Induction Of Endoplasmic Reticulum Stress Research Articles

Understanding the impact of ER stress on lung physiology

Human lungs consist of a distinctive array of cell types, which are subjected to persistent challenges from chemical, mechanical, biological, immunological, and xenobiotic stress throughout life. The disruption of endoplasmic reticulum (ER) homeostatic function, triggered by various factors, can induce ER stress. To overcome the elevated ER stress, an adaptive mechanism known as the unfolded protein response (UPR) is activated in cells. However, persistent ER stress and maladaptive UPR can lead to defects in proteostasis at the cellular level and are typical features of the lung aging. The aging lung and associated lung diseases exhibit signs of ER stress-related disruption in cellular homeostasis. Dysfunction resulting from ER stress and maladaptive UPR can compromise various cellular and molecular processes associated with aging. Hence, comprehending the mechanisms of ER stress and UPR components implicated in aging and associated lung diseases could enable to develop appropriate therapeutic strategies for the vulnerable population.

NIR-715 photodynamic therapy induces immunogenic cancer cell death by enhancing the endoplasmic reticulum stress response

Effectively interfering with endoplasmic reticulum (ER) function in tumor cells and simultaneously activating an anti-tumor immune microenvironment to attack the tumor cells are promising strategies for cancer treatment. However, precise ER-stress induction is still a huge challenge. In this study, we synthesized a near-infrared (NIR) probe, NIR-715, which induces tumor cell death and inhibits tumor growth without causing apparent side effects. NIR-715 triggers severe ER stress and immunogenic cell death (ICD) after visible light exposure. NIR-715 induced ICD-associated HMGB1 release in vitro and anti-tumor immune responses, including increased cytotoxic T lymphocyte (GZMB+ CD8+ T cell) infiltration and decreased numbers of exhausted T lymphocytes (PD-L1+ CD8+ T cell). These findings suggest that NIR-715 may be a novel agent for “cold” tumor photodynamic therapy (PDT).Schematic illustration of NIR-715 photodynamic therapy for visible light-triggered, endoplasmic reticulum-targeting antitumor therapy.

Canagliflozin differentially modulates carfilzomib-induced endoplasmic reticulum stress in multiple myeloma and endothelial cells.

Carfilzomib (CFZ), a second-generation proteasome inhibitor, is a key treatment for multiple myeloma (MM), but its use is associated with significant cardiovascular adverse events (CVAEs), including heart failure and hypertension. Endothelial dysfunction is believed to contribute to these CVAEs. Building on our previous findings that CFZ induces endothelial toxicity and that canagliflozin protects against CFZ-induced endothelial apoptosis, this study aimed to evaluate CFZ-induced endoplasmic reticulum (ER) stress and autophagy in endothelial and MM cells, as well as the impact of canagliflozin on these processes and its impact on the anticancer effects of CFZ in MM cells. Endothelial cells (HUVECs and EA.hy926) and multiple myeloma cells (RPMI8226) were treated with 0.5µM CFZ, either alone or in combination with canagliflozin (5-20µM), to assess the effects on ER stress and autophagy in both cell types. CFZ induced ER stress in endothelial and MM cells. In endothelial cells, canagliflozin mitigated CFZ-induced markers of ER stress, while unexpectedly upregulating CFZ-induced CHOP. Whereas, in MM cells, canagliflozin did not alter CFZ-induced ER stress, but instead further upregulated CFZ-induced ATF-4. In addition, CFZ induced autophagy in endothelial cells while inhibiting it in MM cells. Canagliflozin abrogated CFZ-induced autophagy in endothelial cells. In striking contrast to its effects in endothelial cells, canagliflozin enhanced the cytotoxic effects of CFZ in MM cells. Intriguingly, in an innovative co-culture system, canagliflozin enhanced CFZ-induced apoptosis in MM cells while protecting endothelial cells. These findings underscore the dual role of canagliflozin in reducing CFZ-induced endothelial toxicity, while enhancing its cytotoxic effect in MM.

Oxidative and Endoplasmic Reticulum Stress Mediate Testicular Injury in a Rat Model of Varicocele.

