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Fast and Efficient Genome Editing of Human FOXP3⁺ Regulatory T Cells.

ABSTRACT: FOXP3⁺ regulatory T cells (Tregs) are central for maintaining peripheral tolerance and immune homeostasis. Because of their immunosuppressive characteristics, Tregs are a potential therapeutic target in various diseases such as autoimmunity, transplantation and infectious diseases like COVID-19. Numerous studies are currently exploring the potential of adoptive Treg therapy in different disease settings and novel genome editing techniques like CRISPR/Cas will likely widen possibilities to strengthen its efficacy. However, robust and expeditious protocols for genome editing of human Tregs are limited. Here, we describe a rapid and effective protocol for reaching high genome editing efficiencies in human Tregs without compromising cell integrity, suitable for potential therapeutic applications. By deletion of IL2RA encoding for IL-2 receptor α-chain (CD25) in Tregs, we demonstrated the applicability of the method for downstream functional assays and highlighted the importance for CD25 for in vitro suppressive function of human Tregs. Moreover, deletion of IL6RA (CD126) in human Tregs elicits cytokine unresponsiveness and thus may prevent IL-6-mediated instability of Tregs, making it an attractive target to potentially boost functionality in settings of adoptive Treg therapies to contain overreaching inflammation or autoimmunity. Thus, our rapid and efficient protocol for genome editing in human Tregs may advance possibilities for Treg-based cellular therapies.

SUBMITTER: Van Zeebroeck L

PROVIDER: S-EPMC8365355 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundRegulatory T cells are known to play a key role to counter balance the protective immune response and immune mediated pathology. However, the role of naturally occurring regulatory cells CD4+CD25+Foxp3+ in malaria infection during the disease pathogenesis is controversial. Beside this, ICOS molecule has been shown to be involved in the development and function of regulatory T cell enhance IL-10 production. Therefore, possible involvement of the ICOS dependent regulatory CD4+ICOS+Foxp3+ T cells in resistance/susceptibility during malaria parasite is explored in this study.Methods5 × 105 red blood cells infected with non-lethal and lethal parasites were inoculated in female Balb/c mice by intra-peritoneal injection. Infected or uninfected mice were sacrificed at early (3rd day post infection) and later stage (10th day post infection) of infection. Harvested cells were analysed by using flow cytometer and serum cytokine by Bioplex assay.ResultsThin blood films show that percentages of parasitaemia increases with disease progression in infections with the lethal malaria parasite and mice eventually die by day 14th post-infection. Whereas in case of non-lethal malaria parasite, parasitaemia goes down by 7th day post infection and gets cleared within 13th day. The number of CD4+ ICOS+ T cells increases in lethal infection with disease progression. Surprisingly, in non-lethal parasite, ICOS expression decreases after day 7th post infection as parasitaemia goes down. The frequency of CD4+ICOS+FoxP3+ Tregs was significantly higher in lethal parasitic infection as compared to the non-lethal parasite. The level of IL-12 cytokine was remarkably higher in non-lethal infection compared to the lethal infection. In contrast, the level of IL-10 cytokines was higher in lethal parasite infection compared to the non-lethal parasite.ConclusionTaken together, these data suggest that lethal parasite induce immunosuppressive environment, protecting from host immune responses and help the parasite to survive whereas non-lethal parasite leads to low frequencies of Treg cells seldom impede immune response that allow the parasite to get self-resolved.

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Fast and Efficient Genome Editing of Human FOXP3⁺ Regulatory T Cells.

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OmicsDI is part of the ELIXIR infrastructure

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Dataset Information

Fast and Efficient Genome Editing of Human FOXP3+ Regulatory T Cells.

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OmicsDI is part of the ELIXIR infrastructure

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Fast and Efficient Genome Editing of Human FOXP3⁺ Regulatory T Cells.