Journal: International Journal of Neuropsychopharmacology

Article Title: Mesenchymal Stem Cell–Derived Extracellular Vesicles Alleviate M1 Microglial Activation in Brain Injury of Mice With Subarachnoid Hemorrhage via microRNA-140-5p Delivery

doi: 10.1093/ijnp/pyab096

Figure Lengend Snippet: Activin-like kinase 5 (ALK5) silencing reduces NADPH oxidase 2 (NOX2) expression to contribute to suppression of M1 microglia activation in subarachnoid hemorrhage (SAH) mice. A, Quantification of CD16-positive M1 microglia and CD206-positive M2 microglia. B, Western blot analysis of NOX2 protein expression in SAH mice after silencing ALK5. * P <.05 vs sham-operated mice; # P <.05 vs SAH mice treated with shRNA negative control (sh-NC). C, NOX2 knockdown efficiency confirmed by western blot analysis along with quantification of CD16-positive M1 microglia and CD206-positive M2 microglia. * P <.05 vs wild type (WT) mice. D, Western blot analysis of ALK5 protein expression in NOX2−/− mice pretreated with oe-ALK5 lentiviral vector. E, Quantification of CD16-positive M1 microglia and CD206-positive M2 microglia. n = 6. * P <.05 vs NOX2−/− mice pretreated with vector. The data were measurement data, which were expressed by mean ± standard deviation. The comparison between the two groups was analyzed by unpaired t-test. Comparison among multiple groups was analyzed by one-way variance of analysis.

Article Snippet: Then, sections/cells were incubated with primary antibodies to CD16 (1:200, A13980, ABclonal), CD206 (1:100, ab195191, Abcam), and Iba 1 (1:100, A12391, ABclonal) overnight at 4°C, and incubated with corresponding fluorescent secondary antibodies (AS011 and AS007, ABclonal).

Techniques: Expressing, Activation Assay, Western Blot, shRNA, Negative Control, Knockdown, Plasmid Preparation, Standard Deviation, Comparison

Journal: International Journal of Neuropsychopharmacology

Article Title: Mesenchymal Stem Cell–Derived Extracellular Vesicles Alleviate M1 Microglial Activation in Brain Injury of Mice With Subarachnoid Hemorrhage via microRNA-140-5p Delivery

doi: 10.1093/ijnp/pyab096

Figure Lengend Snippet: miR-140-5p shuttled by MSC-derived EVs represses brain injury and microglial M1 activation in mice after subarachnoid hemorrhage (SAH). A, The expression of miR-140-5p in EVs from MSCs transfected with miR-140-5p mimic or miR-140-5p inhibitor detected by real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR). * P <.05 vs EVs from mimic-NC-transfected MSCs. # P <.05 vs EVs from inhibitor-NC-transfected MSCs. Sham-operated mice were used as control, and SAH mice were treated or not treated with EVs or EVs-miR-140-5p. B, Quantification of neurological function score (n = 6). C, TdT-mediated dUTP-biotin nick end labeling with NeuN (TUNEL/NeuN) immunofluorescence staining of the cerebral cortex (scale bar = 100 μm; DAPI: blue; TUNEL: green; NeuN: red) and the number of the TUNEL/NeuN positive cells (n = 6). D, The expression of IL-6, IL-1β, and TNF-α in serum detected by enzyme-linked immunosorbent assay (ELISA) (n = 6). E, Representative immunofluorescence micrographs of microglia polarization the cerebral cortex and the number of CD16-positive M1 microglia and CD206-positive M2 microglia (scale bar = 25 μm; CD16/CD206: red, Iba 1: green, DAPI: blue). F, The expression of miR-140-5p in SAH mice detected by RT-qPCR. G, Western blot analysis of the expression of ALK5 and NADPH oxidase 2 (NOX2) in SAH mice. * P <.05 vs SAH mice; # P <.05 vs SAH mice treated with EVs. The data were measurement data, which were expressed by mean ± standard deviation. Comparison among multiple groups was analyzed by one-way variance of analysis. n = 6..

