bims-mideyd 2022-05-01 papers

Issue of 2022‒05‒01
four papers selected by
Raji Shyam
Indiana University Bloomington

Stem Cell Reports. 2022 Apr 14. pii: S2213-6711(22)00156-4. [Epub ahead of print]

Sleep deprivation induces corneal epithelial progenitor cell over-expansion through disruption of redox homeostasis in the tear film.

Sanming Li, Liying Tang, Jing Zhou, Sonia Anchouche, Dian Li, Yiran Yang, Zhaolin Liu, Jieli Wu, Jiaoyue Hu, Yueping Zhou, Jia Yin, Zuguo Liu, Wei Li.

Sleep deficiency, a common public health problem, causes ocular discomfort and affects ocular surface health. However, the underlying mechanism remains unclear. Herein, we identified that short-term sleep deprivation (SD) resulted in hyperproliferation of corneal epithelial progenitor cells (CEPCs) in mice. The expression levels of p63 and Keratin 14, the biomarkers of CEPCs, were upregulated in the corneal epithelium after short-term SD. In addition, SD led to elevated levels of reactive oxygen species (ROS), and subsequent decrease in antioxidant capacity, in the tear film. Exogenous hydrogen peroxide (H2O2) could directly stimulate the proliferation of CEPCs in vivo and in vitro. Topical treatment of antioxidant L-glutathione preserved the over-proliferation of CEPCs and attenuated corneal epithelial defects in SD mice. Moreover, the activation of the phosphoinositide 3-kinase (PI3K)/AKT signaling pathway is essential to ROS-stimulated cell proliferation in CEPCs. However, long-term SD ultimately led to early manifestation of limbal stem cell deficiency.

Keywords: ROS; cell proliferation; cornea; dry eye; epithelial progenitor cell; lacrimal gland; limbal stem cell; sleep deprivation; stem cell; tear film

DOI: https://doi.org/10.1016/j.stemcr.2022.03.017

Autophagy. 2022 Apr 26. 1-20

Increased LCN2 (lipocalin 2) in the RPE decreases autophagy and activates inflammasome-ferroptosis processes in a mouse model of dry AMD.

In dry age-related macular degeneration (AMD), LCN2 (lipocalin 2) is upregulated. Whereas LCN2 has been implicated in AMD pathogenesis, the mechanism remains unknown. Here, we report that in retinal pigmented epithelial (RPE) cells, LCN2 regulates macroautophagy/autophagy, in addition to maintaining iron homeostasis. LCN2 binds to ATG4B to form an LCN2-ATG4B-LC3-II complex, thereby regulating ATG4B activity and LC3-II lipidation. Thus, increased LCN2 reduced autophagy flux. Moreover, RPE cells from cryba1 KO, as well as sting1 KO and Sting1Gt mutant mice (models with abnormal iron chelation), showed decreased autophagy flux and increased LCN2, indicative of CGAS- and STING1-mediated inflammasome activation. Live cell imaging of RPE cells with elevated LCN2 also showed a correlation between inflammasome activation and increased fluorescence intensity of the Liperfluo dye, indicative of oxidative stress-induced ferroptosis. Interestingly, both in human AMD patients and in mouse models with a dry AMD-like phenotype (cryba1 cKO and KO), the LCN2 homodimer variant is increased significantly compared to the monomer. Sub-retinal injection of the LCN2 homodimer secreted by RPE cells into NOD-SCID mice leads to retinal degeneration. In addition, we generated an LCN2 monoclonal antibody that neutralizes both the monomer and homodimer variants and rescued autophagy and ferroptosis activities in cryba1 cKO mice. Furthermore, the antibody rescued retinal function in cryba1 cKO mice as assessed by electroretinography. Here, we identify a molecular pathway whereby increased LCN2 elicits pathophysiology in the RPE, cells known to drive dry AMD pathology, thus providing a possible therapeutic strategy for a disease with no current treatment options.Abbreviations: ACTB: actin, beta; Ad-GFP: adenovirus-green fluorescent protein; Ad-LCN2: adenovirus-lipocalin 2; Ad-LCN2-GFP: adenovirus-LCN2-green fluorescent protein; LCN2AKT2: AKT serine/threonine kinase 2; AMBRA1: autophagy and beclin 1 regulator 1; AMD: age-related macular degeneration; ARPE19: adult retinal pigment epithelial cell line-19; Asp278: aspartate 278; ATG4B: autophagy related 4B cysteine peptidase; ATG4C: autophagy related 4C cysteine peptidase; ATG7: autophagy related 7; ATG9B: autophagy related 9B; BLOC-1: biogenesis of lysosomal organelles complex 1; BLOC1S1: biogenesis of lysosomal organelles complex 1 subunit 1; C57BL/6J: C57 black 6J; CGAS: cyclic GMP-AMP synthase; ChQ: chloroquine; cKO: conditional knockout; Cys74: cysteine 74; Dab2: DAB adaptor protein 2; Def: deferoxamine; DHE: dihydroethidium; DMSO: dimethyl sulfoxide; ERG: electroretinography; FAC: ferric ammonium citrate; Fe2+: ferrous; FTH1: ferritin heavy chain 1; GPX: glutathione peroxidase; GST: glutathione S-transferase; H2O2: hydrogen peroxide; His280: histidine 280; IFNL/IFNλ: interferon lambda; IL1B/IL-1β: interleukin 1 beta; IS: Inner segment; ITGB1/integrin β1: integrin subunit beta 1; KO: knockout; LC3-GST: microtubule associated protein 1 light chain 3-GST; C-terminal fusion; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; LCN2: lipocalin 2; mAb: monoclonal antibody; MDA: malondialdehyde; MMP9: matrix metallopeptidase 9; NLRP3: NLR family pyrin domain containing 3; NOD-SCID: nonobese diabetic-severe combined immunodeficiency; OS: outer segment; PBS: phosphate-buffered saline; PMEL/PMEL17: premelanosome protein; RFP: red fluorescent protein; rLCN2: recombinant LCN2; ROS: reactive oxygen species; RPE SM: retinal pigmented epithelium spent medium; RPE: retinal pigment epithelium; RSL3: RAS-selective lethal; scRNAseq: single-cell ribonucleic acid sequencing; SD-OCT: spectral domain optical coherence tomography; shRNA: small hairpin ribonucleic acid; SM: spent medium; SOD1: superoxide dismutase 1; SQSTM1/p62: sequestosome 1; STAT1: signal transducer and activator of transcription 1; STING1: stimulator of interferon response cGAMP interactor 1; TYR: tyrosinase; VCL: vinculin; WT: wild type.

