Journal: Investigative ophthalmology & visual science

Article Title: Selective Serotonin Reuptake Inhibitors Aggravate Depression-Associated Dry Eye Via Activating the NF-κB Pathway.

doi: 10.1167/iovs.18-25572

Figure Lengend Snippet: FIGURE 1. SSRI treatment is closely related to depression-associated dry eye disease and causes increased tear serotonin levels with a serious inflammatory response and cell apoptosis at the ocular surface. (A) The clinical indicators based on the baseline data and data collected after 90 days of treatment in the SSRI and control groups. The P value means the difference between ‘‘change’’ in SSRI and control groups. (B) ELISA analysis showing the tear serotonin levels of the patients with SSRI treatment and the control group. (C, D) Protein chip analysis showing the inflammatory cytokine content following SSRI treatment, based on examination of 507 proteins, including TLR4, IL-6, and IL-10. (E) qRT-PCR analysis showing the conjunctival TLR2 and TLR4 and inflammatory cytokine IL1b, IL10, and TNFa mRNA levels between the patients with SSRI treatment and the control group. (F) qRT-PCR analysis showing the conjunctival mRNA levels of the proapoptotic genes AIF, BAD, and BAX and the antiapoptotic genes BCL2 and XIAP between the patients with SSRI treatment and the control group. *P < 0.05, **P < 0.01.

Article Snippet: The membranes were incubated overnight at 48C with primary antibodies, including IL-1b (1:1000; Abcam), IL-6 (1:1000; Proteintech, Wuhan, China), IL-10 (1:500; Abcam), TNF-a (1:500; Abcam), TLR2 (1:1000; Abcam), TLR4 (1:200; Santa Cruz Biotechnology), AIF (1:1000; Proteintech), BAD (1:1000; Abcam), BAX (1:1000; Proteintech), P-p65 (1:2000; Abcam), p65 (1:2000; Abcam), IjBa (1:1000; Abcam), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, 1:12000; Proteintech).

Techniques: Control, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR

Journal: Investigative ophthalmology & visual science

Article Title: Selective Serotonin Reuptake Inhibitors Aggravate Depression-Associated Dry Eye Via Activating the NF-κB Pathway.

doi: 10.1167/iovs.18-25572

Figure Lengend Snippet: FIGURE 4. Serotonin can induce corneal epithelial cell inflammation and apoptosis. (A) qRT-PCR analysis showing the mRNA levels of the inflammatory cytokines TLR4, TLR2, IL1b, IL6, IL10, and TNFa with the serotonin level. (B, C) Western blot analysis showing the protein levels in the growth plate of TLR4, TLR2, IL-1b, IL-6, IL-10, and TNF-a with the serotonin level. (D) ELISA analysis of the related IL-1b, IL-10, and TNF-a content in the cell supernatants in cells with increased serotonin levels. (E, F) Western blot analysis showing the protein levels in the growth plate of the proapoptotic proteins BAX, BAD, and AIF with the serotonin level. (E) Flow cytometry showing apoptosis in corneal epithelial cells with serotonin treatment. *P < 0.05, **P < 0.01.

Article Snippet: The membranes were incubated overnight at 48C with primary antibodies, including IL-1b (1:1000; Abcam), IL-6 (1:1000; Proteintech, Wuhan, China), IL-10 (1:500; Abcam), TNF-a (1:500; Abcam), TLR2 (1:1000; Abcam), TLR4 (1:200; Santa Cruz Biotechnology), AIF (1:1000; Proteintech), BAD (1:1000; Abcam), BAX (1:1000; Proteintech), P-p65 (1:2000; Abcam), p65 (1:2000; Abcam), IjBa (1:1000; Abcam), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, 1:12000; Proteintech).

Techniques: Quantitative RT-PCR, Western Blot, Enzyme-linked Immunosorbent Assay, Flow Cytometry

Journal: Investigative ophthalmology & visual science

Article Title: Selective Serotonin Reuptake Inhibitors Aggravate Depression-Associated Dry Eye Via Activating the NF-κB Pathway.

doi: 10.1167/iovs.18-25572

Figure Lengend Snippet: FIGURE 5. Serotonin receptors are indispensable for serotonin-induced cell inflammation and apoptosis in corneal epithelial cells. (A) qRT-PCR analysis showing the HTR1A, 1B, 1D, 1E, 1F, 2A, 2B, 2C, 3A, 3B, 4, 5A, 6, and 7 mRNA levels in corneal epithelial cells. (B, C) Western blot assay showing the expression levels of the proinflammatory response element TLR4, inflammatory cytokines IL-6 and IL-10, and proapoptotic proteins BAX, BAD, and AIF in asenapine maleate–treated corneal epithelial cells. (D) ELISA analysis of the related IL-1b, IL-10, and TNF-a content in the supernatants of cells with accumulated serotonin treated with asenapine maleate. (G) Flow cytometry showing apoptosis in corneal epithelial cells with serotonin treated with asenapine maleate. *P < 0.05, **P < 0.01.

Article Snippet: The membranes were incubated overnight at 48C with primary antibodies, including IL-1b (1:1000; Abcam), IL-6 (1:1000; Proteintech, Wuhan, China), IL-10 (1:500; Abcam), TNF-a (1:500; Abcam), TLR2 (1:1000; Abcam), TLR4 (1:200; Santa Cruz Biotechnology), AIF (1:1000; Proteintech), BAD (1:1000; Abcam), BAX (1:1000; Proteintech), P-p65 (1:2000; Abcam), p65 (1:2000; Abcam), IjBa (1:1000; Abcam), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, 1:12000; Proteintech).

Techniques: Quantitative RT-PCR, Western Blot, Expressing, Enzyme-linked Immunosorbent Assay, Flow Cytometry

Journal: Investigative ophthalmology & visual science

Article Title: Selective Serotonin Reuptake Inhibitors Aggravate Depression-Associated Dry Eye Via Activating the NF-κB Pathway.

doi: 10.1167/iovs.18-25572

Figure Lengend Snippet: FIGURE 6. Serotonin induces cell inflammation and apoptosis by activating the NF-jB pathway. (A, B) Western blot analysis showing the p65 phosphorylation level and the IjBa protein level with the serotonin level. (C, D) Western blot analysis showing the P-p65, IjBa, proinflammatory response element TLR4, inflammatory cytokine IL-1b, and proapoptotic protein BAX, BAD, and AIF protein levels in JSH-23–treated corneal epithelial cells. (E) ELISA analysis of the related IL-1b, IL-10, and TNF-a content in the supernatants of cells treated with the NF-jB signaling inhibitor. (F) Flow cytometry showing apoptosis in corneal epithelial cells with accumulated serotonin following JSH-23 treatment. *P < 0.05, **P < 0.01.

Article Snippet: The membranes were incubated overnight at 48C with primary antibodies, including IL-1b (1:1000; Abcam), IL-6 (1:1000; Proteintech, Wuhan, China), IL-10 (1:500; Abcam), TNF-a (1:500; Abcam), TLR2 (1:1000; Abcam), TLR4 (1:200; Santa Cruz Biotechnology), AIF (1:1000; Proteintech), BAD (1:1000; Abcam), BAX (1:1000; Proteintech), P-p65 (1:2000; Abcam), p65 (1:2000; Abcam), IjBa (1:1000; Abcam), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, 1:12000; Proteintech).

Techniques: Western Blot, Phospho-proteomics, Enzyme-linked Immunosorbent Assay, Flow Cytometry