Effects Of DHA And ARA Supplementation During Lactation And Weaning On The Development Of Oral Tolerance in Allergy-Prone Rats: Mechanistic Insights Based On Intestinal Immune Regulation

Food allergies have become one of the major health issues among children globally, closely linked to defects in the oral tolerance (OT) mechanism. Long-chain polyunsaturated fatty acids (LCPUFAs) such as arachidonic acid (ARA) and docosahexaenoic acid (DHA) play a critical role in immune system development. However, the regulatory mechanisms of early-life supplementation of ARA+DHA on the intestinal immune microenvironment and OT development remain unclear. A study published in 2024 in The Journal of Nutrition (DOI: 10.1016/j.tjnut.2024.10.021) utilized an allergy-prone Brown Norway rat model to elucidate how ARA+DHA supplementation during lactation and weaning promotes OT development through modulating gut cytokines and immunoglobulins.

I. Research Background and Experimental Design

(1) Scientific Question and Hypothesis

The establishment of OT relies on the induction of immune tolerance to food antigens by gut-associated lymphoid tissue (GALT). An imbalance in mucosal immunity (such as Th2 skewing) is central to food allergies. Previous studies have shown that early supplementation with ARA+DHA can promote OT and reduce allergy risk, but the underlying mechanisms at the gut level are not fully understood. This study hypothesized that ARA+DHA supplementation during lactation and weaning improves gut fatty acid composition, enhances mucosal immune tolerance-related cytokines (such as TGF-β, IL-2), and secretory immunoglobulin A (sIgA), thereby promoting OT development.

(2) Experimental Methods

- Animal Model and Grouping: Allergy-prone pregnant Brown Norway rats were randomly divided into control (0% ARA, 0% DHA) and ARA+DHA groups (0.45% ARA, 0.8% DHA) for intervention during lactation (0-3 weeks). Post-weaning (3-8 weeks), offspring were further divided into control and ARA+DHA groups (0.5% ARA, 0.5% DHA), forming a 2×2 factorial design (lactation × post-weaning intervention).

- Allergen Challenge: Offspring at 3 weeks old received intragastric ovalbumin (OVA) or sucrose for five consecutive days, followed by intraperitoneal OVA challenge at 7 weeks old to induce allergic reactions.

- Detection Indicators:

- Gut fatty acid composition: Levels of ARA and DHA in ileal phospholipids and triglycerides.

- Gut structure: Villus height/crypt depth (V/C) ratio in the jejunum.

- Mucosal immunity: Levels of sIgA, IL-2, IL-6, TGF-β, and fecal calprotectin (an inflammation marker).

II. Key Research Findings

(1) Dynamic Changes in Gut Fatty Acid Composition

- Specific Effects of Post-weaning Supplementation: In the post-weaning ARA+DHA group, the proportion of DHA in ileal phospholipids and triglycerides significantly increased (+32%-41%, P<0.001), while ARA levels remained unaffected. The effects of lactation supplementation did not persist into the post-weaning period but significantly reduced the proportion of saturated fatty acids (SFA) (-9.6%, P=0.01) and increased monounsaturated fatty acids (MUFA) (+15.5%, P=0.034), potentially enhancing gut barrier function by improving membrane fluidity.

(2) Promotion of Mucosal Immune Tolerance

- sIgA and Cytokine Regulation:

- Supplementation with ARA+DHA during lactation and/or weaning significantly increased mucosal sIgA levels after OVA stimulation (+28%, P=0.033). As the first line of defense in gut immunity, sIgA neutralizes antigens to reduce allergic reactions.

- Increased concentrations of IL-2 (+21%, P=0.049) and TGF-β (+19%, P=0.04) in the supplemented groups promoted the differentiation of regulatory T cells (Treg), inhibiting Th2-type inflammatory responses (e.g., IL-6 decreased by 23%, P=0.026).

- Inflammatory Marker Changes: No significant differences in fecal calprotectin levels were observed among groups, but IL-6 elevation was more pronounced in the control group post-OVA stimulation, suggesting suppressed gut inflammation in the supplemented groups.

