Conditions Of Cellular Stress Research Articles

Temperature influence on a linear array of coupled Sal'nikov oscillators: the muscle fiber scenario

Muscles are thermogenic organs for adult mammals and birds but can also be thermosensitive. In this respect, recent research has shown the excitability of cardiac muscle tissue when irradiated with infrared laser light. Likewise, intrinsic muscle function should give place to local thermal gradients, either because of Ca2+-ATPase thermoregulatory mechanisms or, specifically in the skeletal muscle, after intense exercise. Alongside internal thermal gradients, muscle fatigue characterizes by stressful cellular conditions. Similar to stress-like conditions has been documented in myocytes from rats, the emergence of oscillations of important biochemical species [1]. We show how a thermal gradient or a thermal pulse influences the dynamics of biochemical oscillations in a simplified biochemical model of muscle fiber and discuss the consequences in a living muscle. For this purpose, each simplified sarcomere behavior is governed according to a modified Sal'nikov model, as proposed in a previous paper [2].

Influence of oxidative stress on drug-DNA binding: Microcalorimetric and mechanistic insights with anticancer drugs

Oxidative stress causing the chronic inflammation, mediates to deadly diseases such as cancer, diabetes, cardiovascular disease. Maintaining the ratio of reactive oxygen species generation to scavenging, thus the oxidative stress is important to have healthy cells. Oxidative stress not only causes the deadly diseases but also affect the conformation of biomolecules such as protein and DNA, even at lower concentrations. Binding of drugs to DNA is susceptible to these DNA structural or conformational changes. Here, we have reported the influence of oxidative stress on binding of anti-cancer drugsdoxorubicin hydrochloride (DOX),mitoxantrone dihydrochloride (MTX), capecitabine (CPT), and ethidium bromide (EtBr) with DNA. Quantitative insights of association of these anticancer drugs with DNA in presence of different environments, in vitro (pH – 7.4), cellular (100:1 ratio of GSH to GSSG) and oxidative stress condition (1:1 ratio of GSH to GSSG) have been compared. Microcalorimetry has been used for determination of thermodynamic parameters, binding constant Kb, molar enthalpy of interaction (ΔHm), molar entropy of interaction (ΔSm) & stoichiometry (n). Fluorescence spectroscopy has been used to illustrate the mode of drug-DNA binding and also to complement the ITC results. Insignificant changes in thermodynamic parameters for drug-DNA binding in in vitro and in presence of cellular condition were observed whereas significant decrease in binding affinity and stoichiometry was observed in the presence of oxidative stress conditions. Quantitative understanding of drug-DNA binding and influence of oxidative stress in terms of thermodynamic signature assists in the structure-based drug designing and development.

Intestinal Apc-inactivation induces HSP25 dependency.

The majority of colorectal cancers (CRCs) present with early mutations in tumor suppressor gene APC. APC mutations result in oncogenic activation of the Wnt pathway, which is associated with hyperproliferation, cytoskeletal remodeling, and a global increase in mRNA translation. To compensate for the increased biosynthetic demand, cancer cells critically depend on protein chaperones to maintain proteostasis, although their function in CRC remains largely unexplored. In order to investigate the role of molecular chaperones in driving CRC initiation, we captured the transcriptomic profiles of murine wild type and Apc-mutant organoids during active transformation. We discovered a strong transcriptional upregulation of Hspb1, which encodes small heat shock protein 25 (HSP25). We reveal an indispensable role for HSP25 in facilitating Apc-driven transformation, using both invitro organoid cultures and mouse models, and demonstrate that chemical inhibition of HSP25 using brivudine reduces the development of premalignant adenomas. These findings uncover a hitherto unknown vulnerability in intestinal transformation that could be exploited for the development of chemopreventive strategies in high-risk individuals.

