Download Cell Structure Tumor Microenvironment

Cell Structure
&
Tumor Microenvironment
Semra Aygun-Sunar, PhD
Department of Cell Stress Biology
[email protected]
RPN-530 Oncology for Scientist-I
October 4, 2012
Overview of this lecture:
• Visualization of cells-“Microscopes”
• Eukaryotic membrane-bound organelles, cell membrane,
Extracellular matrix (ECM) and cytoskeleton in normal and cancer
cells
• Cell-cell and cell-matrix adhesion in normal and cancer cells
• Epithelial-mesenchymal transition (EMT) in tumor progression
and tumor microenvironment
• Components of solid tumor
• Heterotypic signaling (Autocrine/paracrine signaling)
• Extracellular matrix remodeling
• Tumor hypoxia
• Tumor associated-angiogenesis in tumor microenvironment
• Tumor stroma-organ specific metastasis
• Therapeutic targeting the tumor microenvironment
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Reading
Book
• “The Biology of Cancer”, Chapter 13 Robert Weinberg,
Garland Science, 2006
• “Molecular Biology of the Cell” (Fifth Edition). Bruce Alberts,
Alexander Johnson, Julian Lewis, Martin Raff, Keith Roberts,
Peter Walter; 2008
Review
• Hanahan & Weinberg, Hallmarks of Cancer: the next
generation. Cell. 2011. 144: 646-674
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Cell
“is the functional and smallest unit in every living organism ”
• In 1665, the cell was discovered by
Robert Hooke.
• In 1839, the cell theroy was developed
by Matthias J. Schleiden and Theodor
Schwann
(Ref: The Molecular Probes® Handbook-11th Edition, 2010, Invitrogen)
Modern concept of cell theory
• The cell is the essential unit of structure and function in living organisms
• All cells arise from pre-existing cells by division
• All cells have basically the same composition
• Energy flow occurs within cells
• Cells contain hereditary information (DNA) which is passed from cell to cell
during cell division
• Organisms can be “unicellular” or “multi-cellular”
• The activity of an organism depends on the total activity of independent
cells
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Types of Cells
1. Prokaryotic cells: Archae and bacteria
(no nucleus, no membrane-bound organelle, contain primitive
cytoskeleton and circular-shaped DNA)
2. Eukaryotic cells: Protists, fungi, plants, animal (including human)
(contain various membrane-bound organelles, cytoskeleton,
chromatin and chromosome)
Prokaryotic cell
Cell Structure & Tumor Microenvironment
vs
Eukaryotic cell
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Visualization of Cells
Light microscopy
Ø Principle of light microscopy
• use visible light (ranges from 0.4 µm (for violet to 0.7 µm (for deep red)
• light is transmitted through or reflected from specimen
• used to visualize living cell to see their compartments.
Ø Types of light microscopes
1. Bright field
microscopy
Passes light
directly through
specimen
2. Dark-field
microscopy
Light scattered
by specimen
3. Phase-contrast
microscopy
Alter light waves
to enhance the view
of specimen
4. Differential interference
contrast microscopy
(DIC or Nomarski)
Give specimens a 3D
appearance
The same fibroblast cell in culture shown using different types of light microcopies.
Ref: “Molecular Biology of The Cell”, 5th edition. Alberts et al., 2008. Chapter 9, p.584
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Visualization of Cells (cont’d)
Electron microscopy
Ø Principle of electron microscopy
• use beam of highly energetic electron to illuminate the specimen
• has high magnification and high resolution
• specimen require specifically preparation:
- can be cut or sectioned
- is coated in gold metal
Ø Types of electron microscopes
* Transmission Electron Microscopy (TEM)
• uses the electrons that pass through specimen
• is used for analyzing sections
• shows only structure of cell
• gives a 2D views
• magnification: 1,000-500,000X
* Scanning Electron Microscopy (SEM)
• uses the electrons reflected from a specimen
• is used for analyzing surface of the specimen
• shows defined organelles inside cell
• gives a 3D views
• magnification: 10-300,000X
Cell Structure & Tumor Microenvironment
TEM vs SEM
T
E
M
S
E
M
Cilia lining the trachea
under TEM and SEM
http://www.med.nus.edu.sg/ant/
histonet/tacsem/tac20.sem.gif
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Visualization of Cells (cont’d)
Fluorescence microscopy & Confocal microscopy
• are used for studying material that can be made to fluorescence either
in its natural form (such as chlorophyll has endogenous auto-fluorescence) or
when labeled or tagged with fluorescent dyes or probes
• can visualize and localize specific proteins and other molecules inside
living cells.
