Journal: Neural Regeneration Research

Article Title: Effects of electroacupuncture on the functionality of NG2-expressing cells in perilesional brain tissue of mice following ischemic stroke

doi: 10.4103/1673-5374.330611

Figure Lengend Snippet: Effects of EA stimulation on NG2-expressing cell types in the perilesional striatum and the corpus callosum of mice at 21 days after MCAO . Photomicrographs (A) and histograms (B and C) showing labeling and quantification of NG2 (green) -, CC1 (red) -, CD31 (red) - and CD68 (red) -positive cells in the perilesional striatum and the corpus callosum of MCAO mice. EA stimulation significantly increased the NG2 and CD68double positive cells in the perilesional striatum. n = 6. All data are shown as mean ± SEM. ## P < 0.01, vs . MCAO group; & P < 0.05, vs . MCAO + EA1 group (one-way analysis of variance with Tukey's post hoc tests). Scale bar in A: 20 μm. DAPI: 4′,6-Diamidino-2-phenylindole; EA1: electroacupuncture at 1 mA; MCAO: middle cerebral artery occlusion; NG2: neural/glial antigen 2.

Article Snippet: The samples were then incubated with following primary antibodies: mouse anti-green fluorescent protein (GFP; 1:100, Cat# 2955, Cell Signaling), rabbit anti-GFP (1:100, Cat# G10362, Invitrogen), rat anti-BrdU (1:100, Cat# ab6326, Abcam), rabbit anti-CC1 (1:100, Cat# ab40778, Abcam), rat anti-CD31 (1:100, Cat# 550274, BD Biosciences, San Jose, CA, USA), mouse anti-CD68 (1:100, Cat# MCA1957, AbD Serotec), rabbit anti-BDNF (1:100, Cat# ab108319, Abcam), or mouse anti-GSK3β (1:100, Cat# ab93926, Abcam), and counterstained with 4′,6-diamidino-2-phenylindole (DAPI, H3570, Invitrogen) to label cell nuclei.

Techniques: Expressing, Labeling

Journal: Neural Regeneration Research

Article Title: Effects of electroacupuncture on the functionality of NG2-expressing cells in perilesional brain tissue of mice following ischemic stroke

doi: 10.4103/1673-5374.330611

Figure Lengend Snippet: Effects of EA stimulation on cellular phenotype and the expression of BDNF and GSK3β in GFP + /DAPI + NG2-expressing cells, or whole cells in ipsilateral perilesional striatum of NG2-mEGFP mice at 21 days after MCAO . Quantitative analysis of the flow cytometry data (A and C) and histograms showing the percentage of GFP + /DAPI + cells also expressing BrdU, CC1, CD31, CD68, and GSK3β, as well as the percentage of DAPI + cells also expressing BDNF (B and D, respectively). The numbers of GFP + cells that also express BrdU, CC1, CD31and GSKβ were significantly increased after EA1 stimulation. n = 5. All data are shown as mean ± SEM. # P < 0.05, ## P < 0.01, and ### P < 0.01, vs . MCAO group (independent samples t -test). BDNF: Brain-derived neurotrophic factor; BrdU: bromodeoxyuridine; DAPI: 4′,6-Diamidino-2-phenylindole; EA1: Electroacupuncture at 1 mAEA1: electroacupuncture at 1 mA; GFP: green fluorescent protein; GSK3β: glycogen synthase kinase 3 beta; MCAO: middle cerebral artery occlusion; mEGFP: mutated enhanced green fluorescent protein; NG2: neural/glial antigen 2.

Article Snippet: The samples were then incubated with following primary antibodies: mouse anti-green fluorescent protein (GFP; 1:100, Cat# 2955, Cell Signaling), rabbit anti-GFP (1:100, Cat# G10362, Invitrogen), rat anti-BrdU (1:100, Cat# ab6326, Abcam), rabbit anti-CC1 (1:100, Cat# ab40778, Abcam), rat anti-CD31 (1:100, Cat# 550274, BD Biosciences, San Jose, CA, USA), mouse anti-CD68 (1:100, Cat# MCA1957, AbD Serotec), rabbit anti-BDNF (1:100, Cat# ab108319, Abcam), or mouse anti-GSK3β (1:100, Cat# ab93926, Abcam), and counterstained with 4′,6-diamidino-2-phenylindole (DAPI, H3570, Invitrogen) to label cell nuclei.