Dual role of an essential HtrA2/Omi protease in the human malaria parasite: Maintenance of mitochondrial homeostasis and induction of apoptosis-like cell death under cellular stress

Members of the HtrA family of serine proteases are known to play roles in mitochondrial homeostasis as well as in programmed cell death. Mitochondrial homeostasis and metabolism are crucial for the survival and propagation of the malaria parasite within the host. Here we have functionally characterized a Plasmodium falciparum HtrA2 (PfHtrA2) protein, which harbours trypsin-like protease activity that can be inhibited by its specific inhibitor, ucf-101. A transgenic parasite line was generated, using the HA-glmS C-terminal tagging approach, for localization as well as for inducible knock-down of PfHtrA2. The PfHtrA2 was localized in the parasite mitochondrion during the asexual life cycle. Genetic ablation of PfHtrA2 caused significant parasite growth inhibition, decreased replication of mtDNA, increased mitochondrial ROS production, caused mitochondrial fission/fragmentation, and hindered parasite development. However, the ucf-101 treatment did not affect the parasite growth, suggesting the non-protease/chaperone role of PfHtrA2 in the parasite. Under cellular stress conditions, inhibition of PfHtrA2 by ucf-101 reduced activation of the caspase-like protease as well as parasite cell death, suggesting the involvement of protease activity of PfHtrA2 in apoptosis-like cell death in the parasite. Under these cellular stress conditions, the PfHtrA2 gets processed but remains localized in the mitochondrion, suggesting that it acts within the mitochondrion by cleaving intra-mitochondrial substrate(s). This was further supported by trans-expression of PfHtrA2 protease domain in the parasite cytosol, which was unable to induce any cell death in the parasite. Overall, we show the specific roles of PfHtrA2 in maintaining mitochondrial homeostasis as well as in regulating stress-induced cell death.

Pericytes take up and degrade α-synuclein but succumb to apoptosis under cellular stress

Parkinson’s disease (PD) is characterised by the progressive loss of midbrain dopaminergic neurons and the presence of aggregated α-synuclein (α-syn). Pericytes and microglia, two non-neuronal cells contain α-syn in the human brain, however, their role in disease processes is poorly understood. Pericytes, found surrounding the capillaries in the brain are important for maintaining the blood–brain barrier, controlling blood flow and mediating inflammation. In this study, primary human brain pericytes and microglia were exposed to two different α-synuclein aggregates. Inflammatory responses were assessed using immunocytochemistry, cytometric bead arrays and proteome profiler cytokine array kits. Fixed flow cytometry was used to investigate the uptake and subsequent degradation of α-syn in pericytes. We found that the two α-syn aggregates are devoid of inflammatory and cytotoxic actions on human brain derived pericytes and microglia. Although α-syn did not induce an inflammatory response, pericytes efficiently take up and degrade α-syn through the lysosomal pathway but not the ubiquitin–proteasome system. Furthermore, when pericytes were exposed the ubiquitin proteasome inhibitor—MG132 and α-syn aggregates, there was profound cytotoxicity through the production of reactive oxygen species resulting in apoptosis. These results suggest that the observed accumulation of α-syn in pericytes in human PD brains likely plays a role in PD pathogenesis, perhaps by causing cerebrovascular instability, under conditions of cellular stress.

TRAF2 regulates the protein stability of HIPK2

A nuclear serine/threonine kinase homeodomain-interacting protein kinase 2 (HIPK2) is a critical regulator of development and DNA damage response. HIPK2 can induce apoptosis under cellular stress conditions and thus its protein level is maintained low by constant proteasomal degradation. In the present study, we present evidence that TNF receptor-associated factor 2 (TRAF2) regulates the protein stability of HIPK2. Overexpression of TRAF2 decreased while its knockdown increased the HIPK2 protein level. The TRAF2-mediated decrease in HIPK2 protein expression was blocked by proteasomal inhibitor. In addition, TRAF2 decreased the protein half-life of HIPK2. We found that HIPK2 and TRAF2 co-immunoprecipitated. Interestingly, the co-immunoprecipitation was reduced while HIPK2 protein level increased following TNFα treatment, suggesting TNFα induced dissociation of TRAF2 from HIPK2 to accumulate HIPK2. Inhibition of HIPK2 partially suppressed TNFα-induced cell death, indicating that the accumulated HIPK2 may contribute to the TNFα-induced cell death. Our results suggest that TRAF2 can regulate proapoptotic function of HIPK2 by promoting proteasomal degradation.