Evidence of endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR) have been increasingly reported in varicocele (VCL)-affected testes. However, the mechanisms by which oxidative stress (OS) and ER stress contribute to male infertility in VCL remain unclear. In this study, male Sprague-Dawley rats were divided into a control group, which underwent sham surgery, and a VCL group, in which VCL was surgically induced. Eight weeks postoperatively, the VCL group exhibited significant testicular damage and sperm abnormalities. Western blot analysis revealed upregulation of ER stress-related proteins and downstream apoptotic markers in the VCL group. For further investigation, we developed an in vitro oxidative stress model using GC-2 cells treated with 400 µM hydrogen peroxide (H2O2) for 12h. Cells were also treated with either an ER stress inducer or inhibitor. We found that treatment with H2O2 and the ER stress inducer significantly reduced GC-2 cell viability and increased reactive oxygen species (ROS) production, ER stress, and apoptosis. Conversely, treatment with the ER stress inhibitor 4-phenylbutyric acid (4-PBA) alleviated these effects. Our findings suggest a strong association between VCL-induced redox imbalance and ER stress-related injury driven by the UPR in this rat model. Furthermore, 4-PBA effectively reduced germ cell damage induced by ROS-mediated ER stress, offering potential therapeutic insights for treating VCL-related infertility.

In vitro anticancer effects in hepatocellular carcinoma (HCC) and protein interaction study of xanthoangelol.

Xanthoangelol (C25H28O4), a natural flavonoid derived from chalcones, has shown potential pharmacological activities. However, its primary interaction mechanism with proteins and cells is not well understood. In the present study, we focus on the anticancer effects of xanthoangelol against hepatocellular carcinoma (HCC) as well as its binding affinity with a plasma drug carrier protein, α2-macroglobulin. The anticancer effects of xanthoangelol on human HCC cell line HepG2 cells were assayed using MTT, LDH, qPCR, and caspase activity assays. Efficient binding of the xanthoangelol with α2-macroglobulin was established by experimental and molecular docking studies. It was found that xanthoangelol significantly mitigates cell viability through upregulating intrinsic (Bax/Bcl-2, caspase-9) and extrinsic (caspase-8) apoptotic pathways. Moreover, it was detected that xanthoangelol induces ER stress through the upregulation of CHOP in HepG2 cells. Fluorescence spectra show that xanthoangelol strongly interacts with α2-macroglobulin mediated by a static quenching mechanism and Trp1237 and Tyr1323 residues were exposed to the solvent with the addition of xanthoangelol. Meanwhile, both experimental and theoretical studies display that hydrophilic forces play a key role in the formation of xanthoangelol-α2-macroglobulin complex, leading to a slight conformational change in α2-macroglobulin. In conclusion, our findings suggest that xanthoangelol, which has a high binding affinity for a plasma carrier protein, may inhibit the viability of HCC by inducing apoptosis and ER stress.

Synthesis, Biological Evaluation, and Mechanistic Insights of Rubrolide Analogues as Antitumor Agents.

Marine natural products and their analogues have as of now been acknowledged as an important source of bioactive molecules for the treatment of cancer. Rubrolides, a unique group of γ-butenolides derived from marine microorganisms, have shown strong cytotoxic activity against various tumor cells. In this study, we synthesized and characterized 21 rubrolide analogues (including 16 new compounds) and investigated their antitumor activities in order to screen more active molecules and elucidate their mechanism of action. Primary MTT assay showed that compounds 1 and 4-9 all exhibited excellent antiproliferative activities. In particular, compound 7 showed broad-spectrum cytotoxic activity against six tumor cell lines, with IC50 values mostly ranging from 2.5 to 0.2 μM. Further mechanistic studies revealed that compound 7 could penetrate HCT116 and Hela cells, localize in the endoplasmic reticulum, and upregulate the PERK-eIF2α-CHOP pathway, inducing ER stress and increasing intracellular reactive oxygen species (ROS) levels to ultimately trigger apoptosis in tumor cells. Additionally, compound 7 was found to upregulate Cyclin B1 protein expression, causing cell cycle reticulum at the G2/M phase. In vivo studies further demonstrated that liposomal delivery of compound 7 exhibited a potent antitumor efficacy against Hela xenograft tumors. Based on these results, marine-derived rubrolide analogues show significant potential as novel lead compounds for antitumor drug development.

