# From muscle to meat: physiological and metabolic basis

Michel Jacques M.J. Duclos, Cécile Berri

► To cite this version:

Michel Jacques M.J. Duclos, Cécile Berri. From muscle to meat: physiological and metabolic basis. 1. Advencia academy: "Nutritionist: a challenging job! Feeding an ecosystem to transform feedstuffs into animal proteins", European Association of Animal Production (EAAP). ESP., May 2017, Barcelone, Spain. 37 diapos. hal-01604313

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Submitted on 5 Jun 2020

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INRA
SCIENCE & IMPACT

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# From muscle to meat: physiological and metabolic basis

M. J. DUCLOS & C. BERRI

Unité de Recherches Avicoles (Avian Research Unit)

URA, INRA, Nouzilly, France

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INRA
SCIENCE & IMPACT
Increasing demand in poultry meat worldwide (+2%/year)

Rapid evolution of the market share

<figure><img src="image_12.png" /><figcaption>Market share, NCC (2012)</figcaption></figure>

## Many quality expectations:

* Meat yields

* Functional properties

* Storage ability

* Sensory perception

* Nutritional value

* Image (animal welfare)

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Consequences for broiler production

Strain

1957

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34 g

1978

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42 g

2005

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44 g

0 d

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316 g

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632 g

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1,396 g

28 d

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56 d

905 g

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1,808 g

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4,202 g

Source: Zuidhof et al., 2014

○ Feed efficiency ↑

○ Growth rate ↑

○ Breast meat yield ↑

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Consequences for broiler production

<table><thead><tr><th>Year</th><th>Strain</th><th>Live weight, g</th><th>Age, j</th><th>Breast, g</th><th>Breast, % LW</th></tr></thead><tbody><tr><td>1957</td><td>Athens</td><td>2078</td><td>57</td><td>280</td><td>13,5</td></tr><tr><td>2001</td><td>Ross 308</td><td>2207</td><td>43</td><td>349</td><td>15,8</td></tr><tr><td>2007</td><td>Ross 308</td><td>2200</td><td>36</td><td>410</td><td>18,6</td></tr><tr><td>2012</td><td>Ross 308</td><td>2200</td><td>35</td><td>464</td><td>21,1</td></tr></tbody></table>

(Petracci et al., 2015)

Strain

1957

0 d

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1977

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2005

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28 d

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56 d

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Source: Zuidhof et al., 2014

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# Consequences for broiler production

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* Decreasing age at slaughter (< 2.2kg)

* Production of heavy and high-yielded broilers (> 3kg)

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# Muscle hypertrophy : fiber size increase

Muscle fiber surface, µm²

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# Consequences of older broiler production

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## Production of heavy and high-yielded broilers (> 3kg)

- Increased muscle fiber size (Baéza et al., 2012)

- Increased lipid and protein content (Baéza et al., 2012; Janisch et al., 2013)

- Modified post-mortem metabolism (Baéza et al., 2012)

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The post-mortem pH fall: a key element for meat quality

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# Postmortem metabolism and meat quality

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# Muscle growth and meat quality

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# Impact of increasing muscle growth on meat pH

(Berri et al., 2001)

Selection

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pHu

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Genotype

BW: 1.3kg → 3.0 kg
BMY: 11.5% → 18.5%

(Baéza et al., 2012)

Age at
slaughter

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pHu

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Age at slaughter

BW: 1.9 kg → 4.2 kg
BMY: 18.7% → 22.2%

(Berri et al., 2008)

Nutrition

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pHu

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Digestible Lysine

BW: 2.6 kg → 2.7 kg
BMY: 22.0% → 22.9%

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# Consequences for broiler production

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## Production of heavy and high-yielded broilers (> 3kg)

- Increased muscle fiber size (Baéza et al., 2012)

- Increased lipid and protein content (Baéza et al., 2012; Janisch et al., 2013)

- Modified metabolism (Baéza et al., 2012)
# Metabolic Changes

(Berri et al., 2001)

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128 µM

Glycogen

87µM

2.4 ppm

Iron content

1.8 ppm

5.8 mM/g

HAD activity

4.7 mM/g

- Lower carbohydrate reserve

- Lower blood supply

- Lower oxidative activity

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# Muscle blood supply and Metabolism

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Lower blood supply

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(Alnahhas et al., 2015)

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Lower glycogen in muscle

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20% difference in breast
muscle glycogen
(P. major)

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(Alnahhas et al., 2014,2015)

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# Metabolomic profile (muscle & serum)

(Beaucercq et al., J. Proteome Res. 2016, 15, 1168–1178)

♂ **pHu-**/*pHu+* (n = 40) [**6.3** vs **5.5** – glycogen **195** vs **145**]

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## Modeling by oPLS-DA analysis

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a.

