tissue engineering & stem cell therapy

Question 1

what problems may there be for craniofacial reconstruction?

Answer 1

Insufficient host tissue for adequate repair of defect

Donor site morbidity

If indogenous disease - may not be suitable

Question 2

what does craniofacial reconstruction and repair require a combination of altering

Answer 2

Skin

Soft tissue

Cartilage

Bone

Neural

Vascular regeneration

Question 3

what is the simplifed steps of the routine exercise in craniofacial reconstruction

Answer 3

1. Clear pathological site to remove damage and pathological tissue
2. Use natural bone mineral tissues to graft on wound site
3. After surgery

• Can do dental implants

Question 4

What are the three approaches in tissue engineering?

Answer 4

conduction

induction

cell transplantation

Question 5

What is conduction

Answer 5

• Guided tissue regeneration (GTR).
• Guided bone regeneration (GBR).
• using a 3D scaffold - usually nanomaterials
  
  • on which cells can attach, migrate, proliferate and differentiate
  • promotes proliferation
• the scaffold can also function as a barrier to promote repopulation of the defect site with a selected cell population

Question 6

what is induction

Answer 6

• using bioactive signals to induce cell migration and control cell differentiation and behaviour
• delivered in form of proteins
  
  • cytokines
  • growth factors
• can be delivered using gene therapy
  
  • transfer of specific genetic information to host cells

Question 7

what are the steps of cell transplantation

Answer 7

1. biopy from donour source (allogenic) or from patient’s own tissue (autologous) is obtained
2. isolate and expand donor cells in vitro
3. usually seeding cells onto scaffold such as fibre mesh or hydrogel
4. cells attach to scaffold and proliferate
5. formation of new tissue which may be implanted into defect site of the patient

Question 8

where it regulation crucial during cell transplantation?

Answer 8

it is important to precisely control the lineage specific differentiation of the transplanted stem cells

Question 9

which of the three approaches in tissue engineering lead to the most successful reconstruction?

Answer 9

all three factors in combination

Question 10

steps of making mucosa :

Answer 10

1. Small piece of tissue taken

• Separated into its components
• Then into individual cells

2. These are grown in plastic dishes

• In organotypic air-nutrient fluid system
• One cell can divide to make several thousand cells

3. Collagen matrix containing fibroblasts is made
4. Epithelial cells are plated onto it and the construct raised to the air-liquid interface

= organotypic culture forms tough compound mucosa

• is elastic and strong

Question 11

how does skin reconstruction occur naturally?

Answer 11

• basal cells do symmetrical cell division forming identical daughter cells
• cells begin to slightly tilt
• triggers differentiation
• one cell will stay in basal layer
• one cell will initate differentiation outward
• eventually cells lose nuclei and flake off surface
• skin tissue is replaced

Question 12

what cellular requirements are there for tissue regeneration?

Answer 12

1. directed cell migration

• ideal migration from surrounding tissues rather than transplant into centre of the wound site
  
  • pathological environment and toxic factors in wound site could kill the stem cells

2. organised cell division

• to achieve sufficient cell numbers
• rate
  
  • determines effectiveness
• orientation
  
  • determines daughter cell fate

3. targeted cell differentiation
   * how we control the lineage specific differentiation of the stem cells once they are transplanted into the recipient

Question 13

what are totipotent stem cells

Answer 13

Produced from fusion of egg and sperm cell

Also cells produced by the first few divisions of the fertilised egg

These cells can differentiate into embryonic and extraembryonic cell types

Question 14

what are pluripotent stem cells

Answer 14

Stem cells are the descendants of totipotent cells

can differentiate into cells derived from any of the 3 germ layers

• Ectoderm (outer layer)
• Mesoderm (middle layer)
• Endoderm (inner layer)

Question 15

what are multipotent stem cells?

Answer 15

Stem cells can produce only cells of closely related family of cells

E.g. haematopoietic stem cells differentiate into red blood cells, white blood cells, platelets, etc.)

Question 16

what are unipotent stem cells

Answer 16

Cells can only produce one cell type

Have property of self-renewal

• Distinguishes from non-stem cells
• Becomes a terminally differentiated cell

Question 17

what are the differences between embryonic stem cells and adult stem cells?

Answer 17

embryonic stem cells –

• . Pluripotent.
• . Drug screening.
• . Replicable in labs.
• . Limited tests on human (Geron – later BioTime, Califonia USA)
• Ethical problems

adult stem cells –

• . Unipotent.
• . Exist in limited organs.
• Limited sources to gain them from
• . Technically difficult (DPSCs, oral mucosal progenitor cells).
• . Can use own stem cells, but not suitable for genetic disorders, (Huntington’s).
• . Well used clinically.

Question 18

how are embryonic stem cells obtained?

Answer 18

From blastocysts.

• fertilized embryos
• (ethical problem)

Somatic Cell Nuclear Transfer (therapeutic cloning).

• unfertilized embryos

Induced Pluripotent Stem Cells

• reprogramme somatic cells into stem cells (tumorgenesis)
• By reprogramming - risk of inducing tumour into the genome

Question 19

what determines the cell fate after transplantation?

Answer 19

Hematopoietic system

• No intrinsic differences after stem cell has been grafting
• Environmental differences
  
  • Depends on where the stem cell is injected into
  • E.g. Liver tissue - affected by liver environment
  • = Becomes liver progenitor cells

Nervous systems

• Intrinsic growth factor differences to determine cell fate after transplantation
• Gradient of growth factor of spinal cord tissue
• E.g.
  
  • Influences what the stem cell will differentiate into
  • BMP - astrocytes, interneurons
  • Shh - oligodendrocytes, motors neurons

Question 20

what are the advantages of using dental pulp stem cells?

Answer 20

• easy access to the cells and produces low morbidity
  
  • wisdom teeth extraction
  • no ethical issues
• have extensive differentiation ability
  
  • many possible clincial applications
• demonstrated interactivity with biomaterials
  
  • ideal for tissue reconstruction

Question 21

should we use mixed population or single cell derived dental pulp cells?

what are the advantages of a mixed population?

what are the disadvantages of a mixed population?

Answer 21

Heterogenous

• Variety of growth factors, cytokines and chemokines released from stem cells
• Support each other in their roles
• Aiding survival
• Disadvantages
  
  • Lot of unknown factors we cannot define

Question 22

should we use mixed population or single cell derived dental pulp cells?

what are the advantages of a single cell population?

what are the disadvantages of a single cell population?

Answer 22

Has purity of stem cell population

• Need to know all elements of what we transplant into human body for clinical use - therefore very useful
• Can precisely regulate and characterise

Disadvantages

• Does not have supportive tissue to work together
• Harder to survive pathological situations after transplantation

Question 23

what are disadvantages of tissue engineering?

Answer 23

• Stem cells from blastocysts
  
  • Destroy fertilised eggs
• Stem cells by somatic cell nuclear transfer
  
  • Require eggs and still involve cloning
• Stem cells by reprogramme somatic cells
  
  • Retroviruses used to insert genes can cause tumour