C/EBP Homologous Protein Research Articles

Coordinated stimulation of axon regenerative and neurodegenerative transcriptional programs by ATF4 following optic nerve injury.

Stress signaling is important for determining the fates of neurons following axonal insults. Previously we showed that the stress-responsive kinase PERK contributes to injury-induced neurodegeneration (Larhammar et al., 2017). Here we show that PERK acts primarily through Activating Transcription Factor-4 (ATF4) to stimulate not only pro-apoptotic but also pro-regenerative responses following optic nerve damage. Using conditional knockout mice, we find an extensive PERK/ATF4-dependent transcriptional response that includes canonical ATF4 target genes and modest contributions by C/EBP Homologous Protein (CHOP). Overlap with c-Jun-dependent transcription suggests interplay with a parallel stress pathway that orchestrates regenerative and apoptotic responses. Accordingly, neuronal knockout of ATF4 recapitulates the neuroprotection afforded by PERK deficiency, and PERK or ATF4 knockout impairs optic axon regeneration enabled by disrupting the tumor suppressor PTEN. These findings reveal an integral role for PERK/ATF4 in coordinating neurodegenerative and regenerative responses to CNS axon injury.

Taraxasterol attenuates zearalenone-induced kidney damage in mice by modulating oxidative stress and endoplasmic reticulum stress

Taraxasterol is one of the bioactive ingredients from traditional Chinese herb Taraxacum, which exhibits multiple pharmacological activities and protective effects. However, the underlying influence and mechanism of its use against kidney damage caused from zearalenone (ZEA) remain unexplored. The ZEA-induced kidney damage model of mice was established by feeding diets containing ZEA (2 mg/kg), and taraxasterol (5 and 10 mg/kg) was administered by gavage for 28 days. Results demonstrated taraxasterol increased average daily gain (ADG) and average daily feed intake (ADFI), reduced feed-to-gain ratio (F/G) and kidney index of mice induced by ZEA. Taraxasterol alleviated histopathological changes of kidney, reduced ZEA residue and the levels of blood urea nitrogen (BUN), uric acid (UA), and creatinine (CRE). Concurrently, taraxasterol reduced the contents of oxidative stress indicator reactive oxygen species (ROS) and malondialdehyde (MDA), and increased the activities of antioxidant enzymes catalase (CAT), total superoxide dismutase (T-SOD), and glutathione peroxidase (GSH-Px). Further, taraxasterol up-regulated the mRNA and protein expression of nuclear factor erythroid-2-related factor 2 (Nrf2), GSH-Px, NAD(P)H quinone oxidoreductase 1 (NQO1), and heme oxygenase-1 (HO-1), and down-regulated the mRNA and protein expression of KELCH like ECH associated protein (Keap1) in Nrf2/Keap1 pathway. Taraxasterol down-regulated the mRNA and protein expression of immunoglobulin binding protein (Bip), C/EBP homologous protein (CHOP), Bcl-2 associated X (Bax), cysteine protease (Caspase)-12, and Caspase-3, and up-regulated B-cell lymphoma 2 (Bcl-2) expression in endoplasmic reticulum stress pathway. This study suggests that taraxasterol attenuates ZEA-induced mouse kidney damage through the modulation of Nrf2/Keapl pathway to play antioxidant role and endoplasmic reticulum stress pathway to enhance anti-apoptotic ability. It will provide a basis for taraxasterol as a potential drug to prevent and treat ZEA-induced kidney damage.

J147 treatment protects against traumatic brain injury by inhibiting neuronal endoplasmic reticulum stress potentially via the AMPK/SREBP-1 pathway

Endoplasmic reticulum (ER) stress is recognized as a crucial contributor to the progression of traumatic brain injury (TBI) and represents a potential target for therapeutic intervention. This study aimed to assess the potential of J147, a novel neurotrophic compound, in alleviating ER stress by modulating related signaling pathways, thereby promoting functional recovery in TBI. To this end, adult mice underwent controlled cortical impact (CCI) injury to induce TBI, followed by oral administration of J147 one-hour post-injury, with daily dosing for 3 to 7 days. Multiple behavioral assessments were conducted over 35 days, revealing a significant, dose-dependent improvement in neurofunctional recovery with J147 treatment. The neuropathological analysis demonstrated reduced acute neurodegeneration (observed at three days through FJC staining), enhanced long-term neuron survival (H&E and Nissl staining), and improved neuroplasticity (Golgi staining) at 35 days post-TBI. At the molecular level, TBIinduced AMP-activated protein kinase (AMPK) dephosphorylation, sterol regulatory element binding protein-1 (SREBP-1) activation, and upregulation of ER stress marker proteins, including phosphorylated eukaryotic initiation factor-2α (p-eIF2a), activating transcription factor 4 (ATF4), and C/EBP homologous protein (CHOP) in perilesional cortex neurons at three days post-injury. Notably, the J147 treatment significantly attenuated AMPK dephosphorylation, SERBP-1 activation, and expression of the ER stress markers. In summary, this study reveals the therapeutic promise of J147 in mitigating secondary brain damage associated with TBI and improving long-term functional recovery by modulating ER stress pathways.

