Journal: Neural Regeneration Research

Article Title: Neural stem cell-derived exosome as a nano-sized carrier for BDNF delivery to a rat model of ischemic stroke

doi: 10.4103/1673-5374.346466

Figure Lengend Snippet: The features of hNSCs and engineered exosomes (Exo) . (A) Immunofluorescence showed the expression of Nestin (red), Sox2 (green), MOG (red), Tuj1 (red), and GFAP (green). Nuclei were stained by DAPI (blue). Scale bars: 100 μm in the MOG panel; 50 μm in the Tuj1 panel; 20 μm in Nestin, Sox2 and GFAP panels. (B) hNSC-Exo and BDNF-hNSC-Exo showed cup-shaped exosome morphology under TEM. Scale bars: 200 μm. (C) NTA analysis indicated a similar size range for hNSC-Exo and BDNF-hNSC-Exo. (D) Western blot analysis of exosomal marker proteins including CD81, HSP70, and TSG101 and calnexin (as a negative control). (E) Representative micrographs of exosome uptake in NSCs incubated with PKH67-labeled exosomes (green) for 48 hours. Nuclei were stained by DAPI (blue). Scale bars: 20 μm. BDNF: Brain-derived neurotrophic factor; GFAP: glial fibrillary acidic protein; hNSC: human neural stem cell; HSP70: heat shock protein 70; MOG: myelin oligodendrocyte glycoprotein; TSG101: tumor susceptibility 101; Tuj1: β-tubulin III.

Article Snippet: Fluorescent labeling of hNSC-Exo and BDNF-hNSC-Exo was performed using the PKH67 Green Fluorescent Cell Linker Kit (MilliporeSigma, Cat# MINI67) following the manufacturer’s instructions.

Techniques: Immunofluorescence, Expressing, Staining, Western Blot, Marker, Negative Control, Incubation, Labeling, Derivative Assay

Journal: Neural Regeneration Research

Article Title: Neural stem cell-derived exosome as a nano-sized carrier for BDNF delivery to a rat model of ischemic stroke

doi: 10.4103/1673-5374.346466

Figure Lengend Snippet: BDNF-hNSC-Exo promotes the survival and differentiation of neural stem cells in vitro . (A) The expression level of BDNF was measured by western blotting after the addition of BDNF-hNSC-Exo to cultured neural stem cells. Data were normalized to β-actin expression. (B) The differentiation efficiency of BDNF-hNSC-Exo was evaluated using quantitative reverse transcription-polymerase chain reaction. Data were normalized to GAPDH mRNA expression. Cells were treated with a 500 μM solution H 2 O 2 for 5 hours. (C) Western blot analysis of Tuj1 in neural stem cells. Cells were treated with a 500 μM solution H 2 O 2 for 5 hours. Data were normalized to GAPDH expression. (D) Immunofluorescence showed that the ratio of Tuj1-positive cells (red) in the BDNF-hNSC-Exo group was higher than that in the control group. The percentage of Tuj1-positive cells was obtained by counting cells in three visual fields. Cells were treated with a 500 μM solution H 2 O 2 for 5 hours. Nuclei were stained by DAPI (blue). Scale bars: 50 μm. (E) BDNF-hNSC-Exo remarkably increased cell survival in the H 2 O 2 stress model (CCK8 assay). Cells were treated with a 500 μM solution H 2 O 2 for 5 hours. (F) TUNEL-positive cells. The proportion of TUNEL-positive cells in the BDNF-hNSC-Exo group was remarkably lower than that in the PBS group. Cells were treated with a 500 μM solution H 2 O 2 for 5 hours. Nuclei were stained by DAPI (blue). Scale bars: 50 μm. (G) Western blotting showed that the expressions of apoptosis-related proteins cleaved-caspase-3 and Bax were significantly decreased in the BDNF-hNSC-Exo group compared with the other two groups. Cells were treated with a 500 μM solution H 2 O 2 for 5 hours. Data were normalized to GAPDH expression. All experiments were conducted in triplicate. All data are presented as mean ± SEM. * P < 0.05 (one-way analysis of variance followed by Tukey’s post hoc test). Bax: Bcl2 associated X, apoptosis regulator; BDNF: brain-derived neurotrophic factor; Exo: exosomes; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; hNSC: human neural stem cell; Tuj1: β-tubulin III.

Article Snippet: Fluorescent labeling of hNSC-Exo and BDNF-hNSC-Exo was performed using the PKH67 Green Fluorescent Cell Linker Kit (MilliporeSigma, Cat# MINI67) following the manufacturer’s instructions.

