Journal: Nature Communications

Article Title: Paclitaxel drives TREM2 + macrophage expansion underlying its inferior therapeutic efficacy compared to Nab-paclitaxel

doi: 10.1038/s41467-026-69060-5

Figure Lengend Snippet: a Schematic of the experimental setup with THP1 or Raw264.7 cells with indicated treatment. Created in BioRender. Xing, Y. (2026) https://BioRender.com/nsp747p b Quantification of proportion of TREM2 in THP1 and Raw264.7 cells with PTX. n = 3 biological independent samples. c Western blot of THP1 and Raw264.7 cells with indicated treatments. The experiment was independently repeated three times with similar results. d Schematic of the experimental strategy. CM was collected from tumor cells with the indicated treatment. TREM2 expression in macrophages incubated with the CM was assessed. Created in BioRender. Xing, Y. (2026) https://BioRender.com/nsp747p e Quantification of proportion of TREM2 in THP1 incubated with CM of BT549 cells, SUM159 cells and MDA-MB-231 cells treated with PTX, in BMDM incubated with CM of Py8119 treated with PTX, in Raw264.7 incubated with CM from 4T1 and Py8119 cells treated with PTX, respectively. n = 3 biological independent samples. f Western blot analysis of TREM2 and related proteins in THP1, BMDMs, and Raw264.7 cells incubated with CM from BT549, SUM159, Py8119, and 4T1 cells, respectively. The experiment was independently repeated three times with similar results. g Representative immunofluorescence staining of THP1 cells incubated with BT549 CM. n = 3 biological independent samples. h Quantification of proportion of TREM2 in THP1 incubated with CM from BT549, SUM159, and MDA-MB-231 cells treated with Nab-PTX, respectively. n = 3 biological independent samples. i Quantification of the proportion of TREM2 in BMDM incubated with CM of Py8119 treated with Nab-PTX. n = 3 biological independent samples. j Cytokine array analysis of CM from BT549 cells treated indicated treatment. k Western blot analysis of the indicated proteins in Raw264.7 cells and BMDMs treated with recombinant FGF2. The experiment was independently repeated three times with similar results. l Quantification of the proportion of TREM2 in Raw264.7 cells and BMDMs treated with recombinant FGF2. n = 3 biological independent samples. m Schematic of the experimental strategy. CM was collected from tumor cells with the indicated treatment, and then pretreatment using an FGF2 neutralizing antibody. TREM2 expression in macrophages incubated with the CM was assessed. Created in BioRender. Xing, Y. (2026) https://BioRender.com/nsp747p n Western blot analysis of the indicated proteins in Raw264.7 and BMDMs incubated with the indicated CM. The experiment was independently repeated three times with similar results. o Quantification of the proportion of TREM2 in Raw264.7 and BMDMs incubated with the indicated CM. n = 3 biological independent samples. p Representative multiplex immunofluorescence staining of CD68, TREM2 and FGF2 in tumors treated with PTX or Nab-PTX ( n = 3 mice per group). The experiment was independently repeated three times with similar results. Data are shown as means ± S.D. and were analyzed by two-sided unpaired Student’s t test ( b , e , h and i ) and two-sided one-way ANOVA followed by Tukey’s test ( l and o ). Source data are provided as a Source Data file.

Article Snippet: The FGF2 protein (HY-P73052AF) was purchased from MedChemExpress and used with indicated concentration.

Techniques: Western Blot, Expressing, Incubation, Immunofluorescence, Staining, Recombinant, Multiplex Assay

Journal: Nature Communications

Article Title: Paclitaxel drives TREM2 + macrophage expansion underlying its inferior therapeutic efficacy compared to Nab-paclitaxel

doi: 10.1038/s41467-026-69060-5

Figure Lengend Snippet: a Overlap of RNA-seq ( n = 3) and PROMO public database analyses to predict transcription factors regulating TREM2 expression. b Correlations between FGF2 and ATF3 expression in breast cancer using TCGA databases. c qPCR analysis of Atf3 expression in the indicated cells. n = 4 (DMSO and PTX) or 3 (PBS and Nab-PTX) biological independent samples. d Western blot analysis of the indicated proteins in Py8119 cells transfected with ATF3-expressing or control vectors. The experiment was independently repeated three times with similar results. e qPCR analysis of Fgf2 expression in Py8119 cells transfected with Atf3 -expressing or control vectors. n = 3 biological independent samples. f Luciferase activity in HEK293T cells transfected with the indicated reporters and Atf3 -expressing or control vectors. n = 3 biological independent samples. g Abundance of Atf3 bound to the Trem2 promoter in Py8119 cells, as assessed by ChIP-qPCR. n = 3 biological independent samples. h ATAC-seq tracks showing the chromatin accessibility in the ATF3 loci for BT549 cells treated by PTX or Nab-PTX. n = 2 samples per group. Data are shown as means ± S.D. and were analyzed by two-sided unpaired Student’s t test ( c, e and f ) and two-sided two-way ANOVA followed by Šídák’s test ( g ). Source data are provided as a Source Data file.