Apoptosis of Pancreatic Cancer Cells after Co-Treatment with Eugenol and Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand.

Pancreatic cancer is a refractory cancer with limited treatment options. Various cancer types are resistant to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Eugenol, the main component of clove oil, exhibits anticancer, anti-inflammatory, and antioxidant effects. However, no studies have reported that eugenol increases TRAIL sensitivity by upregulating death receptor (DR) expression. Here, we aimed to investigate eugenol as a potent TRAIL sensitizer. Increased apoptosis and inhibition of cell proliferation was observed in pancreatic cancer cells treated with eugenol and TRAIL compared with those treated with eugenol alone. Eugenol upregulated the expression of DR5, inhibited the FLICE-inhibitory protein (FLIP), an anti-apoptotic protein, and increased p53, a tumor suppressor protein. In addition, eugenol induced the generation of reactive oxygen species (ROS) and caused endoplasmic reticulum (ER) stress. C/EBP-homologous protein (CHOP) knockdown using siRNA decreased the expression of DR5 and reduced the combined effects of eugenol and TRAIL. These results demonstrate that eugenol enhances TRAIL-induced apoptosis by upregulating DR5 through the ROS-mediated ER stress-CHOP pathway, which enhances ER stress by inducing p53 and downregulating FLIP expression. This suggests that eugenol has the potential to treat pancreatic cancer by increasing cell sensitivity to TRAIL.

Polydatin ameliorates early brain injury after subarachnoid hemorrhage through up-regulating SIRT1 to suppress endoplasmic reticulum stress.

This study aims to investigate the inhibitory effect of Polydatin (PD) on endoplasmic reticulum (ER) stress following subarachnoid hemorrhage (SAH) and to elucidate the underlying mechanisms. A standard intravascular puncture model was established to mimic SAH in mice. Neurological functions were assessed using neurological scoring, Grip test, and Morris water maze. Brain edema and Evans blue extravasation were measured to evaluate blood-brain barrier permeability. Western blot and quantitative real-time polymerase chain reaction (PCR) analyses were performed to examine protein and mRNA expressions related to ER stress. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining was used to detect cell apoptosis, and transmission electron microscopy was used to observe the ultrastructure of the endoplasmic reticulum. The results indicated that PD significantly reduced brain edema and Evans blue extravasation after SAH, improving neurological function. Compared to the SAH group, the expression levels of ER stress-related proteins including glucose-regulated protein 78 (GRP78), phosphorylated protein kinase R-like endoplasmic reticulum kinase (p-PERK), phosphorylated eukaryotic initiation factor 2α (p-eIF2α), activating transcription factor 4 (ATF4), and C/EBP homologous protein (CHOP), were significantly lower in the PD-treated group. Moreover, PD significantly enhances the protein expression of Sirtuin 1 (SIRT1). Validation with sh-SIRT1 confirmed the critical role of SIRT1 in ER stress, with PD's inhibitory effect on ER stress being dependent on SIRT1 expression. Additionally, PD attenuated ER stress-mediated neuronal apoptosis and SAH-induced ferroptosis through upregulation of SIRT1. PD alleviates ER stress following SAH by upregulating SIRT1 expression, thereby mitigating early brain injury. The protective effects of PD are mediated through SIRT1, which inhibits ER stress and reduces neuronal apoptosis and ferroptosis.

Palmitic acid-induced cell death: impact of endoplasmic reticulum and oxidative stress, mitigated by L-citrulline.

Palmitic acid (PA), the most abundant saturated free fatty acids, induces apoptosis in bovine mammary epithelial cells. It is suggested that oxidative stress and endoplasmic reticulum (ER) stress are key mechanisms underlying PA-induced cell death. This study aimed to investigate the interaction between ER stress and oxidative stress during PA-induced cell death in MAC-T cells. Additionally, we examined whether L-citrulline can protect against PA-induced damage of MAC-T cells. MAC-T cells were treated with 4-phenyl butyric acid (4-PBA) or N-acetyl-L-cysteine (NAC) to inhibit PA-induced ER stress and oxidative stress, respectively. MAC-T cells were pretreated with or without L-citrulline for 48 h followed by PA treatment. Cell viability was measured with MTT assays. Intracellular reactive oxygen species (ROS) levels in MAC-T cells were assessed using 5-(and-6)-chloromethyl- 2`,7`-dichlorodihydrofluorescein diacetate acetyl ester dye. Real-time qPCR was used to explore the regulation of genes associated with oxidative stress, and ER stress genes. Western blotting analysis was also carried out. 4-PBA significantly reduced PA-induced mRNA expressions of activating transcription factor 4 (ATF4), C/EBP homologous protein (CHOP), nuclear factor (erythroid-derived 2)-like 2 (NRF2), and intracellular ROS levels. Furthermore, NAC dramatically reduced PA-induced ROS levels and the mRNA expressions of NRF2, ATF4, and CHOP. L-citrulline pretreatment effectively rescued cell viability decreased by PA. Moreover, L-citrulline pretreatment significantly downregulated the PA-induced upregulation of GRP78, ATF4, and CHOP mRNA expression, and protein expression of p-PERK and cleaved caspase-3. PA increased intracellular ROS levels and NRF2 mRNA expression, whereas L-citrulline pretreatment remarkably reduced these levels. Both ER and oxidative stresses interact during PA-induced cell death in MAC-T cells, and L-citrulline could attenuate this cell death by inhibiting ER and oxidative stresses. Therefore, L-citrulline may be a promising supplement for protecting against PA-induced cell death in bovine MECs during the lactation period of dairy cows.

