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

Journal:

Article Title: Medial Ganglionic Eminence-derived Neural Stem Cell Grafts Ease Spontaneous Seizures and Restore GDNF Expression in a Rat Model of Chronic Temporal Lobe Epilepsy

doi: 10.1002/stem.446

Figure Lengend Snippet: Distribution and differentiation of cells derived from MGE-NSC grafts in the chronically epileptic hippocampus after 3-months of grafting. The figures A1–A11 illustrate the location of transplants and transplant-derived cells (shown in pink color based on Chlorodeoxyuridine+ [CldU+]) immunoreactivity) with respect to hippocampal cell layers and subfields in a chronically epileptic rat. These tracings, performed using the Neurolucida software (Microbrightfield Inc), represent every tenth 30-µm thick section through a chronically epileptic hippocampus that received four MGE-NSC grafts. Note that grafts and graft-derived cells were mostly located in the CA3 subfield and lateral ends of the CA1 subfield and the dentate gyrus. The core of transplants partially projected ventrally into the thalamus only at certain levels. The figures B1–B3 illustrate the distribution of CldU+ graft-derived cells in the host hippocampus. B2 is a magnified view of a region from B1 and B3 is an enlarged view of a region from B2. Both B2 and B3 show engraftment of graft-derived cells into strata oriens (SO), pyramidale (SP) and radiatum (SR) of the CA3 subfield. Scale bar, A1–A11, 1000 µm; B1, 500µm; B2, 200 µm; B3, 100 µm. Figures C1–F1 illustrates differentiation of MGE-NSC graft-derived CldU+ cells into NeuN+ mature neurons (C1), S-100β+ mature astrocytes (D1), NG2+ oligodendrocyte precursors (E1) and GABA+ neurons (F1), visualized through dual immunolabeling for CldU (red) and markers of neurons/glia (green) and Z-section analyses in a confocal microscope. Scale bar, C1–E1, 20 µm; F1, 10 µm; orthogonal inset of F1, 5 µm. The bar chart in G1 depicts the percentages of graft-derived cells that differentiate into NeuN+ neurons, GABA+ neurons, S-100β+ astrocytes and NG2+ oligodendrocyte precursors.

Article Snippet: The primary antibodies used were anti CldU from Serotech, anti NeuN, anti NG2, anti S-100β and anti GDNF from Chemicon, and anti GABA from Sigma.

Techniques: Derivative Assay, Software, Immunolabeling, Microscopy

Journal:

Article Title: Medial Ganglionic Eminence-derived Neural Stem Cell Grafts Ease Spontaneous Seizures and Restore GDNF Expression in a Rat Model of Chronic Temporal Lobe Epilepsy

doi: 10.1002/stem.446

Figure Lengend Snippet: Expression of glial-derived neurotrophic factor (GDNF) in cells derived from MGE-NSC grafts (A1–A4), and MGE-NSC grafting mediated changes in GDNF expression of host hippocampal astrocytes (B1–E1) and hippocampal neurogenesis (F1–I1). Arrows in figures A1–A3 show examples of graft-derived CldU+ cells (red) that express GDNF (green). A4 shows orthogonal views of a GDNF+ graft derived cell (with CldU+ red nucleus and GDNF+ green cytoplasm). Scale bar, A1–A3 = 40µm; A4 = 5µm. Figures B1–D3 show confocal microscopic images of S-100β+ hippocampal astrocytes (red) that exhibit GDNF expression (green) in an age-matched control rat (B1–B3), a rat with chronic epilepsy alone (C1–C3) and a chronically epileptic rat that received MGE-NSC grafts (D1–D3). The insets in B3, C3, and D3 show orthogonal views of cells indicated by arrows. Note that these cells are positive for both S-100β and GDNF. Arrowheads in C1 and C3 denote S-100β+ astrocytes that lack GDNF expression in the epilepsy alone group. Scale bar, B1–D3 = 10 µm. The bar chart in E1 shows percentages of S-100β+ astrocytes expressing GDNF in the dentate gyrus, and CA1 & CA3 subfields of the hippocampus in different groups. Note that GDNF expression in astrocytes declines substantially with chronic epilepsy in all regions of the hippocampus but recovers dramatically after MGE-NSC grafting. Figures F1–H1 illustrates examples of newly born (doublecortin+) neurons in the dentate gyrus in the three groups. Newly born neurons in the rat with chronic epilepsy alone (G1) and the chronically epileptic rat that received MGE-NSC grafts (H1) exhibit abnormal polarity and basal dendrites, in comparison to normal orientation of dendrites in newly born neurons of the age-matched control rat (F1). DH, dentate hilus; GCL, granule cell layer. Scale bar, F1–H1 = 25µm. The bar chart in I1 compares numbers of newly born neurons between the three groups. Note that rats with chronic epilepsy alone (red) and chronically epileptic rats that received MGE-NSC grafts (green) exhibit greatly declined hippocampal neurogenesis compared to age-matched control rats (blue).

