Hypoxia

Question 1

Describe the role of hypoxia in disease.

Define hypoxia & its roles (1) (1/8)

Answer 1

• condition of body where oxygen is N/A in sufficient amounts to maintain adequate homeostasis ∴ = starved of oxygen
• can be result of inadequate oxygen delivery to T due to
  1) low blood supply
  2) low [O2] in blood (hypoxemia)
• can have a range of physiological implications for a range of T & cell types
  
  • peripheral smooth muscle cells vasodilate
  • pulmonary smooth muscle cells vasoconstrict
• due to differences in signalling events
• regulates many canonical pathways
  
  • acute H: sig pathways that promote cell survival upregulated
  • chronic H: sig pathways that promote cell death upregulated
• functions controlled by Hypoxia Inducible Factors (HIFs)
  
  • family of TCF that = stabilised during H, degraded during normoxia
  • stabilised & dimerised HIF translocate to nucleus to bind promotes of HIF-pathway target genes via the Hypoxia Response Element
  • allows HIF-DNA-binding complex → gene TC
  • plethora of changes in cell behavioural output
• acute mediated via HIF1α
• chronic mediated via HIF2α
• both = reported to be ass with tumour progression & aggressive cancer pheno
• chronic has also been reported to contribute to cancer regression, therefore role of hypoxia in disease = complicated

Question 2

Describe the role of hypoxia in disease.

Define a disease (1) (2/8)

Answer 2

• cancer = group of diseases instigated by uncontrollable division of cells & their subsequent spread into surrounding T
• Hypoxic zones arise due to imbalance b/w O2 supply & consumption in a solid malignant tumour, where O2 <2%
• several mech can contribute to development of H in tumour microenviro
  
  • Perfusion-restricted hypoxia (aka acute/intermittent): transient w/ insuff delivery of O2 due to aberrant BV undergoing cycles of dilation/constriction → heterogenous cell population in tumour
  • Diffusion-restricted hypoxia (aka chronic): sustained restriction in delivery & diffusion due to abnormal vascular network
• acute can occur when tumour cells prolif & expand beyond 70 µm from pre-existing BV → inadequate delivery
• @ > 180 µm, cells = necrotic
• Anemic H in tumours can be therapy-induced
• tumours ≠ compensate for ↓ [O2], so = susceptible to anemic H
• proliferating cells pass & grow beyond the diffusion limit
• during H conditions, >100 genes = activated & TC’d that are involved in
  
  • angiogenesis, metabolism, apoptosis, prolif
• H can modify tumour microenviro & select for cells that can survive during periods of oxygen & nutrient deprivation
• encouraging metastasis

Question 3

Describe the role of hypoxia in disease.

Define hypoxia in cancer and how it contributes to the pathophysiology of the disease (involvement in cancer stages; intermittent H involved; failures of other studies) (1) (3/8)

Answer 3

• upregulation of HIF-1α = implicated in tumour growth, invasion, angio, therapy-R & metastasis in a number of cancer types
  
  • ovarian, breast, lung cancer
• metastasis = rare & has many bottlenecks
• it is in the final stage in cancer development
• spread of cancer cells from 1° site → distinct 2° location
  
  • intravasation & extravasation through BV
• establish a new microenviro where they may elicit induction of angiogenesis → nutrient-rich niche
• @ some point the tumour will be so large that centre is too far from BV, so H pathways may respond
• defining what makes tumour cells susceptible to metastasis = vital area of research
  
  • try & modify associated targetable pathways/ genes/ processes
  • however limited studies (inc the specific one) discuss/observe the involvement of EMT, wh/ = necessary for cell mobility (discussed later)
• study was conducted to investigate the different consequences of exposing murine models of breast cancer to intermittent & chronic H.
• findings (below) demonstrated that intermittent H, but not chronic, induced a number of genetic, molecular, biochemical and cellular changes that facilitated tumour cell survival, colonisation & creation of a permissive microenviro, that allows metastatic growth:
  
  • promoted clonal diversity
  • upregulated metastasis-associated gene XPS’ion
  • induced a pro-tumorigenic secretory profile
  • increased stem cell-like marker XPS’ion
  • higher freq of tumour-initiating cells
• Prior attempts to characterise the impact of H had failed because they had inappropriate models
• one study done earlier used immune-deficient mice in intermittent H
  
  • IS has been obvs to be a strong regulator of metastasis, ∴ ≠ a ✓
    model

Question 4

Describe the role of hypoxia in disease.

