Research on donkey milk · Study 21

Donkey milk, Paneth cells & lysozyme

In a mouse model of experimental ileitis, Yvon et al. examined how donkey milk was related to inflammatory findings, Paneth cell-associated antimicrobial defence substances and selected gut bacteria.

Important: This is preclinical animal research. The study is not a clinical study in humans and demonstrates neither the treatment nor the prevention of a human intestinal disease. Nor was it proven that lysozyme alone was responsible for the observed effects.
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DON DONKEY’S® donkey milk
Study 21
Publication Yvon et al.
2018
Model Experimental
ileitis in mice
Cell type Paneth cells
of the small intestine
Focus Lysozyme · defensins
inflammation · microbiota

The research question

What happens to the body’s own intestinal defence during ileitis?

Paneth cells are among the specialised cells of the small intestinal mucosa. They are located deep in the intestinal crypts and produce, among other things, antimicrobial defence substances such as defensins and lysozyme.

The researchers examined whether administration of donkey milk to mice with experimentally induced ileitis was associated with changes in inflammatory findings, Paneth cell markers and selected microbial groups.

Important for understanding The study did not exclusively concern the lysozyme contained in donkey milk. A central part of the work dealt with the body’s own antimicrobial defence of the Paneth cells.

Study design

From the ileitis model to the analysis of intestinal defence

Step 01

Mouse model

The study was carried out as a preclinical animal experiment.

Step 02

Ileitis

Inflammation of the small intestine was induced experimentally with indomethacin.

Step 03

Milk comparison

The researchers compared, among other things, untreated and heated donkey milk.

Step 04

Gut analysis

Tissue findings, inflammatory markers, Paneth cell markers, lysozyme and selected bacterial groups were analysed.

Below the surface

Deep in the intestinal crypt sit the Paneth cells.

Paneth cells are located in the lower part of the crypts of the small intestine. There they form components of the body’s own antimicrobial defence.

These include defensins and lysozyme. These molecules are part of the complex interaction between the intestinal epithelium, immune defence and microbial environment.

In the experimental ileitis model, the corresponding defence parameters were altered. The researchers examined how these parameters looked with donkey milk.

Intestinal crypt · schematic representation Animation
Intestinal lumen
LYS
DEF
LYS
LYS = lysozyme · DEF = defensins. Schematic biological illustration – not a microscope image from the study and not a representation of a directly observed molecular mechanism.

An interesting comparison

Untreated and heated donkey milk did not behave the same in the model.

The different findings make the processing of the milk scientifically interesting. However, they do not prove which single component is responsible.

Untreated donkey milk

More favourable findings in the model

For several parameters altered by the experimental ileitis, more favourable or normalised findings were described with untreated donkey milk.

Heated donkey milk

Not the same finding

The heated donkey milk did not show a corresponding effect. This makes heat-sensitive components scientifically interesting but does not identify a single responsible substance.

Important for interpretation

Two sources of lysozyme must not be confused.

Donkey milk itself contains lysozyme. At the same time, Paneth cells in the small intestine produce the body’s own lysozyme.

Changes in Paneth cell defence therefore do not mean that ingested milk lysozyme simply replaces the body’s own lysozyme.

LYS
Milk lysozyme A component of donkey milk and a possible candidate for further research.
≠
LYS
Paneth cell lysozyme The body’s own component of antimicrobial intestinal defence.

Key observations

What was observed in the experimental ileitis model

01 · Intestinal tissue

Changes in the small intestine

In the model, indomethacin caused, among other things, changes in intestinal length and greater macroscopic and microscopic tissue damage.

02 · Inflammation

Lipocalin-2

In the ileitis model, the inflammation-associated marker lipocalin-2 rose. According to the authors, untreated donkey milk was associated with more favourable findings for several of these changes.

03 · Paneth cells

Defensins & lysozyme

The ileitis was accompanied by reduced α-defensin expression and changes in lysozyme in the Paneth cells.

04 · Enzymatic level

Lysozyme activity

The measured lysozyme activity in the faeces was also reduced in the experimental ileitis model and was among the parameters examined.

05 · Microbiota

Selected bacterial groups

The researchers also documented changes in selected microbial groups in the faeces, thereby also looking at the gut environment.

06 · Processing

Heat as a research approach

The difference between untreated and heated donkey milk makes heat-sensitive milk components interesting. However, this does not provide causal proof for lysozyme or any other single substance.

Mechanistic research question

How could the relationship be investigated further biologically?

Starting point Bioactive components of donkey milk
Possible level Interaction with the gut environment
Possible level Mucosa and immune response
Measurable endpoint Paneth cell and inflammatory parameters
Scientific limit: This representation is a mechanistic working hypothesis. Study 21 does not fully prove this chain of effects and in particular does not show that lysozyme from donkey milk alone causes the observed changes.

Critical assessment

What Study 21 shows – and what it does not

What was observed

✓ In the mouse model examined, untreated donkey milk was associated with more favourable findings for several ileitis-associated changes.
✓ Paneth cell-associated antimicrobial markers such as α-defensins and the body’s own lysozyme were part of the study.
✓ Enzymatic lysozyme activity and selected microbial parameters were also examined.
✓ Untreated and heated donkey milk did not show the same finding in the model.

What was not proven

× No treatment, cure or prevention of human intestinal diseases.
× No proof that milk lysozyme alone causes the observed changes.
× The heat comparison does not identify a single responsible milk component.
× The results do not demonstrate a corresponding effect of the DON DONKEY’S® product.

The next research question

How could DON DONKEY’S® investigate this question further experimentally?

A sensible first step would be product-specific analysis. It would need to examine separately how much lysozyme is present and how much of it is actually enzymatically active.

Stage 01 Analyse fresh milk Determine lysozyme amount and enzymatic activity in defined fresh milk batches.
Stage 02 Compare processing Analytically compare fresh milk and reconstituted powder from comparable starting batches.
Stage 03 Validate biologically With suitable analytical results, the samples could then be compared in a validated preclinical model.
Stage 04 Test lysozyme specifically Targeted removal or inactivation followed by add-back would allow a possible causal contribution of lysozyme to be investigated much more precisely.
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Discover our donkey milk as a food. On our research pages we show which properties of donkey milk are being studied scientifically and where the limits of the evidence to date lie.

Transparency: The DON DONKEY’S® product offered here was not examined in Study 21. Results from the mouse model are not transferred to our product or to humans. Product-specific statements on lysozyme amount or enzymatic activity require corresponding analyses of our own.
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Original publication

Donkey milk consumption exerts anti-inflammatory properties by normalizing antimicrobial peptides levels in Paneth’s cells in a model of ileitis in mice

Yvon S. et al.
European Journal of Nutrition · 57 · 155–166 · 2018

DOI: 10.1007/s00394-016-1304-z

PubMed: PMID 27581119

The publication describes preclinical animal research. No statements about the treatment, cure or prevention of human diseases can be derived from it. Nor can any single component of donkey milk be identified from the results as the sole cause of the observed effects.