Journal: Pigment Cell & Melanoma Research

Article Title: β-Catenin regulates melanocyte dendricity through the modulation of PKCζ and PKCδ

doi: 10.1111/j.1755-148x.2010.00695.x

Figure Lengend Snippet: Figure 3. (A) Effect of b-catenin on the protein levels for dendrite formation- related factors in normal human epidermal melanocytes (NHEM). NHEM were transduced with 10 multiplicity of infection (MOI) of adenovirus expressing GFP-fused b-catenin (GFP-bCat) for 12 h, then replenished with fresh medium and further incubated for 3 days. Protein levels for Rho family GTPases (RhoA, Rac1 and Cdc42) and Rab27a were verified by Western blot analysis. The adenovirus expressing GFP was used as negative control. (B) Effect of b-catenin on the intracellular signaling pathway. Phosphorylation of p38 MAPK and ERK was determined by Western blot analysis. The protein levels for cell cycle-related proteins p53 and PCNA were also detected by Western blot analysis. (C) Effects of MAPK inhibitors on melanocyte dendricity. After adenoviral transduction, NHEM were treated with p38 MAPK inhibitor (SB203580, 10 lM) and MEK1 inhibitor (PD98059, 10 lM) for 3 days. (D) Knockdown by miR RNAi expression system with EmGFP. Cells were transduced with adenovirus expressing Em-GFP-miR-p38 MAPK and ⁄ or Em-GFP- miR-ERK1 ⁄ 2 for 12 h. Cells were replenished with fresh medium and further incubated for 3 days. Adenovirally transduced cells were verified under the fluorescent microscope. In EmGFP- miR-p38 MAPK-transduced cells, dendrites were markedly reduced (green arrows). While in EmGFP-miR-p38 MAPK-non- transduced cells, dendrites were not affected (red arrowheads). Adenovirus expressing EmGFP-miR-scrambled was used as negative control. Lower panels show the knockdown of p38 MAPK and ERK1 ⁄ 2 by Western blot.

Article Snippet: The following primary antibodies were used in this study: GFP, b-catenin, MITF, p53, PCNA, Rac1, Cdc42, Rab27a, RhoA, PKCf and PKCd (Santa Cruz Biotechnologies, Santa Cruz, CA, USA), phospho p38, total p38, phospho ERK, and total ERK (Cell Signaling Technology, Danvers, MA, USA), tyrosinase, TRP-1, and DCT (kindly provided by Dr Hearing VJ, NIH, Bethesda, MD), actin (Sigma).

Techniques: Transduction, Infection, Expressing, Incubation, Western Blot, Negative Control, Phospho-proteomics, Knockdown, Microscopy

Journal: eLife

Article Title: Genome-wide interrogation of extracellular vesicle biology using barcoded miRNAs

doi: 10.7554/eLife.41460

Figure Lengend Snippet:

Article Snippet: Antibody , rabbit polyclonal anti-Rab27A , Synaptic Systems , 168 013 , (1:1000).

Techniques: Recombinant, Software

Journal: CNS Neuroscience & Therapeutics

Article Title: Rab27a‐dependent exosomes protect against cerebral ischemic injury by reducing endothelial oxidative stress and apoptosis

doi: 10.1111/cns.13902

Figure Lengend Snippet: Ischemia increases Rab27a expression and brain EXs secretion. (A) Heat map of proteins involved in EXs biogenesis or secretion by mRNA sequence in brain tissue of control mouse and IS mouse. (B) Comparison of the top five elevated genes including Rab5a, Pdcd6ip, Ykt6, Rab27a, Rab5c by QRT‐PCR. Data were expressed as mean ± SEM ( n = 3 mice/group). (C) Rab27a and CD63 expression in brain of control mouse and IS mouse were detected by Western blotting. Data were expressed as mean ± SEM ( n = 3 mice/group). (D) The level of brain EXs were detected by structured illumination microscopy in WT mice after sham or tMCAO surgery. EXs (CD63, green), cerebral vessels (CD31, red). Data were expressed as mean ± SEM, n = 20 fields from 4 mice per group. (E) The level of brain EXs were detected by NTA assay in WT mice after sham or tMCAO surgery. Data were expressed as mean ± SEM, n = 20 fields from 4 mice per group. * p < 0.05, ** p < 0.01 compared with the sham group

Article Snippet: The male adult (8–10 weeks and weight 22–24 g) C57BL/ 6 WT and Rab27a knockout (Rab27a −/− ) mice were purchased from Cyagen Biosciences Company (Guangzhou, China) and housed in the Animal Care Facility at the Guangdong Medical University.

