WHY WE AGE AND HOW TO STOP IT

Dr. Sandra Kaufmann

Master's in Tropical Ecology and Evolutionary

biology @ University of Connecticut

Medical School @ University of Maryland

Fellowship in Pediatric Anesthesia @ Johns

Hopkins

Chief of Pediatric Anesthesia @ Joe DiMaggios

Children's Hospital
 What the Protocol is Not

● It is not about diet and exercise

● It is not a treaty on weight loss
● It is not hormone replacement
● It is not just for older people
● Does not offer specific treatments for disease
states
 What the Protocol IS
● Comprehensive Interpretation of Cellular Aging
● All information is derived from real science
● Evidence is extrapolated from individual cells and animal
models to the human form
● My theory is presented in a unique factory model
● Practical applications to decelerate the aging process
 What is aging?
● “Aging is associated with a generalized decline in all physiological
functions, and between 30 and 70 we are likely to observe a 25-30%
reduction on most functional capacities.” (Barbieri 2015)

● “...a deterioration in the maintenance of homeostatic processes

over time leading to functional decline and increased risk of death and
disease…” (Barzailai 2012)

● “Aging is a complex multifactorial process of molecular and

cellular decline that affects tissue function over time, rendering
organisms frail and susceptible to disease and death.” (Carmona 2016)
The Body as a Factory

● Company Operating Manual

● Energy source
● Pathways…Assembly lines
● Quality Control
● Security systems
● Work Force
● Waste Management
 Analogy Applied to cells
● Company operating manual DNA
● Energy Source Mitochondria
● Pathways Pathways:
AMPKinase, sirtuins, mTOR
● Quality control DNA and Protein
repair mechanisms
● Security Immune system
● Workforce Individual cell
requirements
 Tenet 1: DNA
● Understand the science of DNA packaging

● Epigenetics

● Telomeres and telomerase

 Epigenetics
Decoration or Methylation of DNA
Phosphates, acetyl groups or
methylation of Histones
 Epigenetics

● Predictable pattern changes over time - Horvath

clock

● Epigenetic Drift- more specific to the individual

● Bee example
● Foods and Agents that positively alter
the epigenetic pattern
Telomeres & Telomerase
● Positioned on the ends of DNA, once considered non-sense DNA
● Serve as protective caps
● Lose length with cell divisions and stresses
● High correlation between length of life and length of telomeres
● Telomerase, useful enzyme that increases the length in some cells,
esp stem cells
● Over time: lose telomere length, and lose activity of telomerase
Tenet 2: Energy…..Mitochondria

● The Powerhouse of the cell

● Different numbers in different cell types

● Understand mitochondrial dynamics…fission and fusion

 Mitochondria
● Functions via the Electron Transport Chain

● Byproduct of aerobic metabolism is radicalized Oxygen

● Lead to Mitochondrial Superoxide Theory of Aging

● Decline in endogenous scavengers over time; SOD, catalase,

glutathione, etc

● Need for additional Free radical Scavengers

● Uncontrolled radicals cause increase in DNA, protein and lipid

damage
 Mitochondria
● Electron Transport Chain functions by
passing electrons/ protons down chain
to create gradient

● Rate limiting molecule over time is Nicotinamide adenine

Dinucleotide or NAD/NADH

● Humans develop a severe deficiency of NAD with age

● Thus, energy production declines with aging

 Tenet 3: Aging Pathways

● Based on Caloric restriction

● Discovered SIRTUIN pathways

● AMP Kinase pathway

● MTOR pathway
Caloric Restriction

● Caloric restriction: a 20 to 50% caloric decrease from a

standard diet

● Prolongs the mean and the maximum lifespan in dogs,

rodents, worms, flies, fish and even yeast.
 Caloric Restriction
“Caloric restriction with adequate nutrition is the only non-
genetic, and the most consistent non-pharmacological
intervention that extends lifespan in model organisms from yeast to
mammals, and protects against the deterioration of biological
functions, delaying or reducing the risk of many age-related
diseases.” (Testa 2014)

“Caloric restriction is by far the most effective environmental

manipulation that can extend maximum lifespan in many different
species.” (Yuanyang 2011)
 Sirtuins
● Silent Information Regulator gene

