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Role of Small Intestine and Gut Microbiome in Plant-Based Oral Tolerance for Hemophilia.

ABSTRACT: Fusion proteins, which consist of factor VIII or factor IX and the transmucosal carrier cholera toxin subunit B, expressed in chloroplasts and bioencapsulated within plant cells, initiate tolerogenic immune responses in the intestine when administered orally. This approach induces regulatory T cells (Treg), which suppress inhibitory antibody formation directed at hemophilia proteins induced by intravenous replacement therapy in hemophilia A and B mice. Further analyses of Treg CD4⁺ lymphocyte sub-populations in hemophilia B mice reveal a marked increase in the frequency of CD4⁺CD25^-FoxP3^-LAP⁺ T cells (but not of CD4⁺CD25⁺FoxP3⁺ T cells) in the lamina propria of the small but not large intestine. The adoptive transfer of very small numbers of CD4⁺CD25^-LAP⁺ Treg isolated from the spleen of tolerized mice was superior in suppression of antibodies directed against FIX when compared to CD4⁺CD25⁺ T cells. Thus, tolerance induction by oral delivery of antigens bioencapsulated in plant cells occurs via the unique immune system of the small intestine, and suppression of antibody formation is primarily carried out by induced latency-associated peptide (LAP) expressing Treg that likely migrate to the spleen. Tolerogenic antigen presentation in the small intestine requires partial enzymatic degradation of plant cell wall by commensal bacteria in order to release the antigen. Microbiome analysis of hemophilia B mice showed marked differences between small and large intestine. Remarkably, bacterial species known to produce a broad spectrum of enzymes involved in degradation of plant cell wall components were found in the small intestine, in particular in the duodenum. These were highly distinct from populations of cell wall degrading bacteria found in the large intestine. Therefore, FIX antigen presentation and Treg induction by the immune system of the small intestine relies on activity of a distinct microbiome that can potentially be augmented to further enhance this approach.

SUBMITTER: Kumar SRP

PROVIDER: S-EPMC7251037 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Role of Small Intestine and Gut Microbiome in Plant-Based Oral Tolerance for Hemophilia.

Kumar Sandeep R P SRP Wang Xiaomei X Avuthu Nagavardhini N Bertolini Thais B TB Terhorst Cox C Guda Chittibabu C Daniell Henry H Herzog Roland W RW

Frontiers in immunology 20200520

Fusion proteins, which consist of factor VIII or factor IX and the transmucosal carrier cholera toxin subunit B, expressed in chloroplasts and bioencapsulated within plant cells, initiate tolerogenic immune responses in the intestine when administered orally. This approach induces regulatory T cells (Treg), which suppress inhibitory antibody formation directed at hemophilia proteins induced by intravenous replacement therapy in hemophilia A and B mice. Further analyses of Treg CD4<sup>+</sup> ly ...[more]

PMID: 32508814

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Project description:The main complication of hemophilia A treatment is the development of neutralizing antibodies (inhibitors) against factor VIII (FVIII). Immune tolerance induction (ITI) is the prescribed treatment for inhibitor eradication, although its working mechanism remains unresolved. To clarify this mechanism, we compared blood samples of hemophilia A patients with and without inhibitors for presence of immunoregulatory cells and markers, including regulatory B-cells (Bregs), regulatory T-cells (Tregs), myeloid-derived suppressor cells (MDSCs), and expression of regulatory markers on T-cells (programmed cell death protein 1 [PD1], inducable T-cell costimulator, cytotoxic T-lymphocyte-associated protein 4 [CTLA4]), by use of flow cytometry. By cross-sectional analysis inhibitor patients (N = 20) were compared with inhibitor-negative (N = 28) and ex-inhibitor (N = 17) patients. In another longitudinal study, changes in immunoregulatory parameters were evaluated during ITI (N = 12) and compared with inhibitor-negative hemophilia A patients (N = 36). The frequency of Bregs, but not of Tregs nor MDSCs, was significantly reduced in inhibitor patients (3.2%) compared with inhibitor-negative (5.9%) and ex-inhibitor patients (8.9%; P < 0.01). CTLA4 expression on T-cells was also reduced (mean fluorescence intensity 133 in inhibitor versus 537 in inhibitor-negative patients; P < 0.01). Fittingly, in patients followed during ITI, inhibitor eradication associated with increased Bregs, increased Tregs, and increased expression of CTLA4 and PD1 on CD4+ T-cells. In conclusion, inhibitor patients express significantly lower frequency of Bregs and Tregs marker expression, which are restored by successful ITI. Our findings suggest that an existing anti-FVIII immune response is associated with deficits in peripheral tolerance mechanisms and that Bregs and changes in immunoregulatory properties of CD4+ T-cells likely contribute to ITI in hemophilia A patients with inhibitors.

Dataset Information

Role of Small Intestine and Gut Microbiome in Plant-Based Oral Tolerance for Hemophilia.

Publications

Role of Small Intestine and Gut Microbiome in Plant-Based Oral Tolerance for Hemophilia.

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