Levosimendan Relaxes Thoracic Aortic Smooth Muscle in Mice by Inhibiting PKC and Activating Inwardly Rectifying Potassium Channels.

ABSTRACT: Studies have examined the therapeutic effect of levosimendan on cardiovascular diseases such as heart failure, perioperative cardiac surgery, and septic shock, but the specific mechanism in mice remains largely unknown. This study aimed to investigate the relaxation mechanism of levosimendan in the thoracic aorta smooth muscle of mice. Levosimendan-induced relaxation of isolated thoracic aortic rings that were precontracted with norepinephrine (NE) or KCl was recorded in an endothelium-independent manner. Vasodilatation by levosimendan was not associated with the production of the endothelial relaxation factors NO and PGI2. The voltage-dependent K+ channel (KV) blocker (4-aminopyridine) and selective KCa blocker (tetraethylammonium) had no effect on thoracic aortas treated with levosimendan, indicating that KV and KCa channels may not be involved in the levosimendan-induced relaxation mechanism. Although the inwardly rectifying K+ channel (Kir) blocker (barium chloride) and the KATP channel blocker (glibenclamide) significantly inhibited levosimendan-induced vasodilation in the isolated thoracic aorta, barium chloride had a much stronger inhibitory effect on levosimendan-induced vasodilation than glibenclamide, suggesting that levosimendan-induced vasodilation may be mediated by Kir channels. The vasodilation effect and expression of Kir 2.1 induced by levosimendan were further enhanced by the PKC inhibitor staurosporine. Extracellular calcium influx was inhibited by levosimendan without affecting intracellular Ca2+ levels in the isolated thoracic aorta. These results suggest that Kir channels play a more important role than KATP channels in regulating vascular tone in larger arteries and that the activity of the Kir channel is enhanced by the PKC pathway.

Biosynthesis-inspired mining and identification of untapped alkaloids in Camptotheca acuminate for enzyme discovery using ultra-high performance liquid chromatography coupled with quadrupole-time of flight-mass spectrometry.

Leaves, flowers, fruits and stems (44 sample groups) were collected from mature Camptotheca acuminate during 2017.3-2018.3 and classified by ultra-high performance liquid chromatography coupled with quadrupole-time of flight-mass spectrometry based metabolomics. One hundred metabolites including forty-seven alkaloids, fifteen terpenes, thirty-two polyphenols and six other metabolites were rapidly identified through the in-house database alignment at first glance. Thirty-three alkaloids classified into five groups including camptothecin group (CG1-13), pumiloside group (PG1-5), strictosidinic acid group (SG1-3), vincosamide group (VG1-7), and a new hybrid group, vincosamide-camptothecin group (VC1-5) were mined and further characterized by MS/MS analyses. The identification of two untapped biosynthetic precursors, 2-hydroxypumiloside (PG2) and 16­hydroxy­15, 16-dihydrocamptothecoside (CG3), along with sixteen new alkaloids enables us for a better understanding of camptothecin biogenetic reasoning. The underlying enzymes involved in camptothecin biosynthesis were also proposed according to the guiding metabolic map, thus purposefully mining of enzymes involved in the downstream biosynthetic pathway of camptothecin could be initiated with the help of this map.

Possible clues for camptothecin biosynthesis from the metabolites in camptothecin-producing plants.

The plant derived camptothecin (CPT) is a pentacyclic pyrroloquinoline alkaloid with unique antitumor activity. Successive discoveries of new CPT-producing plants occurred in recent years due to market demands. The scattered distribution among angiosperms drew researchers' attention. The aim of this review is to appraise the literature available to date for CPT distribution and the phytochemistry of these CPT-producing plants. Metabolite comparative analyses between the plants were also conducted for tracking of possible clues for CPT biosynthesis. Forty-three plant species in total were reported to possess CPT-producing capability, and one hundred twenty-five alkaloids classified into three major categories are summarized herein. Metabolite comparative analysis between these plants suggests the probability that the formation of the central intermediate for CPT biosynthesis has multiple origins. A more complete biogenetic reasoning for CPT and its structural homolog was delineated based on this fragmentary phytochemical evidence from a chemical point of view. Furthermore, an in-house compound database was constructed for further metabolomic analysis.