1. The Effects of Aging on Human Bone Marrow Stromal Cells
Alec Richardson
3/17/09

2. What is Aging? Changes in an organism over time
Can be physiological or behavioral or both
Key component is time – Aging seen as the result of numerous changes over time
Viewed as a decline in function – perhaps better to think of it as a change in function
Affects people differently and the same (m/f)
Different vulnerabilities to disease
Aging results in a higher probability of death
No proven biological basis

3. Aging Theories
Inside: mechanisms that protect against disease become intrinsically disrupted
Changes in transcription/translation, increased cell death, disruption of organ function
Outside: environment affects genomic integrity or other physiologic parameters
Results in similar changes that lead to vulnerability and malfunction
Probably a mixture of both inside and outside factors
Statistics: Billions of bodily events occur every second
These events can be harmless or harmful based on the genetic makeup of the organism and ‘luck’
e.g. tumor suppression mechanisms
As the number of events accumulates, the probability of mistakes and problems increases

4. Aging in Bone Focus on one particular organ
Intimate relationship between bone and other organs
How does bone change over time in humans?
Disease, disruption in synergy between other organs, changes in cell #, vulnerability to fracture, load bearing capacity
Production of cells, support, storage, detox
Mineralization, stem cells, hormones, immune system, ions, homeostasis

5. Aging Models Models of accelerated aging
E.g. w/t mice
Differences between young and old organisms
Artificially aging of organisms

6. Human Bone Marrow Stromal Cells
Precursor cells in bone are necessary to replenish supplies of vital osteogenic cell types such as osteoblasts
Hypothesis: The aging process results in a decrease in the number of HBMSCs and/or a decrease in the ability of such cells to differentiate and produce mineralization.
If true, this theory could explain why aging results in vulnerability to fracture and diseases such as osteoporosis in older populations

7. Possible mechanisms of age-related bone loss
BMPs
Collagen
Osteocalcin
Osteopontin
Collagenase
Other NCPs
Mineralization
Histone
Collagen
TGFβ1
Osteopontin
Alk Phos
BSP
Collagen
IGF-I, PGE2
Vitamin D
Steroids
BMPs
TGFβ
Stem
Cell
Mesenchymal
Stem Cell
Osteoprogenitor
Pre-osteoblast
Osteoblast
BMPs
PTH
Runx2
Osx
Adipocyte
Chondrocytes
Myocytes
Fibroblasts
Osteoblast senescence
Bone-
Lining
cell
Osteocyte
MSC senescence
Lineage switching
Transdifferentiation

8. Experimental Model CFU-F Assay
Previous papers show CFU-Fs to contain osteoprogenitor cells
Human donors
Young=Age 34-49, Old=Age 70-85
Stains
Crystal Violet, Alkaline Phosphatase, Alizarin Red
14 days, 4 re-feedings
Differentiation Factors added to all plates
Β-Glycerophosphate, Ascorbic Acid, Dexamethasone
Seeding density for all plates: 40 cells/cm^2
Image J software – used for all stains except Aliz. Red

9. Crystal Violet (+ D.F.)
Age 77
Age 43

10. Alizarin Red
Age 77
Age 43

11. Alkaline Phosphatase
Age 77
Age 43

16. Future Directions Compare passaged and un-passaged cells
Ficoll gradient to isolate mononuclear cells
Passage to P1 or P2