Journal: Frontiers in Cellular Neuroscience

Article Title: Hypoxic culture of umbilical cord mesenchymal stem cell-derived sEVs prompts peripheral nerve injury repair

doi: 10.3389/fncel.2022.897224

Figure Lengend Snippet: Cellular localization and uptake of fluorescently labeled sEVs with SCs. (A) Cellular colocalization: SP8 was used to photograph sEVs prelabeled with PKH67 (green fluorescence) with Hoechst 33,342 (blue fluorescence)-stained SC nuclei. (B) Western blot: Detection of ERK1/2, ZEB2, and c-JUN expression levels in NC Schwann cells and after 48-h treatment with hypoxia sEVs. (C) Real-time PCR: Inflammatory and other restoration-related factors (IL-1β, IL-6, TNF-α, etc.) were detected. (D) ELISA test: GDNF NDF and NT-3 in the supernatant of Schwann cell preparation. Statistical significance, * p < 0.05,** p < 0.01, and **** p < 0.0001. *Significant difference.

Article Snippet: The following ELISA kits were: mouse glial cell line-derived neurotrophic factor (GDNF), ELISA kit (CSB-E07341m, CUSABIO), mouse neurotrophin 3 (NT-3), ELISA kit (CSB-E04687m, CUSABIO), and mouse nerve growth factor (NGF), ELISA kit (CSB-E04684m, CUSABIO).

Techniques: Labeling, Fluorescence, Staining, Western Blot, Expressing, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Plasmid map for GDNF production

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Plasmid Preparation

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Bone-marrow derived macrophages were transfected by electroporation using four different conditions described in Experimental section. Then, cells were washed and cultured in complete media for up to 6 days. The GDNF expression levels in cells (solid bars), and EVs collected from conditioned media (stripped bars) was assessed by ELISA on day 1 (white bars), day 4 (grey bars), and day 6 (black bars). Successful transfection was accomplished with three conditions (#2 - #4). N = 4, *p < 0.05, compared to sham-transfected macrophages (dashed line, condition #1).

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Derivative Assay, Transfection, Electroporation, Cell Culture, Expressing, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Primary macrophages were transfected with GDNF-encoding p DNA by electroporation (condition #4), and EV-GDNF were collected from conditioned media on day 6. EV-GDNF were characterized for size, zeta potential, and morphology by ZetaView QUATT Nanoparticle Tracking Microscope PMX-420 ( A ), and AFM ( B ). The presence of EV-specific membrane proteins was EV-GDNF was confirmed by Wes ( C ) and quantified using Compass SW software ( D ). The bar: 1 µm.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Transfection, Electroporation, Zeta Potential Analyzer, Microscopy, Membrane, Software

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: EVs samples from sham-transfected (white bars), and GDNF-transfected (black bars) macrophages were digested ( N =3) with trypsin and examined by nano-liquid chromatography tandem MS (nanoLC–MS/MS) with multiple reaction monitoring (MRM). Samples of 20 µg total protein were used, and 0.08 µg (0.4 % of the sample) was injected. No significant differences in specific proteins expression were found between sham EVs and EV-GDNF (t-tests, p < 0.05). Peptide identification is shown in . A CD81 peptide employed in other studies was not detected in these analyses. Values are means ± SD.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Transfection, Liquid Chromatography, Tandem Mass Spectroscopy, Targeted Proteomics, Injection, Expressing

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Macrophages were transfected with GDNF-encoding p DNA by electroporation, and the levels of GDNF-DNA in the cells ( A ) and EVs released by these cells ( B ) were assessed. A significant amount of GDNF-DNA was detected in parent cells, as well as in the EVs. Statistical significance was assessed by One Way ANOVA corrected for multiple comparisons using the FDR. ** p < 0.01, or **** p < 0.0001.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Transfection, Electroporation

