Journal: Oncotarget

Article Title: CCL2-CCR2 axis promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2-MMP2/9 pathway

doi: 10.18632/oncotarget.6695

Figure Lengend Snippet: ( A ) Representative of CCL2 expression and CCR2 expression in NPC tumor tissue and non-tumor tissue detected by IHC. ( B ) Upregulations of CCL2 and CCR2 were observed in different NPC cell lines compared with non-tumor control. NP69 was set as an control. ( C ) Result from ELISA analyses of human sera, the mean serum CCL2 concentration of 50 NPC patients was significantly higher than the mean serum CCL2 concentration of 50 non-NPC patients’. *P < 0.05. ( D ) Kaplan-Meier analysis indicates both upregulation of CCL2 and upregulation of CCR2 were significantly associated with poorer overall survival and distant metastasis-free survival of NPC patients ( p = 0.011, p = 0.039, p = 0.004, p = 0.016, respectively).

Article Snippet: In immunohistochemical (IHC) analysis of CCL2 and CCR2, the paraffin-embedded slices were deparaffinized, rehydrated, and blocked by 5% bovine serum albumin (BSA) at room temperature for 20 minutes, then incubated with rabbit polyclonal antibody against CCL2 (bs-1955R, Bioss) /CCR2 (bs-0562R, Bioss) at a dilution of 1:100 at 4°C overnight, and subsequently incubated with horseradish peroxidase (HRP) anti-rabbit/mouse immunoglobulin at a concentration of 1:100 for 30 minutes at 37°C, then detected the primary antibodies followed by 3, 3-diaminobenzidine substrate visualization and counterstaining with hematoxylin (GTVision III Detection System/Mo & Rb).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Concentration Assay

Journal: Oncotarget

Article Title: CCL2-CCR2 axis promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2-MMP2/9 pathway

doi: 10.18632/oncotarget.6695

Figure Lengend Snippet: Clinicopathological correlation of CCL2 expression and CCR2 expression in NPC

Article Snippet: In immunohistochemical (IHC) analysis of CCL2 and CCR2, the paraffin-embedded slices were deparaffinized, rehydrated, and blocked by 5% bovine serum albumin (BSA) at room temperature for 20 minutes, then incubated with rabbit polyclonal antibody against CCL2 (bs-1955R, Bioss) /CCR2 (bs-0562R, Bioss) at a dilution of 1:100 at 4°C overnight, and subsequently incubated with horseradish peroxidase (HRP) anti-rabbit/mouse immunoglobulin at a concentration of 1:100 for 30 minutes at 37°C, then detected the primary antibodies followed by 3, 3-diaminobenzidine substrate visualization and counterstaining with hematoxylin (GTVision III Detection System/Mo & Rb).

Techniques: Expressing

Journal: Oncotarget

Article Title: CCL2-CCR2 axis promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2-MMP2/9 pathway

doi: 10.18632/oncotarget.6695

Figure Lengend Snippet: Cox proportional hazard regression analyses for 10-year OS and 10-year DMFS

Article Snippet: In immunohistochemical (IHC) analysis of CCL2 and CCR2, the paraffin-embedded slices were deparaffinized, rehydrated, and blocked by 5% bovine serum albumin (BSA) at room temperature for 20 minutes, then incubated with rabbit polyclonal antibody against CCL2 (bs-1955R, Bioss) /CCR2 (bs-0562R, Bioss) at a dilution of 1:100 at 4°C overnight, and subsequently incubated with horseradish peroxidase (HRP) anti-rabbit/mouse immunoglobulin at a concentration of 1:100 for 30 minutes at 37°C, then detected the primary antibodies followed by 3, 3-diaminobenzidine substrate visualization and counterstaining with hematoxylin (GTVision III Detection System/Mo & Rb).

Techniques:

Journal: Oncotarget

Article Title: CCL2-CCR2 axis promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2-MMP2/9 pathway

doi: 10.18632/oncotarget.6695

Figure Lengend Snippet: ( A ) Relatively high expressions of CCL2 and CCR2 were respectively confirmed by Western blotting in CCL2/CCR2- overexpressed S26 and SUNE1 cells compared with vector control cells. ( B ) Cell growth rates between CCL2-, CCR2- and empty vector-transfected cells were compared by XTT assay. ( C ) Representatives and summaries of foci formations in monolayer culture induced by CCL2 and CCR2 and its vector control. ( D ) Representatives of soft agar assays for colony formation induced by CCL2 and CCR2 and its vector control. ( E ) Decreased expressions of CCL2 and CCR2 were respectively confirmed by Western blotting in CCL2/CCR2- silenced S18 and 58F cells compared with scramble shRNA control cells. ( F ) Cell growth rates between CCL2-, CCR2- and scramble shRNA control cells were compared by XTT assay. ( G ) Representatives and summaries of foci formations in monolayer culture induced by CCL2 and CCR2 and its scramble shRNA control cells. ( H ) Representatives of soft agar assays for colony formation induced by CCL2 and CCR2 and its scramble shRNA control cells.

Article Snippet: In immunohistochemical (IHC) analysis of CCL2 and CCR2, the paraffin-embedded slices were deparaffinized, rehydrated, and blocked by 5% bovine serum albumin (BSA) at room temperature for 20 minutes, then incubated with rabbit polyclonal antibody against CCL2 (bs-1955R, Bioss) /CCR2 (bs-0562R, Bioss) at a dilution of 1:100 at 4°C overnight, and subsequently incubated with horseradish peroxidase (HRP) anti-rabbit/mouse immunoglobulin at a concentration of 1:100 for 30 minutes at 37°C, then detected the primary antibodies followed by 3, 3-diaminobenzidine substrate visualization and counterstaining with hematoxylin (GTVision III Detection System/Mo & Rb).

Techniques: Western Blot, Plasmid Preparation, Transfection, XTT Assay, shRNA

Journal: Oncotarget

Article Title: CCL2-CCR2 axis promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2-MMP2/9 pathway

doi: 10.18632/oncotarget.6695

Figure Lengend Snippet: ( A – D ) Overexpression of CCL2 and overexpression of CCR2 respectively promotes the migration and invasion of poorly metastatic NPC cells. Representative pictures and summaries showed that both CCL2 and CCR2 could promote cell migration and cell invasion in S26 and SUNE1 cells compared with vector control cells. * P < 0.05, ** P < 0.01. ( B – E ) Overexpression of CCL2 and overexpression of CCR2 respectively increases distant metastasis in vivo . Representative pictures of lungs derived from mice injected with CCL2-, CCR2- and empty vector-transfected S26 cells and SUNE1 cells. Visible tumor nodules were counted and summarized. * P < 0.05. ( C – F ) H&E staining was performed on pulmonary sections derived from mice. Original magnification: 20 × objective.

Article Snippet: In immunohistochemical (IHC) analysis of CCL2 and CCR2, the paraffin-embedded slices were deparaffinized, rehydrated, and blocked by 5% bovine serum albumin (BSA) at room temperature for 20 minutes, then incubated with rabbit polyclonal antibody against CCL2 (bs-1955R, Bioss) /CCR2 (bs-0562R, Bioss) at a dilution of 1:100 at 4°C overnight, and subsequently incubated with horseradish peroxidase (HRP) anti-rabbit/mouse immunoglobulin at a concentration of 1:100 for 30 minutes at 37°C, then detected the primary antibodies followed by 3, 3-diaminobenzidine substrate visualization and counterstaining with hematoxylin (GTVision III Detection System/Mo & Rb).

Techniques: Over Expression, Migration, Plasmid Preparation, In Vivo, Derivative Assay, Injection, Transfection, Staining

Journal: Oncotarget

Article Title: CCL2-CCR2 axis promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2-MMP2/9 pathway

doi: 10.18632/oncotarget.6695

Figure Lengend Snippet: ( A – D ) Silencing CCL2 and silencing CCR2 respectively inhibits the migration and invasion of highly metastatic NPC cells. Representative pictures and summaries showed that silencing CCL2 and silencing CCR2 respectively could inhibit cell migration and cell invasion in S18 and 58F cells compared with scramble shRNA control cells. * P < 0.05, ** P < 0.01, *** P < 0.001. ( B – E ) Silencing CCL2 and silencing CCR2 respectively decreases distant metastasis in vivo . Representative pictures of lungs derived from mice injected with shCCL2-, shCCR2- and scramble shRNA control S18 cells and 58F cells. Visible tumor nodules were counted and summarized. * P < 0.05, *** P < 0.001. ( C – F ) H & E staining was performed on pulmonary sections derived from mice. Original magnification: 20 × objective.

Article Snippet: In immunohistochemical (IHC) analysis of CCL2 and CCR2, the paraffin-embedded slices were deparaffinized, rehydrated, and blocked by 5% bovine serum albumin (BSA) at room temperature for 20 minutes, then incubated with rabbit polyclonal antibody against CCL2 (bs-1955R, Bioss) /CCR2 (bs-0562R, Bioss) at a dilution of 1:100 at 4°C overnight, and subsequently incubated with horseradish peroxidase (HRP) anti-rabbit/mouse immunoglobulin at a concentration of 1:100 for 30 minutes at 37°C, then detected the primary antibodies followed by 3, 3-diaminobenzidine substrate visualization and counterstaining with hematoxylin (GTVision III Detection System/Mo & Rb).

Techniques: Migration, shRNA, In Vivo, Derivative Assay, Injection, Staining

Journal: Oncotarget

Article Title: CCL2-CCR2 axis promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2-MMP2/9 pathway

doi: 10.18632/oncotarget.6695

Figure Lengend Snippet: ( A ) Overexpression of CCL2/CCR2 axis further facilitates the migration and invasion of poorly metastatic NPC cells. Representative pictures and summaries showed that adding exogenous CCL2 to CCR2-overexpressed cells could intensively promote cell migration and cell invasion in SUNE1 cells compared with vector control cells and individual CCL2/CCR2-overexpressed cells. * P < 0.05, ** P < 0.01, *** P < 0.001. ( B ) The suppressive motility of cell migration and invasion of highly metastatic NPC cells inhibited by silencing CCR2 were failed to be rescued by exogenous CCL2. Representative pictures and summaries showed that adding exogenous CCL2 to CCR2-silenced cells could not promote cell migration and cell invasion in 58F cells compared with scramble shRNA control cells, and individual shCCL2/shCCR2 cells. * P < 0.05, ** P < 0.01, *** P < 0.001. ( C ) Luciferase reporter assay was performed separately in cells of CCL2/CCR2 overexpressed integrally (left) and respectively (right). Remarkable activation of ERK1/2 signaling pathway is observed in CCL2/CCR2 axis integrally upregulated while no transcriptional activity is distinctive when CCL2/CCR2 respectively overexpressed. ( D ) Western blotting showed that five phosphorylated key-proteins of ERK1/2 signaling pathway were increased in CCL2/CCR2 axis overexpressed SUNE1 cells compared with vector control cells and individual CCL2 or CCR2-overexpressed cells (left). Total ERK1/2 was set as an internal control. And only two of the five phosphorylated key-proteins of ERK1/2 signaling pathway were detected in 58F scramble shRNA control cells and all five were under detectable in 58F CCL2/CCR2-silenced cells (right). ( E ) MMP2 and MMP9 were compared between CCL2/CCR2 axis-, individual CCL2/CCR2- and vector-transfected cells in SUNE1 (left), or between shCCL2/CCR2- and shCCR2-cultured with exogenous CCL2, and scramble shRNA control cells (right) in 58F by Western blotting. β-Tubulin was used as a loading control. ( F ) Schematic plot of CCL2/CCR2 axis in promoting metastasis: CCL2 binds CCR2 to form activated CCL2/CCR2 axis, which phosphorylates ERK1/2 and consequently upregulates MMP2/MMP9, then increases cell migration and invasion and eventually promotes tumor metastasis.

