Journal: European Heart Journal

Article Title: Increased expression of pigment epithelium-derived factor in aged mesenchymal stem cells impairs their therapeutic efficacy for attenuating myocardial infarction injury ‡

doi: 10.1093/eurheartj/ehr131

Figure Lengend Snippet: Localization of engrafted mesenchymal stem cells in the infarcted heart. ( A ) The representative image of infarcted heart 1 day post-injection of adenoviral vector transduced mesenchymal stem cells (haematoxylin and eosin staining). ( B ) The fluorescent image of consecutive sections of Figure 3A . Many green cells were dispersed in the infarct region. ( C ) Bar graph shows no significant differences ( P = 0.708) in the green fluorescent intensity of infarcted hearts among Y& Ad.Null , Y& Ad.PEDF , O& Ad.shctrl, and O& Ad.shPEDF groups. Data expressed as means ± SEM ( n = 5 mice/group).

Article Snippet: Furthermore, PEDF levels were also determined by a human or mouse PEDF ELISA kit (USCN Life.

Techniques: Injection, Plasmid Preparation, Staining

Journal: European Heart Journal

Article Title: Increased expression of pigment epithelium-derived factor in aged mesenchymal stem cells impairs their therapeutic efficacy for attenuating myocardial infarction injury ‡

doi: 10.1093/eurheartj/ehr131

Figure Lengend Snippet: Mesenchymal stem cells alter the cellular profile in the myocardial infarction area through pigment epithelium-derived factor. ( A–D ) Representative confocal microscopic images of CD31- or vimentin-, or αSMA- or F4/80-positive cells (red fluorescence, white arrows point to representative positive cells) in infarct region 7 days after administration of saline, young mesenchymal stem cells, or older mesenchymal stem cells (bar: 20 µm). Nuclei were stained by DAPI (blue fluorescence). The bar graph shows the cell density ( n = 5 mice/group). ( E–H ) Representative confocal microscopic images of CD31- or vimentin- or αSMA- or F4/80-positive cells (red fluorescence) in infarct region 7 days after administration of Y& Ad.Null , Y& Ad.PEDF , O& Ad.shctrl, or O& Ad.shPEDF groups (bar: 20 µm). Injected mesenchymal stem cells were green fluorescent protein-positive (green fluorescence). Nuclei were stained by DAPI (blue fluorescence). White arrows point to representative CD31-, vimentin-, αSMA-, or F4/80 positive cells. Yellow arrows show representative CD31- or vimentin- or αSMA- or F4/80 and green fluorescent protein double positive cells. Bar graph shows the cell density ( n = 5 mice/group). All data are expressed as means ± SEM. * P < 0.05, ** P < 0.01, *** P ≤ 0.001.

Article Snippet: Furthermore, PEDF levels were also determined by a human or mouse PEDF ELISA kit (USCN Life.

Techniques: Derivative Assay, Fluorescence, Saline, Staining, Injection

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 2. PEDF mediates the anti-cellular senescence effects of C2C12-CM. (A–C) MEFs were cultured in the presence of C2C12-CM treated with a control or PEDF antibody for 3 days. (A) Cell numbers were counted and relative changes in cell numbers in 3 days were plotted. (B) Total RNA was isolated from MEFs and the expression of Ink4a and Arf was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (C) p16INK4a and p19ARF levels were analyzed by immunoblotting. β-Actin was used as the loading control. (D) MEFs were cultured in the presence of a recombinant of PEDF (100 ng/mL) for 3 days. Changes in cell numbers were plotted. (E) Cell viability was determined by the trypan blue exclusion assay. (F) The expression of Ink4a, Arf, and p21 was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (G) p16INK4a, p19ARF, and p21 levels were analyzed by immunoblotting. β-Actin was used as a loading control. (H) Cells were stained for SA-β-gal. Scale bar, 100 μm. (I) The percentage of SA-β-gal-positive cells was plotted. (J) Cells were stimulated with the indicated concentrations of recombinant PEDF for 3 days. Intracellular ROS levels were analyzed in each sample, and relative values were plotted against the average of the control sample. Values represent means ± SD. Data were analyzed by the Student’s t-test (A, B, D–F, I) or a one-way ANOVA and Tukey’s post-hoc analysis (J). *P <0.05, **P <0.01, and ***P <0.001.

