Tissue EngineeringDr. Siti Sunarintyas

Research on several different types of tissues and organs : Skin Blood Vessels Bone Muscle Nerves Liver Kidney etc. DefinitionTissue Engineering : the in vitro development of tissues or organs to replace the function of injured body parts.

How it works ??Three basic steps in tissue engineering :1. getting the base cells to work with2. putting the altered cells into a scaffold in order to incubate the cells3. putting the newly created cells or organ into use.

Step 1- Creating the MaterialsThe materials that are used for tissue engineering are actually living cells that were taken from somewhere else.These cells are then modified to whatever kind of cell is needed.There are 7 sources from where you can get these cells.

7 Sources of CellsAutologous- from the person himself.Allogenic- from a body with the same species.Xenogenic- from a different species.Syngenic- from genetically identical people.(Twins).Primary- from any organism.Secondary- from a cell bank.Stem cells- Undifferentiated cells.

Step 2-Bioreactors and ScaffoldsBioreactors are used to create physiological environments so that the cells can grow in a natural manner.Once the cells are obtained, then put into a scaffold to let them grow into the new forms.The problem of creating new organs : they would die before they placed into the organism.This problem has been solved with the developments in 3D printers, which allows for blood vessels to be put into the new organ.

8 Types of ScaffoldsNano fiber Self-AssemblyTextilesSolvent Casting & Particulate LeachingGas FoamingEmulsification/Freeze-dryingThermally Induced Phase SeparationElectrospinningCAD/CAM

Step 3- UsageThis step obviously is when the newly created cells are put into use.Whether they are being used to create a new organ via the 3D Organ Printer, or they are being used as new skin.

FutureTissue engineering is a very promising field in Biomedical engineering.It can solve many problems that people experience today.It will eventually continue to grow and make the need for a donor list obsolete as they will be able to just grow organs specifically for people.

Tissue Organization

Before a tissue can be developed in vitro, we must understand how tissues are organized. The basic tenant is :

all tissues are comprised ofseveral levels of structural hierarchy

These structural levels exist from the macroscopic level (centimeter range) to the molecular level (nanometer range)

Organization of the Tendon

Organization of the Kidney

Cellular CommunicationsSoluble Signals:small proteins (15-20 kDa) which are chemically stable with long half-lives (unless specifically degraded)growth factors, steroids, hormones, cytokines, chemokinesbind to membrane receptors usually with high affinity (low binding constants: 10-100 pM)can diffusion long distances

Cell-to-Cell Contact:some membrane receptors are adhesive moleculesadherent junctions and desmosomes other serve to create junctions between adjacent cells allowing for direct cytoplasmic communicationgap junctions1.5-2 nm diameter and only allow transport of small molecules ~1 kDa

Cell-ECM Interactions:ECM is multifunctional and also provides a substrate that cells can communicatesince cells synthesize the ECM, they can modify the ECM to elicit specific cellular responsesseveral specialized receptors that allow for cell-ECM interactionsintegrins, CD44, etc.also a mechanism by with cells respond to external stimuli (mechanical transducers)

Tissue Engineering ScaffoldsBiomaterial Scaffolds Materials:Polymeric : chitosan, alginate, hydrogels, fibresNatural : collagen, elastin, fibrinCeramic : calcium phosphate based for bone tissue engineeringpermanent versus resorbabledegradation typically by hydrolysismust match degradation rate with tissue growthChemical and Physical Modifications:attachment of growth factors, binding sites for integrins, etc.nanoscale physical features

Culturing of CellsTypes of Cell Culturemonolayer (adherent cells)suspension (non-adherent cells)three-dimensional (scaffolds or templates)

Sterilization Methodsultra-violet light, 70% ethanol, steam autoclave, gamma irradiation, ethylene oxide gasGrowth Conditionssimulate physiological environmentpH 7.4, 37C, 5% CO2, 95% relative humidityculture (growth) media replenished periodicallyCulture (Growth) Mediaappropriate chemical environmentpH, osmolality, ionic strength, buffering agentsappropriate nutritional environmentnutrients, amino acids, vitamins, minerals, growth factors, etc.

Cell Sources

primary cellstissue biopsy, low cellular yield , potential age-related problemspassaged cellsserial expansion of primary cells (can increase population by 100-1000X)tendency to either lose potency or de-differentiate with too many passagesstem cellsundifferentiated cellsself-renewal capability (unlimited?)can differentiate into functional cell typesvery rare

Stem CellsStem cells naturally exist in some tissues (especially those that rapidly proliferate or remodel) and are present in the circulation.

There are two predominant lineages of stem cells:mesenchymalgive rise to connective tissues (bone, cartilage, etc.)although found in some tissues, typically isolated from bone marrowhematopoieticgive rise to blood cells and lymphocytesisolated from bone marrow, blood (umbilical cord)

Stem cells are rare; bone marrow typically has:a single mesenchymal stem cell for every 1,000,000 myeloid cellsa single hematopoietic stem cell for every 100,000 myeloid cells

Stem Cells (Mesenchymal)

Stem Cells (Hematopoietic)

Colony-Forming Units (CFUs)

Bioreactorsa) Spinner Flask:semi-controlled fluid shearcan produce turbulent eddies which could be detrimental

b) Rotating Walllow shear stresses, high mass transfer ratecan balance forces to stimulate zero gravity

c) Hollow Fibreused to enhance mass transfer during the culture of highly metabolic cells

d) Perfusionmedia flows directly through construct

e) Controlled Mechanicsto apply physiological forces during culture

Tissue model construction

Tissue engineering requires three things:

CellsSignals

Scaffold

The scaffold refers to the matrix within the tissue model constructThe signals refer to molecular signaling molecules, also known as growth factors

Basic scaffold criteria:Portions need to be biodegradableUsually designed in the shape of the tissue product the researcher is working on