CST Guide:

Section I: Research Areas
chapter 06: Development and differentiation
Elevated PDGF
Receptor β expression
is found within stromal
cells of many forms
of cancer.
PDGF Receptor β (28E1) Rabbit mAb
#3169: IHC analysis of paraffin-embedded
human colon carcinoma using #3169.
CD73 expression on
tumor cells promotes
angiogenesis.
NT5E/CD73 (D7F9A) Rabbit mAb
#13160: IHC analysis of paraffin-embedded
human lung carcinoma using #13160.
Pericyte Signaling
Pericytes are support cells that provide structural stability for newly formed blood vessels, promote EC
survival, and regulate vasoconstriction and dilation. This is done through a reciprocal signaling mechanism
in which PDGF-BB secreted into the matrix by ECs acts as a ligand for PDGFR-β located on the
pericyte membrane. In return, pericytes produce and secrete VEGF that signals through the endothelial
VEGF receptor.
Tumor Signaling
Tumor angiogenesis occurs when cancer cells stimulate new blood vessel growth in order to bring
oxygen and nutrients to a tumor. As a tumor grows in size, diffusion is no longer sufficient to oxygenate
the cells at the center of the mass, creating a hypoxic environment. Hypoxia stabilizes levels of HIF-1α,
a transcription factor that responds to changing oxygen levels. Under hypoxic conditions, HIF-1α binds
HIF-1β to activate transcription of angiogenesis-promoting genes. Cancer cells also secrete a variety
of growth factors and cytokines that stimulate classical angiogenic signaling pathways, extracellular
matrix remodeling, and an inflammatory response that leads to new blood vessel formation.
Under normoxia conditions, HIF-1α is
hydroxylated at Pro564, marking it for degradation.
PathScan ® Hydroxy-HIF-1α (Pro564) Sandwich ELISA Kit #13201:
Treatment of HeLa cells with the hydroxylase inhibitor dimethyloxaloylglycine
(DMOG) results in decreased hydroxylation of HIF-1α, as detected by
#13201, but does not affect the level of total HIF-1α detected by PathScan ®
Total HIF-1α Sandwich ELISA Kit #13127. Absorbance at 450 nm is shown
in the top figures, while corresponding western blots using a total HIF-1α
antibody (left) and Hydroxy-HIF-1α (Pro564) (D43B5) XP ® Rabbit mAb #3434
(right) are shown in the bottom figures. Treatment of HeLa cells with the
proteasome inhibitor MG-132 #2194 stabilizes HIF-1α protein.
Untreated
MG-132-treated
MG-132-treated,
DMOG-treated
Total HIF-1α
Hydroxy-HIF-1α
(Pro564)
Angiogenesis in Cancer
Cancer cells secrete a variety of growth factors and cytokines that stimulate classical angiogenesis
signaling pathways and extracellular matrix remodeling. Cytokines can also induce an inflammatory
response that initiates angiogenesis and consequent vascularization of the tumor. Sprouting angiogenesis
is the first step in this neovascularization process. In response to stimuli released by tumor cells,
a single EC migrates toward the angiogenic factors and proliferates to form sprouts of ECs surrounding
a lumen that is connected to the mother blood vessel. Tumors may also be vascularized by intussusceptive
angiogenesis. This process proceeds much more quickly than sprouting and is essentially the
splitting of an existing blood vessel into two new vessels. The ECs, pericytes, and basement membrane
associated with a tumor exhibit abnormalities leading to tortuous, poorly organized, and leaky vasculature.
However, this sub-optimal structure is often sufficient to supply the tumor with oxygen, nutrients,
and soluble factors that help it survive and expand. It is through this vascular system that some tumor
cells are able to escape and travel through the circulatory system to new locations. Such metastatic
colonization is a key factor in the fatal outcomes of many types of cancer, and the density of tumor
angiogenesis has been linked to tumor metastasis and patient survival rates. Accordingly, angiogenesis
is under investigation as an important prognostic indicator in cancer, and angiogenesis inhibitors are
attractive as candidate therapeutics.
Absorbance 450nm
3.5
3.0
2.5
2.0
1.5
1.0
0.5
0
– + + – + + MG-132
– – + – – + DMOG
Angiogenesis Resources
Please visit our website for additional resources and products relating to the study of Angiogenesis.
www.cellsignal.com/cstangiogenesis
In ECs, VE-Cadherin signaling, expression, and localization
correlate with vascular permeability and tumor angiogenesis.
