Cordain_MS_talk

Loren Cordain, Ph.D.

Colorado State UniversityFort Collins, CO USA

Potential Therapeutic Characteristics of Pre-agricultural Diets in the

Prevention and Treatment of Multiple Sclerosis

Homo sapiens

H. neanderthalensis

H. antecessor

H. heidelbergensis

H. erectus

H. ergaster

Au. rudolfensis

Au.bahrelghazali

Au. anamensis

Australopithecus habilis

Au. garhi

Au. africanus

Au. afarensis

P. robustus

Paranthropus boisei

Ardipithecus ramidus

Orrorintugenensis

Sahelanthropustchadensis

Kenyanthropus platyops

P. aethiopicus

0

1

2

3

4

5

6

7

8

Mill

ions

of Y

ears

The Hominin Fossil Record:Plio-Pleistocene Diets

As many as 20 hominin species may have existed since the evolutionary split between hominins and pongids (5-7 MYA)

No universal diet existed, but rather varied by ecologic niche, season, geographic locale, availability of edible foods

Wood B. Palaeoanthropology: hominid revelations for Chad. Nature 2002:418:133-35

Minimally Processed, Wild Plants

Highly Processed, Refined Foods

What are the HealthImplications?

Minimally Processed, Wild Animals

Plio-Pleistocene Hominin Diet: The Known – Foods That Couldn’t Have Been Eaten

These foods comprise (>70% energy) in typical Western Diets

But were virtually unknown in Ancestral Human Diets

Breads, Cereals, Rice and Pasta Dairy Products Added Salt

Refined Vegetable Oils Refined Sugars(except honey) Alcohol

Cordain et al. Am J Clin Nutr 2001;71:682-92

Fatty Meats

Refined sugars, grains, vegetable oils and dairy = 70.9% of energy in the U.S. food supply

Refined sugars, grains, vegetable oils and dairy represent Neolithic & Industrial era foods that were not present in traditional ancestral human diets

By default, their inclusion displaces minimally processed, wild plant and animal foods.

15.71.4

3.1

3.3

4.8

0.8

10.623.9

17.8

18.6Refined Sugars

Refined Vegetable Oils

Vegetables

Fruits

Grains

Nuts, SeedsLegumes

Eggs

Dairy

Meats, Fish

Miscellaneous

Gerrior S, Bente I. 2002. Nutrient Content of the U.S. Food Supply, 1909-99: A Summary Report.U.S.D.A, Center for Nutrition Policy and Promotion. Home Economics Research Report No. 55

Evolution of the Western Diet:Neolithic (10,000 to 5,500 yrs ago) Food

Introductions 10

,000

9,00

0

8,00

0

7,00

0

6,00

0

5,00

0

4,00

0

3,00

0

2,00

0

1,00

0

Years ago

066100133167200233267300333HumanGenerations

pres

ent

33

SUCROSE

WHEAT & BARLEY DOMESTICATED ~10,000 YRS AGO

FIRST DAIRYING EVIDENCE

SHEEP, GOATS, COWS DOMESTICATED

WINE & BEER

FIRST SALT MINES

Evolution of the Western Diet:Industrial Revolution (~200 yrs ago)

1797

1827

1857

1887

1917

1947

1977Year

0234567HumanGenerations

2007

1

REFINED GRAINS

HFCS

HYDROGENATED OILS

SUCROSE

REFINED VEGETABLE OILS

FEEDLOT PRODUCED MEATS

Generations (~30 yrs) in the Evolution of Humanity

Generations % TotalHomo habilis (1st Homo species ) 76,667 100.0Homo erectus (modern body size) 60,000 78.2Modern Homo sapiens (cranial size) 6,666 8.7Agricultural Revolution (cereals) 333 0.4 Advent of Dairying (milk, cheese etc) 200 0.26Industrial Revolution (refined sugars, 7 0.009 refined cereals, oils, canned food) Food Processing Industry (junk food) 4 0.005

Conclusion: 99.6% of all Homo generations had noevolutionary experience with commonly consumed modern foods introduced during the Neolithic!

Neolithic and Industrial Era Foods: Nutritional Implications

As Neolithic & Industrial Era foods displace minimally processed, wild plant and animal foods, they adversely affect the following nutritional factors:

1. The Glycemic Load 2. The Fatty Acid Balance 3. The Macronutrient Balance 4. The Trace Nutrient Density 5. The Acid/Base Balance 6. The Sodium/potassium

Balance 7. The Fiber Content

Disruption of these 7 nutritionalcomponents fundamentally underliesmuch of the chronic diseases in the Western World

Item % total energy

Whole grains 3.5 Refined grains 20.4 TOTAL:23.9

85 % of all grains are consumed as refined grains


Contribution of CerealsTo Total Energy in the U.S. Diet


(Cereals)

Cereal grains which are the seeds of grasses (Gramineae) in their wild state are:

1. Small 2. Difficult to harvest3. Minimally digestible without

(a) grinding to break down cell walls (b) cooking to gelatinize starch granules

Cordain L. Cereal grains: humanity’s double edged sword. World Review of Nutrition and Dietetics 1999;84:19-73

Ancestral Human Diet: Foods That Couldn’t Have Been Eaten

(Cereals)

Bar-Yosef O. The Natufian culture in the Levant, threshold to the origins of agriculture. Evol Anthropol 1998; 6:159-177.

