Journal: Frontiers in Immunology

Article Title: Low-intensity pulsed ultrasound ameliorates partial infraorbital nerve ligation-induced trigeminal neuropathic pain through inhibiting Schwann cell Pannexin 1 channel

doi: 10.3389/fimmu.2025.1712759

Figure Lengend Snippet: LIPUS treatment attenuates partial infraorbital nerve ligation (pIONL)-induced trigeminal neuropathic pain (TNP) and inflammation in infraorbital nerve (ION). (A) Duration and application schedule of LIPUS therapy. (B) Pain response measurements using 0.02g and 0.16g von Frey filaments. Statistical analysis performed via mixed-effects (Group×Time), primary ΔAUC {0–21 d} (95% CI) (n=5 per group), with results presented as mean ± SD. (C) Timeline of LIPUS intervention. (D) Quantitative PCR assessment of Panx 1, F4/80, TNF-α, and IL-1β mRNA levels in the infraorbital nerve three days post-pIONL. Student’s t-test (n=5 per group); data shown as mean ± SD. (E) Immunofluorescence staining of Panx 1 and S100β in the infraorbital nerve. Scale bar: 20μm. Analysis included at least three sections from three animals per group. Student’s t-test (n=3 per group); data expressed as mean ± SD. * p<0.05, ** p<0.01, *** p<0.001 (LIPUS vs. pIONL).

Article Snippet: The sections were incubated with following primary antibodies overnight at 4°C: S100β antibody specifically labels Schwann cells, as validated in previous studies , rabbit anti-Iba1 (1:1000, 19741, Wako, Osaka, Japan), mouse anti-S100β (1:500, bsm-10832M, Bioss), and rabbit anti-Panx 1 (1:200, A00915-2, Boster, China), rabbit anti-IL-1β (1: 200, A00101, Boster, China), rabbit anti-TNF-α (1: 400, BA0131, Boster, China), rabbit anti-P2X7 (1:200, 28207-1-AP, proteintech, USA).

Techniques: Ligation, Real-time Polymerase Chain Reaction, Immunofluorescence, Staining

Journal: Frontiers in Immunology

Article Title: Low-intensity pulsed ultrasound ameliorates partial infraorbital nerve ligation-induced trigeminal neuropathic pain through inhibiting Schwann cell Pannexin 1 channel

doi: 10.3389/fimmu.2025.1712759

Figure Lengend Snippet: Mechanistic validation of LIPUS efficacy in vivo and in vitro . (A) LIPUS treatment protocol. (B) RT-qPCR of TNF-α-stimulated Schwann cells revealed LIPUS-mediated suppression of pro-nociceptive channels and inflammatory factors (Tukey’s test, n=3 per group; mean ± SD). *p < 0.05, ***p < 0.001 (TNF-α vs. DMEM); #p < 0.05, ##p < 0.01, ###p < 0.001 (TNF-α vs. LIPUS). (C) Western blot analysis of Panx1, TNF-α, IL-1β, and IL-6 protein expression (n=3 per group). (D) Timeline for BzATP injections and LIPUS intervention. (E) Behavioral pain scores in response to 0.02g filament stimulation (statistics as in Figure 2 with primary ΔAUC_{0–3 d} (95% CI), n=5 per group; mean ± SD). (F) RT-qPCR of BzATP-treated infraorbital nerve with/without LIPUS (day 3; Tukey’s test, n=5–8 per group; mean ± SD). *p < 0.05, **p < 0.01, ***p < 0.001 (BzATP vs. saline); #p < 0.05, ##p < 0.01, ###p < 0.001, n.s. (BzATP vs. LIPUS+BzATP). (G) Immunofluorescence co-staining of S100β/P2X7 and S100β/Panx1 in infraorbital nerve (scale bar: 20 μm). Analysis included at least three sections from three animals per group.

Article Snippet: The sections were incubated with following primary antibodies overnight at 4°C: S100β antibody specifically labels Schwann cells, as validated in previous studies , rabbit anti-Iba1 (1:1000, 19741, Wako, Osaka, Japan), mouse anti-S100β (1:500, bsm-10832M, Bioss), and rabbit anti-Panx 1 (1:200, A00915-2, Boster, China), rabbit anti-IL-1β (1: 200, A00101, Boster, China), rabbit anti-TNF-α (1: 400, BA0131, Boster, China), rabbit anti-P2X7 (1:200, 28207-1-AP, proteintech, USA).

Techniques: Biomarker Discovery, In Vivo, In Vitro, Quantitative RT-PCR, Western Blot, Expressing, Saline, Immunofluorescence, Staining

Journal: NPJ Regenerative Medicine

Article Title: Harnessing 3D collagen hydrogel-directed conversion of human GMSCs into SCP-like cells to generate functionalized nerve conduits

doi: 10.1038/s41536-021-00170-y

Figure Lengend Snippet: a GMSCs were encapsulated in 40 µl of 3D-collagen hydrogel at different concentrations (2, 4, 6 mg/mL) and a cell density of 2 × 10 6 /mL and then filled into AxoGuard Nerve protector or connector (NGC) (10 mm in length and 2 mm in inner diameter). Then, the constructs (NGC containing 3D collagen hydrogel encapsulated with GMSCs) were cultured for 24 h in complete α-MEM. Portions of this figure were made using templates from SMART SERVIER MEDICAL ART ( https://smart.servier.com ). b Before harvesting, the NGC constructs were labeled with 10 µM calcein-AM at 37 °C for 30 min. Cryosections were cut and the migrated cells labeled with calcein-AM (green color) in the wall matrix were observed under a fluorescence microscope. Nuclei were counterstained with 4′, 6-diamidino-2-phenylindole (DAPI; blue). c Quantification of IF intensity of calcein-AM. d Cryosections of NGCs containing cell-free collagen hydrogel (Empty) or GMSC-laden collagen hydrogel at a concentration of 4 mg/mL (Cell-laden) were prepared for dual-color immunostaining for human nuclei (hNuclei; red color), SOX10 (red color), S-100β (green color) or GFAP (green color). Nuclei were counterstained with 4′, 6-diamidino-2-phenylindole (DAPI; white or blue). Scale bar = 50 µm ( b , d ). *** p < 0.0001. Data represent the mean ± SD, n = 3 biological replicates. One-way ANOVA with the Tukey’s post test ( c ). NGC, nerve guidance conduit.

Article Snippet: Cryosections prepared from 3D-collagen gel or GMSC-seeded nerve conduits were blocked and permeabilized for 1 h at room temperature in PBS with 2.5% goat serum and 0.5%Triton X‐100, followed by incubation with the following primary antibodies at the appropriate dilution overnight at 4 °C: p75 (mouse IgG, 1:200, Sigma), SOX-9 (rabbit IgG, 1:200, Cell Signal Tech), SOX-10 (mouse IgG, 1:200, R & D), S-100β (rabbit IgG, 1:200, Boster Biological Tech), NOTCH3 (rabbit IgG, 1:200, Abcam), HES1 (rabbit IgG, 1:200, Cell Signaling Tech), vinculin (mouse IgG, 1:400, Millipore), TRITC-conjugated phalloidin (1:400, Millipore), BDNF (rabbit IgG, 1:200, Abcam), GDNF (rabbit IgG, 1:200, Abcam), or NGF (rabbit IgG, 1:200, Abcam).

Techniques: Construct, Cell Culture, Labeling, Fluorescence, Microscopy, Concentration Assay, Immunostaining