Techniques: In Vitro, Expressing, Western Blot, Cell Culture, Reverse Transcription Polymerase Chain Reaction, Immunofluorescence, Staining, CCK-8 Assay, TUNEL Assay, Derivative Assay

Journal: Neural Regeneration Research

Article Title: Neural stem cell-derived exosome as a nano-sized carrier for BDNF delivery to a rat model of ischemic stroke

doi: 10.4103/1673-5374.346466

Figure Lengend Snippet: BDNF-hNSC-Exo improves neurological deficits and brain damage in rats following cerebral ischemia . (A) Experimental design. (B) BDNF-hNSC-Exo increases the expression of BDNF in the infarct areas. Data were normalized to GAPDH expression. (C–E) Evaluation of the behavioral function of MCAO rats at 1, 3, 7, 14, and 28 days after BDNF-hNSC-Exo treatment by rotarod, postural reflex, and mNSS tests. n = 7–10 rats/group. (F) TTC staining showed that BDNF-hNSC-Exo significantly reduced the volume of cerebral infarction. White indicates infact region. n = 3 rats/group. All data are mean ± SEM. * P < 0.05 (one-way analysis of variance followed by Tukey’s post hoc test). BDNF: Brain-derived neurotrophic factor; Exo: exosomes; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; hNSC: human neural stem cell; MCAO: middle cerebral artery occlusion; mNSS: modified neurological severity score; PBS: phosphate-buffered saline; TTC: 2,3,5-triphenyltetrazolium chloride.

Article Snippet: Fluorescent labeling of hNSC-Exo and BDNF-hNSC-Exo was performed using the PKH67 Green Fluorescent Cell Linker Kit (MilliporeSigma, Cat# MINI67) following the manufacturer’s instructions.

Techniques: Expressing, Staining, Derivative Assay, Modification

Journal: Neural Regeneration Research

Article Title: Neural stem cell-derived exosome as a nano-sized carrier for BDNF delivery to a rat model of ischemic stroke

doi: 10.4103/1673-5374.346466

Figure Lengend Snippet: BDNF-hNSC-Exo inhibit neuroinflammation and promote neurogenesis in the peri-infarct zone of rats . (A) Immunofluorescence double-labeling demonstrated that more functional neurons in the peri-infarct area were generated in the BDNF-hNSC-Exo and hNSC-Exo groups than in the PBS group. BrdU (red) co-localized with Tuj1 (green). Nuclei were stained by DAPI (blue). Scale bar: 20 μm. n = 5 rats/group. (B) Double immunofluorescence staining showed that in the BDNF-hNSC-Exo group, the proportion of BrdU/GFAP double-positive cells in the peri-infarct area was lower than that in the hNSC-Exo group and the PBS group on day 28 after treatment. BrdU (red) co-localized with GFAP (green). Nuclei were stained by DAPI (blue). Scale bar: 20 μm. n = 5 rats/group. (C) Immunofluorescence showed that BDNF-hNSC-Exo significantly reduced the expression of Iba1 (red), indicating reduced neuroinflammation. Nuclei were stained by DAPI (blue). Scale bar: 20 μm. n = 5 rats/group. All data are shown as the mean ± SEM. * P < 0.05 (one-way analysis of variance followed by Tukey’s post hoc test). BDNF: Brain-derived neurotrophic factor; BrdU: bromodeoxyuridine; Exo: exosomes; GFAP: glial fibrillary acidic protein; hNSC: human neural stem cell; Iba1: induction of brown adipocytes 1; MCAO: middle cerebral artery occlusion; PBS: phosphate-buffered saline; Tuj1: β-tubulin.

Article Snippet: Fluorescent labeling of hNSC-Exo and BDNF-hNSC-Exo was performed using the PKH67 Green Fluorescent Cell Linker Kit (MilliporeSigma, Cat# MINI67) following the manufacturer’s instructions.

Techniques: Immunofluorescence, Labeling, Functional Assay, Generated, Staining, Double Immunofluorescence Staining, Expressing, Derivative Assay

Journal: PLoS ONE

Article Title: Omega-3 Fatty Acid Deficiency during Brain Maturation Reduces Neuronal and Behavioral Plasticity in Adulthood

doi: 10.1371/journal.pone.0028451

Figure Lengend Snippet: Reductions in plasma membrane DHA may disrupt signaling of membrane embedded receptors, such as the BDNF receptor TrkB ( B) and the insulin receptor (IR; A). Dysfunction in TrkB signaling may influence downstream BDNF cascades such as CaMKII and CREB ( C), leading to further dysfunction in the BDNF system, and ultimately increasing vulnerability for anxiety-like behavior. Given the interaction between foods, BDNF, synaptic plasticity, and metabolic pathways, the n-3 dietary deficiency may disrupt events related to the insulin receptor (IR) signaling pathways elements such as the IR substrate IRS-1, Akt and mTOR ( B), which in turn, can affect BDNF-related synaptic plasticity leading to increased anxiety-behavior. Our results show that metabolic and behavioral pathways are both impacted by DHA deficiency.

