Cholesterol and oxysterols in retinal neuron-glia interactions: relevance for glaucoma.

Cholesterol is an essential component of cellular membranes, crucial for maintaining their structural and functional integrity. It is especially important for nervous tissues, including the retina, which rely on high amounts of plasma membranes for the transmission of the nervous signal. While cholesterol is by far the most abundant sterol, the retina also contains cholesterol precursors and metabolites, especially oxysterols, which are bioactive molecules. Cholesterol lack or excess is deleterious and some oxysterols are known for their effect on neuron survival. Cholesterol homeostasis must therefore be maintained. Retinal glial cells, especially Müller cells, the principal glial cells of the vertebrate retina, provide mechanical, nutritional, and metabolic support for the neighboring neurons. Several pieces of evidence indicate that Müller cells are major actors of cholesterol homeostasis in the retina, as it is known for other glial cells in the brain. This process is based on a close cooperation with neurons, and sterols can be signaling molecules participating in glia-neuron interactions. While some implication of cholesterol in age-related macular degeneration is now recognized, based on epidemiological and laboratory data, evidence for its role in glaucoma is still scarce. The association between cholesterolemia and glaucoma is controversial, but experimental data suggest that sterols could take part in the pathological processes. It has been demonstrated that Müller glial cells are implicated in the development of glaucoma through an ambivalent reactive retinal gliosis process. The early steps contribute to maintaining retinal homeostasis and favor the survival of ganglion cells, which are targeted during glaucoma. If gliosis persists, dysregulation of the neuroprotective functions, cytotoxic effects of gliotic Müller cells and disruption of glia-neuron interactions lead to an acceleration of ganglion cell death. Sterols could play a role in the glial cell response to glaucomatous injury. This represents an understudied but attractive topic to better understand glaucoma and conceive novel preventive or curative strategies. The present review describes the current knowledge on i) sterol metabolism in retinal glial cells, ii) the potential role of cholesterol in glaucoma, and iii) the possible relationships between cholesterol and oxysterols, glial cells and glaucoma. Focus is put on glia-neuron interactions.

Retinal cholesterol metabolism is perturbated in response to experimental glaucoma in the rat.

Alterations of cholesterol metabolism have been described for many neurodegenerative pathologies, such as Alzheimer's disease in the brain and age-related macular degeneration in the retina. Recent evidence suggests that glaucoma, which is characterized by the progressive death of retinal ganglion cells, could also be associated with disruption of cholesterol homeostasis. In the present study we characterized cholesterol metabolism in a rat model of laser-induced intraocular hypertension, the main risk factor for glaucoma. Sterol levels were measured using gas-chromatography and cholesterol-related gene expression using quantitative RT-PCR at various time-points. As early as 18 hours after the laser procedure, genes implicated in cholesterol biosynthesis and uptake were upregulated (+49% and +100% for HMG-CoA reductase and LDLR genes respectively, vs. naive eyes) while genes involved in efflux were downregulated (-26% and -37% for ApoE and CYP27A1 genes, respectively). Cholesterol and precursor levels were consecutively elevated 3 days post-laser (+14%, +40% and +194% for cholesterol, desmosterol and lathosterol, respectively). Interestingly, counter-regulatory mechanisms were transcriptionally activated following these initial dysregulations, which were associated with the restoration of retinal cholesterol homeostasis, favorable to ganglion cell viability, one month after the laser-induced ocular hypertension. In conclusion, we report here for the first time that ocular hypertension is associated with transient major dynamic changes in retinal cholesterol metabolism.

Is 24(S)-hydroxycholesterol a potent modulator of cholesterol metabolism in Müller cells? An in vitro study about neuron to glia communication in the retina.