Keywords: ATG4B; autophagy; dry age-related macular degeneration; ferroptosis; inflammasome; iron; lipocalin 2; monoclonal/neutralizing antibody; oxidative stress; retinal pigmented epithelial cells

DOI: https://doi.org/10.1080/15548627.2022.2062887

Oxid Med Cell Longev. 2022 ;2022 7299182

Biliverdin/Bilirubin Redox Pair Protects Lens Epithelial Cells against Oxidative Stress in Age-Related Cataract by Regulating NF-κB/iNOS and Nrf2/HO-1 Pathways.

Yang Huang, Jinglan Li, Wenzhe Li, Nanping Ai, Haiying Jin.

Age-related cataract (ARC) is the leading cause of vision impairment globally. It has been widely accepted that excessive reactive oxygen species (ROS) accumulation in lens epithelial cells (LECs) is a critical risk factor for ARC formation. Biliverdin (BV)/bilirubin (BR) redox pair is the active by-product of heme degradation with robust antioxidative stress and antiapoptotic effects. Thus, we purpose that BV and BR may have a therapeutic effect on ARC. In the present study, we determine the expression levels of enzymes regulating BV and BR generation in human lens anterior capsule samples. The therapeutic effect of BV/BR redox pair on ARC was assessed in hydrogen peroxide (H2O2)-damaged mouse LECs in vitro. The NF-κB/inducible nitric oxide synthase (iNOS) and nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) signaling pathways were evaluated to illustrate the molecular mechanism. The results revealed that the mRNA expressions of Nrf2, HO-1, and biliverdin reductase A (BVRA) were all decreased in human samples of age-related nuclear cataract. BV/BR redox pair pretreatment protected LECs against H2O2 damage by prohibiting NF-κB p65 nuclear trafficking, ameliorating iNOS expression, reducing intracellular and mitochondrial ROS levels, and restoring glutathione (GSH) and superoxide dismutase (SOD) levels. BV and BR pretreatment also regulated the expression of apoptotic molecules (Bax, Bcl-2, and cleaved caspase-3), thus decreasing the apoptosis of LECs. In addition, BV/BR pair promoted Nrf2 nuclear accumulation and HO-1 induction, whereas the knockdown of BVRA counteracted the effect of BV on activating Nrf2/HO-1 pathway and antiapoptosis. These findings implicated that BV/BR redox pair protects LECs against H2O2-induced apoptosis by regulating NF-κB/iNOS and Nrf2/HO-1 pathways. Moreover, BVRA is responsible for BV-mediated cytoprotection by reductive conversion of BV to BR. This trial is registered with ChiCTR2000036059.

DOI: https://doi.org/10.1155/2022/7299182

Invest Ophthalmol Vis Sci. 2022 Apr 01. 63(4): 22

Repressed miR-34a Expression Dictates the Cell Fate to Corneal Endothelium Failure.

Junji Hamuro, Kazuko Asada, Morio Ueno, Tomoko Yamashita, Atsushi Mukai, Tomoko Fujita, Eiko Ito, Nao Hiramoto, Munetoyo Toda, Chie Sotozono, Shigeru Kinoshita.

Purpose: To reveal the mechanism triggering the functional disparity between degenerated and non-degenerated corneal endothelium cells in the water efflux from corneal stroma to the anterior chamber.Methods: The varied levels of the microRNA (miR)-34, miR-378, and miR-146 family in human corneal endothelium and cultured cells thereof were investigated using 3D-Gene Human miRNA Oligo Chips. Concomitantly, CD44, p53, c-Myc, matrix metalloprotease (MMP)-2 expression, and Ras homolog gene family member A (Rho A) activity was correlated to the expression intensities of these microRNAs, partly complemented with their altered expression levels with the transfection of the corresponding mimics and inhibitors. The levels of miRs were further associated with intracellular pH (pHi) and mitochondrial energy homeostasis.
Results: P53-inducible miR-34a/b repressed CD44 expression, and CD44 was repressed with the elevated c-Myc. The repressed miR-34a activated the CD44 downstream factors Rho A and MMP-2. MiR-34a mimics downregulated pHi, inducing the skewing of mitochondrial respiration to oxidative phosphorylation. The oxidative stress (H2O2) induced on human corneal endothelial cells, which repressed miR-34a/b expression, may account for the impaired signaling cascade to mitochondrial metabolic homeostasis necessary for an efficient water efflux from the corneal stroma.
Conclusions: The upregulated expression of CD44, through repressed miR-34a/b by reactive oxygen species and elevated c-Myc by oxidative stress, may impair mitochondrial metabolic homeostasis, leading to human corneal endothelial failure.

DOI: https://doi.org/10.1167/iovs.63.4.22