Article Snippet: Samples were analyzed using a Mouse PEDF, PEDF ELISA Kit (CSB-E08820m, Cusabio Technology, LLC).

Techniques: Cell Culture, Control, Isolation, Expressing, Real-time Polymerase Chain Reaction, Western Blot, Recombinant, Trypan Blue Exclusion Assay, Staining

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 3. Voluntary exercise up-regulates PEDF and suppresses cellular senescence. Six-month-old wild-type mice were subjected to voluntary exercise for 8 weeks. (A) Total RNA was isolated from skeletal muscles (TA and SOL) and Pedf levels were analyzed by real-time PCR. Values were normalized to 18S rRNA in each sample. (B) Serum PEDF levels were analyzed by ELISA. (C) The expression of Ink4a, Arf and p21 in the lungs was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (D) Lung sections were stained for SA-β- gal. Arrowheads indicate SA-β-gal-positive cells. Scale bar, 100 μm. (E) The number of SA-β-gal-positive cells was counted in each sample. (F) The expression of Il-1b, Il-6, and Mmp-12 in lung total RNA was analyzed by real-time PCR. Values were normalized to 18S-rRNA in each sample. Values represent means ± SEM. Data were analyzed by the Student’s t-test. *P <0.05, **P <0.01, and ***P <0.001.

Article Snippet: Samples were analyzed using a Mouse PEDF, PEDF ELISA Kit (CSB-E08820m, Cusabio Technology, LLC).

Techniques: Isolation, Muscles, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Expressing, Staining

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 4. PEDF suppresses cellular senescence in lung tissues. A recombinant PEDF protein (10 μg/kg body weight) was intraperitoneally injected twice a week for 4 weeks. (A) The expression of Ink4a, Arf, and p21 in lung total RNA was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (B) Lung tissues were stained for SA-β-gal. Arrowheads indicate SA-β-gal-positive cells. Scale bar, 50 μm. (C) The number of SA-β-gal-positive cells was counted in each sample. Values represent means ± SEM. Data were analyzed by the Student’s t-test. *P <0.05 and **P <0.01.

Article Snippet: Samples were analyzed using a Mouse PEDF, PEDF ELISA Kit (CSB-E08820m, Cusabio Technology, LLC).

Techniques: Recombinant, Injection, Expressing, Real-time Polymerase Chain Reaction, Staining

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 5. PEDF protected lung tissues from PPE-induced emphysema. (A) Experimental design. A recombinant PEDF protein was injected intraperitoneally twice a week for 4 weeks. PPE (5 units) was intranasally administered 2 weeks after the first dose of PEDF. Mice were analyzed 3 weeks after the PPE treatment. (B) Total RNA was isolated from lung tissues and the expression of Ink4a, Arf, and p21 was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (C) Representative images of control and PEDF-treated mouse lung sections. Sections were stained with hematoxylin and eosin. Scale bar, 100 μm. (D) Alveolar mean linear intercepts were measured. (E–H) A pulmonary function test was performed. Lung compliance (E), capacity (F), tissue damping (G), and tissue elastance (H) are shown. Values represent means ± SEM. Data were analyzed by the Student’s t-test (B), or a one-way ANOVA and Tukey’s post-hoc analysis (D–H). *P <0.05, **P <0.01, and ***P <0.001.

Article Snippet: Samples were analyzed using a Mouse PEDF, PEDF ELISA Kit (CSB-E08820m, Cusabio Technology, LLC).

Techniques: Recombinant, Injection, Isolation, Expressing, Real-time Polymerase Chain Reaction, Control, Staining

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 6. Involvement of miR-127 in PEDF signaling. (A) An RNA sequencing analysis was performed on MEFs stimulated with a recombinant PEDF protein. A volcano plot of RNA sequencing data is shown. A full list of RNA sequencing data is available in the Gene Expression Omnibus database (accession number; GSE241459). (B) The expression of miR-127 (-loop, -5p, and -3p) in MEFs was analyzed by real-time PCR. Values were normalized to U6 snRNA in each sample. (C) The expression of BCL-6 in MEFs was analyzed by immunoblotting. β- Actin was used as a loading control. (D, E) miR-127 levels in the lungs of mice subjected to the PEDF treatment (D, Figure 4) or voluntary wheel running (E, Figure 3) were analyzed by real-time PCR. miR-127 levels were normalized to U6 snRNA in each sample. Values represent means ± SD (B) or means ± SEM (D, E). Data were analyzed by the Student’s t-test. *P <0.05, **P <0.01, and ***P <0.001.