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: The effect of EV-GDNF on motor functions and activity was assessed in Wire hanging test, and Rotarod test ( A, B ), as well as in OFA tests ( C, D ). ( A, B ) Transgenic mice were i.n . injected with EV-GDNF (triangles, 3×10 9 particles/10 µL/mouse), or sham EVs (empty circles, 3×10 9 particles/10 µL/mouse), or saline (filled circles, 10 µL/mouse). Wild type mice were i.n . injected with saline (filled squares, 10 µL/mouse) were used as controls. Wire hanging test ( A ), and Rotarod test ( B ) demonstrated significant improvements in motor functions upon treatment with EV-GDNF. ( C, D ) OFA tests at 12 mo. demonstrated improved behavior in EV-GDNF treated PD mice (striped bars) compared to PD mice treated with saline (white bars) that was similar as in healthy WT mice (black bars) including decreases in the hyperactivity and anxiety-like behavior. The differences between sham EVs and saline in PD mice were inconclusive. Values are means ± SEM ( N = 10), * p < 0.05, ** p < 0.005, and # p < 0.05, as compared to WT control.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Activity Assay, Transgenic Assay, Injection, Saline, Control

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Transgenic mice (4 mo. old, N = 10) were i.n . injected with: saline (10 µL/mouse), or EV-GDNF (3×10 9 particles/10 µL/mouse), or sham EVs (3×10 9 particles/10 µL/mouse). Wild type control mice were intranasally injected with saline (10 µL/mouse). Animals were sacrificed at mo. 16, and brain slides were stained with TH, a marker for dopaminergic neurons ( A ); or Ab to CD11b for activated microglia ( B ). The images indicate significant preservation of TH-positive neurons and decrease in microglial activation in Parkin Q311(X)A mice upon EV-GDNF treatment compared to PD mice treated with saline. The administration of sham EVs did not cause significant therapeutic effects.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Transgenic Assay, Injection, Saline, Control, Staining, Marker, Preserving, Activation Assay

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Transgenic mice (4 mo. old) were intranasally injected with: saline (10 µL/mouse), or EV-GDNF (3×10 9 particles/10 µL/mouse), or sham EVs (3×10 9 particles/10 µL/mouse). Wild type control mice were intranasally injected with saline (10 µL/mouse). Animals were sacrificed at mo. 16, brains were removed post-mortem, and homogenized in cell lysis buffer. Elevated cytokine levels in the brain, were recorded in PD mice treated with saline and Sham EVs. Administration of EV-GDNF significantly decreased pro-inflammatory molecules in the brain compared with PD mice treated with saline. N = 4, # p < 0.05 compared to healthy WT animals; * p < 0.05 compared to PD mice treated with saline, $ p <0.05 compared to PD mice treated with saline and sham EVs.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Transgenic Assay, Injection, Saline, Control, Lysis

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Transgenic mice (4 mo. old) were intranasally injected with: saline (10 µL/mouse), or ( 3 ) EV-GDNF (3×10 9 particles/10 µL/mouse), or sham EVs (3×10 9 particles/10 µL/mouse). Wild type control mice were intranasally injected with saline (10 µL/mouse). Animals were sacrificed at mo. 16, brain slides were stained with Nissl staining ( A – D ) and H&E staining ( E – H ). The obtained bright light images show lower number of Nissl bodies with neuronal shrinkage ( B ) and damages tissues with degeneration in the neurons ( F ) in PD mice treated with saline when compared to WT mice ( A, E ). Histological analysis indicate neuroprotective effects in the brain of PD mice treated with EV-GDNF with healthy morphology in tissue structure and high integrity of neurons ( C, G ) when comparted to PD mice treated with saline ( B, F ). The administration of sham EVs did not have significant therapeutic effect in PD mice ( D, H ). Black arrows, degenerated neurons; blue arrows, elongated irregular nuclear morphology.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Transgenic Assay, Injection, Saline, Control, Staining

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Transgenic mice (4 mo. old) were intranasally injected with: saline (10 µL/mouse), or EV-GDNF (3×10 9 particles/10 µL/mouse), or sham EVs (3×10 9 particles/10 µL/mouse) weekly three times. Wild type control mice were intranasally injected with saline (10 µL/mouse). Animals were sacrificed at mo. 16, brain slides were stained with Nissl staining. The obtained bright light images show lower number of Nissl bodies with neuronal shrinkage and damages tissues with degeneration in the neurons in PD mice treated with saline when compared to WT mice. Histological analysis indicates neuroprotective effects in the brain of PD mice treated with GDNF-EVs with healthy morphology in tissue structure and high integrity of neurons when comparted to PD mice treated with saline. The administration of sham EVs did not have significant therapeutic effect in PD mice.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Transgenic Assay, Injection, Saline, Control, Staining

