Tissue Engineering - Quiz 1

  1. What stage are toitopotent stem cells present?
    eight-cell stage of the embryo
  2. Stem cells are distinguished from other cell types by two important characteristics:
    • They are unspecialized cells capable of renewing themselves through cell division
    • Under certain conditions, they can be induced to become tissue- or organ-specific cells with special functions.
  3. What are unique properties of all stem cells?
    • Capable of dividing and renewing themselves for long periods
    • Unspecialized
    • Can give rise to specialized cell types.
  4. What is an iPSC?
    Induced pluripotent stem cell.

    Adult (somatic) cells that have been genetically reprogrammed to an embryonic stem cell-like state by being forced to express genes and factor important for maintaining the defining properties of embryonic stem cells.
  5. What is a teratoma?
    A multi-layered benign tumor that grows from pluripotent cells.
  6. What are the two cell types and the associated sources.
    Primary (direct from tissue) and cell line (immortalized or tumor derived)

    Once cells are modified in any manner, they are considered cell lines.
  7. What are the advantages and disadvantages of primary cells?
    Advantages - protentially retain all differentiated function

    Disadvantages - difficult to maintain viability and differentiated function; may not proliferate

    Once cells are modified in any manner, they are considered cell lines.
  8. What are the advantages and disadvantages of cell lines?
    Advantages: Relatively easy to maintain; prolifer readily for many generations

    Disadvantages: Do not necessarily retain function of normal cells; safety concerns

    Once cells are modified in any manner, they are considered cell lines.
  9. What are the three cell sources?
    • Autologous - patient source
    • Allogeneic - other human source
    • Xenogeneic - other animal source
  10. What are some advantages and disadvantages to autologous cells?
    No immune rejection

    • Difficulty/inabilty to grow up cells
    • Impractical for certain tissues
  11. What are some advantages and disadvantages to allogeneic cells?
    Human source

    • Scarce for some tissues
    • Immune incompatibility
    • Transfer adventitious agents
  12. What are some advantages and disadvantages to xenogeneic cells?
    Readily available

    • Immune incompatibility
    • Function not necessarily identical
    • Transfer adventitious agents
  13. What are the two responses to foreign materials? What are the acute and late responses?
    • Toxic response
    • -Acute
    • --Hemolysis
    • --Temperature increase
    • --Inflammation
    • --Anaphylactic shock
    • --Tissue damage
    • -Late
    • --Malignant neoplastic formation
    • --delayed allergy
    • --teratogenesis
    • --tissue necrosis

    • Foreign body response
    • -Acute
    • --Complement activation
    • --Blood coagulation
    • --Thrombus formation
    • --Phagocytosis
    • -Late
    • --Encapsulation
    • --Neointima formation
    • --calcification
    • --Hyperplasia
    • --Atrophy
  14. What is mixed chimerism? What are the advantages?
    Mixed chimerism involves giving the patient antibodies from the donor to prevent an immune reaction.

    Usually chimerism develops accidentally in transplant patients when immature cells from the donor organ are incorporated into the recipient's blood.

    This triggers the production of white blood cells matching the donor organ, thus protecting the organ from being destroyed
  15. What are some advantages of established cell lines?
    • Many kinds of cell lines
    • Generally easy to grow and manipulate
    • Proliferate indefinitely

    Disadvantages include ploidy problems; loss of biochemical properties of parent tissue.
  16. Describe Primary cell cultures.
    • Cells derived directly from a tissue
    • Limited growth potential
    • Limited Life span
    • Cells are used within days from culture
    • May give rise to a cell strain or be immortalized

    Cell strain - a lineage of cells originating from one primary culture
  17. Describe transformed cells (cell lines)
    • Derived from tumors
    • Arise spontaneously in culture
    • Primary cells transformed with viruses, radiation etc
    • Grow indefinitely in culture
    • Usually unstable complement of chromosomes
    • Cell line - cells derived from a single parental transformed cell
  18. (T/F) Cells must be grown at the biological pH
  19. (T/F) The medium is selected depending on the cell type.
  20. (T/F) The best growth is not always the fastest and the optimized media for the experiment must be used.
  21. (T/F) CO2 concentration is important for the bicarbonate buffering system in the blood.

    • Too high in CO2 will change the pH of the cell culture.
    • Too much oxygen may overwhelm the cells (oxidative stress)
  22. List some pros and cons of using antibotics in cell cultures
    • Reduce freqnecy of contamination
    • Encourage development of antibiotic resistant strains
    • Hide low level cryptic contaminations
    • Anti-metabolic effect
    • Encourage poor aseptic technique
  23. Trypsin is a highly active, relatively non-specific, cheap protease derived from pancreas. List some uses in cell cultures.
    • Used in concentration of 0.05 to 0.25%
    • Cleaves proteins on the cell surface and extracellular matrix
    • Detaches cells
    • Inactivated by serum or soybean trypsin inhibitor
    • Cells needs to be washed withPBS (phosphate buffered saline) before trypsin treatment to remove serumwhich is a trypsin inhibitor
    • Over-trypsinization causes cell injury
    • Trypsin has to be deactivated after use
  24. What are some sources of contamination in cell cultures?
    • Bacteria, mold, yeast
    • Mycoplasma
    • Cross contamination
  25. Describe totipotent stem cells.
    Totipotent (a.k.a omnipotent) stem cells can differentiate into embryonic and extraembryonic cell types. Such cells can construct a complete, viable organism.