Urolithin A Protects Hepatocytes from Palmitic Acid-Induced ER Stress by Regulating Calcium Homeostasis in the MAM

An ellagitannin-derived metabolite, Urolithin A (UA), has emerged as a potential therapeutic agent for metabolic disorders due to its antioxidant, anti-inflammatory, and mitochondrial function-improving properties, but its efficacy in protecting against ER stress remains underexplored. The endoplasmic reticulum (ER) is a cellular organelle involved in protein folding, lipid synthesis, and calcium regulation. Perturbations in these functions can lead to ER stress, which contributes to the development and progression of metabolic disorders such as metabolic-associated fatty liver disease (MAFLD). In this study, we identified a novel target protein of UA and elucidated its mechanism for alleviating palmitic acid (PA)-induced ER stress. Cellular thermal shift assay (CETSA)-LC-MS/MS analysis revealed that UA binds directly to the sarcoplasmic/endoplasmic reticulum Ca2+-ATPase (SERCA), an important regulator of calcium homeostasis in mitochondria-associated ER membranes (MAMs). As an agonist of SERCA, UA attenuates abnormal calcium fluctuations and ER stress in PA-treated liver cells, thereby contributing to cell survival. The lack of UA activity in SERCA knockdown cells suggests that UA regulates cellular homeostasis through its interaction with SERCA. Collectively, our results demonstrate that UA protects against PA-induced ER stress and enhances cell survival by regulating calcium homeostasis in MAMs through SERCA. This study highlights the potential of UA as a therapeutic agent for metabolic disorders associated with ER stress.

The Effects of Microplastics on Musculoskeletal Disorder; A Narrative Review.

The physical health impacts of microplastics have received increasing attention in recent years. However, limited data impedes a full understanding of the internal exposure to microplastics, especially concerning the musculoskeletal system. The purpose of this review is to summarize the recent literature regarding the effects of microplastics on the musculoskeletal system. Microplastics have been shown to cause abnormal endochondral ossification and disrupt the normal function of pre-osteoblasts, osteocyte-like cells, and pre-osteoclasts through gene mutations, endoplasmic reticulum stress induction, and reduced autophagosome formation in bone growth areas. Although there are few reports on their effects on muscle, it has been noted that microplastics inhibit energy and lipid metabolism, decrease type I muscle fiber density, impair muscle angiogenesis, cause muscle atrophy, and increase lipid deposition. Only a few recent studies have shown that microplastics interfere with the normal function of bone growth-related cells and reduce muscle mass and quality. This review underscores the need for further research into other parts of the musculoskeletal system and studies using human tissues at the disease level.

EP152R-mediated endoplasmic reticulum stress contributes to African swine fever virus infection via the PERK-eIF2α pathway.

African swine fever virus (ASFV) is a large, icosahedral, double-stranded DNA virus in the Asfarviridae family and the causative agent of African swine fever (ASF). ASFV causes a hemorrhagic fever with high mortality rates in domestic and wild pigs. ASFV contains an open reading frame named EP152R, previous research has shown that EP152R is an essential gene for virus rescue in swine macrophages. However, the detailed functions of ASFV EP152R remain elusive. Herein, we demonstrate that EP152R, a membrane protein located in the endoplasmic reticulum (ER), induces ER stress and swelling, triggering the PERK/eIF2α pathway, and broadly inhibiting host protein synthesis invitro. Additionally, EP152R strongly promotes immune evasion, reduces cell proliferation, and alters cellular metabolism. These results suggest that ASFV EP152R plays a critical role in the intracellular environment, facilitating viral replication. Furthermore, virus-level experiments have shown that the knockdown of EP152R or PERK inhibitors efficiently affects viral replication by decreasing viral gene expression. In summary, these findings reveal a series of novel functions of ASFV EP152R and have important implications for understanding host-pathogen interactions.

The Role of Protein Disulfide Isomerase Inhibitors in Cancer Therapy.