Techniques: Expressing, Sequencing, Control, Comparison, Quantitative RT-PCR, Western Blot, Microscopy

Journal: CNS Neuroscience & Therapeutics

Article Title: Rab27a‐dependent exosomes protect against cerebral ischemic injury by reducing endothelial oxidative stress and apoptosis

doi: 10.1111/cns.13902

Figure Lengend Snippet: Levels of brain EXs were significantly reduced in Rab27a −/− mice. (A) Rab27a expression in brain tissue of WT mice and Rab27a −/− mice was measured by Western blotting. (B) EXs‐specific marker CD63 and TSG101 were detected by Western blotting. (C) TEM was used to detect the size and morphology of EX WT or EX Rab27a−/− . (D) The concentration and size of EXs from brain tissue of WT mice (EX WT ) or Rab27a −/− mice (EX Rab27a−/− ) were detected by NTA. (E) The purification efficiency of CD63+ EXs in the total brain vesicles; data were expressed as mean ± SEM, n = 3 mice per group. (F) The level of brain EXs in WT mice and Rab27a −/− mice under normal or IS condition were observed using structured illumination microscopy. EXs (CD63, red), cerebral vessels (CD31, green). (G) The level of brain EXs in WT mice and Rab27a −/− mice under normal or IS condition were detected by using NTA. * p < 0.05, ** p < 0.01 compared with the WT group; ns p >0.05 compared to the sham group; all data were expressed as mean ± SEM, n = 20 fields from 4 mice per group

Article Snippet: The male adult (8–10 weeks and weight 22–24 g) C57BL/ 6 WT and Rab27a knockout (Rab27a −/− ) mice were purchased from Cyagen Biosciences Company (Guangzhou, China) and housed in the Animal Care Facility at the Guangdong Medical University.

Techniques: Expressing, Western Blot, Marker, Concentration Assay, Purification, Microscopy

Journal: CNS Neuroscience & Therapeutics

Article Title: Rab27a‐dependent exosomes protect against cerebral ischemic injury by reducing endothelial oxidative stress and apoptosis

doi: 10.1111/cns.13902

Figure Lengend Snippet: Rab27a −/− mice displayed aggravated cerebral ischemic injury. (A) Representative images and summarized data showing the infarct volume in WT mice and Rab27a −/− mice after tMCAO surgery. (B) Summarized data showing the NDS of WT mice and Rab27a −/− mice after tMCAO surgery. (C) Representative images and summarized data showing the CBF in WT mice and Rab27a −/− mice after tMCAO surgery. (D) Representative images and summarized data showing the cMVD in WT mice and Rab27a −/− mice after tMCAO surgery. ## p < 0.01 or ns p >0.05 compared to the sham group in WT mice; * p < 0.05, ** p < 0.01 compared with the tMCAO group in WT mice; ++ p < 0.01 compared with the sham group in Rab27a −/− mice; ns p > 0.05 compared with the sham group in WT mice. All data were expressed as mean ± SEM ( n = 10 mice/group)

Article Snippet: The male adult (8–10 weeks and weight 22–24 g) C57BL/ 6 WT and Rab27a knockout (Rab27a −/− ) mice were purchased from Cyagen Biosciences Company (Guangzhou, China) and housed in the Animal Care Facility at the Guangdong Medical University.

Techniques:

Journal: CNS Neuroscience & Therapeutics

Article Title: Rab27a‐dependent exosomes protect against cerebral ischemic injury by reducing endothelial oxidative stress and apoptosis

doi: 10.1111/cns.13902

Figure Lengend Snippet: Pre‐infusion of EXs increased the brain EXs level in Rab27a −/− mice. (A) Schematic depicting the workflow for the in vivo and in vitro experiments. (B) Immunofluorescence of CD31 (green) and CD63 (red) in the peri‐infarct area of Rab27a −/− mice using super‐resolution microscopy. Scale bar: 5 μm. (C) EXs levels in the peri‐infarct brain tissues in different EX infusion groups measured by super‐resolution microscopy and NTA. * p < 0.05, ** p < 0.01 compared with the vehicle group; # p < 0.05, ## p < 0.01 compared with the EX WT group; + p < 0.05, ++ p < 0.01 compared with the L‐EX Rab27a−/− group. All data were expressed as mean ± SEM, n = 20 fields from 4 mice per group

Article Snippet: The male adult (8–10 weeks and weight 22–24 g) C57BL/ 6 WT and Rab27a knockout (Rab27a −/− ) mice were purchased from Cyagen Biosciences Company (Guangzhou, China) and housed in the Animal Care Facility at the Guangdong Medical University.