Wright K. The origins and development of ground stone assemblages in Late Pleistocene Southwest Asia. Paleorient 1991;17:19-45

Thus, the appearance of crude grindstones and mortars in the Middle East (Natufians) and elsewhere (10-15,000 years ago) heralds the beginnings of humanity’s use of cereal grains as a staple food


(Cereals)

High Glycemic Foods ALMOST ALL REFINED GRAINS

HAVE HIGH GLYCEMIC INDICES Rice Chex Cereal 89 Corn flakes 84 Pretzels 83 Rice Krispie Cereal 82 Rice Cakes 82 Rye bread 76 Waffles 76 Total Cereal 76 Graham crackers 74 Cheerios 74 Bagels 72 Short grain white rice 72 Corn chips 72 White bread 70 Whole Wheat bread 69

HIGH G.I. FOODS > 70MEDIUM G.I. FOODS 55-70LOW G.I. FOODS < 55

Foster-Powell K et al. Am J Clin Nutr 2002;76:5-56

High Glycemic Load Carbohydrates Promote Diseases of Insulin Resistance

Type 2 Diabetes Hypertension Coronary Heart Disease (CHD) Dyslipidemia (Reduced serum HDL

cholesterol, elevated triglycerides, elevated VLDL, elevated small dense LDL cholesterol)

Obesity Gout

Liu S et al. Dietary glycemic load and atherothrombotic risk. Curr Atherosclerosis Rep 2002;4:454-61

Ludwig DS. The glycemic index. Physiological mechanisms relating to obesity, diabetes and cardiovascular disease. JAMA 2002;287:2414-23.

The Metabolic Syndrome

Item % total energy

Whole milk 1.6 Low fat milks 2.1 Cheese3.2 Butter 1.1 Other 2.6 TOTAL:10.6


Contribution of Dairy ProductsTo Total Energy in the U.S. Diet


(Dairy)


(Dairy)

Ever tried to approach a wild animal?How About Milking It?


(Dairy)10

,000

9,00

0

8,00

0

7,00

0

6,00

0

5,00

0

4,00

0

3,00

0

2,00

0

1,00

0

Years ago

O100150200250300350400450500HumanGenerations

pres

ent

50

FIRST DAIRYING EVIDENCE

SHEEP, GOATS, COWS DOMESTICATED

Hiendleder S et al. Proc R Soc Lond B 2002;269:893-904 (SHEEP); Luikart G et al. ProcNatl Acad Sci 2001;98:5927-32 (GOATS); Loftus RT et al. Mol Ecol 1999 8:2015-22 (COWS)

Copley MS et . Proc Natl Acad Sci 2003;100:1524-29

Evolution of the Western Diet:Neolithic Food Introductions (Dairy)

ALMOST ALL REFINED GRAINS HAVE HIGH GLYCEMIC INDICES

Rice Chex Cereal 89 Corn flakes 84 Pretzels 83 Rice Krispie Cereal 82 Rice Cakes 82 Rye bread 76 Waffles 76 Total Cereal 76 Graham crackers 74 Cheerios 74 Bagels 72 Short grain white rice 72 Corn chips 72 White bread 70 Whole Milk 27 Yogurt 24

Foster-Powell K et al. Am J Clin Nutr 2002;76:5-56

Despite a low glycemic load,dairy products paradoxicallyhave insulin indices similar towhite bread

Ostman EM et al. Am J Clin Nutr 2001;74:96-100

The Displacement of Game Meats & Fish by Dairy Foods Increases Saturated Fats at the Expense of

Polyunsaturated Fats & Monounsaturated Fats

2.8 3.7

35.728.7

33.128.9

36.2

23.6

63.7 62.2

23.6

38

010203040506070

Cheese Whole Milk Salmon Venison

Polyunsaturated Fat Monounsaturated Fat Saturated Fat

% T

otal

Fa t

s

Total Fat (74% fat) (49% fat)) (46.0% fat) (19.0 %) (by energy)

Increased Saturated Fat = Increased risk for Syndrome X,CHD, certain cancers

Hot Dogs82 % Fat, 14 % Protein

Salami74 % Fat, 22 % Protein Ground Beef

64 % Fat, 33 % Protein

T-bone Steak68 % Fat, 30 % Protein

Bacon77 % Fat, 21 % Protein

Pork Ribs72 % Fat, 26 % Protein

Plio-Pleistocene Hominin Diet: The Known – Foods That Were Rarely Eaten

(Fatty Meats)

http://thingsihate.org/images/bacon-hormel-big.jpg

Wild vs. Domestic Animals

Body fat in wild animals waxes and wanes seasonally

With the advent of animal husbandry 10,000 years ago, it became possible to attenuate or prevent the seasonal decline in body fat % by provisioning captive animals with plant food

