ADULT STEM CELLS

Question 1

Give me an introduction about the adult stem cells

Answer 1

• characteristics about the adult stem cells
• where can we find them
• stem cell niche

Question 2

Hematopoietic stem cells

Answer 2

Basic definition of hematopoietic stem cells
Main sources: bone marrow, peripheral blood and cord blood (differences)
Isolation: differences of markers between mHSCs and hHSCs
Manipulation: long-term culture assays, assays in mouse (including serial transplantation assays) and xenotransplantation assays in humans
Main clinical application: HSC transplantation
HSC niche: different cell types with which the HSCs interact

Question 3

Mesenchymal stromal/stem cells

Answer 3

Explain why we talk about stromal/stem cells: not all the MSCs are able to differentiate into the mesodermal lineages, osteocytes, adipocytes and condrocytes.
They explain a strong variability in their phenotype, proliferation capacity and expression of cell-surface markers: intra-population heterogeneity and inter-population heterogeneity.
Discovery by Friedenstein
Isolation (from bone marrow aspirates or lipoaspirates) and selection (adherence to substrate, colony-forming units, differentiation into the mesenchymal lineages)
Roles in vivo: HSC niche maintenance and immunomodulatory role
Paper of Paolo Bianco, 2007: they are not dispersed in the bone marrow stroma but they are located in an adventitial location + CD146 as marker
Clinical applications: senescence during long-term culture, decline in the differentiation potential, shortening of the telomeres, morphological alterations, increased probability of malignant transformation, they are cultured in 10% heatin-activated FBS (FBS proteins can be immunogenic in vivo).
Paradigms of exploiting MSCs for their therapeutic potential

Question 4

Epithelial stem cells

Answer 4

Description of the epithelia and how do they form
Epithelial stem cells are specified during development and controlled by epithelial-mesenchymal interactions
Role in vivo: maintenance of the tissue homeostasis if activated by physiological stimuli or tissue repair and regeneration if activated by pathological stimuli (injury, trauma) > from G0 to transiently amplifying cells
Identification through pulse/chase experiments

Question 5

Intestinal stem cells

Answer 5

How the intestinal epithelium is made
Villi-cripts units
How we can find the stem cells and which cells they can generate
Infection by Eligsomoides polygyrus to study how they can shift from a tissue-maintenance program to a tissue-repair program

Question 6

Skin stem cells

Answer 6

Pilosebaceous units
Interfollicular epidermis: squamous stratified epithelium which a basal layer of proliferative cells that can differentiate and continuously replenish the suprabasal layers > maintenance of the homeostasis.
During an injury, their activity increases significantly enabling the wound healing.
Multiple locations: bulge stem cells and hair germ stem cells with their continuous cycle of regression and regeneration > best model to study how the niche regulates stem cell behavior and function.
Firstly described in 70s by Reinwhald and Green > culture conditions to grow human skin stem cells in vitro.
Then, used to propagate human keratinocytes from burned patients and to graft as sheets of autologous cultured cells.

Question 7

Corneal stem cells

Answer 7

Corneal epithelium
Where do the corneal stem cells reside? Limbal region
Quiescent state, activated by stimuli
Importance of the niche: they receive a lot of stimuli from different cell types and the ECM > nutrients, oxygen, mechanical cues, biochemical factors
Difference between physiological and pathological activation: they lose plasticity
Ocular surface pathologies and damages that can affect the niche > invasion by the conjunctival cells and corneal blindness
Pellegrini and De Luca studies on ocular burn: approval of Holoclar (first European stem-cell based medicinal product)

Question 8

Neuronal stem cells

Answer 8

Dogma of Ramon Cajal
1960: identification of thymidine-labeled neurons (proliferating neurons) under the ependymal layer of the ventricular wall of the brain
2004: first paper reporting neural stem cells in the SVZ and SGZ of the adult brain > adult neurogenesis
Neurosphere assays
In vivo evidence (demonstrated only in rodents, in humans only indirect evidence of the presence of adult neurogenesis from post-mortem studies)
Rodent model architecture of SVZ with different cell types and markers
Characteristics of the SVZ in human
Adult neurogenesis in pathological conditions: epilepsy, cerebral ischemia and neurodegenerative diseases