1. The Biology of Stress Lance Baumgard, Sara Stoakes and Rob Rhoads
Iowa State University
Department of Animal Science

2. Hypothesis
Leaky gut explains the negative consequences of heat stress and off-feed events (all farm animals), and ketosis (dairy cows) problems

3. Heat Stress: Economics and Food Security
Cost: (lost productivity, mortality, product quality, health care etc.)
American Agriculture: > $4 billion/year
Global Agriculture: > $150 billion/year
It will get worse in the future if:
Climate change continues as predicted
Genetic selection continues to emphasize milk synthesis, lean tissue accretion, piglets/sow etc..
Heat producing processes
St. Pierre et al., 2003; Baumgard and Rhoads, 2013

4. Temperature Humidity Index (THI)
Easy way to measure and evaluate heat stress
Based on cows only under shade….solar radiation is incredibly potent
72 thought to be when cows become susceptible
Based on 60 year old data when cows were producing lb/d
Modern cows are stressed at THI of 68

5. Heat Stress

6. Heat Stress Questions??
Does the decrease in feed intake explain the reduced milk yield during heat stress?
Indirect vs. direct effects of heat
If we have a better understanding of the biological reasons WHY heat stress reduces production, we’ll have a better idea of how to alleviate it.

7. Effects of Heat Stress on Feed Intake
Lactation:
Effects of Heat Stress on Feed Intake
Heat -stressed
Pair-fed
Heat stress  feed intake by ~30 %
Rhoads et al., 2009

8. Effects of Heat Stress on Milk Yield
Heat stress  yield ~45%
Pair-feeding  yield by ~19%
Thus,  feed intake only accounts for ~50% of the reductions in milk yield
Rhoads et al., 2009
Wheelock et al., 2010
Baumgard et al., 2011

9. Body Weight Loss
Pair-fed
Heat Stress
Rhoads et al., 2009

10. Heat Stress Reduces Back-fat Mobilization
Rhoads et al., 2009
Wheelock et al., 2010
Baumgard and Rhoads, 2013

11. Something other than the mammary gland is utilizing ~400 g more/day?
Milk Sugar Output
Heat Stress Cows Secrete ~400 g less lactose/day than Pair-Fed Thermal Neutral Controls
Something other than the mammary gland is utilizing ~400 g more/day?
Rhoads et al., 2009
Wheelock et al., 2010

12. Gastro-Intestinal Tract (GIT) Review Guts

13. Reminder: Intestinal Functions
GIT is a tube running from the mouth to the anus
Everything inside of the tube is technically “outside” of the body
Digest and absorb nutrients
GIT lumen is a inhospitable environment
Prevent parasites, pathogens, enzymes, acids, toxins etc.. From infiltrating “self”
Barrier function

14. Human GIT Surface Area:
Lungs
GIT
~50X
~150 X
Skin
~2 m2
That’s an enormous amount of area to “defend”!
No wonder 70% of the immune system resides in GIT

15. Biology of Heat Stress Symptoms

16. Heat Stress and Gut Health
Diversion of blood flow to skin and extremities
Coordinated vasoconstriction in intestinal tissues
Reduced nutrient and oxygen delivery to enterocytes
Hypoxia increases reactive oxygen species (ROS)
Reduced nutrient uptake increases rumen and intestinal osmolarity in the intestinal lumen
Multiple reasons for increased osmotic stress
Baumgard and Rhoads, 2013

17. Intestinal Morphology
Thermal Neutral
Heat Stress
Pair-fed
Pearce et al., 2011

18. Heat Stress and Gut Health
Lipopolysaccharide (LPS) stimulates the immune system
LPS promotes inflammation production….catabolic condition
TNF, IL-1 etc..
Reduced appetite
Stimulates fever
Causes muscle breakdown
Induces lethargy
....reduces productivity
LPS can cause liver damage