Article Snippet: Then, sections/cells were incubated with primary antibodies to CD16 (1:200, A13980, ABclonal), CD206 (1:100, ab195191, Abcam), and Iba 1 (1:100, A12391, ABclonal) overnight at 4°C, and incubated with corresponding fluorescent secondary antibodies (AS011 and AS007, ABclonal).

Techniques: Derivative Assay, Activation Assay, Expressing, Transfection, Reverse Transcription, Polymerase Chain Reaction, Quantitative RT-PCR, Control, End Labeling, TUNEL Assay, Immunofluorescence, Staining, Enzyme-linked Immunosorbent Assay, Western Blot, Standard Deviation, Comparison

Journal: International Journal of Neuropsychopharmacology

Article Title: Mesenchymal Stem Cell–Derived Extracellular Vesicles Alleviate M1 Microglial Activation in Brain Injury of Mice With Subarachnoid Hemorrhage via microRNA-140-5p Delivery

doi: 10.1093/ijnp/pyab096

Figure Lengend Snippet: miR-140-5p shuttled by MSC-EVs diminishes Activin-like kinase 5 (ALK5) expression to suppress brain injury and microglial M1 activation in subarachnoid hemorrhage (SAH) mice. SAH mice were treated with EVs, EVs-miR-140-5p, or EVs-miR-140-5p + ALK5. A, The expression of IL-6, IL-1β, and TNF-α in serum detected by enzyme-linked immunosorbent assay (ELISA) (n = 6). B, The number of CD16-positive M1 microglia and CD206-positive M2 microglia. C, The expression of miR-140-5p in SAH mice detected by real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR). D, Western blot analysis of the expression of ALK5 and NADPH oxidase 2 (NOX2) in SAH mice. * P <.05 vs SAH mice treated with EVs; # P <.05 vs SAH mice treated with EVs-miR-140-5p. The data were measurement data, which were expressed by mean ± standard deviation. Comparison among multiple groups was analyzed by one-way variance of analysis. n = 6..

Article Snippet: Then, sections/cells were incubated with primary antibodies to CD16 (1:200, A13980, ABclonal), CD206 (1:100, ab195191, Abcam), and Iba 1 (1:100, A12391, ABclonal) overnight at 4°C, and incubated with corresponding fluorescent secondary antibodies (AS011 and AS007, ABclonal).

Techniques: Expressing, Activation Assay, Enzyme-linked Immunosorbent Assay, Reverse Transcription, Polymerase Chain Reaction, Quantitative RT-PCR, Western Blot, Standard Deviation, Comparison

Journal: Oncotarget

Article Title: Exosomal miR-141-3p regulates osteoblast activity to promote the osteoblastic metastasis of prostate cancer

doi: 10.18632/oncotarget.22014

Figure Lengend Snippet: (A) pRNAT-U6.1/Neo expression vector. (B) Two shRNA constructs and the corresponding primers. (C) Two shRNA constructs stably silence DLC1 in osteoblasts. (D) DLC1 KD activated RhoA and CDC42 and affected p38MAPK phosphorylation. (E) CDC42 inhibition significantly affected p38MAPK phosphorylation.

Article Snippet: The blotting membrane was blocked with bovine serum albumin and incubated with rabbit monoclonal anti-CD63 antibody (1:1000; ab134045; Abcam), rabbit monoclonal anti-GM130 (1:1000; ab52649; Abcam), rabbit polyclonal anti-DLC1 antibody (1:1000; ab126257; Abcam), rabbit polyclonal Cdc42 antibody (1:1000; A13984; Thermo Fisher), rabbit polyclonal anti-P38 antibody (1:1000; ab27986; Abcam), rabbit phospho-p38 MAPK alpha polyclonal antibody (1:1000; 44-684G; Thermo Fisher), rabbit polyclonal anti-RhoA antibody (1:500; ab86297; Abcam) and RhoA/Rac1/Cdc42 Combo Activation Assay (1:1000; ab211168; Abcam), followed by incubation with horseradish peroxidase (HRP)-coupled goat anti-rabbit IgG H&L (1:5000; ab6721; Abcam) or HRP-coupled goat anti-mouse IgG H&L (1:5000; ab97040; Abcam).

