Protein and amino acids for dairy cows

around calving

Johan Osorio, Ph.D.

Assistant Professor
Dairy and Food Science Department
South Dakota State University

Advanced Dairy Solutions

Outline
• Peripartal Period

• Methionine and the Peripartal Period

• Liver Function

• Immune system

• Histone methylation

• Remarks
Peripartal or Transition period
Tissue mobilization

Energy balance

Pre-calving Post-calving
Body condition score
~3-4 wk Calving ~3-4 wk

Energy intake

4% Fat corrected milk

(Ingvartsen, 2006)
(Block et al., 2001)
 Cow in negative
Liver
Fat or Adipose
Tissue
energy balance

NEFA NEFA NEFA

NE, Epi Insulin

TG Mitochondria
CO2

Ketone
Bodies

Milk
Fat TG TG
Mammary VLDL
Gland

VLDL = very-low density lipoproteins (modified from Drackley, 1999)

 Both body fat and protein are mobilized

-61
-46

-21 -21

Increased milk by 3 kg/d

(Komaragiri and Erdman, 1997)
with no change in intake
Metabolizable protein and amino acids
likely limiting around calving
Metabolizable Protein

Bell et al. (2000)

Needed for milk protein, glucose synthesis, synthesis of

other compounds (e.g. SAM, glutathione, taurine)
Net liver uptake of Methionine and Histidine increases
after calving

(Larsen and Kristensen, 2013)

Net uptake of Met by liver can be enhanced by supplemental RP-Met

……..also prevents decrease in blood Met postpartum (Dalbach et al., 2011)
Metabolizable protein
Amino Acid Profile
Outline
• Peripartal Period

• Methionine and the Peripartal Period

• Liver Function

• Immune system

• Histone methylation

• Remarks
Methionine and the Peripartal Period

Dietary Osorio et al., 2013 Zhou et al., 2016 Batistel et al., 2017
component Control Met Control Met Control Met
CP, % of DM 17.4 17.4 17.2 17.3 17.7 17.7
MP supplied (g/d) 1,563 1,840 2,090 2,374 2,425 2,640
MP balance (g/d) -574 -616 -434 -573 -118 -160
Lys (% of MP) 6.17 6.07 6.33 6.24 6.40 6.38
Met (% of MP) 1.81 2.15 1.79 2.30 1.70 2.24
Lys:Met 3.43:1 2.82:1 3.54:1 2.71:1 3.78:1 2.88:1
 Methionine and the Peripartal Period
50

40
Milk yield (kg/d)

30

+ 3.4 kg + 3.8 kg
20

10 Control
Methionine

0
0 5 10 15 20 25 30 35
Day relative to parturition
Adapted from Osorio et al., 2013 Adapted from Zhou et al., 2016

Milk Yield

+ 4.2 kg

Adapted from Batistel et al., 2017

 Methionine and the Peripartal Period
22 Diet P = 0.06
Time P < 0.01
20 DxT P = 0.78
18
DMI (kg)

16

14

12
+ 2.1 kg + 2.0 kg
10

8
0 0 5 10 15 20 25 30

Day relative to parturition

Adapted from Osorio et al., 2013
Adapted from Zhou et al., 2016

Intake

+ 1.6 kg

Adapted from Batistel et al., 2017

 Methionine and the Peripartal Period
3.4 + 0.18% 3.4
+ 0.18%

3.2
Milk protein (%)

3.2

Milk protein (%)

3.0
3.0

2.8
2.8

2.6
Methionine 2.6
Control Control Methionine
Adapted from Osorio et al., 2013 Adapted from Zhou et al., 2016

3.6
+ 0.16%
3.4

Milk protein (%)

Milk protein % 3.2

3.0

2.8

2.6
Control Methionine

Adapted from Batistel et al., 2017

 Methionine and the Peripartal Period
1.6
1.4 + 0.12 kg/d 1.5 + 0.18 kg/d

Milk protein yield (kg/d)

1.3 1.4
Milk protein yield (kg/d)

1.3
1.2
1.2
1.1
1.1
1.0
1.0
0.9 0.9

0.8 0.8
Control Methionine Control Methionine
Adapted from Osorio et al., 2013 Adapted from Zhou et al., 2016

1.5

1.4
+ 0.17 kg/d

Milk protein yield (kg/d)

1.3

Milk protein yield 1.2

1.1

1.0

0.9

0.8
Control Methionine
Adapted from Batistel et al., 2017
Methionine
Milk yield & Performance

Antioxidant

Liver function

DNA & Histone methylation

Reduced immunosuppression
Outline
• Peripartal Period

• Methionine and the Peripartal Period

• Liver Function

• Immune system

• Histone methylation

• Remarks
 Methionine Cycle
DNA & RNA VLDL
Apolipoproteins
Folate
Choline
Milk
yield DNA & Histone
Met Methylation
Cycle