Protein disulfide isomerase (PDI) is a member of the mercaptan isomerase family, primarily located in the endoplasmic reticulum (ER). At least 21 PDI family members have been identified. PDI plays a key role in protein folding, correcting misfolded proteins, and catalyzing disulfide bond formation, rearrangement, and breaking. It also acts as a molecular chaperone. Dysregulation of PDI activity is thus linked to diseases such as cancer, infections, immune disorders, thrombosis, neurodegenerative diseases, and metabolic disorders. In particular, elevated intracellular PDI levels can enhance cancer cell proliferation, metastasis, and invasion, making it a potential cancer marker. Cancer cells require extensive protein synthesis, with disulfide bond formation by PDI being a critical producer. Thus, cancer cells have higher PDI levels than normal cells. Targeting PDI can induce ER stress and activate the Unfolded Protein Response (UPR) pathway, leading to cancer cell apoptosis. This review discusses the structure and function of PDI, PDI inhibitors in cancer therapy, and the limitations of current inhibitors, proposing especially future directions for developing new PDI inhibitors.

A WFS1 variant disrupting acceptor splice site uncovers the impact of alternative splicing on beta cell apoptosis in a patient with Wolfram syndrome.

Wolfram syndrome 1 (WS1) is an inherited condition mainly manifesting in childhood-onset diabetes mellitus and progressive optic nerve atrophy. The causative gene, WFS1, encodes wolframin, a master regulator of several cellular responses, and the gene's mutations associate with clinical variability. Indeed, nonsense/frameshift variants correlate with more severe symptoms than missense/in-frame variants. As achieving a genotype-phenotype correlation is crucial for dealing with disease outcome, works investigating the impact of transcriptional and translational landscapes stemming from such mutations are needed. Therefore, we sought to elucidate the molecular determinants behind the pathophysiological alterations in a WS1 patient carrying compound heterozygous mutations in WFS1: c.316-1G>A, affecting the acceptor splice site (ASS) upstream of exon 4; and c.757A>T, introducing a premature termination codon (PTC) in exon 7. Bioinformatic analysis was carried out to infer the alternative splicing events occurring after disruption of ASS, followed by RNA-seq and PCR to validate the transcriptional landscape. Patient-derived induced pluripotent stem cells (iPSCs) were used as an in vitro model of WS1 and to investigate the WFS1 alternative splicing isoforms in pancreatic beta cells. CRISPR/Cas9 technology was employed to correct ASS mutation and generate a syngeneic control for the endoplasmic reticulum stress induction and immunotoxicity assays. We showed that patient-derived iPSCs retained the ability to differentiate into pancreatic beta cells. We demonstrated that the allele carrying the ASS mutation c.316-1G>A originates two PTC-containing alternative splicing transcripts (c.316del and c.316-460del), and two open reading frame-conserving mRNAs (c.271-513del and c.316-456del) leading to N-terminally truncated polypeptides. By retaining the C-terminal domain, these isoforms sustained the endoplasmic reticulum stress response in beta cells. Otherwise, PTC-carrying transcripts were regulated by the nonsense-mediated decay (NMD) in basal conditions. Exposure to cell stress inducers and proinflammatory cytokines affected expression levels of the NMD-related gene SMG7 (>twofold decrease; p<0.001) without eliciting a robust unfolded protein response in WFS1 beta cells. This resulted in a dramatic accumulation of the PTC-containing isoforms c.316del (>100-fold increase over basal; p<0.001) and c.316-460del (>20-fold increase over basal; p<0.001), predisposing affected beta cells to undergo apoptosis. Cas9-mediated recovery of ASS retrieved the canonical transcriptional landscape, rescuing the normal phenotype in patient-derived beta cells. This study represents a new model to study wolframin, highlighting how each single mutation of the WFS1 gene can determine dramatically different functional outcomes. Our data point to increased vulnerability of WFS1 beta cells to stress and inflammation and we postulate that this is triggered by escaping NMD and accumulation of mutated transcripts and truncated proteins. These findings pave the way for further studies on the molecular basis of genotype-phenotype relationship in WS1, to uncover the key determinants that might be targeted to ameliorate the clinical outcome of patients affected by this rare disease. The in silico predicted N-terminal domain structure file of WT wolframin was deposited in the ModelArchive, together with procedures, ramachandran plots, inter-residue distance deviation and IDDT scores, and Gromacs configuration files (doi/10.5452/ma-cg3qd). The deep-sequencing data as fastq files used to generate consensus sequences of AS isoforms of WFS1 are available in the SRA database (BioProject PRJNA1109747).