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Transgenic mice (4 mo. old) were intranasally injected with: saline (10 µL/mouse), or EV-GDNF (3×10 9 particles/10 µL/mouse), or sham EVs (3×10 9 particles/10 µL/mouse) weekly three times. Wild type control mice were intranasally injected with saline (10 µL/mouse). Animals were sacrificed at mo. 16, brain slides were stained with Nissl staining. The obtained bright light images show damaged tissues with degeneration in the neurons in PD mice treated with saline when compared to WT mice. Histological analysis indicates neuroprotective effects in the brain of PD mice treated with GDNF-EVs with healthy morphology in tissue structure when comparted to PD mice treated with saline. The administration of sham EVs did not have significant therapeutic effect in PD mice.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Transgenic Assay, Injection, Saline, Control, Staining

Journal: bioRxiv

Article Title: Using Extracellular Vesicles Released by GDNF-transfected Macrophages for Therapy of Parkinson’s Disease

doi: 10.1101/2022.05.25.493424

Figure Lengend Snippet: Transgenic mice (4 mo. of age) were i.n . injected with saline, or EV-GDNF, or sham EVs (3×10 9 particles/10 µL/mouse, once a week, 3x weeks). At 16 mo. of age total weigh of the animals was recorded. No gross toxicity manifested in the losing weight was detected in mice injected with EV-GDNF and well as sham EVs.

Article Snippet: Human GDNF cDNA (NM_199234) was provided by OriGene (Rockville, MD, USA) that was propagated in DH5α E.coli, followed by purification Giga-prep kits (Qiagen, Valencia, CA, USA).

Techniques: Transgenic Assay, Injection, Saline

Journal: Journal of advanced research

Article Title: A cocktail hydrogel promoting the functional interneurons regeneration of human neural progenitor cells for brain injury therapy.

doi: 10.1016/j.jare.2025.05.063

Figure Lengend Snippet: Fig. 2. The in vitro release behavior of neural inducers and biocompatibility of BGA@GelMA hydrogel. (A to C) The relative release profile of BDNF, GDNF, and cAMP of the BGA@GelMA hydrogel measured with ELISA kit, n = 3. (D) Schematic representation of 3D culture system via the BGA@GelMA and the Matrigel hydrogel. (E) Flow cytometry was used to detect APC of cell apoptosis. (F) Statistical diagram of apoptotic cell distribution, n = 3, ***p = 0.0005. (G) Quantitative analysis of live cells and dead cells per field in the live/dead assay, n = 3. (H) Representative live/dead images of hNPCs co-cultured with the BGA@GelMA hydrogels and the Matrigel hydrogels after 14 days.

Article Snippet: The release kinetics of neural inducers in BGA@GelMA hydrogel were assessed using the human BDNF valukine enzyme-linked immunosorbent assay (ELISA) kit (VAL136, R&D Systems), GDNF ELISA kit (CSB-E04565h, Cusabio) and cAMP ELISA kit (CSBE04488h, Cusabio), respectively.

Techniques: In Vitro, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Live Dead Assay, Cell Culture

Journal: Ecotoxicology and environmental safety

Article Title: SCF/C-kit drives spermatogenesis disorder induced by abscopal effects of cranial irradiation in mice.

doi: 10.1016/j.ecoenv.2024.116504

Figure Lengend Snippet: Fig. 6. SCF/C-kit drives spermatogenesis disorder induced by abscopal effects of cranial irradiation. (A) KEGG analysis of differentially expressed proteins (DEPs). (B-C) The levels of mRNA and protein related to PI3K/Akt pathway in testis detected by qRT-PCR and western blotting, n = 6. The expression of P-Akt (D-E) and C-kit (L-M) in testis detected by IHC staining, Scale bar = 100 μm, n = 3. The levels of SCF and GDNF protein (F) and mRNA (G) in testis detected by ELISA and qRT-PCR. n = 6. The expression of VIM (H-J) and BrdU (O-P) in testis detected by IF staining, Scale bar = 50 μm, n= 3. The level of C-kit protein in testis detected by western blotting (K), n = 6. The level of PLZF mRNA in testis detected by qRT-PCR (N), n = 6. W: weeks.