Article Snippet: The FGF2 protein (HY-P73052AF) was purchased from MedChemExpress and used with indicated concentration.

Techniques: RNA Sequencing, Expressing, Western Blot, Transfection, Control, Luciferase, Activity Assay, ChIP-qPCR

Journal: Nature Communications

Article Title: Paclitaxel drives TREM2 + macrophage expansion underlying its inferior therapeutic efficacy compared to Nab-paclitaxel

doi: 10.1038/s41467-026-69060-5

Figure Lengend Snippet: a Overlap of GeneCards and EDCODE public databases analyses to predict transcription factors that regulate TREM2. b Western blot analysis of the indicated proteins in BMDMs and Raw264.7 cells incubated with FGF2 (left) and with CM from Py8119 cells treated with PTX (right). The experiment was independently repeated three times with similar results. c Western blot analysis of the indicated proteins in Raw264.7 cells incubated with the indicated treatment. The experiment was independently repeated three times with similar results. d qPCR analysis of Trem2 expression in BMDMs transfected with Egr1 -expressing vectors. n = 3 biological independent samples. e Western blot analysis of TREM2 expression in BMDMs transfected with Egr1 -expressing vectors. The experiment was independently repeated three times with similar results. f Luciferase activity of HEK293T cells transfected with the indicated reporters and EGR1 -expressing or control vectors. n = 3 biological independent samples. g Abundance of EGR1 bound to the TREM2 promoter in BMDMs assessed by ChIP-qPCR. n = 3 biological independent samples. h Schematic of the Transwell assay. The first CM was collected from tumor cells with indicated treatment, and the second CM was collected from macrophages incubated with the first CM. The migration and invasion capabilities of macrophages incubated with the first CM were assessed. Created in BioRender. Xing, Y. (2026) https://BioRender.com/nsp747p i Quantification of migration and invasion of Py8119 cells induced by BMDM CM incubated with CM from Py8119 cells treated with PTX, and BT549, SUM159, and MDA-MB-231 cells induced by THP1 CM incubated with CM from BT549, SUM159, and MDA-MB-231 cells treated with PTX (left) or Nab-PTX (right), respectively. n = 5 biological independent samples. j Cytokine array analysis of CM from BMDMs with or without TREM2. k Quantification of migration and invasion of Py8119 cells incubated with indicated proteins. n = 3 biological independent samples. l Quantification of migration and invasion of Py8119 cells induced by CM from BMDMs ( Trem2 +/+ ) with CDE-096. n = 3 biological independent samples. m Western blot analysis of EMT- stimulating proteins in BMDMs incubated with indicated proteins. The experiment was independently repeated three times with similar results. n Schematic illustration showing that FGF2 promotes ERK1/2 phosphorylation to upregulate EGR1, which increases TREM2 expression in macrophages. Upregulated TREM2 enhances the secretion of Serpin E1, HGF, CCL3, and CXCL2 from macrophages to tumor cells, facilitating tumor metastasis via EMT. Created in BioRender. Xing, Y. (2026) https://BioRender.com/nsp747p . Data are shown as means ± S.D. and were analyzed by two-sided unpaired Student’s t test ( d, f, i and l ), two-sided one-way ANOVA followed by Tukey’s test ( k ) and two-sided two-way ANOVA followed by Šídák’s test ( g ). Source data are provided as a Source Data file.

Article Snippet: The FGF2 protein (HY-P73052AF) was purchased from MedChemExpress and used with indicated concentration.