● Discovered in 2000

● Yeast that had additional copy of SIRT1, lived 30% longer

● NAD dependent genes and enzymes that sense environmental and

nutritional stressors

● Activated Sirtuins:
○ Regulate the bodies metabolic and growth pathways
○ Trigger the transcription of specific proteins that enhance metabolic
efficiency
○ Increase anti-oxidant pathways
○ Facilitates DNA damage repair
 Sirtuins
● Seven mammalian sirtuins

● SIRT 1:
Located in the nucleus

○ Circadian rhythm regulation

○ Mitochondrial DNA transcription
○ Oxidative stress
○ Inflammatory pathways (NF-kB)
○ Sarcopenia
 Sirtuin families
SIRT2:
Located in the cytoplasm and nucleus

Mitosis
cellular reorganization during cell replication and division

Known to affect histones @H4K16

...thus an epigenetic modifier
 Sirtuin families
SIRT3:
Located in mitochondria

○ Orchestrates mitochondrial function

○ Increases production of superoxide dismutase
○ Apoptosis (getting rid of useless dead cells)
○ Effects brown fat expression
○ Known to affect histones @ H3K9, H3K56
SIRT4:
Located in the mitochondria
TCA or Krebs cycle

SIRT5:
Located in the mitochondria
Uric Acid cycle

SIRT6:
Located in the nucleus
Controls inflammation through effects on NF-kB
Telomeric preservation
Prevents diet-induced obesity
DNA repair
Affects Histones H3K9, H3K56
SIRT7:
Located in the nucleus
Controls nucleolar maintenance during cellular stress
Sirtuins

● Decline with age

● NAD dependent…and NAD declines with age

● Artificially activate sirtuin gene family

Are there SIRT activators? Of course

AMP Kinase
● Adenosine Monophosphate-activated Protein Kinase

● Central regulator of cellular and organismal metabolism

● Plays a critical role in maintaining energy homeostasis

● Otherwise known as the Metabolic Master Switch

○ Promotes catabolic mechanisms that generate ATP, while

simultaneously inhibiting anabolic systems that require ATP.
 AMP Kinase

In order to INCREASE ATP production:

1) Increases cellular uptake of glucose

2) Increases glycolysis

3) Increases Fatty Acid Oxidation

4) Triggers the acute destruction of defective mitochondria while

stimulating new mitochondria to be produced. (Autophagy)
AMP Kinase
In order to DECREASE ATP utilization

1) Decrease fatty acid synthesis

2) Decrease steroid synthesis

3) Decrease glycogen storage

4) Decrease protein production

5) Decrease cellular growth

What happens with the loss of AMP Kinase?

● Decrease in autophagy

● Increasing oxidative stress

● Increasing inflammation

● Increasing fat deposition

● Hyperglycemia

Are there AMP Kinase activators? Of course

mTOR: mechanistic target of Rapamycin
● mTOR is a serine/threonine protein kinase

● Senses the environment and promotes anabolic processes …its essential to

the biosynthesis of proteins and lipids

● Get hyper-functioning of cells…contributes to high blood pressure,

osteoporosis and hypercoagulability

● Pathway becomes obsolescent

● Blocking mTOR been shown to increase longevity…Rapamycin

 Rapamycin
● First inhibitor of the mTOR complex 1
central regulator of RNA translation,
cellular growth and metabolism

● Cancer treatment
Renal cell CA, neuroendocrine tumors, some breast cancers,
some leukemias, and lymphomas.

● Potent immunosuppressant after kidney transplant

● Drug-eluting cardiovascular stents

Rapamycin- the positives
● Delay in stem cell loss

● Delay in cognitive decline

● Delayed heart failure

● Delayed liver degeneration

● Less tendon stiffening

● Less decline in physical activity

● Some aspects of cancer prevention

 Rapamycin- the negatives
Side effects with doses from immune suppression:
Immunosuppression
Edema
Mouth ulcers
Alopecia
Testicular function
Fertility

Even smaller doses

Hippocampal neurons...memory
Sarcopenia
“Consolidation and reconsolidation of fear memory, spacial memory, and
modulation of auditory cortex-dependent memory require activation of
mTORCH1 and possibly mTORCH2.” (Bockaert)