Article Snippet: Samples were analyzed using a Mouse PEDF, PEDF ELISA Kit (CSB-E08820m, Cusabio Technology, LLC).

Techniques: RNA Sequencing, Recombinant, Gene Expression, Expressing, Real-time Polymerase Chain Reaction, Western Blot, Control

Journal: Viruses

Article Title: Comparative Characterization and Pathogenicity of a Novel Porcine Epidemic Diarrhea Virus (PEDV) with a Naturally Occurring Truncated ORF3 Gene Coinfected with PEDVs Possessing an Intact ORF3 Gene in Piglets

doi: 10.3390/v13081562

Figure Lengend Snippet: Virus isolation and purification of the PEDV strains with different ORF3 genotypes. CPE formation in Vero cells infected with the PEDV strains of ( a ) 17GXCZ-1ORF3d or ( b ) 17GXCZ-1ORF3c showing rounded and clustered at 48 hpi (200×). ( c ) Vero cells control (200×). ( d ) Detection and amplification of the ORF3 gene in PEDV strains. M: DL 2000 marker; Lane 1: negative control; Lane 2: 17GXCZ-1ORF3c (~740 bp); Lane 3: 17GXCZ-1ORF3d with a large genomic deletion (~358 bp); Lane 4: CV777 strain (~740 bp). The solid arrows indicate the predicted products (740 bp) and the dashed arrows indicate the products of the PEDV variants with a large genomic deletion (~358 bp).

Article Snippet: All animals were diagnosed as negative for PEDV, transmissible gastroenteritis virus (TGEV), rotavirus (PoRV), porcine deltacoronavirus (PDCoV), classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV) and pseudorabies virus (PRV) by a virus-specific RT-PCR analysis of rectal swabs and determined to be free of antibodies to PEDV by a commercial PEDV antibody ELISA kit (Biovet Inc., St-Hyacinthe, QC, Canada).

Techniques: Virus, Isolation, Purification, Infection, Control, Amplification, Marker, Negative Control

Journal: Viruses

Article Title: Comparative Characterization and Pathogenicity of a Novel Porcine Epidemic Diarrhea Virus (PEDV) with a Naturally Occurring Truncated ORF3 Gene Coinfected with PEDVs Possessing an Intact ORF3 Gene in Piglets

doi: 10.3390/v13081562

Figure Lengend Snippet: Plaque purification and viral growth kinetics of the PEDV strains 17GXCZ-1ORF3d and 17GXCZ-1ORF3c. Plaque purification of the PEDV strains ( a ) 17GXCZ-1ORF3d and ( b ) 17GXCZ-1ORF3c. Vero cell monolayers were inoculated with 10-fold serially diluted PEDV strains of 17GXCZ-1ORF3d and 17GXCZ-1ORF3c, respectively. After incubation for 1 h, the cells were overlaid with 1% agarose. Plaques were stained with crystal violet at 3–4 dpi. ( c ) The plaque diameters of 17GXCZ-1ORF3d and 17GXCZ-1ORF3c. The diameters of ten randomly selected plaques were measure using a ruler in the ImageJ 1.8.0 software. *** represents that the mean diameters of plaques formed by 17GXCZ-1ORF3d (1.18 mm) were significantly smaller than those of 17GXCZ-1ORF3c (1.70 mm), with p values of <0.001. ( d ) Viral growth kinetics of 17GXCZ-1ORF3d and 17GXCZ-1ORF3c in Vero cells. Vero cells were inoculated with the 17GXCZ-1ORF3d and 17GXCZ-1ORF3c strains at MOI = 0.001, respectively. The infected cells were collected at 6, 12, 24, 48, 60 and 72 hpi for virus titrations. The virus titers at each time point were determined in triplicate using plaque assays.