Techniques: Western Blot, Incubation, Expressing, Transfection, Luciferase, Activity Assay, Control, ChIP-qPCR, Transwell Assay, Migration, Phospho-proteomics

Journal: Nature Communications

Article Title: Paclitaxel drives TREM2 + macrophage expansion underlying its inferior therapeutic efficacy compared to Nab-paclitaxel

doi: 10.1038/s41467-026-69060-5

Figure Lengend Snippet: PTX, but not Nab-PTX, promotes lung metastasis by inducing TREM2 + macrophage recruitment. Mechanistically, PTX enhances the ATF3-FGF2 axis in breast cancer cells; secreted FGF2 activates the EGR1–TREM2–EMT cytokine axis in macrophages. Created in BioRender. Xing, Y. (2026) https://BioRender.com/6hxlbow .

Article Snippet: The FGF2 protein (HY-P73052AF) was purchased from MedChemExpress and used with indicated concentration.

Techniques:

Journal: Heliyon

Article Title: Triptolide attenuates cardiac remodeling by inhibiting pyroptosis and EndMT via modulating USP14/Keap1/Nrf2 pathway

doi: 10.1016/j.heliyon.2024.e24010

Figure Lengend Snippet: TP inhibits EndMT in cardiac remodeling models. (A – D) Western blot analysis of CD31, VE-Cadherin and α-SMA protein expression in mouse heart tissue (n = 6). (E – H) Western blot analysis of Slug, Snail and Twist protein expression in mouse heart tissue (n = 6). (I – L) Western blot analysis of CD31, VE-Cadherin and α-SMA in EN (HCAEC) (n = 6). (M – P) Western blot analysis of Slug, Snail and Twist in EN (HCAEC) (n = 6). All the values are presented as mean ± SD. # P < 0.05, ## P < 0.01 compared with Sham or Ctl group; *P < 0.05, **P < 0.01 compared with TAC or TGF-β1 group. The original blots were provided in supplementary data as .

Article Snippet: In our study, recombinant human TGF‐β1 (10 ng/ml, MCE) was added to stimulate the cells for 6d to induce EndMT.

Techniques: Western Blot, Expressing

Journal: Heliyon

Article Title: Triptolide attenuates cardiac remodeling by inhibiting pyroptosis and EndMT via modulating USP14/Keap1/Nrf2 pathway

doi: 10.1016/j.heliyon.2024.e24010

Figure Lengend Snippet: TP inhibits EndMT via Nrf2 pathway. (A – B) Western blot analysis of Nrf2 in EN (HCAEC) ( n = 6). (C – D) Western blot analysis of Nrf2 expression in the presence or absence of Nrf2 siRNA in EN (HCAEC) ( n = 6 ) . (E – H) CD31, VE-Cadherin and α-SMA expression elicited by TGF-β1 in the presence or absence of Nrf2 siRNA in EN (HCAEC) (n = 6) (I – L) Slug, Snail and Twist expression elicited by TGF-β1 in the presence or absence of Nrf2 siRNA in EN (HCAEC) (n = 6). All the values are presented as mean ± SD. # P < 0.05, ## P < 0.01 compared with Ctl group; *P < 0.05, **P < 0.01 compared with siRNA-NC group or TGF-β1 group; & P < 0.05, && P < 0.01 compared with TGF-β1+TP + scRNA group. The original blots were provided in supplementary data as .

Article Snippet: In our study, recombinant human TGF‐β1 (10 ng/ml, MCE) was added to stimulate the cells for 6d to induce EndMT.

Techniques: Western Blot, Expressing

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: Killing efficiency of different physicochemical treatments on recombinant L. lactis strains displaying different binders of proinflammatory cytokines on their surface. The killing efficiency was determined by culturing and counting colonies on agar plates. Complete killing of bacteria i.e., 0 CFU/mL (no bacterial colonies on agar plates) was considered as 100% efficiency. pSD-ZIL6, L. lactis displaying anti-IL6 affibody; pSD-ZTNF, L. lactis displaying anti-TNF affibody; pSD-Fyn17, L. lactis displaying anti-IL17 fynomer; pSD-EVA, L. lactis displaying anti-IL8 evasin; pNZ8148, L. lactis harboring empty plasmid. The results presented are means ± standard deviation (SD) of three technical replicates from a representative experiment.