Antioxidant Effect of Coenzyme Q10 in the Prevention of Oxidative Stress in Arsenic-Treated CHO-K1 Cells and Possible Participation of Zinc as a Pro-Oxidant Agent.

Oxidative stress is an imbalance between levels of reactive oxygen species (ROS) and antioxidant enzymes. Compounds with antioxidant properties, such as coenzyme Q10 (CoQ10), can reduce cellular imbalance caused by an increase in ROS. CoQ10 participates in modulating redox homeostasis due to its antioxidant activity and its preserving mitochondrial functions. Thus, the present study demonstrated the protective effects of CoQ10 against oxidative stress and cytotoxicity induced by arsenic (As). Antioxidant capacity, formation of hydroperoxides, generation of ROS, and the effect on cellular viability of CoQ10, were investigated to determine the protective effect of CoQ10 against As and pro-oxidant compounds, such as zinc. Cell viability assays showed that CoQ10 is cytoprotective under cellular stress conditions, with potent antioxidant activity, regardless of the concentration tested. Zn, when used at higher concentrations, can increase ROS and show a pro-oxidant effect causing cell damage. The cytotoxic effect observed for As, Zn, or the combination of both could be prevented by CoQ10, without any decrease in its activity at cellular levels when combined with Zn.

Bioluminescent test systems based on firefly luciferase for studying stress effects on living cells.

The bioluminescent luciferin-luciferase reaction is based on the oxidation of D-luciferin by oxygen in the presence of ATP and magnesium ions, catalyzed by firefly luciferase. The possibilities of using this reaction to study the influence of external effectors of a physical and chemical nature (temperature exposure, additions of drugs, membrane-active compounds, etc.) on living cells (prokaryotes and eukaryotes) are considered. Examples of the use of test systems based on living cells producing thermostable firefly luciferase for monitoring cellular homeostasis are given. The study of the kinetics of changes in the concentration of ATP and luciferase inside and outside cells made it possible to determine in dynamics the metabolic activity, cytotoxicity, and survival of cells under conditions of cellular stress, to study the processes of ATP synthesis/hydrolysis, and to evaluate the effectiveness of lytic agents in changing the permeability of the cell membrane.

Profiling the Site of Protein CoAlation and Coenzyme A Stabilization Interactions.

Coenzyme A (CoA) is a key cellular metabolite known for its diverse functions in metabolism and regulation of gene expression. CoA was recently shown to play an important antioxidant role under various cellular stress conditions by forming a disulfide bond with proteins, termed CoAlation. Using anti-CoA antibodies and liquid chromatography tandem mass spectrometry (LC-MS/MS) methodologies, CoAlated proteins were identified from various organisms/tissues/cell-lines under stress conditions. In this study, we integrated currently known CoAlated proteins into mammalian and bacterial datasets (CoAlomes), resulting in a total of 2093 CoAlated proteins (2862 CoAlation sites). Functional classification of these proteins showed that CoAlation is widespread among proteins involved in cellular metabolism, stress response and protein synthesis. Using 35 published CoAlated protein structures, we studied the stabilization interactions of each CoA segment (adenosine diphosphate (ADP) moiety and pantetheine tail) within the microenvironment of the modified cysteines. Alternating polar-non-polar residues, positively charged residues and hydrophobic interactions mainly stabilize the pantetheine tail, phosphate groups and the ADP moiety, respectively. A flexible nature of CoA is observed in examined structures, allowing it to adapt its conformation through interactions with residues surrounding the CoAlation site. Based on these findings, we propose three modes of CoA binding to proteins. Overall, this study summarizes currently available knowledge on CoAlated proteins, their functional distribution and CoA–protein stabilization interactions.