Article Snippet: All animals were diagnosed as negative for PEDV, transmissible gastroenteritis virus (TGEV), rotavirus (PoRV), porcine deltacoronavirus (PDCoV), classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV) and pseudorabies virus (PRV) by a virus-specific RT-PCR analysis of rectal swabs and determined to be free of antibodies to PEDV by a commercial PEDV antibody ELISA kit (Biovet Inc., St-Hyacinthe, QC, Canada).

Techniques: Purification, Incubation, Staining, Software, Infection, Virus

Journal: Viruses

Article Title: Comparative Characterization and Pathogenicity of a Novel Porcine Epidemic Diarrhea Virus (PEDV) with a Naturally Occurring Truncated ORF3 Gene Coinfected with PEDVs Possessing an Intact ORF3 Gene in Piglets

doi: 10.3390/v13081562

Figure Lengend Snippet: Comparisons of the nucleotide sequences of the genome organization of the 2017 Guangxi isolates and their genogroup representative PEDV strains.

Article Snippet: All animals were diagnosed as negative for PEDV, transmissible gastroenteritis virus (TGEV), rotavirus (PoRV), porcine deltacoronavirus (PDCoV), classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV) and pseudorabies virus (PRV) by a virus-specific RT-PCR analysis of rectal swabs and determined to be free of antibodies to PEDV by a commercial PEDV antibody ELISA kit (Biovet Inc., St-Hyacinthe, QC, Canada).

Techniques:

Journal: Viruses

Article Title: Comparative Characterization and Pathogenicity of a Novel Porcine Epidemic Diarrhea Virus (PEDV) with a Naturally Occurring Truncated ORF3 Gene Coinfected with PEDVs Possessing an Intact ORF3 Gene in Piglets

doi: 10.3390/v13081562

Figure Lengend Snippet: Genotyping and origin of the PEDV strains 17GXCZ-1ORF3d and 17GXCZ-1ORF3c based on different genes. Phylogenetic trees were constructed based on the aligned full-length genomes ( a ) S protein and ( b ) ORF3 protein ( c ) by using the neighbor-joining method from MEGA 5.2, with 1000 bootstrap replicates. Scale bars represent the branch lengths measured by the number of substitutions per site. Each PEDV strain is indicated in the following format: country of origin (three letter code: CHN, China; JPN, Japan; KOR, Korea; MEX, Mexico; SUI, Switzerland; and USA, the United States)/strain name/year of sample collection (Genbank accession number). Group 1, G2a subgroup, G2b subgroup and Group 3 are coded in red, green, pink and yellow, respectively. The triangle symbols represent the strains obtained in this study.

Article Snippet: All animals were diagnosed as negative for PEDV, transmissible gastroenteritis virus (TGEV), rotavirus (PoRV), porcine deltacoronavirus (PDCoV), classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV) and pseudorabies virus (PRV) by a virus-specific RT-PCR analysis of rectal swabs and determined to be free of antibodies to PEDV by a commercial PEDV antibody ELISA kit (Biovet Inc., St-Hyacinthe, QC, Canada).

Techniques: Construct

Journal: Viruses

Article Title: Comparative Characterization and Pathogenicity of a Novel Porcine Epidemic Diarrhea Virus (PEDV) with a Naturally Occurring Truncated ORF3 Gene Coinfected with PEDVs Possessing an Intact ORF3 Gene in Piglets

doi: 10.3390/v13081562

Figure Lengend Snippet: Pathogenicity analysis of the PEDV strains 17GXCZ-1ORF3d and 17GXCZ-1ORF3c. ( a ) The average body weight changes in each group. ( b ) Fecal scores of piglets in different groups. Rectal swabs were collected at different timepoints after PEDV infection, scores standard: 0, normal; 1, pasty stool; 2, semiliquid diarrhea; and 3, liquid diarrhea. ( c ) The survival rate of piglets in each group. After oral inoculation with PEDV, one piglet died at 96 hpi and one at 108 hpi in the 17GXCZ-1ORF3c group, while two piglets died at 96 hpi and one piglet died at 108 hpi in the 17GXCZ-1ORF3d group. Each data point in the graph represents the average value from four or all of the surviving animals ± SEM. The asterisks indicate significant differences between the groups of 17GXCZ-1ORF3d and 17GXCZ-1ORF3c (* p < 0.05; ** p < 0. 01; *** p < 0 .001). Gross lesions found in piglets of ( d ) 17GXCZ-1ORF3d, ( e ) 17GXCZ-1ORF3c and ( f ) mock groups at necropsy. The piglets were necropsied upon death, whereas all surviving piglets from the challenged and mock groups were euthanized at 5 dpi for post-mortem examinations.

