• Somatic Cells = all body cells that are not germ cells

• Germ Cells = sex cells ( ovary / egg , and sperm )

• Totipotent = everything , including extraembryonic ( placental ) tissue

• Pluripotent = everything except extraembryonic tissue

• Multipotent = everything in a particular class of cells. Hematopoietic ➡️ Any blood cell

• Differentiated = final form , specialized cell

Stem Cells Types

• Embryonic :

  • Pluripotent

  • derived from inner cell mass of mammalian blastocysts

  • proliferate infinitely while retaining pluripotency

  • Collection :

    1. Wait for morula to divide into blastocyst and form Inner Cell Mass ( ICM )

    2. Collect ICM cells ( pluripotent )

    3. Culture

• Fetal :

  • Multipotent

• Umbilical :

  • Multipotent

  • you bank them at birth and then don't have to worry about donor rejection problems

  • expensive over time

• Adult :

  • Multipotent

  • Hematopoietic ( HSCs )

    • blood regeneration

    • derived from bone marrow

    • you can induce these just like iPSCs

      • created by reprogramming mature RBCs with growth factors

      • provide patient-specific blood cells without the need for donors

  • Mesenchymal ( MSCs )

    • tissue regeneration

    • derived from bone marrow

  • Neural ( NSCs )

  • Epithelial

  • Muscle ( Satellite Cells )

• iPSCs / ES-like :

  • derived from somatic cells

  • created by nuclear reprogramming

  • pluripotent

  • gets rid of ethical issues

• ES Cell - Single Blasomere Removal :

  • remove 1 of the 8 cells in the morula

  • similar to IVF

  • advantage = no embryo is killed

iPSCs

• Production :

  • Reprogramming somatic cells to a pluripotent state by introducing specific transcription factors.

  • 1 factor is not enough , minimum of 4 are needed

    • in order to obtain ES-like cells , designated iPS cells for induced pluripotent stem cells

  • Yamanaka Factors :

    • Oct4 : Stem cell self-renewal gene.

    • Sox2 : Maintains self-renewal and survival.

    • c-Myc : Promotes proliferation.

    • Klf4 : Enhances cell survival.

  • Typically introduced via retroviral vectors

• Advantages :

  • Avoids ethical issues associated with embryonic stem cells.

  • Potential for patient-specific therapies.

• Disadvantages :

  • integration into the genome is issue when using retroviruses

• Morphology of iPSCs cells are indistinguishable from ES Cells

  • can't tell the difference

• transplanted anywhere other than the placenta ( blastocysts ) , they produce teratomas

• they showed it was technically possible in-vitro only to only need VPA and then 2 of the factors ( Oct4 and Sox2 )

  • suggests chemical reprogramming might be possible

  • but may provide cells for transplantation therapy and drug screening.

SCNT : Somatic Cell Nuclear Transfer

• Production :

  1. Remove nucleus from a egg / oocyte ( enucleation )

     • remove the polar body and chromosomes

  2. Remove the nucleus from a somatic cell.

  3. Transfer the somatic nucleus into the perivitelline space of enucleated egg.

  4. Oocyte and donor cell are fused and activated by an electric pulse to begin cell division

  5. Developed embryos transferred to surrogate females or grown in culture

• Applications :

  • Creation of embryonic stem cells matching the donor's DNA.

  • Potential for personalized regenerative medicine.

• Challenges :

  • Ethical concerns regarding embryo use.

  • Technical difficulties and low success rates.

Cloning

• How to Clone :

  1. Remove Nucleus from an egg , keeping only the cytoplasm

  2. Take some somatic cell ( fibroblast )

  3. Fuse them together by shocking them

  4. Grow and divide until blastocyst stage

  5. Implant blastocyst in uterus

• In Vitro Fertilization ( IVF ) :

  • NOT reproductive cloning

• Animal Models :

  • Mice = straight forward , no real problems

  • Rats = stop dividing 4 cells unless they are given a secondary media

  • Mouflon Sheep = first successful clone using DNA from dead cells

• To clone primates , you have to remove epigenetic marks

  • de-acetylation of DNA

Misc

• Stem cells let us clone things

• Clone = genetic copy with exact same DNA

• Monozygotic = single egg splits into identical twins

• Dizygotic = two separate eggs = not identical

• Inbred Strain = clone

• Outbred Strain = not a clone

• Everything morula ( 8 cells ) and smaller are totipotent

• Lineage Determination :

  • signaling and positioning in the yolk sack determine fate and differentiation

  • also timing of exposure to different growth factors

• To date , only Bone Marrow stem cells have been transferred effectively

• Somatic cells can be reprogrammed by transferring their nuclear content into oocytes or by fusion with ES cells

  • proves cells have some kind of factors that allow them to be totipotent

• Trophoblast organoids :

  • pseudo 3D micro environment