Article Snippet: The primary antibodies used were anti CldU from Serotech, anti NeuN, anti NG2, anti S-100β and anti GDNF from Chemicon, and anti GABA from Sigma.

Techniques: Expressing, Derivative Assay

Journal: ACS Omega

Article Title: Platelet-Rich Plasma Gel-Loaded Collagen/Chitosan Composite Film Accelerated Rat Sciatic Nerve Injury Repair

doi: 10.1021/acsomega.2c05351

Figure Lengend Snippet: COL/CS composite film improved nerve morphology following SNI in rats. (A) Micrographs of the sciatic nerve stained by hematoxylin and eosin (H&E) at 8 weeks after nerve injury (×200 magnification and bottom right is ×400 magnification). The red arrow indicates nerve fiber vacuolation; the black arrow indicates myelin sheath. Immunohistochemical analysis of (B) S-100β and (C) GFAP. (D) Quantitative analysis of the S-100β mean gray value. (E) Quantitative analysis of the GFAP mean gray value. Values represent the mean ± SD, n = 5, * p < 0.05.

Article Snippet: The slides were then incubated at 4 °C overnight with monoclonal anti-S-100β (1:200, Cat no: 66616-1-Ig, Proteintech) or polyclonal anti-GFAP antibody (1:200, Cat no: 16825-1-AP, Proteintech).

Techniques: Staining, Immunohistochemical staining

Journal: Neural Regeneration Research

Article Title: Folic acid contributes to peripheral nerve injury repair by promoting Schwann cell proliferation, migration, and secretion of nerve growth factor

doi: 10.4103/1673-5374.243718

Figure Lengend Snippet: Purity of cultures determined by immunofluorescence staining for the Schwann cell marker S-100. (A) Schwann cells were identified by S-100β immunofluorescence staining. (B) The nuclei of all cells were labeled with 4′,6-diamidino-2-phenylindole (DAPI). (C) Merged image of Schwann cells and nuclei of all cells. Scale bar: 100 μm. (D) Schwann cell culture purity was equal to the proportion of cells stained with fluorescein isothiocyanate (FITC) (green) among all cell nuclei stained with DAPI (blue). The purity of Schwann cells was 93–98%.

Article Snippet: The purity of the cultures was determined by immunofluorescence staining for anti-S-100 (S-100β; Boster, Wuhan, China) as a Schwann cell marker.

Techniques: Immunofluorescence, Staining, Marker, Labeling, Cell Culture

Journal: Scientific Reports

Article Title: Biocompatibility between Silicon or Silicon Carbide surface and Neural Stem Cells

doi: 10.1038/s41598-019-48041-3

Figure Lengend Snippet: Differentiation of Dental Pulp Stem Cells into Neural Stem Cells. ( A ) Representative photomicrograph of neuronal-like stem cells derived from DPSC in bright field taken from randomly selected slides scanned by a Nikon Ti Eclipse inverted microscope (scale bar 50 μm). ( B ) Gene expression changes following differentiation. Quantitative real-time PCR assay was performed to assess gene expression changes after 16 days of RA-treatment. mRNA levels were normalized to the amount of β-actin mRNA and represented in a heatmap (*P < 0.05 and ***P < 0.001 as determined by two-way ANOVA followed by Sidack post hoc test). ( C , D ): representative photomicrographs of DP-NSCs show the fluorescent distribution of neuronal marker NF and MAP2. Nuclei were counterstained with DAPI (scale bar 50 μm).

Article Snippet: The following primary antibodies were incubated overnight at 4 °C: mouse monoclonal anti-NF-H antibody (1:200; Abcam, Cambridge, UK), mouse monoclonal anti-MAP2 antibody (1:1000; Merck Millipore, Billerica, Massachusetts, USA), mouse monoclonal anti-S-100β antibody (1:100; Sigma), rabbit polyclonal anti-nestin antibody (1:200; AbCam), rabbit polyclonal anti-GFAP (1:1000; Dako).

Techniques: Derivative Assay, Inverted Microscopy, Gene Expression, Real-time Polymerase Chain Reaction, Marker

Journal: Scientific Reports

Article Title: Biocompatibility between Silicon or Silicon Carbide surface and Neural Stem Cells

doi: 10.1038/s41598-019-48041-3

Figure Lengend Snippet: List of used primers pairs.

Article Snippet: The following primary antibodies were incubated overnight at 4 °C: mouse monoclonal anti-NF-H antibody (1:200; Abcam, Cambridge, UK), mouse monoclonal anti-MAP2 antibody (1:1000; Merck Millipore, Billerica, Massachusetts, USA), mouse monoclonal anti-S-100β antibody (1:100; Sigma), rabbit polyclonal anti-nestin antibody (1:200; AbCam), rabbit polyclonal anti-GFAP (1:1000; Dako).

Techniques: