Corpus cavernosal smooth muscle relaxation effect of a novel AMPK activator, beta-lapachone.
BACKGROUND
Adenosine monophosphate-activated protein kinase (AMPK) activation is suggested to relax smooth muscle by endothelial nitric oxide synthase (eNOS) phosphorylation.


OBJECTIVE
To assess the mechanism and effect of a novel AMPK activator, beta-lapachone, upon cavernosal smooth muscle relaxation and the therapeutic potential for erectile dysfunction.


METHODS
Human umbilical vein endothelial cells (HUVECs) were treated with beta-lapachone. The lysates were blotted with specific antibodies for phosphorylated AMPK (p-AMPK) or phosphorylated eNOS (p-eNOS). The membranes were re-blotted for total AMP total eNOS, or beta-actin. The eNOS activity was measured by the conversion of L-14C-arginine to L-14C-citrulline in HUVECs lysates. In a separated experiment, cavernosal strips from New Zealand white rabbits were harvested for organ bath study and the relaxation effect of beta-lapachone on phenylephrine-induced contracted strips was evaluated and compared with sodium nitroprusside, zaprinast, metformin, and aminoimidazole carboxamide ribonucleotide (AICAR). Methylene blue and L-NAME were used to assess the inhibition of cyclic guanosine monophosphate/nitric oxide pathway. Zinc-protoporphyrin-IX (ZnPP) was also used to investigate the contribution of mevalonate pathway.


METHODS
The expression of p-AMPK, p-eNOS, AMPK and eNOS induced by beta-lapachone in HUVECs study and the percent relaxation of cavernosal tissue in organ bath study.


RESULTS
Beta-lapachone clearly induced AMPK phosphorylation and, as a consequence, eNOS phosphorylation in HUVECs. Beta-lapachone-induced upregulation of eNOS activity was also observed in HUVECs and steadily increased up to 1 hour. In organ bath study, beta-lapachone significantly relaxed the phenylephrine pretreated strips in a dose-dependent manner. This relaxation effect was not totally blocked by methylene blue or L-NAME. After removing endothelium, the relaxation was totally blocked by ZnPP.


CONCLUSIONS
A novel AMPK activator, beta-lapachone has a strong relaxation effect on precontracted cavernosal smooth muscle strips in the rabbit. And phosphorylation of AMPK and eNOS strongly related to the action of beta-lapachone. Mevalonate pathway also might be considered as a suggestive mechanism.
Bae
Jae Hyun
JH
Department of Urology, Korea University Medical Center and Korea University Institute of Regenerative Medicine, Seoul, Korea.
Kim
Jin Wook
JW
Kweon
Gi Ryang
GR
Park
Myoung Gyu
MG
Jeong
Kyeong-Hoon
KH
Kim
Je Jong
JJ
Moon
Du Geon
du G
D055372:AMP-Activated Protein Kinases / Q000378:metabolism*; D000818:Animals; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D008297:Male; D009126:Muscle Relaxation / Q000187:drug effects; D009130:Muscle, Smooth / Q000187:drug effects; D009285:Naphthoquinones / Q000494:pharmacology*; D009465:Neuromuscular Agents / Q000494:pharmacology*; D052250:Nitric Oxide Synthase Type III / Q000096:biosynthesis / Q000378:metabolism*; D010413:Penis / Q000187:drug effects* / Q000378:metabolism; D010766:Phosphorylation; D011817:Rabbits
D009285:Naphthoquinones; D009465:Neuromuscular Agents; C014638:beta-lapachone; D052250:Nitric Oxide Synthase Type III; D055372:AMP-Activated Protein Kinases