Journal: Investigative Ophthalmology & Visual Science

Article Title: PTEN Inhibition Accelerates Corneal Endothelial Wound Healing through Increased Endothelial Cell Division and Migration

doi: 10.1167/iovs.61.8.19

Figure Lengend Snippet: Differential expressions of PTEN in the human and rat corneal endothelium. ( a ) Immunofluorescence staining of PTEN ( red ) and actin detected by phalloidin antibody ( green ) in human and rat corneas. Cell nuclei were stained with DAPI ( blue ). Scale bar = 20 µm. N = 3. ( b ) Flat-mount staining of PTEN ( red ) in human and rat corneal endothelium. Cell nuclei were stained with DAPI ( blue ). Scale bar = 10 µm. N = 3. ( c, d ) Total protein was extracted from the corneal endothelium detached from human corneoscleral rims and rat corneas. Cell lysates were analyzed by western blotting and immunoblotted using an anti-PTEN antibody. Data are presented as the mean ± SD from three independent experiments and plotted as PTEN protein expression compared with the respective GAPDH control. ** P < 0.01 by the Student's t -test. ( e ) Total protein extracted from human ( N = 9) and rat ( N = 6) corneal endothelium tissue were analyzed by ELISA. Data are presented as the mean ± SD from three independent experiments and plotted as concentration of PTEN in the corneal endothelium (ng/mL). *** P < 0.001 by the Student's t -test.

Article Snippet: The quantity of PTEN in human and rat corneal endothelium tissues was measured using Human and Rat PTEN ELISA Kits (ELK Biotechnology, Wuhan, China), respectively, according to the manufacturer protocol.

Techniques: Immunofluorescence, Staining, Western Blot, Expressing, Control, Enzyme-linked Immunosorbent Assay, Concentration Assay

Journal: Investigative Ophthalmology & Visual Science

Article Title: PTEN Inhibition Accelerates Corneal Endothelial Wound Healing through Increased Endothelial Cell Division and Migration

doi: 10.1167/iovs.61.8.19

Figure Lengend Snippet: Inhibition of PTEN with bpV(pic) contributed to alleviate the TGF-β2-induced G1-arrest. Cell cycle synchronization of B4G12 cells was achieved by starvation in DMEM without FBS for 24 hours. B4G12 cells were then treated with 10 ng/mL TGF-β2, 10 µM bpV(pic), or the combination of the two supplemented in DMEM with 5% FBS. ( a ) After treatment for another 24 hours, cells were collected and underwent flow cytometry analysis. ( b ) Quantification data are presented as the mean ± SD from three independent experiments. *** P < 0.001 by 1-way ANOVA. ( c ) BrdU immunofluorescence staining ( green ) was used to label S-phase cells after drug treatment ( blue signals show nuclei counterstaining with DAPI). ( d ) The percentage of BrdU-positive cells are presented as the mean ± SD from three independent experiments. Scale bar = 50 µm. * P < 0.05, ** P < 0.01, *** P < 0.001 by 1-way ANOVA.

Article Snippet: The quantity of PTEN in human and rat corneal endothelium tissues was measured using Human and Rat PTEN ELISA Kits (ELK Biotechnology, Wuhan, China), respectively, according to the manufacturer protocol.

Techniques: Inhibition, Flow Cytometry, Immunofluorescence, Staining

Journal: Investigative Ophthalmology & Visual Science

Article Title: PTEN Inhibition Accelerates Corneal Endothelial Wound Healing through Increased Endothelial Cell Division and Migration

doi: 10.1167/iovs.61.8.19

Figure Lengend Snippet: Inhibition of PTEN with bpV(pic) enhanced the promotion effect of TGF-β2 on CEC migration through activating the PI3K/Akt signaling pathway. ( a ) A scratch on the confluent B4G12 monolayer was made, followed by treatment with different drugs. Wound healing was recorded during the next 6 hours. Scale bar = 100 µm. N = 3. ( b ) Cell invasion assay was conducted and after drug treatment for 12 hours, the migrating cells were visualized through crystal violet staining. Scale bar = 100 µm. N = 3.

Article Snippet: The quantity of PTEN in human and rat corneal endothelium tissues was measured using Human and Rat PTEN ELISA Kits (ELK Biotechnology, Wuhan, China), respectively, according to the manufacturer protocol.

Techniques: Inhibition, Migration, Invasion Assay, Staining

Journal: Investigative Ophthalmology & Visual Science

Article Title: PTEN Inhibition Accelerates Corneal Endothelial Wound Healing through Increased Endothelial Cell Division and Migration

doi: 10.1167/iovs.61.8.19

Figure Lengend Snippet: A model for the mechanism of PTEN regulating corneal endothelial wound healing. PTEN inhibition with bpV(pic) released CECs from TGF-β2-induced G1-arrest and promoted CEC migration in synergy with TGF-β2 through regulating p27Kip1 and PI3K/Akt pathway, thus improved corneal endothelial regeneration in situ in a rat corneal endothelial injury model.

Article Snippet: The quantity of PTEN in human and rat corneal endothelium tissues was measured using Human and Rat PTEN ELISA Kits (ELK Biotechnology, Wuhan, China), respectively, according to the manufacturer protocol.

Techniques: Inhibition, Migration, In Situ

Journal: Microbiology Spectrum

Article Title: Clarithromycin Inhibits Pneumolysin Production via Downregulation of ply Gene Transcription despite Autolysis Activation

doi: 10.1128/Spectrum.00318-21

Figure Lengend Snippet: Neither CLR nor ERY affected the PLY-induced leakage of elastase from neutrophils. (A) Hemolytic activity of recombinant PLY was determined in the presence or absence of 5 μg/ml of CLR or ERY. (B) Human neutrophils (2 × 10 5 cells in 200 μl) were exposed to recombinant PLY (10 μg/ml) in the presence or absence of 5 μg/ml CLR or ERY for 2 h. NE activity of the culture supernatant was evaluated using a human NE activity assay kit. Data represent the mean ± SD of triplicate experiments and were evaluated using one-way analysis of variance with Tukey’s multiple-comparison test. *, Significantly different from the untreated group at P < 0.05. CLR, clarithromycin; Ctrl, control; ERY, erythromycin; NE, neutrophil elastase; PLY, pneumolysin; SD, standard deviation.

Article Snippet: Subsequently, human neutrophils (2 × 10 5 cells in 200 μl) were exposed to recombinant PLY (10 μg/ml) in the presence or absence of 5 μg/ml CLR or ERY for 2 h. The culture supernatant was evaluated for NE activity using a human NE activity assay kit (Cayman Chemical, Ann Arbor, MI, USA) according to the manufacturer’s instructions.

Techniques: Activity Assay, Recombinant, Comparison, Control, Standard Deviation