Fluorescent molecules absorb light at
one wavelength called as Excitation,
and emit it at another long wavelength
called as Emission
Fluorescence probes
Epithelial cells stained with
DAPI (blue) and two antibodies
(green and red) via
immunofluorescence
Cell Structure & Tumor Microenvironment
Protein probe
Ex
(nm)
Em
(nm)
Em
color
FITC
488
525
Green
Nucleic acid probe
Ex (nm)
Em
(nm)
Em color
DAPI
350
470
Blue
Hoechst 33342
343
483
Blue
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Visualization of Cells (cont’d)
Fluorescence microscopy & Confocal microscopy (cont’d)
Ø Principle of fluorescence microscopy
• uses a much higher intensity light (UV mercury vapor lamp) that
excites fluorescence in the specimen
• UV light is filtered to select excitation light to pass through
• All parts of the specimen in the optical path are excited at the same
time and the resulting fluorescence is detected by the microscope's
camera including a large unfocused background part.
Ø Principle of confocal microscopy
uses point illumination and a pinhole in an optically conjugate plane in
front of the detector to eliminate out-of-focus signal (the name "confocal"
stems from this configuration)
As only light produced by fluorescence very close to the focal plane can
be detected, the image's optical resolution, particularly in the sample
depth direction.
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Visualization of Cells (cont’d)
Fluorescence microscopy vs Confocal microscopy
Wide-field fluorescence vs Confocal microscopy
Ref: http://www.itg.uiuc.edu/technology/atlas/microscopy/confocal.htm
•works with very thin specimens or
when a thick specimen is cut into
sections.
Cell Structure & Tumor Microenvironment
• performs optical sectioning of thick
samples,
• ability to control depth of field,
• provides 3D-image reconstruction,
• detects very weak fluorescent
signals,
• generate high-resolution images,
• more color possibilities.
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The major intracellular compartments of animal cell
http://www.rkm.com.au/CELL/animalcell.html
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Eukaryotic Cell Organelles
1. Nucleus
Ø Structure
!
I. Nuclear envelope
* Composed of inner
and outer membrane
separated by a
perinuclear space and
having nuclear pores
which connect with ER
* Each pore is a ring of 8
proteins with an
opening in the center of
the ring
* Numerous openings
for nuclear traffic
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Eukaryotic Cell Organelles (cont’d)
Ø Structure (cont’d)
1. Nucleus (cont’d)
II. Nucleoplasm - fluid of the nucleus
III. Nucleolus
• Spherical shaped structure and not have membrane
• Area of condensed DNA
DNA is organized with proteins to form “chromatin” in chromosomes
(Chromatin-contains DNA and histones)
(Chromosome – fiber of DNA with proteins attached
• Ribosomal subunits are produced
Ribosomes -small particles of RNA and protein that are involved in
protein synthesis.
Nucleosome:
11 nm in diameter that
consists of 146 base
pairs of DNA wrapped
around eight histone
molecules
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Eukaryotic Cell Organelles (cont’d)
1. Nucleus (cont’d)
“contains the genetic information of the cell”
Ø Function
* Storage of hereditary material
* Production of messenger RNA and ribosomes that needed for
protein synthesis.
* Storage of proteins and RNA in the nucleolus.
* During the cell division, chromatins are arranged into
chromosomes in the nucleus.
* Selective transportation of regulatory factors and energy molecules
through nuclear pores.
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Eukaryotic Cell Organelles (cont’d)
2. Mitochondria
Ø Structure
• Surrounded by 2 membranes:
- smooth outer membrane (permeable)
- folded inner membrane (impermeable)
with layers called “cristae”
• Contains two internal compartments:
- mitochondrial matrix is within the
inner membrane (contain ribosomes
mitochondrial DNA (mtDNA) and
enyzmes)
- intermembrane space is located
between the two membranes
The structure of mitochondria.
Ref:microbewiki.kenyon.edu/index.php/
Mitochondria
Cell Structure & Tumor Microenvironment
mtDNA:
• inherited from mother
• not protected by histones
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Eukaryotic Cell Organelles (cont’d)
2. Mitochondria (cont’d)
“ Energy producing organelle”
Ø Function
•
!
•
“Oxidative phosphorylation”--ATP synthesis
!
!
!
---Reactive Oxygen Species (ROS)/Free
!
!
!
!
