# Incorporation of dairy lipids and Lactobacillus fermentum in infant formulas: long-term effects on intestinal permeability, mucosal immunity and metabolism in a minipig model

Marion Lemaire, Gaëlle Boudry, Stéphanie Ferret-Bernard, Isabelle Nogret, Laurence Le Normand, Gwenaëlle Randuineau, Michele Formal, Sylvie Guerin, Armelle Cahu, Véronique Rome, et al.

► To cite this version:

Marion Lemaire, Gaëlle Boudry, Stéphanie Ferret-Bernard, Isabelle Nogret, Laurence Le Normand, et al. Incorporation of dairy lipids and Lactobacillus fermentum in infant formulas: long-term effects on intestinal permeability, mucosal immunity and metabolism in a minipig model. 4. International Conference on Nutrition & Growth Conference, Mar 2017, Amsterdam, Netherlands. **hal-01530771**

HAL Id: hal-01530771

https://hal.science/hal-01530771v1

Submitted on 5 Jun 2020

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# Incorporation of dairy lipids and *Lactobacillus fermentum* in infant formulas: long-term effects on intestinal permeability, mucosal immunity and metabolism in a minipig model

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M. Lemaire<fnref n="1" />⁻², G. Boudry<fnref n="1" />, S. Ferret-Bernard<fnref n="1" />, I. Nogret<fnref n="1" />, L. Le Normand<fnref n="1" />, G. Randuineau<fnref n="1" />, M. Formal<fnref n="1" />, S. Guérin<fnref n="1" />, A. Cahu<fnref n="1" />, V. Romé<fnref n="1" />, P. Le Ruyet², I. Cuinet², C. Baudry², S. Blat<fnref n="1" />, I. Le Huërou-Luron<fnref n="1" />

<figure><img src="image_3.png" /></figure>

¹ INRA, INSERM, Univ Rennes 1, Univ Bretagne Loire, Nutrition Metabolisms and Cancer (NuMeCan), Rennes, France, ² Lactalis R&D, 35240 Retiers, France

marion.lemaire@inra.fr

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## Context and objective

Incorporation of dairy lipids (DL) in infant formulas (IF) has been associated with short-term benefits on digestion, gut physiology and microbiota in a piglet model. Clinical studies showed that the probiotic *Lactobacillus fermentum* CECT 5716 (Lf) might prevent infections in infants. The objective of this study was to investigate the long-term effects of addition of both DL and Lf in IF on growth, intestinal barrier function and metabolic and immunologic status of adult Yucatan minipigs. We hypothesized that gut microbiota could be a key factor of this programming.

## Methods

<figure><img src="image_5.png" /></figure>

### 26

piglets received from postnatal day (PND) 2 to 28 a formula containing as lipids:

- only plant lipids (PL)

- a half-half mixture of PL and DL (DL)

- a half-half mixture of PL and DL supplemented with Lf (DL+Lf).

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Pigs were subsequently fed:

- a standard diet for 1 month

- then challenged with a hyperenergetic diet (HE) for 3 months

- euthanized at PND140.

<figure><img src="image_10.png" /></figure>

<figure><img src="image_11.png" /></figure>

### Analyses at PND140:

- Intestinal permeability (Ussing chambers)

- Mucosal immunity (cytokine secretion of ileal explants challenged with LPS)

- Entero-insular axis (density of GLP-1 secreting cells, meal test, pancreas anatomy)

- Metabolism (lipid profile, intravenous glucose tolerance test (IVGTT))

- Gut microbiota metabolism (faecal metabolome)

- Gut microbiota composition (16S RNA sequencing)

<figure><img src="image_12.png" /></figure>

### Statistics:

- ANOVA testing diet, gender and replication factors followed by post-hoc tests

- * : p < 0.05 and # : p < 0.1

- Data are presented as boxplot or mean ± SEM

## Results

### 1. Intestinal permeability

<figure><img src="image_13.png" /><figcaption>Paracellular permeability
Ileum, 90 min</figcaption></figure>

<figure><img src="image_14.png" /><figcaption>LPS passage
Jejunum, 120 min</figcaption></figure>

<figure><img src="image_15.png" /></figure>

The IF composition modulated gut barrier function. Ileal trans- and paracellular permeabilities were increased in DL ± Lf compared to PL, though jejunal LPS passage was decreased in DL+Lf compared to PL.

### 2. In vitro secretion of LPS-stimulated ileal explants

<figure><img src="image_16.png" /></figure>

Pro-inflammatory cytokine (TNFα and IL-8) secretions of LPS-stimulated ileal explants were decreased in DL ± Lf LPS-responders compared to PL.

### 3. Entero-insular axis

<figure><img src="image_17.png" /><figcaption>Caecum GLP-1</figcaption></figure>

<figure><img src="image_18.png" /><figcaption>Plasma GLP-1
Secretory response
to meal stimulation</figcaption></figure>

<figure><img src="image_19.png" /></figure>

Basal and meal-stimulated GLP-1 secretory capacities were increased in DL+Lf. GLP-1 secreting cells density and pancreas anatomy (data not shown) were similar between groups.

### 4. Host metabolism

<figure><img src="image_20.png" /><figcaption>Plasma lipid profile</figcaption></figure>

<figure><img src="image_21.png" /><figcaption>HOMA-IR</figcaption></figure>

<figure><img src="image_22.png" /></figure>

FFA: free fatty acids, HOMA-IR: homeostasis model assessment of insulin resistance

<figure><img src="image_23.png" /><figcaption>Insulinemia</figcaption></figure>

<figure><img src="image_24.png" /></figure>

Growth, food intake and metabolic dysfunctions (dyslipidemia and insulin resistance) induced by the HE diet were similar between groups.

### 5. Gut microbiota metabolism and composition<fnref n="1" />

<figure><img src="image_25.png" /><figcaption>OTU's discrimination</figcaption></figure>

<figure><img src="image_26.png" /><figcaption>Main families</figcaption></figure>

<figure><img src="image_27.png" /></figure>

Fu: Fusobacteria, A: Actinobacteria, P: Proteobacteria, B: Bacteroidetes, Fi: Firmicutes

Gut microbiota metabolism and composition at the operational taxonomic unit (OTU) level were different between groups. It might explain the long-term effects observed on gut physiology.

## Conclusion

This study highlights a long-term programming effect of early consumption of DL+Lf on intestinal epithelial barrier and endocrine function and of DL (± Lf) on immune orientation. These long-term effects might be mediated by changes in gut microbiota (composition and metabolism), which are different with DL alone or DL+Lf early consumption. In-depth analysis of changes in microbiota composition along with analysis of the potential mechanisms leading to the observed long-term effects are ongoing.

This work was founded by Lactalis Group. The authors acknowledge all the members of the porcine experimental facilities of the INRA (Saint-Gilles) and all the technical staff of the NGB team for their expert assistance and help. The authors wish to thank the Nutrition, Chemical Food Safety and Consumer Behaviour research division of the INRA for financing the metabolomics study and the analytical platform for metabolomics and toxicology (MetaToul-AXIOM, INRA, UMR1331 Toxalim, Toulouse, France) for their help in interpreting these results.

<footnote n="1">¹ Collaboration with P. Gérard and M. Rhimi (INRA, Micalis, Jouy-en-Josas)</footnote>