Thapsigargin: An Unlikely Drug in the Battle Against Viral Pandemics

Antiviral resistance is an increasingly categorized phenomenon, and with the high pandemic potential of emergent pathogens such as coronaviruses, the need for novel therapeutic agents is dire. Thapsigargin, a potent endoplasmic reticulum stress inducer found in the Mediterranean region, has shown significant promise in combatting replication of several viruses including coronaviruses by harnessing the host's innate immune system, even being more efficacious than some antivirals currently used. There is a major gap in knowledge regarding the exact mechanism of action that permits Thapsigargin to be an incredible endoplasmic reticulum stress inducer and an potent antiviral. We hypothesize that interferon 1 production and SERCA-inhibition caused by thapsigargin mediates the antiviral effects observed. This review outlines what is currently known about this novel compound, the gap in research, and details methodologies including gene expression analyses, protein quantification, and mammalian cell culturing, to address the gap and answer the hypothesis.

Unveiling genome plasticity as a mechanism of non-antifungal-induced antifungal resistance in Cryptococcus neoformans.

Cryptococcus neoformans, a critical priority pathogen designated by the World Health Organization, poses significant therapeutic challenges due to the limited availability of treatment options. The emergence of antifungal resistance, coupled with cross-resistance, further hampers treatment efficacy. Aneuploidy, known for its ability to induce diverse traits, including antifungal resistance, remains poorly understood in C. neoformans. We investigated the impact of tunicamycin, a well-established ER stress inducer, on aneuploidy formation in C. neoformans. Our findings show that both mild and severe ER stress induced by tunicamycin lead to the formation of aneuploid strains in C. neoformans. These aneuploid strains exhibit diverse karyotypes, with some conferring resistance or cross-resistance to antifungal drugs fluconazole and 5-flucytosine. Furthermore, these aneuploid strains display instability, spontaneously losing extra chromosomes in the absence of stress. Transcriptome analysis reveals the simultaneous upregulation of multiple drug resistance-associated genes in aneuploid strains. Our study reveals the genome plasticity of C. neoformans as a major mechanism contributing to non-antifungal-induced antifungal resistance.

Characterization of the Ubiquitin-Modified Proteome of Recombinant Chinese Hamster Ovary Cells in Response to Endoplasmic Reticulum Stress.

Chinese hamster ovary (CHO) cells remain the most widely used host cell line for biotherapeutics production. Despite their widespread use, understanding endoplasmic reticulum (ER) stress conditions in recombinant protein production remains limited, often creating bottlenecks preventing improved production titers and product quality. Ubiquitination not only targets substrates (e.g., misfolded proteins) for proteasome degradation but also has important regulatory control functions including cell cycle regulation, translation, apoptosis, autophagy, etc. and hence is likely to be central to understanding and controlling the productivity of recombinant biotherapeutics. This study aimed to uncover differentially expressed ubiquitinated proteins following artificial induction of ER-stress in recombinant CHO cells. CHO cells were treated with the stress inducer tunicamycin and the proteasome inhibitor MG132, followed by LC-MS/MS proteomic analysis. We identified >4000 ubiquitinated peptides from CHO-DP12 cells under ER stress conditions and proteasome inhibition. Moreover, data analysis showed altered abundance levels of >900 ubiquitinated proteins under the combination of ER stress and proteasome inhibition compared to untreated controls. Gene Ontology (GO) analysis of these ubiquitinated proteins resulted in a significant enrichment of key pathways involving the proteasome, protein processing in the ER, N-glycan biosynthesis, and ubiquitin-mediated proteolysis. ER stress response proteins such as GRP78, HSP90B1, ATF6, HERPUD1, and PDIA4 were found to be highly ubiquitinated and exhibited a significant increase in abundance following induction of ER-stress conditions. This study broadens our comprehension of the roles played by protein ubiquitination in CHO cell stress responses, potentially revealing targets for tailored cell line engineering aimed at enhancing stress tolerance and production efficiency.