19. Compromised TJs Healthy TJs
Lumen
Hypoxia
Actin
HIF-1α
Myosin
MLCK P
MLCK
TJs
PGE2
TNFα
IL-1β
IL-16
INFᵞ
APP
IkB
TLR4
NFkB
P60
P65
Blood stream
Submucosa

20. The effects are rapid!
Plasma LPS & LBP
Pearce et al., 2015

21. Heat Stress Summary Direct and indirect effects Hyperinsulinemia
↓DMI only accounts for 50% of reduced milk yield
Hyperinsulinemia
Blunted adipose mobilization
Liver remains sensitive to catabolic signals
Leaky gut
Inflammation and acute phase protein response
Unknown whereabouts of 400 g of glucose

22. Leaky Gut and Ketosis?

23. Transition Period Disorders: Mediated Largely by NEFA?
Transition Period
Metabolic shift
NEBAL
Negative effects on future production
Dystocia
Milk fever
Retained placenta
Metritis
Ketosis
Displaced abomasum
Fatty liver
Lameness
Death
Only 50% of cows in North America complete the transition period without experiencing one of these problems
Drackley, 1999

24. Dogma
Excess adipose tissue mobilization causes fatty liver and ketosis
This is worse in high producing cows
Industry Goal: Reduce blood NEFA

25. Correlation Studies Many studies associate NEFA and BHBA with:
Increased risk of ketosis, decreased milk yield, LDA, metritis, retained placenta, laminitis, or poor reproduction
Chapinal et al., 2011; Huzzey et al., 2011; Ospina et al., 2010a, 2010c; Duffield et al., 2009; LeBlanc et al., 2005
Plasma NEFA are markedly increased (>700 mEq/L) following calving in almost all cows
~15-20% get clinical ketosis
What makes these cows more susceptible to ketosis?
Predisposition to developing fatty liver?

26. ↑ Inflammatory response ↑ Acute Phase Proteins: Serum Amyloid A
Gut Lumen
LPS/LBP Complex
TLR4
LPS
Immune Cell
Liver
↑ Inflammatory response
↑ Acute Phase Proteins:
Serum Amyloid A
Haptoglobin
LBP
follow LPS as it is translocated across the intestinal epithelium into the portal circulation where it must be bound to LBP in order to engage the TLR4 receptor of immune cells where it elicits an immune response (which includes production and increased circulation of acute phase proteins). Therefore an animal will increase circulating levels of LBP when there are high amounts of LPS in order to process and clear the bacteria.
LBP
Portal Circulation
Sara Stoakes

27. Objectives
Measure biomarkers of leaky gut in cows that were retrospectively classified as ketotic (only diagnosed problem) and healthy herd mates
n = 8 ketotic cows
n = 8 “healthy” cows
Initial experiment had non-ketotic objectives
Nayeri et al., 2015

28. Increased LBP in Ketotic Cows
Nayeri et al., 2013

29. Objectives
Confirm that the biomarkers of leaky gut increase during the transition period for clinically ketotic cows
A compromised GIT barrier and subsequent endotoxin (LPS) infiltration may play a causative key role in ketosis development

30. Trt: P = 0.02

31. Similar to our previous data

32. Transition Cow Problems are Associated with Biomarkers of Leaky Gut
Intestinal barrier becomes leaky
Endotoxin induced immune activation
Immune system has energetic cost
Reprioritization of nutrients away from milk synthesis……………..$$$ problem

33. Objectives: Determine Feed Restriction’s Impact on Leaky Gut “Out of Feed Events” are common in EVERY animal agriculture industry

34. 12 hours of feed restriction (40% of ad libitum intake) causes leaky gut in the pig
Lipopolysaccharide (LPS)
LPS-BP b a
b,c
a,b
a,c b a a
Pearce et al., 2015

35. The Effects of 7 day Feed Restriction on Inflammation in the cow
BUN-???
We saw linear tendencies for acute phase proteins LBP and SAA which both increased with the severity of feed restriction
We saw a significant cubic effect in the acute phase protein haptoglobin, we saw the highest level of haptoglobin only in the treatment receiving 40% ad lib