!
radical generation
Regulation of apoptotic death
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Eukaryotic Cell Organelles (cont’d)
2. Mitochondria (cont’d)
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Eukaryotic Cell Organelles (cont’d)
2. Mitochondria (cont’d)
ETC may leak electron to oxygen partially reducing oxygen to superoxide
anion that is the precursor of ROS
Ref: Murphy MP. Biochem J, 2009
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Eukaryotic Cell Organelles (cont’d)
2. Mitochondria (cont’d)
Apaf-1: Apoptotic
protease activating
factor 1
Ref: http://www.reading.ac.uk/nitricoxide/intro/apoptosis/mito.htm
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Eukaryotic Cell Organelles (cont’d)
3. Endoplasmic reticulum (ER)
“a network of membranes throughout the cytoplasm of the cell”
Ø Structure
Cisternae
• Cisternae (sac-like structures)
• Cisternal space (or lumen)
• ER membrane is
continuous with nuclear
envelope
Ø Types of ER
• Smooth-type (SER) :
•Ribosome-free
•Contains enzyme for lipid
biosynthesis
•Involved in attachment of
receptors on cell membrane
proteins
• Rough-type (RER) :
•Ribosomes embedded in
surface
•Involved in protein
synthesis
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Eukaryotic Cell Organelles (cont’d)
3. Endoplasmic reticulum (ER) (cont’d)
ØFunction
• Protein folding, modification and secretion
• Cellular protein quality control by extracting and degrading unfolded
proteins (known as a ER-associated protein degradation-ERAD)
• Lipid and sterol biosynthesis
• Storage of calcium ions in the ER lumen and their regulated release
into the cytosol (calcium homeostasis)
• Detoxification of drugs
• Core oligosaccharide biosynthesis
• Apoptosis
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3. Endoplasmic reticulum (ER) (cont’d)
Major responses to ER stress
ERAD : ER Associated
Degradation
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Unfolded Protein Response
http://www.pdbj.org/eprots/index_en.cgi?PDB%3A2RIO
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Eukaryotic Cell Organelles (cont’d)
Ø Structure
4. Golgi Apparatus
• Composed of numerous group of
flat membranes called cisternea
forming a sac
---a complex network of tubules and
vesicles are located at the edges of
cisternea
---help proteins and cytoplasmic
components travel between different
parts of the cell
• Cis face (Cis Golgi network): ER to
Golgi apparatus
• Golgi stack: Main processing area
• Trans face (Trans Golgi network): Golgi
apparatus to target
ØFunction
• Receives, sorts, modifies, packs and ships the biochemical for use
inside and outside the cell
• Produce specialist vesicles or vessels for transport of the product
• Protects cells from apoptosis
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Eukaryotic Cell Organelles (cont’d)
5. Lysosomes
Ø Structure
“suicide bags” and “recycling”
• are spherical bodies about 50-70 nm in diameter that bounded by a
single membrane
•arise from the golgi apparatus
• are found in all animal cells, but mostly in disease-fighting cells, such as
white blood cells.
•contain hydrolytic enzymes that degrade proteins, nucleic acid and lipids
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Eukaryotic Cell Organelles (cont’d)
Types of lysosomes
5. Lysosomes (cont’d)
Peroxisomes: * a vesicle containing oxidases and catalase and located by the
smooth ER
*functions in oxidizing amino acids and fatty acids and also detoxifing alcohol.
Proteasome: a tiny barrel-shaped structure and contain proteases
Ø Function
• Phagocytosis: Degrade the
products of ingestion such as
bacteria
• Autophagy: Degrade damaged
organelles such as mitochondria.