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Recombinant, Bacteria, Plasmid Preparation, Standard Deviation

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: Cytokine binding ability of recombinant L. lactis bacteria displaying different binders of proinflammatory cytokines on their surface upon exposure to bacteria-killing treatments. ELISA-determined concentrations of recombinant IL6 (A) , TNF (B) , IL17 (C) , and IL8 (D) that remained in the solution following incubation with the corresponding strain of bacteria before and after treatment with heat (70 °C, 40 min or 100 °C, 30 min), sonication, ethanol, UV, and gamma irradiation. pSD-ZIL6, L. lactis displaying ZIL6 affibody; pSD-ZTNF, L. lactis displaying anti-TNF affibody; pSD-Fyn17, L. lactis displaying anti-IL17 fynomer; pSD-EVA, L. lactis displaying anti-IL8 evasin; Ctrl: L. lactis control cells containing empty plasmid pNZ8148. The results are presented as means ± SD of three technical replicates of a representative experiment. ns, p = 0.09; *, p ≤ 0.05; **, p = 0.002; ***, p < 0.001 (unpaired two-tailed t -test).

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Binding Assay, Recombinant, Bacteria, Enzyme-linked Immunosorbent Assay, Incubation, Sonication, Irradiation, Control, Plasmid Preparation, Two Tailed Test

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: Determination of binding affinity of ZIL6-displaying L. lactis for human IL6. A constant number of 8 × 10 6 CFU bacteria was incubated with increasing concentrations of human IL6 for 2 h. MFI values were measured by flow cytometry and binding curves were fitted to the data using the Hill equation in the GraphPad Prism v.10.3.1. The dissociation constants (Kd) values were calculated from the binding curves for non-treated, live ZIL6-displaying L. lactis bacteria and bacteria treated by heat (70 °C, 40 min or 100 °C, 30 min), sonication, ethanol, and UV irradiation. Data are means ± SD of three technical replicates from a representative experiment.

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Binding Assay, Bacteria, Incubation, Flow Cytometry, Sonication, Irradiation

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: Quantification of the maximum binding capacity of ZIL6-displaying L. lactis . Increasing concentrations of recombinant human IL6 (0.45, 4.5, 45 and 450 ng) were incubated with a constant number of bacteria (4.5 × 10 7 CFU equivalent to 0.1 mg dry cell weight) in 450 μl. Residual IL6 in the supernatants was quantified by ELISA for live non-treated bacteria and the cells treated by heat (70 °C, 40 min or 100 °C, 30 min), sonication, ethanol, and UV irradiation. The percentage of bound IL6 was calculated from the difference measured in the presence of ZIL6-displaying L. lactis and control bacteria (carrying empty plasmid pNZ8148). The data are means ± SD of two biological replicates. The asterisks denote statistically significant differences between treated and non-treated bacteria for each concentration. ***, p < 0.001 (one-way ANOVA with Dunnett multiple comparison test).

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Binding Assay, Recombinant, Incubation, Bacteria, Enzyme-linked Immunosorbent Assay, Sonication, Irradiation, Control, Plasmid Preparation, Concentration Assay, Comparison

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: (A) Inhibition of STAT3 signaling in HEKblue-IL6R cells by non-viable ZIL6-displaying L. lactis bacteria in comparison to live non-treated strain. ZIL6-displaying L. lactis cells (1 × 10 8 CFU/mL or 1 × 10 9 CFU/mL) were preincubated with IL6 (1 ng/mL) for 2 h. After removal of bacterial cells, the cell-free supernatant containing remainder of IL6 was added to HEKblue-IL6R cells to induce the reporter system. The percentage of STAT3 inhibition was calculated relative to the STAT3 signaling induced by IL6 in the absence of bacteria. The inhibition of STAT3 signaling was determined for live non-treated bacteria and the cells treated by heat (70 °C, 40 min or 100 °C, 30 min), sonication, ethanol, UV, and gamma irradiation. pSD-ZIL6, L. lactis displaying ZIL6 affibody; Ctrl: L. lactis control cells containing empty plasmid pNZ8148. An anti-IL6 monoclonal antibody (Ab) was used as a positive control. The data are means ± SD of four biological replicates. The asterisks denote statistically significant differences between treated and non-treated bacteria for each concentration. ***, p < 0.001 (one-way ANOVA with Dunnett multiple comparison test). (B) The effect of lactic acid production by L. lactis on viability of HEKblue-IL6R cells. HEKblue-IL6R cells (100 000 cells/well) were incubated with ZIL6-displaying L. lactis (2 × 10 7 bacteria/well) for 6, 12, and 24 h, and the viability of HEKblue-IL6R cells was determined with trypan blue. The data are means ± SD of three technical replicates of a representative experiment.

