Search

The activation of mitochondrial calcium-sensitive potassium (mBKCa) channels is crucially involved in cardioprotection induced by preconditioning. For milrinone (Mil)-induced preconditioning, the involvement of mBKCa-channels and further mitochondrial signaling is unknown. We hypothesize that (1) Mil-induced preconditioning is concentration-dependent and (2) that the activation of mBKCa-channels, release of reactive oxygen species (ROS), and the mitochondrial permeability transition pore (mPTP) could be involved. Isolated hearts of male Wistar rats were perfused with Krebs-Henseleit buffer and underwent 33 min of ischemia followed by 60 min of reperfusion. For determination of a concentration-dependent effect of Mil, hearts were perfused with different concentrations of Mil (0.3&ndash, 10 &micro, M) over 10 min before ischemia. In a second set of experiments, in addition to controls, hearts were pretreated with the lowest protective concentration of 1 &micro, M Mil either alone or combined with the mBKCa-channel blocker paxilline (Pax + Mil), or paxilline alone (Pax). In additional groups, Mil was administered with and without the ROS scavenger N-2-mercaptopropionylglycine (MPG + Mil, MPG) or the mPTP inhibitor cyclosporine A (MPG + Mil + CsA, CsA + Mil), respectively. Infarct sizes were determined by triphenyltetrazolium chloride (TTC) staining. The lowest and most cardioprotective concentration was 1 &micro, M Mil (Mil 1: 32 &plusmn, 6%, p &lt, 0.05 vs. Con: 63 &plusmn, 8% and Mil 0.3: 49 &plusmn, 6%). Pax and MPG blocked the infarct size reduction of Mil (Pax + Mil: 53 &plusmn, 6%, MPG + Mil: 59 &plusmn, 7%, 0.05 vs. Mil: 34 &plusmn, 6%) without having an effect on infarct size when administered alone (Pax: 53 &plusmn, 7%, MPG: 58 &plusmn, 5%, ns vs. Con). The combined administration of CsA completely restored the MPG-inhibited cardioprotection of Mil (MPG + Mil + CsA: 35 &plusmn, 7%, p &lt, 0.05 vs. MPG + Mil). Milrinone concentration-dependently induces preconditioning. Cardioprotection is mediated by the activation of mBKCa-channels, release of ROS and mPTP inhibition.

Accumulating evidence has shown the beneficial health effects of extra virgin olive oil (EVOO) consumption in reducing blood pressure and preventing the risk of developing hypertension. Some studies associate the hypotensive activity of EVOO to a minor component&mdash, the phenols. This study was designed to investigate the effects of EVOO phenols on the rat resistance mesenteric artery (MA) and to find out the possible vascular pathways involved. The experiments were carried out using a pressurized myograph, which allowed the effects of phenols on isolated MA to be tested under different conditions: (a) with endothelium removed, (b) with inhibition of nitric oxide synthase by N&omega, Nitro-l-arginine methyl ester hydrochloride (l-NAME, 10-4 M) + N&omega, Nitro-l-arginine (l-NNA, 10-4 M), (c) with inhibition of cyclooxygenase by indomethacin (10&minus, 5 M), (d) with inhibition of guanylate cyclase by 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one (ODQ,10&minus, 5 M) or adenylate cyclase by 9-(Tetrahydro-2&prime, furyl)adenine (SQ, 10&minus, (e) with depolarization by high potassium chloride (40 mM), and (f) with inhibition of the large conductance Ca2+&ndash, potassium channels (BKCa2+) with paxilline (10&minus, 5 M). EVOO phenols induce vasodilation of the endothelium, mediated by a direct effect on smooth muscle cells (SMC) by activation of BKCa2+ channels, an action by which phenols can regulate the vascular tone of the resistance artery. Phenols can be regarded as bioactive molecules that may contribute to the antihypertensive effects of EVOO.

