Sun Woong Kim1, Yilu Xie2, Paul Q Nguyen2, Vickie T Bui2, Kelly Huynh2, Jonathan S Kang2, Donald J Brown2, James V Jester3. 1. Gavin Herbert Eye Institute, University of California Irvine, Irvine, CA, United States; Department of Ophthalmology, Yonsei University Wonju College of Medicine, Wonju, South Korea. 2. Gavin Herbert Eye Institute, University of California Irvine, Irvine, CA, United States. 3. Gavin Herbert Eye Institute, University of California Irvine, Irvine, CA, United States. Electronic address: JJester@uci.edu.
Abstract
PURPOSE: To evaluate the role of PPARγ in regulating meibocyte differentiation and lipid synthesis in a human meibomian gland epithelial cell line (hMGEC). METHODS: HMGEC were exposed to the PPARγ agonist, Rosiglitazone, from 10-50 μM. Cultures were also exposed to specific PPARγ antagonist, T0070907, to block PPARγ receptor signaling. Cells were then stained with Ki-67 and LipidTox to determine the effects on cell cycling and lipid synthesis, respectively. Expression of meibocyte differentiation related proteins, ADFP, PPARγ, ELOVL4, and FABP4, were evaluated by quantitative PCR and western blotting. A human corneal epithelial cell line (hTCEpi) was used as a control. RESULT: Rosiglitazone significantly decreased Ki-67 staining within 2 days in a dose-dependent manner (P = 0.003) and increased lipid accumulation in hMGEC in a dose dependent manner. T0070907 suppressed both lipid droplet synthesis and cell cycle exit. Rosiglitazone significantly upregulated expression of ADFP, PPARγ, ELOVL4, and FABP4 by 9.6, 2.7, 2.6, and 3.3 fold on average (all P < 0.05 except for FABP4, P = 0.057) in hMGEC. T0070907 significantly abrogated rosiglitazone-induced upregulation of these genes when treated prior to rosiglitazone treatment (all P < 0.05). The observed lipogenic differentiation response was not duplicated in hTCEpi after exposure to rosiglitazone. CONCLUSION: Rosiglitazone induced cell cycle exit and upregulation of lipogenic gene expression leading to lipid accumulation in hMGEC. These effects were suppressed by PPARγ antagonist indicating that PPARγ signaling specifically directs lipogenesis in hMGEC. These findings suggest that PPARγ plays a critical role in meibocyte differentiation.
PURPOSE: To evaluate the role of PPARγ in regulating meibocyte differentiation and lipid synthesis in a human meibomian gland epithelial cell line (hMGEC). METHODS: HMGEC were exposed to the PPARγ agonist, Rosiglitazone, from 10-50 μM. Cultures were also exposed to specific PPARγ antagonist, T0070907, to block PPARγ receptor signaling. Cells were then stained with Ki-67 and LipidTox to determine the effects on cell cycling and lipid synthesis, respectively. Expression of meibocyte differentiation related proteins, ADFP, PPARγ, ELOVL4, and FABP4, were evaluated by quantitative PCR and western blotting. A humancorneal epithelial cell line (hTCEpi) was used as a control. RESULT: Rosiglitazone significantly decreased Ki-67 staining within 2 days in a dose-dependent manner (P = 0.003) and increased lipid accumulation in hMGEC in a dose dependent manner. T0070907 suppressed both lipid droplet synthesis and cell cycle exit. Rosiglitazone significantly upregulated expression of ADFP, PPARγ, ELOVL4, and FABP4 by 9.6, 2.7, 2.6, and 3.3 fold on average (all P < 0.05 except for FABP4, P = 0.057) in hMGEC. T0070907 significantly abrogated rosiglitazone-induced upregulation of these genes when treated prior to rosiglitazone treatment (all P < 0.05). The observed lipogenic differentiation response was not duplicated in hTCEpi after exposure to rosiglitazone. CONCLUSION:Rosiglitazone induced cell cycle exit and upregulation of lipogenic gene expression leading to lipid accumulation in hMGEC. These effects were suppressed by PPARγ antagonist indicating that PPARγ signaling specifically directs lipogenesis in hMGEC. These findings suggest that PPARγ plays a critical role in meibocyte differentiation.
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