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Tissue eng > Lecture 3 > Flashcards

Lecture 3 Flashcards

1
Q

what are the 3 pillars of tissue engineering

A
  • cells : stem cells, differentiated
  • scaffold: ECM
  • signals: GF, environmetntal cues(e.g. mechanical, from ECM, vascular flow)
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2
Q

What is cell theory?

A

o All living organism are composed of one or more cells
o The cell is the basic unit of structure and organisation in organisms
o Cells come from pre-existing cells (hereditary information is passed from cell to cell)

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3
Q

what are the main classes of tissue in the body

A
  1. epithelial
  2. connective
  3. muscular
  4. nervous
  5. blood
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4
Q

How is tissue assembled

A

-Epithelium cells have cell adhesion molecules between themselves and between them and the basement membrane
-Mesenchymal cells which form the connective cells are attached to the ECM using cell adhesion molecules - so they can still communicate with each other

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5
Q

Describe how mesenchymal cells connect to the ECM.

A

Cells adhere to the ECM via cell adhesion molecules, the main one being integrin, which allows cells to sense mechanical forces eg changes in pressure and respond accordingly

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6
Q

what are the different types of epithelia?

A
  • simple/stratified
  • squamous/cuboidal/columnar
  • Stratified is usually used when there’s more than one layer
  • Pseudostratified is the term used to describe an epithelium consisting of closely packed cells which appear to be arranged in layers but all of which are in fact attached to the basement membrane.
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7
Q

A characteristic of epithelial cells?

A

polarised: apical surface (facing lumen or external surface) + basal (facing basement membrane)

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8
Q

Function of cytoskeleton in epithelial cells

A
  • Cytoskeleton are a network of tubules and filaments
  • give the cell shape
  • allow it to communicate with other cells
  • hold firm to the ECM
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9
Q

What structures make up the cytoskeleton in epithelial cells?

A
  • Microtubules - composed of tubulin
  • Microfilaments - composed of actin
  • Intermediate filaments - in epithelial cells are composed of keratins
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10
Q

Which cytoskeleton components are involved in formation of junctional complexes

A

microfilaments and intermediate filaments

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11
Q

Describe the main cell junctions in epithelium cells and their functions

A

Starting from apical:
Tight junctions - Prevent movement of solute (i.e. water movement)
Adherens junctions - Holds cells together
Desmosomes - Holds cells together
Gap junction - Enables solute flow between cells
Hemidesmosome - Anchors cell to the basal lamina.

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12
Q

what is the function of ECM?

A

-organise cells into tissue
-coordinate cellular function (e.g. division, signalling, motility) by activating cellular signalling pathways that control growth proliferation + gene expression

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13
Q

what are the 3 main types of ECM?

A
  • proteoglycans (Heparan sulfate)
  • fibrous ECM (collagen 1,4,7/elastin)
  • multiadhesive matrix proteins (fibronectin,laminin)
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14
Q

Why is mechanotransduction important

A

Mesenchymal focal adhesion structures
sends information about the type of ECM they are on and in doing so causes them to differentiate into different types of cells depending on the ECM they are on.
(In other words: Mesenchymal stem cells grown of substrates/ECM of varying stiffness, will differentiate into different cell types. (Outside in Signaling)) -
this is achieved by the focal adhesion (the use of integrin to join the cell to the ECM). - this process of ECM influencing the behaviour of the cell in a process is known as mechanotransduction

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15
Q

what are the different methods of tissue culture?

A
  • organ culture
  • explant culture - chop it up
  • dissociated cell culture (uses enzymes to break bonds
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16
Q

How to digest organ tissue to form culture for (dissociated cell cultures)

A
  • To digest an organ and make a culture of its cells:
    use Dispase which is an enzyme that digests collagen 4 which is found in the basement membrane, in doing so it separates the mesenchymal cells from the epithelial cells
17
Q

How to culture mesenchymal cells

A

Needs to isolate individual mesenchymal cells so use collagenase to breakdown the collagen IV.
Then individual mesenchymal calls plated in cell culture with bits of ECM for them to adhere to

18
Q

How to culture epithelial cells

A

To isolate the cells for culture, use trypsin enzyme to break the adhesion molecules between them, separating them.
Then either:
* Coat the dish with ECM and place the epithelial cells on them (the reason you coat the dish with ECM (can be collagen 1, fibronectin or laminin etc) is because you need something for the epithelial cells to hold on to and by providing the ECM they can form adhesion with the ECM)
Or
place a layer of inactivated cells (called a feeder layer) and this is the structure which the epithelial cells will form adhesions with

19
Q

What is cell culture focal adhesions

A

in mesenchymal cells, in culture, you get focal adhesions in which integrin attaches to the extracellular matrix on one side and the actin filaments within the cell.

20
Q

what is the difference between 2D vs 3D cell culture

A

2D cultures have no physical or physiological similarity to tissue of origin

21
Q

How does moving from a 3D environment to a 2D environment affect cell behaviour?

A

Cell doesn’t have apical and basal surface, you lose the orientation and in the case of the acini cells made in vitro, you don’t produce milk

22
Q

Problems of 2d cultures and solution

A

-As soon as you remove the cells from the body to put them in culture, the change in their shape will dramatically alter their behavior.
-Build an environment which resembles the environment the cells are used to in vivo. Architecture outside the cell influences behaviour,

23
Q

Using three pillar of term: engineering skin

A

Scaffold: collagen matrix/decellularised matrix
Cells: allogenic fibroblasts/keratinocytes
Signal: epidermal growth factor released from scaffold/fibroblasts/ added to medium

24
Q

Engineering a trachea

A

Cells: Epithelial cells, stems cells differentiated into chondrocytes (autologous)
Scaffolds: decellularised trachea from donor
sIGNALS: insulin, parathyroid hormone ect

25
Q

describe the stages of hair follicle development

A
  • via ectodermal appendage development

induction = interaction between 2 tissues → responding tissue undergoes differentiation

  • signal comes from dermine
  • 1st signal: likely Wnt
    • unipotent epidermis + dermis → placode
  • 2nd signal: promotes condensate formation
    • placode development → condensate formation
    • condensate = cells at double the density of surrounding dermis
    • get cluster of cells below placode
  • 3rd signal: promotes epithelail down growth
    • epithelium grows down forming hair germ, hair peg and later the hair follicle
    • epithelium engulfs condesate formig forming dermal papilla of adult follicle

Tissue eng (3 decks)

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