Techniques: Expressing, Flow Cytometry, Derivative Assay

Journal: Frontiers in Cellular Neuroscience

Article Title: Activity-dependent oligodendrocyte calcium dynamics and their changes in Alzheimer’s disease

doi: 10.3389/fncel.2023.1154196

Figure Lengend Snippet: Neuronal activity-dependent Ca 2+ activities in oligodendrocytes (OCs). (A,B) Representative images (A) and quantification (B) of GCaMP expression in the motor cortex of PLP-GCaMP mice and co-localization with markers for OC + oligodendrocyte precursor cell (OPC) (Olig2), OC (CC1), OPC (PDGFRα, NG2), neuron (NeuN), astrocyte (S100β) (Olig2 [83.41 ± 3.899%], CC1 [72.65 ± 3.464%], PDGFRα+ [9.979 ± 0.9322%], NG2 [6.736 ± 0.2401%], NeuN [1.481 ± 0.7246%], and S100β [8.156 ± 0.5504%]). Scale bar, 30 μm. (C,D) Representative images of MBP immunostaining in the motor cortex of PLP-GCaMP6 mice. Scale bars: in (C) , 100 μm; in (D) , 30 μm. (E) Quantitative analysis of the co-localization areas of MBP and GCaMP’ + ve processes based on immunostaining data. The proportion of MBP’ + ve processes in the processes of GCaMP’ + ve cells was about 15%. (F) Experimental protocol of Ca 2+ imaging in OCs. The first craniotomy was performed one week before Pre-imaging (Pre) using two-photon microscopy. After Pre-imaging, a second craniotomy was performed and TTX was applied for 30 min, followed by a second imaging (TTX) of the same cells. (G,H) Representative image of GCaMP’ + ve cells of the motor cortex in PLP-GCaMP6 mice before and after TTX application. Red spots indicate the Ca 2+ activated areas (spot). Scale bar = 30 μm. Representative Ca 2+ traces from spots of typical GCaMP’ + ve cells are shown. (I) Changes in Ca 2+ spots, Ca 2+ events and total area under the curve (AUC) of GCaMP’ + ve cells between before and after TTX application. The number of Ca 2+ spots and Ca 2+ events and total AUC were significantly decreased after TTX application. Pre: n = 6 mice, 13 imaging fields (cells); TTX: n = 6 mice, 13 imaging fields (cells). * p < 0.05, ** p < 0.01, Mann–Whitney U test. Data are presented as mean ± standard error of mean. For detailed data, check the source data file. (J) Changes in AUC, Amplitude, and Latency of GCaMP’ + ve cells between before and after TTX application. AUC was not significantly changed, Amplitude was significantly decreased, and Latency was significantly increased after TTX application. Pre: n = 6 mice, 426 events; TTX: n = 6 mice, 189 events. N.S., not significant, * p < 0.05, *** p < 0.001, Mann–Whitney U test. Violin plots show median (black line) and distribution of the data. For detailed data, check the source data file. (K) Proportions of AUC and Latency were not significantly changed between before and after TTX application. The proportion of lower Amplitude was significantly increased after TTX application. N.S., not significant, ** p < 0.01, Kolmogorov–Smirnov test. For detailed data, check the source data file.

Article Snippet: The primary antibodies used in this study were as follows: anti-Olig2 (rabbit; Millipore, AB9610; 1:1000), anti-adenomatous polyposis coli (clone CC1) (mouse; Calbiochem, OP80; 1:500), anti-PDGFRα (goat; R&D systems, AF1062; 1:200), anti-NG2 (rabbit; Millipore, AB5320; 1:500), anti-NeuN (mouse; Millipore, MAB377; 1:500), anti-S100β (rabbit; Abcam, ab52642; 1:1000), anti-MBP (mouse; BioLegend, clone SMI 99; 1:100), and anti-GFP (chicken; Novus biologicals, NB100-1614; 1:2000).

Techniques: Activity Assay, Expressing, Immunostaining, Imaging, Microscopy, MANN-WHITNEY