Article Snippet: In immunohistochemical (IHC) analysis of CCL2 and CCR2, the paraffin-embedded slices were deparaffinized, rehydrated, and blocked by 5% bovine serum albumin (BSA) at room temperature for 20 minutes, then incubated with rabbit polyclonal antibody against CCL2 (bs-1955R, Bioss) /CCR2 (bs-0562R, Bioss) at a dilution of 1:100 at 4°C overnight, and subsequently incubated with horseradish peroxidase (HRP) anti-rabbit/mouse immunoglobulin at a concentration of 1:100 for 30 minutes at 37°C, then detected the primary antibodies followed by 3, 3-diaminobenzidine substrate visualization and counterstaining with hematoxylin (GTVision III Detection System/Mo & Rb).

Techniques: Over Expression, Migration, Plasmid Preparation, shRNA, Luciferase, Reporter Assay, Activation Assay, Activity Assay, Western Blot, Transfection, Cell Culture

Journal: Cell reports

Article Title: MyD88 signaling in myeloid cells induces gastrointestinal tract injury and systemic inflammation after West Nile virus infection

doi: 10.1016/j.celrep.2025.116852

Figure Lengend Snippet: WT C57BL/6J mice were treated with anti-IFNAR1 mAb plus isotype control or anti-CCR2 mAb (25 μg/mouse) and subcutaneously inoculated with 10 2 FFU of WNV 1 day later. Two additional doses of isotype control or anti-CCR2 mAb were administered at 1 and 3 dpi. At 5 dpi, mice were administered fluorescently conjugated dextrans by oral gavage. (A-B and E-F) Confocal microscopy images of duodenal sections at 5 dpi. (A) WNV antigen (red), monocytes (CCR2, green), EpCAM (white), and nuclei (Hoechst 33258, blue). Scale bars, 100 μm (upper panel) or 50 μm (lower panel). (B) Quantification of CCR2-positive cells in the GI tract, expressed as percentage of total Hoechst 33258-positive cells per field. (C and D) Serum levels of 10 (C) and 250 (D) kDa dextran. (E and F) Quantification of WNV antigen-positive cells in the GI tract, expressed as percentage of total Hoechst 33258-positive cells per field (E) or as percentage of total EpCAM-positive epithelial cells per field (F). (G and H) Hematoxylin and eosin staining of liver sections from mice at 5 dpi. Scale bars, 100 μm (G). (H) Quantitation of liver injury. (I–K) Confocal microscopy images of cerebral cortex sections at 5 dpi showing Iba1 (white) and nuclei (Hoechst 33258, blue). Scale bars, 100 μm (upper panel) or 50 μm (lower panel) (I). (J) Iba1-positive area was quantified from at least 15 microglia/macrophage cells per field and normalized to the number of microglia/macrophage nuclei counted. (K) Numbers of Iba1-positive microglia/macrophages per field. Each data point is derived from at least three independent fields per mouse brain (J–K). (L–O) Cytokine levels of TNF (L and M) and IL-6 (N and O) in serum (L and N) and brain homogenates (M and O). (P–S) WNV RNA levels in the serum (P), spleen (Q), liver (R), and brain (S) at 5 dpi. Bars indicate median values from two experiments; dotted lines show LODs; n = 10 mice (A–F and I–S); n = 6 mice (G–H). Statistical analysis: Mann-Whitney test (ns: non-significant, ** p < 0.01, *** p < 0.001, and **** p < 0.0001).

Article Snippet: Sections were incubated overnight at 4°C with rat anti-WNV hyperimmune serum (1:750 dilution), rabbit anti-EpCAM polyclonal serum (1:2000, Abcam) and goat anti-Iba1 polyclonal serum (1:500, ThermoFisher) or rabbit anti-CCR2 polyclonal serum (1:500, Novus Biologicals) diluted in 1X TBS containing 1% BSA, 3% normal donkey serum, and 0.1% Triton X-100.

