Omega-3 polyunsaturated fatty acid derived lipid mediators: a comprehensive update on their application in anti-cancer drug discovery.

INTRODUCTION: ω-3 Polyunsaturated fatty acids (PUFAs) have a range of health benefits, including anticancer activity, and are converted to lipid mediators that could be adapted into pharmacological strategies. However, the stability of these mediators must be improved, and they may require formulation to achieve optimal tissue concentrations. AREAS COVERED: Herein, the author reviews the literature around chemical stabilization and formulation of ω-3 PUFA mediators and their application in anticancer drug discovery. EXPERT OPINION: Aryl-urea bioisosteres of ω-3 PUFA epoxides that killed cancer cells targeted the mitochondrion by a novel dual mechanism: as protonophoric uncouplers and as inhibitors of electron transport complex III that activated ER-stress and disrupted mitochondrial integrity. In contrast, aryl-ureas that contain electron-donating substituents prevented cancer cell migration. Thus, aryl-ureas represent a novel class of agents with tunable anticancer properties. Stabilized analogues of other ω-3 PUFA-derived mediators could also be adapted into anticancer strategies. Indeed, a cocktail of agents that simultaneously promote cell killing, inhibit metastasis and angiogenesis, and that attenuate the pro-inflammatory microenvironment is a novel future anticancer strategy. Such regimen may enhance anticancer drug efficacy, minimize the development of anticancer drug resistance and enhance outcomes.

Sishen Wan enhances intestinal barrier function via regulating endoplasmic reticulum stress to improve mice with diarrheal irritable bowel syndrome.

BACKGROUND: Diarrheal irritable bowel syndrome (IBS-D), characterized primarily by the presence of diarrhea and abdominal pain, is a clinical manifestation resulting from a multitude of causative factors. Furthermore, Sishen Wan (SSW) has demonstrated efficacy in treating IBS-D. Nevertheless, its mechanism of action remains unclear. METHODS: A model of IBS-D was induced by a diet containing 45 % lactose and chronic unpredictable mild stress. Additionally, the impact of SSW was assessed by measuring body weight, visceral sensitivity, defecation parameters, intestinal transport velocity, intestinal neurotransmitter levels, immunohistochemistry, and transmission electron microscopy analysis. Immunofluorescent staining was used to detect the expression of Mucin 2 (MUC2) and Occludin in the colon. Western blotting was used to detect changes in proteins related to tight junction (TJ), autophagy, and endoplasmic reticulum (ER) stress in the colon. Finally, 16S rRNA amplicon sequencing was used to monitor the alteration of gut microbiota after SSW treatment. RESULTS: Our study revealed that SSW administration resulted in reduced visceral sensitivity, improved defecation parameters, decreased intestinal transport velocity, and reduced intestinal permeability in IBS-D mice. Furthermore, SSW promotes the secretion of colonic mucus by enhancing autophagy and inhibiting ER stress. SSW treatment caused remodeling of the gut microbiome by increasing the abundance of Blautia, Muribaculum and Ruminococcus torques group. CONCLUSION: SSW can improve intestinal barrier function by promoting autophagy and inhibiting ER stress, thus exerting a therapeutic effect on IBS-D.

Network pharmacology, bioinformatics, and experimental validation to identify the role of Hedyotis diffusa willd against gastric cancer through the activation of the endoplasmic reticulum stress.

Background: Globally, gastric cancer (GC) is recognized as the third leading cause of cancer-related deaths and the fifth most prevalent malignant disease. Multiple studies have indicated that Hedyotis diffusa Willd, in pinyin, called Bai Hua She Cao (BHSSC), a traditional Chinese medicine (TCM) is an herbal remedy for cancer treatment. However, the specific mechanisms underlying its anti-tumor properties and mode of action are still unclear. Methods: To determine the role of BHSSC in GC, candidate target genes were selected from The Encyclopedia of Traditional Chinese Medicine (ETCM) and analyzed using network pharmacology, bioinformatics, and experimental validation. Differentially expressed genes (DEGs) associated with gastric cancer were obtained from RNA sequencing (RNA-seq) data sourced from The Cancer Genome Atlas-Stomach adenocarcinoma (TCGA-STAD). The Reactome Pathway was examined using Analysis Tools, while KEGG pathways were analyzed using KOBAS. Gene Ontology (GO) evaluations were performed using WebGestalt and DAVID. The relationships between proteins were investigated using the STRING database. Furthermore, cell viability, colony formation, and cell migration ability were conducted in gastric cancer cells, BGC-823 and MGC-803. Results: Network pharmacology and bioinformatics analyses revealed a significant association between BHSSC and metabolic pathways. In vitro experiments demonstrated that BHSSC effectively suppressed gastric cancer cell proliferation and colony formation, inhibited cell migration, and activated the endoplasmic reticulum (ER) stress. Furthermore, it was found that enhancement of the expression of IRE1α and BIP is the mechanism by which BHSSC activates ER stress. Conclusions: The findings suggest that BHSSC exerts its effects through modulation of metabolic pathways, leading to the suppression of cell proliferation, inhibition of cell migration, and activation of the endoplasmic reticulum. These results provide valuable insights into the mechanisms underlying the therapeutic effects of BHSSC in GC and support its potential as a novel treatment option.

