Jiawen Cui,Zhenda Liang,Yuhao Liu,Zhiyu Hao,You Tang,Li Zhou,Xiaohua Teng
Environmental endocrine disrupting chemical 4-tert-butylphenol (4-tBP), a widely-utilized surfactant in various industries, poses potential risks to aquatic organisms. Our previous sequencing results suggested that 4-tBP-induced common carp liver injury might be associated with Ca 2+ signaling and autophagy. However, the intricate involvement of these pathways in 4-tBP-induced cytotoxic mechanisms remained unexplored. To bridge these knowledge gaps, this study focused on epithelioma papulosum cyprini (EPC) cells, a significant cell type in fish biology. Initial observations showed that 4-tBP induced a dose-dependent perturbation in Ca 2+ levels. Further investigations, with siRNA and L-type Ca 2+ channel agonist (BAY K8644), identified L-type calcium channel gene CACNA1D as a critical regulator of 4-tBP-induced Ca 2+ overload. Predictive analysis using miRanda platform suggested a potential interaction between miR-363 and CACNA1D, which was subsequently verified through dual-luciferase reporter gene assays. We then established miR-363 mimic/inhibitor models, along with miR-363 and CACNA1D co-suppression models in EPC cells. Through TEM observation, immunofluorescence assay, Ca 2+ staining, and qRT-PCR analysis, we evaluated the role of miR-363/CACNA1D axis in modulating the effects of 4-tBP on Ca 2+ signaling and autophagy. Results showed that miR-363 inhibitor exacerbated 4-tBP-induced increase in CALM2, CAMKII, Calpain2, and p62 expression and also led to decrease in ATG5, ATG7, and LC3b expression. In contrast, miR-363 mimic notably alleviated these changes. Notably, siRNA CACNA1D effectively modulating miR-363 inhibitor's effect. Our study revealed that 4-tBP induced Ca 2+ overload and subsequent autophagy impairment via miR-363/CACNA1D axis. These findings illuminated a profound understanding of molecular mechanisms underlying 4-tBP-induced cytotoxicity and spotlighted a potential therapeutic target.
Guo-Xuan Liu,Fan Diao,Guang Lu,Qiang Zheng,Bin Fu,Hua-Chen Jiao,Ke-Zhou Wang,Dong-Hai Liu
Alectinib is the first-line therapy for anaplastic lymphoma kinase rearranged non-small cell lung cancer globally. Sinus bradycardia, as the major adverse cardiac events of alectinib, still widely impact patient’s quality of life. However, its underlying mechanism remains elusive. The aim of this study was to reveal the pathogenesis of the alectinib induced sinus bradycardia (AISB) in a rat model, including the electrophysiology alterations and the molecular mechanism. SD rats were administered alectinib (10 mg / kg / day) by gavage for 7 to 10 days to mimic the clinical AISB. 3-days alectinib treatment did not change heart rate and sinus node recovery time (SNRT) as assessed through in vivo electrophysiology study. Also, alectinib didn’t influence automaticity in isolated heart or single sinus node cardiomyocytes, indicating alectinib cannot decrease sinus node function rapidly. The decreased heart rate and prolonged SNRT was found after 7-days alectinib treatment. The inducibility of atrial fibrillation was not affected under the same condition. The RNA-seq assay revealed the transcriptomic alterations in sinus node of alectinib treated rats, and the dysregulation of genes in cardiac function were observed. The decreased expression of L-type calcium channel Cacna1d was confirmed among the channel candidates identified from RNA-seq assay. Subsequently, the patch-clamp test revealed the reduction in the corresponding L-type calcium current density in 7-days alectinib treated rats. These findings revealed that the AISB was caused by the reduction in Cacna1d expression which resulted in the electro-dysfunction mediated by the suppressed I CaL .
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