• Receptor-Mediated
endocytosis: Degrade
macromolecules
Autophagy roles in the cellular
response to oxidative stress
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Plasma membrane (or cell membrane)
“separates the cell from the environment”
Ø Structure
• Fluid phospholipid bilayer
• Hydrophilic polar heads face outside, and hydrofobic nonpolar tails face
each other
• Contains integral (embedded in the membrane) and peripheral proteins
(found on the inner membrane surface)
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Plasma membrane (cont’d)
ØFunction
• Protects cells
• Diffusion barrier
• Regulation of transport
• Detection of signals
• Cell-cell communication
• Cell identity
• Helps in cell movement
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Cytoskeleton
“the movers and shapers in the cell”
is found underlying the cell membrane in
the cytoplasm
Ø Structure
• the motor proteins: kinesin, dynein and myosin
• the protein filaments: actin filaments,
intermediate filaments and microtubules
The cytoskeleton in animal cells
The blue areas are the nucleus
of the cells, the green areas show
cytoskeleton microtubules and red
areas show actin filaments
(Ref: www.bscb.org)
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Major Structures of the Cytoskeleton
I. Actin filaments
Microfilament structure and assembly
http://micro.magnet.fsu.edu/cells/microfilaments/microfilaments.html
• occurs in every cell
• composed of actin
proteins
• interacts specifically with
myosin helical polymers
made of actin flexible,
organized into 2D networks
and 3D gels
II. Intermediate filaments
Intermediate filament structure
http://micro.magnet.fsu.edu/cells/microfilaments/microfilaments.html
Cell Structure & Tumor Microenvironment
• occurs only in
animal cells
• composed of
heterogenous group
filamentous proteins
called keratin
• rope-like structure
• size: 8 -12 nm
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Major Structures of the Cytoskeleton (cont’d)
III. Microtubules
Microtubule helical structure
http://micro.magnet.fsu.edu/cells/microfilaments/microfilaments.html
Cell Structure & Tumor Microenvironment
• structural support of Cilia
and Flagella
• composed of alpha and
beta-tubulin
• rigid, long, straight
• length: 200 nm-25 µm
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Cytoskeleton (cont’d)
Ø Function
• Cell shape,
• Cell polarity,
• Cell motility,
Cell cycle (Cell division and chromosomal separation by actin and
tubulin cytoskeletal structures)
• Cell morphology,
• Cell migration,
• Cell adhesion,
• Phagocytosis (are driven by actin cytoskeleton)
• Provides a machinery for muscle contraction (by actin and myosin )
• Wound healing
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Epithelial Cells
vs
Mesenchymal cells
• are fairly uniform small spindle• are polygonal in shape
• have three membrane domains:
apical, lateral and basal,
• have tight junctions between
apical and lateral domains,
• apical-basal polarity,
• have adherens junctions,
• express cell-cell adhesion
markers such as E-cadherin,
• lack of mobility.
Cell Structure & Tumor Microenvironment
shaped cells,
• do not make mature cell-cell
contacts, and can invade through
the ECM,
• are connected to other cells within
a 3D- cellular network,
• bipolar (because of having different
cytoskeleton arrangement and distinct
organelle distribution inside them)
• are a special type of
undifferentiated connective tissue,
• express markers such as Ncadherin, Twist, snail, fibronectin
• can migrate easily,
• function in the progression of
tissue development.
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Epithelial-mesenchymal transition (EMT)
• is a process by which epithelial cells lose their polarity and are
converted to a mesenchymal phenotype,
• function in embryonic development, organ formation, tissue
regeneration, wound healing, organ fibrosis and cancer progression
and metastasis.
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Cell-cell adhesion
Cadherins
• Expresses on the plasma
membrane
• Interacts in a zipper-like fashion
• Stabilized by catenin complex
• Are divided into Type-I and
Type-II cadherins
Type-I: * E-cadherin (epithelial)
* P-cadherin (placental)
* N-cadherin (neuronal)
Homophilic E-cadherin interaction and
homotypic cell adhesion within the
epithelium.
(Ref: Mohamet et al., Journal of Oncology, 2011)
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Cell-cell adhesion (cont’d)
“intercellular junctions”
Plasma
membrane
Intracellular
space
Tight junction
proteins
Tight
junction
Plasma
membrane
Protein
filaments
Intracellular
space
Desmosome
Intracellular
space
Plasma
membrane
Protein
channels
Gap
junction
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Cell Structure & Tumor Microenvironment
• Tight junction:
- fused membranes of adjacent cells
- form a continuous belt around cells
- impermeable
- ex: intestine, kidneys, epithelium of
skin
• Desmosomes
- fastening adjacent cells together
by connecting cell membrane to
cytoskeleton proteins
- binding spots between cells with
cadherins
-ex: stomach, bladder, heart
• Gap junction
- allow small molecules to pass
directly from cell to cell
in heart muscle, the flow of ions
through gap junctions coordinates
the contraction
- chemical communication between
animal embryos is essential for
development
- ex: heart muscle, animal embryos
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Extracellular matrix (ECM)
Is the defining feature of connective tissue in animals
•
•
•
•
includes the interstitial
matrix and basement
membrane
Adhesive glycoproteins:
fibronectin and laminins
Protein-polysachharide
complexes: proteoglycans
Structural proteins:
collagens and elastins
**They can be mixed up in different
Plasma
membrane
Ø Structure
proportions for different functions
•
•
consisting of various cell
types (i.e. fibroblasts,
epithelial cells)
contains secreted proteins
(cytokines)
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Extracellular matrix (ECM) (cont’d)
Ø Function
• Cell shape,
• Cell attachment,
• Adhesion,
• Migration (example: wound healing),
• Cell proliferation,
• Polarity,
• Differentiation,
• Survival & apoptosis,
• Motility,
• Management of growth factors,
• Embryonic development.