S107: SOD2 PROMOTES ACUTE LEUKEMIA ADAPTATION TO AMINO ACID STARVATION THROUGH THE N-DEGRON PATHWAY

Background: The ability of cells to tolerate amino acid starvation is fundamental for survival under cellular stress conditions. Some cancer cells are vulnerable to asparagine depletion, which is exploited therapeutically using asparaginase. However, the mechanisms of adaptation to amino acid starvation in leukemia cells remain incompletely understood. Aims: We recently performed a genome-wide CRISPR/Cas9 loss-of-function screen in the resistant T-ALL cell line CCRF-CEM to identify molecular pathways that promote asparaginase resistance. We found that Wnt-dependent stabilization of proteins (Wnt/STOP) induces a profound therapeutic vulnerability to asparaginase in acute leukemias and colorectal cancers (Hinze et. al., 2019; Hinze et al., 2020). Another unrelated gene on the top of the screen included SOD2, a mitochondrial superoxide dismutase. Intriguingly, to date, SOD2 activity has not been linked to a cellular amino acid starvation response, whose biologic basis we thus sought to further investigate. Methods: To evaluate the significance of SOD2 in mediating an asparaginase response, we employed genetic epistasis experiments as well as phenotypic assays including short hairpin RNA (shRNA) mediated knockdowns, quantitative PCRs, Western blots, amino acid starvation, and viability assays. Results: Knockdown of SOD2 (shSOD2) resulted in a profound sensitivity to asparaginase in several T-ALL and B-ALL cell lines (p<0.0001), and an increase in apoptosis, as assessed by caspase 3/7 activity (p<0.001). The sensitization was rescued by either overexpressing SOD2 cDNA (p<0.0001), or by adding the functional SOD2 mimetic MnTBAP (p<0.01). Of note, shSOD2 mediated sensitization was selective to asparaginase, as it could not be observed for other commonly used chemotherapeutic agents including vincristine, doxorubicin, dexamethasone, and 6-mercaptopurine (p=ns). Due to the selectivity to asparagine depletion, we then investigated whether SOD2 inhibition mediates a broader amino acid starvation response. Indeed, culturing SOD2-inhibited T-ALL cells in the absence of essential amino acids (EAA) or non-EAA, induced a significant decrease in cell viability (p<0.05). Sensitization appeared to be specific to the SOD2 isoform, and distinct from known SOD2-associated pathways including reactive oxygen species, cell cycle changes, alterations of mTOR signaling, or glutamine anaplerosis. To better understand the molecular underpinnings of SOD2 in regulating an amino acid starvation response, we leveraged the Bioplex Interactome database (Huttlin et al., 2020), and identified UBR2, an E3 ubiquitin ligase in the N-degron pathway, as a unique binding partner of SOD2. Ubiquitin E3 ligases target their substrates for ubiquitination, leading to proteasome-mediated degradation (Yang et al., 2010). Indeed, SOD2 and UBR2 were co-immunoprecipitated, suggesting the formation of a complex that can drive proteasome-dependent protein catabolism. In line, inhibition of SOD2 significantly decreased ubiquitin levels, suggesting that SOD2 positively regulates catabolic protein degradation through the N-degron pathway to promote cancer cell fitness in amino acid starved conditions. Summary/Conclusion: The interaction of SOD2 and the N-degron pathway represents a previously unknown molecular adaptation of cancer cells in response to amino acid starvation. These results serve as a strong proponent for an in-depth characterization of the N-degron pathway in mediating leukemia cell fitness upon amino acid starvation and thus provide a basis for therapeutic intervention in refractory leukemias.

A novel maladaptive unfolded protein response as a mechanism for small bowel resection-induced liver injury.