Cysteine

Taurine Immunosuppression

Glutathione
Immune
cells
Antioxidant
 Liver function
Genetics

Management

Nutrition

Performance

Prepartum Postpartum
 Methionine plays several roles in liver
Peripartum

Acetate ↑ NEFA

Esterification
+

Choline + FA + TG TG

Phospholipid
synthesis +
VLDL,
Met Apoprotein secreted
synthesis from liver
Amino acids (Met, et al.)
Glutathione
Taurine
(antioxidants) Major secretory protein
Albumin synthesized by liver
 Liver function
Carnitine metabolism
 Liver function
Carnitine metabolism

Lysine Methyllysine Dimethyllysine Trimethyllysine

Methyl Methyl Methyl

 Liver function
Carnitine metabolism
 Liver function
Carnitine metabolism

180
Free carnitine CON

Free carnitine (nmoles/g of wet tissue)

160
Met P < 0.01 MS
140 SM
120

100

80

60

40

20

0
7 21

Day relative to calving

Osorio et. al., 2014

 Liver function
ApoB-100
30
Diet, P = 0.16
Time, P = 0.97
25 D × T, P = 0.75
Contrast, P = 0.14
ApoB 100 (ng/mL)

20

15

10

0
Control
VLDL = Very-low density lipoproteins
Methionine
Adapted from Osorio et. al., 2013
540
ApoB-100
VLDL
0.58
520
Diet, P = 0.001
Diet, P = 0.008
ApoB 100, ug/mL

0.56

0.54

VLDL, mmol/L
500
0.52

0.50
480

0.48

460 0.46

0.44
Control Methionine Control Methionine
Adapted from Sun et. al., 2016 Adapted from Sun et. al., 2016
 Inflammatory response
Metritis

+ APP:
Serum amyloid A (SAA)
Local acute Haptoglobin
Inflammatory Ceruloplasmin
response

Liver
Mastitis - APP:
Albumin
Apolipoproteins

Kuby Immunology, 2007

 Inflammatory response
Acute
Phase
Proteins

Metritis Mastitis

Liver

Potential Pathogen Infection

 Cow in negative
Fat or Adipose
Tissue energy balance Liver

NEFA NEFA
NE, Epi Insulin

TG

NEFA Inflammatory
Response
Signal

(modified from Drackley, 1999)

 Inflammatory response
Acute Albumin
Phase
Proteins

Metritis Mastitis

Liver

Potential Pathogen Infection

 Liver function

Diet, P = 0.28
Time, P = 0.04
38 D × T, P = 0.20 Milk yield +6.5 kg/d in Upper
Contrast, P = 0.15
37
Albumin (g/L)

36

35

34 CON
MS
SM
33
-21 -10 7 14 21

Day relative to parturition

Osorio et. al., 2014 Bertoni et. al., 2008. J Dairy Sci
 Inflammatory response
Ceruloplasmin Acute Albumin
Serum Amyloid A Phase
Proteins

Metritis Mastitis

Liver

Potential Pathogen Infection

 Liver function
Ceruloplasmin Serum Amyloid A

3.2 - 0.33 umol/L Diet, P = 0.03 80

- 18.9 ug/L Diet, P = 0.17
Time, P < 0.01 Time, P = 0.24
D × T, P = 0.67 D × T, P = 0.47
3.0 Contrast, P < 0.01 Contrast, P = 0.06
Ceruloplasmin (µmol/L)

60

SAA (ug/mL)
2.8

2.6 40

2.4
20
2.2

2.0 0
CON MS SM CON MS SM

CON = Control
Osorio et. al., 2014 MS = Metasmart
SM = Smartamine M
 Methionine Cycle
DNA & RNA VLDL
Apolipoproteins
Folate
Choline
Milk
yield DNA & Histone
Met Methylation
Cycle

Cysteine

Taurine Immunosuppression

Glutathione
Immune
cells
Antioxidant
Dairy and Food Science Department
 Liver function
Oxidative Stress: ORAC and Glutathione

Avg. 0.69 mol/L higher Avg. 0.36 mM higher

14 2.5
3.5
Glutathione
CON
3.0 Met P = 0.04
MS
2.0
13 SM

Glutathione (mM)
(mM)
2.5
ORAC (mol/L)

1.5
2.0

Glutathione
12

1.5
1.0

11 1.0
0.5
0.5

10 0.0
0.0
Control Metasmart Smartamine M Control
-10 Metasmart
7 Smartamine21
M

Day relative to parturition

Osorio et. al., 2014

ORAC = Oxygen radicals absorbance capacity
 Liver function
Glutathione Priming Effect

3.5
Glutathione
CON
3.0 Met P = 0.04
MS
SM
Batistel et al., 2018

Glutathione (mM)
2.5

2.0

1.5

1.0

0.5

0.0
-10 7 21

Day relative to parturition

Osorio et. al., 2014

Zhou et al., 2016

Outline
• Peripartal Period

• Methionine and the Peripartal Period

• Liver Function

• Immune system

• Histone methylation

• Remarks
 Inflammatory response

Immune
Mastitis cells
Metritis
Ceruloplasmin
Serum Amyloid A Acute
Phase Albumin
Proteins

Liver

Potential Pathogen Infection

Peripheral blood polymorphonuclear cells
Phagocytosis and oxidative burst
 1024
Monocytes
Phagocytosis 793