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Cell-ECM adhesion
Integrins
• are heterodimeric cell-surface molecules
• function as transmembrane linkers between actin cytoskeleton
and the ECM (mediate cell-matrix interaction)
• function as signal transducers, activating various intracellular
signaling pathways when activated by matrix binding
The regulation of the extracellular binding activity of a cell's integrins Ref: Molecular Biology of the Cell, 4th edition
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Normal cells vs Tumor cells
Normal cells
• Controlled cellular growth
• Balanced cell division
• Perform a special function
Cell Structure & Tumor Microenvironment
Tumor cells (as Disease)
• Uncontrolled cellular growth
• Unbalanced cell division
• Loss of special function (abnormal
organelles and cell components)
• Cellular invasion into adjacent tissues
• Potential to metastasize
• Abnormal vessel wall structure.
• High glycolytic rate
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Tumors as 'wounds that do not heal'
Normal skin tissue
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Invasive carcinoma
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Tumor types
Bening (non-cancerous) tumor vs malignant (cancerous) tumor
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Normal Cells vs Cancer Cells
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Nuclear structure in cancer cells
Normal
N
e
ucl
oli
Nuclear Nuclear
lamina matrix
PML
Cancer P
e
comrinuc
par leola
tme r
nt
PML: Promyelocytic
leukaemia
NMP: nuclear matrix
proteins
NMP
(Ref: Zink et al.,
Nature reviews, 2004)
heterochromatin
Nucleus in normal cell
• single and small nucleus & single
nucleolus
•Fine chromatin granules in the
nucleus
•Clear nuclein stain
•The smooth nuclear boarder
•Stain artifact in one cell where
another small white blood cell
overlaps.
Cell Structure & Tumor Microenvironment
Nucleus in cancer cell
• multiple and large nucleolus &
multiple nuclei
• large chromatin clumps in the nucleus
• the dark staining of the nucleus
irregular nuclear boarder.
• Nuclei can become irregular and
begin to fold
• Nucleoli can be enlarged and PML
bodies can mislocalized.
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Mitochondria in cancer cells
• mtDNA mutations inhibit oxidative phosphorylation: Increase ROS level and tumor
cell proliferation
Apoptosis-resistant mitochondria and cancer
Ref: Indran et al., BBA, 2011
•Crosstalk between nucleus and mitochondria contribute to oncogenesis and
tumor progression
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Unfolded protein response (UPR) & Endoplasmic
reticulum (ER) stress in cancer cells
Ref: Wang et al., Am J Transl Res, 2010
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Lysosome alterations & Autophagy in cancer cells
Normal
Cancer
Lysosomes in normal versus cancer cells.
Visualization of the lysosomal compartment
(using lysosome-associated membrane
protein 1) monoclonal
antibodies, red) and the actin cytoskeleton
(using anti-β-actin monoclonal antibodies,
green) in murine embryonic fibroblasts.
Note the perinuclear and peripheral
localization of lysosomes in control and
transformed cells, respectively.
Lysosomal alterations increase expression
and altered trafficking of lysosomal
enzymes participates in tissue invasion,
angiogenesis and lysosomal death
pathway.
Autophagy act as both a “tumor suppressor”
by preventing the accumulation of damaged
proteins and organelles and as a “mechanism
of cell survival” that can promote the growth of
established tumors.