It also became feasible to consistently slaughter the animal at peak body fat %

Caribou

Increased Saturated Fat = Increased risk CHD, Syndrome X, certain cancers

Total -3 Fatty Acids in Wild, Grass and Grain Fed Animals Muscle Meat

(100 g sample)

178

225 216

6146

0

50

100

150

200

250

Elk Deer Antelope Pasture fedCow

Grain fedCow

Tota

l n- 3

fatty

aci

ds (m

g)

Cordain et al. Eur J Clin Nutr 2002;56:181-91

Diseases linked to reduced ω-3 fatty acids: Syndrome X,CHD,cancer, autoimmune diseases, all inflammatory (“itis”) diseases

Total Salt (NaCl) in the U.S. Diet (Grams per Day) Source grams/day

Added in processed foods 7.2 Table salt and cooking use 1.4 Naturally occurring in foods 1.0 TOTAL: 9.6


Plio-Pleistocene Hominin Diet: The Known – Foods That Were Rarely Consumed

(Added Salt)

Salt was known to be gathered on a dry lake bed in China ~ 8,000 years ago

First inland salt mines appear in Europe ~ 6,000 years ago

Hunter gatherers living near the ocean dipped food in seawater and used dried sea salt

Inland hunter-gatherers rarely used salt on a regular basis

The first known salt mine in Europe(6,200 - 5,600 years ago)

The Mountain of Salt (Cardona, Catalonia, Spain)

Diseases linked to salt consumption: Hypertension, stroke, osteoporosis, kidneystones, Menierre’s Syndrome, stomach cancer, insomnia, motion sickness, asthma,exercise induced asthma

Weller O. Antiquity 2002;76:317-18.

Plio-Pleistocene Hominin Diet: The Known – Foods That Were Rarely Consumed

(Added Salt)

Contribution of Refined Sugars to Total Energy in the U.S. Diet

Item % total energy

Sucrose 8.0 High fructose corn syrup 7.8 Glucose 2.6 Syrups 0.1 Other 0.1 TOTAL:18.6


U.S.D.A. Economic Research Service, 2002. Food consumption (percapita) data system, sugarsSweeteners, Washington D.C.

Plio-Pleistocene Hominin Diet: The Known – Foods That Were Not Consumed

(Refined Sugars)

http://www.howstuffworks.com/gif/diet-oreos.jpg

Evolution of the Western Diet:Industrial Era Food Introductions (Refined Sugars)

0

20

40

60

80

100

120

1745

1760

1775

1790

1805

1820

1835

1850

1865

1880

1893

1905

1915

1922

1930

Year

Per c

apita

con

sum

ptio

n (lb

s.)

1937

Per Capita Sugar (Sucrose) Consumption in the Netherlands (1745-1937)

Diseases linked to refined sugars:

Metabolic Syndrome: (Type 2 diabetes, CHD, dyslipidemia, obesity, gout, hypertension

Dental caries Certain cancers


(Refined Sugars)

Cordain L et al. Hyperinsulinemic diseases of civilization: more than just syndrome X. Comp Biochem Physiol Part A 2003;136:95-112.

Contribution of Refined Vegetable Oils to Total Energy in the U.S. Diet

Item % total energy

Salad, Cooking Oils 8.8 Shortening 6.6 Margarine 2.4 TOTAL:17.8



(Refined Vegetable Oils)

05

1015202530

1909-19

1920-29

1930-39

1940-49

1950-59

1960-69

1970-79

1980-89

1990-99

Margarine Shortening Salad, cooking oils Total Vegetable Oils

kgPer Capita Change in Refined

Vegetable Oils in the U.S. (1909-99)

Total vegetable oil consumption has increased 459 % since 1909


Salad, Cooking Oil consumption has increased 1340 % since 1909Margarine consumption has increased 488 % since 1909Shortening consumption has increased 237 % since 1909

Vegetable oils are high in ω-6 fatty acids, but low in ω-3

Diseases linked to high ω-6/ ω-3

Metabolic Syndrome: (Type 2 diabetes, CHD, dyslipidemia, obesity, gout, hypertension), cancers, autoimmune diseases, virtually all inflammatory (“itis”) diseases


(Refined Vegetable Oils)

Recommendations for a Contemporary Diet Based Upon

Paleolithic Food Groups

Fresh FruitsFresh Veggies Nuts/Seeds

FishSeafood

Lean Meats

Neolithic Food Introductions: Potential Factors underlying Multiple Sclerosis

Milk and Dairy ?

Dietary Lectins ?

Legumes:Peanuts

Legumes:Beans

Tomatoes

Whole Grains:Wheat

Multiple Sclerosis: is an autoimmune disease in which the immune system destroys the covering (myelin sheaths) of nerve cells in the brain + spinal cord (CNS)

Activated T cells: attack proteins within the myelin which appear to be foreign

While healing may occur (remission), plaques (scarring) frequently develop throughout the CNS which impair motor and sensory function

M.S. afflicts ~ 1 in 1,000 people or ~300,000 people in the U.S.