VE-Cadherin (D87F2) XP ® Rabbit
mAb #2500: Confocal IF analysis of
HUVEC (VE-Cadherin positive; left) and
HeLa cells (VE-Cadherin negative; right)
using #2500 (green). Actin filaments
have been labeled with DY-554 phalloidin
(red). Blue pseudocolor = DRAQ5 ®
#4084 (fluorescent DNA dye).
FGFR1 binds acidic FGF and basic FGF, both potent promoters of angiogenesis.
A
B
Events
60
50
40
60
50
40
30
FGF Receptor 1
FGF Receptor 1 (D8E4) XP ® Rabbit mAb #9740: IHC analysis of paraffin-embedded human breast carcinoma (A) using
#9740. Flow cytometric analysis of A-204 cells (B) using #9740 (blue) compared to concentration matched Rabbit (DA1E)
mAb IgG XP ® Isotype Control #3900 (red). WB analysis of extracts from A-204 (FGFR1 positive), KG-1a (FGFR1 oncogenic
partner-FGFR1 fusion), A172 (FGFR1 low expression), and HT-29 (FGFR1 negative) cells (C) using #9740 (upper) and β-Actin
(D6A8) Rabbit mAb #8457 (lower).
Angiogenesis Modulators
Select Reviews
Benazzi, C., Al-Dissi, A., Chau, C.H., et al. (2014) Scientific World Journal 2014, 919570. • Claesson-Welsh, L. (2003)
Biochem. Soc. Trans. 31, 20–24. • Ferrara, N., Gerber, H.P., and LeCouter, J. (2003) Nat. Med. 9, 699–676. • Jain, R.K.
(2005) Science 307, 58–62. • Karkkainen, M.J. and Petrova, T.V. (2000) Oncogene 19, 5598–5605. • Koch, S., Tugues, S.,
Li, X., Gualandi, L., and Claesson-Welsh, L. (2011) Biochem. J. 437, 169–183. • Lu, X. and Kang, Y. (2010) Clin. Cancer Res.
16, 5928–5935. • Makanya, A.N., Hluchchuk, R., and Djonov, V.G. (2009) Angiogenesis 12, 113–123. • Meyer, M., Clauss,
M., Lepple-Wienhues, A., et al. (1999) EMBO J. 18, 363–374. • Patel-Hett, S. and D’Amore, P.A. (2011) Int. J. Dev. Biol. 55,
353–363. • Rahimi, N., Dayanir, V., and Lashkari, K. (2000) J. Biol. Chem. 275, 16986–16992. • Ribatti, D., Nico, B., and
Crivellato, E. (2011) Int. J. Dev. Biol. 55, 261–268. • Robinson, C.J. and Stringer, S.E. (2001) J. Cell Sci. 114, 853–865. •
Saharinen, P., Eklund, L., Pulkki, K., et al. (2011) Trends Mol. Med. 17, 347–362. • Sakurai, T. and Kudo, M. (2011) Oncology
81 Suppl 1, 24–29. • Senger, D.R. and Davis, G.E. (2011) Cold Spring Harb. Perspect Biol. 3, a005090. • Weis, S.M. and
Cheresh, D.A. (2011) Nat. Med. 17, 1359–1370.
C
kDa
200
140
100
80
1 2 3 4
Promoters of Angiogenesis
Inhibitors of Angiogenesis
Acidic fibroblast growth factor (aFGF) #5234
ADAMTS1
Angiogenin
Angiostatin
Basic fibroblast growth factor (bFGF) #8910
Endostatin
Epidermal growth factor (EGF) #8916 Interferons (alpha) #8927
Granulocyte colony-stimulating factor (GM-CSF) #8922 Platelet factor 4
Hepatocyte growth factor (HGF)
Prolactin 16 kDa fragment
Interleukin 8 (IL-8) #8921
Thrombospondin
Placental growth factor (PGF)
Tissue inhibitor of metalloproteinase-1 (TIMP-1)
Platelet-derived endothelial growth factor (PEGF)
Tissue inhibitor of metalloproteinase-2 (TIMP-2)
Transforming growth factor alpha (TGF-α) #5495
Tissue inhibitor of metalloproteinase-3 (TIMP-3)
Tumor necrosis factor alpha (TNF-α) #8902
Vascular endothelial growth factor (VEGF) #8908, #8065
FGFR1
FOP-
FGFR1
β-Actin
Lanes
1. A-204
2. KG-1a
3. A172
4. HT-29
168 For Research Use Only. Not For Use in Diagnostic Procedures. See pages 302 & 303 for Pathway Diagrams, Application, and Reactivity keys.
www.cellsignal.com/cstangiogenesis
169