Enhanced Lipid Deposition and Foam Cell Formation in Coronary Artery - - PowerPoint PPT Presentation

Fan Zhang, Min Xu, Krishna M Boini, Min Xia and Pin-lan Li Department of Pharmacology & Toxicology Medical College of Virginia Campus Virginia Commonwealth University

Enhanced Lipid Deposition and Foam Cell Formation in Coronary Artery Wall by Deletion of CD38 Gene

NAADP and Lysosomal TRP-ML1

• NAADP (Nicotinic Acid Adenine Dinucleotide Phosphate)

is an newly

• discovered

intracellular Ca2+ second messenger that participates in a variety

• f

pathophysiological processes.

• This nucleotide has been found to release Ca2+ from

Lysosomal TRP-ML1 (Transient Receptor Potential- Mucolipin 1) channel.

• NAADP is mainly produced through an enzyme, CD38

ADP - ribosylcyclase, by catalyzing the exchange of nicotinamide in NADP as a substrate with nicotinic acid.

Lysosomal TRP-ML1 and Lipids Deposition

• TRP-ML1 is a specific lysosome membrane - located

non-specific Ca2+ release channel.

• Clinically, TRP-ML1 mutation has been confirmed as a

critical genetic determinant for the lysosomal lipids storage diseases, such as Mucolipidosis type IV.

Lysosomal Cholesterol Accumulation in Atherosclerosis

• Advanced

atherosclerotic foam cell formation has features of an acquired lysosomal storage disorder.

• The

accumulated lipids in lysosomes are rich in cholesteryl ester and free cholesterol, and the retarded free cholesterol efflux has been found to play a critical pathogenic role in lysosome lipids deposition.

Abnormal CD38-TRP-ML1 regulation

• f

lysosome function importantly contributes to lipid deposition and foam cell formation in coronary artery wall.

Hypothesis

Histological Evidence of Atherosclerosis in CD38-/- Mouse Coronary Artery

Western Diet Normal Diet CD38+/+ CD38-/- Western Diet Normal Diet CD38+/+ CD38-/-

A B

40 x

Electron Microscope Evidence of Atherosclerosis in CD38-/- Mouse Coronary Artery

+/+

Normal Diet CD38+/+ Western Diet Western Diet CD38-/- CD38+/+ X 12,000

A B C

Conversion Rate of NADP (nmol/min/mg protein) CD38+/+ CD38-/- CD38-/-+Rescue 0.00 0.04 0.08 0.12 0.16

NAADP Conversion Rate Was Decreased in CD38-/- Macrophage by HPLC Assay

NADP+ Nicotinic Acid NAADP CD38

*

Enhanced Lipids Deposition in CD38-/- Macrophages

2+

CD38+/+ CD38-/- Basal

• xLDL

CD38+/+ Relative Oil Red Extract Absorption 1 2 3 4 5 6

Basal

• x-LDL

CD38-/-

*

A B

NAADP Signaling Antagonists Increased Lipids Deposition in CD38+/+ Macrophages

Basal

• xLDL

PPADS+oxLDL Nicot+oxLDL Baf+oxLDL

Ox-LDL 1 2 3 4 5 6 7

* * *

#

Relative Oil Red Extract Absorption

A B

Ctrl Nicot PPADS Baf Relative Fluorescence Intensity 1 2 3 4 TRP-ML1 shRNA

* * * * Further Confirm NAADP Signaling Antagonists Increased Lipids Deposition in CD38+/+ Macrophages A B

CD38 Gene Transfection Attenuated Foam Cells Formation in CD38-/- Macrophages

CD38-/- CD38-/- CD38+/+

CD38-GFP GFP GFP

CD68/Alexa Fluor 594 Overlay Fluorescence Intensity Ratio CD68-Fluor 594/GFP CD38+/+ CD38-/- CD38-/- + CD38-GFP 0.0 0.5 1.0 1.5 2.0 2.5 3.0 GFP

* A B

NAADP Signaling Antagonists Increased Foam Cell Formation in CD38+/+ Macrophages

Relative CD68-Fluor 594 Intensity 2 4 6 8 10 12 14 16

Vehicle

• x-LDL

Nicot+

• x-LDL

PPADS+

• x-LDL

Baf+

• x-LDL

TRP-ML1shRNA+

• x-LDL

A B * * * *

• xLDL

NAADP Increased Cholesterol Efflux from Lysosomes in CD38-/- Macrophages

Cholesterol /Filipin Overlay Lamp-1/ Alexa Fluor 555 Basal

• xLDL
• xLDL
• xLDL+

TRP-ML1shRNA

• xLDL

+CD38 rescue NAADP(100nM) Normalized Cholesterol/Filipin Fluorescence Intensity 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 NAADP(100nM)

*

#

A B

Conclusion

CD38-NAADP-TRP-ML1 signaling pathway play a critical role in the regulation of lysosomal lipids deposition in macrophages and foam cell formation.

Clinical Relevance

CD38-NAADP/TRP-ML1 pathway may act as an important signaling in the pathogenesis of atherosclerosis. This signaling pathway could be an important therapeutic target for the prevention and treatment of atherosclerosis.

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