Study on the Antitumor Mechanism of Tanshinone IIA In Vivo and In Vitro through the Regulation of PERK-ATF4-HSPA5 Pathway-Mediated Ferroptosis.

As a traditional Chinese medicine, Salvia miltiorrhiza Bunge was first recorded in the Shennong Materia Medica Classic and is widely used to treat "the accumulation of symptoms and masses". The main active ingredient of Salvia miltiorrhiza Bunge, Tanshinone IIA (TIIA), has shown anti-inflammatory, antitumor, antifibrosis, antibacterial, and antioxidative activities, etc. In this study, the results showed that TIIA could inhibit the proliferation and migration of HepG2 cells and downregulate glutathione (GSH) and Glutathione Peroxidase 4 (GPX4) levels; besides, TIIA induced the production of Reactive Oxygen Species (ROS), and upregulated the total iron content. Based on network pharmacology analysis, the antitumor effect of TIIA was found to be focused on the endoplasmic reticulum (ER)-mediated ferroptosis signaling pathway, with protein kinase R (PKR)-like ER kinase (PERK)-activating transcription factor 4 (ATF4)-heat shock 70 kDa protein 5 (HSPA5) as the main pathway. Herein, TIIA showed typical ferroptosis characteristics, and a ferroptosis inhibitor (ferrostatin-1) was used to verify the effect. The antitumor effects of TIIA, occurring through the inhibition of the PERK-ATF4-HSPA5 pathway, were further observed in vivo as significantly inhibited tumor growth and the improved pathological morphology of tumor tissue in H22-bearing mice. In summary, the antitumor mechanism of TIIA might be related to the downregulation of the activation of PERK-ATF4-HSPA5 pathway-mediated ferroptosis.

[Huangpu Tongqiao Capsule improves cognitive impairment in rats with Wilson disease by inhibiting endoplasmic reticulum stress-mediated apoptosis pathway].

OBJECTIVE: To investigate the neuroprotective effect of Huangpu Tongqiao Capsule (HPTQ) in a rat model of Wilson disease (WD) and explore the underlying mechanisms. METHODS: SD rat models of WD were established by feeding of coppersupplemented chow diet and drinking water for 12 weeks, and starting from the 9th week, the rats were treated with low-, moderate- and high-dose HPTQ, penicillamine, or normal saline by gavage on a daily basis for 3 weeks. Copper levels in the liver and 24-h urine of the rats were detected, and their learning and memory abilities were evaluated using Morris water maze test. HE staining was used to observe morphological changes of CA1 region neurons in the hippocampus, and neuronal apoptosis was detected with TUNEL staining. Hippocampal expressions of endoplasmic reticulum stress (ERS)-mediated apoptosis pathway-related proteins GRP78, CHOP, caspase-12, cleaved caspase-9, and cleaved caspase-3 at both the mRNA and protein levels were detected using RT-qPCR, immunofluorescence assay or Western blotting. RESULTS: Compared with normal control rats, the rat models with copper overload-induced WD exhibited significantly increased copper levels in both the liver and 24-h urine, impaired learning and memory abilities, obvious hippocampal neuronal damage in the CA1 region and increased TUNEL-positive neurons (P<0.01), with also lowered mRNA and protein expressions of GRP78, CHOP, caspase-12, cleaved caspase-9, and cleaved caspase-3 in the hippocampus (all P<0.01). Treatments with HPTQ and penicillamine significantly lowered copper level in the liver but increased urinary copper level, improved learning and memory ability, alleviated neuronal damage and apoptosis in the hippocampus, and decreased hippocampal expressions of GRP78, CHOP, caspase-12, cleaved caspase-9, and cleaved caspase-3 in the rat models (P<0.01 or 0.05). CONCLUSION: HPTQ Capsule has neuroprotective effects in rat models of WD possibly by inhibiting ERS-mediated apoptosis pathway.

Dried tangerine peel polysaccharide (DTPP) alleviates hepatic steatosis by suppressing TLR4/MD-2-mediated inflammation and endoplasmic reticulum stress.

Non-alcoholic fatty liver disease (NAFLD) is a complex pathogenic metabolic syndrome characterized by increased inflammation and endoplasmic reticulum stress. In recent years, natural polysaccharides derived from traditional Chinese medicine have shown significant anti-inflammatory effects, making them an attractive therapeutic option. However, little research has been conducted on the therapeutic potential of dried tangerine peel polysaccharide (DTPP) - one of the most important medicinal resources in China. The results of the present study showed that DTPP substantially reduced macrophage infiltration in vivo and suppressed the expression of pro-inflammatory factors and endoplasmic reticulum stress-related genes. Additionally, surface plasmon resonance analysis revealed that DTPP had a specific affinity to myeloid differentiation factor 2, which consequently suppressed lipopolysaccharide-induced inflammation via interaction with the toll-like receptor 4 signaling pathway. This study provides a potential molecular mechanism underlying the anti-inflammatory effects of DTPP on NAFLD and suggests DTPP as a promising therapeutic strategy for NAFLD treatment.

