Dental Stem Cells and Regenerative Approaches in Dentistry

• What are stem cells?

  - Undifferentiated cells

  - capable of self-renewal OR under certain physiological or experimental conditions, they can be induced to become tissue (or organ) specific cells with special functions
• What are the different types of stem cells?

  - embryonic (pluripotent)

  - non-embryonic adult

  - induced pluripotent
• Where do embryonic stem cells come from?

  inner cell mass of the blastocyst (early developing embryo)
• What is characteristic of embryonic stem cells (ESCs) in appearance?

  Particular cell surface markers

  (Oct4, Nanog...)
• How can you stimulate ESCs to differentiate into specialised cells in vitro (culturing conditions)?

  - Change the composition of the media (liquid cells are given for nourishment)

  - Alter surface of the dishes

  - Insert genes into cells
• what 3 layers does the embryonic stem cells in the inner mass of the bastocyst form?

  - ecotoderm

  - mesoderm

  - endoderm
• what does it mean when we say embryonic stem cells are pluripotent?

  Pluripotent cells can give rise to all of the cell types that make up the embryo.
• Where are adult stem cells found?

  Present in many organs and tissues & reside in stem cell niches
• when are adult stem cells activated?

  - they remain non-dividing for a long time

  - they are activated when needed to maintain tissue or disease/injury
• what does it mean when we say adult stem cells are Multipotent?


  they can develop into more than one cell type, but are more limited than pluripotent cells;
• What are iPSCs (induced pluripotent stem cells)?

  these are artifical stem cells:

  they are adult cells that have been genetically re-programmed to an embryonic cell-like state by introduction of certain embryonic genes into a somatic cell

  
• give examples of reprogramming factors that induce the formation of induced pluripotent cells

  cMyc, Oct4, Sox2, Klf4, Nanog, LIN28
• what are the different types of stem cells from the teeth?

  - SCAP - stem cells of apical papilla

  - DPSCs- from dental pulp

  - SHED - from deciduous teeth

  - PDLSCs - from PDL

  - GMSCs- from gingiva

  - from dental follicle

  
• What do the different dental stem cells look like under Alizarin red staining?

  (Alizarin red staining binds to the calcium)

  
• What are SHED (stem cells from human exfoliated dentition) cells?

  Stem cells from human exfoliated deciduous teeth:

  
  

  They are highly proliferative, clonogenic cells capable of differentiating into a variety of cell types including neural cells, adipocytes, and odontoblasts
• What is gingival tissue a source of?

  - Mesenchymal: gingival fibroblast stem cells

  - epithelial: gingival epithelial stem cells

  
• Why is the tooth a perfect source of stem cells?

  - it's accessible and would otherwise be discarded

  - it gives the possibility of authologus (obtained from the same individual) and/or allogenic (given) source of stem cells
• dental pulp stem cells

  
  

  describe dental pulp stem cells

  - high proliferating and colony forming

  - able to produce sporadic, but densely calcified nodules

  - capable of differentiating into adipocytes. osteoblasts, and endotheliocytes

  - have a broad capacity for differentiation and this might be due to their developmental origin as neural crest derived cells
• dental pulp stem cells

  
  

  what is the dual origin of dental pulp stem cells?

  pericytes (perivascular cells) & glial cells
• What are the 2 different methods to regenerate and repair dental tissues/teeth?

  1. Tissue repair & engineering-

  regenerating/repairing dental tissues

  
  

  2. Organ engineering-

  whole tooth replacement
• What should you always consider before repairing dental tissue?

  - understand the specificity of tissue and know the cellular events: presence/absence of cells

  
  

  - understand the response upon injury of the tissue: we need to know the capacity to limited repair and re-vascularisation

  
  

  - we need to know how to stimulate a repair response and what the limits are of the repair of certain tissues/organs
• How do the subsequent cellular events of the tooth vary depending on the extent of injury?

  1. if there is an early stage of injury/ a relatively low grade injury:

  - the pulp tissue will be upregulated with the secretory activity of the odontoblasts at the site of injury. we will have focal deposition of reactionary dentine at the pulp dentin border.

  ------> this is formed by the existing primary odontoblasts, this dentin has a regular tubular structure and tubular continuity with the existing dentine

  
  

  2. if there is a progression of a disease or intense injury:

  - there will be local death of odontoblasts at the site of the injury and inflammatory process will intensity

  ------> there will be a deposition of reparative dentine at the pulp dentine border

  
• By what method can you recruit cells to become odontoblasts?

  1. inside the dental pulp (wound healing?)

  OR

  2. outside the dental pulp (engineering?)
• How do you switch on the natural wound healing pathway?

  Wnt signalling is activated by wounding & participates in every subsequent stage of the healing process from the control of inflammation & programmed cell death, to the mobilisation of stem cell reservoirs within the wound site

  
• What does the activation of Wnt signalling as an immediate early response to tissue damage provide?

  A potential route for enhancing natural repair by overstimulating the pathway

  - this allows us to direct and tailor the repair
• What are the different methods of periodontal repair & regeneration?

  1. cell base theory

  (repair via stem cell transplantation)

  
  

  2. scaffold fabrication techniques

  (scaffolds serve as a temporary framework to support cell growth, proliferation, and tissue formation)

  ---> there are various fabrication methods to create scaffolds with specific characteristics tailored to the requirements of the target tissue. For example: 3D printing, Electrospinning, Hydrogel-based Techniques...

  
  

  3. gene therapy

  ( genetic material is introduced into cells to treat or prevent diseases)

  e.g. Gene Replacement Therapy, Gene Editing, Gene Regulation Therapy...

  
• what is a bioengineered tooth?

  artificial genetically engineered tooth
• Describe the process of bioengineering teeth.

  - Culture cells

  - Transplantation

  - Bioengineered tooth primordia (early) development

  - Bioengineered tooth

  
• Outline the experimental process of trying to bioengineer human teeth.

  - Human adult gingival epithelial cells (HGEP cells) were grown as clusters & recombined with mouse embryonal tooth mesenchyme (MM)

  - these were cultured in collagen and implanted in the mice

  - after 6 weeks, microCT scans were done and histology analysis.

  
  

  results: tooth grew and all the tooth's components were present.

  -> the tooth DID contain human cells

  
• How long does mouse tooth development take compared to human tooth development?

  Mouse tooth development: 5 days

  Human tooth development: 42 days (8 times slower than mouse)
• What are the clinical uses of oral mucosa epithelial cells?

  They're used in corneal reconstruction

  
• are stem cells grow/expanded in vitro (outside living organism) the same as the ones in vivo (inside living organism)?


  In vivo:

  stem cells are influenced by their microenvironment, known as the stem cell niche, which provides signals and cues that regulate their behavior, including self-renewal and differentiation. This niche includes neighboring cells, extracellular matrix components, and soluble factors.

  
  

  In vitro:

  culture conditions attempt to mimic aspects of the stem cell niche, but they may not fully replicate the complex interactions found in vivo. Consequently, stem cells cultured in vitro may behave differently in terms of proliferation rate, differentiation potential, and other characteristics compared to their counterparts in vivo.