Article Snippet: All animals were diagnosed as negative for PEDV, transmissible gastroenteritis virus (TGEV), rotavirus (PoRV), porcine deltacoronavirus (PDCoV), classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV) and pseudorabies virus (PRV) by a virus-specific RT-PCR analysis of rectal swabs and determined to be free of antibodies to PEDV by a commercial PEDV antibody ELISA kit (Biovet Inc., St-Hyacinthe, QC, Canada).

Techniques: Infection

Journal: Viruses

Article Title: Comparative Characterization and Pathogenicity of a Novel Porcine Epidemic Diarrhea Virus (PEDV) with a Naturally Occurring Truncated ORF3 Gene Coinfected with PEDVs Possessing an Intact ORF3 Gene in Piglets

doi: 10.3390/v13081562

Figure Lengend Snippet: Pathogenicity analysis of the PEDV strains 17GXCZ-1 ORF3d and 17GXCZ-1 ORF3c found in China, 2017 †.

Article Snippet: All animals were diagnosed as negative for PEDV, transmissible gastroenteritis virus (TGEV), rotavirus (PoRV), porcine deltacoronavirus (PDCoV), classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV) and pseudorabies virus (PRV) by a virus-specific RT-PCR analysis of rectal swabs and determined to be free of antibodies to PEDV by a commercial PEDV antibody ELISA kit (Biovet Inc., St-Hyacinthe, QC, Canada).

Techniques: Cell Culture

Journal: Viruses

Article Title: Comparative Characterization and Pathogenicity of a Novel Porcine Epidemic Diarrhea Virus (PEDV) with a Naturally Occurring Truncated ORF3 Gene Coinfected with PEDVs Possessing an Intact ORF3 Gene in Piglets

doi: 10.3390/v13081562

Figure Lengend Snippet: H&E and IHC staining of the jejunum from the PEDV-infected and mock piglets. Different tissue samples including the duodenum, jejunum, ileum, cecum, MLNs and stomach from each group were taken and then processed for H&E and IHC staining, and representative images of the jejunum are shown (200×). H&E staining of the jejunum in piglets from the ( a ) 17GXCZ-1ORF3d, ( b ) 17GXCZ-1ORF3c and ( c ) mock groups. The jejunum from the PEDV-infected piglets was characterized by the shortening, atrophy or even the shedding of intestinal villus. IHC staining of the jejunum from the ( d ) 17GXCZ-1ORF3d, ( e ) 17GXCZ-1ORF3c and ( f ) mock groups. PEDV antigen appears as a brown stain and was detected in the epithelial cells of the jejunum in 17GXCZ-1ORF3d and 17GXCZ-1ORF3c infected piglets. The intestinal tissue sections were stained with an anti-PEDV spike protein monoclonal antibody followed by the incubation with an HRP conjugated goat anti-mouse antibody and then visualized using a fluorescent microscope.

Article Snippet: All animals were diagnosed as negative for PEDV, transmissible gastroenteritis virus (TGEV), rotavirus (PoRV), porcine deltacoronavirus (PDCoV), classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV) and pseudorabies virus (PRV) by a virus-specific RT-PCR analysis of rectal swabs and determined to be free of antibodies to PEDV by a commercial PEDV antibody ELISA kit (Biovet Inc., St-Hyacinthe, QC, Canada).