Evaluation of Endoplasmic Reticulum Stress in an Experimental Intestinal Ischemia-Reperfusion Model in Rats: The Role of Ozone Therapy and Trimetazidine.

The objective of the study was to assess the impact of ozone (O3) and trimetazidine on the intestines following ischemia-reperfusion (I/R) injury through the investigation of endoplasmic reticulum stress. Forty Sprague Dawley rats were separated into five groups. The groups were named as follows: control, O3, I/R, I/R + trimetazidine (TMZ), and I/R + O3. The control group had laparotomy and exploration of the superior mesenteric artery (SMA) only. Furthermore, alongside laparotomy and SMA exploration, an intraperitoneal (i.p.) administration of a 0.7 mg/kg ozone-oxygen (O3-O2) combination was given to the O3 group. In the experimental groups, the SMA was blocked with the silk suture ligation technique for a duration of 1 h and then restored to normal blood flow for another hour. In the I/R + O3 group, ozone was delivered i.p. at a dosage of 0.7 mg/kg, 30 min after ischemia. In the I/R + TMZ group, a dose of 20 mg/kg/day of trimetazidine was administered orally via gavage for a duration of 7 days, beginning 1 week prior to the induction of ischemia. Intestinal tissues were taken to assess indicators of intestinal mucosal injury and oxidative stress. The level of the lipid peroxidation marker malondialdehyde (MDA) was significantly reduced in the experimental groups as compared to the I/R group (p < 0.05). The experimental groups had considerably greater levels of glutathione (GSH), which reflects antioxidant capacity, compared to the I/R group (p < 0.05). Nevertheless, the concentration of GSH was observed to be increased in the I/R + O3 group in comparison to the I/R + TMZ group (p < 0.05). The histopathological damage score showed a substantial decrease in the experimental groups as compared to the I/R group (p < 0.05). The I/R + O3 group had the lowest injury score. The experimental groups exhibited significantly reduced positivity of the endoplasmic reticulum (ER) stress markers C/EBP homologous protein (CHOP) and glucose-regulated protein (GRP)-78 compared to the I/R group (p < 0.05). The findings provide evidence for the potential advantages of utilizing ozone therapy in the treatment of intestinal ischemia-reperfusion injury. Additionally, they propose that ozone should be assessed in more extensive clinical trials in the future as a therapeutic agent that can disrupt endoplasmic reticulum stress.

The renoprotective effects of cannabidiol on lipopolysaccharide-induced systemic inflammation model of rats.

Sepsis-induced renal damage poses a significant threat, necessitating effective therapeutic strategies. Cannabidiol (CBD) has beneficial effects on tissues and their functions by exhibiting antioxidant and anti-inflammatory effects. This study investigates the potential protective effects of CBD in mitigating lipopolysaccharide (LPS)-induced renal injury in Wistar Albino rats. Thirty-two Wistar Albino rats were categorized into control, LPS (5mg/kg i.p.), LPS + CBD, and CBD (5mg/kg i.p.) groups. After the experiment, samples were collected for biochemical, genetic, histopathological, and immunohistochemical analyses. Oxidative stress markers as total oxidant status (TOS) and total antioxidant status (TAS), oxidative stress index (OSI), superoxide dismutase (SOD), glutathione peroxidase (GPx), malondialdehyde (MDA), immune staining as tumor necrosis factor alpha (TNF-α), interleukin-10 (IL-10), caspase-3, gene expressions as nuclear factor erythroid 2-related factor 2 (NRF2), C/EBP homologous protein (CHOP), caspase-9, glucose-regulating protein 78 (GRP78), B-cell leukemia/lymphoma 2 (Bcl2), and tissue histology have been examined. The LPS-exposed group exhibited significant renal abnormalities, mitigated by CBD intervention in the LPS + CBD group. CBD reduced immunoexpression scores for TNF-α, caspase-3, and IL-10. Biochemically, CBD induced a positive shift in the oxidative balance, increasing TAS, SOD, and GPx, while decreasing TOS, OSI, and MDA levels. Genetic analyses highlighted CBD's regulatory impact on NRF2, CHOP, caspase-9, GRP78, and Bcl2, providing molecular insights into its protective role against LPS-induced renal damage. This study underscores CBD as a promising protective agent against sepsis-induced renal damage. Our findings could provide valuable insights into potential therapeutic avenues for addressing renal complications in sepsis.