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Inhibition, Bacteria, Comparison, Sonication, Irradiation, Control, Plasmid Preparation, Positive Control, Concentration Assay, Incubation

Journal:

Article Title:

doi:

Figure Lengend Snippet: Data: mean ± SD. ( A , B ) Intraperitoneal injection of paclitaxel produced mechanical allodynia as evidenced by the increases in paw withdrawal frequencies in response to 0.07 g ( A ) and 0.4 g ( B ) von Frey filaments on the left side. n = 12 mice/time point. * P < 0.05, ** P < 0.01 versus the vehicle-treated group at the corresponding time points by two-way ANOVA with repeated measures followed by post hoc Tukey test. ( C , D ) The levels of Nt3 mRNA ( C ) and NT3 protein ( D ) in L3/4 DRGs on different days as indicated after the first injection of paclitaxel or vehicle. D: representative Western blots (left) and a summary of the densitometric analysis (right) are shown. n = 3 biological repeats (3 mice)/time point. * P < 0.05, ** P < 0.01 by two-way ANOVA followed by post hoc Tukey test. ( E ) The amount of NT3 protein in the L3/4 spinal cord on different days as indicated after the first injection of paclitaxel. n = 3 biological repeated (3 mice)/time point. P > 0.05 by one-way ANOVA followed by post hoc Tukey test. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

Techniques: Injection, Produced, Western Blot

Journal:

Article Title:

doi:

Figure Lengend Snippet: Data: mean ± SD. ( A , B ) Representative images of in situ hybridization histochemistry (ISHH) for Nt3 mRNA (red) and immunohistochemistry for β-tubulin III (green; A ) or glutamine synthetase (GS; green; B ) in the L3/4 DRGs 14 days after the first paclitaxel or vehicle injection. Cellular nuclei were labeled by 4’, 6-diamidino-2- phenylindole (DAPI; blue). n = 9–12 sections from three mice/group. * P < 0.05 by two-tailed unpaired Student t test. Scale bars: 50 µm. ( C ) Size distribution of Nt3 mRNA-labeled neuronal somata in the L3/4 DRGs 14 days after the first paclitaxel injection. Small, 25%, medium, 36%; large, 39%. ( D – H ) Representative images of ISHH for Nt3 mRNA (red) and immunohistochemistry for different DRG cell markers (calcitonin gene-related peptide (CGRP; green; D ), neurofilament 200 (NF200; green; E ), isolectin B4 (IB4; green; F ), or tyrosine hydroxylase (TH; green; G )) in the L3/4 DRGs 14 days after the first paclitaxel injection. Arrows: double-labeling neurons. Statistical summary of number of double-labeling neurons ( H ). n = 9–12 sections from three mice. Scale bar: 50 µm. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

Techniques: In Situ Hybridization, Immunohistochemistry, Injection, Labeling, Two Tailed Test

Journal:

Article Title:

doi:

Figure Lengend Snippet: Data: mean ± SD. ( A ) Expression of NT3 protein in the ipsilateral L3/4 DRGs 14 days after the first injection of paclitaxel (PTX) or vehicle (Veh) from the NT3 siRNA- or NC siRNA-microinjected male mice. n = 3 biological repeats (6 mice)/group. * P < 0.05, ** P < 0.01 by two-way ANOVA followed by post hoc Tukey test. ( B – H ) Paw withdrawal frequency to 0.07 g ( B , F ) and 0.4 g ( C , G ) von Frey filament stimuli and paw withdrawal latency to heat ( D , H ) and cold ( E ) stimuli on the ipsilateral ( B – E ) and contralateral ( F – H ) side on days as indicated from male mice with DRG microinjection of NT3 siRNA or NC siRNA 7 days after the first injection of paclitaxel (PTX) or vehicle. n = 9 mice/group. * P < 0.05, ** P < 0.01 versus the paclitaxel plus NC siRNA-treated group at the corresponding time points by three-way ANOVA with repeated measures followed by post hoc Tukey test. ( I ) Levels of p-ERK1/2, total ERK1/2, and GFAP in the ipsilateral L3/4 dorsal horn 14 days after the first injection of paclitaxel (PTX) or vehicle (Veh) from male mice with DRG microinjection of NT3 siRNA or NC siRNA. n = 3 mice/group. * P < 0.05, ** P < 0.01 by two-way ANOVA followed by Tukey post hoc test. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

Techniques: Expressing, Injection, Microinjection

Journal:

Article Title:

doi:

Figure Lengend Snippet: Data: mean ± SD. ( A ) Expression of NT3 protein in the ipsilateral L3/4 DRGs 14 days after the first injection of paclitaxel (PTX) or vehicle (Veh) from the NT3 siRNA- or NC siRNA-microinjected female mice. n = 3 biological repeats (6 mice)/group. * P < 0.05, ** P < 0.01 by two-way ANOVA followed by post hoc Tukey test. ( B – H ) Paw withdrawal frequency to 0.07 g ( B , F ) and 0.4 g ( C , G ) von Frey filament stimuli and paw withdrawal latency to heat ( D , H ) and cold ( E ) stimuli on the ipsilateral ( B – E ) and contralateral ( F – H ) side on the days as indicated from female mice with DRG microinjection of NT3 siRNA or NC siRNA on day 7 after the first injection of paclitaxel or vehicle. n = 9 mice/group. * P < 0.05, ** P < 0.01 versus the paclitaxel plus NC siRNA-treated group at the corresponding time points by three-way ANOVA with repeated measures followed by post hoc Tukey test. ( I ) Levels of p-ERK1/2, total ERK1/2, and GFAP in the ipsilateral L3/4 dorsal horn 14 days after the first injection of paclitaxel (PTX) or vehicle (Veh) from female mice with DRG microinjection of NT3 siRNA or NC siRNA. n = 3 mice/group. ** P < 0.01 by two-way ANOVA followed by Tukey post hoc test. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

Techniques: Expressing, Injection, Microinjection

Journal:

Article Title:

doi:

Figure Lengend Snippet: Data: mean ± SD. ( A , B ) Levels of Nt3 mRNA ( A ) and NT3 protein ( B ) in the L3/4 DRGs 28 days after the first intraperitoneal (i.p.) injection of paclitaxel (PTX) or vehicle (Veh) from the tamoxifen-treated NT3 f/f mice and NT3 cKD mice. n = 3 biological repeated (3 mice)/group. * P < 0.05, ** P < 0.01 by two-way ANOVA followed by post hoc Tukey test. ( C – I ) Paw withdrawal frequency to 0.07 g ( C , G ) and 0.4 g ( D , H ) von Frey filament stimuli and paw withdrawal latency to heat ( E , I ) and cold ( F ) stimuli on the left ( C – F ) and right ( G – I ) side on days as indicated from NT3 f/f mice and NT3 cKD with i.p. injection of tamoxifen (Tam) daily for 7 consecutive days starting at day 7 after the first i.p. injection of paclitaxel (PTX) or vehicle. n = 9 mice/group. * P < 0.05, ** P < 0.01 versus the paclitaxel-treated NT3 f/f mice at the corresponding days by three-way ANOVA with repeated measures followed by post hoc Tukey test. ( J , K ) Spontaneous ongoing pain as assessed by the CPP test 21 days after the first i.p. injection of paclitaxel (PTX) or vehicle in the tamoxifen-treated NT3 f/f mice and NT3 cKD mice. Time spent in each chamber ( J ) and difference scores for chamber preferences calculated by subtracting preconditioning (Pre) preference time from postconditioning (Post) time spent in the lidocaine-paired chamber ( K ). n = 9 mice/group. ** P < 0.01 by three-way ANOVA with repeated measures followed by post hoc Tukey test ( J ) or by two-way ANOVA followed by Tukey post hoc test ( K ). ( L ) Levels of p-ERK1/2, total ERK1/2 and GFAP in the L3/4 dorsal horn 28 days after the first i.p. injection of paclitaxel (PTX) or vehicle in the tamoxifen-treated NT3 f/f mice and NT3 cKD mice. n = 3 mice/group. * P < 0.05, ** P < 0 . 01 by two-way ANOVA followed by post hoc Tukey test. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