Effect of Calcium Supplementation and TMEM16A Inhibition on Endoplasmic Reticulum Stress Induced by Dental Fluorosis in Mice.

BACKGROUND: Dental fluorosis is a discoloration of the teeth caused by the excessive consumption of fluoride. It represents a distinct manifestation of chronic fluorosis in dental tissues, exerting adverse effects on the human body, particularly on teeth. The transmembrane protein 16a (TMEM16A) is expressed at the junction of the endoplasmic reticulum and the plasma membrane. Alterations in its channel activity can disrupt endoplasmic reticulum calcium homeostasis and intracellular calcium ion concentration, thereby inducing endoplasmic reticulum stress (ERS). This study aims to investigate the influence of calcium supplements and TMEM16A on ERS in dental fluorosis. METHODS: C57BL/6 mice exhibiting dental fluorosis were subjected to an eight-week treatment with varying calcium concentrations: low (0.071%), medium (0.79%), and high (6.61%). Various assays, including Hematoxylin and Eosin (HE) staining, immunohistochemistry, real-time fluorescence quantitative polymerase chain reaction (qPCR), and Western blot, were employed to assess the impact of calcium supplements on fluoride content, ameloblast morphology, TMEM16A expression, and endoplasmic reticulum stress-related proteins (calreticulin (CRT), glucose-regulated protein 78 (GRP78), inositol requiring kinase 1α (IRE1α), PKR-like ER kinase (PERK), activating transcription factor 6 (ATF6)) in the incisors of mice affected by dental fluorosis. Furthermore, mice with dental fluorosis were treated with the TMEM16A inhibitor T16Ainh-A01 along with a medium-dose calcium to investigate the influence of TMEM16A on fluoride content, ameloblast morphology, and endoplasmic reticulum stress-related proteins in the context of mouse incisor fluorosis. RESULTS: In comparison to the model mice, the fluoride content in incisors significantly decreased following calcium supplements (p < 0.01). Moreover, the expression of TMEM16A, CRT, GRP78, IRE1α, PERK, and ATF6 were also exhibited a substantial reduction (p < 0.01), with the most pronounced effect observed in the medium-dose calcium group. Additionally, the fluoride content (p < 0.05) and the expression of CRT, GRP78, IRE1α, PERK, and ATF6 (p < 0.01) were further diminished following concurrent treatment with the TMEM16A inhibitor T16Ainh-A01 and a medium dose of calcium. CONCLUSIONS: The supplementation of calcium or the inhibition of TMEM16A expression appears to mitigate the detrimental effects of fluorosis by suppressing endoplasmic reticulum stress. These findings hold implications for identifying potential therapeutic targets in addressing dental fluorosis.

Effects of electroacupuncture of different intensities and durations on PERK/ATF4/CHOP signaling pathway in liver of non-alcoholic fatty liver disease rats. / ä¸åŒå¼ºåº¦åŠæ—¶ç¨‹ç”µé’ˆå¹²é¢„å¯¹éžé ’ç²¾æ€§è„‚è‚ªè‚ç— å¤§é¼ è‚è„PERK/ATF4/CHOP通路的影响.