Safflower Yellow Injection Alleviates Myocardial Ischemia/Reperfusion Injury by Reducing Oxidative and Endoplasmic Reticulum Stress.

Safflower yellow is an extract of the famous Chinese medicine Carthamus tinctorious L, and safflower yellow injection (SYI) is widely used clinically to treat angina pectoris. However, there are few studies on the anti-myocardial ischemia/reperfusion (I/R) injury effect of SYI, and its mechanisms are unclear. In the present study, we aimed to investigate the protective effect of SYI on myocardial I/R injury and explore its underlying mechanisms. Male Sprague Dawley rats were randomly divided into a control group, sham group, model group, and SYI group (20 mg/kg, femoral vein injection 1 h before modeling). The left anterior descending coronary artery was ligated to establish a myocardial I/R model. H9c2 cells were exposed to oxygen-glucose deprivation/reoxygenation (OGD/R) after incubation with 80 μg/mL SYI for 24 h. In vivo, TsTC, HE, and TUNEL staining were performed to evaluate myocardial injury and apoptosis. A kit was used to detect superoxide dismutase (SOD) and malondialdehyde (MDA) to assess oxidative stress. In vitro, flow cytometry was used to detect the reactive oxygen species (ROS) content and apoptosis rate. Protein levels were determined via Western blotting. Pretreatment with SYI significantly reduced infarct size and pathological damage in rat hearts and suppressed cardiomyocyte apoptosis in vivo and in vitro. In addition, SYI inhibited oxidative stress by increasing SOD activity and decreasing MDA content and ROS production. Myocardial I/R and OGD/R activate endoplasmic reticulum (ER) stress, as evidenced by increased expression of activating transcription factor 6 (ATF6), glucose-regulated protein 78 (GRP78), cysteinyl aspartate-specific proteinase caspase-12, and C/EBP-homologous protein (CHOP), which were all inhibited by SYI. SYI ameliorated myocardial I/R injury by attenuating apoptosis, oxidative damage, and ER stress, which revealed new mechanistic insights into its application.

1,25-D3 ameliorates ischemic brain injury by alleviating endoplasmic reticulum stress and ferroptosis: Involvement of vitamin D receptor and p53 signaling

Endoplasmic reticulum stress (ERS) and ferroptosis are linked to cerebral ischemia reperfusion injury (CIRI). The neuroprotective properties of 1α, 25-dihydroxyvitamin D3 (VitD3 or 1,25-D3) have been well established; however, the mechanism by which VitD3 treats CIRI through ERS and ferroptosis has not been examined. Hence, we developed middle cerebral artery occlusion/reperfusion (MCAO/R) model in SD rats to ascertain if VitD3 preconditioning mediates ERS and ferroptosis involving of p53 signaling. In this study, we observed that VitD3 can reduce infarction volume and cerebral edema, which leads to the improvement of nerve function. HE, Nissl and Tunel staining showed that VitD3 treatment significantly improved the morphology of neuronal cells and reduced their death. The expression and activation of Vitamin D receptor (VDR), PKR-like ER kinase (PERK), C/EBP-homologous protein (CHOP), p53, nuclear factor erythroid 2-related factor 2 (Nrf2), glutathione peroxidase 4 (GPX4) and reactive oxygen species (ROS) in the ischemic penumbral area were detected by real-time qPCR, Western-blotting and Elisa. The results showed that after VitD3 treatment, VDR increased, ERS-related indices (PERK, CHOP) significantly decreased and ferroptosis-related indices (Nrf2, GPX4) increased. As a VDRs antagonist, pyridoxal-5-phosphate (P5P) can partially block the neuroprotective effects of VitD3. Therefore, CIRI can induce ERS and ferroptosis in the ischemic penumbra area and VitD3 may ameliorate nerve damage in CIRI rats by up-regulating VDR, alleviating p53-associated ERS and ferroptosis.

Ferroptosis is involved in Staphylococcus aureus-induced mastitis through autophagy activation by endoplasmic reticulum stress

Cell death caused by severe Staphylococcus aureus (S. aureus) infection is a fatal threat to humans and animals. However, whether ferroptosis, an iron-dependent form of cell death, is involved in S. aureus-induced cell death and its role in S. aureus-induced diseases are unclear. Using a mouse mastitis model and mammary epithelial cells (MMECs), we investigated the role of ferroptosis in the pathogenesis of S. aureus infection. The results revealed that S. aureus-induced ferroptosis in vivo and in vitro as demonstrated by dose-dependent increases in cell death; the level of malondialdehyde (MDA), the final product of lipid peroxidation; and dose-dependent decrease the production of the antioxidant glutathione (GSH). Treatment with typical inhibitors of ferroptosis, including ferrostatin-1 (Fer-1) and deferiprone (DFO), significantly inhibited S. aureus-induced death in MMECs. Mechanistically, treatment with S. aureus activated the protein kinase RNA-like ER kinase (PERK)-eukaryotic initiation factor 2, α subunit (eIF2α)-activating transcription factor 4 (ATF4)-C/EBP homologous protein (CHOP) pathway, which subsequently upregulated autophagy and promoted S. aureus-induced ferroptosis. The activation of autophagy degraded ferritin, resulting in iron dysregulation and ferroptosis. In addition, we found that excessive reactive oxygen species (ROS) production induced ferroptosis and activated endoplasmic reticulum (ER) stress, manifesting as elevated p-PERK-p-eIF2α-ATF4-CHOP pathway protein levels. Collectively, our findings indicate that ferroptosis is involved in S. aureus-induced mastitis via ER stress-mediated autophagy activation, implying a potential strategy for the prevention of S. aureus-associated diseases by targeting ferroptosis. In conclusion, the ROS-ER stress-autophagy axis is involved in regulating S. aureus-induced ferroptosis in MMECs. These findings not only provide a new potential mechanism for mastitis induced by S. aureus but also provide a basis for the treatment of other ferroptotic-related diseases.