In the hippocampus: “When mTORCH1 was inhibited by chronic intracerebral

ventricular infusion of rapamycin the phosphorylation of mTOR substrates were
also inhibit(ed) as well as the learning-induced enhancement of protein
synthesis and the acquisition of learning.” (Garza-Lombo)

In the Amygdala: “It was found that rapamycin increases neuronal activity and
anxiety-related behavior, impairs both consolidation and reconsolidating of an
auditory fear memory, and produces impairment of IA (inhibitory avoidance)
memory.” (Garza-Lombo)

Inhibiting the mTOR system thus may adversely effect long term memory
 Sarcopenia

“Patients taking rapamycin for more than 6 months for the treatment
of renal cell carcinoma or paracrine neuroendocrine tumors
demonstrated an increase in sarcopenia.” (Walter and Cox)
 Tenet 4: Quality Control
● DNA: Four primary repair mechanisms

● Proteins: Four primary repair mechanisms

● Autophagy
 DNA repair
● Single strand break: 5 - 10,000 per cell, per day

● Double strand breaks: 10 per cell, per day

● Errors in substitutions, deletions, strand crossing and linking

● Inclusively up to 105 DNA errors per cell/ per day

 “Double-stranded breaks have been studied
extensively and are known to arise from ROS, gamma
irradiation, mechanical stress, defective telomere
processing, chemotherapeutic drugs and replication fork
collapse.” (Moehrle 2016)
 DNA Repair Mechanisms
Step one:

● PARPs: Poly ADP-ribose polymerases

● an enzyme that deconstructs the NAD molecule, pieces together
the ADP-riboses, and discards the nicotinamide

Step two:
1) Base excision repair (BER)
2) Nucleotide excision repair (NER)

3) Homogenous Recombination.
4) Non-homogenous recombination
How does DNA damage cause aging?
● DNA damage causes cells to become senescent or undergo
apoptosis

● Senescence causes chronic inflammation

● DNA damage causes cancer

“Insufficient DNA repair mechanisms are also linked with cardiovascular

disease and neurodegeneration. In specific, they play a role in such
diseases as Alzheimer’s, Huntington's, and Parkinson’s.” (Panish 2015)
Tenet 5: Security systems
Immune system and Inflammatory cascade

Three main issues central to aging

○ The body gets put in a chronic state of inflammation

○ Infection risk rises

○ Increase in cancers, especially in the cells that originate

from bone marrow.
 Chronic Inflammation
“The aging process is a chronic smoldering oxidative and inflammatory
stress that leads to the damage of cellular components, including proteins,
lipids, and DNA, contributing to the age-related decline of physiological
functions.” (Szarc 2015)

Inflammaging
Factors highly correlated to aging:
Interleukin-1, Interleukin-6, Interleukin-18, C-reactive protein (CRP), Tumor
Necrosis Factor-alpha (TNF-a), serum Amyloid, Soluble vascular cell
adhesion molecule-1 (sVCAM-1), and Monocyte chemoattractant
protein-1( MCP-1).
 Infection Risk

● Less robust cell production from declining stem cells

● Less efficacious macrophages, killer cells, and B cells

● Less response to vaccines with age

Tenet 6: Work Force
● Individual employees…...Translates into individual cells

● Several determinants require needs

Cell life length
Mobility...circulate vs stationary

● Fast turnover cells require

Optimized stem cells
High nutrient availability

● Long lived cells require optimized niche and nutrient delivery

 Individual cells
● Categorized by length of existence

○ Short lived…hours to days…

live hard and die fast
○ Medium….years….goldilock cell

○ Long….entire lifespan

Stem cells, neurons

Short- lived cells

● Originate from stem cells

● High nutrient requirement

● Don’t have waste or accumulation issues

● Don’t suffer DNA or protein error issues

 Middle-aged cells

● Replaced every few years depending on tissue type

● Tend to be stationary, thus niche dependent

● Some issues with waste accumulation and damages

Long-lived cells
● Cells not replaced

● Protein and DNA damage accumulation

● Accumulation of waste products

● Can become senescent

● Mitochondrial dysfunction

● Prone to Epigenetic remodeling

Stem Cells
● Absolute number of viable stem cells decline with age

● Differentiation potential declines

● Compromised DNA repair mechanisms

● Decline in Telomerase activity

● Mitochondria dysfunction

● Prone to Epigenetic remodeling

● Niche dependent
Tenet 7: Waste Management
● Glucose precipitates AGEs and rAGEs

● Autophagy creates Lipofuscin

● Oxygen causes Free Radical formation

Advanced Glycation End products
● Result of Glucose and oxidative stress

● Non-Enzymatic, multi-step reaction

● Creates AGEs, ALEs, or DNA-AGEs

Do?

