A general lecture that was given to dermatologists on the importance of the human skin lipidome and the means of thei quantitative analysis

1. Quantitative Analysis of Structural
and Signaling Lipids
A networks approach towards investigating the
human skin lipidome
Dayanjan Wijesinghe, Ph.D.

2. The human skin
 Largest organ of the body
 Protective barrier function against
 Allergens
 Irritants
 Microorganisms
 Water loss
 These functions are primarily through the SC
 Cells (corneocytes) embedded in a highly organized lipid matrix
 Primary lipids are
 Ceramides
 Cholesterols
 Free Fatty Acids

3. There are times when wounds heal…

4. There are times when they don't…
102.821.4
9.8
7.4
11.0
11.3
1.0
Worldwide Wound Incidence
( In Millions !!!)
Surgical Lacerations Burns Pressure Ulcer
Venous Ulcer Diabetic Ulcer Other

5. 1. Snowballing national and international health problem!
2. Very little focused research
3. Externally accessible
4. Treatments easily testable
5. Very fast translatability
6. Multiple research opportunities
a) Diagnostics
b) Treatments
c) Dressings
7. Multiple avenues of funding
a) NIH – NIGMS, NIDDK, NHLBI, NIA
b) DoD – DMRDP(CDMRP) “to advance medical research and
development (R&D) for wounded warriors ..”
c) Department of Veterans Affairs – PU, DFU.
d) Public-Private partnerships
Why study wound healing?
An area where the combination of analytical chemistry, nanotechnology, and
medicine will yield outstanding translational results!

6. Importance of the lipidome
Lipidome
Structural
and Barrier
Signaling
Energetics
Membrane
protein
Interactions

7. Genome
Transcriptome
Proteome
Lipidome
Metabolome
•Total metabolic
status
• Inflammatory status
• Healing potential
• Wound
nutritional status
• Redox balance
• Bacterial load
• Hormone status
•Perfusion status
Why Study the lipidome?
Microbes
Environment
Lifestyle
Phenome
a) Dysregulation of biochemical networks that prevent healing?
b) How can those networks be modified to enable healing?
c) Is what I am doing actually working?

8. Diversity and Variability of
the Skin Lipidome and
Inherent Problems in their
Analysis

9. Janssens et al, 2012
Structure of the skin ceramides

10. Lamellar and lateral lipid organization in human SC
Janssens et al, 2012

11. Imokawa et al, 2008
Tape Stripping for studying the skin lipidome

12. Diversity of the lipidome (>180,000)

13. Isobaric Overlapment
Experiment: EMS scan of Bovine Heart Extract (BHE)
• Lipidomic spectra
are incredibly
complex
• MS spectra alone
is not sufficient for
iidentification
• MS/MS spectra
generated on
precursors in
zones of isobaric
overlap will
contain product
ions form other
isobaric species

14. The need for analytical specificity
Prostaglandin E2
Prostaglandin D2

15. SLIDE COURTESY: Dr. Greg Schultz, Ph.D., Director Wound Healing Institute, University of
Florida

16. Chemometrics
(data reduction)
Identifying
Chemicals of
Interest
Understanding
Biochemical
Relevance
Understanding
Therapeutic
Requirements
Identifying
Diagnostic
Molecules
Mass Spectrometric Characterization of Wounds
Analytical CharacterizationGenome
Transcriptome
Proteome
Lipidome
Metabolome
•Metabolic status
• Inflammatory status
• Healing potential
• Redox balance
•Perfusion status
•Wound nutrition
•Bacterial load
•Hormone status
•Signaling status

17. Lipidomic changes skin wound healing
Dayanjan S Wijesinghe, Charles E Chalfant. (2013) Systems level lipid analysis methodologies for
qualitative and quantitative investigation of lipid signaling events during wound healing. Advances in
Wound Care.

18. Lipids during human wound healing
Wijesinghe DS, Brentnall M, Mietla JA, Hoeferlin LA, Diegelmann RF,
Boise LH, Chalfant CE. (2014) Ceramide kinase is required for a normal
eicosanoid response and the subsequent orderly migration of fibroblasts.
Journal of Lipid Research.

