IGF-1-Mediated Survival from Induced Death of Human Primary Cultured Retinal Pigment Epithelial Cells Is Mediated by an Akt-Dependent Signaling Pathway
Abstract:Degeneration of the human retinal pigmented epithelium (hRPE) is involved in several eye disorders such as age-related macular degeneration (AMD). In this study, we investigated the protective effect of IGF-1 on human primary cultured RPE cells and its underlying mechanism. IGF-1 dose- and time-dependently promoted the survival of RPE cells from serum deprivation. Western blot showed that IGF-1 stimulated the activation of the PI3K/Akt and MAPK pathways in hRPE. Inhibition of the PI3K/Akt pathway by the PI3K-s… Show more
“…In line with our previous results (Fig. 1c, d ), this included increased phosphorylation of proline-rich AKT1 substrate 40 (PRAS40), whose expression is induced by IGF-1 48 , 49 , while Sandalore ® reduced phosphorylation of p53 (S46), which is highly phosphorylated in apoptotic cells 50 (IGF-1 is a key apoptosis suppressor 51 – 53 ). Fig.…”
Olfactory receptors are expressed by different cell types throughout the body and regulate physiological cell functions beyond olfaction. In particular, the olfactory receptor OR2AT4 has been shown to stimulate keratinocyte proliferation in the skin. Here, we show that the epithelium of human hair follicles, particularly the outer root sheath, expresses OR2AT4, and that specific stimulation of OR2AT4 by a synthetic sandalwood odorant (Sandalore®) prolongs human hair growth ex vivo by decreasing apoptosis and increasing production of the anagen-prolonging growth factor IGF-1. In contrast, co-administration of the specific OR2AT4 antagonist Phenirat® and silencing of OR2AT4 inhibit hair growth. Together, our study identifies that human hair follicles can engage in olfactory receptor-dependent chemosensation and require OR2AT4-mediated signaling to sustain their growth, suggesting that olfactory receptors may serve as a target in hair loss therapy.
“…In line with our previous results (Fig. 1c, d ), this included increased phosphorylation of proline-rich AKT1 substrate 40 (PRAS40), whose expression is induced by IGF-1 48 , 49 , while Sandalore ® reduced phosphorylation of p53 (S46), which is highly phosphorylated in apoptotic cells 50 (IGF-1 is a key apoptosis suppressor 51 – 53 ). Fig.…”
Olfactory receptors are expressed by different cell types throughout the body and regulate physiological cell functions beyond olfaction. In particular, the olfactory receptor OR2AT4 has been shown to stimulate keratinocyte proliferation in the skin. Here, we show that the epithelium of human hair follicles, particularly the outer root sheath, expresses OR2AT4, and that specific stimulation of OR2AT4 by a synthetic sandalwood odorant (Sandalore®) prolongs human hair growth ex vivo by decreasing apoptosis and increasing production of the anagen-prolonging growth factor IGF-1. In contrast, co-administration of the specific OR2AT4 antagonist Phenirat® and silencing of OR2AT4 inhibit hair growth. Together, our study identifies that human hair follicles can engage in olfactory receptor-dependent chemosensation and require OR2AT4-mediated signaling to sustain their growth, suggesting that olfactory receptors may serve as a target in hair loss therapy.
“…In particular, the stimulators of PI3K/Akt could represent a promising therapeutic tool for the prevention of the degeneration of RPE, and theoretically, for the treatment of eye disorders, such as AMD [53, 54]. …”
Background/Aims: the anti-vascular endothelial growth factors (VEGF), Aflibercept and Ranibizumab, are used for the treatment of macular degeneration. Here we examined the involvement of nitric oxide (NO), mitochondria function and of apoptosis/autophagy in their antioxidant effects in human retinal pigment epithelium cells (RPE). Methods: RPE were exposed to Ranibizumab/Aflibercept in the absence or presence of NO synthase (NOS) inhibitor and of autophagy activator/blocker, rapamicyn/3-methyladenine. Specific kits were used for cell viability, NO and reactive oxygen species detection and mitochondrial membrane potential measurement, whereas Western Blot was performed for apoptosis/ autophagy markers and other kinases detection. Results: In RPE cultured in physiological conditions, Aflibercept/Ranibizumab increased NO release in a dose and time-dependent way. Opposite results were obtained in RPE pretreated with hydrogen peroxide. Moreover, both the anti-VEGF agents were able to prevent the fall of cell viability and of mitochondrial membrane potential. Those effects were reduced by the NOS inhibitor and 3-methyladenine and were potentiated by rapamycin. Finally, Aflibercept and Ranibizumab counteracted the changes of apoptosis/autophagy markers, NOS, Phosphatidylinositol-3-Kinase/Protein Kinase B and Extracellular signal–regulated kinases 1/2 caused by peroxidation. Conclusion: Aflibercept and Ranibizumab protect RPE against peroxidation through the modulation of NO release, apoptosis and autophagy.
“…RPE65 staining was performed as described by Zheng et al . (). The eyes were harvested 24 h after light exposure and fixed for 24 h in 4% paraformaldehyde, dehydrated in 15 and 30% sucrose solution overnight, embedded in Tissue‐Tek optimal cutting temperature compound, and 6 μm frozen sections were collected.…”
Amiodarone is one of the most effective anti-arrhythmic drugs available, but its clinical applications are limited by toxic side effects including optic toxicity. The purpose of this study was to investigate the toxic effect of amiodarone on D407 cells (a human retinal pigmented epithelial (RPE) cell line) and the mechanisms of the protective effect of insulin-like growth factor-1 (IGF-1).
EXPERIMENTAL APPROACHThe involvement of the kinases, Akt and ERK, was analysed by Western blot. Intracellular accumulation of ROS was measured using fluorophotometric quantification. A pharmacological approach with inhibitors was used to investigate the pathways involved in the protective action of IGF-1.
KEY RESULTSAmiodarone concentration-dependently augmented the production of ROS, lipid peroxidation and apoptosis in D407 cells. IGF-1 time-and concentration-dependently reversed these effects of amiodarone and protected D407 cells from amiodarone-mediated toxicity. Amiodarone inhibited the pAkt but not pErk, and IGF-1 reversed this inhibitory effect of amiodarone. However, IGF-1 failed to suppress amiodarone-induced cytotoxicity in the presence of PI3K/Akt inhibitor LY294002 suggesting the direct involvement of the PI3K/Akt pathway. Furthermore, in vivo rat flash electroretinogram (FERG) recordings showed that IGF-1 reverses the amiodarone-induced decrease in a-and b-waves. The immunocytochemistry findings confirmed that vitreous IGF-1 injections promote the survival of RPE cells in rat retina treated with amiodarone.
CONCLUSION AND IMPLICATIONSIGF-1 can protect RPE cells from amiodarone-mediated injury via the PI3K/Akt pathway in vivo and in vitro. IGF-1 has potential as a protective drug for the prevention and treatment of amiodarone-induced optic toxicity.Abbreviations FERG, flash electroretinogram; IGF-1, insulin-like growth factor-1; INL, inner nuclear layer; IPL, inner plexiform layer; RGCs, retinal ganglionic cells; RPE, retinal pigmented epithelium
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