: The primary antibodies included PDCD4 (1:1000, Anti‐PDCD4 rabbit polyclonal antibody; Sangon Biotech BBI, CHINA), GAPDH (1:1000, rabbit pAb to GAPDH; Abcam,).

Techniques:

Journal: Journal of Clinical Laboratory Analysis

Article Title: Generation of MiRNA sponge constructs targeting multiple MiRNAs

doi: 10.1002/jcla.24527

Figure Lengend Snippet: Expression levels of hsa‐miR‐21‐5p, hsa‐miR‐31‐5p and hsa‐miR‐155‐5p in lung cancer cells lines and BEAS‐2B cells and the expression levels of genes targeted by hsa‐miR‐21‐5p, hsa‐miR‐31‐5p and hsa‐miR‐155‐5p. (A) The relative expression levels of hsa‐miR‐21‐5p in lung cancer cells lines and BEAS‐2B cells which is derived from normal human bronchial epithelium. (B) The relative expression levels of hsa‐miR‐31‐5p in lung cancer cells lines and BEAS‐2B cells which is derived from normal human bronchial epithelium. (C)The relative expression levels of hsa‐miR‐155‐5p in lung cancer cells lines and BEAS‐2B cells which is derived from normal human bronchial epithelium. (D) The relative mRNA level of RhoBTB1 in CRL‐5810 cells with 16xmiR‐31 sponges and 16xmiR‐31&16xmiR‐155 sponges. (E) The relative mRNA level of SGK3 in CRL‐5810 cells with 16xmiR‐155 sponges and 16xmiR‐31&16xmiR‐155 sponges. (F)The relative mRNA level of RACK1 in CRL‐5810 cells with 16xmiR‐155 sponges and 16xmiR‐31&16xmiR‐155 sponges. (G)The relative mRNA level of PTEN in A549 cells with 16xmiR‐21 sponges and 16xmiR‐21&16xmiR‐31 sponges. (H)The relative mRNA level of PDCD4 in A549 cells with 16xmiR‐21 sponges and 16xmiR‐21&16xmiR‐31 sponges. (I)The relative mRNA level of ARID1A in A549 cells with 16xmiR‐31 sponges and 16xmiR‐21&16xmiR‐31 sponges

Article Snippet: The primary antibodies included PDCD4 (1:1000, Anti‐PDCD4 rabbit polyclonal antibody; Sangon Biotech BBI, CHINA), GAPDH (1:1000, rabbit pAb to GAPDH; Abcam,).

Techniques: Expressing, Derivative Assay

Journal: Journal of Clinical Laboratory Analysis

Article Title: Generation of MiRNA sponge constructs targeting multiple MiRNAs

doi: 10.1002/jcla.24527

Figure Lengend Snippet: Cell growth of lung cancer cells infected with miRNA sponges or inhibitors. (A)The relative protein levels of PDCD4 in A549 cells with 16xmiR‐21 sponges and 16xmiR‐21&16xmiR‐31 sponges. (B)The relative mRNA level of TIAM1 in A549 cells with 16xmiR‐21 sponges 、16xmiR‐31 sponges and 16xmiR‐21&16xmiR‐31 sponges. (C) The cell growth of CRL‐5810 cells infected with miR‐31 or miR‐155 sponges separately or both. (D)The cell growth of CRL‐5810 cells infected with miR‐31 or miR‐155 inhibitors separately or both. (E)The cell growth of A549 cells infected with miR‐21 or miR‐31 sponges separately or both. (F)The cell growth of A549 cells infected with miR‐21 or miR‐31 inhibitors separately or both

Article Snippet: The primary antibodies included PDCD4 (1:1000, Anti‐PDCD4 rabbit polyclonal antibody; Sangon Biotech BBI, CHINA), GAPDH (1:1000, rabbit pAb to GAPDH; Abcam,).

Techniques: Infection

Journal: Frontiers in Immunology

Article Title: Elevated microRNA-21 Is a Brake of Inflammation Involved in the Development of Nasal Polyps

doi: 10.3389/fimmu.2021.530488

Figure Lengend Snippet: The primer sequences used in qPCR.

