Ch 3 Stem cells and regenerative medicine Flashcards
4 classes of stem cells
stem cells are responsible for growth, maintenace and repair
embryonic ESC
Adult ASC
fetal/perinatal PSC
Induced pluripotent (IPSC) artificial
potenency types
Totipotent; differentiate into cells from each of the germ layer lineages (zygote)
pluripotent; more than one germ layer lineage but not the extraembryonic (ESC and PSC)
multipotent; restricted to the germ layer lineage from which they originate (ASC)
embryonic stem cells
What is? What problem?
harvested from the inner cell mass of a blastocyst, esulting in the destruction of the embryo
strong tendency to expand and differentiate in a dysregulated manner > resulting in the formation of teratomas
tumorigenic potential of undifferentiated embryonic stem cells has raised serious concerns
adult stem cells
What is? List 3 types of
source of replacement cells that drives tissue maintenance and repair
purified from dissociated tissue and expanded in culture to isolate and attain
hematopoietic stem cells, mesenchymal stem cells, and neural stem cells
Perinatal SC
Where from? 3
arvested from the umbilical cord, amniotic fluid, and fetal membranes
uperior expansion potential, increased plasticity, and may possess superior immune privilege
induced pluripotent SC
What is? How generated?
directly reprogramming adult somatic (i.e., terminally differentiated) cells. Epigenetic reprogramming alters or “resets” the gene expression
First induced pluripotent stem cells were generated using a retroviral vector
risk of causing insertional mutagenesis or oncogenic transformation
mesencymal SC
where located?
properties affected by wat variables?
what differntiate into? (3) COT
heterogeneous population of spindle-shaped, plastic adherent cells
reside in a perivascular location
> derived from either the bone marrow or adipose
cells within an individual colony of mesenchymal stem cells are morphologically diverse: small fast cells, large slow cells
capable of differentiating into cells such as osteoblasts, chondrocytes, and tenocytes
Properties of MSC? (4)
1) adhere to tissue culture plastic and exhibit a spindle-shaped
(2) form colonies of cells from single parent cells when cultured in low-density “clonal” cultures (CFU)
(3) express a specific set of cell surface marker proteins (exclude from hematopoietic lineages)
(4) possess the ability to differentiate into osteoblast, adipocytes, and chondrocytes (i.e., tri-lineage differentiation) using defined in vitro differentiation assays
msc mechanisams of action (4)
what anti-inflamm agents?
how immunosuppress?
terminally differentiating into various somatic cells “mesengenic process”
homing to sites of injury and contribute to tissue repair (migration and invasion) - paracrine effect.
immunosuppressive effects of MSC via:
- stimulation of conversion of M1 macrophages to M2 (anti-inflammatory)
- inhibition of monocyte development into dendritic cells
- inhibition of proliferation and cytotoxicity of NK cells
- suppression of T cell proliferation and modulation of T cell response
Volk et al
canine MSC require what to differentiate to bone in vitro
bone morphogenetic protein-2 (BMP-2)
important differences between mesenchymal stem cells isolated from various species,
MSC sources (5)
Autologus vs allogenic (donar)
bone marrow (dogs)
adipose tissue (dos)
synovium
muscle
teeth
btained from different donors and tissue sources exhibit substantial differences in regard to their properties and function, and the source of mesenchymal stem cells should be thoughtfully considered.
ability of MsC
differentiate into a variety of connective tissues (bone and cartilage) > suggests that they have great potential for orthopedic applications
enhance the inherent repair of tissues through secretion of trophic factors
how generate msc population
culture expanded
purification and in vitro expansion in culture to attain clinically relevant numbers of cells
pros: expand sufficient cell numbers in culture, the potential to cryopreserve large quantities of cells for readministration, the ability to accurately assess dosage and viability, and, most important, the ability to perform in vitro characterization assays prior to administration
unprocessed, minimally manipulated preparations of tissues that can be rapidly generated at the point of care
one marrow concentrate (BMC)141 or the stromal vascular fraction (SVF) of tissue digests
not possible to ascertain the absolute mesenchymal stem cell number, cell dose, or, in many cases, viability prior to clinical application
Licensing
by exposing them to inflammatory cytokines (like IFN-γ, TNF-α, and IL-1β) or other stimuli that mimic the inflammatory environment where they are intended to be used
MSC-based therapies hold promise for treating a wide range of diseases, but their efficacy can be variable. Licensing MSCs offers a potential strategy to improve their therapeutic efficacy and address some of the challenges associated with MSC-based therapies
hat are the options of MSC administration?
