Although artesunate has been reported to be a promising candidate for colorectal cancer (CRC) treatment, the underlying mechanisms and molecular targets of artesunate are yet to be explored. Here, we report that artesunate acts as a senescence and autophagy inducer to exert its inhibitory effect on CRC in a reactive oxygen species (ROS)-dependent manner. In SW480 and HCT116 cells, artesunate treatment led to mitochondrial dysfunction, drastically promoted mitochondrial ROS generation, and consequently inhibited cell proliferation by causing cell cycle arrest at G0/G1 phase as well as subsequent p16- and p21-mediated cell senescence. Senescent cells underwent endoplasmic reticulum stress (ERS), and the unfolded protein response (UPR) was activated via IRE1α signaling, with upregulated BIP, IRE1α, phosphorylated IRE1α (p-IRE1α), CHOP, and DR5. Further experiments revealed that autophagy was induced by artesunate treatment due to oxidative stress and ER stress. In contrast, N-Acetylcysteine (NAC, an ROS scavenger) and 3-Methyladenine (3-MA, an autophagy inhibitor) restored cell viability and attenuated autophagy in artesunate-treated cells. Furthermore, cellular free Ca²⁺ levels were increased and could be repressed by NAC, 3-MA, and GSK2350168 (an IRE1α inhibitor). In vivo, artesunate administration reduced the growth of CT26 cell-derived tumors in BALB/c mice. Ki67 and cyclin D1 expression was downregulated in tumor tissue, while p16, p21, p-IRE1α, and LC3B expression was upregulated. Taken together, artesunate induces senescence and autophagy to inhibit cell proliferation in colorectal cancer by promoting excessive ROS generation.

More than 300 million people suffer from depressive disorders globally. People under early-life stress (ELS) are reportedly vulnerable to depression in their adulthood, and synaptic plasticity can be the molecular mechanism underlying such depression. Herein, we simulated ELS by using a maternal separation (MS) model and evaluated the behavior of Sprague–Dawley (SD) rats in adulthood through behavioral examination, including sucrose preference, forced swimming, and open-field tests. The behavior tests showed that SD rats in the MS group were more susceptible to depression- and anxiety-like behaviors than did the non-MS (NMS) group. Nissl staining analysis indicated a significant reduction in the number of neurons at the prefrontal cortex and hippocampus, including the CA1, CA2, CA3, and DG regions of SD rats in the MS group. Immunohistochemistry results showed that the percentages of synaptophysin-positive area in the prefrontal cortex and hippocampus (including the CA1, CA2, CA3, and DG regions) slice of the MS group significantly decreased compared with those of the NMS group. Western blot analysis was used to assess synaptic-plasticity protein markers, including postsynaptic density 95, synaptophysin, and growth-associated binding protein 43 protein expression in the cortex and hippocampus. Results showed that the expression levels of these three proteins in the MS group were significantly lower than those in the NMS group. LC–MS/MS analysis revealed no significant differences in the peak areas of sex hormones and their metabolites, including estradiol, testosterone, androstenedione, estrone, estriol, and 5β-dihydrotestosterone. Through the application of nontargeted metabolomics to the overall analysis of differential metabolites, pathway-enrichment results showed the importance of arginine and proline metabolism; pantothenate and CoA biosyntheses; glutathione metabolism; and the phenylalanine, tyrosine, and tryptophan biosynthesis pathways. In summary, the MS model caused adult SD rats to be susceptible to depression, which may regulate synaptic plasticity through arginine and proline metabolism; pantothenate and CoA biosyntheses; glutathione metabolism; and phenylalanine, tyrosine, and tryptophan biosyntheses.

Sinisan (SNS) consists of four kinds of herbs, which is the core of antidepressant prescription widely used in traditional Chinese medicine clinic treatment for depression induced by early life stress. However, the role and precise mechanism of SNS antidepressant have not yet been elucidated. This study aimed to investigate the mechanism SNS on antidepressant of regulating mitochondrial function to improve hippocampal synaptic plasticity. 90 Sprague-Dawley (SD) rats male pups on Post-Natal Day (PND) 0 were randomly divided into Control group (ddH₂0), Model group (ddH₂0), Fluoxetine group (5.0 mg/kg fluoxetine), and SNS-L group (2.5 g/kg SNS), SNS-M group (5.0 g/kg SNS) and SNS-H group (10.0 g/kg SNS), 15 animals per group. Maternal separation (MS) from PND1 to PND21, drug intervention from PND60 to PND90, and behavior tests including sucrose preference test, open field test and forced swimming test from PND83 to PND90 were performed. Synaptic structure and mitochondrial structure were observed by TEM. The expression levels of PSD-95 and SYN were detected by immunohistochemistry and western blot test, the adenosine triphosphate (ATP) content in the hippocampus was detected by assay kits, and the expression levels of Mfn2, Drp1 and Fis1 protein were detected by western bolt test. SNS can alleviate depression-like and anxiety-like behaviors in MS rats, improve the damage of synapses and mitochondria, reduce the decrease of ATP in hippocampus, and reverse the expression levels of PSD-95, SYN, Mfn2, Drp1, and Fis1 proteins. SNS reduced the risk of early life stress induced depression disorder via regulating mitochondrial function and synaptic plasticity. Targeting mitochondrial may be a novel prospective therapeutic avenue for antidepressant.

Adverse stress in early life negatively influences psychiatric health by increasing the risk of developing depression and suicide in adulthood. Clinical antidepressants, such as fluoxetine, exhibit unsatisfactory results due to their low efficacy or intolerable side effects. SiNiSan (SNS), a traditional Chinese herbal formula, has been proven to have affirmatory antidepressive effects. However, the underlying mechanism remains poorly understood. Therefore, this study aimed to explore the impact and molecular mechanism of SNS treatment in rats exposed to neonatal maternal separation (MS)-combined young-adult chronic unpredictable mild stress (CUMS). Seventy-two neonatal male Sprague-Dawley rats were randomly divided into six groups of 12 rats each: control + ddH2O, model + ddH2O, positive (fluoxetine: 5 mg/kg), SNS-low dose (2.5 g/kg), SNS-medium dose (5 g/kg), and SNS-high dose (10 g/kg). Behavioral tests included sucrose preference test, open-field test, and forced swimming test. Calcium sensitive receptor (CaSR), protein kinase C (PKC), ERK1/2, and synapse-associated proteins (PSD-95, GAP-43, and synaptophysin [Syn]) in the hippocampus (HIP) and prefrontal cortex (PFC) were assayed using Western blot. CaSR and Syn protein expression was measured by immunohistochemistry. MS-combined CUMS rats exhibited depression-like behavior. SNS exerted antidepressant effects on stress-induced depression-like behavior. The levels of CaSR, PKC, and p-ERK1/2 in the HIP and PFC decreased in stressed rats. SNS treatment significantly upregulated the expression of CaSR, PKC, and p-ERK1/2 in the HIP and PFC of adult stressed rats. MS-combined CUMS could develop depression-like behavior in adult. SNS exhibited antidepressive effects accompanied by improving synaptic plasticity by activation of the CaSR-PKC-ERK signaling pathway.