Omega-3 From Fish Oil Augments Gvhd Through the Enhancement of Chemotherapy Conditioning Regimen and Selective Foxp3 Depletion

Omega-3 has been reported to enhance the effects of some cancer chemotherapeutic agents and to play a role in the immune system as immunoregulator and immunosuppressant. The effect of a diet rich in omega-3 during immunosuppressed states of anticancer treatment and on bone marrow transplantation (BMT) outcome is not well known.

To study the effect of omega-3 on busulfan-cyclophosphamide (Bu-Cy) conditioning regimen. Moreover, to evaluate the effect of omega-3 on the outcome of BMT after Bu-Cy conditioning.

We evaluated the effects of menhadenfish oil (omega-3 rich) on the myeloablative treatment with Bu-Cy as well as on the outcome of BMT in mice compared to the effects of a diet containing corn oil (omega-6 rich) or to standard chow.

Animals were divided into three groups, Group 1, Group 2 and Group 3. In all the groups, female BALB/c mice, 8–12 weeks old, were randomly selected to receive a diet enriched with fish oil, a diet enriched with corn oil or standard chow. The mice were fed for two weeks prior to conditioning. Thereafter, all the mice were injected IP with 80 mg/kg busulfan (Bu) for four days (day –7 to day –4) followed by 200 mg/kg cyclophosphamide (Cy) for two days (days –3 and −2). The mice in groups 1 and 2 were killed on day 0 (the proposed day for transplantation) and spleen and femur bone marrow cells were harvested. Organs were analysed with flow cytometry to evaluate the lymphoid and myeloid cells phenotype. Also, regulatory T cell number and function were studied. Mice in Group 3 were transplanted on day 0 with allogeneic bone marrow cells from 8–12 weeks old male C57BL/6 mice (2.0 × 10⁷ bone marrow cells and 3.0 × 10⁷spleen cells).

Fish oil-enriched chow significantly suppressed the function (Figure 1) of CD4+CD25+FoxP3+ regulatory T (T_reg) cells compared to standard chow and corn oil fed mice at day 0. However, the percentage of FoxP3+ cells was significantly elevated in mice fed corn oil compared to standard chow. In concert with myeloablative chemotherapy, fish oil enhanced the suppressive effect of Bu-Cy on CD11b+ myeloid and CD11c+CD86+ dendritic cell populations in the bone marrow and spleen compared to the other dietary groups.

Recipient mice fed fish oil had lower survival after BMT (Figure 2). A higher survival was observed for corn oil-fed recipients, but this survival rate was still inferior to that of recipients fed standard chow. Feeding recipients omega-3 enriched diet was associated with a more pronounced acute graft versus host disease (aGVHD). BM and spleen from mice fed corn oil demonstrated lower chimerism.

Fish oil enhances the immunosuppressive effect of the conditioning regimen (Bu-Cy) in mice. BMT recipients fed a diet rich in polyunsaturated fatty acids either have lower engraftment (corn oil) or react with a lethal aGVHD (fish oil) associated with a suppression of Treg cells. These findings may have clinical implications. PUFA supplementation should be reconsidered and caution is advised in patients undergoing stem cell transplantation.

Figure 1.

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Mixed lymphocyte reaction assay to study the regulatory T cell function. The ratio between the responder cells and Treg cells was 1 to 0.5. The stimulator was anti-CD3 in 3μg/ml and 10μg/ml.

Figure 1.

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Mixed lymphocyte reaction assay to study the regulatory T cell function. The ratio between the responder cells and Treg cells was 1 to 0.5. The stimulator was anti-CD3 in 3μg/ml and 10μg/ml.

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Figure 2.

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Survival curve after SCT in mice received Bu-Cy conditioning and fed omega-3, corn oil or standard food.

Figure 2.

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Survival curve after SCT in mice received Bu-Cy conditioning and fed omega-3, corn oil or standard food.

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No relevant conflicts of interest to declare.