Microdissection of the Rodent Eye

Mohan, K. Varsha; Sinha, Preema; Swami, Bharati; Muniyasamy, Abaranjitha; Nagarajan, Priyadharsini; Upadhyay, Pramod

doi:10.3791/64414

Cited by 1 publication

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“…The eyeballs were micro-dissected for further analysis. Only the retina was micro-dissected from the eyecup and was mechanically disrupted in sterile PBS to obtain single-cell suspension [41]. The suspension was filtered through a 70-micron nylon filter and centrifuged at 300g for 10 minutes.…”

Section: Enucleationmentioning

confidence: 99%

iTreg mediated TGF-Β1 therapy improves functional engraftment of cell therapy in rd1 Retinitis Pigmentosa mouse model

Mohan,

Mishra,

Sinha

et al. 2024

Preprint

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Purpose: Retinitis Pigmentosa (RP) is a progressive and hereditary disease that primarily affects the retina, leading to partial or complete vision loss. In addition to the direct impact on vision, the degeneration of the retina in RP also leads to inflammation in the eye, which can further damage the retina and make it difficult to treat the condition with cell therapy. This inflammation led to oxidative stress and cell death, creating an unfavourable environment for the introduction of new cells via cell therapy. Methods: The potential of Transforming Growth Factor-Beta1 (TGF-Β1) as an anti-inflammatory agent to treat ocular inflammation was investigated done by administering TGF-B1 intravitreally to the eyes of rd1 mice. However, due to the transient effect of TGF-B1 injection, the in-vitro-induced Treg (iTregs) cells that secrete TGF-Β1, were generated and transplanted into the conjunctiva of 4 weeks old rd1 mice to achieve a sustained release of TGF-B1. After administering iTregs, Retinal Neuron-Like Cells (RNLCs) were transplanted into the rd1 mouse retina as a form of cell therapy to improve vision perception. Flow cytometry was used to estimate the number of Qtracker labelled RNLCs post 30 days of transplantation. The potential of iTregs as an adjunct transplantation with RNLCs to improve cell therapy survival and vision rescue was investigated by conducting Electroretinography and behavioural studies. Results: The study found that ocular inflammation can be reduced by treating with TGF-Β1. After 30 days, mice transplanted with iTregs showed a significant increase in the number of transplanted RNLCs that survived compared to the mice who only received RNLCs. In the total fluid of the eye (aqueous plus vitreous), there was a significant increase in the levels of anti-inflammatory cytokines TGF-Β1 and IL-10, and some decrease in the levels of pro-inflammatory cytokines Monocyte Chemoattractant Protein-1 (MCP1). The adjunct therapy of iTregs transplantation resulted in improvement in ERG wave functions and vision preservation compared to the group without adjunct iTregs. Conclusions: The administration of TGF-Β1-secreting iTregs to the affected eye reduced the inflammatory environment, which enabled transplanted RNLCs to stay longer compared to without TGF-B1. The iTregs mediated sustained anti-inflammatory adjunct therapy can improve the outcome of cell therapy for RP.

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Section: Enucleationmentioning

confidence: 99%