TGF-ß and SHH Regulate Pluripotent Stem Cell Differentiation into Brain Microvascular Endothelial Cells in Generating an In Vitro Blood-Brain Barrier Model.

Blood-brain barrier (BBB) models are important tools for studying CNS drug delivery, brain development, and brain disease. In vitro BBB models have been obtained from animals and immortalized cell lines; however, brain microvascular endothelial cells (BMECs) derived from them have several limitations. Furthermore, obtaining mature brain microvascular endothelial-like cells (BME-like cells) from human pluripotent stem cells (hPSCs) with desirable properties for establishing BBB models has been challenging. Here, we developed an efficient method for differentiating hPSCs into BMECs that are amenable to the development and application of human BBB models. The established conditions provided an environment similar to that occurring during BBB differentiation in the presence of the co-differentiating neural cell population by the modulation of TGF-ß and SHH signaling. The developed BME-like cells showed well-organized tight junctions, appropriate expression of nutrient transporters, and polarized efflux transporter activity. In addition, BME-like cells responded to astrocytes, acquiring substantial barrier properties as measured by transendothelial electrical resistance. Moreover, the BME-like cells exhibited an immune quiescent property of BBB endothelial cells by decreasing the expression of adhesion molecules. Therefore, our novel cellular platform could be useful for drug screening and the development of brain-permeable pharmaceuticals.

A Critical Role for ERO1α in Arterial Thrombosis and Ischemic Stroke.

BACKGROUND: Platelet adhesion and aggregation play a crucial role in arterial thrombosis and ischemic stroke. Here, we identify platelet ERO1α (endoplasmic reticulum oxidoreductase 1α) as a novel regulator of Ca2+ signaling and a potential pharmacological target for treating thrombotic diseases. METHODS: Intravital microscopy, animal disease models, and a wide range of cell biological studies were utilized to demonstrate the pathophysiological role of ERO1α in arteriolar and arterial thrombosis and to prove the importance of platelet ERO1α in platelet activation and aggregation. Mass spectrometry, electron microscopy, and biochemical studies were used to investigate the molecular mechanism. We used novel blocking antibodies and small-molecule inhibitors to study whether ERO1α can be targeted to attenuate thrombotic conditions. RESULTS: Megakaryocyte-specific or global deletion of Ero1α in mice similarly reduced platelet thrombus formation in arteriolar and arterial thrombosis without affecting tail bleeding times and blood loss following vascular injury. We observed that platelet ERO1α localized exclusively in the dense tubular system and promoted Ca2+ mobilization, platelet activation, and aggregation. Platelet ERO1α directly interacted with STIM1 (stromal interaction molecule 1) and SERCA2 (sarco/endoplasmic reticulum Ca2+-ATPase 2) and regulated their functions. Such interactions were impaired in mutant STIM1-Cys49/56Ser and mutant SERCA2-Cys875/887Ser. We found that ERO1α modified an allosteric Cys49-Cys56 disulfide bond in STIM1 and a Cys875-Cys887 disulfide bond in SERCA2, contributing to Ca2+ store content and increasing cytosolic Ca2+ levels during platelet activation. Inhibition of Ero1α with small-molecule inhibitors but not blocking antibodies attenuated arteriolar and arterial thrombosis and reduced infarct volume following focal brain ischemia in mice. CONCLUSIONS: Our results suggest that ERO1α acts as a thiol oxidase for Ca2+ signaling molecules, STIM1 and SERCA2, and enhances cytosolic Ca2+ levels, promoting platelet activation and aggregation. Our study provides evidence that ERO1α may be a potential target to reduce thrombotic events.

Hepatic PTP4A1 ameliorates high-fat diet-induced hepatosteatosis and hyperglycemia by the activation of the CREBH/FGF21 axis.