36. Effects of Feed Withdrawal (100%) on Milk Yield
22% b
39% c
Milk yield really took a hit in both LPS and LPS-Eu cows. Interestingly, glucose did not recover milk yield like we thought it might. Milk yield decreased at both time points for both treatments by about 85%.
Stoakes et al., 2015

37. Could 12 h of FR causes leaky gut in cattle?
Don’t know….depends upon mechanism
Decreased luminal nutrient delivery….then likely not
Continues out-flow of ruminal nutrients
Psychological stress
Hunger-induced ACTH-Cortisol action…..then possible
Cortisol causes leaky gut in multiple models
Intestinal Mast cells are responsive to nerves
Upon CRF stimulation they release proteases and TNF
Both proteases and TNF cause tight junction breakdown

38. Can “leaky gut” explain suboptimal production frequently observed in animal agriculture?
Heat Stress
Inadequate feed intake
“off-feed event”
The negative effects on growth and milk yield are bioenergetically unexplainable by reduced feed intake
Transition period
Cause of ketosis?
Weaning
Shipping
Overcrowding
Unpalatable feed

39. $$ Billion Dollar Question $$
Can the Feed or Animal Health Industry do anything about leaky gut????
Targets:
Direct action at intestine
Indirect via:
Increased feed intake
Rumen acidosis prevention
Hind gut acidosis prevention
Improved immune function

40. Potential nutritional strategies to ameliorate intestinal permeability
Baumgard et al., 2014

41. Lactating Dairy Cow Metabolic Adaptation to Heat Stress and Maladaptation to Lactation
Summary

42. Successful Transition
Metabolic Flexibility:
Decreased Insulin Sensitivity
Baumgard and Rhoads, 2013

43. Unsuccessful Transition
Fatty Liver, Excessive Ketone Synthesis
LPS
Metabolic Flexibility:
Decreased Insulin Sensitivity
Glucose redirected to immune system
Baumgard and Rhoads, 2013

44. Metabolic Inflexibility Remains Insulin Sensitive
LPS
Metabolic Inflexibility
Remains Insulin Sensitive
Baumgard and Rhoads, 2013

45. Ketosis: When to intervene?
Treat:
High ketones
Not coming into milk
Not aggressively eating
Looks sick
Has a mild fever
Don’t mess with
High ketones….but she’s
Eating like a champ
Milking like a world-record holder
Looks great
No fever

46. Summary
Heat stress and ketotic cows have a similar metabolic and endocrine fingerprint
Leaky gut is a common denominator in both
The activated immune system utilizes an enormous amount of glucose.
Dietary Strategies

47. Conclusions
Leaky gut may play an important role in suboptimal milk yield commonly observed during “stress”
Strategies that can improve intestinal integrity need to be researched…in a “stressed model”
If leaky gut is the fundamental cause of many typical on-farm problems….then it is a financial problem that dwarfs all others combined

48. Acknowledgments Funding Support USDA NRI/AFRI # 2005-35203-16041
Dr. Victoria Sanz-Fernandez
Dr. Sarah Pearce
Dr. Jay Johnson
Amir Nayeri
Nathan Upah
Sam Lei
Erin Laughlin
Erin Nolan
David Valko
Dr. Pat Gorden
Mohannad Abumajieh
Mohammad MalQaisi
Johanna Mayorga Lozano
Jake Siebert
Mr. Howard Green
Dr. Chel Moore
Dr. Mark McGuire
Dr. Arnaldo Burgos
Dr. Howard Green
Dr. Matt Waldron
Dr. Lynn Davis
Dr. Jeff DeFrain
Dr. Mark Wilson
Dr. Mike Socha
Funding Support
USDA NRI/AFRI # # # # #
Zinpro Inc.
Elanco Animal Health
Kemin Inc.