Fig 1: hAMSC-GDNF promotes therapeutic effects in PD models. (A) Representative immunofluorescence staining pictures of TH in the different groups, scale bar, 200 µm. (B–E) Immunostaining assays showed that the volume of grafts, the number, and density of TH-positive cells in the PD/hAMSC-GDNF group were significantly higher than the other groups. (F–I) The number of TH- and NeuN-positive cells in the PD/hAMSC-GDNF group was larger than in the PD/hAMSC-vector group in 6-OHDA lesioned mouse models. Scale bar, 50 µm. Error bars represent SEM. *P < 0.05, **P < 0.01, ***P < 0.001. GDNF: glial cell line-derived neurotrophic factor; GFP: green fluorescent protein; hAMSCs: human primary adipose-derived mesenchymal stem cells; OD: optical density; PD: Parkinson’s disease; SEM: standard error of the mean; TH: tyrosine hydroxylase.
Fig 2: Laser confocal fluorescence microscopic images.The impact of DPSC-CM on trigeminal ganglion survival and neurite length after 4 days of culture in the presence of specific neutralising antibodies. Antibodies against GDNF (1 µg/mL; R&D systems, Oxon, UK; AF-212-NA) and NT-3 (2 µg/mL; R&D systems; Cat# AF1404) were used to test the impact of their absence on TG neuronal survival using NeuN (neuronal nucleus immune stain marker; Cat# ab128886 at a 1:1000 dilution; Abcam, Cambridge, MA, USA) and neurite outgrowth extension using MAP-2 (mature neuronal immune stain marker; Cat# ab11267 at a 1:500 dilution; Abcam). Immunolocalization of neuronal markers NeuN in neuronal cell body and MAP-2 in neuronal cell body and processes showing the difference in the number of surviving neurons and the neurite length, respectively. DAPI (blue) was used as a nuclear staining. Anti-GDNF antibody completely blocks CM mediated neurite outgrowth effect while anti-NT-3 markedly reduce the number of surviving neurons. Scale bars: 100 µm. Unpublished data. CM: Concitioned medium; DAPI: 4',6-diamidino-2-phenylindole; GDNF: glial cell line-derived neurotrophic factor; MAP-2: microtubule-associated protein 2; NeuN: neuronal nuclear protein; NT-3: neurotrophin-3.
Fig 3: Neuritic degeneration after polybrene treatment in vivo and in vitro. a Performance on the accelerating rotating rod apparatus showed that the mice with polybrene injection had significantly shorter step length and less performance on the rotating rod (n = 4, **P< 0.01). b–d Altered NeuN and GFAP immunostaining after ICV injection with polybrene. b NeuN and GFAP immunofluorescent staining in sagittal sections from 3-month mouse brain 3 days after ICV injection of polybrene or saline. The regions enclosed by the white dotted rectangles are displayed at higher magnification at right. Quantification is shown in c and d. Scale bar: 100 µm (n = 4, ***P < 0.001). e–g Fraction of neurites with beads and fragmented neurites in neurons treated with polybrene for 24 h. The neurites with beads are indicated by arrow. Scale bar: 25 µm (n = 4, ***P < 0.001)
Fig 4: ANT-DBS reduced BDNF expression in the hippocampal neurons of epileptic monkeys. Immunofluorescence staining of BDNF and NeuN in hippocampal CA1 (A) and CA3 (C) neurons. NeuN is a specific neuronal marker. BDNF expression was elevated in EP and EP-sham-DBS groups in hippocampal CA1 and CA3 neurons and reduced by ANT-DBS. Immunofluorescent intensity of BDNF in hippocampal CA1 (B) and CA3 (D) neurons was quantified. (n=3 in each group; in each monkey, a total of thirty cells in CA1 or CA3, the average immunofluorescent intensity of these cells was recorded) *P < 0.05; **P < 0.01; NS, P > 0.05. Data were presented as mean ± SD.
Fig 5: LRCH1 knockdown worsens tissue damage and locomotor function after SCI. a H&E staining of spinal cords on day 7 after SCI. Sham-saline, sham-operated rats injected with saline. LC, rats receiving LC-infected microglia. LL, rats receiving LL-infected microglia. The arrow indicates the tissue deficit probably caused by saline injection. b, c Double staining of TUNEL and NeuN in T12 on day 7 after SCI. Representative fluorescent images are shown in b. Please note that the green color of NeuN is a pseudocolor added by the imaging software, because Alexa Fluor 647 is a far-red fluorescent dye invisible to the eyes. Statistics for the proportions of TUNEL+NeuN+ cells in NeuN+ cells was shown in c. n = 3 in 3 independent experiments. d BBB scores. n = 8. e mRNA levels of indicated cytokines in T12 on day 7 after SCI. n = 6 in 3 independent experiments. *p < 0.05; **p < 0.01; ***p < 0.001
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