Ref: Kroemer and Jäättelä nature Reviews, 2005, 5:
886-897
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Plasma membrane structure in cancer cells
Changes in the plasma membrane fluidity of tumor cells
Komizu et al,
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Cytoskeletons in the cancer cells
Actin cytoskeleton in the invasion/ metastatic process
Vignjevic and Montagnac, Seminars in Cancer Biology, 2008
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Integrins and cadherins in the cancer cells
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Blood vessels & flow in cancer cells
Diagrammatic representation of the vascular system. A. Normal tissue. B. Solid tumor. Red represents well-oxygenated
arterial blood, blue represents poorly oxygenated venous blood, and green represents lymphatic vessels. (Ref: Tredan et al.,
JNCI, 2007, 99:1441-1454)
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Cancer classifications
Based on the location or origin of the malignant tumor
Carcinoma
Malignant tumors
of epithelial cells
Sarcoma
Lymphoma
Malignant tumors
Malignant tumors
of lymphocytes
of supporting tissue
(non-epithelial tumors:
mesenchymal origin)
Solid tumor
Cell Structure & Tumor Microenvironment
Leukemia
Malignant tumors
of blood cells
Non-solid tumor
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Solid tumor
“organ-like structure”
Consisting of Tumor cells and Stroma (microenvironment)
Tumor cells:
• Epithelial cells
• Mesenchymal cells
• Hematopoietic cells
Stroma:
• Neoplastic cells (cancer stem
(macrophage,
neutrophil, mast
cell)
An assemblage of various cell types constitutes most solid tumors (Ref:
Hanahan & Weinberg, Cell. 2011. 144: 646-674)
cells)
• Stromal cells (fibroblasts, myofibroblast, endothelial cells, pericytes,
adipose and inflammatory cells)
• Secreted soluble factors include “cytokines” (i.e. CXCR-4 and CXCL-12,
TNF-α), “matrix-altering enzymes” (i.e. matrix metalloproteinases (MMPs)) and
growth factors (i.e. VEGF, FDG, PDGF, TGF-β)
• The extracellular matrix
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Different tumor microenvironments
CSC
CC
ICs
ECM
ECM
CAF
ICC
EC
PC
Multistep of tumorogenesis and tumor microenvironment.
(Ref: Hanahan & Weinberg, Cell. 2011. 144: 646-674)
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Cancer stem cells
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Stromal cells
Fibroblasts
• are the major stromal cell types
• are principal cellular component of connective tissue.
• secrete the proteins needed for fiber synthesis and components of the
ECM.
• are involved in the production and remodeling of the ECM in both
normal and cancer tissue
• produce different types of fibers like collagen, reticular and elastic
• function in wound healing, tumor development, tumor angiogenesis
and metastasis
NIH/3T3 Fibroblasts in cell culture
(Ref: http://en.wikipedia.org/wiki/Fibroblast)
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Stromal cells (cont’d)
Myofibroblasts
• are a type of cell that is between a fibroblast and a smooth muscle cell
in differentiation. • express alpha smooth muscle actin (α-SMA)
• are often observed in the stroma of various human carcinomas
• function in wound healing and tumor development,
• stimulate angiogenesis
• involved in ECM remodeling because they secrete key ECM molecules
and several growth factors and cytokines
α-smooth muscle actin staining
(reddish brown) of a 3 day
excisional wild-type wound reveals
a myofibroblast presence in the
wound granulation tissue (asterisk)
(Martin et al., Curr Biol. 2003. 13:1122-8)
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Carcinoma-Associated fibroblasts (CAF)
Within tumor, fibroblasts acquire an “activated” phenotype within the
tumor characterized by expression of myofibroblast markers (α-SMA) and
increasing proliferation and motility. Under these conditions, fibroblasts
are called carcinoma-associated fibroblasts (CAF) or tumorassociated fibroblast or reactive stromal fibroblast
CAF are consisting of both fibroblasts and myofibroblast
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Stromal cells (cont’d)
Endothelial cells & Pericytes
“are participate in the construction of the tumor-associated vasculature”
Endothelial cells:
• are line the walls of capillaries and large blood vessels, and lymphatic
ducts.
Pericytes:
• are vascular connective tissue cells that occur in small blood vessels
• are adjacent to endothelial cells and embedded within the vascular
basement membrane of blood microvessels
• are relatively undifferentiated cells but differentiate into a fibroblast,
macrophage or smooth muscle cell
Endothelial–pericyte interactions in microvessels.
Ref: Armulik et al.,
Circulation
Research, 2005
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Stromal cells (cont’d)
Tumor-associated endothelial cells
Ø Function
• tumor development and progression
• development and function of blood and lymph vessels (angiogenesis/
lymphangiogenesis),
• controlling leukocyte recruitment,
• tumor cell behavior,
• metastasis formation.
Pericytes - Endothelial cells signaling network can contribute to tumor
development and metastasis.
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Stromal cells (cont’d)
Immune inflammatory cells
Many tumor-associated immune cells such as macrophages, dendritic cells,
and cytotoxic T cells may either promote tumor growth and progression or
protect the cancer cell from apoptosis
Macrophages
• are white blood cells
• are crucial member of tumor stromal cells.
• are phagocyte of cell debris and pathogens
• function in wound healing, inflammatory
response
Tumor associated macrophages
ØFunction
A macrophage of a mouse
stretching its
"arms" (Pseudopodia) to engulf
two particles, possibly pathogens
[Obli at en.wikipedia CC-BYSA-2.0.]
