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SMP - SDR > Lecture 21 & 22 - Stem Cells > Flashcards

Lecture 21 & 22 - Stem Cells Flashcards

1
Q

How have stem cells been used for decades now?

A

Bone marrow transplants

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2
Q

What are stem cells?

A

Cells that can give rise to cells like themselves or other cell types and present in constantly reviewing tissues:

  1. Lining of GIT
  2. Epidermis of skin
  3. Seminiferous epithelium of males
  4. Ovaries in females
  5. Marrow cavities of the long bones
  6. Neuronal stem cells in brain
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3
Q

What is the ultimate stem cell? What do we call these?

A

Single cell embryo or inner cell mass of an embryo = embryonic stem cells

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4
Q

Difference between embryonic and adult stem cells?

A
  1. Embryonic: derived from early embryos, aka the blastocyst (excluding the placenta) and capable of becoming a wide variety of adult cell types
  2. Adult: difficult to identify and isolate, also includes progenitor cells => all multipotent cells capable of becoming a limited number of cell types
  3. Adult cells induced to become pluripotent = iPSCs
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5
Q

2 types of embryonic stem cells?

A
  1. Totipotent: 1-8 cell stage embryo => can form all cells, including placental
  2. Pluripotent => can form all cells except the placenta
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6
Q

Can multipotent cells give rise to neurons?

A

NOPE

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7
Q

How can you construct a transgene?

A
  1. Use vector and restriction enzymes to insert cloned gene into multiple cloning site
  2. Cut gene out and run it on an agarose get to separate it
  3. Insert using a needle in an embryo held by a holding needle with microscope
  4. Deliver gene into pronucleus
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8
Q

Use of transgenic mouse models? Example?

A
  1. Every type of solid tumor cancer has been modeled using transgenic mice
  2. Hundreds of diseases have been modeled by transgenic mice (e.g., Huntington’s, Parkinson’s, many types of heart disease, skin diseases, etc.)

Example: used to understand virtually every signal involved in epidermis formation including hair follicle development, which has been directly used for skin grafts

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9
Q

2 methods to allow “3-parent babies”? Describe them. Purpose?

A
  1. Embryo repair: fertilizing mother and donor eggs => let pronuclei dissolve to allow chromosomes to mix => mother’s nucleus removed and put in place of the donor nucleus => embryo transferred to mother
  2. Egg repair: mother’s nucleus is isolated and donor nucleus is removed => mother’s nucleus is placed into donor egg => fertilization => embryo transferred to mother

Purpose = cut out risk of certain genetic disorders that the mother may pass on through her mitochondrial DNA

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10
Q

What is an alternative method to retrieving totipotent cells? Issue?

A

Preimplantation genetic diagnosis (PGD): take a couple blastomeres to test for genetic diseases like cystic fibrosis

BUT a lot of the time the embryo collapses when this is done

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11
Q

How do ESCs grow?

A

In colonies

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12
Q

When were ESCs first isolated?

A

1998 in IVF program

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13
Q

Can ESCs become a baby?

A

Not with current technologies, NOPE because they are pluripotent, not totipotent

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14
Q

Where does the controversy with ESCs arise from?

A

How ESCs are obtained => destroying the embryo

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15
Q

What could stem cells be used for?

A
  1. Cure diseases that are caused by a single gene malfunction or damage to a single cell type: bone marrow cells may be used to treat leukemia, nerve cells (motor neurons) to treat diseases such as amyotrophic lateral sclerosis (ALS) or Lou Gehrig’s disease, heart muscle cells repair heart attacks, retinal pigment epithelial cells to cure Stargardt’s macular dystrophy, and pancreatic cells to cure diabetes
  2. Drug development and toxicity tests: for example, if we coax stem cells to become liver cells, the effects of a particular drug on the liver can be predicted by its impact on these liver cells
  3. Assess the mechanisms by which specific genes alter development of particular organs - for example, if one wishes to determine how a gene produces colon cancer, one can put the cancer gene in embryonic stem cells, coax them to become colon cells, and determine the means by which the gene makes them cancerous
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16
Q

How does cloning work? What do we call this? Benefit over stem cells?

A

Somatic cell nuclear transfer:

Take sick patient somatic cell => remove nucleus (spindle apparatus from metaphase II) => form patient-derived cloned blastocyst => isolate ESCs => correct gene => expand and culture cells => differentiate them => inject mature corrected cells

Benefit: no need for immunosupressants

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17
Q

What signaling mechanisms regulate stem cell differentiation? Why is this important?

A

Unknown

We need to know these because:

  • we need the ability to correct the problem
  • if things go wrong, FDA will probably want to know if it was ES cell related or user related
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18
Q

Main issues when using iPSCs and ESCs?

A
  1. Risk of teratomas
  2. Immunogenicity (ESCs only)
  3. Ethical dilemmas (ESCs only)
  4. Full understanding of terminal differentiation (quality control)
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19
Q

What diseases have been cured using ESCs?

