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Crucial role of milk proteins on adhesion of intestinal microbes

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HAL Id: hal-02504336

https://hal.univ-lorraine.fr/hal-02504336

Submitted on 10 Mar 2020

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Crucial role of milk proteins on adhesion of intestinal microbes

Sarah Barrau, C. Gaiani, Claire Retourney, C. Mangavel, Yves Le Roux, Claire Soligot-Hognon, Matthieu Delannoy, Joël Scher, J. Burgain

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Results and Discussion

Crucial role of milk proteins on adhesion of intestinal microbes

S. Barrau

1

, C. Gaiani

1

, C. Retourney

1

, C. Mangavel

1

, Y. Le Roux

2

, C. Soligot-Hognon

2

, M. Delannoy

2

, J. Scher

1

, J. Burgain

1

Introduction

Aim of the study

jennifer.burgain@univ-lorraine.fr ; sarah.barrau@univ-lorraine.fr

Materials and Methods

1

Université de Lorraine, LIBio, F-54000 Nancy, France

2

Université de Lorraine, Inra, URAFPA, F-54000 Nancy, France

Use of differentiated cell

monolayer obtained after

22 culture days

❖ Identification SpaCBA pili role in interaction with intestinal cells

and GPs

❖ Establishment of interactions and benefits of GPs on bacteria

adhesion

❖ Improved understanding of the role of milk in the adhesion of

bacteria to epithelial intestinal cells

Materials used

Intestinal epithelial cells

Caco-2 TC7 cells

HT29-CL16E cells

(producing mucus)

LGG WT LGG spaCBA

LGG WT

LGG spaCBA

OPN

Lf

Milk GPs

Osteopontin

Lactoferrin

Probiotic bacteria

L. rhamnosus GG (LGG) WT

LGG pili depleted (spaCBA)

Caco-2 TC7 cells

HT29-CL16E cells

Acknowledgments

[1] Bron et al. Emerging molecular insights into the interaction between probiotics and the host intestinal mucosa. Nat. Rev. Microbiol., 10 (2012), 66-78.

[2] Lebeer et al. Host interactions of probiotic bacterial surface molecules: comparison with commensals and pathogens. Nat. Rev. Microbiol., 8 (2010), 171-184.

[3] Burgain et al. Significance of bacterial surface molecules interactions with milk proteins to enhance microencapsulation of Lactobacillus rhamnosus GG. Food Hydrocoll. 41 (2014), 60–70.

[4] Christensen and Sørensen, Structure, Function and Nutritional Potential of Milk Osteopontin, International

Dairy Journal, 57 (2016), 1–6

[5] Guerin et al. Adhesive interactions between milk fat globule membrane and Lactobacillus rhamnosus GG inhibit bacterial attachment to Caco-2 TC7 intestinal cell, Colloids and Surfaces B: Biointerfaces, 167 (2018), 44-53.

[6] Tarazanova et al. Cell Surface Properties of Lactococcus lactis Reveal Milk Protein Binding Specifically Evolved in Dairy Isolates. Front. Microbiol. 8 (2017), 1691.

[7] Guerin et al. Pili of Lactobacillus rhamnosus GG mediate interaction with β-lactoglobulin. Food Hydrocoll. 58 (2016), 35–41.

[8] Liu et al. Protective effects of Lactoferrin on injured intestinal epithelial cells, Journal of Pediatric Surgery (2019)

Main source of nutrition and

immunological protection for the neonate

Contain glycoproteins (GPs)

Use of probiotic bacteria

Beneficial for health and

intestinal diseases

Conclusion / Perspectives

mucus

(1) Without GPs: bacteria

adhesion with epithelial cells

Receptor

to pilus

SpaCBA pilus

Cell adhesion site

(2) Bacterial

adhesion with GPs

❖ Impact of mucus on bacterial adhesion (compared to Caco-2 cells)

❖ SpaCBA and WT: almost similar adhesion with GPs

OPN: best adhesion results of both cell types in all present cases

Bacteria

(3) With GPs: increase

of bacteria adhesion to

epithelial cells

GPs

Bacteria

Bacteria

Num

be

r

of

ad

he

ren

t

bac

terial

cells

Milk

Potentially bind bacteria' pili to adhesion

site of model intestinal cells

Glycoproteins

Cell plate with Caco-2 TC7 or HT29-CL16E

Bacterial culture on fresh MRS broth: end of

exponential phase (12h)

Epithelial monolayer incubated for 2h

with a) Bacteria suspension or b)

Bacteria + GPs

Sequential washing of intestinal cells

Counting of bacteria attached

to epithelial cells

“Generally Recognized as Safe”

(GRAS)

What are the different existing interactions ?

Mucus layer: potentially

impact the bacterial adhesion

LGG WT LGG spaCBA

Num

be

r

of

ad

he

ren

t

bac

terial

cells

(Norma

lised

)

Key role of SpaCBA

pilus in adhesion of

LGG to Caco-2 cells

LGG WT

❖ GPs improve LGG adhesion

❖ OPN: major impact on bacterial

adhesion

Lf

Lf

OPN

OPN

Lf + 23.4 %*

OPN + 51.4 %*

Lf + 14.7 %*

OPN + 70.6 %*

Addition

of GPs

* Percentage increase in cell adhesion

compared to results obtained without GPs

Ø

Ø

LGG WT LGG spaCBA

Identification of all interactions

Future works: Identification of molecules engaged in

adhesion like Peptins (adhesins, GPs, surface molecules…)

Caco-2

HT29

❖Arla Foods and Glanbia for providing milk GPs

❖ LUE Impact Biomolécules for funding COMPET project.

References

Références

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