(3) Association Between Gut Structure and OT

No significant differences in jejunal V/C ratios were found among groups, indicating that ARA+DHA's promotion of OT depends more on immune regulation than structural remodeling. This aligns with previous findings that n-3 LCPUFAs improve allergies through immune rather than mechanical barriers.

III. Mechanisms and Key Pathways

(1) Interaction Between Fatty Acid Metabolism and Immune Regulation

DHA integrates into intestinal cell membrane phospholipids, potentially altering the membrane fluidity of immune cells (e.g., dendritic cells, T cells), suppressing pro-inflammatory mediators (e.g., prostaglandin E2), and enhancing Treg cell suppressive functions. ARA, as a precursor to inflammatory mediators, may exert dual regulatory effects when supplemented in appropriate amounts by generating anti-inflammatory metabolites (e.g., lipoxins), but no changes in ARA proportions were observed in this study, suggesting DHA as the primary active component.

(2) Remodeling of the Cytokine Network

Supplementation with ARA+DHA upregulated IL-2 and TGF-β, leading to:

1. Activation of Treg Cells: TGF-β induces Foxp3+ Treg cell differentiation, inhibiting excessive activation of Th2 cells.

2. IgA Class Switching: IL-2 synergizes with TGF-β to promote B-cell differentiation into IgA-secreting plasma cells, enhancing mucosal immune tolerance.

3. Negative Feedback Regulation of Inflammation: Inhibition of pro-inflammatory cytokines like IL-6 blocks the positive feedback loop of Th2 cell polarization.

(3) "Programming Effect" of Lactation

Lactation supplementation with ARA+DHA had no lasting effect on ileal fatty acid composition but induced long-term changes in mucosal immune markers (e.g., IL-2, sIgA), suggesting that early-life nutritional interventions might program the gut immune microenvironment via epigenetic mechanisms, laying the groundwork for subsequent OT establishment.

IV. Research Significance and Clinical Implications

(1) Window of Early Nutritional Intervention

Lactation and weaning are critical periods for gut immune system development. Supplementation with ARA+DHA during these periods can induce lasting tolerance phenotypes through the "metabolism-immunity" axis. This provides a theoretical basis for designing infant formulas targeting high-risk infants, maintaining an appropriate DHA/ARA ratio (1:1 in this study during weaning).

(2) Precision Nutrition Strategies

- Dose Optimization: Post-weaning DHA content needs to reach above 0.5% to significantly alter gut fatty acid composition, while lactation ARA+DHA's "programming effect" may be achieved through breast milk transfer (e.g., a fourfold increase in breast milk DHA concentration).

- Population Screening: Early supplementation with ARA+DHA may be more effective in Th2-biased individuals (e.g., infants with a family history of atopic dermatitis).

(3) Limitations and Future Directions

This study did not explore the role of gut microbiota, although previous research suggests LCPUFAs can modulate symbiotic bacterial abundance (e.g., promoting bifidobacteria growth). Future studies should integrate microbiomics to understand the tripartite interaction between "fatty acids-microbiota-immunity." Additionally, long-term safety and dose variations across different ethnic populations need verification.

This study confirms that ARA+DHA supplementation during lactation and weaning increases gut DHA deposition, upregulates mucosal sIgA, and anti-inflammatory cytokines (IL-2, TGF-β), creating an intestinal immune microenvironment conducive to OT development. These findings provide new mechanisms for preventing food allergies through early nutritional interventions, emphasizing the importance of nutrition during the first 1000 days of life in programming the immune system. With deeper understanding of "nutrition-immunity-gut" interactions, precision nutrition strategies targeting LCPUFAs could become a core approach in allergy prevention and control.

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Reference:

Feeding Docosahexaenoic Acid and Arachidonic Acid during Suckling and Weaning Contributes to Oral Tolerance Development by Beneficially Modulating the Intestinal Cytokine and Immunoglobulin Levels in an Allergy-Prone Brown Norway Rat Model https://doi.org/10.1016/j.tjnut.2024.10.021