FSC-A
562

Neutrophils
331

100
0 1 2 3 4
10 10 10 10 10
SSC-A

60 55.1 a
Phagocytosis at 21 d (%)

50 a
45.8

40 38.5 b
Diet P = 0.07
30 SEM = 5.6

20

10

0
Control MetaSmart Smartamine
Osorio et. al., 2013
 Met on immune function
Phagocytosis 6.55% higher
68 Met P = 0.01 With Met
Day P = 0.10 Without Met
66
Met*Day P = 0.73
64
Phagocytosis (%)

62

60

58

56

54

52

50
1 4 14 28

Day relative to parturition

Zhou et. al., 2016

 Met on immune function
Oxidative Burst 8.24 % higher

70 With MET
Met P=0.03 Without MET
Day P=0.45
65 Met*Day P=0.57
Oxidative Burst (%)

60

55

50

45
1 4 14 28

Day relative to parturition

 Osorio Experimental Design
Overfed + SM = Smartamine M (0.07% DM)
Overfed = 1.54 Mcal/kg DM
Control = 1.24 Mcal/kg DM

Overfed +SM Lactation+SM

Overfed Lactation
Far-off Control Lactation
Far-off Close-up Lactation

-50 d -21 d Calving +30 d

 Met on immune function
Cellular receptors
14

12
SELL TRT P = 0.07 Blood vessel
Time P = 0.96
10 TRT*Time P = 0.06 a
8

4 b
Relative mRNA expression

0 CON
b
-2 OVE
OVE+SM
-4

ITGAM TRT P = 0.07

20 Time P = 0.96
TRT*Time P = 0.06
Infection site
a
15
a Metritis Mastitis
10
Metal detector
ab
b
5 b
b
0

-10 3 21
Day relative to parturition
Osorio et. al., 2014
 Met on immune function
6.0
5.8
SCC Diet, P = 0.06
60
Milk yield
Time, P < 0.01 50
5.6 D × T, P = 0.90

Milk yield (kg/d)

40
Log10(SCC)

5.4
5.2 30

5.0 20 CON
OVE
4.8 10 OVE+SM
4.6 0
4.4 24
DMI
4.2 22
20
0 1 2 3 4 5 18

DMI (kg/d)
16
Week relative to parturition 14
12
CON = Control 10

OVE = Overfed
8
6
OVE+SM = OVE + Smartamine M 4
0 5 10 15 20 25 30

Li et. al., 2016 Day relative to parturition

 Met on immune function
22 TRT P = 0.33
SOD1 Time P = 0.36
20
TRT*Time P = 0.10
18
16
a
14
12
10
Cytosol
8
6
ab
CON
4 b
OVE
2 OVE + MET
Glutathione
1.5
SOD2
1.0 TRT P = 0.06
a Time P = 0.04 a
TRT*Time P = 0.08
0.5
ab
0.0 Mitochondrial
-0.5
b b
-1.0

-1.5 b
-2.0
-10 3 21
Day relative to parturition
Modified from Li et. al., 2016
Outline
• Peripartal Period

• Methionine and the Peripartal Period

• Liver Function

• Immune system

• Histone methylation

• Remarks
Methionine (AA), glucose, and lipids

Bioactive molecules
Methionine and Nutrigenomics
Milk yield & Performance

Antioxidant

Liver function

Reduced immunosuppression

Gene Expression -> RNA

Methionine supplementation alters the liver transcriptome
2,663 genes with diet × time effect
5

0.9

0.8

0.7

0.6

0.5

0.4

0.3

0.2 Control Methionine Gene expression

-10.0 7.0 21.0 -10.0 7.0 21.0

-10 7 21 -10 7 21
Day relative to parturition
 Methionine and Nutrigenomics
A model for gene regulation of Methionine in dairy cows
 NUTRIGENOMICS
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(Russian)
• яйца куриные 6 шт.
• сливочный сыр Маскарпоне
500 г
• сахар 150 г
• печенье Савоярди 250 г
• кофе эспрессо 300 мл
• какао-порошок 1-2 ст. ложки
• коньяк (по желанию) 30-50 г
Ingredientes
• 2 xicaras de café forte e frio
• 2 colheres de vinho marsala
• 2 colheres de rum
• 5 gemas
• 200 g de açúcar
• 200 g de queijo mascarpone em
temperatura ambiente
Tiramisu
• 150 mL de crème de leite fresco
• 300 g de biscoito de tipo inglês
• 4 colheres de cacaum em pó para
polvilhar
Methionine and Nutrigenomics
Histone Methylation

Tiramisu
 Histone methylation
Fluorescent Protein (FP)

FRET
Fluorescent Microscopy
Qualitative vs. Quantitative
 Histone methylation

Control 500 uM Met

Mean Intensity
0.9

0.8

0.7

125 uM Met 0.6

0.5

0.4

0.3

0.2

0.1

0
250 uM Met
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
0
1
2
3
4
5
6
7
8
9

Control 125 uM 250 uM 500 uM

 Remarks
Methionine

Performance

Performance
Metabolism

Gene expression

?
Methionine
Histone Model
Nutrigenomic effect
Methylation

Prepartum Postpartum
Advanced Dairy Solutions