● Create inflammatory response

● Sticks to almost everything made of collagen/ structural integrity

● Lose protein or DNA function

 Lipofuscin
● Byproduct of cellular recycling in the lysosomes

● Accumulations accurately age crustaceans

● Cause space occupying issues in long lived cells

● Prevents lysosomes from efficient recycling

● Get negative spiral of cell

Summation of Aging
● Company operating manual…DNA

● Energy Source…Mitochondria

● Pathways…Aging pathways, i.e. AMP Kinase, sirtuins, MTOR

● Quality control…DNA and Protein repair mechanisms

● Security…Immune system

● WorkForce…Individual cell requirements

● Waste Management…AGE’s, lipofuscin

Protocol Chart
 Astaxanthin

0.3.0.0.2.0.0
(Kaufmann Rating Number)

xanthophyll carotenoid
3,3’-dihydroxy-beta,beta-carotene-4,4’-
dione
 Astaxanthin
● Substance made by a unicellular
biflagellate, Haematococcus
pluvialis, under stressful
conditions
● Xanthophyll carotinoid
● Extremely red
● Responsible for most red found
in and around water...salmon,
crabs, lobster, roseate
spoonbills
Astaxanthin
0.3.0.0.2.0.0

Tenet #2: Mitochondria

Powerful free radical scavenger and anti-oxidant

“It is well worth mentioning that astaxanthin can act as a safeguard

against oxidative damage through various mechanisms, by quenching of
singlet oxygen, scavenging of radicals, inhibiting lipid preoccupation,
and regulating gene expression related to oxidative stress.” (Wu 2015)

Stimulates production of the endogenous antioxidant enzymes:

catalase, superoxide dismutase, and peroxidase.
 Astaxanthin
0.3.0.0.2.0.0

Tenet# 5 Security/ Immune system

Reduces activation of NF-Kb, which then suppresses

the production of IL-1B, IL-6 and TNF-a

Inhibits cyclooxygenase 2 (COX-2), prostaglandin E2,

and C-Reactive Protein (CRP)
 Astaxanthin
Vision
In 2009, Japanese researchers administered 6 mg of
astaxanthin daily to middle aged people (46-65) for one
month. Remarkably, 60% of the subjects had visual
improvements, especially in the categories of “difficulty to see
near objects,” “eye strain” and “blurred vision.” (Yuan and Kajita 2009)
Astaxanthin
Skin
In human cell lines, especially skin fibroblasts and
melanocytes, astaxanthin was shown to reduce
DNA damage that was precipitated by UVA
radiation.

In human skin studies:

Topical astaxanthin demonstrated improvements
in crows feet, age spot size, elasticity, skin
texture, and the moisture content of
corneocytes.
 Astaxanthin
Fitness
Prevents exercise related increases in Free
Radicals

Decreases DNA and protein damage with exertion

Increases exercise capacity

 Carnosine

0.3.0.0.0.0.3
KRN

L-histidine and B-alanine dipeptide

 Carnosine
0.3.0.0.0.0.3
● Present in all muscle

● Identified in 1900 by a Russian scientist, V.S. Gulewitch

● The amino acids come from our diet

● Amount on the body varies with age and gender

● Men have more than women

● Youth have more than the aged

 Cool experiment
Senescent human fibroblast cells were put into a bath of carnosine
Very quickly, the old cells exhibited a rejuvenated appearance. But they
didn’t just look younger, they acted younger….the senescent cells
reverted to juvenile phenotypes.

● If the carnosine was removed, the cells quickly became old again.

● If the carnosine was reintroduced, the transformation recurred.