OBJECTIVES: To analyze the effects of electroacupuncture (EA) at "Fenglong" (ST40) and "Zusanli" (ST36) of different intensities and durations on rats with non-alcoholic fatty liver disease (NAFLD) based on the protein kinase R-like endoplasmic reticulum kinase (PERK)-activating transcription factor 4 (ATF4)-C/EBP homologous protein (CHOP) signaling pathway, so as to explore its mechanism underlying improvement of NAFLD. METHODS: SD rats were randomly divided into normal diet group, high-fat model group, sham EA group, strong stimulation EA (SEA) group, and weak stimulation EA (WEA) group, with 15 rats in each group. Each group was further divided into 2, 3, and 4-week subgroups. NAFLD rat model was established by feeding a high-fat diet. After successful modeling, rats in the SEA and WEA groups received EA at bilateral ST40 and ST36 with dense and sparse waves (4 Hz/20 Hz) at current intensities of 4 mA (SEA group) and 2 mA (WEA group), lasting for 20 minutes, once a day, 5 days a week with 2 days of rest. The sham EA group only had the EA apparatus connected without electricity. Different duration subgroups were intervened for 2, 3, and 4 weeks. After the intervention, the contents of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in rats were detected by an automatic biochemical analyzer；liver morphological changes were observed by Oil Red O staining；real-time fluorescence quantitative PCR and Western blot were used to detect the expression of PERK, ATF4, and CHOP mRNAs and proteins in the rat liver tissue. RESULTS: In the high-fat model group, there was a significant accumulation of red lipid droplets in the liver cells, which was reduced significantly in the SEA group at the 4th week. Compared with the normal diet group with the same treatment duration, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs and proteins in the liver tissue were elevated (P<0.01) in the high-fat model group . Compared with the high-fat model group with the same treatment duration, the contents of serum ALT, AST, and the expression of PERK, ATF4, CHOP mRNAs and proteins in the liver tissue were decreased (P<0.01, P<0.05) in the SEA and WEA groups. Compared with the sham EA group with the same treatment duration, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs were decreased (P<0.01, P<0.05) in the SEA and WEA groups, the expression of PERK, ATF4, and CHOP proteins in the liver tissue was decreased (P<0.01) in the SEA group at the 2nd, 3rd, and 4th week, the expression of PERK and CHOP proteins at the 2nd, 3rd, 4th week and ATF4 protein at 2nd week in the liver tissue were decreased (P<0.01, P<0.05) in the WEA group. Compared with the SEA group with the same treatment duration, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs and proteins in the liver tissue were elevated (P<0.05, P<0.01) in the WEA group. Compared with the 2-week time point within the groups, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs and PERK proteins in the liver tissue were decreased (P<0.01, P<0.05) in the SEA and WEA groups at 3rd and 4th week, the expression of ATF4 proteins in the liver tissue was decreased (P<0.01) in the SEA group at 3rd and 4th week, and the expression of CHOP proteins in the liver tissue was decreased (P<0.01) in the SEA group at 4th week and in the WEA group at 3rd and 4th week. Compared with the 3-week time point within the groups, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs were significantly decreased (P<0.05, P<0.01) in the SEA and WEA groups at 4th week, the expression of PERK and CHOP proteins in the liver tissue was decreased (P<0.01) in the SEA and WEA groups at 4th week, and the expression of ATF4 protein in the liver tissue was decreased (P<0.05) in the SEA group at 4th week. CONCLUSIONS: EA at ST40 and ST36 can significantly improve liver function in NAFLD rats, and its mechanism of action may involve inhibiting PERK expression thereby targeting the downstream ATF4/CHOP signaling pathway to suppress endoplasmic reticulum stress, exerting a liver protective effect；the optimal effect was observed with EA intensity of 4 mA for 4 weeks.

Effects of Letrozole Treatment and Vitamin C Supplementation on Morphology, Endoplasmic Reticulum Stress, Programmed Cell Death, and Oxidative Stress in the Small Intestine of Adult Male Rats.

Estrogens are hormones that play an important role in the digestive tract, including in men. Letrozole is an inhibitor of cytochrome P450 aromatase, an enzyme converting androgens to estrogens. The use of letrozole may cause oxidative stress and endoplasmic reticulum stress in the cells. Factors modulating cellular stress may include vitamin C. The purpose of this study was to examine whether letrozole and/or vitamin C supplementation can affect the morphology of the small intestine, the parameters of endoplasmic reticulum stress, programmed cell death markers, and oxidative damage. Three-month-old male rats were divided into four groups and treated with the following: (I) CTRL-water; (II) CTRL+C-L-ascorbic acid; (III) LET-letrozole; and (IV) LET+C-letrozole + L-ascorbic acid. The morphometrical measurements included epithelial thickness, crypt and lumen area, crypt perimeter, nuclei number in the crypt, and the cell size of crypts. The expression levels of PERK, caspase-3, and catalase were determined. Significant differences in the morphometrical measurements and immunoexpression were observed. This may indicate that chronic treatment with letrozole can affect morphology and induce ER stress, oxidative stress, and programmed cell death in the epithelial cells of the small intestine of adult male rats. Vitamin C supplementation exerts an effect on some parameters of the molecular processes.

Discovery of novel peptide-dehydroepiandrosterone hybrids inducing endoplasmic reticulum stress with effective in vitro and in vivo anti-melanoma activities.

Steroid hybrids have emerged as a type of advantageous compound as they could offer improved pharmacological and pharmaceutical properties. Here, we report a series of novel peptide-dehydroepiandrosterone hybrids, which would effectively induce endoplasmic reticulum stress (ERS) and lead to apoptosis with outstanding in vitro and in vivo anti-melanoma effects. The lead compound IId among various steroids conjugated with peptides and pyridines showed effective in vivo activity in B16 xenograft mice: in medium- and high-dose treatment groups (60 and 80 mg/kg), compound IId would significantly inhibit the growth of tumours by 98%-99% compared to the control group, with the highest survival rate as well. Further mechanism studies showed that compound IId would damage the endoplasmic reticulum and upregulate the ERS markers C/EBP homologous protein (CHOP) and glucose-regulated protein 78 (GRP78), which could further regulate caspase and Bcl-2 family proteins and lead to cell apoptosis. The compound IId was also proven to be effective in inhibiting B16 cell migration and invasion.

Lycopene Alleviates Endoplasmic Reticulum Stress in Steatohepatitis through Inhibition of the ASK1-JNK Signaling Pathway.