What is Multiple Sclerosis ?

Damaged Myelin in Multiple Sclerosis

Nerve Cell

Normal MyelinNerve Fiber

Muscle Fiber

Damaged Myelin(plaques)

Brain signalis blocked because of damagedmyelin

BrainSignal:normal

T Cells

It is well recognized that both genetic endowment and environment play important roles in MS pathogenesis

“The former being stronger the latter still unidentified” 1.

The MHC locust where the HLA DRB1*1501, DQA1*0102, and DQB1*0602 haplotype is well validated as the major genetic risk factor for MS2.

The low concordance of MS in monozygotic twins (25.3%) 3 clearly is indicative that environmental factors must be involved.

1. Poser CM. Clin Neurol Neurosurg 2006;108: 227-332. Hafler DA et al. Nat Rev Immunol 2005;5:83-913. Willer C et al. Proc Natl Acad Sci 2003;100:12877-82

What Causes Multiple Sclerosis: (Genetics + Environment)

Infection: Epstein Barr virus, Chlamydia pneumoniae, Mycoplasma pneumoniae, viruses causing influenza, measles, rubella

Geography: Higher latitude increases risk; sunlight; UV exposure – vitamin D?

Physical trauma:, surgery, psychological stress, certain toxins, occupational exposures, cigarette smoking

Vaccinations Dietary Factors: Saturated/PUFA,

antioxidants, milk consumption, wheat consumption

1. Poser CM. Clin Neurol Neurosurg 2006;108: 227-332. Hafler DA et al. Nat Rev Immunol 2005;5:83-913. Willer C et al. Proc Natl Acad Sci 2003;100:12877-82

Multiple Sclerosis Etiology: Suspected Environmental Factors

Epstein Barr virus

WHY do T cells not recognize myelin proteins as “self” and attack it like a foreign protein?

[I] Because they become sensitized (outside the CNS) to foreign proteins which structurally resemble myelin: MOLECULAR MIMICRY

Damaged Myelin in Multiple Sclerosis

Muscle Fiber

Damaged Myelin(plaques)

T Cells

Nerve Cell

Multiple Sclerosis Etiology: Key Questions?

f s w g a e g

f g w g a e l

q r

n m

Molecular Matches

Myelin Basic Protein:

Escherichia coli bacteria:

WHERE: do T cells become sensitized to foreign proteins that structurally resemble myelin?

[II] They become sensitized (outside the CNS):

A. Gut, mouth, nose (mucosal linings)

B. Direct entry into blood By: Viral, bacterial and

dietary proteins (antigens)

T Cells


f s w g a e g

f g w g a e l

q r

n m

Molecular Matches



GutBacteria

HOW: do T cells become sensitized (activated) to foreign proteins that structurally resemble myelin?

[III] There is no single foreign antigen that activates T cells in Multiple Sclerosis, but rather multiple factors acting synergistically:

1. The immune system must first be primed by a prior viral or bacterial infection whose molecular structure mimics a myelin protein (many viruses and bacteria mimic various myelin proteins)

2. In most, but not all cases, there must be genetic susceptibility (E.g. HLA DRB1*1501, DQA1*0102, and DQB1*0602 haplotypes)


f s w g a e g

f g w g a e l

q r

n m

Molecular Matches



3. There must be a secondary and continued stimulation of T cells outside the CNS by foreign proteins which mimic a myelin protein or proteins

4. In addition to the mimicking proteins (immunogens), an adjuvant (immune system booster) must also be simultaneously present to develop immunity (autoimmunity)

Multiple Sclerosis: Key Questions?

GutBacteria

T Cells

ImmunogenOnly

No Immunity

Immunogen +Adjuvant

Development ofImmunity


[III] cont.

4. If the foreign protein is derived from the gut , it must cross the gut barrier and reach circulation to activate T cells in the lymph

5. Activated T cells must be able to then cross the blood brain barrier (BBB) to destroy myelin tissue

6. There must be continual and chronic activation of other immune components (inflammatory cytokines) for myelin destruction by T cells

Multiple Sclerosis: Key Questions?


Gut Cross Section:

Gut bacterial proteins: cannot normally cross gutInto circulation

circulation

circulation

Brain &CNS

BBB

[III] cont.

