title: Retinal ischemia/reperfusion injury is mediated by Toll-like receptor 4 activation of NLRP3 inflammasomes.
abstract: OBJECTIVE
  Retinal ischemia/reperfusion (IR) is common in eye disorders. Pattern-recognition receptors (PRRs) are reported to initiate sterile inflammatory response. The role of PRRs in retinal IR injury is currently unknown. Thus, we investigated the expression and function of membrane and cytoplasmic PRRs during retinal IR.
  
  
  METHODS
  Retinal IR was induced in adult Brown Norway rats by clipping the retinal vessels for 30 minutes. RNA and proteins were extracted during the course of reperfusion, and the expression levels of the following proteins were determined: Toll-like receptor 2 (TLR2), TLR4, myeloid differentiation factor 88 (MyD88), TNF receptor-associated factor 6 (TRAF6), nuclear factor-κB (NF-κB), nucleotide-binding oligomerization domain-like receptor with pyrin domain protein 1 (NLRP1), NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), caspase-1, IL-1β, and IL-18. TLR4 expression in the retina was studied using immunohistochemistry. In addition, a TLR4 inhibitor was injected into the vitreous body as a therapeutic agent. After the treatment of TLR4 inhibitors, the levels of the above factors were evaluated, the apoptosis of cells in the retina, expression of cleaved-caspase-3 (c-casp-3), death of retinal ganglion cells, and the retina electroretinography was assessed.
  
  
  RESULTS
  After releasing the artery clamp, the retinal vessels were reperfused in 5 minutes. During the reperfusion, TLR4, MyD88, TRAF6, NF-κB, NLRP1, NLRP3, mature IL-1β, and IL-18 were upregulated, but not TLR2. In the IR model, TLR4 was highly expressed in ganglion cell and glia cell. Additionally, the inhibition of TLR4 significantly downregulated the activation of NLRP3, but not NLRP1, and the secretion of mature IL-1β and IL-18 also were inhibited. Moreover, the TLR4 inhibitor partially attenuated the injury of the retina, including alleviated retina apoptosis, downregulated c-casp-3 expression, rescued retinal ganglion cells death, and restored retina function.
  
  
  CONCLUSIONS
  These findings suggest that TLR4-signaling activation, triggered by damage-associated molecular patterns, regulates the activation of the NLRP3 inflammasomes and is responsible for the function of the retina in retinal IR injury.
authors.affiliation: Key Laboratory of Vision Loss and Restoration, Ministry of Education; Beijing Key Laboratory of Diagnosis and Therapy of Retinal and Choroid Diseases; Department of Ophthalmology, Peking University People's Hospital, Beijing, China.
authors.forename: Yun
authors.initials: Y
authors.lastname: Qi
authors.affiliation: Key Laboratory of Vision Loss and Restoration, Ministry of Education; Beijing Key Laboratory of Diagnosis and Therapy of Retinal and Choroid Diseases; Department of Ophthalmology, Peking University People's Hospital, Beijing, China.
authors.forename: Min
authors.initials: M
authors.lastname: Zhao
authors.affiliation: Key Laboratory of Vision Loss and Restoration, Ministry of Education; Beijing Key Laboratory of Diagnosis and Therapy of Retinal and Choroid Diseases; Department of Ophthalmology, Peking University People's Hospital, Beijing, China.
authors.forename: Yujing
authors.initials: Y
authors.lastname: Bai
authors.affiliation: Key Laboratory of Vision Loss and Restoration, Ministry of Education; Beijing Key Laboratory of Diagnosis and Therapy of Retinal and Choroid Diseases; Department of Ophthalmology, Peking University People's Hospital, Beijing, China.
authors.forename: Lvzhen
authors.initials: L
authors.lastname: Huang
authors.affiliation: Key Laboratory of Vision Loss and Restoration, Ministry of Education; Beijing Key Laboratory of Diagnosis and Therapy of Retinal and Choroid Diseases; Department of Ophthalmology, Peking University People's Hospital, Beijing, China.
authors.forename: Wenzhen
authors.initials: W
authors.lastname: Yu
authors.affiliation: Department of Ophthalmology and Visual Sciences, University of Michigan Kellogg Eye Center, Ann Arbor, Michigan, United States.
authors.forename: Zongmei
authors.initials: Z
authors.lastname: Bian
authors.affiliation: Key Laboratory of Vision Loss and Restoration, Ministry of Education; Beijing Key Laboratory of Diagnosis and Therapy of Retinal and Choroid Diseases; Department of Ophthalmology, Peking University People's Hospital, Beijing, China.
authors.forename: Mingwei
authors.initials: M
authors.lastname: Zhao
authors.affiliation: Key Laboratory of Vision Loss and Restoration, Ministry of Education; Beijing Key Laboratory of Diagnosis and Therapy of Retinal and Choroid Diseases; Department of Ophthalmology, Peking University People's Hospital, Beijing, China.
authors.forename: Xiaoxin
authors.initials: X
authors.lastname: Li
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002352:Carrier Proteins; D002462:Cell Membrane / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D007150:Immunohistochemistry; D058847:Inflammasomes / Q000378:metabolism*; D008297:Male; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D051192:Receptors, Pattern Recognition / Q000378:metabolism / Q000502:physiology*; D015427:Reperfusion Injury / Q000378:metabolism*; D012164:Retinal Diseases / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D051197:Toll-Like Receptor 4 / Q000502:physiology*
chemical_list: D015415:Biomarkers; D002352:Carrier Proteins; D058847:Inflammasomes; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; C573346:Nlrp3 protein, rat; D018160:Receptors, Cytoplasmic and Nuclear; D051192:Receptors, Pattern Recognition; D051197:Toll-Like Receptor 4
keywords: IL-18; IL-1β; Toll-like receptor 4; inflammasomes; ischemia-reperfusion; retina