Lycopene has been proven to alleviate nonalcoholic steatohepatitis (NASH), but the precise mechanisms are inadequately elucidated. In this study, we found a previously unknown regulatory effect of lycopene on the apoptosis signal-regulating kinase 1 (ASK1) signaling pathway in both in vivo and in vitro models. Lycopene supplementation (3 and 6 mg/kg/day) exhibited a significant reduction in lipid accumulation, inflammation, and fibrosis of the liver in mice fed with a high-fat/high-cholesterol diet or a methionine-choline-deficient diet. RNA sequencing uncovered that the mitogen-activated protein kinases signaling pathway, which is closely associated with inflammation and endoplasmic reticulum (ER) stress, was significantly downregulated by lycopene. Furthermore, we found lycopene ameliorated ER swelling and decreased the expression levels of ER stress markers (i.e., immunoglobulin heavy chain binding protein, C/EBP homologous protein, and X-box binding protein 1s). Especially, the inositol-requiring enzyme 1α involved in the ASK1 phosphorylation was inhibited by lycopene, resulting in the decline of the subsequent c-Jun N-terminal kinase (JNK) signaling cascade. ASK1 inhibitor DQOP-1 eliminated the lycopene-induced inhibition of the ASK1-JNK pathway in oleic acid and palmitic acid-induced HepG2 cells. Molecular docking further indicated hydrophobic interactions between lycopene and ASK1. Collectively, our research indicates that lycopene can alleviate ER stress and attenuate inflammation cascades and lipid accumulation by inhibiting the ASK1-JNK pathway.

Effect of moxibustion combined with intraperitoneal injection of benazepril on endoplasmic reticulum stress-related protein phosphorylation in rats with chronic heart failure. / è‰¾ç¸è”åˆè´é‚£æ™®åˆ©å¯¹æ ¢æ€§å¿ƒåŠ›è¡°ç«­å¤§é¼ å¿ƒè‚Œå† è´¨ç½‘åº”æ¿€ç›¸å ³è›‹ç™½ç£·é ¸åŒ–è¡¨è¾¾çš„å½±å“.

OBJECTIVES: To observe the effect of moxibustion at "Xinshu"(BL15) and "Feishu"(BL13) combined with intraperitoneal injection of benazepril on cardiac function and phosphorylation of protein kinase R-like endoplasmic reticulum kinase (PERK) and eukaryotic initiation factor 2α (elF2α) proteins in myocardium of rats with chronic heart failure (CHF), so as to explore its potential mechanism underlying improvement of CHF. METHODS: A total of 42 male SD rats were randomly assigned to blank control (n=10), CHF model (n=7), medication (benazepril, n=8), moxibustion (n=8) and moxibustion+benazepril (n=9) groups, after cardiac ultrasound model identification and elimination of the dead. The CHF model was established by intraperitoneal injection of doxorubicin hydrochloride (DOX), once every week for 6 weeks. Mild moxibustion was applied to bilateral BL15 and BL13 regions for 20 min, once daily for 3 weeks. The rats of the medication group and moxibustion+benazepril group (benazepril was given first, followed by moxibustion) received intraperitoneal injection of benazepril (0.86 mg/kg) solution once daily for 3 weeks . The cardiac ejection fraction (EF) and left ventricular fractional shortening (FS) were measured using echocardiography. Histopathological changes of the cardiac muscle tissue were observed under light microscope after hematoxylin-eosin (H.E.) staining. Serum contents of B-type brain natriuretic peptide (BNP) and angiotensin Ⅱ (AngⅡ) were measured by enzyme-linked immunosorbent assay (ELISA). The expressions of phospho-PERK (p-PERK) and phospho-elF2α (p-elF2α) in the myocardium were detected by Western blot. RESULTS: Compared with the blank control group, the EF and FS of the left cardiac ventricle were significantly decreased (P<0.01), while the contents of serum BNP and AngⅡ, and expression levels of p-PERK and p-eIF2α significantly increased in the model group (P<0.01). In comparison with the model group, both the decreased EF and FS and the increased BNP and AngⅡ contents as well as p-PERK and p-elF2α expression levels were reversed by moxibustion, medication and moxibustion+benazepril (P<0.01). The effects of moxibustion+benazepril were markedly superior to those of simple moxibustion and simple medication in raising the levels of EF and FS rate and in down-regulating the contents of BNP, Ang Ⅱ, levels of p-PERK and p-elF2α (P<0.01, P<0.05). Outcomes of H.E. staining showed irregular arrangement of cardiomyocytes, cell swelling, vacuole and inflammatory infiltration in the model group, which was relatively milder in the 3 treatment groups. The effects of moxibustion+benazepril were superior to those of moxibustion or benazepril. CONCLUSIONS: Moxibustion combined with Benazepril can improve the cardiac function in CHF rats, which may be related to its functions in down-regulating the expression levels of myocardial p-PERK and p-elF2α to inhibit endoplasmic reticulum stress response.

Forsythoside A protects against Zearalenone-induced cell damage in chicken embryonic fibroblasts via mitigation of endoplasmic reticulum stress.

