Diltiazem Inhibits Coronary Spasm via Inhibition of Cav1.2Phosphorylation and Protein Kinase C Activation in a Mouse Model of Coronary Spastic Angina.

Calcium antagonists are used for coronary spastic angina (CSA) treatment. We previously identified a phospholipase C (PLC) -δ1 gene variant that results in enhanced PLC activity in patients with CSA and developed a CSA animal model by generating vascular smooth muscle cell-specific human variant PLC-δ1 overexpression (PLC-TG) mice. In this study, we investigated the molecular mechanism of CSA using the PLC-TG mice and the inhibitory effect of a calcium antagonist, diltiazem hydrochloride (DL).We treated the PLC-TG and wild-type (WT) mice with oral DL or trichlormethiazide (TM) (control) for 2 weeks. Ergometrine injection-induced coronary spasm was observed on the electrocardiogram in all 5 PLC-TG mice treated with TM, but only in 1 of 5 PLC-TG mice treated with DL. Voltage-dependent calcium channel (Cav1.2) phosphorylation and protein kinase C (PKC) activity were enhanced in the aortas of PLC-TG mice treated with TM. DL treatment significantly inhibited Cav1.2 phosphorylation and PKC activity. Although total Cav1.2 expression was similar between WT and PLC-TG mice treated with TM, DL treatment significantly increased its expression in PLC-TG mice. Furthermore, its expression remained high after DL discontinuation. DL and PKC inhibitor suppressed intracellular calcium response to acetylcholine in cultured rat aortic smooth muscle cells transfected with variant PLC-δ1.These results indicate that enhanced PLC activity causes coronary spasm, presumably via enhanced Cav1.2 phosphorylation and PKC activity, both of which were inhibited by DL. Enhanced total Cav1.2 expression after DL discontinuation and high PKC activity may be an important mechanism underlying the calcium antagonist withdrawal syndrome.

Leishmanicidal active constituents from Nepalese medicinal plant Tulsi (Ocimum sanctum L.).

In the course of screening leishmanicidal active compounds from Asian and South American medicinal plants, a Nepalese medicinal plant, Tulsi (Ocimum sanctum L.), showed strong activity. We therefore studied the isolation and structural elucidation of the active constituents from O. sanctum L. From the ethyl acetate soluble fraction of the plant, seven new novel neolignan derivatives were isolated along with 16 known compounds. The structures of the new compounds (1-7) were elucidated as 6-allyl-3',8-dimethoxy-flavan-3,4'-diol (1), 6-allyl-3-(4-allyl-2-methoxyphenoxy)-3',8-dimethoxyflavan-4'-ol (2), 5-allyl-3-(4-allyl-2-methoxyphenoxymethyl)-2-(4-hydroxy-3-methoxyphenyl)-7-methoxy-2,3-dihydrobenzofuran (3), 1,2-bis(4-allyl-2-methoxyphenoxy)-3-(4-hydroxy-3-methoxyphenyl)-3-methoxypropane (4), 1-(4-hydroxy-3-methoxyphenyl)-1,2,3-tris(4-allyl-2-methoxyphenoxy)propane (5), 1-allyl-4-(5-allyl-2-hydroxy-3-methoxyphenoxy)-3-(4-allyl-2-methoxyphenoxy)-5-methoxybenzene (6), and 3-(5-allyl-2-hydroxy-3-methoxyphenyl)-1-(4-hydroxy-3-methoxyphenoxy)-prop-1-ene (7) by means of (1)H-NMR, (13)C-NMR, and 2D-NMR spectral data. Some of these compounds showed leishmanicidal activity.

Evaluation of human peroxisome proliferator-activated receptor (PPAR) subtype selectivity of a variety of anti-inflammatory drugs based on a novel assay for PPAR delta(beta).

The nuclear receptor PPAR (peroxisome proliferator-activated receptor) has three subtypes named alpha, delta(beta), and gamma that may act as receptors for a range of compounds including antihyperglycaemic drugs, insulin sensitizers, and non-steroidal anti-inflammatory drugs (NSAIDs). Although profiling of the subtype selectivity of the compounds for PPAR is indispensable to elucidate their pharmacological action, the absence of an appropriate transactivation assay for PPAR delta led us to develop a sensitive and reproducible method. We found that co-expression of PPAR delta, retinoid X receptor (RXR) alpha, and coactivators such as CBP and SRC-1 enhanced basal and agonist-dependent activation of PPAR responsive element (PPRE)-driven transcription by PPAR delta, rendering a PPRE-driven reporter assay reliable and sensitive. Utilizing this assay for PPAR delta, we re-evaluated the subtype selectivity of a variety of anti-inflammatory drugs for human PPAR. The PPAR agonists tested included two leukotriene (LT) D(4) antagonist, seven NSAIDs, and two anti-rheumatoid drugs. We found that a novel LTD(4) antagonist, FK011 ([2-(((2-(4-tert-butyl-1,3-thiazol-2-yl)-1-benzofuran-5-yl)oxy)methyl)phenyl]acetic acid), showed marked agonistic activity for PPAR gamma. NSAIDs were classified into the following three groups: those showing no activity for all subtypes, those that were selective for PPAR gamma such as indomethacin and diclofenac, and those showing agonistic activity for the delta and gamma subtypes such as ibuprofen. These results will be important to studies on the molecular mechanisms of pharmacological actions of LTD(4) antagonists and NSAIDs.