A

None yet

20
Q

Why does Zika cause microcephaly?

A

Underdevelopment of cerebral cortex

21
Q

2 theories to use adult stem cells as treatments? Pros and cons?

A
  1. Used adult stem cells from healthy donor (e.g. bone marrow transplants to cure leukemias)
    - pros: already done today
    - cons: need for immunosuppressants, limited donors and painful procedures
  2. Adult stem cells harvested from sick patient => dedifferentiation into ES like state => genetically modified to correct the disease => differentiate to form necessary cell => cells injected back into patient OR simply take sick adult stem cells and directly transdifferentiate them
    - pros: no need for immunosuppressants, no shortage of cells
    - cons: cannot form ALL cell types because only multipotent (e.g. derived myocytes often cause arrhythmias), not done in humans successfully
22
Q

Example of adult stem cells that do not differentiate well?

A

Mesenchymal stem cells when injected into the heart

23
Q

Easily obtained and purified:

  1. ESCs?
  2. ASCs?
A
  1. Yes

2. Yes/depends on cell type

24
Q

Maintain their multipotential lineage capacity?

  1. ESCs?
  2. ASCs?
A
  1. Yes

2. Limited

25
Q

Show directed differentiation?

  1. ESCs?
  2. ASCs?
A
  1. Yes

2. Yes

26
Q

Autologous to the patient?

  1. ESCs?
  2. ASCs?
A
  1. IPSCs: yes/ESCs: no (except if they are clones)

2. Yes/depends on cell type

27
Q

Possible to genetically modify cells?

  1. ESCs?
  2. ASCs?
A
  1. Yes

2. Depends on cell line

28
Q

Possible to expand cell numbers in culture?

  1. ESCs?
  2. ASCs?
A
  1. Yes

2. Limited

29
Q

Non-tumorigenic?

  1. ESCs?
  2. ASCs?
A
  1. Normally no but technologies are emerging

2. Yes

30
Q

How can stem cells be injected to the heart? Which one seems most promising?

A
  1. Direct injection in the heart: epicardial, endocardial
  2. Delivery via LAD or coronary sinus* (enabling cell delivery into myocardial areas subserved by occluded coronary vessels)
  3. IV
31
Q

Purpose of stem cells with a myocardial infarction?

A

Prevent infarction from growing because secrete paracrine factors, but avoid beating cells

32
Q

Could stem cells be used to cure Alzheimer’s?

A

Potentially

33
Q

4 genes necessary to create iPSCs?

A
  1. cMyc (dispensable)
  2. KLF4
  3. SOX2
  4. OCT4
34
Q

Problems with iPSCs?

A
  1. Retroviruses or adenoviruses are needed
  2. Genes may be oncogenes (cMyc: can be replaced by Lin 28)
  3. Genes are not controlled by endogenous promoters
  4. Genes may insert (randomly) into genome replacing a tumor suppressor
35
Q

Different ways that iPSCs can cause cancer?

A
  1. Can form teratoma
  2. Genes inserted could eventually cause cancer
  3. Where the genes insert could replace a tumor suppressor gene
36
Q

What 3 important cells can iPSCs be used to make?

A
  1. Neurons
  2. Beta-cells
  3. Cardiomyocytes
37
Q

4 similarities between iPSCs and hESCs?

A
  1. Both capable of going germline
  2. Both form teratomas
  3. Both use similar protocols for generating desired cell type
  4. Gene expression very similar*
38
Q

5 differences between iPSCs and hESCs?

A
  1. *Subtle differences in gene expression may be significant
  2. Chromosomal abnormalities in iPSCs (genetic instability)
  3. Methylation patterns of genes are different (iPSCs different from donor)
  4. Differentiation potential-neural differentiation
  5. iPSCs harbor a residual epigenetic memory, which restricts their fates (should use cells from same germ layer to generate them) - ESCs do not harbor this memory
39
Q

What is great about spermatogonial stem cells?

A

Pluripotent but do not cause teratomas

40
Q

Do spermatogonial stem cells need to 4 genes to be pluripotent?

A

NOPE, they’re endogenously expressed

41
Q

Do ESCs electrically connect with heart tissue?

A

YUP

42
Q

Do adult stem cells electrically connect with heart tissue?

A

NOPE

43
Q

2 advantages of cloning?

A
  1. DNA is identical to that of host so rejection is unlikely and immune responses of any type are minimized
  2. Recently thought to be perfected by caffeine because it releases calcium in a pulsatile manner
44
Q

Why is therapeutic cloning controversial?

A

The major societal concern reflects the possibility that blastocysts produced by nuclear transplantation will be placed in a woman’s womb and, theoretically, produce a human cloned child

45
Q

Are adult stem cells and progenitor cells the same?

A

NOPE, because progenitors can’t go backwards to become the original stem cell they came from

SMP - SDR (23 decks)

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