Lectin: Latin verb (legere) to select Originally defined by ability to

agglutinate (clump) erythrocytes More recent definition: ability to

reversibly bind a specific mono or oligosaccharide

Lectins are omnipresent in plant kingdom & likely evolved as toxic defensive mechanisms to ward off predators

Most dietary lectins are benign and non toxic to humans

Primary exceptions: Grain and legume lectins which bind to gut tissue

Legumes

Whole Grains

Multiple Sclerosis Etiology: Dietary Lectins

Common Food Lectins which May Bind Gut Tissue and Their Concentrations

Wheat germ: 300 – 350 mg/kg wheat germ agglutinin (WGA) (1)

Whole wheat flour: 30-50 mg/kg WGA (2)

White flour: 4.4 mg/kg WGA (2)

Kidney beans (Phaseolus vulgaris): 1,000-10,000 mg/kg phytohemagglutinin (PHA) (3)

Soybeans: 200 – 2,000 mg/kg soybean agglutinin (SBA) (3)

Tomatoes: <10 mg/kg tomato lectin (TL) (3)

Peanuts: 110 mg/kg peanut agglutinin (PNA) (1)

1. Vincenzi S, et al. J Agric Food Chem. 2002 Oct 23;50(22):6266-70.2. Matucci A et al. Food Control 2004;15: 391-953. Peumans WJ, Van Damme EJM. Trends Food Sci Technol 1996;7:132-39

Necessary Qualities of Lectins to Influence Multiple Sclerosis Must survive cooking and

processing Must survive digestive enzymatic

degradation Must bind gut tissue Must cross gut tissue barrier Must resist immunological and

hepatic disposal Must arrive in peripheral circulation

intact in physiological concentrations

Must interact with one or more mechanisms known to influence multiple sclerosis

Peumans WJ, Van Damme EJM. Trends Food Sci Technol 1996;7:132-39

Ingested Peanut Lectin Rapidly Enters Peripheral Blood in Humans

-1

0

1

2

3

4

5

0 0.5 1 1.5 2 4 24 48 72

Subj 1Subj 2Subj 3Subj 4Subj 5Subj 6Subj 7Im

mun

oblo

t det

ecta

ble

PNA

(µg/

mL

seru

m)

Time after peanut ingestion (h)

Wang Q, Yu LG, Campbell BJ, Milton JD, Rhodes, JM. Identification of intactpeanut lectin in peripheral venous blood. Lancet 1998;352:1831-32

Appearance of PNA in serum following consumption of either 200 g whole raw peanuts (n=2) or 200 g roasted salted peanuts (n=5)

Gut Binding Lectins Demonstrated to Enter Peripheral Circulation

Yes 4 Yes 1 Yes 1

Yes 2

Yes 3

Yes 4 Yes 5

The entry of dietary lectins into peripheral circulation has been sparsely examined in both humans and animals

It is quite likely that all lectins capable of binding gut epithelial tissue enter circulation

There may be multiple common entry pathways:

1. “M” cells 2. Paracellular entry 3. Trans-membrane

receptors

1. Kilpatrick DC, et al. FEBS Lett. 1985;185:299-305.2. Wang Q, et al. Lancet. 1998;352:1831-23. Pusztai A, et al. Biochem Soc Trans 1989;17:481-2.4. Lochner N, et al. Pharm Res. 2003 May;20(5):833-9.5. Pusztai A, et al. Br J Nutr. 1993 Jul;70(1):313-21.

In Vitro Animals Humans

TL

PNA

PHA

WGA

How Do Lectins Cross Gut Barrier

Lectins arriving intact in gut must get past:

1. Mucus lining 2. Glycocalyx: Trans-membrane

glycoconjugate layer extending from apical surface of epithelial cells

Glycocalyx dimensions: (400-500 nm thick) & pore

diameter: (7.4 – 28.8 nm) Function: Size selective

diffusion barrier that excludes bacteria, viruses & foreign material from contacting enterocyte membrane

3. Cell membrane (Either absorptive enterocytes, “M” cells, goblet cells, Paneth cells, or enteroendocrine cells)

Glycocalyx

Microvilli

or “Brush

Border”

Mucus

http://www.uni-mainz.de/FB/Medizin/Anatomie/workshop/EM/eigeneEM/Colon/Col66b.jpg

How do Lectins Cross Gut Barrier

Specialized epithelial cell found only in lymphoid follicle

Function: take up luminal material (microorganisms, dietary antigens) & transport it to underlying lymphoid tissue

Lack dense villi Glycocalyx thin or lacking Maintain apical surface

glycoproteins which bind gut antigens

Entry pathway exploited by certain pathogens

M

EM M M

E

M

Single Lymphoid Follicle:Within a Peyer’s

PatchM

cell

Villus

Crypt

EnterocytesT & B lymphocytes

E

1. Endocytosis of Lectins by “M” Cells

GUT LUMEN

TightJunctions

Enterocytes

BrushBorder (Villi)

Dietary Lectins(WGA/PHA)

2. Paracellular Transport of Lectins via IncreasedEnterocyte Permeability “Leaky Gut”

1. Sjolander A et al. The effect of concanavalin A and wheat germ agglutinin on the ultrastructureand permeability of rat intestine. Int Arch Allergy Appl Immunol 1984; 75, 230–236.2. Greer F & Pusztai A (1985) Toxicity of kidney bean (Phaseolus vulgaris) in rats: changes inintestinal permeability. Digestion32, 42–46.3. Pellegrina CD et al. Plant lectins as carriers for oral drugs: Is wheat germ agglutinin a suitable candidate? Toxicol Appl Pharmacol 2005;207:170-78