Glycosylation of Ceramide Synthase 6 is Required for Its Activity

Sphingolipids play key roles in membrane structure and cellular signaling. Ceramide Synthase (CerS)-generated ceramide is implicated in cellular stress responses and induction of apoptosis. Ceramide and other sphingolipids are linked to the induction of ER stress response pathways. However, the mechanisms by which ceramide modulates ER stress signaling are not well understood. Here, we show that the ER stress inducer Brefeldin A (BFA) causes increased glycosylation of CerS6, and that treatment with BFA causes increased endogenous ceramide accumulation. To our surprise, we found that CerS6 activity was not affected by BFA-induced glycosylation. Instead, our data show that basal glycosylation of CerS6 at Asn18 is required for CerS6 activity. We used a robust HCT116 CRISPR-Cas9 CerS6 KO with reintroduction of either wild type CerS6 or a mutant CerS6 with a point mutation at asparagine-18 to an alanine (N18A) which abrogated glycosylation at that residue. Our data show that cells stably expressing the N18A mutant CerS6 had significantly lower activity in vitro and in situ as compared to WT CerS6 expressing cells. Further, the defective CerS6 with N18A mutation also had defects in GSK3β, AKT, JNK, and STAT3 signaling. Despite being required for CerS6 activity, Asn18 glycosylation did not influence ER stress response pathways. Overall, our study provides vital insight into the regulation of CerS6 activity by post-translational modification at Asn18 and identifies glycosylation of CerS6 to be important for ceramide generation and downstream cellular signaling pathways.

Zelenirstat Inhibits N-Myristoyltransferases to Disrupt Src Family Kinase Signalling and Oxidative Phosphorylation Killing Acute Myeloid Leukemia Cells.

Acute myeloid leukemia (AML) is a hematological malignancy with limited treatment options and a high likelihood of recurrence after chemotherapy. We studied N-myristoylation, the myristate modification of proteins linked to survival signaling and metabolism, as a potential therapeutic target for AML. N-myristoylation is catalyzed by two N-myristoyltransferases (NMTs), NMT1 and NMT2, with varying expressions in AML cell lines and patient samples. We identified NMT2 expression as a marker for AML patient survival, and low NMT2 expression was associated with poor outcomes. We used the first-in-class pan-NMT inhibitor, zelenirstat, to investigate the role of N-myristoylation in AML. Zelenirstat effectively inhibits myristoylation in AML cell lines and patient samples, leading to degradation of Src family kinases (SFKs), induction of endoplasmic reticulum (ER) stress, apoptosis, and cell death. Zelenirstat was well tolerated in vivo and reduced the leukemic burden in an ectopic AML cell line and in multiple orthotopic AML patient-derived xenograft models. The leukemia stem cell (LSC)-enriched fractions of the hierarchical OCI-AML22 model were highly sensitive to myristoylation inhibition. Zelenirstat also impairs mitochondrial complex I and oxidative phosphorylation, which are critical for LSC survival. These findings suggest that targeting N-myristoylation with zelenirstat represents a novel therapeutic approach for AML, with promise in patients with currently poor outcomes.

Small hepatitis B virus surface antigen (SHBs) induces dyslipidemia by suppressing apolipoprotein-AII expression through ER stress-mediated modulation of HNF4α and C/EBPγ.