Techniques: In Vivo, In Vitro, Immunofluorescence, Super-Resolution Microscopy

Journal: CNS Neuroscience & Therapeutics

Article Title: Rab27a‐dependent exosomes protect against cerebral ischemic injury by reducing endothelial oxidative stress and apoptosis

doi: 10.1111/cns.13902

Figure Lengend Snippet: Pre‐infusion of EXs decreased the cerebral vascular ROS overproduction and apoptosis in peri‐infarct area of Rab27a −/− mice. (A) Immunofluorescence of EX WT (PKH26, red) or PKH26‐PBS merged with ECs (CD31, green). Scale bar: 20 μm. (B) Immunofluorescence of ROS (DHE, red) with ECs (CD31, green) in peri‐infarct area of Rab27a −/− mice. Scale bar: 20 μm. (C) Immunofluorescence of apoptotic cells (Tunnel staining, red) with ECs (CD31, green) in peri‐infarct area of Rab27a −/− mice. Scale bar: 20 μm. (D) Summarized data showing the cerebral vascular ROS production in the peri‐infarct area of Rab27a −/− mice. (E) Summarized data showing the cerebral ECs apoptosis in the peri‐infarct area of Rab27a −/− mice. * p < 0.05, ** p < 0.01 compared to the vehicle group; # p < 0.05, ## p < 0.01 compared with the EX WT group; + p < 0.05, ++ p < 0.01 compared with the L‐EX Rab27a−/− group. All data were expressed as mean ± SEM ( n = 10 mice/group)

Article Snippet: The male adult (8–10 weeks and weight 22–24 g) C57BL/ 6 WT and Rab27a knockout (Rab27a −/− ) mice were purchased from Cyagen Biosciences Company (Guangzhou, China) and housed in the Animal Care Facility at the Guangdong Medical University.

Techniques: Immunofluorescence, Staining

Journal: CNS Neuroscience & Therapeutics

Article Title: Rab27a‐dependent exosomes protect against cerebral ischemic injury by reducing endothelial oxidative stress and apoptosis

doi: 10.1111/cns.13902

Figure Lengend Snippet: Pre‐infusion of EXs protected Rab27a −/− mice from ischemic injury. (A) Representative images and summarized data showing the infarct volume in tMCAO operated Rab27a −/− mice. (B) Summarized data showing the NDS of Rab27a −/− mice after tMCAO surgery. (C) Immunofluorescence of cMVD (CD31, green) in peri‐infarct area of Rab27a −/− mice. Scale bar: 40 μm. Summarized data showing the cMVD in the peri‐infarct area of Rab27a −/− mice. (D) Representative images and summarized data showing the CBF in tMCAO operated Rab27a −/− mice. * p < 0.05, ** p < 0.01 compared with the vehicle group; # p < 0.05, ## p < 0.01 compared with the EX WT group; + p < 0.05, ++ p < 0.01 compared with the L‐EX Rab27a−/− group. All data were expressed as mean ± SEM ( n = 10mice/group)

Article Snippet: The male adult (8–10 weeks and weight 22–24 g) C57BL/ 6 WT and Rab27a knockout (Rab27a −/− ) mice were purchased from Cyagen Biosciences Company (Guangzhou, China) and housed in the Animal Care Facility at the Guangdong Medical University.

Techniques: Immunofluorescence

Journal: CNS Neuroscience & Therapeutics

Article Title: Rab27a‐dependent exosomes protect against cerebral ischemic injury by reducing endothelial oxidative stress and apoptosis

doi: 10.1111/cns.13902

Figure Lengend Snippet: Pre‐incubation of EXs decreased ECs ROS overproduction and apoptosis under OGD condition. (A–C) Representative bands and summarized data showing the protein level of NOX and cleaved caspase‐3 in ECs. (D, F) Representative images and summarized data showing the apoptosis of ECs pre‐incubated with various EXs. (E, G) Representative images and summarized data showing the ROS production in ECs pre‐incubated with various EXs. − p < 0.05, − − p < 0.01 compared with the control group; * p < 0.05, ** p < 0.01 compared with the culture medium group; # p < 0.05, ## p < 0.01 compared with the EX WT group; + p < 0.05, ++ p < 0.01 compared with the L‐EX Rab27a−/− group. All data were expressed as mean ± SEM ( n = 3/group)

Article Snippet: The male adult (8–10 weeks and weight 22–24 g) C57BL/ 6 WT and Rab27a knockout (Rab27a −/− ) mice were purchased from Cyagen Biosciences Company (Guangzhou, China) and housed in the Animal Care Facility at the Guangdong Medical University.

Techniques: Incubation, Control