Fig 1: The effect of OECs‐Exo treatment on neuronal survival and preservation of axons after SCI. (a) Representative images of NeuN‐immunostaining in transverse sections in PBS‐ or OECs‐Exo‐treated rats at 4 wpi. The area of 3000, 2000, 1000 μm rostral and caudal to the lesion, as well as epicenter was selected to analyze. Scale bar = 300 μm. (b) Quantification of NeuN‐positive cells. Note that OECs‐Exo treatment significantly increased the numbers of NeuN‐stained neurons. N = 6/group, *p < 0.05. (c,d) Representative images of AAV2/5‐GFP‐labeled axons in longitudinal sections at 4 wpi in different groups. The high magnification images (C1–C3, D1–D3) were from the low magnification images in epicenter, rostral, and caudal areas of 2000 μm adjacent to lesion. Scale bar = 800 μm in low magnification images and 300 μm in high magnification images. Note that there are some axons present at caudal to the epicenter, demonstrating that our SCI model was incomplete. (e) Quantification of AAV2/5‐GFP‐labeled axons. N = 6/group, *p < 0.05. (f) The area on which quantification was performed. (g) Expression and quantification of NF200 in sham, SCI + PBS, or SCI + OECs‐Exo treatment. N = 3, *p < 0.05, **p < 0.01
Fig 2: S100A9 blockade improves neuronal cell survival after SCI. Immunofluorescence of (A) MAP2 and (B) NF200 at the lesion site on day 28 following surgery among the different experimental groups. (C) Nissl staining for neuronal morphology at the lesion site 28 days post-OR among groups. n=5. Scale bar, 200 µm. **P<0.01, ***P<0.001. ##P<0.01. S100A9, S100 calcium-binding protein A9; SCI, spinal cord injury; MAP2, microtubule associated protein 2; NF200, neurofilament 200; post-OR, after the operation; High-ABR, high ABR dose; Low-ABR, low ABR dose.
Fig 3: Trypsin-3 causes internalisation of PAR2 in human submucosal neurons. Representative images showing that the expression of PAR2 in human submucosal neurons changes in the presence of trypsin-3 (10 nM). Upper panel shows the clustered staining of PAR2 (arrowheads, red) on the plasma membrane of submucosal neurons (asterisks, NF200, green). Trypsin-3 (2 hours) caused internalisation of PAR2 (red) inside the neuron (asterisk, NF200, green). PAR2 staining appeared diffuse or clustered (arrowheads) inside the neuron. Pre-incubation with GB83 (10 μM, 30 min before trypsin-3) inhibited the internalisation of PAR2, which was expressed on the plasma membrane (arrowheads, red) of neurons (asterisks, NF200, green). Right panels show merged images. Scale bars: 20 μm.
Fig 4: Biomimetic periosteum promotes nerve regeneration and bone regeneration of bone defects. (A–C) NF200, OCN staining immunofluorescence detection of bone defect in PCL, PPE, PPEA group. (D) The quantitative analysis of nerve regeneration in A-C. (E) The quantitative analysis of bone regeneration in A-C. Scale bar = 200 μm in whole slice, 50 μm in enlarge.N = 5, ∗p＜0.05, ∗∗p＜0.01.
Fig 5: HUCMSCs-derived exosomal miR-146 b attenuates PC12 nerve cell injury by targeting NF-κB signaling. PC12 cells were stimulated with LPS for 24 h, followed by treatment with hUCMSCs -derived exosomes and NF-κB inhibitor PDTC another 24 h. (A) Cell viability was measured by CCK-8. (B) Cell apoptosis was checked by TUNEL assay. (C) The protein levels of Bcl-2, Bax, cleaved caspase-3, caspase-3, GFAP, and NF200 were detected by western blotting. (D–F) The release of TNF-α, IL-6, and IL-1β were measured by ELISA assay. **p < 0.01. Scale bar, 50 μm.
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