Intermittent hypoxia induces a greater metastatic potential (1) (4/8)

Answer 4

• greater ability to colonise the lung & grow into overt tumour loci after being injected into experimental models of metastasis
• lungs of injected mice contained a ↑ # & larger metastases vs normoxia controls or chronic H-treatment groups
• integrity of the H response pathway is maintained by intermittent-H-treated cells
• Intermittent H ≠ interfere w/ cells ability to respond to H
• Any cells that are challenged w/ H will undertake usual pathways
  
  • also means tumour sizes that exceed diffusion limit will be subject to H conditions
  • according to this paper, this increases the chance of tumour metastasis

Question 5

Describe the role of hypoxia in disease.

Intermittent hypoxia induces senescence at a lower frequency compared to chronic-hypoxic cells (1) (5/8)

Answer 5

• +ive staining for senescence associated β-galactosidase obvs in chronic & intermittent H-exposed cells
• % of β-galactosidase +ive cells ↑’ed after complete chronic hypoxic treatment
• qRT-PCR of mRNA for senescence markers ✓ this obvs
• Intermittent H may mediate lung colonisation by inducing a senescence-associated pro-tumorigenic pheno
• ≈ generate a heterogenous tumour by encouraging clonal diversity

Question 6

Describe the role of hypoxia in disease.

Intermittent hypoxia induces changes in expression of pro-metastatic genes & increases secretion of pro-tumorigenic cytokines (↑ MMP9 & CD1; ↓ IFN-γ and TIMP-1) (1) (6/8)

Answer 6

• IH ↑ XPS’ion of genes previously described to predict lung metastasis in breast cancer
• both IH- & CH-treated cells had sig ↑ XPS’ion of genes referred to for their oncogenic nature
  
  • ISG20, ↑’ed in both b/ ↑er in IH-treatment
  • ~ have a role in clear cell renal cell carcinoma where it ≈ promote prolif & metastasis via upregulation of MMP9 & cyclin D1
    
    • MMP9 involved in breakdown of ECM (necessary for meta), ∴ ↑’ed XPS’ion may promote migration & invasion of breast cancer cells
    • Cyclin D1 = regulator of CDK kinases that mediate the cell cycle. Mutations, amplifications & over-XPS’ion of CD1 have been obvs in a variety of tumours & contributes to tumorigenesis
• SUGGESTS H induces TC’ional Δ’s that ≈ contribute to metastatic capabilities
• significant ↓ in secretion of IFN-γ & TIMP-1 following IH- or CH-treatment
• IH = significant ↑ in IL6 secretion ≈ potentially indicative of IH inducing an immuno-suppressive enviro (b/c IL6 = stimulates acute phase responses, hematopoiesis, & immune reactions)
• despite cells ≠ showing an overt EMT pheno, they still showed ↑ MMP9, so may be able to migrate somehow

Question 7

Describe the role of hypoxia in disease.

Intermittent hypoxia induces tumour-initiating capability (1) (7/8)

Answer 7

• CH- and IH-treatment led to ↑ XPS’ion of mammary stem cell genes
  
  • Aldh1, Pai
• and luminal lineage genes
  
  • ELF5, GATA3
• compared to normoxic cells
• XPS’ion of GATA3 & Pai = sig ↑er after IH exposure
• IL6, Notch1, & Nanog gene XPS’ion was increased only by IH treatment
• to assess whether the ↑ in XPS’ion of stem-cell markers in IH would impact cancer-forming capacity in vivo, limiting dilution assays were performed
  
  • mice = injected w/ a decreasing # of cells
  • only IH cells had a greater ability to generate tumours > normoxic
  • IH induced a stat sig 3- to 4-fold ↑ in tumour-initiating cells > normoxic & CH-treatments

Question 8

Describe the role of hypoxia in disease.

Hypoxic cells only partially establish EMT (1) (8/8)

Answer 8

• an overt EMT pheno ≠ obvs in either CH/IH
• other studies have reported H-induced snail XPS’ion (inducer of EMT)
• has been reported that H induces XPS’ion of snail in breast cancer cell lines
• after LT exposure, Snail translocates → nucleus & exerts EMT-inducing functions
  
  • downreg of E-cadherin
  • upreg of vimentin lvls
• but these results vary across diff cancer cell lines, ∴ Snail may be an important regulator of EMT in some contexts, but inferior to other EMT regulators (Slug, Twist) in other cell lines
• Maybe then, the specific study did not observe the 3D culture for long enough to observe the EMT-inducing func of Snail, or perhaps it was an inferior regulator
• OR maybe Snail induces a migratory pheno independent of EMT induction in breast cancer cell lines (BCCL).
  
  • Snail induces a migratory pheno in epithelial-like & mesenchymal cell lines
  • OverXPS’ion of Snail in one BCCL resulted in an ↑’ed migratory ability despite maintenance of E-cadherin
    
    • E-cadherin XPS’ion ~ ≠ exclude migratory behaviour
• ∴ the obvs of maintained E-cadherin XPS’ion in the specific study may mean that Snail is driving migration, however it is independent of an EMT induction in this cancer cell line