Attenuation of Microglial Activation and Pyroptosis by Inhibition of P2X7 Pathway Promotes Photoreceptor Survival in Experimental Retinal Detachment.

Purpose: Photoreceptor (PR) death is the ultimate cause of irreversible vision loss in retinal detachment (RD). Although microglial infiltration in the subretinal space (SRS) was observed after RD, the molecular mechanism underlying microglial activation and the outcomes of infiltrating microglia remain unclear. We aimed to uncover the mechanism of initiation of microglial activation to help explore potential therapy to promote PR survival. Methods: An RD model was conducted by injecting sodium hyaluronate into SRS of C57BL/6J wild type mice. Adenosine triphosphate (ATP) was measured by a ATP Microplate Assay Kit. Bioinformatics analysis was used to evaluate the upregulated receptor relating to ATP binding in human datasets and mouse transcriptomes of RD. Expression of P2X7, its downstream signaling pathways, and microglial pyroptosis were confirmed by qPCR, WB, and immunofluorescence in vivo and in vitro. The cell viability of PR was measured by cell counting kit-8. Brilliant Blue G, a P2X7 antagonist, was subretinally or intraperitoneally injected to inhibit microglial activation in vivo and was applied for microglia cell line treatment in vitro. The decrease in microglial activation and pyroptosis was detected by immunofluorescence and WB. The protective effect on PR was measured by hematoxylin and eosin staining, TUNEL assay, and electroretinogram analysis. Results: The results showed that extracellular ATP released in the SRS after RD triggered P2X7 activation and attracted microglia. The downstream cascade of inflammasome activation induced by P2X7 activation contributed to microglial pyroptosis and then to PR death. ATP-activated microglia led to PR death in vitro. P2X7 blockade rescued PR morphologically and functionally by inhibiting microglial activation and pyroptosis. Conclusions: These results elucidate that ATP-induced P2X7-mediated microglial activation leads to microglial pyroptosis, contributing to PR death. Appropriate inhibition of microglial pyroptosis might serve as a pharmacotherapeutic strategy for decreasing PR death in RD.

Bioinformatics analyses of retinoblastoma reveal the retinoblastoma progression subtypes.

INTRODUCTION: Retinoblastoma (RB) is one common pediatric malignant tumor with dismal outcomes. Heterogeneity of RB and subtypes of RB were identified but the association between the subtypes of RB and RB progression have not been fully investigated. METHODS: Four public datasets were downloaded from Gene expression omnibus and normalization was performed to remove batch effect. Two public datasets were explored to obtain the RB progression gene signatures by differentially expression analysis while another two datasets were iterated for RB subtypes identification using consensus clustering. After the RB progressive subtype gene signatures were identified, we tested the diagnostic capacity of these gene signatures by receiver operation curve. RESULTS: Three hundreds and forty six genes that were enriched in cell cycle were identified as the progression signature in RB from two independent datasets. Four subtypes of RB were stratified by consensus clustering. A total of 21 genes from RB progression signature were differentially expressed between RB subtypes. One subtype with low expression cell division genes have less progression of all four subtypes. A panel of five RB subtype genes (CLUL1, CNGB1, ROM1, LRRC39 and RDH12) predict progression of RB. CONCLUSION: Retinoblastoma is a highly heterogeneous tumor and the level of cell cycle related gene expression is associated with RB progression. A subpopulation of RB with high expression of visual perception has less progressive features. LRRC39 is potentially the RB progression subtype biomarker.

Identifying circRNA-associated-ceRNA networks in retinal neovascularization in mice.

Retinal neovascularization is a complication which caused human vision loss severely. It has been shown that circular RNAs (circRNAs) play essential roles in gene regulation. However, circRNA expression profile and the underlying mechanisms in retinal neovascular diseases remain unclear. In the present study, we identified altered circRNAs in the retinas of oxygen-induced retinopathy (OIR) mouse model by microarray profiling. Microarray analysis revealed that 539 circRNAs were significantly altered in OIR retinas compared with controls. Among them, 185 up-regulated and 354 down-regulated circRNAs were identified. The expression levels of 4 altered circRNAs including mmu_circRNA_002573, mmu_circRNA_011180, mmu_circRNA_016108 and mmu_circRNA_22546 were validated by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Bioinformatic analysis with validated circRNAs such as competing endogenous RNA (ceRNA) regulatory networks with Gene Ontology (GO) enrichment analysis demonstrated that qRT-PCR validated circRNAs were associated with cellular process, cell part and phosphoric ester hydrolase activity. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis demonstrated that MAPK signaling pathway and renin-angiotensin system were related to validated circRNAs, suggesting these pathways may participate in pathological angiogenesis. The results together suggested that circRNAs were aberrantly expressed in OIR retinas and may play potential roles in retinal neovascular diseases.

Attenuation of periostin in retinal Müller glia by TNF-α and IFN-γ.

AIM: To investigate the regulation and mechanisms of periostin expression in retinal Müller glia, and to explore the relevance to retinal neovascularization. METHODS: The oxygen-induced retinopathy (OIR) mouse model and the human Moorfield/Institute of Ophthalmology-Müller 1 (MIO-M1) cell line were used in the study. Immunofluorescence staining was used to determine the distribution and expression of periostin and a Müller glial cell marker glutamine synthetase (GS). Cytokines TNF-α and IFN-γ were added to stimulate the MIO-M1 cells. ShRNA was used to knockdown periostin expression in MIO-M1 cells. Quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) was conducted to assess the mRNA expression of periostin. RESULTS: Immunofluorescence staining showed that periostin was expressed by MIO-M1 Müller glia. GS-positive Müller glia and periostin increased in OIR retinas, and were partially overlaid. The stimulation of TNF-α and IFN-γ reduced the mRNA expression of periostin significantly and dose-dependently in MIO-M1 cells. Knockdown of periostin reduced mRNA expression of vascular endothelial growth factor A (VEGFA) in MIO-M1 cells, while VEGFA expression was not changed in periostin knock-out OIR retinas. CONCLUSION: Müller glia could be one of the main sources of periostin in the retina, and might contribute to the pathogenesis of retinal neovascularization. Proinflammatory cytokines TNF-α and IFN-γ attenuate the periostin expression in retinal Müller glia, which provides a potential and novel method in treating retinal neovascular diseases.