Early Gene Expression Profile in Retinal Ganglion Cell Layer After Optic Nerve Crush in Mice

Ueno, Satoru; Yoneshige, Azusa; Koriyama, Yoshiki; Hagiyama, Man; Shimomura, Yoshikazu; Ito, Akihiko

doi:10.1167/iovs.17-22438

Cited by 21 publications

(23 citation statements)

References 107 publications

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“…Atf3 was previously found to be significantly changed in ONC GCL [12]. This result was in accord with our results.…”

Section: Discussionsupporting

confidence: 93%

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Laser capture microdissection-based single-cell RNA sequencing reveals optic nerve crush-related early mRNA alterations in retinal ganglion cells

Pan

Chang

et al. 2020

Preprint

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Retinal ganglion cells (RGC) are the primary cell type injured in a variety of diseases of the optic nerve, and the early changes of RGC's RNA profiling may be important to understand the mechanism of optic nerve injury and axon regeneration. Here we employed the optic nerve crush (ONC) model to explore early mRNA alterations in RGCs using laser capture microdissection (LCM) and single-cell RNA sequencing. We successfully established an optimal LCM protocol using 30 μm-thick retinal tissue sections mounted on glass slides and laser pressure catapulting (LPC) to collect RGCs and obtain high-quality RNA for single-cell sequencing. Based on our protocol, we identified 8744 differentially expressed genes that were involved in ONC-related early mRNA alterations in RGCs. Candidate genes included Atf3, Lgals3, LOC102551701, Plaur, Tmem140 and Maml1. The LCM-based single-cell RNA sequencing allowed new insights into the early mRNA changes in RGCs, highlighting new molecules associated with ONC.

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“…Atf3 was previously found to be significantly changed in ONC GCL [12]. This result was in accord with our results.…”

Section: Discussionsupporting

confidence: 93%

“…LCM is often used to study the expression of both minimally or abundantly expressed genes in different brain regions [5][6][7][8][9] and occasionally in retinas [10][11][12]. In our study, we adopted an optimal LCM protocol with single-cell sequencing to examine the early mRNA changes in RGCs in an optic nerve crush model.…”

Section: Discussionmentioning

confidence: 99%

Laser capture microdissection-based single-cell RNA sequencing reveals optic nerve crush-related early mRNA alterations in retinal ganglion cells

Pan

Chang

et al. 2020

Preprint

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“…Atf3 was previously found to be significantly changed in ONC RGC layer. 12 , 15 This was in accord with our results, and it also proved the reliability of our results. Atf3 was reported to be involved in axonal regeneration, because Atf3 expression was activated after crush, and axonal regeneration was enhanced in various types of neurons as a result.…”

Section: Discussionsupporting

confidence: 93%

“…LCM is often used to study the expression of both minimally or abundantly expressed genes in different brain regions 5 – 9 and occasionally in retinas. 10 – 12 In our study, we adopted an optimal LCM protocol with microsequencing to examine the early mRNA changes in RGC layer in an optic nerve crush model.…”

Section: Discussionmentioning

confidence: 99%

Laser Capture Microdissection-Based RNA Microsequencing Reveals Optic Nerve Crush-Related Early mRNA Alterations in Retinal Ganglion Cell Layer

Pan

Chang

et al. 2020

Trans. Vis. Sci. Tech.

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Purpose To establish a method of laser capture microdissection (LCM) and RNA microsequencing for exploring optic nerve crush (ONC)–related early mRNA alterations in retinal ganglion cell (RGC) layer. Methods An LCM protocol was developed using retinal tissue sections to obtain high-quality RNA for microsequencing. Cells in the RGC layer were collected by laser pressure catapulting (LPC) using a PALM Zeiss UV LCM system. The effect of section thickness and slide type on tissue capture success and RNA yield and the integrity after LCM were evaluated. The optimal LCM protocol was used to explore ONC-related early mRNA alterations in the RGC layer. Candidate genes were validated by real-time polymerase chain reaction of the RGC layer tissue dissected by “cut and LPC” using the same LCM system. Results We successfully established an optimal LCM protocol using 30-µm–thick retinal tissue sections mounted on glass slides and laser pressure catapulting (LPC) to collect cells in the RGC layer and to obtain high-quality RNA for microsequencing. On the basis of our protocol, we identified 8744 differentially expressed genes that were involved in ONC-related early mRNA alterations in the RGC layer. Candidate genes included Atf3, Lgals3, LOC102551701, Plaur, Tmem140, and Maml1. Conclusions The LCM-based single-cell RNA sequencing allowed a new sight into the early mRNA changes of RGCs highlighting new molecules associated to ONC. Translational Relevance This technique will be helpful for more accurate transcriptome analysis of clinical pathological samples of ophthalmology and provide important reference for the discovery of new pathological diagnosis indicators and drug development targets.

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“…While the majority of studies on the role of LCN-2 in ocular health focus on intraocular inflammation, a few studies demonstrated that LCN-2 is involved in a number of optic nerve disorders. For example, LCN-2 expression was increased > 80 fold in the ganglion cells following optic nerve crush-a popular model for glaucoma and neuronal apoptosis due to axonal degeneration [102]. LCN-2 was also found to regulate inflammation in the optic nerve during the progression of optic neuritis [103].…”

Section: Role Of Lcn-2 In Ocular Diseases: Focus On Age-related Maculmentioning

confidence: 99%

The role of lipocalin-2 in age-related macular degeneration (AMD)

Ghosh

Stepicheva

Yazdankhah

et al. 2020

Cell. Mol. Life Sci.

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Lipocalins are a family of secreted adipokines which play important roles in various biological processes. Lipocalin-2 (LCN-2) has been shown to be involved in acute and chronic inflammation. This particular protein is critical in the pathogenesis of several diseases including cancer, diabetes, obesity, and multiple sclerosis. Herein, we discuss the general molecular basis for the involvement of LCN-2 in acute infections and chronic disease progression and also ascertain the probable role of LCN-2 in ocular diseases, particularly in age-related macular degeneration (AMD). We elaborate on the signaling cascades which trigger LCN-2 upregulation in AMD and suggest therapeutic strategies for targeting such pathways.Keywords Lipocalin-2 (LCN-2) · Age-related macular degeneration (AMD) · Inflammation · AKT2 signaling · Retinal degeneration Cellular and Molecular Life SciencesElectronic supplementary material The online version of this article (https ://doi.Publisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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Early Gene Expression Profile in Retinal Ganglion Cell Layer After Optic Nerve Crush in Mice

Abstract: In response to ONC, the expression of apoptotic genes was stimulated soon after crush. Atf3, Lcn2, and Tnfrsf12a might be key molecules responsible for RGC loss in glaucoma.

Cited by 21 publications

References 107 publications

Laser capture microdissection-based single-cell RNA sequencing reveals optic nerve crush-related early mRNA alterations in retinal ganglion cells

Laser capture microdissection-based single-cell RNA sequencing reveals optic nerve crush-related early mRNA alterations in retinal ganglion cells

Laser Capture Microdissection-Based RNA Microsequencing Reveals Optic Nerve Crush-Related Early mRNA Alterations in Retinal Ganglion Cell Layer

The role of lipocalin-2 in age-related macular degeneration (AMD)

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