Intravascular infusion (cytotherapy) - less than 5%of cells successfully migrate to location of injury and survive
Direct implantation/injection (eg, seed onto a scaffold, intra-articular injection) ‘tissue engineering’
main limitations/concerns regarding MSC therapy (8)
Gaps in knowledge
Inherent variability in product
Access to facilities and expertise
Increasing federal oversight (FDA has banned in humans in many states)
Unknown safety - stimulate growth of neoplasms?
Increased risk of infection?
Administration of IV bolus has risk of microvascular embolisation and ischaemia
Current lack of strong evidence
2005, Volk et al…..
Evaluated osteogenic differentiation potential of marrow-derived MSC in 19 dogs - necessary to supply with BMP-2
2007 Black et al…
Evaluated effect of single intra-articular injection of stromal vascular fraction celss in 18 dogs with hip OA - significant improvement in lameness and function at 3m
Follow-up study - Single injection in 14 dogs with elbow OA - outcomes improved 30-40% comapred to baseline
Nishida et al…
Weekly injections of bone marrow derived MSC into spinal cord lesion causing lack of deep nociception. No complications but no improvement
Olsen 2019 VCOT
intravenously administered
allogeneic mesenchymal stem cells were well tolerated
by dogs with elbow osteoarthritis. While some subjective
outcome evaluations found significant improvements with
this treatment, a raft of different objective outcome measures
failed to demonstrate any significant improvement.
Pavarotti 2020 VCOT
lack of a placebo control group
Intra-articular injection of autologous adipose tissue may
therefore represent a promising treatment for osteoarthritis
in dogs. The one-step procedure appeared safe, minimally
invasive and simple, with early and sustainable clinical
effects up to day 180.
Scoping review of the use of mesenchymal stem
and stromal cell products in cats, Part 2: current scope
and efficacy
Tracy L. Webb 2024
discuss the variability in cell source, administration routes, dose range, and the lack of a control group
in most studies. Despite these significant limitations, few adverse events were reported across the studies
and several studies have demonstrated therapeutic effects supporting the need for further investigation.
Pueckler 2022 VCOT
future studies could track labelled MSC to prove efficacy
achilles tx
2 murine studies have provided data that licensing can enhance MSC therapy of an Achilles tendon defect and stimulate tenocyte expression of tendon-relevant genes.
depompeo 2020 Vet surg
Additional studies are required to determine the relative
concentrations of viable AD-MSC obtained via traditionally
and laparoscopically harvested falciform tissue.
Furthermore, complete characterization of the cells and evaluation
of maintenance of multipotency and viability through
passage numbers
Regenerative medicine
- interdisciplinary field
- focused on repair, replacement, or regeneration of cells, tissues, and organs
- reduction of inflammation and pain, improved healing of injured tissues
- goal of restoring structure and function
- including stem cell therapy, tissue engineering, gene therapy, and platelet-rich plasma or autologous conditioned sera.
quality of evidence
- lack of control groups make efficacy claims problematic (39.7% and 79% dogs in trials may experience placebo effect)
- many are case series, involve small numbers, insufficiently powered to make conclusions regarding efficacy.
- utilize subjective outcome measures that are more likely to be affected by bias
immunosuppressive effects of MSC via:
- stimulation of conversion of M1 macrophages to M2 (anti-inflammatory)
- inhibition of monocyte development into dendritic cells
- inhibition of proliferation and cytotoxicity of NK cells
- suppression of T cell proliferation and modulation of T cell response
published uses:
- semitendinosus myopathy – improved gait/lameness, gait analysis
- supraspinatus tendinopathy – MSC combined with PRP – gait analysis similar to contralateral limb in 88%, improved fibre pattern/echogenicity in 90.6%
- cranial cruciate ligament rupture – MSC +/- PRP > reduction of inflammation
12/13 subjective improvement, 9/13 intact CrCL on second look arthroscopy (initial partial)
osteoarthritis:
- elbow: subjective improvements in lameness, pain, function, owner-assessment scores (Olsen 2019)
- hip: subjective improvement in lameness, pain, ROM, objective improvements in limb function vs placebo
(variable effect across time points)
