Hydrogen sulfie donor, GYY4137, exhibits anti-atherosclerotic activity in high fat fed apolipoprotein E knockout mice

Prof. Yong Ji, Ph.D., M.D

Atherosclerosis Research Center

Nanjing Medical University

Ross R. N Engl J Med. 1999.

These risk factors most frequently include hypertension, tobacco smoking, diabetes mellitus, obesity, and genetic predisposition; the molecular details of how they work are not yet known.

Atherosclerosis

John W.etal. Antioxid. Redox Signal. 2009.

Endogenously produced gases

John W. et al. Antioxid. Redox Signal. 2009.

Cardiovascular actions of hydrogen Sulfide

Cardiovascular actions of hydrogen Sulfide

Elsey DJ. et al. Cell Biochem Funct 2010.

CBS

H2S

Pyridoxal-5’-phosphate

H2SH2S

John W. et al. Antioxid. Redox Signal. 2009.

3-MST CSE

L-cysteine3-mercaptopyruvate

neurons astrocytes

Biosynthesis of H2S

Role of Hydrogen Sulfide in the Development of Atherosclerotic Lesions in Apolipoprotein

E Knockout Mice

Wang Y, et al. Arterioscler Thromb Vasc Biol 2009.

H2S and Atherosclerosis

ACS14 may prevent the progression of atherosclerosis by downregulating macrophage CX3CR1 expression.

H2S and Atherosclerosis

Zhang H, et al. Eur J Pharmacol 2012.

H2S and Atherosclerosis

H2S inhibited atherogenic modification of purified LDL induced by hypochlorite

H2S induced apoptosis of human aorta smooth muscle cells.

H2S inhibit homocysteine-induced superoxide production in rat thoracic aortic smooth muscle cells.

H2S inhibit adhesion to TNF-α-activated HUVEC.

Yang G. et al. FASEB J. 2004.

Yan et al. BBRC. 2006.

Lagger et al. Free Radic Res. 2007

Pan LL, et al. Plos one 2011.

Controversial findings

H2S induces proinflammatory cytokines synthesis in human monocyte cell line.

H2S regulates leukocyte trafficking in cecal ligation and

puncture-induced sepsis.

Zhang H et al.J Leukoc Biol. 2007.

Zhi L, et al. J Leukoc Biol. 2007.

Limitations

In aqueous solution, NaHS releases large amount of H2S over a period of a few seconds.

Nevertheless, this salt gives poor satisfaction for clinical uses , because the rapid release of H2S may cause adverse effects, such as acute and excessive lowering of blood pressure.

Ideal H2S-donors for therapeutic purposes should generate H2S with slow releasing rates.

Notably, considerable emphasis has also been placed on the use of NaHS as a “tool” to model the biological effects of endogenous H2S.

Characterization of a Novel, Water-Soluble Hydrogen Sulfide Releasing Molecule

(GYY4137): New Insights Into the Biology of Hydrogen Sulfide

Li L, et al. Circulation. 2008.

GYY4137

GYY4137

NaHS GYY4137

Li L et al. Circulation. 2008.

GYY4137 and NaHS

Li L et al. Free Radic Biol Med, 2009.

0-1000umol/L

Anti-inflammation

Macrophages

LPS

GYY4137

NaHS Inflammation

Effect of hydrogen sulfide donors on LPS-induced formation of inflammatory mediators in macrophages

AS

Oxide stress Inflammation Endothelial dysfunction

Hypothesis

GYY4137 inhibited lipid accumulation and experssion of LOX-1 protein induced by Ox-LDL in RAW264.7 Cells

A

B

C

*P <0.05,***P < 0.001 vs CTL ; #P<0.05, ##P<0.01vs ox-LDL treated group. n=4

Effects of GYY4137 on H2S system in RAW264.7 Cells

A B

+P<0.05, c.f. no treatment, *P<0.05, c.f. treatment with oxLDL. n=4

?

ZYJ1122, a structural analogue of GYY4137

ZYJ1122, a structural analogue of GYY4137 , is unable to release H2S or affect lipid accumulation in RAW264.7 Cells

BC

+P<0.05, c.f. no treatment, n=4

ZYJ1122, a structural analogue of GYY4137 , is unable to release H2S or affect lipid accumulation in RAW264.7 Cells

A B

C

+P<0.05, c.f. control, n=4

ZYJ1122, a structural analogue of GYY4137 , is unable to inhibit lipid accumulation in in RAW264.7 Cells

A

B

C

+P<0.05, c.f. no treatment, *P<0.05, c.f. treatment with ox-LDL.n=4-6

GYY4137, but not ZYJ1122, inhibited lipid accumulation in human peripheral blood moncyte-derived macrophages

*P <0.05, **P<0.01 vs CTL ; #P<0.05vs GYY4137.n=6-9.

healthy donors. patient with coronary heart disease.

GYY4137 inhibited chemokine releasing in ox-LDL-treated RAW 264.7 cells

CXCL2 CXCL10 CXCR-4

+P<0.05, c.f. no treatment, *P<0.05, c.f. treatment with oxLDL.n=4-6

GYY4137 prevented ox-LDL induced macrophage iNOS expression

A B

+P<0.05, c.f. no treatment, *P<0.05, c.f. treatment with oxLDL.n=4

GYY4137 prevented oxLDL-induced macrophage NF-κB activation

A B C

D E

*P <0.05, ,**P < 0.01, ***P < 0.001 vs no treatment ; #P<0.05, ##P<0.01 ###P<0.001vs ox-LDL treated group

GYY4137 prevented oxLDL-induced macrophage ICAM-1 and VCAM-1 Experssion

+P<0.05, c.f. no treatment, *P<0.05, c.f. treatment with oxLDL.n=4

Eight-week-old

ApoE-/- mice

High-fat diet

for four weeks

Oil-red O–stained aortic root lesions of mice

Normal food for 30days

GYY4137( 50mg/kg body weight ) or NS, (i.p. )

Anesthetized and sacrificed .

Animal experiment

Effects of GYY4137 on H2S system in apoE-/- mice

‡ P<0.05, c.f. WT mice, §P<0.05, c.f. ApoE-/- + NS 。（ n=5-8 ）

A B

?

GYY4137 has no effect on body weight and plasma lipids

TC:total cholesterol, TG:triglyceride, HDL-C: high-density lipoprotein-cholesterol, LDL-C: low-density lipoprotein-cholesterol. *P ＜ 0.05, **P ＜ 0.01, ***P ＜ 0.001 vs WT. n=5-8 animals/group.

GYY4137 decreases atherosclerotic lesion size in apoE-/- mice

A B

**P ＜ 0.01, ***P ＜ 0.001 vs WT, ###P ＜ 0.01 vs apoE-/- + NS. n=5-8 animals/group.

GYY4137 improved endothelium-dependent relaxation function and activated PI3K/Akt/eNOS Pathway

A B

C D

*P ＜ 0.05, ***P ＜ 0.001 vs WT. #P ＜ 0.05, ##P ＜ 0.01,vs apoE-/- + NS. n=6 animals/group.

?

GYY4137 reduced superoxide formation and LOX-1 expression in aortas of apoE-/- mice

A a b

B

**P ＜ 0.01 vs WT mice. #P ＜ 0.05 vs apoE-/- + NS. n=6 animals/group.

GYY4137 reduced TNF-α, IL-6 and ICAM-1 mRNA expression in aortas of apoE-/- mice

**P ＜ 0.01 vs WT mice. #P ＜ 0.05 vs apoE-/- + NS. n=6 animals/group.

Summary