Precise regulation of kinases and phosphatases is crucial for human metabolic homeostasis. This study aimed to investigate the roles and molecular mechanisms of protein tyrosine phosphatase type IVA1 (PTP4A1) in regulating hepatosteatosis and glucose homeostasis. Method: Ptp4a1-/- mice, adeno-associated virus encoding Ptp4a1 under liver-specific promoter, adenovirus encoding Fgf21, and primary hepatocytes were used to evaluate PTP4A1-mediated regulation in the hepatosteatosis and glucose homeostasis. Glucose tolerance test, insulin tolerance test, 2-deoxyglucose uptake assay, and hyperinsulinemic-euglycemic clamp were performed to estimate glucose homeostasis in mice. The staining, including oil red O, hematoxylin & eosin, and BODIPY, and biochemical analysis for hepatic triglycerides were performed to assess hepatic lipids. Luciferase reporter assays, immunoprecipitation, immunoblots, quantitative real-time polymerase chain reaction, and immunohistochemistry staining were conducted to explore the underlying mechanism. Results: Here, we found that deficiency of PTP4A1 aggravated glucose homeostasis and hepatosteatosis in mice fed a high-fat (HF) diet. Increased lipid accumulation in hepatocytes of Ptp4a1-/- mice reduced the level of glucose transporter 2 on the plasma membrane of hepatocytes leading to a diminution of glucose uptake. PTP4A1 prevented hepatosteatosis by activating the transcription factor cyclic adenosine monophosphate-responsive element-binding protein H (CREBH)/fibroblast growth factor 21 (FGF21) axis. Liver-specific PTP4A1 or systemic FGF21 overexpression in Ptp4a1-/- mice fed an HF diet restored the disorder of hepatosteatosis and glucose homeostasis. Finally, liver-specific PTP4A1 expression ameliorated an HF diet-induced hepatosteatosis and hyperglycemia in wild-type mice. Conclusions: Hepatic PTP4A1 is critical for regulating hepatosteatosis and glucose homeostasis by activating the CREBH/FGF21 axis. Our current study provides a novel function of PTP4A1 in metabolic disorders; hence, modulating PTP4A1 may be a potential therapeutic strategy against hepatosteatosis-related diseases.

The EDN1/EDNRA/ß­arrestin axis promotes colorectal cancer progression by regulating STAT3 phosphorylation.

Endothelin receptor A (EDNRA) has been reported to play various crucial physiological roles and has been shown to be associated with the pathology of several diseases, including colorectal cancer (CRC). However, the molecular mechanisms of EDNRA in the development of human CRC have not been fully elucidated to date. In this context, the present study was performed to investigate biological functions and novel downstream signaling pathways affected by EDNRA, during CRC progression. First, using public data repositories, it was observed that the EDRNA expression levels were markedly increased in CRC tissues, as compared to normal tissues. Patients with CRC with an increased EDNRA expression exhibited a significantly decreased survival rate in comparison with those with a lower EDNRA expression. Furthermore, a positive correlation between the levels of EDNRA and its ligand, EDN1, was found in CRC tissues. The ectopic expression of EDNRA or its ligand, EDN1, promoted, whereas the silencing of EDNRA or EDN1 decreased cell proliferation and migration in vitro. To elucidate the signaling pathways involved in the regulation of EDNRA expression in CRC cells, a phosphokinase array analysis was performed, and it was observed that the knockdown of EDNRA substantially suppressed the phosphorylation of signal transducer and activator of transcription 3 (STAT3) in CRC cells. Of note, STAT3 silencing simultaneously decreased EDN1 and EDNRA expression, with the expression of EDN1 and/or EDNRA appearing to be directly regulated by binding STAT3 to their promoter region, according to chromatin immunoprecipitation and promoter assays, ultimately indicating a positive feedback loop in the expression of EDNRA and EDN1. It was also observed that treatment with an EDNRA antagonist (macitentan), alone or in combination with cisplatin, suppressed cell growth and migration ability, and induced cell apoptosis. Collectively, these data suggest a critical role of the EDN1/EDNRA signaling pathway in CRC progression. Thus, the pharmacological intervention of this signaling pathway may prove to be a potential therapeutic approach for patients with CRC.

Endothelial PTP4A1 mitigates vascular inflammation via USF1/A20 axis-mediated NF-κB inactivation.

