Dr. Jeffrey Escobar - Does Immune Activation Alter Growth Potential and Nutrient Digestibility?

Alterations in amino acid digestion and metabolism

during diseaseJeffery Escobar, Ph.D.

Executive ManagerNutritional Physiology Research

Agenda

• Immune activation and metabolism alteration

• Plasma amino acids

– Changes during LPS challenge and their balance

• Amino acid digestibility

– Apparent, basal ENL, standardized

– Salmonella challenge

– Fate of undigested CP/AA

• Conclusions

LPS induce inflammatory response

• Inflammatory cytokines

– TNF-α, IL-1b, IL-6

• Induce fever

• Reduce feed intake

• Stimulate – Skeletal muscle proteolysis

– Liver protein synthesis

• Produce acute phase proteins

Kawai and Akira, 2010

Protein synthesis and accretion in immune challenged pigs

Orellana et al., 2002; Escobar et al., 2004

* P < 0.05

Immune activation and metabolism

mg/kg BWAA Nitrogen

Phe 0 0Trp 2 <1Val 10 5Ser 11 1Cys 12 1Tyr 16 1Pro 25 3Met 26 2Thr 28 4Gly 39 7Ile 41 4Asp+Asn 61 12Ala 66 10His 68 11Leu 72 8Arg 83 28Lys 104 20Glu+Gln 140 15

Excess release of AA and nitrogen from skeletal muscle protein over incorporation into mixed acute-phase proteins

– Increase in plasma urea nitrogen

Reeds et al., 1994

• Compare AA composition of skeletal muscle protein and a mixture of acute-phase proteins

• Assuming a typical increase in the synthesis of a mixture of acute-phase protein of 850 mg/kg BW

• The amount of skeletal muscle protein needed to be mobilized = 1,980 mg

• A difference of 1,150 mg/kg BW

• For a 100 kg pig:

– About 200 g of muscle protein mobilized

– About 13 g of excess nitrogen excreted

LPS, cytokines, and plasma urea NP

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g/m

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Pla

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g/d

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Hour after-injectionHour after-injection

Webel et al., 1997

12-h feed-deprived pigs

Immune activation in fasted vs fed state

• Most acute immune activation studies with LPS have been conducted in the fasted state

– Normal food deprivation period: 8 - 24 h

• Although sick, pigs normally continue to eat, digest, and absorb nutrients

• Does acute immune activation in the fasted and fed state result in similar changes

– Digestion, absorption, metabolism, excretion

Arrival

24 h

LPS

-10 0

Surgery

Fasting Blood sampling

Feed

Recovery-2

Blood urea nitrogen in fed pigs

Price, 2011

Changes in plasma Phe with LPS

Price, 2011

Changes in plasma Phe and urea nitrogen with LPS in fed pigs

Price, 2011

Changes in plasma amino acids with LPS

Price, 2011

Price, 2011

Area underthe curve

Price, 2011

Area under the curve

Estimating plasma amino acid balance

• Calculate the area under the curve for each AA

– Saline and LPS treated pigs

• Use correction factor of 0.075 L plasma/kg BW to convert AUC to µmolekg BW-1

h-1 as a unit of AA metabolism (Talbot and Swenson, 1970)

• Apply the following equation:

AA balance = (AUCLPS- AUCSAL) 0.075 L 1kg BW h2××

Price, 2011

Negative AA balance: SAL > LPS

Price, 2011

Positive AA balance: LPS > SAL

Price, 2011

Summary of AA metabolism

Is lysine the most limiting AA during immune activation?

• Acute immune activation via LPS

– Induce similar changes in plasma urea nitrogen during the fasted and postprandial state.

