Aconitine is the main bioactive ingredient of Aconitum plants, which are well-known botanical herbs in China. Aconitine is also notorious for its high cardiotoxicity, as it can induce life-threatening ventricular arrhythmias. Unfortunately, there are few effective antidotes to aconitine toxicity. This study aimed to evaluate the potent protective effects of the ingredients from V. baillonii on aconitine toxicity on H9c2 cell line. Cell viability was assessed by methylthiazoltetrazolium bromide (MTT). Intracellular Ca2+ concentration alteration and reactive oxygen species (ROS) generation were observed by confocal microscopy and flow cytometry, respectively. Cellular oxidative stress was analyzed by measuring malondialdehyde (MDA) and superoxide dismutase (SOD) levels. Mitochondrial membrane potential (ΔΨ) was determined using JC-1 kit. RT-PCR and Hoechst staining techniques were conducted to determine the levels of autophagy/apoptosis. The mRNA levels of dihydropyridine receptor (DHPR), ryanodine receptors (RyR2) and sarcoplasmic reticulum Ca2+-ATPase (SERCA) were measured by RT-PCR. We screened six components from V. baillonii, among which, sweroside exhibited the strongest protective effects on aconitine-induced cardiac toxicity. Sweroside suppressed the aconitine-induced mRNA expressions of NaV1.5 (encoded by SCN5A), RyR2 and DHPR, and reversed the aconitine-induced decrease in mRNA level of SERCA, thus preventing the aconitine-induced persistent intracellular Ca2+ accumulation and avoiding intracellular Ca2+ overload. We further found that sweroside restabilized the aconitine-disrupted mitochondrial membrane potential (ΔΨ) and reversed the aconitine-induced increase in the mRNA levels of cell autophagy-related factors (Beclin-1, Caspase-3, and LC3- II) in H9c2 cells. In the whole-animal experiments, we observed that sweroside (50 mg/kg) alleviated effectively aconitine-induced arrhythmias by analysis of electrocardiogram (ECG) recording in rats. Our results demonstrate that sweroside may protect cardiomyocytes from aconitine toxicity by maintaining intracellular Ca2+ homeostasis, restabilizing mitochondrial membrane potential (ΔΨ) and avoiding cell autophagy/apoptosis.

The participation of large-conductance Ca2+ activated K+ channels (BKs) in chloroquine (chloro)-induced relaxation of precontracted airway smooth muscle (ASM) is currently undefined. In this study we found that iberiotoxin (IbTx, a selective inhibitor of BKs) and chloro both completely blocked spontaneous transient outward currents (STOCs) in single mouse tracheal smooth muscle cells, which suggests that chloro might block BKs. We further found that chloro inhibited Ca2+ sparks and caffeine-induced global Ca2+ increases. Moreover, chloro can directly block single BK currents completely from the intracellular side and partially from the extracellular side. All these data indicate that the chloro-induced inhibition of STOCs is due to the blockade of chloro on both BKs and ryanodine receptors (RyRs). We also found that low concentrations of chloro resulted in additional contractions in tracheal rings that were precontracted by acetylcholine (ACH). Increases in chloro concentration reversed the contractile actions to relaxations. In the presence of IbTx or paxilline (pax), BK blockers, chloro-induced contractions were inhibited, although the high concentrations of chloro-induced relaxations were not affected. Taken together, our results indicate that chloro blocks BKs and RyRs, resulting in abolishment of STOCs and occurrence of contraction, the latter will counteract the relaxations induced by high concentrations of chloro.

Background: Placenta-specific 9 (Plac9) is a putative secreted protein that was first discovered in the context of embryogenesis. The expression pattern of Plac9 during embryogenesis, together with the results of recent reports, suggest that Plac9 may play a role in the liver development. The present study was conducted to investigate the secretory characteristics of Plac9 and its potential role in liver cell physiology. Methods: Immunofluorescence was employed to identify the subcellular distribution of Plac9. Cellular proliferative activity was analyzed by MTT assay and cell colony formation. The cell cycle distribution of Plac9 was analyzed by flow cytometry, and a functional analysis was performed using L02 cells following their stable infection with a lentivirus over-expressing Plac9. Results: Plac9 is a novel protein that is localized to the cytoplasm and may be secreted through the classic endoplasmic reticulum-Golgi route. The overexpression of Plac9 inhibits cell growth and induces G2/M phase arrest. Conclusion: Our findings reveal a novel role for Plac9 in regulating cell growth.

Cortex phellodendri is used to reduce fever and remove dampness and toxin. Berberine is an active ingredient of C. phellodendri. Berberine from Argemone ochroleuca can relax airway smooth muscle (ASM); however, whether the nonberberine component of C. phellodendri has similar relaxant action was unclear. An n-butyl alcohol extract of C. phellodendri (NBAECP, nonberberine component) was prepared, which completely inhibits high K+- and acetylcholine- (ACH-) induced precontraction of airway smooth muscle in tracheal rings and lung slices from control and asthmatic mice, respectively. The contraction induced by high K+ was also blocked by nifedipine, a selective blocker of L-type Ca2+ channels. The ACH-induced contraction was partially inhibited by nifedipine and pyrazole 3, an inhibitor of TRPC3 and STIM/Orai channels. Taken together, our data demonstrate that NBAECP can relax ASM by inhibiting L-type Ca2+ channels and TRPC3 and/or STIM/Orai channels, suggesting that NBAECP could be developed to a new drug for relieving bronchospasm.

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