WGA

Enterocyte Cytosol

Gut Lumen(Inside)

WGA

WGA

PHA

Into Lymph

Into Circulation

WGA and PHA gain access to circulation via enterocyte endocytosis at the EGF-R

The gut EGF-R is expressed luminally in humans 1 and binds galactose specific PHA 2

and neuraminic acid specific WGA 3

Other legume, cereal and tomato lectins likely use this pathway because of common sugar binding affinities

EGF R

eceptor

EGF R

eceptor

EGF Receptor

1. Hormi K et al. Cell Tissue Res 1994;278:439-502. Rebbaa A et al. J Neurochem 1996;67:2265-22723. Lochner N, et al. Pharm Res. 2003 May;20(5):833-9.

or


3. Entry via Trans Membrane Receptor

Necessary Qualities of Lectins to Influence Multiple Sclerosis

Must survive cooking and processing

Must survive digestive enzymatic degradation

Must bind gut tissue Must cross gut tissue barrier Must resist immunological and

hepatic disposal Must arrive in peripheral

circulation intact in physiological concentrations

Must interact with one or more mechanisms known to influence multiple sclerosis

Peumans WJ, Van Damme EJM. Trends Food Sci Technol 1996;7:132-39

WGA

Enterocyte Cytosol

Gut Interior

WGA

WGA

Into Lymph

Into Circulation

Lectins simultaneously bind bacterial and/or food antigens in the gut + EGF Receptor

Both the (Lectin + bacteria or food antigen) enter the lymph/ circulation where:

They may activate T Cells Multiple gut bacteria display

molecular mimicry with myelin proteins

EGF R

eceptor

EGF R

eceptor

EGF Receptor

[I] &[II] T – Cells must become sensitized (outside the CNS) to foreign proteins which structurally resemble myelin: MOLECULAR MIMICRY

Lectins Must Interact With Mechanisms Influencing MS: Molecular Mimicry

B/FB/F

B/F

B/F

B/F

Westall FC. Molecular mimicry revisited:gut bacterial and multiple sclerosis. J ClinMicrobiol 2006;44:2099-2104

Bacterial/Food PeptidesLectin

Activate T Cells

WGA

3. There must be a secondary and continued stimulation of T cells outside the CNS by foreign proteins which mimic a myelin protein or proteins

Lectin transport mimicking gut proteins into circulation

4. In addition to the mimicking proteins (immunogens), an adjuvant (immune system booster) must also be simultaneously present to develop immunity (autoimmunity)

WGA and Tomato Lectin 1 simultaneously act as adjuvants + vehicles for immunogen transport

Lectins likely transport gut bacterial cell wall proteins (adjuvants)2 into circulation

ImmunogenOnly No Immunity

Immunogen +Adjuvant

WG

A

Development ofImmunity

[III]

Lectins Must Interact with Mechanisms Known to Influence MS: Adjuvants

1. LaVelle EC et al. The identification of plantLectins with mucosal adjuvant activity.Immunology 2001;102:77-86.2. Visser L. Proinflammatory bacterial peptido-glycan as a cofactor for the development of central nervous system autoimmune disease.J Immunol 2005;174:808-816.

Lectin

Lectins Must Interact with Mechanisms Known to Influence MS: Inflammation

Pathogenic mechanisms contributing to MS include:

1. Leukocyte chemotaxis into the CNS (Crossing the BBB)

2. Production of inflammatory mediators resulting in:

Oligodendrocyte damage, demyelination and neuronal injury

MS characteristically elevated cytokines: IFNγ, TNFα, TNFβ IL-2, IL-1β, IL-6, IL-8, IL-12, IL-18

Inflammatory Cytokines(TNFα, IFNγ, IL-12, IL-2)

APC

MHC

Cl

ass

II

Myelin peptide

TCR Naïve Autoreactive

T cell

Activated T cell

Microglial cell

B cell

Autoantibody

Complement Inflammatory cytokines

(TNFα, IFNγ, IL-12, IL-2)

Blood

BBB (Endothelial cells)

CNS

Adhesion Molecules

ICAM-1VCAM-1

Monocyte

Holmes S et al. Exp Rev Mol Med 2005;7: 1-17

Lectins Stimulate Inflammatory Cytokines in Vitro Mononuclear Cells

6

3.1

5.2

002468

LCA(lentil)

PHA PSA (pea) WGA

58.637.4

70.6

30

0255075

LCA(lentil)

PHA PSA(pea)

WGA

18.15.9

52.2

26

0204060

LCA(lentil)

PHA PSA(pea)

WGA

IL-2 (24 hrs)

IL-12 (48 hrs) INF-γ (24 hrs)

Arb

itrar

y U

/ml

Arb

itrar

y U

/ml

Arb

itrar

y U

/ml

Muraille E et al. Cell Immunol 1999;191:1-9

PHA stimulates TNF α; TNF β, IL-1 β, IL-2, IL-6, IL-8, IL-12 and INF γ 1-4

WGA stimulates IL-12 and INF γ 3 SBA stimulates IL-6 5

1. van den Borne BE et al. J Rheumatol 1997;24:55-602. Isler P et al. Eur Cytokine Netw 1993;4:15-23;3. Muraille E et al. Cell Immunol 1999;191:1-9.4. Baran J et al. Clin Diag Lab Immunol 2001;8:303-13.5. Jenkins DJ et al. Metabolism 2002;51:919-924.

