Clusterin acts as an endogenous statin by controlling the expression and nuclear translocation of sterol regulatory-element binding protein (SREBP) in trabecular meshwork.

Abstract

Purpose : The membrane remodeling of the TM is important for mechanosensing and aqueous humor (AH) drainage. Lipids play a critical role in the cell membrane fluidity and mechanobiology. Membrane–bound transcription factor SREBP contains a lipid-sensing domain that, when activated, facilitates cleavage of its transcription factor domain via the enzyme SREBP cleavage-activating protein (SCAP) and entry into the nucleus to activate genes involved in lipid metabolism. Clusterin or Apolipoprotein J is associated with lipid metabolism. Our studies show constitutive clusterin expression lowers intraocular pressure (IOP) with an unclear mechanism. We hypothesize that clusterin regulates lipid homeostasis in TM and thereby AH drainage. Therefore, we investigated the role of clusterin in the regulation of SREBP.

Methods : Quantitative-polymerase chain reaction (qPCR) and immunofluorescence assays were performed on human TM (HTM) cells in culture to assess – a) expression of transcript variants of SREBP and SCAP in human trabecular meshwork (HTM) cells, b) effects of constitutive clusterin expression using adenovirus-mediated clusterin (AdCLU) in comparison to 20µM fatostatin, an inhibitor of SREBP activation, on SREBP transcription and translocation into the nucleus and active form of transforming growth factor b-2 (AdTGFb2^mut), a factor known to increase IOP. Students t-test was used for statistical analyses and results were significant if p<0.05 with a sample size of n=3-6.

Results : Transcript variants of SREBP-F1V1, V2, V3 and -F2V1 and SCAP are expressed in HTM cells. Immunofluorescence analyses showed that SREBP and SCAP puncta were colocalized on endoplasmic reticulum (ER) membrane and Golgi as visualized by ER marker -KDEL and Golgi marker -GM130. AdCLU expression lowered the expression of SREBP1V2 significantly (p=0.008, n=6) and similar to the treatment with 20µM fatostatin (p=0.002, n=6). Interestingly, AdTGFβ2^mut increased SREBF1V2 expression (p=0.046, n=3).

Conclusions : Our preliminary studies identify clusterin as an endogenous lipid modulator in the TM by regulating the homeostasis of lipid biogenesis. Further studies on the role of clusterin on lipid metabolism in TM pathway will have a significant impact on potential regulation of the membrane properties and AH outflow resistance.

This is a 2020 ARVO Annual Meeting abstract.