Enterovirus 3A protein disrupts endoplasmic reticulum homeostasis through interaction with GBF1.

Enteroviruses are single-stranded, positive-sense RNA viruses causing endoplasmic reticulum (ER) stress to induce or modulate downstream signaling pathways known as the unfolded protein responses (UPR). However, viral and host factors involved in the UPR related to viral pathogenesis remain unclear. In the present study, we aimed to identify the major regulator of enterovirus-induced UPR and elucidate the underlying molecular mechanisms. We showed that host Golgi-specific brefeldin A-resistant guanine nucleotide exchange factor 1 (GBF1), which supports enteroviruses replication, was a major regulator of the UPR caused by infection with enteroviruses. In addition, we found that severe UPR was induced by the expression of 3A proteins encoded in human pathogenic enteroviruses, such as enterovirus A71, coxsackievirus B3, poliovirus, and enterovirus D68. The N-terminal-conserved residues of 3A protein interact with the GBF1 and induce UPR through inhibition of ADP-ribosylation factor 1 (ARF1) activation via GBF1 sequestration. Remodeling and expansion of ER and accumulation of ER-resident proteins were observed in cells infected with enteroviruses. Finally, 3A induced apoptosis in cells infected with enteroviruses via activation of the protein kinase RNA-like endoplasmic reticulum kinase (PERK)/C/EBP homologous protein (CHOP) pathway of UPR. Pharmaceutical inhibition of PERK suppressed the cell death caused by infection with enteroviruses, suggesting the UPR pathway is a therapeutic target for treating diseases caused by infection with enteroviruses.IMPORTANCEInfection caused by several plus-stranded RNA viruses leads to dysregulated ER homeostasis in the host cells. The mechanisms underlying the disruption and impairment of ER homeostasis and its significance in pathogenesis upon enteroviral infection remain unclear. Our findings suggested that the 3A protein encoded in human pathogenic enteroviruses disrupts ER homeostasis by interacting with GBF1, a major regulator of UPR. Enterovirus-mediated infections drive ER into pathogenic conditions, where ER-resident proteins are accumulated. Furthermore, in such scenarios, the PERK/CHOP signaling pathway induced by an unresolved imbalance of ER homeostasis essentially drives apoptosis. Therefore, elucidating the mechanisms underlying the virus-induced disruption of ER homeostasis might be a potential target to mitigate the pathogenesis of enteroviruses.

SIRT1 inhibits apoptosis of human lens epithelial cells through suppressing endoplasmic reticulum stress in vitro and in vivo.

To explore the effect of silent information regulator factor 2-related enzyme 1 (SIRT1) on modulating apoptosis of human lens epithelial cells (HLECs) and alleviating lens opacification of rats through suppressing endoplasmic reticulum (ER) stress. HLECs (SRA01/04) were treated with varying concentrations of tunicamycin (TM) for 24h, and the expression of SIRT1 and C/EBP homologous protein (CHOP) was assessed using real-time quantitative polymerase chain reaction (RT-PCR), Western blotting, and immunofluorescence. Cell morphology and proliferation was evaluated using an inverted microscope and cell counting kit-8 (CCK-8) assay, respectively. In the SRA01/04 cell apoptosis model, which underwent siRNA transfection for SIRT1 knockdown and SRT1720 treatment for its activation, the expression levels of SIRT1, CHOP, glucose regulated protein 78 (GRP78), and activating transcription factor 4 (ATF4) were examined. The potential reversal of SIRT1 knockdown effects by 4-phenyl butyric acid (4-PBA; an ER stress inhibitor) was investigated. In vivo, age-related cataract (ARC) rat models were induced by sodium selenite injection, and the protective role of SIRT1, activated by SRT1720 intraperitoneal injections, was evaluated through morphology observation, hematoxylin and eosin (H&E) staining, Western blotting, and RT-PCR. SIRT1 expression was downregulated in TM-induced SRA01/04 cells. Besides, in SRA01/04 cells, both cell apoptosis and CHOP expression increased with the rising doses of TM. ER stress was stimulated by TM, as evidenced by the increased GRP78 and ATF4 in the SRA01/04 cell apoptosis model. Inhibition of SIRT1 by siRNA knockdown increased ER stress activation, whereas SRT1720 treatment had opposite results. 4-PBA partly reverse the adverse effect of SIRT1 knockdown on apoptosis. In vivo, SRT1720 attenuated the lens opacification and weakened the ER stress activation in ARC rat models. SIRT1 plays a protective role against TM-induced apoptosis in HLECs and slows the progression of cataract in rats by inhibiting ER stress. These findings suggest a novel strategy for cataract treatment focused on targeting ER stress, highlighting the therapeutic potential of SIRT1 modulation in ARC development.