Techniques: Control, Confocal Microscopy, Staining, Quantitation Assay, Derivative Assay, MANN-WHITNEY

Journal: PLoS ONE

Article Title: Dexamethasone Palmitate Ameliorates Macrophages-Rich Graft-versus-Host Disease by Inhibiting Macrophage Functions

doi: 10.1371/journal.pone.0096252

Figure Lengend Snippet: A: The viability of mouse macrophage-like RAW264.7 cells after dexamethasone sodium phosphate (DSP) or dexamethasone palmitate (DP) treatment (48 hours) was evaluated by using a colorimetric assay (left panel). The percentage viability was calculated as follows: (O.D value in the presence of each concentration of steroid/O.D value without steroid) ×100. The results reflect the mean ± SD of three independent determinations (representative experiment of three performed). The viability of splenic T lymphocytes after exposure to DSP or DP (25 nM each, 48 hours) was assessed by trypan blue exclusion (right panel). Viable cells were determined as Trypan blue- negative cells. The percent viability was calculated as follows: (viability in DSP or DP group/viability in control group) ×100 (%). The results reflect the mean ± SD of three independent determinations (representative experiment of three performed). B: CCR2 expression on the surface of mouse primary peritoneal macrophages after DSP or DP treatment was evaluated by FACS. The results are representative of three independent experiments (left panel). C: The migration of peritoneal macrophages towards CCL2 after DSP or DP treatment was analyzed using transwell assays. For quantitative analysis, four fields were randomly selected, and migrated cells were counted under a light microscope (×200). The results reflect the mean ± SD of four independent determinations. Representative results of three independent experiments are shown (right panel). Statistical significance: * P <0.05 and ** P <0.01.

Article Snippet: After washings with PBS, the cells were incubated for 30 minutes on ice with a rabbit anti-mouse CCR2 polyclonal antibody (pAb) (1: 25 dilution, E68, Novus Biologicals, Littleton, CO) in the presence of an anti-mouse CD16/32 mAb (BioLegend, San Diego, CA) to reduce non-specific binding of a primary antibody to Fc receptor.

Techniques: Colorimetric Assay, Concentration Assay, Control, Expressing, Migration, Light Microscopy

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: CFA induces mechanical hyperalgesia and alters spinal cord protein expression in rats. (a) Paw withdrawal thresholds (PWTs) of rats in each group on different days ( n = 6). (b) Western blot analysis of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord of vehicle-injected and CFA-injected rats on days −1, 1, 3, 5, 7, 10, and 14 after CFA injection. (c) Quantitative analysis of the relative expressions of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cords of vehicle-injected and CFA-injected rats on days 1, 3, 7, and 14 after CFA injection ( n = 6). * p < 0.05, ** p < 0.01 versus vehicle-injected group.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Expressing, Western Blot, Injection

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: Endogenous expression and cellular localization of CCL2 and CCR2 in the spinal cord of CFA-injected rats. (a) and (b) Double-label immunofluorescence assays were used to detect the colocalization of CCL2 and CCR2 with NeuN, Iba1, and GFAP in CFA-injected rats. White arrows indicate the colocalization of CCL2 and CCR2 with GFAP and NeuN. Scale bar = 100 μm.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Expressing, Injection, Immunofluorescence

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: Intrathecal injection of the AP-1 inhibitor T-5224 attenuates CFA-induced inflammatory pain and suppresses the expression of c-Jun, p-c-Jun, CCL2, and CCR2. (a) Effect of intrathecal injection of T-5224 on PWT ( n = 6). (b) Western blot analysis of c-Jun, p-c-Jun, CCL2, and CCR2 in spinal cords following intrathecal injection. (c) Quantitative analysis of the relative expressions of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord ( n = 6). T-5224 (500 μg/20 μL, i.t.) was administered once daily on days 2, 3, and 4 after CFA injection. PWTs were measured 4 h after each injection. L4-L6 spinal tissues were collected 4 h after the last injection. * p < 0.05, ** p < 0.01 versus vehicle-injected group; # p < 0.05, ## p < 0.01 versus CFA-injected group.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Injection, Expressing, Western Blot