Techniques: Immunohistochemistry, Infection, Staining, Incubation, Microscopy

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 2. PEDF mediates the anti-cellular senescence effects of C2C12-CM. (A–C) MEFs were cultured in the presence of C2C12-CM treated with a control or PEDF antibody for 3 days. (A) Cell numbers were counted and relative changes in cell numbers in 3 days were plotted. (B) Total RNA was isolated from MEFs and the expression of Ink4a and Arf was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (C) p16INK4a and p19ARF levels were analyzed by immunoblotting. β-Actin was used as the loading control. (D) MEFs were cultured in the presence of a recombinant of PEDF (100 ng/mL) for 3 days. Changes in cell numbers were plotted. (E) Cell viability was determined by the trypan blue exclusion assay. (F) The expression of Ink4a, Arf, and p21 was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (G) p16INK4a, p19ARF, and p21 levels were analyzed by immunoblotting. β-Actin was used as a loading control. (H) Cells were stained for SA-β-gal. Scale bar, 100 μm. (I) The percentage of SA-β-gal-positive cells was plotted. (J) Cells were stimulated with the indicated concentrations of recombinant PEDF for 3 days. Intracellular ROS levels were analyzed in each sample, and relative values were plotted against the average of the control sample. Values represent means ± SD. Data were analyzed by the Student’s t-test (A, B, D–F, I) or a one-way ANOVA and Tukey’s post-hoc analysis (J). *P <0.05, **P <0.01, and ***P <0.001.

Article Snippet: Human sera (information including age, sex, and pathology are listed in Supplementary Table 3) were purchased from ProteoGenex Inc. and PEDF levels were analyzed using a Human PEDF ELISA kit (RD191114200R, BioVender R&D).

Techniques: Cell Culture, Control, Isolation, Expressing, Real-time Polymerase Chain Reaction, Western Blot, Recombinant, Trypan Blue Exclusion Assay, Staining

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 3. Voluntary exercise up-regulates PEDF and suppresses cellular senescence. Six-month-old wild-type mice were subjected to voluntary exercise for 8 weeks. (A) Total RNA was isolated from skeletal muscles (TA and SOL) and Pedf levels were analyzed by real-time PCR. Values were normalized to 18S rRNA in each sample. (B) Serum PEDF levels were analyzed by ELISA. (C) The expression of Ink4a, Arf and p21 in the lungs was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (D) Lung sections were stained for SA-β- gal. Arrowheads indicate SA-β-gal-positive cells. Scale bar, 100 μm. (E) The number of SA-β-gal-positive cells was counted in each sample. (F) The expression of Il-1b, Il-6, and Mmp-12 in lung total RNA was analyzed by real-time PCR. Values were normalized to 18S-rRNA in each sample. Values represent means ± SEM. Data were analyzed by the Student’s t-test. *P <0.05, **P <0.01, and ***P <0.001.

Article Snippet: Human sera (information including age, sex, and pathology are listed in Supplementary Table 3) were purchased from ProteoGenex Inc. and PEDF levels were analyzed using a Human PEDF ELISA kit (RD191114200R, BioVender R&D).

Techniques: Isolation, Muscles, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Expressing, Staining

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 4. PEDF suppresses cellular senescence in lung tissues. A recombinant PEDF protein (10 μg/kg body weight) was intraperitoneally injected twice a week for 4 weeks. (A) The expression of Ink4a, Arf, and p21 in lung total RNA was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (B) Lung tissues were stained for SA-β-gal. Arrowheads indicate SA-β-gal-positive cells. Scale bar, 50 μm. (C) The number of SA-β-gal-positive cells was counted in each sample. Values represent means ± SEM. Data were analyzed by the Student’s t-test. *P <0.05 and **P <0.01.

Article Snippet: Human sera (information including age, sex, and pathology are listed in Supplementary Table 3) were purchased from ProteoGenex Inc. and PEDF levels were analyzed using a Human PEDF ELISA kit (RD191114200R, BioVender R&D).

Techniques: Recombinant, Injection, Expressing, Real-time Polymerase Chain Reaction, Staining

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 5. PEDF protected lung tissues from PPE-induced emphysema. (A) Experimental design. A recombinant PEDF protein was injected intraperitoneally twice a week for 4 weeks. PPE (5 units) was intranasally administered 2 weeks after the first dose of PEDF. Mice were analyzed 3 weeks after the PPE treatment. (B) Total RNA was isolated from lung tissues and the expression of Ink4a, Arf, and p21 was analyzed by real-time PCR. Values were normalized to Gapdh in each sample. (C) Representative images of control and PEDF-treated mouse lung sections. Sections were stained with hematoxylin and eosin. Scale bar, 100 μm. (D) Alveolar mean linear intercepts were measured. (E–H) A pulmonary function test was performed. Lung compliance (E), capacity (F), tissue damping (G), and tissue elastance (H) are shown. Values represent means ± SEM. Data were analyzed by the Student’s t-test (B), or a one-way ANOVA and Tukey’s post-hoc analysis (D–H). *P <0.05, **P <0.01, and ***P <0.001.