AIMS: The nuclear factor-κB (NF-κB) signalling pathway plays a critical role in the pathogenesis of multiple vascular diseases. However, in endothelial cells (ECs), the molecular mechanisms responsible for the negative regulation of the NF-κB pathway are poorly understood. In this study, we investigated a novel role for protein tyrosine phosphatase type IVA1 (PTP4A1) in NF-κB signalling in ECs. METHODS AND RESULTS: In human tissues, human umbilical artery ECs, and mouse models for loss of function and gain of function of PTP4A1, we conducted histological analysis, immunostaining, laser-captured microdissection assay, lentiviral infection, small interfering RNA transfection, quantitative real-time PCR and reverse transcription-PCR, as well as luciferase reporter gene and chromatin immunoprecipitation assays. Short hairpin RNA-mediated knockdown of PTP4A1 and overexpression of PTP4A1 in ECs indicated that PTP4A1 is critical for inhibiting the expression of cell adhesion molecules (CAMs). PTP4A1 increased the transcriptional activity of upstream stimulatory factor 1 (USF1) by dephosphorylating its S309 residue and subsequently inducing the transcription of tumour necrosis factor-alpha-induced protein 3 (TNFAIP3/A20) and the inhibition of NF-κB activity. Studies on Ptp4a1 knockout or transgenic mice demonstrated that PTP4A1 potently regulates the interleukin 1ß-induced expression of CAMs in vivo. In addition, we verified that PTP4A1 deficiency in apolipoprotein E knockout mice exacerbated high-fat high-cholesterol diet-induced atherogenesis with upregulated expression of CAMs. CONCLUSION: Our data indicate that PTP4A1 is a novel negative regulator of vascular inflammation by inducing USF1/A20 axis-mediated NF-κB inactivation. Therefore, the expression and/or activation of PTP4A1 in ECs might be useful for the treatment of vascular inflammatory diseases.

Particulate matter induces ferroptosis by accumulating iron and dysregulating the antioxidant system.

Particulate matter is an air pollutant composed of various components, and has adverse effects on the human body. Particulate matter is known to induce cell death by generating an imbalance in the antioxidant system; however, the underlying mechanism has not been elucidated. In the present study, we demonstrated the cytotoxic effects of the size and composition of particulate matter on small intestine cells. We found that particulate matter 2.5 (PM2.5) with extraction ion (EI) components (PM2.5 EI), is more cytotoxic than PM containing only polycyclic aromatic hydrocarbons (PAHs). Additionally, PM-induced cell death is characteristic of ferroptosis, and includes iron accumulation, lipid peroxidation, and reactive oxygen species (ROS) generation. Furthermore, ferroptosis inhibitor as liproxstatin-1 and iron-chelator as deferiprone attenuated cell mortality, lipid peroxidation, iron accumulation, and ROS production after PM2.5 EI treatment in human small intestinal cells. These results suggest that PM2.5 EI may increase ferroptotic-cell death by iron accumulation and ROS generation, and offer a potential therapeutic clue for inflammatory bowel diseases in human small intestinal cells. [BMB Reports 2023; 56(2): 96-101].

A Sensitive Immunodetection Assay Using Antibodies Specific to Staphylococcal Enterotoxin B Produced by Baculovirus Expression.

Staphylococcal enterotoxin B (SEB) is a potent bacterial toxin that causes inflammatory stimulation and toxic shock, thus it is necessary to detect SEB in food and environmental samples. Here, we developed a sensitive immunodetection system using monoclonal antibodies (mAbs). Our study is the first to employ a baculovirus expression vector system (BEVS) to produce recombinant wild-type SEB. BEVS facilitated high-quantity and pure SEB production from suspension-cultured insect cells, and the SEB produced was characterized by mass spectrometry analysis. The SEB was stable at 4 °C for at least 2 years, maintaining its purity, and was further utilized for mouse immunization to generate mAbs. An optimal pair of mAbs non-competitive to SEB was selected for sandwich enzyme-linked immunosorbent assay-based immunodetection. The limit of detection of the immunodetection method was 0.38 ng/mL. Moreover, it displayed higher sensitivity in detecting SEB than commercially available immunodetection kits and retained detectability in various matrices and S. aureus culture supernatants. Thus, the results indicate that BEVS is useful for producing pure recombinant SEB with its natural immunogenic property in high yield, and that the developed immunodetection assay is reliable and sensitive for routine identification of SEB in various samples, including foods.

