Neurovascular Cell Death and Therapeutic Strategies for Diabetic Retinopathy

Abstract

Diabetic retinopathy (DR) is a major complication of diabetes and a leading cause of blindness worldwide. DR was recently defined as a neurovascular disease associated with tissue-specific neurovascular impairment of the retina in patients with diabetes. Neurovascular cell death is the main cause of neurovascular impairment in DR. Thus, neurovascular cell protection is a potential therapy for preventing the progression of DR. Growing evidence indicates that a variety of cell death pathways, such as apoptosis, necroptosis, ferroptosis, and pyroptosis, are associated with neurovascular cell death in DR. These forms of regulated cell death may serve as therapeutic targets for ameliorating the pathogenesis of DR. This review focuses on these cell death mechanisms and describes potential therapies for the treatment of DR that protect against neurovascular cell death.

Keywords: apoptosis; diabetic retinopathy; ferroptosis; necroptosis; neuroprotection; neurovascular cell death; neurovascular unit; pyroptosis; vasoprotection.

Figure 1

Hypothetic scheme of apoptotic cell death pathways in retinal cells under diabetic stress. The figure is modified and updated from the figure in a previous review [5]. Chronic hyperglycemia is a biochemical injury for retinal cells. Because the cell death mechanisms of the biochemical injury are, in part, common with those of physical injuries, the scheme is mainly made from findings in optic nerve injuries and retinal culture studies [5]. In most neuronal cells, extrinsic pathway signals are thought to be transferred to mitochondria via truncated Bid, probably because most apoptosis-related factors, including pro-caspase-3, may be preserved in the mitochondria in physiological conditions. Thus, regarding neuronal apoptosis, the role of mitochondria may be more critical than that of other cells. In apoptotic cell death, DNA fragmentation is the “point of no return”. AP-1, activator protein-1; Casp-9, caspase-9; Cyto C, cytochrome c; Apaf-1, apoptosis protease-activating factor 1; Bcl-2, B-cell lymphoma 2; Bcl-xL, B-cell lymphoma-extra-large; tBid, truncated Bid; AIF, apoptosis-inducing factor; Casp-3, caspase-3; Casp-8, caspase-8; TRADD, TNF receptor 1-associated death domain protein; FADD, Fas-associated death domain; TNF-α, tumor necrosis factor-α; TNF-R1, tumor necrosis factor receptor 1; ER, endoplasmic reticulum; ROS, reactive oxygen species; IP3R, inositol trisphosphate receptor; IRE1, inositol-requiring ER-to-nucleus signaling protein 1; PERK, protein kinase-like ER eukaryotic initiation factor-2alpha kinase; ATF6, activating transcription factor-6; TRAF2, TNFR-associated factor 2; JNK, c-Jun-N-terminal protein kinase; ASK1, apoptosis signal-regulating kinase 1; eIF-2α, eukaryotic initiation factor-2α; ATF4, activating transcription factor-4; CHOP, CCAAT/enhancer-binding protein homologous protein; Casp-2, caspase-2; PS, phosphatidylserine. SP1/2, site-1/2 protease.

Figure 2

Hypothetical scheme of molecular mechanisms of pyroptosis. The Nomenclature Committee on Cell Death does not recommend the use of alternative terms such as “pyronecrosis” [57]. The central molecule of pyroptosis is gasdermin D (GSDMD). There are two pathways which activate GSDMD, the classical pathway and the non-classical pathway. In the classical pathway, Nod-like receptor family pyrin domain containing 3 (NLRP3) inflammasome is activated under the Toll-like receptor (TLR) signals which results in caspase-1 activation. Caspase-1 cleaves pro-interleukin-1β (pro-IL-1β) and pro-IL-18 in addition to GSDMD. The N-terminal domains of GSDMDs form plasma membrane pores and release low-molecular-damage-associated molecular patterns (DAMPs). On the other hand, in the non-classical pathway, caspase-4/5/11 are activated by endotoxins, such as lipopolysaccharide (LPS). Activated caspase-4/5/11 can cleave GSDMD followed by the formation of GSDMD pores. Further inflammatory signals activate nerve injury-induced protein 1 (NINJ1). Cell-surface NINJ1 mediates further plasma membrane ruptures during pyroptosis followed by releasing high-molecular-weight DMAPs [60]. Thus, pyroptosis can passively exacerbate inflammatory reaction via NINJ1-mediated plasma membrane rupture. MyD88, myeloid differentiation primary response gene 88; TRAF6, TNF receptor-associated factor 6; Ub, ubiquitin; TAK1, TGF-β-activated kinase 1; TAB1/2, TAK1-binding protein 1/2; IκB, inhibitor of κB; NF-κB, nuclear factor-κB; ROS, reactive oxygen species; MEK7, MAPK ERK kinase 7; ASC, apoptosis-associated speck-like protein containing a CARD; P₂X₇R, P₂X₇ receptor.