Zearalenone (ZEA) is a non-steroidal estrogenic mycotoxin that exerts its toxic effects through various damage mechanisms such as oxidative stress, endoplasmic reticulum stress (ERS), mitochondrial damage, cell cycle arrest, and apoptosis. At present, there are few studies on drugs that can rescue ZEA-induced chicken embryonic fibroblasts damage. Forsythoside A (FA) is one of effective ingredients of traditional Chinese medicine that plays a role in various biological functions, but its antitoxin research has not been investigated so far. In this study, in vitro experiments were carried out. Chicken embryo fibroblast (DF-1) cells was used as the research object to select the appropriate treatment concentration of ZEA and examined reactive oxygen species (ROS), mitochondrial membrane potential, ERS and apoptosis to investigate the effects and mechanisms of FA in alleviating ZEA-induced cytotoxicity in DF-1 cells. Our results showed that ZEA induced ERS and activated the unfolded protein response (UPR) leading to apoptosis, an apoptotic pathway characterized by overproduction of Lactate dehydrogenase (LDH), Caspase-3, and ROS and loss of mitochondrial membrane potential. We also demonstrated that FA help to prevent ERS and attenuated ZEA-induced apoptosis in DF-1 cells by reducing the level of ROS, downregulating GRP78, PERK, ATF4, ATF6, JNK, IRE1, ASK1, CHOP, BAX expression, and up-regulating Bcl-2 expression. Our results provide a basis for an in-depth study of the mechanism of toxic effects of ZEA on chicken cells and the means of detoxification, which has implications for the treatment of relevant avian diseases.

Rosmarinic acid mitigates acrylamide induced neurotoxicity via suppressing endoplasmic reticulum stress and inflammation in mouse hippocampus.

BACKGROUND: Acrylamide (ACR) is a widely used compound that is known to be neurotoxic to both experimental animals and humans, causing nerve damage. The widespread presence of ACR in the environment and food means that the toxic risk to human health can no longer be ignored. Rosmarinic acid (RA), a natural polyphenolic compound extracted from the perilla plant, exhibits anti-inflammatory, antioxidant, and other properties. It has also been demon strated to possess promising potential in neuroprotection. However, its role and potential mechanism in treating ACR induced neurotoxicity are still elusive. PURPOSE: This study explores whether RA can improve ACR induced neurotoxicity and its possible mechanism. METHODS: The behavioral method was used to study RA effect on ACR exposed mice's neurological function. We studied its potential mechanism through metabolomics, Nissl staining, HE staining, immunohistochemical analysis, and Western blot. RESULTS: RA pretreatment reversed the increase in mouse landing foot splay and decrease in spontaneous activity caused by 3 weeks of exposure to 50 mg/kg/d ACR. Further experiments demonstrated that RA could prevent ACR induced neuronal apoptosis, significantly downregulate nuclear factor-κB and tumor necrosis factor-α expression, and inhibit NOD-like receptor protein 3 inflammasome activation, thereby reducing inflammation as confirmed by metabolomics results. Additionally, RA treatment prevented endoplasmic reticulum stress (ERS) caused by ACR exposure, as evidenced by the reversal of significant P-IRE1α,TRAF2,CHOP expression increase. CONCLUSION: RA alleviates ACR induced neurotoxicity by inhibiting ERS and inflammation. These results provide a deeper understanding of the mechanism of ACR induced neurotoxicity and propose a potential new treatment method.

D-pinitol alleviates diabetic cardiomyopathy by inhibiting the optineurin-mediated endoplasmic reticulum stress and glycophagy signaling pathway.

Diabetic cardiomyopathy (DCM) is an important complication resulting in heart failure and death of diabetic patients. However, there is no effective drug for treatments. This study investigated the effect of D-pinitol (DP) on cardiac injury using diabetic mice and glycosylation injury of cardiomyocytes and its molecular mechanisms. We established the streptozotocin-induced SAMR1 and SAMP8 mice and DP (150 mg/kg/day) intragastrically and advanced glycation end-products (AGEs)-induced H9C2 cells. H9C2 cells were transfected with optineurin (OPTN) siRNA and overexpression plasmids. The metabolic disorder indices, cardiac dysfunction, histopathology, immunofluorescence, western blot, and immunoprecipitation were investigated. Our results showed that DP reduced the blood glucose and AGEs, and increased the expression of heart OPTN in diabetic mice and H9C2 cells, thereby inhibiting the endoplasmic reticulum stress (GRP78, CHOP) and glycophagy (STBD1, GABARAPL1), and alleviating the myocardial apoptosis and fibrosis of DCM. The expression of filamin A as an interaction protein of OPTN downregulated by AGEs decreased OPTN abundance. Moreover, OPTN siRNA increased the expression of GRP78, CHOP, STBD1, and GABARAPL1 and inhibited the expression of GAA via GSK3ß phosphorylation and FoxO1. DP may be helpful to treat the onset of DCM. Targeting OPTN with DP could be translated into clinical application in the fighting against DCM.

Juan-tong-yin potentially impacts endometriosis pathophysiology by enhancing autophagy of endometrial stromal cells via unfolded protein reaction-triggered endoplasmic reticulum stress.