• induce tumor growth,
• promote angiogenesis,
• enhancement of tumor cell migration and invasion,
• promote metastasis
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Stromal cells (cont’d)
Adipocytes
• are one of the most abundant cell types of the
stromal compartment of the mammary gland
• primarily constitute adipose tissue
• store energy as fat
Ø Function
Section of tumor (mauve) in the
presence of adipocytes (white discs).
The arrows indicate adipocytes
modified by the tumor.
(Credit: Copyright G. Escourrou)
• contribute to tumor progression:
- secrete metalloproteinases, specific cytokines (i.e. adipokines,
adiponectin, resistin, visfatin)
• act as a energy source for the cancer cells
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Cytokines
Transforming growth factor beta (TGF-β):
--- is a secreted protein that principle inducer and regulator of reactive
stroma
--- is secreted by many cell types, including macrophages
--- acts as an antiproliferative factor in normal epithelial cells and at early
stages of oncogenesis
--- induces apoptosis in two ways: through the SMAD pathway or the death
associated protein 6 pathway in numerous cell types.
•TGF-β signaling is involved in many cellular processes such as cell
cycle arrest, angiogenesis, and homeostasis.
•The deformation of TGF-β signaling is crucial in tumorigenesis and
invasion.
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Chemokines in Tumor Microenvironment
Ref: Raman et al., Cancer Letter, 2007
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Metalloproteinases (MMPs)
• are metal-dependent endopeptidases
• are produced by stromal cells rather than by tumor cells in many solid
tumors
Ref: Roy et al., JCO, 2009
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Growth factors
• Fibroblast growth factors (FGF):
----- are heparin-binding proteins that regulates reactive stroma
----- function in proliferation, migration and differentiation of variable cells
----- stimulate angiogenesis and involved in wound healing and regulates
matrix remodeling
----- involved in pathogenesis of cancer
• The platelet derived growth factor (PDGF):
---- is dimeric glycoprotein
---- functions in embryonic development, cell proliferation, cell migration,
and angiogenesis
---- is frequently upregulated in tumors and promote tumor angiogenesis
and metastasis
• Vascular endothelial growth factor (VEGF):
---- is a sub-family of growth factors, specifically the platelet-derived
growth factor
---- stimulates vasculogenesis and angiogenesis
---- is overexpressed in cancer thus cells are able to grow and
metastasize.
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Integrin-growth factor & integrin-cytokine signalling
in tumor stroma
ECM, extracellular matrix; EGF, epidermal growth factor; FAK, focal
adhesion kinase; SDF1, stromal cell-derived factor 1; RTK, receptor
tyrosine kinase.
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Conditions within the tumor microenvironment
• Massive cell death→ results in release of proteins and
additional molecules
• Hypoxia (low oxygen levels)
• Acidic conditions (low pH level)
• Low glucose level
• Abnormal properties of surrounding cells
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Hypoxia in solid tumor
• results from an imbalance between the oxygen supply and consumption
rate
• results in generation of oxygen free radicals, leading to DNA damage
• can induce proteomic and genomic changes that allow the tumor cells
adapt to hypoxic condition
• lead to the activation of genes that are associated within tumor
progression, and metabolic adaptation, and cell survival
• stimulate angiogenesis and inhibite apoptosis
• can induce EMT in tumor cells
B
• induce downregulation of adhesion molecules to promote the tumor cell
detachment
• mediated by transcription factor hypoxia-inducible factor I (Hif1), oxygen
sensor
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Hypoxia in solid tumor (cont’d)
B
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Cross-talk between tumor cells and stroma
• Heterotypic signaling (Autocrine / Paracrine signaling)
• Tumor induced alterations of ECM
• Tumor associated angiogenesis
• Organ-specific metastasis
* is important in tumorogenesis, tumor progression and pathogenesis of
cancer
* influences growth, ability to progress, metastasis
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Heterotypic Signaling
“communication between dissimilar cell types, used to
stimulate proliferation”
• mitogenic growth factors
(such as hepatocyte growth factor (HGF), transforming growth factor-α
(TGF-α) and platelet-derived growth factor (PDGF)
• growth inhibitory signals (such as TGF-β)
• trophic factors (such as insulin-like growth factor -1 and -2 (IGF-1and
IGF-2 )
!
!
!
PDGF
↓
Stimulates fibroblast recruitment
!
!
↓
!
Myofibroblast formation
!
!
!
!
!