Persistent infection with hepatitis B virus (HBV) often leads to disruptions in lipid metabolism. Apolipoprotein AII (apoAII) plays a crucial role in lipid metabolism and is implicated in various metabolic disorders. However, whether HBV could regulate apoAII and contribute to HBV-related dyslipidemia and the underlying mechanism remain unclear. This study revealed significant reductions in apoAII expression in HBV-expressing cell lines, the serum, and liver tissues of HBV-transgenic mice. The impact of HBV on apoAII is related to small hepatitis B virus surface antigen (SHBs). Overexpression of SHBs decreased apoAII levels in SHBs-expressing hepatoma cells, transgenic mice, and the serum of HBV-infected patients, whereas suppression of SHBs increased apoAII expression. Mechanistic investigations demonstrated that SHBs repressed the apoAII promoter activity through a HNF4α- and C/EBPγ-dependent manner; SHBs simultaneously upregulated C/EBPγ and downregulated HNF4α by inhibiting the PI3K/AKT signaling pathway through activating endoplasmic reticulum (ER) stress. Serum lipid profile assessments revealed notable decreases in high-density lipoprotein cholesterol (HDL-C), total cholesterol (TC), and triglycerides (TG) in SHBs-transgenic mice compared to control mice. However, concurrent overexpression of apoAII in these mice effectively counteracted these reductions in lipid levels. In HBV patients, SHBs levels were negatively correlated with serum levels of HDL-C, LDL-C, TC, and TG, whereas apoAII levels positively correlated with lipid content. This study underscores that SHBs contributes to dyslipidemia by suppressing the PI3K/AKT pathway via inducing ER stress, leading to altered expression of HNF4α and C/EBPγ, and subsequently reducing apoAII expression.IMPORTANCEThe significance of this study lies in its comprehensive examination of how the hepatitis B virus (HBV), specifically through its small hepatitis B virus surface antigen (SHBs), impacts lipid metabolism-a key aspect often disrupted by chronic HBV infection. By elucidating the role of SHBs in regulating apolipoprotein AII (apoAII), a critical player in lipid processes and associated metabolic disorders, this research provides insights into the molecular pathways contributing to HBV-related dyslipidemia. Discovering that SHBs downregulates apoAII through mechanisms involving the repression of the apoAII promoter via HNF4α and C/EBPγ, and the modulation of the PI3K/AKT signaling pathway via endoplasmic reticulum (ER) stress, adds critical knowledge to HBV pathogenesis. The research also shows an inverse correlation between SHBs expression and key lipid markers in HBV-infected individuals, suggesting that apoAII overexpression could counteract the lipid-altering effects of SHBs, offering new avenues for understanding and managing the metabolic implications of HBV infection.

Vitamin K2 and the Vitamin-K-Cycle-Enzyme VKORC1L1 improve endothelial regeneration by inhibiting endothelial-mesenchymal transition, inflammatory cell activation and ferroptosis

Abstract Introduction Vascular inflammation is a crucial contributor to atherosclerosis, in which oxidative stress, endoplasmic reticulum (ER) stress and transition of endothelial to mesenchymal cells (EndMT) are of critical importance. Vitamin-K-antagonists (VKA) promote vascular dysfunction, while dietary vitamin K intake was proposed to reduce incidence of coronary artery disease, but the mechanisms remain unresolved. Key enzyme for regeneration of vitamin K is the Vitamin K epoxide reductase complex subunit 1 (VKORC1), which also represents the pharmacological target of VKA. VKOR-like1 (VKORC1L1) is an isoenzyme of VKORC1, which resides at the ER-membrane, exerts antioxidative properties and is involved in vitamin K maintenance. Aim of this study was to investigate the role of Vitamin K2 and the VKOR-enzymes in endothelial cell inflammation. Methods and Results In human coronary artery endothelial cells (HCAEC), Vitamin K2 improves endothelial regeneration by increasing proliferation and cell viability and by reducing both apoptosis and ferroptosis (relative viability: RSL3 0.11 ± 0.01 vs 0.32 ± 0.05 RSL3+K2-1µM vs. 0.44 ± 0.11 RSL3+K2-10µM, p&amp;lt;0.001). Moreover, Vitamin K2-supplementation inhibits EndMT by reducing expression of mesenchymal markers (Calponin, SM22; Fig.1). In silico analyses of the proteome of human coronary atherosclerosis revealed increased expression of VKORC1L1, but not of VKORC1. In vitro induction of oxidative stress and ER-Stress promoted time- and dose-dependent enhanced expression of VKORC1L1, without altering VKORC1 expression (H2O2: 2.3-fold vs. 0.8-fold after 40 minutes, p=0.04; tunicamycin: 1 μg/ml: 1.43-fold vs. 0.8-fold, p&amp;lt;0.01). Likewise, siRNA-mediated VKORC1L1 knockdown induces an inflammatory gene signature with upregulation of NFkB-signalling and endoplasmic reticulum stress (NF-κB: 1.95-fold ± 0.13, GRP78: 1.32-fold ± 0.04 vs. control, p&amp;lt;0.01). Furthermore, VKORC1L1 downregulation increased formation of reactive oxygen species (ROS) and reduced proliferation (EdU signal: 0.56 ± 0.02 vs. control, p &amp;lt; 0.01). In contrast, VKORC1-knockdown does not impair endothelial regeneration and function (Fig.2). Vitamin K2 reduces vascular inflammation and endoplasmic reticulum stress in the case of VKORC1-, but not VKORC1L1-knockdown. In addition, inducing EndMT resulted in a reduction of VKORC1L1, but not of VKORC1, while VKORC1L1-knockdown aggravates EndMT. Conclusion VKORC1 and its isoenzyme VKORC1L1 exhibit divergent effects on endothelial cell inflammation. Further studies are warranted to shed light on the regulation of the enzymes to improve our understanding regarding vascular side effects of VKA treatment and beneficial effects of vitamin K supplementation.