Article Snippet: Sections were then treated with 3% hydrogen peroxide at room temperature for 10min and incubated with rabbit polyclonal antibody against PDCD4 (Proteintech, Wuhan, China) at 4°C overnight.

Techniques:

Journal: Frontiers in Immunology

Article Title: Elevated microRNA-21 Is a Brake of Inflammation Involved in the Development of Nasal Polyps

doi: 10.3389/fimmu.2021.530488

Figure Lengend Snippet: Expression of PDCD4 in patients with CRSwNP. (A) IHC staining of PDCD4 was performed in normal tissues and NPs. (B) Comparison of PDCD4 expression were assessed through mean optical density (MOD) between control subjects (n=6) and NPs (n=20). (C) PDCD4 protein expression was determined by WB in control subjects (n=6) and CRSwNP (n=6), with β-actin expression as a control. (D) Relative PDCD4 protein expression was quantified by densitometry based on immunoblot images. (E) Relative PDCD4 mRNA expression was measured by using qPCR between control subjects (n=15) and CRSwNP (n=43). (F) Correlations between miR-21 and PDCD4 in CRSwNP, R values indicate Spearman correlation coefficients. Mann–Whitney U test was used for comparisons between control subjects and CRSwNP. The asterisk indicates statistical significance, *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: Sections were then treated with 3% hydrogen peroxide at room temperature for 10min and incubated with rabbit polyclonal antibody against PDCD4 (Proteintech, Wuhan, China) at 4°C overnight.

Techniques: Expressing, Immunohistochemistry, Comparison, Control, Western Blot, MANN-WHITNEY

Journal: Frontiers in Immunology

Article Title: Elevated microRNA-21 Is a Brake of Inflammation Involved in the Development of Nasal Polyps

doi: 10.3389/fimmu.2021.530488

Figure Lengend Snippet: Effect of miR-21 on the expression of PDCD4 in vitro . (A) HNEpC was transfected with miR-21 mimics (with mNC as control) and inhibitor (with iNC as control) for 24h. PDCD4 protein expression was determined by WB, normalized to β-actin. (B) Relative PDCD4 protein expression was quantified by densitometry based on immunoblot images. (C) PDCD4 mRNA levels were measured by qPCR. (D) The predicted miR-21 binding sites within the 3′UTR of PDCD4 mRNA. (E) Double luciferase activity assay of HEK 293 cells. After being co-transfected with the analogue of miR-21: control/miR-21 mimic/miR-21 inhibitor, and the following plasmids: pGL3-3′-UTR of control/WT/mutated PDCD4 vector and the pRL-TK vector, the ratio of GFP to RL was determined. Data were obtained in three independent experiments. One-way ANOVA was used to analyze the difference between multiple groups. The asterisk indicates statistical significance, *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: Sections were then treated with 3% hydrogen peroxide at room temperature for 10min and incubated with rabbit polyclonal antibody against PDCD4 (Proteintech, Wuhan, China) at 4°C overnight.

Techniques: Expressing, In Vitro, Transfection, Control, Western Blot, Binding Assay, Luciferase, Activity Assay, Plasmid Preparation

Journal: Frontiers in Immunology

Article Title: Elevated microRNA-21 Is a Brake of Inflammation Involved in the Development of Nasal Polyps

doi: 10.3389/fimmu.2021.530488

Figure Lengend Snippet: Schematic summary of the anti-inflammatory role of miR-21. In nasal epithelial cells, SEB could promote the expression of IL-25, IL-33, and TSLP, while LPS could enhance the expression of IL-1β, IL-6, and IL-8, both by activating NF-κB. However, both SEB and LPS could upregulate miR-21 expression. Elevated miR-21 would decrease PDCD4 expression to suppress the expression of cytokines in HNEpC by inducing the IL-10 production and inhibiting the activation of NF-κB.

Article Snippet: Sections were then treated with 3% hydrogen peroxide at room temperature for 10min and incubated with rabbit polyclonal antibody against PDCD4 (Proteintech, Wuhan, China) at 4°C overnight.

Techniques: Expressing, Activation Assay