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: Intrathecal injection of the CCR2 antagonist RS 504393 attenuates CFA-induced inflammatory pain and suppresses the expression of CCL2 and CCR2. (a) Effect of intrathecal injection of RS 504393 on PWT ( n = 6). (b) Western blot analysis of c-Jun, p-c-Jun, CCL2, and CCR2 in spinal cords following intrathecal injection. (c) Quantitative analysis of the relative expressions of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord ( n = 6). RS 504393 (20 μg/20 μL, i.t.) was administered once daily on days 2, 3, and 4 after CFA injection. PWTs were measured 4 h after each injection. L4-L6 spinal tissues were collected 4 h after the last injection. * p < 0.05, ** p < 0.01 versus vehicle-injected group; # p < 0.05, ## p < 0.01 versus CFA-injected group.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Injection, Expressing, Western Blot

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: Intrathecal injection of the AP-1 inhibitor T-5224 suppresses the expression of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord. (a) Immunofluorescence analysis of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord following intrathecal injection. (b) Mean fluorescence intensity of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord ( n = 6). * p < 0.05, ** p < 0.01 versus vehicle-injected group; # p < 0.05, ## p < 0.01 versus CFA-injected group.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Injection, Expressing, Immunofluorescence, Fluorescence

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: Intrathecal injection of the CCR2 antagonist RS 504393 suppresses the expression of CCL2 and CCR2 in the spinal cord. (a) Immunofluorescence analysis of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord following intrathecal injection. (b) Mean fluorescence intensity of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord ( n = 6). * p < 0.05, ** p < 0.01 versus vehicle-injected group; # p < 0.05, ## p < 0.01 versus CFA-injected group.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Injection, Expressing, Immunofluorescence, Fluorescence

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: Intrathecal injection of recombinant rat CCL2 protein enhances CFA-induced inflammatory pain and increases the expression of CCL2 and CCR2, whereas T-5224 treatment reverses these effects. (a) Repeated intrathecal injections of recombinant rat CCL2 protein significantly increased mechanical pain in rats compared to the CFA-injected group, while T-5224 treatment reversed this effect ( n = 6). (b) Western blot analysis of c-Jun, p-c-Jun, CCL2, and CCR2 in spinal cords following intrathecal injection. (c) Quantitative analysis of the relative expressions of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord ( n = 6). Recombinant rat CCL2 protein (1 µg/10 µL, i.t.) or T-5224 (500 µg/20 µL, i.t.) was administered once daily on days 2, 3, and 4 after CFA injection. PWTs were measured 4 h after each injection. L4-L6 spinal tissues were collected 4 h after the last injection. * p < 0.05, ** p < 0.01 versus CFA + CCL2 group; # p < 0.05, ## p < 0.01 versus CFA-injected group.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Injection, Recombinant, Expressing, Western Blot

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: Mean fluorescence intensity of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord ( n = 6). * p < 0.05, ** p < 0.01 versus CFA + CCL2; # p < 0.05, ## p < 0.01 versus CFA-injected group.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Fluorescence, Injection

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: Intrathecal injection of curcumin attenuates CFA-induced inflammatory pain and suppresses the expression of c-Jun, p-c-Jun, CCL2, and CCR2. (a) Effect of intrathecal injection of curcumin on paw withdrawal thresholds (PWTs; n = 6). (b) Western blot analysis of c-Jun, p-c-Jun, CCL2, and CCR2 in spinal cords following intrathecal injection. (c) Quantitative analysis of the relative expressions of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord ( n = 6). Curcumin (100 µg/20 µL, 500 µg/20 µL, and 1 mg/20 µL, i.t.) was administered once daily on days 2, 3, and 4 after CFA injection. PWTs were measured 4 h after each injection. L4-L6 spinal tissues were collected 4 h after the last injection. * p < 0.05, ** p < 0.01 versus vehicle-injected group; # p < 0.05, ## p < 0.01 versus CFA-injected group.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Injection, Expressing, Western Blot