Nimbolide Induces Cell Apoptosis via Mediating ER Stress-Regulated Apoptotic Signaling in Human Oral Squamous Cell Carcinoma.

Human oral squamous cell carcinoma (OSCC) poses a significant health challenge in Asia, with current therapeutic strategies failing to improve the survival rates for OSCC patients sufficiently. To elucidate the effects of Nimbolide on OSCC cell proliferation and apoptosis, we performed a series of experiments, including cell proliferation assays, annexin V/PI assays, and cell cycle analysis. We further investigated nimbolide's role in modulating endoplasmic reticulum (ER) stress, reactive oxygen species (ROS) production, and mitochondrial dysfunction using flow cytometry. Additionally, Western blotting was used to detect apoptosis-related protein expression. Our findings reveal that nimbolide exerts its anti-proliferative effects on OSCC cells by inducing apoptosis. The nimbolide increased intracellular ROS levels and acceleration of cellular calcium accumulation, respectively promoting endoplasmic reticulum stress and cancer cell apoptosis. Furthermore, nimbolide activates the caspase cascade by altering the mitochondrial membrane potential and apoptotic protein expression, thereby inhibiting the viability of tumor cells. Our data show that Nimbolide suppresses tumor growth through the induction of ROS production, ER stress, and mitochondrial dysfunction, resulting in apoptosis in OSCC cells. Overall, our study highlights nimbolide as a potential natural compound for OSCC therapy.

Endoplasmic Reticulum Stress in Bronchopulmonary Dysplasia: Contributor or Consequence?

Bronchopulmonary dysplasia (BPD) is the most common complication of prematurity. Oxidative stress (OS) and inflammation are the major contributors to BPD. Despite aggressive treatments, BPD prevalence remains unchanged, which underscores the urgent need to explore more potential therapies. The endoplasmic reticulum (ER) plays crucial roles in surfactant and protein synthesis, assisting mitochondrial function, and maintaining metabolic homeostasis. Under OS, disturbed metabolism and protein folding transform the ER structure to refold proteins and help degrade non-essential proteins to resume cell homeostasis. When OS becomes excessive, the endogenous chaperone will leave the three ER stress sensors to allow subsequent changes, including cell death and senescence, impairing the growth potential of organs. The contributing role of ER stress in BPD is confirmed by reproducing the BPD phenotype in rat pups by ER stress inducers. Although chemical chaperones attenuate BPD, ER stress is still associated with cellular senescence. N-acetyl-lysyltyrosylcysteine amide (KYC) is a myeloperoxidase inhibitor that attenuates ER stress and senescence as a systems pharmacology agent. In this review, we describe the role of ER stress in BPD and discuss the therapeutic potentials of chemical chaperones and KYC, highlighting their promising role in future therapeutic interventions.