Article Snippet: Protein samples were separated by electrophoresis on a 10% (12.5% for BDNF) polyacrylamide gel and electrotransferred to a PVDF or nitrocellulose membrane (Millipore, Bedford, MA).

Techniques:

Journal: Molecular Brain

Article Title: Activity-dependent brain-derived neurotrophic factor expression regulates cortistatin-interneurons and sleep behavior

doi: 10.1186/1756-6606-4-11

Figure Lengend Snippet: Effect of sleep deprivation on Bdnf transcripts and protein levels . (A). Relative levels of individual Bdnf transcripts at baseline (Ctrl) and after 12 h of sleep deprivation (SD) in WT and BDNF-KIV animals. Data are quantified and presented as in Figure 1 (n = 3 each condition). (B). Levels of total BDNF protein at baseline (Ctrl) and after 12 h of sleep deprivation (SD) in WT and BDNF-KIV animals. Data is presented at % induction over WT-Ctrl (n = 4 each group).

Article Snippet: Protein levels were normalized using a standard BCA assay (Pierce) and then used for measurement of BDNF protein levels with a commercial BDNF ELISA according to the manufacturer's instructions (Millipore, Billerica, MA).

Techniques:

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Acupuncture Can Play an Antidepressant Role by Regulating the Intestinal Microbes and Neurotransmitters in a Rat Model of Depression

doi: 10.12659/MSM.929027

Figure Lengend Snippet: Statistical graph of the expression of β-CaMKII/NMDAR/BDNF mRNA in the hippocampus of rats. ( A ) Expression of BDNF and β-CaMKII mRNA in the hippocampus by reverse transcription polymerase chain reaction (RT-PCR). ( B ) Expression of NMDAR mRNA in the hippocampus by RT-PCR. ( C ) Analysis of BDNF mRNA. ( D ) Analysis of β-CaMKII mRNA. ( E ) Analysis of NMDAR mRNA. Compared with the control (Con) group, * P <0.05, ** P <0.01, *** P <0.001; compared with the chronic unpredictable mild stress (CUMS) group, # P <0.05, ## P <0.01, ### P <0.001. Data represent mean±standard error of the mean. AP – acupuncture; BDNF – brain-derived neurotrophic factor; β-CaMKII – β isoform of calcium/calmodulin-dependent protein kinase II; Fx – fluoxetine; NMDAR – N-methyl-d-aspartate receptor.

Article Snippet: Diluted anti-BDNF (1: 2000, Millipore, USA), anti-β-CaMKII (1: 2000, Santa Cruz, USA), and anti-NMDAR (1: 2000, Millipore, USA) antibodies were added to the membrane, which was subsequently incubated overnight at 4°C.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Derivative Assay

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Acupuncture Can Play an Antidepressant Role by Regulating the Intestinal Microbes and Neurotransmitters in a Rat Model of Depression

doi: 10.12659/MSM.929027

Figure Lengend Snippet: Statistical graph of β-CaMKII/NMDAR/BDNF protein expression in the rat hippocampus. ( A ) Expression of BDNF proteins in the hippocampus by western blotting. ( B ) Expression of β-CaMKII proteins in the hippocampus by western blotting. ( C ) Expression of NMDAR proteins in the hippocampus by western blotting. ( D ) Analysis of BDNF proteins. ( E ) Analysis of β-CaMKII proteins. ( F ) Analysis of NMDAR proteins. Compared with the control (Con) group, * P <0.05, ** P <0.01, *** P <0.001; compared with the chronic unpredictable mild stress (CUMS) group, # P <0.05, ## P <0.01, ### P <0.001. Data represent mean±standard error of the mean. AP – acupuncture; BDNF – brain-derived neurotrophic factor; β-CaMKII – β isoform of calcium/calmodulin-dependent protein kinase II; Fx – fluoxetine; NMDAR – N-methyl-d-aspartate receptor.

Article Snippet: Diluted anti-BDNF (1: 2000, Millipore, USA), anti-β-CaMKII (1: 2000, Santa Cruz, USA), and anti-NMDAR (1: 2000, Millipore, USA) antibodies were added to the membrane, which was subsequently incubated overnight at 4°C.

Techniques: Expressing, Western Blot, Derivative Assay