Delivery of CDNF by AAV-mediated gene transfer protects dopamine neurons and regulates ER stress and inflammation in an acute MPTP mouse model of Parkinson’s disease

Cerebral dopamine neurotrophic factor (CDNF) and its close structural relative, mesencephalic astrocyte-derived neurotrophic factor (MANF), are proteins with neurotrophic properties. CDNF protects and restores the function of dopamine (DA) neurons in rodent and non-human primate (NHP) toxin models of Parkinson’s disease (PD) and therefore shows promise as a drug candidate for disease-modifying treatment of PD. Moreover, CDNF was found to be safe and to have some therapeutic effects on PD patients in phase 1/2 clinical trials. However, the mechanism underlying the neurotrophic activity of CDNF is unknown. In this study, we delivered human CDNF (hCDNF) to the brain using an adeno-associated viral (AAV) vector and demonstrated the neurotrophic effect of AAV-hCDNF in an acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. AAV-hCDNF resulted in the expression of hCDNF in the striatum (STR) and substantia nigra (SN), and no toxic effects on the nigrostriatal pathway were observed. Intrastriatal injection of AAV-hCDNF reduced motor impairment and partially alleviated gait dysfunction in the acute MPTP mouse model. In addition, gene therapy with AAV-hCDNF had significant neuroprotective effects on the nigrostriatal pathway and decreased the levels of interleukin 1beta (IL-1β) and complement 3 (C3) in glial cells in the acute MPTP mouse model. Moreover, AAV-hCDNF reduced C/EBP homologous protein (CHOP) and glucose regulatory protein 78 (GRP78) expression in astroglia. These results suggest that the neuroprotective effects of CDNF may be mediated at least in part through the regulation of neuroinflammation and the UPR pathway in a mouse MPTP model of PD in vivo.

Flow shear force destabilizes carotid plaques by affecting CHOP and GRP78 proteins

BackgroundVarious factors, including blood, inflammatory, infectious, and immune factors, can cause ischemic stroke. However, the primary cause is often the instability of cervical arteriosclerosis plaque. It is estimated that 18-25% of ischemic strokes are caused by the rupture of carotid plaque.1 Plaque stability is crucial in determining patient prognosis. Developing a highly accurate, non-invasive, or minimally invasive technique to assess carotid plaque stability is crucial for diagnosing and treating stroke.Previous research by our group has demonstrated that the expression levels of CHOP (C/EBP homologous protein) and GRP78 (glucose-regulated protein 78) are correlated with the stability of atherosclerotic plaques.2 ObjectThis research assesses changes in GRP78 and CHOP expressions in human umbilical vein endothelial cells(HUVEC) following experiments within the hemodynamic influencing factors test system. Additionally, it includes conducting an empirical study on the impact of blood flow shear force on the stability of human carotid atherosclerotic plaques. The objective is to explore the implications of blood flow shear force on the stability of carotid atherosclerotic plaques. MethodThe hemodynamic influencing factors test bench system was configured with low (Group A, 4 dyns/cm²), medium (Group B, 8 dyns/cm²), and high shear force groups (Group C, 12 dyns/cm²). Relative expression levels of GRP78 and CHOP proteins in human umbilical vein endothelial cells were measured using Western blot analysis, and quantitative analysis of GRP78 and CHOP mRNA was conducted using RT-qPCR. Meanwhile, plaques from 60 carotid artery patients, retrieved via Carotid Endarterectomy (CEA), were classified into stable (S) and unstable (U) groups based on pathological criteria. Shear force at the carotid bifurcation was measured preoperatively using ultrasound. Western blot and RT-qPCR were used to analyze the relative expression levels of GRP78 and CHOP proteins and mRNA, respectively, in the plaque specimens from both groups. ResultExpression levels of GRP78, CHOP proteins, and their mRNAs were assessed in groups A, B, and C via Western blot and RT-qPCR. Results showed that in the low-shear group, all markers were elevated in group A compared to groups B and C. Statistical analysis revealed significantly lower shear forces at the carotid bifurcation in group U compared to group S. In group U plaques, GRP78 and CHOP expressions were significantly higher in group U than in group S. ConclusionBlood flow shear forces variably affect the expression of GRP78 and CHOP proteins, as well as their mRNA levels, in vascular endothelial cells. The lower the shear force and fluid flow rate, the higher the expression of GRP78 and CHOP, potentially leading to endoplasmic reticulum stress(ERS), which may destabilize the plaque.