Techniques: Injection

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Figure Lengend Snippet: Data: mean ± SD. ( A , B ) Levels of Nt3 mRNA ( A ) and NT3 protein ( B ) in the ipsilateral L3/4 DRGs 8 weeks after microinjection of AAV5-NT3 or control AAV5-GFP in male mice. n = 3 biological repeats (6 mice)/group. ** P < 0.01 by two-tailed unpaired Student t test. ( C – F ) Paw withdrawal frequency to 0.07 g ( C ) and 0.4 g ( D ) von Frey filament stimuli and paw withdrawal latency to heat ( E ) and cold ( F ) stimuli on the ipsilateral and contralateral sides on weeks as indicated after microinjection of AAV5-NT3 or AAV5-GFP into unilateral L3/4 DRGs. n = 9 mice/group. * P < 0.05, ** P < 0.01 versus the AAV5-GFP group on the ipsilateral side at the corresponding time points by three-way ANOVA with repeated measures followed by Tukey post hoc test. ( G , H ) Spontaneous ongoing pain as assessed by the CPP test 8 weeks after microinjection of AAV5-NT3 or AAV5-GFP into unilateral L3/4 DRGs. Time spent in each chamber ( G ) and difference scores for chamber preferences calculated by subtracting preconditioning (Pre) preference time from postconditioning (Post) time spent in the lidocaine-paired chamber ( H ). n = 9 mice/group. ** P < 0.01 by two-way ANOVA with repeated measures followed by post hoc Tukey test ( G ) or by two-tailed unpaired Student t test ( H ). ( I ) Levels of p-ERK1/2, total ERK1/2 and GFAP in the ipsilateral L3/4 dorsal horn 8 weeks after microinjection of AAV5-NT3 or AAV5-GFP into unilateral L3/4 DRGs. n = 3 mice/group. ** P < 0.01 by two-tailed unpaired Student t test. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

Techniques: Microinjection, Control, Two Tailed Test

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Figure Lengend Snippet: Data: mean ± SD. ( A , B ) Levels of Nt3 mRNA ( A ) and NT3 protein ( B ) in the ipsilateral L3/4 DRGs 8 weeks after microinjection of AAV5-NT3 or control AAV5-GFP in female mice. n = 3 biological repeats (6 mice)/group. ** P < 0.01 by two-tailed unpaired Student t test. ( C – F ) Paw withdrawal frequency to 0.07 g ( C ) and 0.4 g ( D ) von Frey filament stimuli and paw withdrawal latency to heat ( E ) and cold ( F ) stimuli on the ipsilateral and contralateral sides on weeks as indicated after microinjection of AAV5-NT3 or AAV5-GFP into unilateral L3/4 DRGs. n = 9 mice/group. * P < 0.05, ** P < 0.01 versus the AAV5-GFP group on the ipsilateral side at the corresponding time points by three-way ANOVA with repeated measures followed by the Tukey post hoc test. ( G , H ) Spontaneous ongoing pain as assessed by the CPP test 8 weeks after microinjection of AAV5-NT3 or AAV5-GFP into unilateral L3/4 DRGs. Time spent in each chamber ( G ) and difference scores for chamber preferences calculated by subtracting preconditioning (Pre) preference time from postconditioning (Post) time spent in the lidocaine-paired chamber ( H ). n = 9 mice/group. ** P < 0.01 by two-way ANOVA with repeated measures followed by post hoc Tukey test ( G ) or by two-tailed unpaired Student t test ( H ). ( I ) Levels of p-ERK1/2, total ERK1/2 and GFAP in the ipsilateral L3/4 dorsal horn 8 weeks after microinjection of AAV5-NT3 or AAV5-GFP into unilateral L3/4 DRGs. n = 3 mice/group. ** P < 0.01 by two-tailed unpaired Student t test. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

Techniques: Microinjection, Control, Two Tailed Test

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Figure Lengend Snippet: Data: mean ± SD. ( A , B ) Levels of Nt3 , TrkC , and p75 NTR mRNAs ( A ) and their respective proteins ( B ) in the ipsilateral L3/4 DRGs 42 days after microinjection of AAV5-NT3 (NT3) or AAV5-GFP (GFP) into unilateral L3/4 DRGs of the mice with co-microinjection of control scrambled siRNA (Scr-si), TrkC siRNA (TrkC-si), or p75 NTR siRNA (p75-si). n = 3 biological repeats (6 mice)/group. * P < 0.05, ** P < 0.01 by one-way ANOVA followed by post hoc Tukey test. ( C – I ) Paw withdrawal frequencies to 0.07 g ( C , G ) and 0.4 g ( D , H ) von Frey filament stimuli and paw withdrawal latencies to heat ( E, I ) and cold stimuli ( F ) on the ipsilateral ( C – F ) and contralateral ( G – I ) sides on days as indicated after AAV-NT3 or AAV-GFP microinjection in the mice with co-microinjection of control scrambled siRNA (Scr-si), TrkC siRNA (TrkC-si), or p75 NTR siRNA (p75-si) starting at 35 days after AAV5 microinjection. n = 9 mice/group. * P < 0.05, ** P < 0.01 versus the AAV5-NT3 plus control scrambled siRNA-treated group at the corresponding days by two-way ANOVA with repeated measures followed by Tukey post hoc test. ( J ) Levels of p-ERK1/2, total ERK1/2 and GFAP in the ipsilateral L3/4 dorsal horn on day 42 after microinjection of AAV5-NT3 (NT3) or AAV5-GFP (GFP) into unilateral L3/4 DRGs of the mice with co-microinjection of control scrambled siRNA (Scr-si), TrkC siRNA (TrkC-si), or p75 NTR siRNA (p75-si). n = 3 mice/group. * P < 0.05, ** P < 0.01 by one-way ANOVA followed by Tukey post hoc test. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