– Reduces the digestion and/or absorption of dietary nutrients or increases AA catabolism

– Alters plasma balance of amino acids

• Lysine was the most negative

• Alanine was the most positive

Phe to Tyr conversion

Harper’s Biochemistry; Kilani et al., 1995; Hsu et al., 2006

Changes in plasma Tyr with LPS

Price, 2011

Changes in plasma Phe to Tyr

Price, 2011

Amino acids:nutritional considerations

Rethink AA limitation order for metabolic purposes

• Dietary amino acids:

– Essential, conditional, nonessential

• “One which cannot be synthesized by the animal organism out of materials ordinarily available to the cells at a speed commensurate with the demands for normal growth” W.C. Rose, 1946

• Essentiality of amino acid for protein synthesis

– Which one is the most limiting AA?, second?

van de Waterbeemd and Gifford, 2003

Digestibility vs. bioavailability

EndogenousFactors

Species

Breed/strain

Gender

Physiological state

ExogenousFactors

Dietary AA balance

Anti-nutritional factors

Intake

Pathogens, disease?

Factors affecting AA digestibility

Salmonella challenge in pigs

• Euthanized pigs (n=12) 24 and 72 h after oral inoculation to collect ileal digesta samples

• Salmonella Typhimurium DT104

– Nalidixic acid: 25 mg/mL

– Novobiocin: 20 mg/mL

• Intra-nasal inoculation 9.8×109 cfu

Lee, 2012

Apparent ileal AA digestibility, %24 h after Salmonella inoculation

Lee, 2012

Basal endogenous AA losses24 h after Salmonella inoculation

Lee, 2012

Standardized ileal AA digestibility, %24 h after Salmonella inoculation

Lee, 2012

Salmonella challenge in ileal cannulated pigs

• 8 growing pigs

• Ileal cannulation and recovery

• Intra-nasal inoculation 1.3×109 cfuSalmonella Typhimurium DT104 (NalRNovR)

• Collect ileal digesta every 8 h from0 to 72 h after inoculation

Lee, 2012

Dynamic changes in AA digestibility in Salmonella challenged pigs

Lee, 2012

Dynamic changes in AA digestibility in Salmonella challenged pigs

Lee, 2012

Summary on digestibility and disease

• Can poor digestion leads to enteric disease?

• Enteric disease results in:

– Lower apparent AA digestibility

– Higher endogenous losses of AA

• Higher gut demand for nutrients and energy

– Production of mucins, digestibe enzymes, enterocytes, etc.

• Provide highly digestible dietary proteins

Anti-nutritional effect of indigestible protein in the hindgut

Hindgut AA fermentation

Meijers and Evenepoel 2011; Wikipedia

Protein fermentation metabolites limiting pig growth

Yokoyama et al., 1982

Dietary protease improvesintestinal environment

Dietary protease reduces protein fermentation and prevents increase in hindgut pH.

Wang et al., 2011

Dietary protease improvesintestinal environment

Protease limits growth of undesirable bacteria and enhances growth of beneficial bacterial species.

Wang et al., 2011

General conclusions

• Immune activation alters digestion, absorption, and metabolism of amino acids

• Fermentation of undigested proteins produce undesirable bacterial metabolites and alter gut environment and microbiota

• Feed pigs highly digestible ingredients or use available technologies to maximize AA digestibility and hence minimize protein fermentation

Acknowledgements

• Students and technicians

– Dr. Kathryn Price

– Dr. Hanbae Lee

– Dr. Matthew Utt

– Elizabeth Ramirez

– Heather Totty

– Greg van Eyk

– Courtney Klotz

– Heather Reeves

– Pat Williams

• Collaborators

– Dr. Rodney Johnson

– Dr. David Baker

– Dr. William van Alstine

– Dr. Teresa Davis

– Dr. Monica Ponder

– Dr. Mark Hanigan

– Dr. Allen Harper

– Dr. Kevin Pelzer

• Virginia Tech Pratt Fellowship

Thank You!

Obrigado Merci

Gracias

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谢谢

Asante sana

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감사합니다ありがとう

Tak! Danke Schoen!