● The cells in carnosine lived longer AND better

● Carnosine cells meanwhile had a 25% increase in the ability

to keep dividing.
Carnosine
0.3.0.0.0.0.3
Tenet #2 Mitochondria

Reduces oxidative damage

Improves endogenous anti-oxidants

Restores depleted levels of glutathione

Increases the basal levels of superoxide dismutase

Carnosine
0.3.0.0.0.0.3
Tenet #7 Waste management

Blocks AGE formation

May actually reverse AGE formation; acts as a
transglycating agent

“Carnosine was found to be effective already in the first step of

AGE formation as well as by reversing glycated protein through
a transglycation mechanism.” (Boldyrev 2013)
Hearing
Protects hearing from loud noises

Vision
Prevents presbyopia and cataract formation (via
carnosine eye drops)

Skin
Improves quality of skin
My subjects have experienced/ reported:

● Higher energy levels

● Decreased rates of infection/ URI’s

● Improved hair growth and color

● Improved skin quality

● Improved vision

● Weight loss

● Decreased joint pain/edema

● Improved sex life

How to choose a protocol
When choosing Regimen:

Age

Medical concerns

Goals
 Age Issues
Physiology different from the age of 35 to 90+

At 35: DNA damage, epigenetic modifications

At 45: Loss of endogenous antioxidants, Sirtuin levels decline, fat levels

increase, vision changes

At 90: No longer preventing…

 Medical Conditions
Diabetes...
More glucose and oxidative stress… increased AGEs…..leads to
elevated Blood Pressure, fragile skin, cataracts, early onset Metabolic
Disease

Do?
Focus on molecular agents that reduce glucose levels, reduce AGE
formation, and potentially lift the AGEs off of tissues
 Goals
● Depends on Age of initiation

● Lifestyle choices (smoking, alcohol, exercise,

overweight, etc)

● Degree of commitment to program

The Kaufmann Protocol: Why we Age and How to Stop it
Website: KaufmannProtocol.com

The App

Facebook: Sandra Kaufmann

Instagram: KaufmannAntiaging
Citations

● Barbieri, Elena, et al. "The pleiotropic effect of physical exercise on mitochondrial dynamics in aging
skeletal muscle." Oxidative medicine and cellular longevity 2015 (2015).
● Barzilai, Nir, et al. "The critical role of metabolic pathways in aging." Diabetes 61.6 (2012): 1315-1322.
● Carmona, Juan José, and Shaday Michan. "Biology of healthy aging and longevity." Rev Invest Clin
68.1 (2016): 7-16.
● Testa, Gabriella, et al. "Calorie restriction and dietary restriction mimetics: a strategy for improving
healthy aging and longevity." Current pharmaceutical design 20.18 (2014): 2950-2977.
● Li, Yuanyuan, Michael Daniel, and Trygve O. Tollefsbol. "Epigenetic regulation of caloric restriction in
aging." BMC medicine 9.1 (2011): 98.
● Moehrle, Bettina M., and Hartmut Geiger. "Aging of hematopoietic stem cells: DNA damage and
mutations?." Experimental Hematology 44.10 (2016): 895-901.
● Panish, U. et al. “Ultraviolet Radiation-Induced Skin Aging: The Role of DNA Damage and Oxidative
Stress in Epidermal Stem Cell Damage Mediated Skin Aging. Stem Cells International. 2016. Art ID#
7370642.
● vel Szi, Katarzyna Szarc, et al. "From inflammaging to healthy aging by dietary lifestyle choices: is
epigenetics the key to personalized nutrition?." Clinical epigenetics 7.1 (2015): 33.
● Bockaert, Joël, and Philippe Marin. "mTOR in brain physiology and pathologies." Physiological reviews 95.4 (2015):
1157-1187.
● Garza-Lombó, Carla, and María E. Gonsebatt. "Mammalian target of rapamycin: its role in early neural development and in
adult and aged brain function." Frontiers in cellular neuroscience 10 (2016): 157.
● Walters, Hannah, and Lynne Cox. "mTORC Inhibitors as Broad-Spectrum Therapeutics for Age-Related Diseases."
International journal of molecular sciences 19.8 (2018): 2325.
● eh, Po-Ting, et al. "Astaxanthin inhibits expression of retinal oxidative stress and inflammatory mediators in
streptozotocin-induced diabetic rats." PloS one 11.1 (2016): e0146438.
?