ETHNOPHARMACOLOGICAL RELEVANCE: Endometriosis (EMs) is characterized by inflammatory lesions, dysmenorrhea, infertility, and chronic pelvic pain. Single-target medications often fail to provide systemic therapeutic results owing to the complex mechanism underlying endometriosis. Although traditional Chinese medicines-such as Juan-Tong-Yin (JTY)-have shown promising results, their mechanisms of action remain largely unknown. AIM OF THE STUDY: To elucidate the therapeutic mechanism of JTY in EMs, focusing on endoplasmic reticulum (ER) stress-induced autophagy. MATERIALS AND METHODS: The major components of JTY were detected using high-performance liquid chromatography-mass spectrometry (HPLC-MS). The potential mechanism of JTY in EMs treatment was predicted using network pharmacological analysis. Finally, the pathogenesis of EMs was validated in a clinical case-control study and the molecular mechanism of JTY was validated in vitro using endometrial stromal cells (ESCs). RESULTS: In total, 241 compounds were analyzed and identified from JTY using UPLC-MS. Network pharmacology revealed 288 targets between the JTY components and EMs. Results of the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses indicated that regulating autophagy, migration, apoptosis, and inflammation were the key mechanisms of JTY in treating EMs. Meanwhile, we found that protein kinase R-like endoplasmic reticulum kinase (PERK), Beclin-1, and microtubule-associated protein light chain 3 B (LC3B) expressions were lower in endometria of patients with EMs than in those with normal eutopic endometria (p < 0.05). Additionally, during in vitro experiments, treatment with 20% JTY-containing serum significantly suppressed ESC proliferation, achieving optimal effects after 48 h. Electron microscopy revealed significantly increased autophagy flux in the JTY group compared with the control group. Moreover, JTY treatment significantly reduced the migratory and invasive abilities of ESCs and upregulated protein expression of PERK, eukaryotic initiation factor 2α (eIF2α)/phospho-eukaryotic initiation factor 2α (p-eIF2α), activating Transcription Factor-4 (ATF4), Beclin-1, and LC3BII/I, while subsequently downregulating NOD-like receptor thermal protein domain associated protein 3 (NLRP3) and interleukin 18 (IL-18) expression. However, administration of GSK2656157-a highly selective PERK inhibitor-reversed these changes. CONCLUSION: JTY ameliorates EMs by activating PERK associated with unfolded protein reaction, enhancing cell ER stress and autophagy, improving the inflammatory microenvironment, and decreasing the migration and invasion of ESCs.

Selenomethionine alleviates chronic heat stress-induced breast muscle injury and poor meat quality in broilers via relieving mitochondrial dysfunction and endoplasmic reticulum stress.

In the present study, the chronic heat stress (CHS) broiler model was developed to investigate the potential protection mechanism of organic selenium (selenomethionine, SeMet) on CHS-induced skeletal muscle growth retardation and poor meat quality. Four hundred Arbor Acres male broilers (680 ± 70 g, 21 d old) were grouped into 5 treatments with 8 replicates of 10 broilers per replicate. Broilers in the control group were raised in a thermoneutral environment (22 ± 2 °C) and fed with a basal diet. The other four treatments were exposed to hyperthermic conditions (33 ± 2 °C, 24 h in each day) and fed on the basal diet supplied with SeMet at 0.0, 0.2, 0.4, and 0.6 mg Se/kg, respectively, for 21 d. Results showed that CHS reduced (P < 0.05) the growth performance, decreased (P < 0.05) the breast muscle weight and impaired the meat quality of breast muscle in broilers. CHS induced protein metabolic disorder in breast muscle, which increased (P < 0.05) the expression of caspase 3, caspase 8, caspase 9 and ubiquitin proteasome system related genes, while decreased the protein expression of P-4EBP1. CHS also decreased the antioxidant capacity and induced mitochondrial stress and endoplasmic reticulum (ER) stress in breast muscle, which increased (P < 0.05) the ROS levels, decreased the concentration of ATP, increased the protein expression of HSP60 and CLPX, and increased (P < 0.05) the expression of ER stress biomarkers. Dietary SeMet supplementation linearly increased (P < 0.05) breast muscle Se concentration and exhibited protective effects via up-regulating the expression of the selenotranscriptome and several key selenoproteins, which increased (P < 0.05) body weight, improved meat quality, enhanced antioxidant capacity and mitigated mitochondrial stress and ER stress. What's more, SeMet suppressed protein degradation and improved protein biosynthesis though inhibiting the caspase and ubiquitin proteasome system and promoting the mTOR-4EBP1 pathway. In conclusion, dietary SeMet supplementation increases the expression of several key selenoproteins, alleviates mitochondrial dysfunction and ER stress, improves protein biosynthesis, suppresses protein degradation, thus increases the body weight and improves meat quality of broilers exposed to CHS.

The role of vitamin E in polyunsaturated fatty acid synthesis and alleviating endoplasmic reticulum stress in sub-adult grass carp (Ctenopharyngodon idella).