Journal: Molecular Pain

Article Title: Curcumin relieves CFA-induced inflammatory pain by inhibiting the AP-1/c-Jun-CCL2-CCR2 pathway in the spinal dorsal horn

doi: 10.1177/17448069251323668

Figure Lengend Snippet: Intrathecal injection of curcumin suppresses the expression of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord. (a) Immunofluorescence analysis of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord following intrathecal injection. (b) Mean fluorescence intensity of c-Jun, p-c-Jun, CCL2, and CCR2 in the spinal cord ( n = 6). * p < 0.05, ** p < 0.01 versus vehicle-injected group; # p < 0.05, ## p < 0.01 versus CFA-injected group.

Article Snippet: Sections were blocked with PBS containing 1% bovine serum albumin for 2 h at room temperature after sectioning and subsequently incubated overnight at 4°C in different antibodies: rabbit anti-c-Jun antibody (9165S, Cell Signaling, 1:100), rabbit anti-p-c-Jun antibody (91952S, Cell Signaling, 1:100), rabbit anti-CCR2 antibody (16153-1-AP, Proteintech, 1:100), rabbit anti-CCL2 antibody (26161-1-AP, Proteintech, 1:500), goat anti-iba1 antibody (microglia marker, ab5076, Abcam, 1: 100), mouse anti-NeuN antibody (neuronal marker, GB11138, Servicebio, 1: 100), and mouse anti-GFAP antibody (astrocyte marker, 3670S, Cell Signaling, 1:100).

Techniques: Injection, Expressing, Immunofluorescence, Fluorescence

Journal: Pharmaceutical Biology

Article Title: Socheongryongtang suppresses COPD-related changes in the pulmonary system through both cytokines and chemokines in a LPS COPD model

doi: 10.1080/13880209.2020.1770808

Figure Lengend Snippet: Primer sequences for RT-PCR.

Article Snippet: Next, the slides were incubated with normal serum so as to block non-specific binding, then incubated for 1 h with primary antibodies (diluted 1:100 to 1:200) such as TNF-α (MyBioSource, San Diego, CA), IFN-γ (sc-74104, Santa Cruz Biotechnology, Dallas, TX), TGF-β (MBS462142, MyBioSource, San Diego, CA), CXCL1 (PAB8798, Abnova, Taipei, Taiwan), CCL-2 (PAB16617, Abnova, Taipei, Taiwan) and CCR2 (Biorbyt, orb137093, Cambridge, UK).

Techniques:

Journal: Pharmaceutical Biology

Article Title: Socheongryongtang suppresses COPD-related changes in the pulmonary system through both cytokines and chemokines in a LPS COPD model

doi: 10.1080/13880209.2020.1770808

Figure Lengend Snippet: Socheongryongtang down-regulated the expression of CXCL1, CCL-2 and CCR2. (A) Socheongryongtang statistically significantly inhibited the DNA level of CXCL1 and trended towards suppressing the levels of CCL-2 and CCR2 . (B) Socheongryongtang significantly suppressed the CXCL1 expression with 150 mg/kg treatment to an extent similar to that by Spiriva treatment. Socheongryongtang down-regulated the expression of CCL2 (C) and CCR2 (D). Each bar represents the mean ± SD ( N = 8). Arrow, positive cells. * p < 0.05 vs. control group; $ p < 0.05 vs. LPS intranasal instillation group; # p < 0.05 vs. Spiriva treatment group. Magnification, ×200.

Article Snippet: Next, the slides were incubated with normal serum so as to block non-specific binding, then incubated for 1 h with primary antibodies (diluted 1:100 to 1:200) such as TNF-α (MyBioSource, San Diego, CA), IFN-γ (sc-74104, Santa Cruz Biotechnology, Dallas, TX), TGF-β (MBS462142, MyBioSource, San Diego, CA), CXCL1 (PAB8798, Abnova, Taipei, Taiwan), CCL-2 (PAB16617, Abnova, Taipei, Taiwan) and CCR2 (Biorbyt, orb137093, Cambridge, UK).

Techniques: Expressing, Control