Depletion of the receptor-interacting protein kinase 3 (RIP3) decreases photoreceptor cell death during the early stages of ocular murine cytomegalovirus infection

Jinxian Xu, Juan Mo, Xinglou Liu, Brendan Marshall, Sally S. Atherton, Zheng Dong, Sylvia Smith, Ming Zhang

Research output: Contribution to journalArticle

Abstract

PURPOSE. The purpose of this study was to determine if the receptor-interacting protein kinase 3 (RIP3) plays a significant role in innate immune responses and death of bystander retinal neurons during murine cytomegalovirus (MCMV) retinal infection, by comparing the innate immune response and cell death in RIP3-depleted mice (Rip3 -/- ) and Rip3 +/+ control mice. METHODS. Rip3 -/- and Rip3 +/+ mice were immunosuppressed (IS) and inoculated with MCMV via the supraciliary route. Virus-injected and mock-injected control eyes were removed at days 4, 7, and 10 post infection (p.i.) and markers of innate immunity and cell death were analyzed. RESULTS. Compared to Rip3 +/+ mice, significantly more MCMV was recovered and more MCMV-infected RPE cells were observed in injected eyes of Rip3 -/- mice at days 4 and 7 p.i. In contrast, fewer TUNEL-stained photoreceptors were observed in Rip3 -/- eyes than in Rip3 +/+ eyes at these times. Electron microscopy showed that significantly more apoptotic photoreceptor cells were present in Rip3 +/+ mice than in Rip3 -/- mice. Immunohisto-chemistry showed that the majority of TUNEL-stained photoreceptors died via mitochondrial flavoprotein apoptosis-inducing factor (AIF)-mediated, caspase 3–independent apoptosis. The majority of RIP3-expressing cells in infected eyes were RPE cells, microglia/macrophages, and glia, whereas retinal neurons contained much lower amounts of RIP3. Western blots showed significantly higher levels of activated nuclear factor–jB and caspase 1 were present in Rip3 +/+ eyes compared to Rip3 -/- eyes. CONCLUSIONS. Our results suggest that RIP3 enhances innate immune responses against ocular MCMV infection via activation of the inflammasome and nuclear factor–jB, which also leads to inflammation and death of bystander cells by multiple pathways including apoptosis and necroptosis.

Original languageEnglish (US)
Pages (from-to)2445-2458
Number of pages14
JournalInvestigative Ophthalmology and Visual Science
Volume59
Issue number6
DOIs
StatePublished - May 2018

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Receptor-Interacting Protein Serine-Threonine Kinases
Muromegalovirus
Photoreceptor Cells
Cytomegalovirus Infections
Protein Kinases
Cell Death
Innate Immunity
Retinal Neurons
In Situ Nick-End Labeling
Apoptosis Inducing Factor
Apoptosis
Inflammasomes
Flavoproteins
Caspase 1
Death Domain Receptors
Microglia
Caspases
Infection
Neuroglia

Keywords

  • Apoptosis
  • Innate immune response
  • Murine cytomegalovirus
  • Retinitis

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

Cite this

Depletion of the receptor-interacting protein kinase 3 (RIP3) decreases photoreceptor cell death during the early stages of ocular murine cytomegalovirus infection. / Xu, Jinxian; Mo, Juan; Liu, Xinglou; Marshall, Brendan; Atherton, Sally S.; Dong, Zheng; Smith, Sylvia; Zhang, Ming.

In: Investigative Ophthalmology and Visual Science, Vol. 59, No. 6, 05.2018, p. 2445-2458.

Research output: Contribution to journalArticle

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abstract = "PURPOSE. The purpose of this study was to determine if the receptor-interacting protein kinase 3 (RIP3) plays a significant role in innate immune responses and death of bystander retinal neurons during murine cytomegalovirus (MCMV) retinal infection, by comparing the innate immune response and cell death in RIP3-depleted mice (Rip3 -/- ) and Rip3 +/+ control mice. METHODS. Rip3 -/- and Rip3 +/+ mice were immunosuppressed (IS) and inoculated with MCMV via the supraciliary route. Virus-injected and mock-injected control eyes were removed at days 4, 7, and 10 post infection (p.i.) and markers of innate immunity and cell death were analyzed. RESULTS. Compared to Rip3 +/+ mice, significantly more MCMV was recovered and more MCMV-infected RPE cells were observed in injected eyes of Rip3 -/- mice at days 4 and 7 p.i. In contrast, fewer TUNEL-stained photoreceptors were observed in Rip3 -/- eyes than in Rip3 +/+ eyes at these times. Electron microscopy showed that significantly more apoptotic photoreceptor cells were present in Rip3 +/+ mice than in Rip3 -/- mice. Immunohisto-chemistry showed that the majority of TUNEL-stained photoreceptors died via mitochondrial flavoprotein apoptosis-inducing factor (AIF)-mediated, caspase 3–independent apoptosis. The majority of RIP3-expressing cells in infected eyes were RPE cells, microglia/macrophages, and glia, whereas retinal neurons contained much lower amounts of RIP3. Western blots showed significantly higher levels of activated nuclear factor–jB and caspase 1 were present in Rip3 +/+ eyes compared to Rip3 -/- eyes. CONCLUSIONS. Our results suggest that RIP3 enhances innate immune responses against ocular MCMV infection via activation of the inflammasome and nuclear factor–jB, which also leads to inflammation and death of bystander cells by multiple pathways including apoptosis and necroptosis.",
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T1 - Depletion of the receptor-interacting protein kinase 3 (RIP3) decreases photoreceptor cell death during the early stages of ocular murine cytomegalovirus infection