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b.

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c.

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pHu+ pHu-

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Polar extract of P. major muscle (1H et 31P NMR): 25 discriminant metabolites

Polar extract of serum: (1H NMR): 20 discriminant metabolites

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# Metabolomic profile (muscle& serum)

## Enrichment analysis on discriminant metabolites

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**pHu -** : Overrepresentation of metabolites related to **carbohydrate turn-over**
**pHu +** : Overrepresentation of metabolites related to **oxidation and protein catabolism**

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# Metabolomic profile (muscle & serum)

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(Beaucercq et al., 2016)

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# Transcriptomic profile (muscle)

♂ **pHu-**/*pHu+* (n = 31)

Agilent microarray (monocolor 8 × 60K)

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Heat map classification

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# Functional Analysis of muscle transcriptome

## Ingenuity Pathway Analysis

pHu-

Glycolysis/neoglucogenesis

ATP Concentration ☐

Amino-Acid Métabolism

Muscle cell organisation

Purine metabolism

Triglyceride Synthesis

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High energetic status
High glycolytic activity

Beaucercq et al, Submitted 2017

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# Histological profile

pHu+

pHu-

Glycogen

Adult fast-myosin

Embryonic and
neonatal fast-myosin

Neonatal fast-myosin

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100 µm

pHu+: Higher ratio of cells expressing embryonic and neonatal developmental fast-myosins, which could be the sign of higher regeneration process

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1H, 31P NMR

Chicken

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Muscle, Serum

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Multivariate
Analysis

Glycogen ↑

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↑Carbohydrates

↑Energy

Glycogen ↓

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↑ROS

↑Oxidation

↑Catabolism

↓Energy

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pHu+

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# Summary : consequences of increased pHu

* Low blood supply → low cell oxygenation, low nutrient supply

* Intensive use of in situ glycogen

* Elevated protein and muscle degradation

* Increased oxidative stress and production of reactive oxidative species (ROS) → altered redox homeostasis

* Muscle dysfunction? Fiber alterations? Link with emergent defects?

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# What is the link between metabolic and structural meat-defects ?

<figure><img src="image_98.png" /><figcaption>White striping</figcaption></figure>

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<figure><img src="image_102.png" /><figcaption>Wooden breast</figcaption></figure>

<figure><img src="image_103.png" /><figcaption>Spongy muscle</figcaption></figure>

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White striping

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(from: Kuttappan et al., 2012)

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# White striping

Transversal cross-section

<figure><img src="image_107.png" /><figcaption>Longitudinal cross-sections</figcaption></figure>

* = muscle fiber necrosis

Fibrophilic and macrophage infiltration

<figure><img src="image_108.png" /><figcaption>NORMAL</figcaption></figure>

Muscle fiber degeneration

<figure><img src="image_109.png" /><figcaption>WHITE STRIPING</figcaption></figure>

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Fibrosis: accumulation of connective tissue

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Lipidose: accumulation of lipids

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# White striping

(unpublished results)

° Global gene expression analysis (microarrays) → functional analysis

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<figure><img src="image_116.png" /><figcaption>Necrosis and Regeneration</figcaption></figure>

<table><thead><tr><th></th><th>Fonctions identifiées par IPA®</th><th>Differential genes</th></tr></thead><tbody><tr><td rowspan="2">Pathways</td><td>Wnt/Ca+: regulation of cellular Ca++, involved in embryonic tissue development</td><td>43</td></tr><tr><td>HGF (hepatocyte Growth Factor): activation of satellite cells during muscle regeneration</td><td>85</td></tr><tr><td>Disorders</td><td>Pathology of connective tissue: hyperplasia of connective tissue and lipodystrophy</td><td>28</td></tr></tbody></table>

<figure><img src="image_117.png" /><figcaption>Inclusion of lipids</figcaption></figure>

<figure><img src="image_118.png" /><figcaption>Extension of connective tissue</figcaption></figure>
# Wooden Breast

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Decreased blood supply?