Vitamin E (VE) is an essential lipid-soluble vitamin that improves the fish flesh quality. However, the underlying molecular mechanisms remain unclear. This study aimed to investigate the effects of VE on growth performance and flesh quality in sub-adult grass carp (Ctenopharyngodon idella). A total of 450 fish (713.53 ± 1.50 g) were randomly divided into six treatment groups (three replicates per treatment) and fed for nine weeks with different experimental diets (dietary lipid 47.8 g/kg) that contained different levels of VE (5.44, 52.07, 96.85, 141.71, 185.66, and 230.12 mg/kg diet, supplemented as dl-α-tocopherol acetate). Notably, the treatment groups that were fed with dietary VE ranging from 52.07 to 230.12 mg/kg diet showed improvement in the percent weight gain, special growth rate, and feed efficiency of grass carp. Moreover, the treatment groups supplemented with dietary VE level of 141.71, 185.66, and 230.12 mg/kg diet showed enhancement in crude protein, lipid, and α-tocopherol contents in the muscle, and the dietary levels of VE ranging from 52.07 to 141.71 mg/kg diet improved muscle pH24h and shear force but reduced muscle cooking loss in grass carp. Furthermore, appropriate levels of VE (52.07 to 96.85 mg/kg diet) increased the muscle polyunsaturated fatty acid content in grass carp. Dietary VE also increased the mRNA levels of fatty acid synthesis-related genes, including fas, scd-1, fad, elovl, srebp1, pparγ, and lxrα, and up-regulated the expression of SREBP-1 protein. However, dietary VE decreased the expression of fatty acid decomposition-related genes, including hsl, cpt1, acox1, and pparα, and endoplasmic reticulum stress-related genes, including perk, ire1, atf6, eif2α, atf4, xbp1, chop, and grp78, and down-regulated the expression of p-PERK, p-IRE1, ATF6, and GRP78 proteins. In conclusion, dietary VE increased muscle fatty acid synthesis, which may be partly associated with the alleviation of endoplasmic reticulum stress, and ultimately improves fish flesh quality. Moreover, the VE requirements for sub-adult grass carp (713.53 to 1590.40 g) were estimated to be 124.9 and 122.73 mg/kg diet based on percentage weight gain and muscle shear force, respectively.

Xingnaojing injection alleviates cerebral ischemia/reperfusion injury through regulating endoplasmic reticulum stress in Vivo and in Vitro.

Background: Xingnaojing (XNJ) injection, an extract derived from traditional Chinese medicine, is commonly used to treat ischemic stroke (IS). Previous studies have shown that XNJ has the ability to alleviate apoptosis in cerebral ischemia-reperfusion injury. However, the potential mechanisms have not been clarified. Objective: To identify the neuroprotective effect of XNJ and explore whether XNJ inhibits cell apoptosis associated with endoplasmic reticulum stress (ERS) after IS. Methods: In this study, cultured hippocampal neurons from mouse embryos and Sprague-Dawley rats were assigned randomly to four groups: sham, model, XNJ, and edaravone. The treatment groups were administered 2 h after modelling. Neurological deficit scores and motor performance tests were performed after 24 h of modelling. Additionally, pathomorphology, cell apoptosis and calcium content were evaluated. To ascertain the expression of ERS proteins, western blotting and polymerase chain reaction were employed. Results: The results indicated that XNJ treatment resulted in a notable decrease in infarct volume, apoptosis and missteps compared with the model group. XNJ also exhibited improvements in neurological function, grip strength and motor time. The calcium content significantly reduced in XNJ group. The XNJ administration resulted in a reduction in the levels of proteins associated with ERS including CHOP, GRP78, Bax, caspase-12, caspase-9, and cleaved-caspase-3, but an increase of the Bcl-2/Bax ratio. Furthermore, the downregulation of mRNA expression of CHOP, GRP78, caspase-12, caspase-9, and caspase-3 was confirmed in both cultured neurons and rat model. Conclusion: These findings suggest that XNJ may alleviate apoptosis by modulating the ERS-induced apoptosis pathway, making it a potential novel therapeutic approach for ischemic stroke.

Betulinic acid induces apoptosis of HeLa cells via ROS-dependent ER stress and autophagy in vitro and in vivo.

Betulinic acid (BA), a naturally occurring lupane-type triterpenoid, possesses a wide range of potential activities against different types of cancer. However, the molecular mechanisms involved in anti-cervical cancer about BA were rarely investigated. Herein, the role of BA in cervical cancer suppression by ROS-mediated endoplasmic reticulum stress (ERS) and autophagy was deeply discussed. The findings revealed that BA activated Keap1/Nrf2 pathway and triggered mitochondria-dependent apoptosis due to ROS production. Furthermore, BA increased the intracellular Ca2+ levels, inhibited the expression of Beclin1 and promoted the expression of GRP78, LC3-II, and p62 associated with ERS and autophagy. Besides, BA initiated the formation of autophagosomes and inhibited autophagic flux by the co-administration of BA with 3-methyladenine (3-MA) and chloroquine (CQ), respectively. The in vivo experiment manifested that hydroxychloroquine (HCQ) enhanced the apoptosis induced by BA. For the first time, we demonstrated that BA could initiate early autophagy, inhibit autophagy flux, and induce protective autophagy in HeLa cells. Thus, BA could be a potential chemotherapy drug for cervical cancer, and inhibition of autophagy could enhance the anti-tumor effect of BA. However, the interactions of signaling factors between ERS-mediated and autophagy-mediated apoptosis deserve further attention.