Summary Regenerative Medicine

Regenerative Medicine
Lecture 1 Introduction

Treating loss of organ function
- Transplantation
- Implantation of prosthetic device
o Occurs in bone
- Autografting
- Stem cells
- In vivo inducting of regeneration in the patient

Regenerative medicine starts with damage
- Acute
o Bone fracture
o Skin cut
o Myocardial infarction
- Chronic
o Non healing wounds (after diabetes for example)
o Hypertrophic scars
o Fibrosis
o Diabetes: hyperglycemia
o Smoking
o Hyperlipidaemia

Damage  repair  regeneration
- Regeneration means maturation of stem cells
- Regeneration: wound healing, rebuilding, training/maturation

Scaffolds can support the ECM
- Scaffold + stem cell keeps the cell in place

Regeneration occurs in lower organisms – absent in higher organisms.
- The differentiation blueprint is present in the stem cells
Salamanders: not low in the sense of regeneration. You can cut off a part of the salamander, but this
part will grow on again. After a few days there will be wound epithelium formed. This will
dedifferentiate (become simpler) and form a blastema. Forming a blastema is not possible in
mammals. This blastema will differentiate and there will be formed a new part again which will
follow the original pattern of the limb.

Patterning and molecular signaling is regulated by molecules
- Regulatory molecules  provide instructive signals for proliferation, differentiation and
function
- Adherence molecules  provide support, instructive signaling and sequestration of
regulatory molecules

Regeneration: the ability of an organism, organ or tissue to regain its original structure and function
after damage
Regenerative medicine
- Stimulates and supports the body’s own self-healing capacity
- Focused on repair, replacement or regeneration of cells, tissues and organs to restore
impaired function

, - Uses several converging technological approaches
- Re-educate the local microenvironment

Matrix
Cells
Adhesion molecules
Growth factors  secreted by focal adhesions




The RM trinity
- Outside the body: engineering
- Inside the body: regenerative medicine

Culturing cells
- Monolayer  adherent cells
- Suspension  non-adherent cells
- Three dimension  scaffolds, templates

Autologous cells
- ‘self’ of the patients
- Can be stem/progenitor cells or somatic cells
- Not rejected
- But the cells can be affected by the disease (are exhausted by repair)
- There is a limited, small number available
- They require individualized culture  expensive
- Can best be used for permanent structure repair
Allogeneic cells
- Best used for temporary repair
- Obtained from various tissues
o Cord blood, ESC, blood, bone marrow, fat
- Can be obtained in large numbers
o Bulk culture  uniform cell batches  less expensive
- But immunological rejection

Ideal: bio-engineered construct accepted without the need of immune suppression
- Thereby prevent risk of cancer
Second-best scenario
- Use immune privileged cells that have a low expression for T-cells
o Embryonic stem cells ESCs  express low MHC I levels and no MHC II levels

, - Cell banking: allows histocompatibility matching
- Use cells with immunoregulatory function
o Mesenchymal stem cells MSC  induce tolerance in grafts


Stem cells
- Unspecialized
- Self-renewal
- Differentiate into various functional phenotypes
- Bone marrow is the place for stem cells to become blood cells
- Embryonic stem cells  derived from inner cell mass of a blastocyst
o Unlimited self-renewal
o Multipotent  can become any cell in the body
o But they are genomically instable
o Tumorigenic  form teratomas (embryonic germ cell tumour)
o Ethical issues
o Important that multipotency is controlled
- Adult stem cells
o Mesenchymal stem cells (MSC): connective tissue stem cell
o Organ-specific stem cells in the gut, skin, bone marrow (all epithelia)
o Pluripotent
 Neural SC: neurons, astrocytes, oligodendrocytes
 Cardiac SC: cardiomyocytes, smooth muscle, endothelium
o Non tumorigenic
o More differentiated
o Patient derived (autologous)  well tolerated
o Disadvantage: disease can affect stem cells too

How to teach stem cells to become specialized cells?
- The substrate on which the cell grows determines what the cell becomes – does not need
growth factors for this
- If you give a cell minimal space it becomes a fat cell, but it you give it more space it becomes
an osteo cell
o Matter of surface area
o Topography (shape of surface)
 Adhesion and differentiation of satellite cells to skeletal muscle is promoted
by topography.
- Softness or stiffness of surface also determines the type of cell it becomes
o Soft  fat cells
o Stiff  bone cells
o But soft substrates sometimes prevents the cell from proliferating
- Soluble factors (paracrine)
o Growth factors
o Chemokines
o Cytokines
o Extracellular matrix components
- Forces (physical environment)
o 2D and 3D scaffolds
o Topography
o Mechanical forces, pressure, tension, shear stress

, Growth factors
- Mesenchymal-epithelial transition (MET)
o Driven by BMP7
o Fibroblast-like cells can differentiate
- Epithelial-mesenchymal transition (EMT)
o Driven by TGF-B
o De-differentiation
o During fibrosis
o A well organized cell becomes less organized
o Fully reversible by BMP7

Physical cues
- 2D versus 3D culturing
- Scaffold roughness/structure
o Porosity – pores are made by proteases
- Mechanical stress
o Shear stress in arteries caused by the tremendous high blood flow. Causes the
endothelial cells to be aligned
o Areas where there is not shear stress in atherosclerotic arteries, the endothelial cells
are not aligned
o So applying shear stress is good

Scaffolds
- Should not be permanent, otherwise they would give problems.
o Biodegradable without adverse inflammation
o Should be replaced by the physiological matrix
- Support biochemical features
o Promote adhesion, survival and function of cells
- Biomaterials/biopolymers
o Natural (collagen, fibronectin) or synthetic
o Mimic ECM
- Synthetic polymers can be processed at higher temperatures.
- Electrospinning: fibers are made out of a polymer solution and by a magnetic field the fibers
are collected.
- Making artificial scaffolds: Take scaffold material and dissolve it in organic solvent and ad
grains, sugar or sold and led the solvent evaporate. When you led that in water the sold or
sugar will solve and you remain with scaffold tissue, with interconnective pores (sponge).


Foreign body reaction FBR once the implant is in the body
- Implantation
o Activation of coagulation
o Coating of implant with serum proteins
- Activation of innate immune system
o Attracting macrophages
- Implant encapsulation by fibroblasts
- Degradation if possible by macrophages

Macrophage function in wound healing
- Matrix synthesis regulation by growth factors, cytokines and enzymes