The unfolded protein response (UPR) is a complex adaptive signaling pathway activated by the accumulation of misfolded proteins in the endoplasmic reticulum (ER). ER stress (ERS) triggers a cascade of responses that converge upon C/EBP homologous protein (CHOP) to drive inflammation and apoptosis. Herein, we sought to determine whether liver injury and fibrosis after small bowel resection (SBR) were mediated by a maladaptive hepatic ERS/UPR. C57BL/6 mice underwent 50% proximal SBR or sham operation. Markers of liver injury and UPR/ERS pathways were analyzed. These were compared with experimental groups including dietary fat manipulation, tauroursodeoxycholic acid (TUDCA) treatment, distal SBR, and global CHOP knockout (KO). At 10 wk, proximal SBR had elevated alanine aminotransferase/aspartate aminotransferase (ALT/AST) (P < 0.005) and greater hepatic tumor necrosis factor-α (TNFα) (P = 0.001) and collagen type 1 α1 (COL1A1) (P = 0.02) than shams. SBR livers had increased CHOP and p-eIF2α, but were absent in activating transcription factor 4 (ATF4) protein expression. Low-fat diet (LFD), TUDCA, and distal SBR groups had decreased liver enzymes, inflammation, and fibrosis (P < 0.05). Importantly, they demonstrated reversal of hepatic UPR with diminished CHOP and robust ATF4 signal. CHOP KO-SBR had decreased ALT but not AST compared with wild-type (WT)-SBR (P = 0.01, P = 0.12). There were no differences in TNFα and COL1A1 (P = 0.09, P = 0.50). SBR-induced liver injury, fibrosis is associated with a novel hepatic UPR/ERS response characterized by increased CHOP and decreased ATF4. LFD, TUDCA, and ileocecal resection rescued the hepatic phenotype and reversed the UPR pattern. Global CHOP KO only partially attenuated liver injury. This underscores the significance of disruptions to the gut/liver axis after SBR and potentiates targets to mitigate the progression of intestinal failure-associated liver disease.NEW & NOTEWORTHY The unfolded protein response (UPR) is a complex signaling cascade that converges upon C/EBP-homologous protein (CHOP). Under conditions of chronic cellular stress, the UPR shifts from homeostatic to proapoptotic leading to inflammation and cell death. Here, we provide evidence that small bowel resection-induced liver injury and fibrosis are mediated by a maladaptive hepatic UPR. Low-fat diet, TUDCA treatment, and ileocecal resection rescued the hepatic phenotype and reversed the UPR pattern.

CRISPR/Cas9-mediated knockout of Lcn2 in human breast cancer cell line MDA-MB-231 ameliorates erastin-mediated ferroptosis and increases cisplatin vulnerability

AimsLipocalin 2 (Lcn2) is an antioxidant-related protein upregulated in various cellular stress conditions, especially cancer. In this study, we abrogated Lcn2 expression in MDA-MB-231 breast cancer cells using the CRISPR/Cas9 technology and evaluated its effect on cellular proliferation, migration, and ferroptotic cell death. Main methodsValidated human Lcn2 CRISPR/Cas9 knockout (KO) and homology-directed repair (HDR) plasmids were co-transfected into MDA-MB-231 breast cancer cells. Lcn2 gene knockout was confirmed at the transcriptional and protein levels using reverse transcription (RT)-PCR and enzyme-linked immunosorbent assay (ELISA). Cell proliferation was measured using Cell Counting Kit-8 (CCK-8) and colony formation assays. Cytotoxicity assay was performed in the presence or absence of erastin, cisplatin (CDDP), and ferrostatin-1 using the CCK-8 method. Ferroptosis level was measured using the malondialdehyde assay lipid peroxidation kit. The migration capacity of the cells was also evaluated using the scratch assay. Key findingsTargeting Lcn2 using CRISPR/Cas9 reduced cellular proliferation and migration capability, and elevated the vulnerability of MDA-MB-231 cells to cisplatin. Furthermore, Lcn2 expression loss effectively promoted erastin-mediated ferroptosis in MDA-MB-231 cells. SignificanceInhibition of Lcn2 is a potentially useful strategy for sensitizing MDA-MB-231 tumor cells to ferroptotic cell death.