Article Snippet: The levels of stem cell factor (SCF) and glial cell line-derived neurotrophic factor (GDNF) in the testis were measured with ELISA kits (E-EL-M0636C; E-EL-M3028; Elabscience).

Techniques: Irradiation, Quantitative RT-PCR, Western Blot, Expressing, Immunohistochemistry, Enzyme-linked Immunosorbent Assay, Staining

Journal: Bioactive Materials

Article Title: Acellular nerve xenografts based on supercritical extraction technology for repairing long-distance sciatic nerve defects in rats

doi: 10.1016/j.bioactmat.2022.03.014

Figure Lengend Snippet: Evaluation of the cytotoxicity of the ANXs scaffold in vitro. (A, D) Living/dead double staining of Schwann cells grown on the ANXs scaffold for 3 days and 7 days (live: green, dead: red). (B, E) SEM images of Schwann cells growing on CD SD and CD + scCO 2 NG scaffolds for 7 days (the picture on the right is an enlarged view of the yellow area in the picture on the left). (C, F) Immunofluorescence images of Schwann cells growing on CD SD and CD + scCO 2 NG scaffolds for 7 days, respectively (S100: red, nucleus: blue). (G) Quantification of the number of live/dead double-stained Schwann cells in each region (0.36 mm 2 ). Data are presented as the mean ± SD (n = 3). (H) The CCK-8 assay was performed after 1, 3, 5 and 7 days of cell culture. Data are presented as the mean ± SD (n = 5). (I, J) Quantitative analysis of the GDNF and NGF expression levels of Schwann cells on the ANXs scaffold. Data are presented as the mean ± SD (n = 5). Statistical analysis: n.s. no significances, **p < 0.01, *p < 0.05.

Article Snippet: In brief, the medium of each group was centrifuged at 1500 rpm and 4 °C for 10 min, the concentration of NGF and BDNF in the supernatant was assessed using ELISA kits, the rat GDNF ELISA kit (EK0363, BOSTER, China) and the rat NGF/NGFβ ELISA kit (EK0471, BOSTER, China), and the absorbance of each well at 450 nm was determined using a spectrophotometer (EPOCH TAKE 3, Bio-Tek, USA).

Techniques: In Vitro, Double Staining, Immunofluorescence, Staining, CCK-8 Assay, Cell Culture, Expressing

Journal: Oncotarget

Article Title: Melatonin promotes goat spermatogonia stem cells (SSCs) proliferation by stimulating glial cell line-derived neurotrophic factor (GDNF) production in Sertoli cells

doi: 10.18632/oncotarget.12720

Figure Lengend Snippet: A. Light field of SSCs cultured with melatonin and GDNF, bar=50 μm. B. Cell density after being cultured with different cell mediums at 48 h. The initial number was 5*10 4 . C. QRT-PCR and western blot analysis of proliferation, self-renewal and Sertoli cell markers. D. Western blot analysis of proliferation and Sertoli cell markers. *, P<0.05,**, P<0.01.

Article Snippet: GDNF levels were determined by using a Human glial cell line-derived neurotrophic factor (GDNF) ELISA Kit (BOSTER).

Techniques: Cell Culture, Quantitative RT-PCR, Western Blot

Journal: Oncotarget

Article Title: Melatonin promotes goat spermatogonia stem cells (SSCs) proliferation by stimulating glial cell line-derived neurotrophic factor (GDNF) production in Sertoli cells

doi: 10.18632/oncotarget.12720

Figure Lengend Snippet: A. ELISA analysis of GDNF levels in the SSCs medium. B. Western Blot analysis of phosphorylation levels of AKT and ERK. *, P<0.05,**, P<0.01.

Article Snippet: GDNF levels were determined by using a Human glial cell line-derived neurotrophic factor (GDNF) ELISA Kit (BOSTER).

Techniques: Enzyme-linked Immunosorbent Assay, Western Blot, Phospho-proteomics