Genetic reprogramming in pathways of colonic cell maturation induced by short chain fatty acids: Com

1
Genetic reprogramming in pathways of colonic cell
maturation induced by short chain fatty acids
Comparison with Trichostatin A, Sulindac, and
Curcumin and cmplications for chemoprevention of
colon cancer.
Presented by Johann Campbell
Mariadason, J., G., Corner, and L. Augenlicht.
2000. Genetic reprogramming in pathways of
colonic cell maturation induced by short chain
fatty acids comparison with trichostatin A,
sulindac, and curcumin and implications for
chemoprevention of colon cancer. Cancer Res.
604561-4572.
2
Background

• Butyrate produced in intestine by microbial
  fermentation
• Butyrate is an inhibitor of HDAC activity
• TSA also inhibits HDAC activity
• Sulindac induces G0-G1 cell cycle arrest and
  apoptosis
• Curcumin induces G2-M cell cycle arrest and
  apoptosis

3
Cell Cycle
4
Investigating genomic activity

• Cell lines used were SW60 and Caco-2
• Microarray technology
• Probe labels included Cy3 and Cy5
• 8063 arrays used out of a total of 18,000
  available
• Western Blot (Western Immunoblot)

5
Microarray Technology

• Micro slide coated with DNA sequences from a
  database of about 18,000
• mRNA from desired genes are labeled fluorescent
  dye
• 2 groups of labeled mRNA used Control and Test
• Labeled mRNA allowed to hybridize on microslide
• Microslide washed to remove unbound mRNA
• Fluorescence measured for ratios of bound
  Control Test

6
Courtesy National Human Genome Institute
7
Microarray Results

• In the control group the distribution of
  sequences remained tightly around the mean
• In the treated test groups, there was a spread of
  ratios
• The spread indicates gene activity
• The width of the spread is co-relational to the
  activity

8
(No Transcript)
9
Gene activity

• A Treeview and Cluster program was used to
  visualize genetic reprogramming
• Red indicates an increase in gene expression
• Green indicates a decrease in gene expression
• Activity among gene clusters are compared

10
(No Transcript)
11
Response to Butyrate

• After 48 hours about 256 sequences were increased
  in expression by butyrate
• About 333 were repressed after 48 hours
• Of the 589 altered sequence, only 345 are named

12
Response to all reagents

• Some reagents displayed similar activity patterns
  at similar times
• Some reagents displayed similar activity at
  different times
• Time similarities/differences were graphed using
  the N-Euclidean distances

13
(No Transcript)
14
Histone Acetylation

• Histone acetylation peaks at different times
• TSA peaks at 2 hours
• Butyrate peaks at 16 hours

15
(No Transcript)
16
Butyrate and Sulindac activity

• Both butyrate and sulindac stimulate similar cell
  cycle arrest
• Both are similar in inducing apoptosis
• Both displayed progressive gene activity with time

17
(No Transcript)
18
Further analysis of Butyrate and Sulindac

• Sulindac and butyrate displayed some similarity
  in the type of gene activity
• About 145 were common
• About 53 showed of the 143 showed opposite
  reactions

19
(No Transcript)
20
Analysis of gene recruitment of butyrate and
sulindac

• Class of genes active in signal transduction and
  transcription alteration
• 2 groups defined group 1 signaling (33 increased
  expression) and group 2 signaling (45 repressed
  expression)

21
(No Transcript)
22
(No Transcript)
23
Cell cycle response

• 61 genes involved in cell cycle regulation were
  chosen
• Euclidean distances calculated for each response
  maximally with time
• Butyrate was similar to TSA in affecting these
  genes
• Butyrate was less like curcumin in affecting
  genes responsible for cell cycle regulation
• The same analysis was done on cells from Caco-2
  lines
• Butyrate response was similar to arrested Caco-2
  cells

24
(No Transcript)
25
Conclusion

• Butyrate was found to play a role in gene
  activity in colonic cells
• Butyrate is also able to induce a cell cycle
  arrest by its activities on cell cycle regulating
  genes
• Butyrate shows similar activities on genes
  similar to TSA, opposite to curcumin
• Butyrate had limited similar effects compared to
  sulindac

26
Questions