Piperine inhibits the proliferation of colorectal adenocarcinoma by regulating ARL3-mediated endoplasmic reticulum stress.

Colorectal adenocarcinoma (COAD) is a significant cause of cancer-related mortality worldwide, necessitating the identification of novel therapeutic targets and treatments. This research aimed to investigate the role of ARL3 in COAD progression and to explore the effects of Piperine on ARL3 expression, cell proliferation, epithelial-mesenchymal transition (EMT), and endoplasmic reticulum (ER) stress. Bioinformatics analysis of The Cancer Genome Atlas (TCGA)-COAD, GSE39582, and GSE44861 datasets assessed ARL3 expression levels. Immunohistochemical data from the Human Protein Atlas (HPA) database confirmed ARL3 overexpression in COAD. The association of ARL3 with COAD clinical parameters and prognosis was also examined. COAD cells were treated with Piperine, and in vitro assays evaluated cell proliferation, apoptosis, EMT marker expression, and ER stress responses. ARL3 overexpression in COAD correlated with poor prognosis and varied across pathological stages. Piperine treatment inhibited COAD cell proliferation in a concentration- and time-dependent manner, as indicated by reduced Ki-67 levels and decreased colony-forming ability. Piperine induced S-phase cell cycle arrest and facilitated apoptosis in COAD cells, evidenced by changes in Bax, Bcl-2, cleaved caspase-3, and cleaved poly (ADP-ribose) polymerase (PARP) levels. Moreover, Piperine downregulated ARL3 expression in COAD cells, thereby suppressing transforming growth factor beta (TGF-β)-induced EMT. Additionally, Piperine attenuated the ARL3-mediated ER stress response, significantly reducing binding immunoglobulin protein (BiP), inositol-requiring enzyme 1 alpha (p-IRE1α), activating transcription factor 6 (ATF6), and C/EBP homologousprotein (CHOP) levels. Piperine exerted anti-cancer effects in COAD by modulating ARL3 expression, disrupting cell cycle progression, inhibiting the EMT pathway, and regulating ER stress. These findings suggest that Piperine holds promise as a therapeutic agent for COAD through its targeting of ARL3.

Apigenin improves testosterone synthesis by regulating endoplasmic reticulum stress

Obesity is a growing epidemic among reproductive-age men, which can cause and exacerbate male infertility by means of associated comorbidities, endocrine abnormalities, and direct effects on the fidelity and throughput of spermatogenesis. A prominent consequence of male obesity is a reduction in testosterone levels. Natural products have shown tremendous potential anti-obesity effects in metabolic diseases. This study aimed to investigate the potential of apigenin (AP) to alleviate testicular dysfunction induced by a high-fat diet (HFD) and to investigate the underlying mechanisms, focusing on endoplasmic reticulum stress (ERS) and testosterone synthesis. A murine model of obesity was established using HFD-fed mice. The effects of AP on obesity, lipid metabolism, testicular dysfunction, and ERS were assessed through various physiological, histological, and molecular techniques. Administration of AP (10 mg/kg) ameliorated HFD-induced obesity and testicular dysfunction in a mouse model, as evidenced by decreased body weight, improved lipid profiles and testicular pathology, and restored protein levels related to testosterone. Furthermore, in vitro studies demonstrated that AP relieved ERS and recovered testosterone synthesis in murine Leydig cells (TM3) treated with free fatty acids (FFAs). It was also observed that AP rescued testosterone synthesis enzymes in TM3 cells, similar to that observed with the inhibitor of the PERK pathway (GSK2606414). In addition, ChIP, qPCR, and gene silencing showed that the C/EBP homologous protein (CHOP) bound directly to the promoter region of steroidogenic STAR and negatively modulated its expression. Collectively, AP has remarkable potential to alleviate HFD-induced obesity and testicular dysfunction. Its protective effects are attributable partly to mitigating ERS and restoring testosterone synthesis in Leydig cells.

Role and mechanism of ginsenoside Rg1 in ameliorating sepsis-induced acute lung injury based on PERK/eIF2α/ATF4/CHOP-induced alveolar epithelial cell apoptosis

