Adaptive and Innate Immune Function Variation in Cloned Pigs

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Adaptive and Innate Immune Function Variation in
Cloned Pigs

• Fall 2006 Research Update
• Erin Luetkemeier

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• The objective of this study is to determine the
  relative influence of genetic makeup and the
  environment on the variation of porcine adaptive
  and innate immune responses.
• Lymphocyte function
• Innate immune function
• Adaptive immunity

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Identical PBMC Response to polyclonal T cell
activators
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Environmental Impact on IFN-a production
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Environmental Impact on IFN-a production
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IFN-a Conclusions

• Both sets of clones and the controls
• demonstrated significant differences when
  comparing their responses to CpG
• Outdoor clones versus indoor clones
• When comparing their coefficient of variations,
  it was observed that the indoor clones
  demonstrated less immune variation than the
  outdoor clones.
• Through F-test evaluation (F-critical 0.05, 3,3
  of 19.16) there were no significant values

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Phenotypic Analysis of PBMC

• CD4CD8-, CD4-CD8, and CD4CD8
• Identify the presence of T and B cells
• CD 29 high and low
• Identify the presence of activated T or memory T
  cells

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PBMC Phenotypes
Dead
Live
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PBMC Phenotypes
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PBMC Phenotypes




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Phenotypic analysis conclusions

• Outdoor reared clones and their age-matched
  controls
• F-test (F-critical 0.05, 4,4 of 9.28) statistical
  analysis was performed
• None of the values obtained were statistically
  significant
• Outdoor versus indoor reared clones
• F test (F-critical value of 19.16 0.05, 3,4)
• significantly less immune variation observed in
  the indoor reared clones as compared to the
  outdoor reared clones (CD 8sp and CD8sphi)

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3. Adaptive immunity

• Variation of Immune responses following
  stimulation to crude extract of Ascaris suum
• Tested using allergy skin test

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Ascaris antigen challenge

• Determine immediate allergic response to Ascaris
• Measured at 30 minutes
• Determine delayed type hypersensitivity reaction
• Measured at 24 hours

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Allergy Comparison
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Ascaris Conclusions

• Basic statistics
• there were no significant differences (F-critical
  0.05, 4,4 is 9.28) between the allergic responses
  observed between the outdoor reared clones with
  the outdoor reared controls
• Cloned pigs reared outdoors respond similarly to
  cell-mediated antigen challenges as age-matched
  out-bred pigs

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Conclusions

• When using cloned animals, if the phenotypic
  analysis is extended to blood parameters, there
  are two overall classes of traits observed
• Class I
• Appears to be related to innate immune responses
• reduced variation seen in cloned pigs, possible
  due to tight genetic control and little
  epigenetic influence
• Class II
• Appears to be related to adaptive immunity
• as much variation seen in the cloned pigs as in
  the control pigs, possibly due to epigenetic
  disregulation from the cloning process or
  susceptibility to environmental stimuli

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Future Aims

• Epigenetics
• Measure innate and adaptive immune function on
  new clones
• All from same uterine environment
• Attempt to show they are epigenetically identical

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Environmental Impact on IFN-a production

• TGE
• Transmissible gastroenteritis virus (coronavirus)
• TLR7 (under speculation)
• acting through a dsRNA produced during an
  infection
• a protein in TGE acts directly on NIP cells
  (Natural Interferon alpha Producing Cells)
• CpG
• DNA methylation motifs
• TLR9 is the receptor for CpG motifs
• Cells respond by production of large quantities
  of INF-a