Lectins Stimulate Inflammatory Cytokines in Vitro Mononuclear Cells (cont.)

Specifically In MS Patients PHA Stimulates: PHA stimulates TNF α; IL-1 β, IL-2, and INF γ 6-10

6. Hollifield RD et al. Autoimmunity 2003;36:133-417. Gusev EI et al. J Neurol 1994;241:500-5108. Chofflon M et al. Eur Cytokine Netw 1992;3:523-319. Martino G et al. J Neuroimmunol 1995;62:169-7610. Guillen C et al. Immunopharmacol Immunotoxicol 1999; 21:527-49

INF γ is pivotal in MS pathogenesis and progression

Relapses are preceded by increased INF γ in cerebrospinal fluid and by peripheral lymphocytes

Administration of recombinant INF γ leads to disease worsening

Conversely, IFN β is a well established immunomodulatory drug for MS treatment

Both PHA and WGA increase INF γ production in vitro

Cosentino M et al. J Neuroimmunol 2005;162:112-121.

Lectins Stimulate Inflammatory Cytokines in Vitro Mononuclear Cells (cont.)

Lectins Must Interact with Mechanisms Known to Influence MS: Leukocyte Chemotaxis

Pro-inflammatory cytokines (TNF α, IL-1 β) induce VCAM-1 and ICAM-1 expression in endothelial cells

Hence increase influx of monocytes and T lymphocytes from blood into peripheral tissue and CNS 1,2

IL-8 is elevated in leukocytes of untreated MS patients 3 and is responsible for BBB disruption and migration of leukocytes into CNS 4

PHA elevates TNF α, IL-1 β and IL-8 in vitro

Inflammatory Cytokines(TNFα, IFNγ, IL-12, IL-2)

APCM

HC

Clas

s II

Myelin peptide

TCR Naïve Autoreactive

T cell

Activated T cell

Microglial cell

B cell

Autoantibody

Complement Inflammatory cytokines

(TNFα, IFNγ, IL-12, IL-2)

Blood

BBB (Endothelial cells)

CNS

Adhesion Molecules

ICAM-1VCAM-1

Monocyte

1. van den Borne BE et al. J Rheumatol 1997;24:55-60;2. Bullard DC et al. J Immunol 2007;178:851-7 3. Lund BT et al. J Neuroimmunol 2004;155:161-714. Mirowska-Guzel DM et al. J Neuroimmunol 2006;176:134-140

PHA stimulates IL-18 production in T cells 1

IL-18 up-regulates VCAM-1 and ICAM on endothelial cells 2

Direct cell to cell contact of T cells with (monocytes/ macrophages) in peripheral tissue

Causes up-regulation of TNF α and IL-1 β 1

Neutralizing antibody to IL-18 prevents monocyte activation in direct cell to cell contact 1 and prevents EAE 3

1. Dai SM et al. Arthritis Rheum 2004; 50: 432-432. Morel JC et al. J Biol Chem 2001;276:37069-753. Fukaura H et al. Nippon Rinsho 2003;61:1416-21

Lectins Stimulate Leukocyte Chemotaxis

Image of Endothelial VCAM-1In Vivo

The endothelial glycocalyx, a highly hydrated mesh of membrane-bound negatively charged proteoglycans, glycosaminoglycans, glycoproteins, and glycolipids, has a thickness of around 500 nm

The “hairy” structures form the endothelial glycocalyx.

Shields the endothelial cells from the flowing blood and forms a mechanical barrier against adhesion of leukocytes to endothelial surface

Electron microscope overview of a alcian blue 8GX-stained rat left

ventricular myocardial capillary.

(bar=1m).

Van den Berg, Vink, Spaan, Circulation Research 2003, 92: 592-594

Lectins May Promote Influx of Monocytes and T CellsInto the Intima by Disrupting the Glycocalyx


Inflammatory agents (TNF α, thrombin) and growth factors (EGF) cause rapid shedding of glycocalyx 1,2

Facilitating Influx of leukocytes

1. van den Berg BM et al. Am J Physiol Heart Circ Physiol 2006;290:H915-H9202. Subramanian SV et al. J Biol Chem 1997;272:14713-20.