Figure 3

Hypothetical scheme of molecular pathways of ferroptosis and Fenton reaction. The late phase of ferroptosis is still considered a “black box”, and thus, it is unclear where the “point of no return” is. However, before membrane integrity disruption, propagation of ferroptosis occurs [83] probably because lipid peroxidation-associated factors, including iron and heme, may be distributed to surrounding cells before the membrane ruptures [84]. The affected points of endogenous inhibitory factors, GPX4 and CoQ₁₀H2, are shown in the figure. The scheme of the Fenton reaction is shown in the green box. Acyl-CoA synthetase long-chain family member 4 (ACSL4) and lysophosphatidylchoiline acyltransferase 3 (LPCAT3) are major regulatory enzymes of ferroptosis [80,82]. ASCL4 catalyzes to connect acyl-CoA with a polyunsaturated fatty acid (PUFA; PUFA-CoA). LPCAT3 catalyzes the translocation of PUFA-CoA into acyl phospholipids which results in synthesis of PUFA-phospholipid (PL)-rich membrane. The PUFA-PL-rich membrane increases the sensitivity of lipid peroxidation followed by facilitation of ferroptosis. NADPH-cytochrome P-450 reductase (POR) peroxidates PUFA-PL (PUFA-PL-OOH) as an electron donor of nicotinamide adenine dinucleotide phosphate (NADPH). Transcription factors NRF2 and BACH1 may control POR via regulating transcription of NADPH quinone dehydrogenase 1, which results in facilitating or inhibiting ferroptosis. TFR, transferrin receptor; STEAP3, six-transmembrane epithelial antigen of the prostate 3; DMT, divalent metal transporter; GSH, glutathione synthase; GSSG, oxidized glutathione; Cys, cysteine; 4-HNE, 4-hydroxynonenal; CoQ₁₀H2, reduced coenzyme q10; GPX4, glutathione peroxidase 4; BH₄, tetrahydrobiopterin; FSP1, ferroptosis suppressor protein 1; HO•, hydroxy radical; PL•, PLOO•, PLO•, phospholipid hydroxy radical; PLOOH, phospholipid hydroperoxide.

Figure 4

Hypothetical scheme of the molecular pathways of necroptosis. Binding TNF-α with TNF-R1 induces complex I formation which includes TRADD, TRAF2, RIPK1, and cellular inhibitors of apoptosis protein 1/2 (cIAP1/2). Once NF-κB target protein synthesis is inhibited, complex IIa is activated followed by caspase-3 activation and induction of apoptosis in an RIPK1-independent manner. The inhibition of RIPK1 ubiquitination or phosphorylation induces complex IIb activation which results in RIPK1-dependent apoptosis. Once caspase-8 is inhibited, RIPK1, RIPK3, and MLKL form the necrosome which results in MLKL phosphorylation and oligomerization. MLKL integrates with the plasma membrane and forms the MLKL pore. TRIF- and ZBP1-mediated necroptosis are independent of RIPK1. RIPK3 is a more critical factor than RIPK1 in the process of necroptosis. FasL, Fas ligand; DR4/5, death receptor 4/5.