the role of cytokines and paracellular permeability

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the role of cytokines and paracellular
permeability
CROHNS DISEASE
Department of Gastrointestinal Sciences,
University of Manchester JEN VIKEBO
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THE GASTROINTESTINAL EPITHELIUM

• The largest interface between a person and their
  external environment
• Allows transport of essential nutrients into the
  bloodstream whilst excluding potentially harmful
  toxins
• Flux through the epithelial epithelium
• TRANSCELLULAR
• PARACELLULAR

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TIGHT JUNCTIONS

• The epithelium is composed of epithelial cells
  connected by tight junctions

Lumen

• Multiprotein complexes
• Determine permeability through paracellular
  pathway
• Barrier function!

Tight Junction
TRANSCELLULAR
PARACELLULAR
Bloodstream
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CROHNS DISEASE

• Inflammatory Bowel disease
• Particularly common in younger people in
  developed countries
• Causes psychological problems
• Major impact on quality of life
• Symptoms severe abdominal pain, diarrhoea,
  weight loss, secondary complications such as skin
  and joint problems

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CROHNS DISEASE

• Multiple factors lead to progression
• Increased intestinal permeability
• Aberrent mucosal immune system
• -High levels of cytokines TNF-? and IL-1?
• Patients are often found to have mutated
  carnitine transporter, OCTN1

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PROJECT AIMS

• To investigate how TNF-? and IL-1? contribute to
  the development of CD using Caco-2 cells
• effects on transepithelial electrical resistance
  (TEER)
• effects on tight junction protein expression
• effects on expression of carnitine transporter
  OCTN1

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New Techniques!
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RESULTS TNF-?

• TNF-? reduces TEER, a marker of tight junction
  integrity

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RESULTS TNF-?
CONTROL
TREATED
Claudin 1

• No detectable changes in expression of tight
  junction proteins claudins 1, 3, 4 and occludin
• No detectable change in expression of carnitine
  transporter OCTN1

Claudin 3
Claudin 4
Occludin
OCTN1
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RESULTS IL-1?

• IL-1? reduces TEER, a marker of tight junction
  integrity

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RESULTS IL-1?
CONTROL
TREATED
CONTROL
TREATED
Claudin 1
OCTN1
OCTN1
Claudin 3

• No detectable changes in expression of Claudins
  1, 3, 4 and Occludin
• Pattern of de-phosphorylation in ZO-1
• Decrease in expression of OCTN1

Claudin 4
Occludin
ZO-1
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CONCLUSIONS TNF-?

• TNF-? causes a decrease in TEER
• TNF-? mechanism of reducing TEER not likely via
  changes in Claudins 1, 3, 4 and Occludin
• No link found between TNF-? and OCTN1

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CONCLUSIONS IL-1?

• IL-1? reduces TEER
• Results suggest a de-phosphorylation in ZO-1
  indicating possible mechanism of this decrease in
  TEER
• ZO-1 phosphorylation previously linked to barrier
  permeability although literature controversial
• Decrease in OCTN1 observed indicating IL-1? could
  putatively regulate the transcription of this
  transporter protein

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THE FUTURE!

• Look at influence of TNF-? and IL-1? on other
  tight junction proteins
• Assess flux of paracellular tracers through
  monolayers
• Look at promoter activity and gene expression of
  SLC22A4 encoding OCTN1 in the presence of IL-1?
• Other work on OCTN1!

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CONCLUDING REMARKS!

• Understanding the underlying molecular
  mechanisms of Crohns Disease is the key to
  understanding the disease itself.
• Hopefully this research will provide insight into
  these mechanisms and lead to new avenues for
  further research and development of therapeutic
  targets.
• There is currently no cure for Crohns Disease!

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ACKNOWLEDGEMENTS

• Huge thanks to everyone in the department of
  Gastrointestinal Sciences especially Cath ONeill
  and Daniel Lambert!