Article Snippet: Human sera (information including age, sex, and pathology are listed in Supplementary Table 3) were purchased from ProteoGenex Inc. and PEDF levels were analyzed using a Human PEDF ELISA kit (RD191114200R, BioVender R&D).

Techniques: Recombinant, Injection, Isolation, Expressing, Real-time Polymerase Chain Reaction, Control, Staining

Journal: Aging

Article Title: Roles of pigment epithelium-derived factor in exercise-induced suppression of senescence and its impact on lung pathology in mice.

doi: 10.18632/aging.205976

Figure Lengend Snippet: Figure 6. Involvement of miR-127 in PEDF signaling. (A) An RNA sequencing analysis was performed on MEFs stimulated with a recombinant PEDF protein. A volcano plot of RNA sequencing data is shown. A full list of RNA sequencing data is available in the Gene Expression Omnibus database (accession number; GSE241459). (B) The expression of miR-127 (-loop, -5p, and -3p) in MEFs was analyzed by real-time PCR. Values were normalized to U6 snRNA in each sample. (C) The expression of BCL-6 in MEFs was analyzed by immunoblotting. β- Actin was used as a loading control. (D, E) miR-127 levels in the lungs of mice subjected to the PEDF treatment (D, Figure 4) or voluntary wheel running (E, Figure 3) were analyzed by real-time PCR. miR-127 levels were normalized to U6 snRNA in each sample. Values represent means ± SD (B) or means ± SEM (D, E). Data were analyzed by the Student’s t-test. *P <0.05, **P <0.01, and ***P <0.001.

Article Snippet: Human sera (information including age, sex, and pathology are listed in Supplementary Table 3) were purchased from ProteoGenex Inc. and PEDF levels were analyzed using a Human PEDF ELISA kit (RD191114200R, BioVender R&D).

Techniques: RNA Sequencing, Recombinant, Gene Expression, Expressing, Real-time Polymerase Chain Reaction, Western Blot, Control

Journal: BMC Ophthalmology

Article Title: Quantitative proteomic analysis of aqueous humor from patients with drusen and reticular pseudodrusen in age-related macular degeneration

doi: 10.1186/s12886-018-0941-9

Figure Lengend Snippet: Differential secretion of selected proteins in hfRPE cell cultures. Total protein concentrations in conditioned media from apical and basal baths (left end). To confirm the biological activity of VEGF, PEDF, and CFH, apical or basal media were collected at 24 h and then analyzed using ELISAs (the rest three graphs). The amount of VEGF protein in the basal media was higher than that in the apical media (2.86 ng/mL vs. 1.51 ng/mL) and was greatly decreased by 100 μM 4-HNE treatment (0.74 ng/mL vs. 0.51 ng/mL). PEDF secretion on the apical side was higher than that on the basal side (4.51 μg/mL vs. 1.74 μg/mL), and the level of PEDF was significantly decreased in a dose-dependent manner at 24 h (0.086 μg/mL and 0.32 μg/mL, in apical and basal baths treated with 100 μM 4-HNE). The ratio of the PEDF concentrations secreted in the apical and basal chambers was reversed in cultures treated with 100 μM 4-HNE. The apical/basal ratios were 2.59 and 1.83 in control cultures and cultures exposed to 50 μM 4-HNE for 24 h; the apical/basal ratio was 0.27 in cultures exposed to 100 μM 4-HNE for 24 h. The level of CFH showed similar changes: it decreased both apically and basally in cultures exposed to oxidative stress compared to control cultures. The apical/basal ratios were 4.19 and 2.64 in control cultures and cultures exposed to 50 μM 4-HNE for 24 h, respectively; the apical/basal ratio was 1.41 in cultures exposed to 100 μM 4-HNE for 24 h

Article Snippet: The levels of VEGF-A (catalog number: DVE00; R&D Systems, MN, USA), PEDF (catalog number: PED613; BioProducts MD, LLC, MD, USA), and complement factor H (CFH) (catalog number: HK342–02; Hycultbiotech, Uden, Netherlands) in conditioned media from hfRPE cells were quantitatively assessed using a sandwich ELISA kit.

Techniques: Activity Assay