KLHL3 deficiency in mice ameliorates obesity, insulin resistance, and nonalcoholic fatty liver disease by regulating energy expenditure.

Obesity is a growing global epidemic that can cause serious adverse health consequences, including insulin resistance (IR) and nonalcoholic fatty liver disease (NAFLD). Obesity development can be attributed to energy imbalance and metabolic inflexibility. Here, we demonstrated that lack of Kelch-like protein 3 (KLHL3) mitigated the development of obesity, IR, and NAFLD by increasing energy expenditure. KLHL3 mutations in humans cause Gordon's hypertension syndrome; however, the role of KLHL3 in obesity was previously unknown. We examined differences in obesity-related parameters between control and Klhl3-/- mice. A significant decrease in body weight concomitant with fat mass loss and improved IR and NAFLD were observed in Klhl3-/- mice fed a high-fat (HF) diet and aged. KLHL3 deficiency inhibited obesity, IR, and NAFLD by increasing energy expenditure with augmentation of O2 consumption and CO2 production. Delivering dominant-negative (DN) Klhl3 using adeno-associated virus into mice, thereby dominantly expressing DN-KLHL3 in the liver, ameliorated diet-induced obesity, IR, and NAFLD. Finally, adenoviral overexpression of DN-KLHL3, but not wild-type KLHL3, in hepatocytes revealed an energetic phenotype with an increase in the oxygen consumption rate. The present findings demonstrate a novel function of KLHL3 mutation in extrarenal tissues, such as the liver, and may provide a therapeutic target against obesity and obesity-related diseases.

Monoclonal antibody K312-based depletion of pluripotent cells from differentiated stem cell progeny prevents teratoma formation.

Human pluripotent stem cells (PSCs) have been utilized as a promising source in regenerative medicine. However, the risk of teratoma formation that comes with residual undifferentiated PSCs in differentiated cell populations is most concerning in the clinical use of PSC derivatives. Here, we report that a monoclonal antibody (mAb) targeting PSCs could distinguish undifferentiated PSCs, with potential teratoma-forming activity, from differentiated PSC progeny. A panel of hybridomas generated from mouse immunization with H9 human embryonic stem cells (hESCs) was screened for ESC-specific binding using flow cytometry. A novel mAb, K312, was selected considering its high stem cell-binding activity, and this mAb could bind to several human induced pluripotent stem cells and PSC lines. Cell-binding activity of K312 was markedly decreased as hESCs were differentiated into embryoid bodies or by retinoic acid treatment. In addition, a cell population negatively isolated from undifferentiated or differentiated H9 hESCs via K312 targeting showed a significantly reduced expression of pluripotency markers, including Oct4 and Nanog. Furthermore, K312-based depletion of pluripotent cells from differentiated PSC progeny completely prevented teratoma formation. Therefore, our findings suggest that K312 is utilizable in improving stem cell transplantation safety by specifically distinguishing residual undifferentiated PSCs. [BMB Reports 2022; 55(3): 142-147].

Anisomycin protects against sepsis by attenuating IκB kinase-dependent NF-κB activation and inflammatory gene expression.

Anisomycin is known to inhibit eukaryotic protein synthesis and has been established as an antibiotic and anticancer drug. However, the molecular targets of anisomycin and its mechanism of action have not been explained in macrophages. Here, we demonstrated the anti-inflammatory effects of anisomycin both in vivo and in vitro. We found that anisomycin decreased the mortality rate of macrophages in cecal ligation and puncture (CLP)- and lipopolysaccharide (LPS)-induced acute sepsis. It also declined the gene expression of proinflammatory mediators such as inducible nitric oxide synthase, tumor necrosis factor-α, and interleukin-1ß as well as the nitric oxide and proinflammatory cytokines production in macrophages subjected to LPS-induced acute sepsis. Furthermore, anisomycin attenuated nuclear factor (NF)-κB activation in LPS-induced macrophages, which correlated with the inhibition of phosphorylation of NF-κBinducing kinase and IκB kinase, phosphorylation and IκBα proteolytic degradation, and NF-κB p65 subunit nuclear translocation. These results suggest that anisomycin prevented acute inflammation by inhibiting NF-κB-related inflammatory gene expression and could be a potential therapeutic candidate for sepsis. [BMB Reports 2021; 54(11): 545-550].