Determining the role of ATF4 in the regulation of mitochondrial remodeling during myotube differentiation and contractile activity

ATF4 is a transcriptional regulator that is selectively induced in response to cellular stress conditions, such as exercise, through the activation of the integrated stress response (ISR). Specifically, in the context of mitochondrial‐specific stress, ATF4 is induced as a key component of the mitochondrial unfolded protein response (UPRmt) and is suggested to upregulate various organellar chaperones and proteases that both preserve, and promote, mitochondrial function. In response to the stress brought about by contractile activity, ATF4 has been implicated in regulating skeletal muscle health by mediating the various signaling events associated with mitochondrial quality control (MQC), including i) mitochondrial biogenesis (expansion), ii) the mitophagy‐lysosomal clearance of damaged and thus potentially harmful organelles, or iii) by activating the mitochondrial unfolded protein response (UPRmt) as an intermediate response to acute cellular stress. However, it remains to be determined whether ATF4 is necessary for mitochondrial adaptations in skeletal muscle. Therefore, our aim was to determine whether ATF4 is required for the maintenance of mitochondrial function and adaptation following an acute 3h bout of contractile activity (ACA), or after repeated bouts (4 days; CCA) in C2C12 myotubes in which ATF4 was either overexpressed (OE) or knocked down (KD) via lentiviral transduction of plasmids containing the ATF4 open reading frame, or siRNA, respectively. Knockdown of ATF4 promoted elongated myotube formation following 5 days of differentiation, whereas ATF4 OE contributed to the opposite effect in which shorter myotubes were observed relative to the control condition. Induction of PGC‐1α mRNA following ACA in ATF4 KD myotubes was attenuated, suggesting diminished drive for mitochondrial biogenesis in the absence of ATF4. The mRNA expression of ATF5, a downstream target of ATF4, was induced both in response to ACA as well as with ATF4 OE, and was reduced in the absence of ATF4. Mitophagy flux, measured by mitochondrial‐localized LC3‐II, was upregulated in ATF4 OE and KD myotubes basally, and was augmented in both control and ATF4 OE cells following ACA, but not in the absence of ATF4. Furthermore, ATF4 OE revealed decrements in mitochondrial content indicated by 20%, and 40%, reductions in VDAC and COX I protein expression relative to control, while 1.2‐1.5‐fold increases in these markers were observed when ATF4 was knocked down. However, following CCA, COX I and VDAC protein content were increased 3‐4‐fold in ATF4 OE cells, while there was no observable increase in mitochondrial content in ATF4 KD myotubes. Together, these data highlight a potential role of ATF4 in regulating basal mitochondrial content and further suggest that ATF4 may be required for contractile activity‐induced increases in mitochondrial content.

Oxidation of the Mycobacterium tuberculosis key virulence factor protein tyrosine phosphatase A (MptpA) reduces its phosphatase activity.

The Mycobacterium tuberculosis tyrosine-specific phosphatase MptpA and its cognate kinase PtkA are prospective targets for anti-tuberculosis drugs as they interact with the host defense response within the macrophages. Although both are structurally well-characterized, the functional mechanism regulating their activity remains poorly understood. Here, we investigate the effect of post-translational oxidation in regulating the function of MptpA. Treatment of MptpA with H2 O2 /NaHCO3 , mimicking cellular oxidative stress conditions, leads to oxidation of the catalytic cysteine (C11) and to a conformational rearrangement of the phosphorylation loop (D-loop) by repositioning the conserved tyrosine 128 (Y128) and generating a temporarily inactive preclosed state of the phosphatase. Thus, the catalytic cysteine in the P-loop acts as a redox switch and regulates the phosphatase activity of MptpA.