Techniques: Expressing, Plasmid Preparation, shRNA, Construct, Stable Transfection, Inhibition

Journal: Antioxidants

Article Title: Superoxide-Mediated Upregulation of MMP9 Participates in BMPR2 Destabilization and Pulmonary Hypertension Development

doi: 10.3390/antiox12111961

Figure Lengend Snippet: ( a ) Western blot analysis for BMPR2 expression in lung tissues from wild-type and MMP9 KO mice exposed to normoxia or Sugen plus hypoxia. ( b ) Western blot analysis for COMP expression in lung tissues from wild-type and MMP9 KO mice exposed to normoxia or Sugen plus hypoxia. ( n = 4–7) * p < 0.05 vs. Nor (WT) using one-way ANOVA with Tukey’s multiple comparison test.

Article Snippet: Specific antibodies were purchased from the companies indicated: anti-MMP9 (Abcam, Cambridge, MA, USA: ab38898), anti-BMPR2 (Abclonal, Woburn, MA, USA: A16778), anti-COMP (Abclonal, Woburn, MA, USA: A13963) anti-GC (Sigma, Burlington, MA, USA: G4405), anti-SIRT3 (Abcam, Cambridge, MA: ab86671), anti-SOD2 (Abclonal, Woburn, MA, USA: A1340) and anti-β-actin (Sigma, Burlington, MA, USA: A5441).

Techniques: Western Blot, Expressing

Journal: World Journal of Gastroenterology

Article Title: Nimbolide inhibits tumor growth by restoring hepatic tight junction protein expression and reduced inflammation in an experimental hepatocarcinogenesis

doi: 10.3748/wjg.v26.i45.7131

Figure Lengend Snippet: Effect of nimbolide on liver function parameters and tumor and cell proliferation markers in diethylnitrosamine and N-nitrosomorpholine induced hepatocellular carcinoma mice. A: Plasma aspartate aminotransferase level in naïve and experimental groups; B: Plasma alanine aminotransferase level in naïve and experimental groups; C: Plasma alkaline phosphatase level in naïve and experimental groups; D: Plasma alpha-fetoprotein level in naïve and experimental groups; E: Representative images of glypican-3 immunostaining of mice liver from naive and experimental groups (200 × magnification); F: Quantification of glypican-3 positive cells in experimental groups by counting five 400 × fields of each liver section; G: Representative images of proliferating hepatocytes by proliferating cell nuclear antigen (PCNA) immunostaining of mice liver from naive and experimental groups (200 × magnification); H: Quantification of PCNA positive nuclei in experimental groups by counting five 400 × fields of each liver section. Scale bar: 200 μm; all the data are expressed as mean ± SEM ( n = 3-6). The comparison between the groups was analyzed by one-way ANOVA followed by Tukey’s multiple comparison post-hoc test or Kruskal-Wallis followed by Dunn’s multiple comparison post-hoc test. c P < 0.0001 compared to naïve group; d P < 0.05, e P < 0.01 compared to hepatocellular carcinoma group. AST: Aspartate aminotransferase; ALT: Alanine aminotransferase; ALP: Alkaline phosphatase; AFP: Alpha-fetoprotein; PCNA: Proliferating cell nuclear antigen; HPF: High-power fields; HCC: Hepatocellular carcinoma.

Article Snippet: Primary antibodies used were rabbit polyclonal anti-glypican-3 (1:100, A13988; Abclonal, Woburn, MA, United States), rabbit polyclonal anti-PCNA (1:100, A0264; Abclonal, Woburn, MA, United States), and rabbit polyclonal anti-4-Hydroxynonenal (4HNE) (1:100, bs-6313R; Bioss, Woburn, Massachusetts, United States).

Techniques: Immunostaining