Immunomagnetic microfluidic integrated system for potency-based multiple separation of heterogeneous stem cells with high throughput capabilities.

Multipotent adult stem cells (MASCs) derived from Pluripotent stem cells (PSCs) have found widespread use in various applications, including regenerative therapy and drug screening. For these applications, highly pluripotent PSCs need to be selectively separated from those that show low pluripotency for reusage of PSCs, and MASCs need to be collected for further application. Herein, we developed immunomagnetic microfluidic integrated system (IM-MIS) for separation of stem cells depending on potency level. In this system, each stem cell was multiple-separated in microfluidics chip by magnetophoretic mobility of magnetic-activated cells based on the combination of two sizes of magnetic nanoparticles and two different antibodies. Magnetic particles had a difference in the degree of magnetization, and antibodies recognized potency-related surface markers. IM-MIS showed superior cell separation performance than FACS with high throughput (49.5%) in a short time (<15 min) isolate 1 × 107 cells, and higher purity (92.1%) than MACS. IM-MIS had a cell viability of 89.1%, suggesting that IM-MIS had no effect on cell viability during isolation. Furthermore, IM-MIS did not affect the key characteristics of stem cells including its differentiation potency, phenotype, genotype, and karyotype. IM-MIS may offer a new platform for the development of multi-separation systems for diverse stem cell applications.

ERO1-PDI Redox Signaling in Health and Disease.

Significance: Protein disulfide isomerase (PDI) and endoplasmic reticulum oxidoreductase 1 (ERO1) are crucial for oxidative protein folding in the endoplasmic reticulum (ER). These enzymes are frequently overexpressed and secreted, and they contribute to the pathology of neurodegenerative, cardiovascular, and metabolic diseases. Recent Advances: Tissue-specific knockout mouse models and pharmacologic inhibitors have been developed to advance our understanding of the cell-specific functions of PDI and ERO1. In addition to their roles in protecting cells from the unfolded protein response and oxidative stress, recent studies have revealed that PDI and ERO1 also function outside of the cells. Critical Issues: Despite the well-known contributions of PDI and ERO1 to specific disease pathology, the detailed molecular and cellular mechanisms underlying these activities remain to be elucidated. Further, although PDI and ERO1 inhibitors have been identified, the results from previous studies require careful evaluation, as many of these agents are not selective and may have significant cytotoxicity. Future Directions: The functions of PDI and ERO1 in the ER have been extensively studied. Additional studies will be required to define their functions outside the ER.

In vivo monitoring platform of transplanted human stem cells using magnetic resonance imaging.

As stem cells show great promise in regenerative therapy, stem cell-mediated therapeutic efficacy must be demonstrated through the migration and transplantation of stem cells into target disease areas at the pre-clinical level. In this study, we developed manganese-based magnetic nanoparticles with hollow structures (MnOHo) and modified them with the anti-human integrin ß1 antibody (MnOHo-Ab) to enable the minimal-invasive monitoring of transplanted human stem cells at the pre-clinical level. Compared to common magnetic resonance imaging (MRI)-based stem cell monitoring systems that use pre-labeled stem cells with magnetic particles before stem cell injection, the MnOHo-Ab is a new technology that does not require stem cell modification to monitor the therapeutic capability of stem cells. Additionally, MnOHo-Ab provides improved T1 MRI owing to the hollow structure of the MnOHo. Particularly, the anti-integrin ß1 antibody (Ab) introduced in the MnOHo targets integrin ß1 expressed in the entire stem cell lineage, enabling targeted monitoring regardless of the differentiation stage of the stem cells. Furthermore, we verified that intravenously injected MnOHo-Ab specifically targeted human induced pluripotent stem cells (hiPSCs) that were transferred to mice testes and differentiated into various lineages. The new stem cell monitoring method using MnOHo-Ab demonstrates whether the injected human stem cells have migrated and transplanted themselves in the target area during long-term stem cell regenerative therapy.