19. What happens when you remove C1P
Wijesinghe DS, Brentnall M, Mietla JA, Hoeferlin LA, Diegelmann RF,
Boise LH, Chalfant CE. (2014) Ceramide kinase is required for a normal
eicosanoid response and the subsequent orderly migration of fibroblasts.
Journal of Lipid Research.

20. Changing C1P alters eicosanoids
Wijesinghe DS, Brentnall M, Mietla JA, Hoeferlin LA, Diegelmann RF,
Boise LH, Chalfant CE. (2014) Ceramide kinase is required for a normal
eicosanoid response and the subsequent orderly migration of fibroblasts.
Journal of Lipid Research.

21. Changing C1P alters eicosanoids
Wijesinghe DS, Brentnall M, Mietla JA, Hoeferlin LA, Diegelmann RF,
Boise LH, Chalfant CE. (2014) Ceramide kinase is required for a normal
eicosanoid response and the subsequent orderly migration of fibroblasts.
Journal of Lipid Research.

22. How does this affect wound healing
Wijesinghe DS, Brentnall M, Mietla JA, Hoeferlin LA, Diegelmann RF,
Boise LH, Chalfant CE. (2014) Ceramide kinase is required for a normal
eicosanoid response and the subsequent orderly migration of fibroblasts.
Journal of Lipid Research.

23. CERK ablation affects fibroblast migration
Wijesinghe DS, Brentnall M, Mietla JA, Hoeferlin LA, Diegelmann RF,
Boise LH, Chalfant CE. (2014) Ceramide kinase is required for a normal
eicosanoid response and the subsequent orderly migration of fibroblasts.
Journal of Lipid Research.

24. Phenotype rescued by exogenous eicosanoids
Wijesinghe DS, Brentnall M, Mietla JA, Hoeferlin LA, Diegelmann RF,
Boise LH, Chalfant CE. (2014) Ceramide kinase is required for a normal
eicosanoid response and the subsequent orderly migration of fibroblasts.
Journal of Lipid Research.

25. Can Lipidomics Be Used to
Predict Successful Wound
Healing?

26. Short answer, YES!

28. Analytical Characterization
Chemometrics
(data reduction)
Identifying
Chemicals of
Interest
Understanding
Biochemical
Relevance
Understanding
Therapeutic
Requirements
Generating synergistic
drug lipid combinations as
nanoliposomes
Delivery of the
therapeutic agent/s
and monitoring the
biochemical
response
Identifying
Diagnostic
Molecules
LCMS Based Diagnostics
Genome
Transcriptome
Proteome
Lipidome
Metabolome
Research Interests - Summary
•Metabolic status
• Inflammatory status
• Healing potential
• Redox balance
•Perfusion status
•Wound nutrition
•Bacterial load
•Hormone status
•Signaling status

29. Research Strategy - Future Directions
Generating synergistic
drug lipid combinations as
nanoliposomes
Delivery of the
therapeutic agent/s
and monitoring the
biochemical
response
Identifying
Diagnostic
Molecules
LCMS Based Diagnostics
Analytical Characterization
Chemometrics
(data reduction)
Identifying
Chemicals of
Interest
Understanding
Biochemical
Relevance
Understanding
Therapeutic
Requirements
Genome
Transcriptome
Proteome
Lipidome
Metabolome

30. Funding and Salary Support
 2014 AB SCIEX Young Investigator Award: “Development of Mass
Spectrometry Applications for Clinical Lipidomic Research”
 U01 – NIH “The utilization of photonics technology to rapidly detect
bioactive lipids associated with preeclampsia development” co-PI
(2015-2019)
 VCU Lipidomics/Metabolomics Core Facility (VLMC) – Deputy Director
 VA CDA1 “Development of Mass Spectrometry Applications for the
Investigation of Wound Healing” 2010-2012
 PI – NRSA T32 Mass spectrometry application for investigating
signaling in tissue injury and repair” 2009-2010
 DoD – “Pre-hospital use of plasma for the treatment of trauma
hemorrhagic shock”

31. Thank You!