Communication between neurons and glia plays a major role in nervous tissue homeostasis. It is thought to participate in tuning cholesterol metabolism to cellular demand, which is a critical issue for neuronal health. Cholesterol is a membrane lipid crucial for nervous tissue functioning, and perturbed regulation of its metabolism has been linked to several neurodegenerative disorders. In the brain, 24(S)-hydroxycholesterol (24S-OHC) is an oxysterol synthesized by neurons to eliminate cholesterol, and 24S-OHC has been shown to regulate cholesterol metabolism in astrocytes, glial cells which provide cholesterol to neurons. In the retina, 24S-OHC is also an elimination product of cholesterol produced by neurons, especially the retinal ganglion cells. However, it is not known whether Müller cells, the major macroglial cells of the retina, play the role of cholesterol provider for retinal neurons and whether they respond to 24S-OHC signaling, similarly to brain glial cells. In the present study, primary cultures of rat Müller cells were treated with 0, 0.5 or 1.5 µM 24S-OHC for 48 hours. The levels of cholesterol, precursors and oxysterols were quantified using gas chromatography coupled to flame-ionization detection or mass spectrometry. In addition, the expression of key genes related to cholesterol metabolism was analyzed using RTq-PCR. Müller cells were shown to express many genes linked to cholesterol metabolism, including genes coding for proteins implicated in cholesterol biosynthesis (HMGCR), cholesterol uptake and export via lipoproteins (LDL-R, SR-BI, ApoE and ABACA1) and regulation of cholesterol metabolism (SREBP2 and LXRß). Cholesterol and several of its precursors and oxidative products were present. CYP27A1, the main retinal enzyme implicated in cholesterol elimination via oxysterol production, was quantified at low transcript levels but neither of its two typical products were detected in Müller cells. Furthermore, our results demonstrate that 24S-OHC has a strong hypocholesterolemic effect in Müller cells, leading to cholesterol depletion (-37 % at 1.5 µM). This was mediated by a decrease in cholesterol synthesis, as illustrated by reduced levels of cholesterol precursors: desmosterol (-38 % at 1.5 µM) and lathosterol (-84 % at 1.5 µM), and strong downregulation of HMGCR gene expression (2.4 fold decrease at 1.5µM). In addition, LDL-R and SR-BI gene expression were reduced in response to 24S-OHC treatment (2 fold and 1.6 fold at 1.5 µM, respectively), suggesting diminished lipoprotein uptake by the cells. On the contrary, there was a dramatic overexpression of ABCA1 transporter (10 fold increase at 1.5 µM), probably mediating an increase in cholesterol efflux. Finally, 24S-OHC induced a small but significant upregulation of the CYP27A1 gene. These data indicate that Müller cells possess the necessary cholesterol metabolism machinery and that they are able to sharply adjust their cholesterol metabolism in response to 24S-OHC, a signal molecule of neuronal cholesterol status. This suggests that Müller cells could be major players of cholesterol homeostasis in the retina via neuron-glia crosstalk.

Cholesterol metabolism and glaucoma: Modulation of Muller cell membrane organization by 24S-hydroxycholesterol.

Glaucoma is a progressive and irreversible blinding neuropathy that is characterized by the loss of retinal ganglion cells (RGCs). Muller Glial Cell (MGC) activation is induced in retinal gliosis. MGCs are the most numerous glial cells in the retina and one of their roles is to sustain cholesterol homeostasis. 24S-hydroxycholesterol (24S-OHC) is one of the form of cholesterol elimination from the retina and is overexpressed during glaucoma. The objective of this study was to determine whether 24S-OHC triggers MGC membrane dynamics involving lipid rafts and contributes to gliosis at early and late time points. A proteomic analysis was carried out by nanoLC-MS/MS in raft and non-raft fractions from MGCs after treatment with 24S-OHC (10µM). The expression of structural and functional proteins was further analyzed by Western-blotting, as well as the levels of GM3 ganglioside by LC-MS. Cholesterol, sphingomyelin, saturated fatty acids and ganglioside GM3 are enriched in the rafts fractions in MGCs. Caveolin-1, flotillin-1, connexin-30 and -43 are localized in the MGCs rafts. Proteins implicated in adhesion or oxidative stress pathways in raft fractions were up and down-regulated by the treatment. Our data showed that 24S-OHC induced early changes in protein distribution in raft microdomains; however, further studies are needed to better characterize the surrounded mechanisms.

High Fructose Diet inducing diabetes rapidly impacts olfactory epithelium and behavior in mice.

Type 2 Diabetes (T2D), a major public health issue reaching worldwide epidemic, has been correlated with lower olfactory abilities in humans. As olfaction represents a major component of feeding behavior, its alteration may have drastic consequences on feeding behaviors that may in turn aggravates T2D. In order to decipher the impact of T2D on the olfactory epithelium, we fed mice with a high fructose diet (HFruD) inducing early diabetic state in 4 to 8 weeks. After only 4 weeks of this diet, mice exhibited a dramatic decrease in olfactory behavioral capacities. Consistently, this decline in olfactory behavior was correlated to decreased electrophysiological responses of olfactory neurons recorded as a population and individually. Our results demonstrate that, in rodents, olfaction is modified by HFruD-induced diabetes. Functional, anatomical and behavioral changes occurred in the olfactory system at a very early stage of the disease.