The effects of Ca2+-activated K+ (BK) channel modulation by Paxilline (PAX) (10&minus, 7&ndash, 10&minus, 4 M), Iberiotoxin (IbTX) (0.1&ndash, 1 &times, 6 M) and Resveratrol (RESV) (1&ndash, 2 &times, 4 M) on cell cycle and proliferation, AKT1pSer473 phosphorylation, cell diameter, and BK currents were investigated in SH-SY5Y cells using Operetta-high-content-Imaging-System, ELISA-assay, impedentiometric counting method and patch-clamp technique, respectively. IbTX (4 &times, 7 M), PAX (5 &times, 5 M) and RESV (10&minus, 4 M) caused a maximal decrease of the outward K+ current at +30 mV (Vm) of &minus, 38.3 &plusmn, 10%, &minus, 31.9 &plusmn, 9% and &minus, 43 &plusmn, 8%, respectively, which was not reversible following washout and cell depolarization. After 6h of incubation, the drugs concentration dependently reduced proliferation. A maximal reduction of cell proliferation, respectively of &minus, 60 &plusmn, 8% for RESV (2 &times, 4 M) (IC50 = 1.50 &times, 4 M), &minus, 65 &plusmn, 6% for IbTX (10&minus, 6 M) (IC50 = 5 &times, 7 M), &minus, 97 &plusmn, 6% for PAX (1 &times, 4 M) (IC50 = 1.06 &times, 5 M) and AKT1pser473 dephosphorylation was observed. PAX induced a G1/G2 accumulation and contraction of the S-phase, reducing the nuclear area and cell diameter. IbTX induced G1 contraction and G2 accumulation reducing diameter. RESV induced G2 accumulation and S contraction reducing diameter. These drugs share common actions leading to a block of the surface membrane BK channels with cell depolarization and calcium influx, AKT1pser473 dephosphorylation by calcium-dependent phosphatase, accumulation in the G2 phase, and a reduction of diameter and proliferation. In addition, the PAX action against nuclear membrane BK channels potentiates its antiproliferative effects with early apoptosis.

The indole-diterpene paxilline is an abundant secondary metabolite synthesized by Penicillium paxilli. In total, 21 genes have been identified at the PAX locus of which six have been previously confirmed to have a functional role in paxilline biosynthesis. A combination of bioinformatics, gene expression and targeted gene replacement analyses were used to define the boundaries of the PAX gene cluster. Targeted gene replacement identified seven genes, paxG, paxA, paxM, paxB, paxC, paxP and paxQ that were all required for paxilline production, with one additional gene, paxD, required for regular prenylation of the indole ring post paxilline synthesis. The two putative transcription factors, PP104 and PP105, were not co-regulated with the pax genes and based on targeted gene replacement, including the double knockout, did not have a role in paxilline production. The relationship of indole dimethylallyl transferases involved in prenylation of indole-diterpenes such as paxilline or lolitrem B, can be found as two disparate clades, not supported by prenylation type (e.g., regular or reverse). This paper provides insight into the P. paxilli indole-diterpene locus and reviews the recent advances identified in paxilline biosynthesis.

Vitamin D (VD) reduces the risk of breast cancer and improves disease prognoses. Potential VD analogs are being developed as therapeutic agents for breast cancer treatments. The large-conductance Ca2+-activated K+ channel KCa1.1 regulates intracellular Ca2+ signaling pathways and is associated with high grade tumors and poor prognoses. In the present study, we examined the effects of treatments with VD receptor (VDR) agonists on the expression and activity of KCa1.1 in human breast cancer MDA-MB-453 cells using real-time PCR, Western blotting, flow cytometry, and voltage-sensitive dye imaging. Treatments with VDR agonists for 72 h markedly decreased the expression levels of KCa1.1 transcripts and proteins in MDA-MB-453 cells, resulting in the significant inhibition of depolarization responses induced by paxilline, a specific KCa1.1 blocker. The specific proteasome inhibitor MG132 suppressed VDR agonist-induced decreases in KCa1.1 protein expression. These results suggest that KCa1.1 is a new downstream target of VDR signaling and the down-regulation of KCa1.1 through the transcriptional repression of KCa1.1 and enhancement of KCa1.1 protein degradation contribute, at least partly, to the antiproliferative effects of VDR agonists in breast cancer cells.