Loss of desmoglein-2 promotes gallbladder carcinoma progression and resistance to EGFR-targeted therapy through Src kinase activation.

Gallbladder carcinoma (GBC) exhibits poor prognosis due to local recurrence, metastasis, and resistance to targeted therapies. Using clinicopathological analyses of GBC patients along with molecular in vitro and tumor in vivo analysis of GBC cells, we showed that reduction of Dsg2 expression was highly associated with higher T stage, more perineural, and lymphatic invasion. Dsg2-depleted GBC cells exhibited significantly enhanced proliferation, migration, and invasiveness in vitro and tumor growth and metastasis in vivo through Src-mediated signaling activation. Interestingly, Dsg2 binding inhibited Src activation, whereas its loss activated cSrc-mediated EGFR plasma membrane clearance and cytoplasmic localization, which was associated with acquired EGFR-targeted therapy resistance and decreased overall survival. Inhibition of Src activity by dasatinib enhanced therapeutic response to anti-EGFR therapy. Dsg2 status can help stratify predicted patient response to anti-EGFR therapy and Src inhibition could be a promising strategy to improve the clinical efficacy of EGFR-targeted therapy.

Polyunsaturated fatty acid biosynthesis pathway determines ferroptosis sensitivity in gastric cancer.

Ferroptosis is an iron-dependent regulated necrosis mediated by lipid peroxidation. Cancer cells survive under metabolic stress conditions by altering lipid metabolism, which may alter their sensitivity to ferroptosis. However, the association between lipid metabolism and ferroptosis is not completely understood. In this study, we found that the expression of elongation of very long-chain fatty acid protein 5 (ELOVL5) and fatty acid desaturase 1 (FADS1) is up-regulated in mesenchymal-type gastric cancer cells (GCs), leading to ferroptosis sensitization. In contrast, these enzymes are silenced by DNA methylation in intestinal-type GCs, rendering cells resistant to ferroptosis. Lipid profiling and isotope tracing analyses revealed that intestinal-type GCs are unable to generate arachidonic acid (AA) and adrenic acid (AdA) from linoleic acid. AA supplementation of intestinal-type GCs restores their sensitivity to ferroptosis. Based on these data, the polyunsaturated fatty acid (PUFA) biosynthesis pathway plays an essential role in ferroptosis; thus, this pathway potentially represents a marker for predicting the efficacy of ferroptosis-mediated cancer therapy.

Dsg2-mediated c-Met activation in anaplastic thyroid cancer motility and invasion.

Anaplastic thyroid cancer (ATC) is a rapidly growing, highly metastatic cancer with limited therapeutic alternatives, thus targeted therapies need to be developed. This study aimed to examine desmoglein 2 (Dsg2) expression in ATC and its biological role and potential as a therapeutic target in ATC. Consequently, Dsg2 was downregulated or aberrantly expressed in ATC tissues. ATC patients with low Dsg2 expression levels also presented with distant metastasis. Dsg2 depletion significantly increased cell migration and invasion, with a relatively limited effect on ATC cell proliferation in vitro and increased distant metastasis in vivo. Dsg2 knockdown induced cell motility through the hepatocyte growth factor receptor (HGFR, c-Met)/Src/Rac1 signaling axis, with no alterations in the expression of EMT-related molecules. Further, specific targeting of c-Met significantly inhibited the motility of shDsg2-depleted ATC cells. Decreased membrane Dsg2 expression increased the metastatic potential of ATC cells. These results indicate that Dsg2 plays an important role in ATC cell migration and invasiveness. Therapies targeting c-Met might be effective among ATC patients with low membrane Dsg2 expression levels, indicating that the analysis of Dsg2 expression potentially provides novel insights into treatment strategies for ATC.

Deficiency of peroxiredoxin 2 exacerbates angiotensin II-induced abdominal aortic aneurysm.