Crystal Structure of TNFαGlycocalyx


It is likely that dietary lectins disrupt and reduce the thickness of the glycocalyx and thereby facilitate influx of leukocytes into the intima

WGA, PNA (neuraminidase), bind myocardial, retinal and brain arteriole glycocalyx 1

Tomato lectin binds variety of human arteriole glycocalyx 2

WGA 3 and PHA 4 bind the EGF-R which causes rapid shedding of the glycocalyx 5

The Arterial Glycocalyx

1. Lawrenson JG et al. J Anat 2000;196:55-602. Debbage PL et al. Histochem Cell Biol 2001;116:349-593. Lochner N, et al. Pharm Res. 2003 May;20(5):833-9.4. Rebbaa A et al. J Neurochem 1996;67:2265-22725. Subramanian SV et al. J Biol Chem 1997;272:14713-20.

Lectins May Promote Influx of Monocytes and T CellsInto the Intima by Disrupting the Glycocalyx


Potential Mechanisms of Lectin Involvement in Multiple Sclerosis

010203040506070

MMP-9 MMP-2

Control WGA MMPs are elevated in MS and

are implicated in disease etiology 1,2 by:

1. Disrupting BBB and facilitating influx of leukocytes

2. Degrading Myelin Basic Protein

PHA up-regulates MMP-9 in human monocytes 3

WGA up-regulates MMP2 in human monocytes, but not MMP-9 4

Lectins Up-Regulate Matrix Metalloproteinases (MMPs)

1. Ram M et al. J Clin Immunol 2006;26:299-3072.Yong VW et al. J Neurol Sci 2007; Mar 22; [Epub ahead of print3. Dubois B et al. FEBS Letter 1998;427:275-278.4. Saja K et al. Mol Cell Biochem 2007;296:185-192

ns

* (p<0.05)

Multiple Sclerosis Etiology: Milk and Dairy

Epidemiological studies have repeatedly associated MS prevalence with milk drinking 1- 4

Milk drinking correlated highly to MS prevalence in 27 countries (Spearman’s rho = 0.84) 3

Epidemiological & experimental studies also link rheumatoid arthritis and type 1 diabetes to milk consumption

Milk and Dairy

1. Agranoff BW et al. Lancet 1974;2:1061-662. Butcher PJ. Med Hypotheses 1986;19:169-1783. Malosse D et al. Neuroepidemiol 1992;11: 304-3124. Lauer K. Neurology 1997; 49:s55-s61

Multiple Sclerosis and Milk: Proposed Mechanisms of Action

In Dark Agouti Rats, immunization with the bovine milk protein, butyrophilin (BTN) triggers EAE 1

Via a class II MHC restricted T cell response that cross reacts (molecular mimicry) with the autoantigen, myelin oligodendrocyte glycoprotein (MOG) peptide sequences 76-87 (IGEGKVALRIQN) 1

Transmucosal (intranasal) administration of BTN or by I.V. suppresses disease activity 1

In MS patients, MOG specific autoantibodies cross react with multiple epitopes of BTN 2

Human Interpretation - Caveats: 1) Will require permissive MHC haplotype, 2) BTN crosses gut barrier intact and induction of oral tolerance must be abrogated

1. Stefferl A et al. J Immunol 2000;165:2859-28652. Guggenmos J et al. J Immunol 2004;172:661-68

Milk and Dairy

Dark Agouti Rat

Multiple Sclerosis and Milk: Proposed Mechanisms of Action (cont.)

0

0.5

1

1.5

2

2.5

MS Patients HealthyControls

An additional bovine milk protein (BSA 193) was encephalitogenic to SJL mice subjected to EAE induction

Structural homology (GLCHMYK) existed between exon 2 of the autoantigen myelin basic protein (MBP) and BSA 193

However, T cell cross reactivity did not occur

1. Winer S et al. J Immunol 2001;166: 4751-56

SJL Mouse

Prol

ifera

tive

T C

ell

Res

pons

e (S

I) Further, MS patients had

significantly (p< 0.0001) higher proliferative T cell responses to BSA 193 than healthy controls

*p<0.0001

Multiple Sclerosis and Milk: Summary Animal EAE and human tissue

studies suggest that at least two bovine milk proteins (BTN and BSA 193) may be involved in MS etiology by molecular mimicry 1-3

Specific permissive human MHC haplotypes are required for cross reactivity 1

Bovine proteins must cross the gut barrier and oral tolerance must be abrogated 1, 3

Neonatal exposure to dietary antigens that cross react with CNS autoantigens may enhance disease susceptibility later in life 4

Milk and Dairy

1. Stefferl A et al. J Immunol 2000;165:2859-28652. Winer S et al. J Immunol 2001;166: 4751-563. Guggenmos J et al. J Immunol 2004;172:661-683. 4. Miller AO et al. Eur J Immunol 1994;24:1026-32

Influence of Diet on Disability in Four Multiple Sclerosis Patients

012345678

-22 -20 -18 -16 -14 -12 -10 -8 -6 -4 -2 0 2 4 6

Subj 1 (F) Subj 2 (F) Subj 3 (F) Subj 4 (M)

Kur

tzke

Exp

ande

d D

isab

ility

Sca

le

Years on diet

Anecdotal data provided by A Embry. Direct MS, Canada: http://www.direct-ms.org/

Years with MS

Diets: grain, legume and dairy free

Thank You!

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