Histone H3K4me3 breadth in hypoxia reveals endometrial core functions and stress adaptation linked to endometriosis

SummaryTrimethylation of histone H3 at lysine 4 (H3K4me3) is a marker of active promoters. Broad H3K4me3 promoter domains have been associated with cell type identity, but H3K4me3 dynamics upon cellular stress have not been well characterized. We assessed this by exposing endometrial stromal cells to hypoxia, which is a major cellular stress condition. We observed that hypoxia modifies the existing H3K4me3 marks and that promoter H3K4me3 breadth rather than height correlates with transcription. Broad H3K4me3 domains mark genes for endometrial core functions and are maintained or selectively extended upon hypoxia. Hypoxic extension of H3K4me3 breadth associates with stress adaptation genes relevant for the survival of endometrial cells including transcription factor KLF4, for which we found increased protein expression in the stroma of endometriosis lesions. These results substantiate the view on broad H3K4me3 as a marker of cell identity genes and reveal participation of H3K4me3 extension in cellular stress adaptation.

Role of ER Stress Mediated Unfolded Protein Responses and ER Stress Inhibitors in the Pathogenesis of Inflammatory Bowel Disease.

Previous investigations have increased the knowledge about the pathological processes of inflammatory bowel diseases. Besides the complex organization of immune reactions, the mucosal epithelial lining has been recognized as a crucial regulator in the commencement and persistence of intestinal inflammation. As the intestinal epithelium is exposed to various environmental factors, the intestinal epithelial cells are confronted with diverse cellular stress conditions. In eukaryotic cells, an imbalance in the endoplasmic reticulum (ER) might cause aggregation of unfolded or misfolded proteins in the lumen of ER, a condition known as endoplasmic reticulum stress. This cellular mechanism stimulates the unfolded protein response (UPR), which elevates the potential of the endoplasmic reticulum protein folding, improves protein production and its maturation, and also stimulates ER-associated protein degradation. Current analyses reported that in the epithelium, the ER stress might cause the pathogenesis of inflammatory bowel disease that affects the synthesis of protein, inducing the apoptosis of the epithelial cell and stimulating the proinflammatory reactions in the gut. There have been significant efforts to develop small molecules or molecular chaperones that will be potent in ameliorating ER stress. The restoration of UPR balance in the endoplasmic reticulum via pharmacological intervention might be a novel therapeutic approach for the treatment of inflammatory bowel diseases (IBDs). This review provides novel insights into the role of chemical chaperone UPR modulators to modify ER stress levels. We further discuss the future directions/challenges in the development of therapeutic strategies for IBDs by targeting the ER stress. Figure depicting the role of endoplasmic reticulum stress-mediated inflammatory bowel disease and the therapeutic role of endoplasmic reticulum stress inhibitors in alleviating the diseased condition.

Periodic Heat Stress Licenses EMSC Differentiation into Osteoblasts via YAP Signaling Pathway Activation.

Background The repair and regeneration of large bone defects represent highly challenging tasks in bone tissue engineering. Although recent studies have shown that osteogenesis is stimulated by periodic heat stress, the thermal regulation of osteogenic differentiation in ectomesenchymal stem cells (EMSCs) is not well studied. Methods and Results In this study, the direct effects of periodic heat stress on the differentiation of EMSCs into osteoblasts were investigated. EMSCs derived from rat nasal respiratory mucosa were seeded onto culture plates, followed by 1 h of heat stress at 41°C every 7 days during osteogenic differentiation. Based on the results of the present study, periodic heating increases alkaline phosphatase (ALP) activity, upregulates osteogenic-related proteins, and promotes EMSC mineralization. In particular, increased YAP nuclear translocation and YAP knockdown inhibited osteogenic differentiation induced by heat stress. Furthermore, the expression and activity of transglutaminase 2 (TG2) were significantly increased after YAP nuclear translocation. Conclusion Together, these results indicate that YAP plays a key role in regulating cellular proteostasis under stressful cellular conditions by modulating the TG2 response.

Transmembrane protein 119 is neither a specific nor a reliable marker for microglia.