Techniques: Microinjection, Control

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Figure Lengend Snippet: Data: mean ± SD. Data: mean ± SD. ( A – C ) Levels of CCL2 and TrkC proteins in the ipsilateral L3/4 DRGs 14 days after the first injection of paclitaxel (PTX) or vehicle (Veh) from the NT3 siRNA- or negative control scrambled siRNA (NC siRNA)-microinjected male ( A ) and female ( B ) mice or 28 days after the first injection of paclitaxel or vehicle from the tamoxifen-treated NT3 f/f mice and NT3 cKD mice ( C ). n = 3 biological repeats (6 mice in A and B and 3 mice in C)/group. * P < 0.05, ** P < 0.01 by two-way ANOVA followed by post hoc Tukey test. ( D ) Level of CCL2 protein in the ipsilateral L3/4 DRGs 42 days after microinjection of AAV5-NT3 (NT3) or AAV5-GFP (GFP) into unilateral L3/4 DRGs of the mice with co-microinjection of control scrambled siRNA (Scr-si), or TrkC siRNA (TrkC-si). n = 3 biological repeats (6 mice)/group. * P < 0.05, ** P < 0.01 by one-way ANOVA followed by post hoc Tukey test. ( E – H ) Levels of Nt3 mRNA ( E , G ), Ccl2 mRNA ( E , G ) and their respective proteins ( F , H ) in the cultured DRG neurons (E-F) or in human neuroblastoma cell lines ( G , H ) with transfection of NT3 siRNA or negative control scrambled siRNA (NC siRNA) plus with exposure to paclitaxel (PTX) or vehicle (Veh). n = 3 biological repeats. * P < 0.05, ** P < 0.01 by one-way ANOVA followed by post hoc Tukey test. ( I , J ) Levels of Nt3 mRNA ( I ), Ccl2 mRNA ( I ) and their respective proteins ( J ) in the cultured DRG neurons with transduction of AAV5-NT3 (NT3) or AAV5-GFP (GFP) plus with transfection of NT3 siRNA or negative control scrambled siRNA (NC siRNA). n = 3 biological repeats. * P < 0.05, ** P < 0.01 by one-way ANOVA followed by post hoc Tukey test. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

Techniques: Injection, Negative Control, Microinjection, Control, Cell Culture, Transfection, Transduction

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Figure Lengend Snippet: Co-expression analysis of Nt3 mRNA, TrkC mRNA and Ccl2 mRNA in individual large (>35 μm in diameter; A ), medium (25–35 µm in diameter; B ), and small (<25 μm in diameter; C ) DRG neurons from naive mice ( n = 2) by single-cell RT-PCR assay. NeuN mRNA is used as a neuronal marker. Tuba-1α mRNA is used as a loading control. M DNA ladder marker, NC no cDNA. n = 5 neurons/size. .

Article Snippet: The primary antibodies included rabbit anti-NT3 (1:250; Alomone Labs, catalog number: ANT-003; Jerusalem, Isreael), rabbit anti-MCP-1 (1:1000, catalog number: ab315478; Abcam; Waltham, MA), rabbit anti-p75NTR (1:1000, catalog number: 07-476; Sigma-Aldrich), rabbit anti-TrkC (1:400, catalog number: ANT-020; Alomone), rabbit anti-ERK1/2 (1:1000, catalog number: 4370S; Cell Signaling Technology), rabbit anti-phosphorylated ERK1/2 (p-ERK1/2; 1:1000; catalog number: 3670S; Cell Signaling Technology), mouse anti-GFAP (1:1000, catalog number: 3670S; Cell Signaling Technology), rabbit anti-GAPDH (1:1000, catalog number: G9545; Sigma-Aldrich) and mouse anti-actin (1:1000, catalog number: sc-47778; Santa Cruz).

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