The study investigates the therapeutic effects and mechanisms of ginsenoside Rg_1(GRg_1) on sepsis-induced acute lung injury(SALI). A murine model of SALI was created using cecal ligation and puncture(CLP) surgery, and mice were randomly assigned to groups for GRg_1 intervention. Survival and body weight changes were recorded, lung function was assessed with a non-invasive lung function test system, and lung tissue damage was evaluated through HE staining. The content and expression of inflammatory factors were measured by ELISA and qRT-PCR. Apoptosis was examined using flow cytometry and TUNEL staining. The activation and expression of apoptosis-related molecules cysteinyl aspartate specific proteinase 3(caspase-3), B-cell lymphoma-2(Bcl-2), Bcl-2 associated X protein(Bax), and endoplasmic reticulum stress-related molecules protein kinase R-like endoplasmic reticulum kinase(PERK), eukaryotic initiation factor 2α(eIF2α), activating transcription factor 4(ATF4), and C/EBP homologous protein(CHOP) were studied using Western blot and qRT-PCR. In addition, an in vitro model of lipopolysaccharide(LPS)-induced lung alveolar epithelial cell injury was used, with the application of the endoplasmic reticulum stress inducer tunicamycin to validate the action mechanism of GRg_1. RESULTS:: indicated that, when compared to the model group, GRg_1 intervention significantly enhanced the survival time of CLP mice, mitigated body weight loss, and improved impaired lung function indices. The GRg_1-treated mice also displayed reduced lung tissue pathological scores, a reduced lung tissue wet-to-dry weight ratio, and lower protein content in the bronchoalveolar lavage fluid. Serum levels of interleukin-6(IL-6), interleukin-1β(IL-1β), and tumor necrosis factor-α(TNF-α), as well as the mRNA expressions of these cytokines in lung tissues, were decreased. There was a notable decrease in the proportion of apopto-tic alveolar epithelial cells, and down-regulated expressions of caspase-3, Bax, PERK, eIF2α, ATF4, and CHOP and up-regulated expression of Bcl-2 were observed. In vitro findings showed that the apoptosis-lowering and apoptosis-related protein down-regulating effects of GRg_1 were significantly inhibited with the co-application of tunicamycin. Altogether, GRg_1 reduces apoptosis of alveolar epithelial cells, inhibits inflammation in the lungs, alleviates lung injury, and enhances lung function, possibly through the PERK/eIF2α/ATF4/CHOP pathway.

The Flavoring Agent Ethyl Vanillin Induces Cellular Stress Responses in HK-2 Cells.

Flavored e-cigarettes are a popular alternative to cigarette smoking; unfortunately, the extrapulmonary effects are not well-characterized. Human proximal tubule cells were cultured for 24 or 48 h with 0-1000 µM ethyl vanillin (ETH VAN) and cytotoxicity evaluated. Mitochondrial health was significantly diminished following 48 h of exposure, accompanied by significantly decreased spare capacity, coupling efficiency, and ATP synthase expression. ETH VAN at 24 h inhibited glycolysis. The endoplasmic reticulum (ER) stress marker C/EBP homologous protein (CHOP) was increased at 100 μM relative to 500-1000 μM. The downstream proapoptotic marker cleaved caspase-3 subsequently showed a decreasing trend in expression after 48 h of exposure. The autophagy biomarkers microtubule-associated proteins 1A/1B light chain 3 (LC3B-I and LC3B-II) were measured by Western blot. LC3B-II levels and the LC3B-II/LC3B-I ratio increased at 24 h, which suggested activation of autophagy. In contrast, by 48 h, the autophagy biomarker LC3B-II decreased, resulting in no change in the LC3B-II/LC3B-I ratio. Mitophagy biomarker PTEN-induced putative kinase 1 (PINK1) expression decreased after 48 h of exposure. The downstream marker Parkin was not significantly changed after 24 or 48 h. These findings indicate that the flavoring ETH VAN can induce energy pathway dysfunction and cellular stress responses in a renal model.

Licochalcone D exhibits cytotoxicity in breast cancer cells and enhances tumor necrosis factor‐related apoptosis‐inducing ligand‐induced apoptosis through upregulation of death receptor 5

AbstractAnticancer strategies using natural products or derivatives are promising alternatives for cancer treatment. Here, we showed that licochalcone D (LCD), a natural flavonoid extracted from Glycyrrhiza uralensis Fisch, suppressed the growth of breast cancer cells, and was less toxic to MCF‐10A normal breast cells. LCD‐induced DNA damage, cell cycle arrest, and apoptosis in breast cancer cells. Furthermore, LCD potentiated tumor necrosis factor‐related apoptosis‐inducing ligand (TRAIL)‐induced cytotoxicity. Mechanistically, LCD was revealed to reduce survival protein expression and to upregulate death receptor 5 (DR5) expressions. Silencing DR5 blocked the ability of LCD to sensitize cells to TRAIL‐mediated apoptosis. LCD increased CCAAT/enhancer‐binding protein homologous protein (CHOP) expression in breast cancer cells. Knockdown of CHOP attenuated DR5 upregulation and apoptosis triggered by cotreatment with LCD and TRAIL. Furthermore, LCD suppressed the phosphorylation of extracellular signal‐regulated kinase and promoted the phosphorylation of c‐Jun amino‐terminal kinase (JNK) and p38 mitogen‐activated protein kinase (MAPK). Pretreatment with JNK inhibitor SP600125 or p38 MAPK inhibitor SB203580 abolished the upregulation of DR5 and CHOP, and also attenuated LCD plus TRAIL‐induced cleavage of poly(ADP‐ribose) polymerase. Overall, our results show that LCD exerts cytotoxic effects on breast cancer cells and arguments TRAIL‐mediated apoptosis by inhibiting survival protein expression and upregulating DR5 in a JNK/p38 MAPK‐CHOP‐dependent manner.