    These cells are produced from the fusion of an egg and sperm cell. Cells produced by the first few divisions of the fertilized egg are also totipotent.
  26. Describe pluripotent stem cells.
    Pluripotent stem cells are the descendants of totipotent cells and can differentiate into nearly all cells, i.e. cells derived from any of the three germ layers.
  27. Describe multipotent stem cells.
    Multipotent stem cells can differentiate into a number of cells, but only those of a closely related family of cells.
  28. Describe oligopotent stem cells.
    Oligopotent stem cells can differentiate into only a few cells, such as lymphoid or myeloid stem cells.
  29. Describe unipotent stem cells.
    Unipotent cells can produce only one cell type, their own, but have the property of self-renewal, which distinguishes them from non-stem cells (e.g., muscle stem cells).
  30. What are the CD59 and decay accelerating factor (DAF)'s functions?
    They mark the cells for destruction.

    By inhibiting this protein, the prevention of a response towards foreign materials may be prevented.
  31. (T/F) Pigs are considered clean and have a relatively low risk of viral transmission.
  32. What is the role of cell culture media?
    • Maintain pH
    • Provide essential nutrients
    • Provide energy source
    • Maintain osmolarity
  33. Why do adult primary cells have a limited lifespan?
    Reduction of telomerase activity which then shortens the length of telomers
  34. (T/F) Serum is well defined and the most widely used as an additive to the culture media.
    It is not well defined and contains insulin and serotonin. It does not inhibit trypsin
  35. Rmbryonic stem cells are _____ stem cells.
    multipotent stem cells.
  36. What are the optimal conditions for cell cultures?
    Always keep the cells in low density and keep them attached to each other.
  37. (T/F) Adenovirus can infect both dividing and non-dividing cells, which express CAR.
  38. (T/F) Adeno associate virus is larger than adenovirus.
  39. Which of the following many be used to deliver a gene to terminally differentiated neurons?
    Adeno-associated virus
    All of them.
  40. (T/F) Under certain physiological or experimental conditions, stem cells can be induced to become cells with special functions.
  41. What are 4 gene delivery methods?
    • A gun to shoot gold particples with DNA
    • Electroporation
    • Biological vehicles
    • Liposomes
  42. What is transfection?
    • Introduction of foreign material into eukaryotic cells
    • -overexpression
    • -Downregulation (RNAi, morpholino)
    • -Protein production (baculovirus, adenovirus)
  43. What are some non-viral methods of DNA delivery?
    • Chemical (such as calcium)
    • Liposomes (surround etc)
    • Electroporation (change electric field outisde the cell membrane to create pores in the membrane)
  44. What are the two transfection categories?
    • Transient transfection - DNA not inserted in host genome (temporary gene expression, DNA lost after cell division)
    • Stable transfection - DNA inserted into host genome (Selection pressure to maintain foeign DNA)
  45. What are some methods of gene delivery?
    • Injection of naked DNA into tumor by simple needle and syringe
    • DNA coated on the surface of gold pellets which are air-propelled into the epidermis (gene gun); mainly non applicable to cancer
    • DNA transfer by liposomes (intravascular, intratracheal, intraperitoneal or intracolonic routes)
    • Biological vehicles (vectors) such as ciruses and bacteria
  46. (T/F) Vector gene delivery should be able to target specific cell types.
  47. Place the following gene delivery methods in decending order for the amount of information they can carry.
    Naked DNA/Liposomes
    • Naked DNA/Liposomes - unlimited
    • HISV-1 40 to 150kb
    • Adenovirus - 30kb
    • Retrovirus - 8kb
    • Adeno-associated - 4kb
  48. What are some advantages and disadvantages of the non-viral DNA carrier cationic liposomes?
    Positively charge lipids interact with negatively charged DNA

    • Advantages
    • Stable complex
    • Can carry large sized DNA
    • Can carry to specific cells
    • Does not induce immunological reactions

    • Disadvantages
    • Low transfection efficiency
    • Transient expression
    • Inhibited by serum
    • Some cell toxicity
  49. What are the main limitations of retrovirus and lentivirus (ie HIV)?
    Only transduces dividing cells, integration might induce creation of cancer

    Main advantage persistent gene transfer in dividing cells
  50. (T/F) Retroviral vectors are able to infect dividing cells only

    In dividing cells, nuclear membranes are broken down so viral genome can enter and integrate into the chromosome

  51. (T/F) Lentiviral vectors are retroviruses that can infect both dividing and non-dividing cells.

    Although retroviruses can infect only dividing cells, lentivuses are apparently the exception.
  52. What are the advantages of lentiviral vectors?
    • Mobilization of a vector designed to inhibit or prevent HIV
    • Replication or pathogenesis has been argued to enhance therapeutic effect

    • Disadvantage
    • Vector spread beyond the intended target tissue may have safety consequences
    • Co-packaging of wt-type HIV RNA and vector RNA may result in recombination
  53. (T/F) Adenoviruses can infect both dividing and non-dividing cells (CAR is necessary for inection)
  54. What gene delivery vectors require CAR for attachement and delivery? Can this virus infect both dividing and non-dividing cells?
    Adenovirus. Can infect both dividing and non-dividing cells.
  55. (T/F) The Adeno-associated virus (AAV) can infect both dividing and non-dividing cells.
  56. (T/F) The adeno-associated virus (AAV) is the smallest virus and can only carry ~ 4kb of DNA.

    The DNA only goes in one spot of the genome.
  57. (T/F) Adeno-associated virus (AAV) does not stimulate inflammation or antibodies in the host.

    Awesome vector but limited in size.
  58. What are the approximate sizes for cells?
    5 - 100 um
  59. What are the approximate sizes for viruses?
  60. What are the approximate sizes for proteins?
    1-10 nm
  61. What are the approximate sizes for bacteria?
    1-5 um
Card Set
Tissue Engineering - Quiz 1
Tissue Engineering - Quiz 1 Stem cells