PKCδ modulates SP1 mediated mitochondrial autophagy to exacerbate diacetylmorphine-induced ferroptosis in neurons.

Diacetylmorphine (DA) is widely implicated in neuronal injury; however, the underlying mechanisms remain unclear. We investigated the role of iron metamorphosis in DA-induced neurotoxicity using Sprague-Dawley rats and PC12 and SH-SY5Y cells. Tandem mass tag proteomics analysis showed that the upregulation of protein kinase C delta (PKCδ) and iron metabolism-related protein transferrin receptor (TFRC) significantly the enriched iron metabolism pathway. Subsequent experiments showed that DA exposure significantly upregulated PKCδ in PC12 cells, which increased the nuclear translocation of specificity protein 1 (SP1), and the intracellular free iron and lipid peroxide levels. In addition, silencing of PKCδ in rats improved behaviour and restored the expression level of glutathione peroxidase 4 (GPX4). In addition, DA exposure activated mitochondrial autophagy in PC12 cells, leading to a decrease in the mitochondrial membrane potential, accumulation of reactive oxygen species (ROS), elevation of LC3 (which plays a key role in autophagy), and a decrease in p62 expression. Following the inhibition of autophagy, the mitochondrial membrane potential and ROS were restored, as was the expression of voltage-dependent anion channel 1 (VDAC1) and GPX4. In conclusion, the present study suggests that PKCδ regulates SP1, further exacerbating DA-induced neuronal ferroptosis. Therefore, inhibition of PKCδ and mitochondrial autophagy or ferroptosis may be a key therapeutic target to ameliorate neurotoxicity following DA exposure.

SNTA1 inhibits the PI3K/AKT signaling pathway leading to increased mitochondrial dysfunction and arrhythmia caused by diacetylmorphine.

Diacetylmorphine (DA) abuse can result in severe arrhythmias and even sudden death. Although previous research has connected ion channel proteins to arrhythmia occurrences, the precise mechanism underlying DA-induced arrhythmias remains poorly understood. This study conducted a comprehensive analysis of the myocardial toxicity of DA by applying proteomic and histopathological approaches and investigated the underlying mechanisms using in vitro experiments. In vivo experiments confirmed that DA induces cardiac arrhythmias, as evidenced by electrocardiographic analyses of rats. Additionally, Masson staining, wheat germ agglutinin staining (WGA) staining, and western blotting of myocardial tissues revealed significant myocardial damage. Tandem mass tag proteomics analysis identified syntrophin alpha 1 (SNTA1) as a pivotal target molecule linked to myocardial toxicity. Ex vivo experiments showed specific upregulation of SNTA1 in rat cardiomyocytes following DA exposure. Furthermore, in vitro experiments indicated that DA caused disruption of potassium channels and activated the arrhythmia-related PI3K/AKT signaling pathway. Silencing and overexpression studies of SNTA1 highlighted its role in ion channel abnormalities and that of the PI3K/AKT signaling pathway expression in cardiomyocytes, underscoring the crucial role of mitochondrial function in cardiac arrhythmias. This research indicates that SNTA1 is integral to arrhythmia development by influencing the PI3K/AKT signaling pathway, leading to mitochondrial dysfunction and ion channel irregularities. SNTA1 is a potential therapeutic target for DA-induced arrhythmias. This study enhances our understanding of DA-induced myocardial toxicity and offers valuable insights for assessing the risks of DA exposure in humans.

Analysis of heroin effects on calcium channels in rat cardiomyocytes based on transcriptomics and metabolomics.

Heroin can cause damage to many human organs, possibly leading to different types of arrhythmias and abnormal electrophysiological function of the heart muscle and the steady state of calcium-ion channels. We explored cardiomyocytes treated with heroin and the effect on calcium-ion channels. Transcriptomics and metabolomics were used to screen for differential genes and metabolite alterations after heroin administration to jointly analyze the effect of heroin on calcium channels in cardiomyocytes. Cardiomyocytes from primary neonatal rats were cultured in vitro and were treated with different concentrations of heroin to observe the changes in morphology and spontaneous beat frequency and rhythm by a patch clamp technique. Transcriptomic studies selected a total of 1,432 differentially expressed genes, 941 upregulated and 491 downregulated genes in rat cardiomyocytes from the control and drug intervention groups. Gene Ontology functional enrichment showed that 1,432 differential genes selected by the two groups were mainly involved in the regulation of the multicellular organismal process, response to external stimulus, myofibril, inflammatory response, muscle system process, cardiac muscle contraction, etc. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis indicated that these genes were mainly concentrated in cardiac muscle contraction, osteoclast differentiation, adrenergic signaling in cardiomyocytes, dilated cardiomyopathy, hypertrophic cardiomyopathy, and other important pathways. Metabolomic testing further suggested that cardiomyocyte metabolism was severely affected after heroin intervention. After the treatment with heroin, the L-type calcium channel current I-V curve was up-shifted, the peak value was significantly lower than that of the control group, action potential duration 90 was significantly increased in the action potential, resting potential negative value was lowered, and action potential amplitude was significantly decreased in cardiomyocytes. In this study, heroin could cause morphological changes in primary cardiomyocytes of neonatal rats and electrophysiological function. Heroin can cause myocardial contraction and calcium channel abnormalities, damage the myocardium, and change the action potential and L-type calcium channel.

CaMKII regulates the proteins TPM1 and MYOM2 and promotes diacetylmorphine-induced abnormal cardiac rhythms.

Although opioids are necessary for the treatment of acute pain, cancer pain, and palliative care, opioid abuse is a serious threat to society. Heroin (Diacetylmorphine) is the most commonly abused opioid, and it can have a variety of effects on the body's tissues and organs, including the well-known gastrointestinal depression and respiratory depression; however, there is little known about the effects of diacetylmorphine on cardiac damage. Here, we demonstrate that diacetylmorphine induces abnormal electrocardiographic changes in rats and causes damage to cardiomyocytes in vitro by an underlying mechanism of increased autophosphorylation of CaMKII and concomitant regulation of myocardial contractile protein TPM1 and MYOM2 protein expression. The CaMKII inhibitor KN-93 was first tested to rescue the toxic effects of heroin on cardiomyocytes in vitro and the abnormal ECG changes caused by heroin in SD rats, followed by the TMT relative quantitative protein technique to analyze the proteome changes. Diacetylmorphine causes increased phosphorylation at the CaMKII Thr287 site in myocardium, resulting in increased autophosphorylation of CaMKII and subsequent alterations in myocardial contractile proteins, leading to myocardial rhythm abnormalities. These findings provide a theoretical basis for the treatment and prevention of patients with arrhythmias caused by diacetylmorphine inhalation and injection.