↓
!
↓
!
↓
SDF-1/CXCL-12
Endothelial precursor recruitment
!
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Autocrine (within the cell) signaling
TGF-β and SDF-1 autocrine signaling
(Ref: Kojima et al., PNAS 2010, 107(46): 20009–20014)
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Paracine (cell-to-cell) signaling
.
Promotion of
tumorogenesis
Promotion of
proliferation
Promotion of
angiogenesis
Promotion
of inflammation
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SDF-1/CXCR4 Paracine and Endocrine signaling
.
EPCs: Endothelial progenitor cells
Ref: Orimo and Weinberg (2006)
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Tumor-associated angiogenesis
A
Molecular mechanisms of tumor-associated angiogenesis. (Ref: Simpson-Haidaris
et al., PPAR Research, 2010)
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Tumor-associated angiogenesis in tumor microenvironment
A
B
Tumor-associated angiogenesis is sustained through stromal microenvironment crosstalk.
(Ref: Simpson-Haidaris et al., PPAR Research, 2010)
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EMT in Tumor Progression
Progression
EMT is characterized by loss of E-cadherin, disruption of cell adhesion, and
induction of cell motility and invasion.
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EMT in tumor microenvironment
Various factors that induce cancer cell EMT in tumor microenvironment. (Ref: Jing et al., Cell & Bioscience 2011, 1:29)
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Tumor stroma & organ-spesific metastasis
Schematic representation
showing the role of
microenvironment in tumor
cell CXCR4 receptor
activation in both the
primary and metastatic
sites
By SDF1 (CXCL12)
chemoattracting of organ
secretion, CXCR4-positive
tumor cells in circulation may
be responsible for the
process of extravasation, and
..organ-specific metastasis.
Ref: Sun et al., Cancer Metastatis
Rev. 2010
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Tumor stroma & Lymph node metastasis
A: Tumor-secreted factors
induce lymphangiogenesis,
lymphatic activation, and preconditioning of lymph nodes
for metastasis.
B: Tumor cells can activate
their surrounding stroma,
while an activated stroma can
also induce increased
tumorigenicity and metastasis
in tumor cells.
C: The stromal microenvironment (ex: immune
cells)
can
induce
lymphangiogenesis through a
variety of signals.
Signals and interactions within the tumor microenvironment could
potentially affect lymph node metastasis
Ref: Journal of Cellular Biochemistry 101:840-850 (2007)
Cell Structure & Tumor Microenvironment
D: The lymphatic endothelium
and/or the lymph node
release factors that recruit
tumor cells
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Heterotypic interactions as therapeutic targeting
“Targeting the tumor microenvironment”
chemokine inhibitors)
Inh
VE ibito
sig GF, Frs of
nal
ing GF
Cell Structure & Tumor Microenvironment
Inhibitors of
HGF /c-Met
Inhi
PDGbitors o
f
sign F
aling
Anti-inflammatory
inhibitors (cytokine,
Inh
ma ibito
trix rs o
tur f
no
ver
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From this lecture you should focus:
• What is the purposes of using different microcopies?
• What are the structure and function of membrane-bound organelles
in eukaryotes?
• How is the cell structure in tumor cells compare to normal cells?
• How cell adhesion molecules help with the cell-cell and cell-matrix
adhesion?
• What is the Epithelial-mesenchymal transition process in tumor
cell?
• How is blood vasculature in tumor tissue compare to normal
tissue?
• How cells interact with both their environment and neighboring cells
in tumor stroma?
• How hypoxia induce the mechanisms for cell survival, invasion and
metastasis
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Sample Questions
1. Match the following
# a-Metalloproteinases
b-Mitochondria#
#
c-VEGF# #
#
d-Integrin##
#
e-Myofibroblasts# #
f-Golgi apparatus# #
1. is a part of endomembrane system
2. express alpha smooth muscle actin
3. cause extracelullar matrix degradation
4. contains own DNA that is not protected by histons
5. stimulates angiogenesis#
6-helps cell-extracellular matrix adhesion
2. Which of the following(s) is/are true?
A. Pericytes acquire an “activated” phenotype within the tumor
characterized by expression of α-SMA and increasing
proliferation and motility.
B. Adipocytes act as a energy source for the cancer cells
C. EMT is characterized by loss of integrin and disruption of
cell adhesion
D. Cancer cells have large variably shaped nuclei and small
cytoplasmic volume relative to nuclei.
3. Briefly explain the structure and function of
mitochondria in normal cells.
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QUESTIONS?????
[email protected]
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