Microglia are the resident innate immune cells of the central nervous system (CNS) parenchyma. To determine the impact of microglia on disease development and progression in neurodegenerative and neuroinflammatory diseases, it is essential to distinguish microglia from peripheral macrophages/monocytes, which are eventually equally recruited. It has been suggested that transmembrane protein 119 (TMEM119) serves as a reliable microglia marker that discriminates resident microglia from blood-derived macrophages in the human and murine brain. Here, we investigated the validity of TMEM119 as a microglia marker in four in vivo models (cuprizone intoxication, experimental autoimmune encephalomyelitis (EAE), permanent filament middle cerebral artery occlusion (fMCAo), and intracerebral 6-hydroxydopamine (6-OHDA) injections) as well as post mortem multiple sclerosis (MS) brain tissues. In all applied animal models and post mortem MS tissues, we found increased densities of ionized calcium-binding adapter molecule 1+ (IBA1+ ) cells, paralleled by a significant decrease in TMEM119 expression. In addition, other cell types in peripheral tissues (i.e., follicular dendritic cells and brown adipose tissue) were also found to express TMEM119. In summary, this study demonstrates that TMEM119 is not exclusively expressed by microglia nor does it label all microglia, especially under cellular stress conditions. Since novel transgenic lines have been developed to label microglia using the TMEM119 promotor, downregulation of TMEM119 expression might interfere with the results and should, thus, be considered when working with these transgenic mouse models.

Serum Anti-heat Shock Protein 27 Antibody Titers in Patients With Dyslipidemia: A Population-based Case-control Study

Background: Heat shock protein 27 (HSP27) is found in several cell types of adults, such as cardiomyocytes, and endothelial cells. It is expressed in response to different cellular stress conditions. HSP27 decreases the levels of reactive oxygen species (ROS) and dyslipidemia is closely associated with increased endothelial production of reactive oxygen species (ROS). Objective: Higher serum HSP27 antigen and anti-HSP27 antibodies have been reported in patients with unstable angina and myocardial infarction Methods: This population-based case-control study was conducted in 2018. We investigated serum anti-HSP27 antibody titers in all participants with dyslipidemia from the Mashhad stroke and heart atherosclerotic disorder (MASHAD) study (n=8141) and those who were healthy in terms of dyslipidemia (n=1637) using an in-house enzyme-linked immune sorbent assay (ELISA) in individuals with dyslipidemia. Findings: Anti-HSP27 titers were significantly lower in individuals with dyslipidemia compared to people without dyslipidemia (P=0.036). Conclusion: Our results revealed that the anti-HSP27 antibody titer was lower in the participants with dyslipidemia than in the negative group. However, there may be a confounding effect of drug therapy. In a subgroup of dyslipidemic subjects, we observed lower anti-HSP27 antibody titers in patients treated with some drugs (statins or corticosteroids, non-steroidal anti-inflammatory drugs [NSAIDs], or anti-diabetic and anti-hypertensive) compared to subjects untreated with these drugs.

Chaperones Skp and SurA dynamically expand unfolded OmpX and synergistically disassemble oligomeric aggregates

Periplasmic chaperones 17-kilodalton protein (Skp) and survival factor A (SurA) are essential players in outer membrane protein (OMP) biogenesis. They prevent unfolded OMPs from misfolding during their passage through the periplasmic space and aid in the disassembly of OMP aggregates under cellular stress conditions. However, functionally important links between interaction mechanisms, structural dynamics, and energetics that underpin both Skp and SurA associations with OMPs have remained largely unresolved. Here, using single-molecule fluorescence spectroscopy, we dissect the conformational dynamics and thermodynamics of Skp and SurA binding to unfolded OmpX and explore their disaggregase activities. We show that both chaperones expand unfolded OmpX distinctly and induce microsecond chain reconfigurations in the client OMP structure. We further reveal that Skp and SurA bind their substrate in a fine-tuned thermodynamic process via enthalpy-entropy compensation. Finally, we observed synergistic activity of both chaperones in the disaggregation of oligomeric OmpX aggregates. Our findings provide an intimate view into the multifaceted functionalities of Skp and SurA and the fine-tuned balance between conformational flexibility and underlying energetics in aiding chaperone action during OMP biogenesis.