AU - Xu, Jinxian

AU - Mo, Juan

AU - Liu, Xinglou

AU - Marshall, Brendan

AU - Atherton, Sally S.

AU - Dong, Zheng

AU - Smith, Sylvia

AU - Zhang, Ming

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N2 - PURPOSE. The purpose of this study was to determine if the receptor-interacting protein kinase 3 (RIP3) plays a significant role in innate immune responses and death of bystander retinal neurons during murine cytomegalovirus (MCMV) retinal infection, by comparing the innate immune response and cell death in RIP3-depleted mice (Rip3 -/- ) and Rip3 +/+ control mice. METHODS. Rip3 -/- and Rip3 +/+ mice were immunosuppressed (IS) and inoculated with MCMV via the supraciliary route. Virus-injected and mock-injected control eyes were removed at days 4, 7, and 10 post infection (p.i.) and markers of innate immunity and cell death were analyzed. RESULTS. Compared to Rip3 +/+ mice, significantly more MCMV was recovered and more MCMV-infected RPE cells were observed in injected eyes of Rip3 -/- mice at days 4 and 7 p.i. In contrast, fewer TUNEL-stained photoreceptors were observed in Rip3 -/- eyes than in Rip3 +/+ eyes at these times. Electron microscopy showed that significantly more apoptotic photoreceptor cells were present in Rip3 +/+ mice than in Rip3 -/- mice. Immunohisto-chemistry showed that the majority of TUNEL-stained photoreceptors died via mitochondrial flavoprotein apoptosis-inducing factor (AIF)-mediated, caspase 3–independent apoptosis. The majority of RIP3-expressing cells in infected eyes were RPE cells, microglia/macrophages, and glia, whereas retinal neurons contained much lower amounts of RIP3. Western blots showed significantly higher levels of activated nuclear factor–jB and caspase 1 were present in Rip3 +/+ eyes compared to Rip3 -/- eyes. CONCLUSIONS. Our results suggest that RIP3 enhances innate immune responses against ocular MCMV infection via activation of the inflammasome and nuclear factor–jB, which also leads to inflammation and death of bystander cells by multiple pathways including apoptosis and necroptosis.

AB - PURPOSE. The purpose of this study was to determine if the receptor-interacting protein kinase 3 (RIP3) plays a significant role in innate immune responses and death of bystander retinal neurons during murine cytomegalovirus (MCMV) retinal infection, by comparing the innate immune response and cell death in RIP3-depleted mice (Rip3 -/- ) and Rip3 +/+ control mice. METHODS. Rip3 -/- and Rip3 +/+ mice were immunosuppressed (IS) and inoculated with MCMV via the supraciliary route. Virus-injected and mock-injected control eyes were removed at days 4, 7, and 10 post infection (p.i.) and markers of innate immunity and cell death were analyzed. RESULTS. Compared to Rip3 +/+ mice, significantly more MCMV was recovered and more MCMV-infected RPE cells were observed in injected eyes of Rip3 -/- mice at days 4 and 7 p.i. In contrast, fewer TUNEL-stained photoreceptors were observed in Rip3 -/- eyes than in Rip3 +/+ eyes at these times. Electron microscopy showed that significantly more apoptotic photoreceptor cells were present in Rip3 +/+ mice than in Rip3 -/- mice. Immunohisto-chemistry showed that the majority of TUNEL-stained photoreceptors died via mitochondrial flavoprotein apoptosis-inducing factor (AIF)-mediated, caspase 3–independent apoptosis. The majority of RIP3-expressing cells in infected eyes were RPE cells, microglia/macrophages, and glia, whereas retinal neurons contained much lower amounts of RIP3. Western blots showed significantly higher levels of activated nuclear factor–jB and caspase 1 were present in Rip3 +/+ eyes compared to Rip3 -/- eyes. CONCLUSIONS. Our results suggest that RIP3 enhances innate immune responses against ocular MCMV infection via activation of the inflammasome and nuclear factor–jB, which also leads to inflammation and death of bystander cells by multiple pathways including apoptosis and necroptosis.

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KW - Innate immune response

KW - Murine cytomegalovirus

KW - Retinitis

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