Decreased glycogen store?

<table><thead><tr><th></th><th>Shear force</th><th>Ultimate pH</th><th>Lightness</th><th>Redness</th></tr></thead><tbody><tr><td>Normal</td><td>8.39</td><td>5.88</td><td>48.74</td><td>0.19</td></tr><tr><td>Wooden breast</td><td><strong>16.24</strong></td><td><strong>6.09</strong></td><td>50.18</td><td><strong>-1.46</strong></td></tr></tbody></table>
# Wooden breast

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Wooden breast

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○ Muscle fibers variable in size

° Regenerative fibers

o Splitting

(Adult Myosin Heavy Chain labelling)

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# Wooden breast

* Change in muscle typing

* Muscle growth factor

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(new fibers)

ad

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(very fast myosin)

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↑ Marker of early stages of development

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WBO

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WI 2

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↑ proliferation and differentiation of muscle fibers

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# Wooden breast

* Smooth muscle formation

* Angiogenesis

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Myocardin

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* Capillaries disconnection: poorer oxygenation, and lower nutrient intake of fiber
# Wooden breast

(Mutryn et al., 2015)

* Global gene expression analysis by RNA-sequencing:

* Abnormal accumulation of intracellular calcium

* Hypoxia

* Increased oxidative stress and accumulation of ROS

* Fiber-type switching (fast to slow)

* Increased cellular repair and regeneration in skeletal muscle

* Downregulation of genes related to coagulation system

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# Wooden breast

* Metabolite profiling (GC/MS; LC-MS/M/z (Abasht et al., 2016))

* Much lower glycogen content (1.7-fold)

* Increased oxidative stress,

* Elevated protein levels,

* Muscle degradation,

* Altered glucose utilization.

* Altered redox homeostasis (↑ hypoxanthine, xanthine)

<figure><img src="image_137.png" /></figure>

Excessive ECM Remodeling; Response to ROS

<figure><img src="image_138.png" /></figure>

WB

Common
regulations
between WB
and pHu+
muscles

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pHu+

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# Major meat quality defects

<figure><img src="image_141.png" /><figcaption>## Muscle 'dystrophic' syndromes</figcaption></figure>

<figure><img src="image_142.png" /><figcaption>## Oxidative damages</figcaption></figure>

<figure><img src="image_143.png" /></figure>

Genetics

Muscle growth

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Carbohydrate (glycogen)
energy storage

Nutrition

Rearing

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Conclusions

° Genomics and metabolomics are very helpful to unravel meat quality determinism and identify pertinent biomarkers for diagnosis purpose

° It highlights some **common regulatory pathways** underlying meat defects related to muscle growth and glycogen depletion...

° Poor muscle fiber vascularity → poor oxygenation of the muscle, anaerobic glycolysis, oxidative stress

° Production of ROS (↓ redox homeostasis)

° Increased protein and amino acid catabolism

° Change in muscle fiber typing

° Increased regeneration process

° But also **specific processes** related to wooden breast and white striping

° Muscle fiber necrosis

° Cell repair processes (fibrosis, Lipidosis)

<figure><img src="image_146.png" /></figure>

Pseudo-hypertrophy of muscle (dystrophic condition)
* Developing non invasive methods to identify breeders prone to develop meat defects:

* Genetic markers useful for selection (genomics)

* Biomarkers such as blood metabolite, enzyme activity, or gene expression

* Ensuring an optimal and harmonious development of broilers with positive impact on animal robustness and meat quality:

* Livestock management, especially during early life

* Preserve glycogen store in muscle,

* Delay the differentiation processes to produce more fibers of smaller size

* Increase the pool of available satellite cells to insure proper cell growth and renewal (limit fibrosis and lipidosis)

* Considering the use of more robust strains with more harmonious body development:

* Gains expected in term of mortality, meat waste at the slaughter plant, increased processing yield
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Thanks for your attention

Special thanks to

- S. BEAUCLERCQ, PhD student

- E. LE BIHAN-DUVAL, senior scientist (genetics)

- Many others at

- URA - INRA Nouzilly

- PEAT - INRA Nouzilly

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