Abdominal aortic aneurysm (AAA) is an inflammatory vascular disease characterized by structural deterioration of the aorta caused by inflammation and oxidative stress, leading to aortic dilatation and rupture. Peroxiredoxin 2 (PRDX2), an antioxidant enzyme, has been reported as a potential negative regulator of inflammatory vascular diseases, and it has been identified as a protein that is increased in patients with ruptured AAA compared to patients with nonruptured AAA. In this study, we demonstrated that PRDX2 was a pivotal factor involved in the inhibition of AAA progression. PRDX2 levels were increased in AAA compared with those in normal aortas in both humans and mice. Ultrasound imaging revealed that the loss of PRDX2 accelerated the development of AAA in the early stages and increased AAA incidence in mice infused with angiotensin II (Ang II). Prdx2-/- mice infused with Ang II exhibited increased aortic dilatation and maximal aortic diameter without a change in blood pressure. Structural deterioration of the aortas from Prdx2-/- mice infused with Ang II was associated with increases in the degradation of elastin, oxidative stress, and intramural thrombi caused by microhemorrhages, immature neovessels, and the activation of matrix metalloproteinases compared to that observed in controls. Moreover, an increase in inflammatory responses, including the production of cell adhesion molecules and the accumulation of inflammatory cells and proinflammatory cytokines due to PRDX2 deficiency, accelerated Ang II-induced AAA progression. Our data confirm that PRDX2 plays a role as a negative regulator of the pathological process of AAA and suggest that increasing PRDX2 activity may be a novel strategy for the prevention and treatment of AAA.

Thermal Contact Conductance-Based Thermal Behavior Analytical Model for a Hybrid Floor at Elevated Temperatures.

Hybrid floors infilled with polymeric materials between two steel plates were developed as a prefabricated floor system in the construction industry. However, the floor's fire resistance performance has not been investigated. To evaluate this, fire tests suggested by the Korean Standards should be performed. As these tests are costly and time consuming, the number of variables were limited. However, many variables can be investigated in other ways such as furnace tests and finite element analysis (FEA) with less cost and time. In this study, furnace tests on heated surface areas smaller than 1 m2 were conducted to investigate the thermal behavior of the hybrid floor at elevated temperatures. To obtain the reliability of the proposed thermal behavior analytical (TBA) model, verifications were conducted by FEAs. Thermal contact conductance including interfacial thermal properties between two materials was adopted in the TBA model, and the values at elevated temperatures were suggested based on thermo-gravimetric analyses results and verified by FEA. Errors between the tests and TBA model indicated that the model was adequate in predicting the temperature distribution in small-scale hybrids. Furthermore, larger furnace tests and analysis results were compared to verify the TBA model's application to different sized hybrid floors.

Selective elimination of human pluripotent stem cells by Anti-Dsg2 antibody-doxorubicin conjugates.

The self-renewal properties of human pluripotent stem cells (hPSCs) contribute to their efficacy in tissue regeneration applications yet increase the likelihood of teratoma formation, thereby limiting their clinical utility. To address this issue, we developed a tool to specifically target and neutralize undifferentiated hPSCs, thereby minimizing tumorigenicity risk without negatively affecting regenerated and somatic tissues. Specifically, we conjugated a monoclonal antibody (K6-1) previously generated in our laboratory against desmoglein 2 (Dsg2), which is highly differentially expressed in undifferentiated hPSCs versus somatic tissues, to the chemotherapeutic agent doxorubicin (DOX). The K6-1-DOX conjugates were selectively targeted and incorporated into Dsg2-positive hPSCs, leading to pH-dependent endosomal release and nuclear localization of DOX with subsequent cytotoxicity via an apoptotic caspase cascade. Conversely, Dsg2-negative fibroblasts showed minimal conjugate uptake or cytotoxicity, suggesting that K6-1-DOX treatment would yield few side effects owing to off-target effects. Selective removal of undifferentiated stem cells was also supported by in vivo studies using a mouse xenograft model, wherein hIgG-DOX- but not K6-1-DOX-pretreated-hPSC injection led to teratoma development. Together, these results validated the ability of the Dsg2-targeted antibody-anticancer drug conjugate to facilitate the safety of stem cell therapies.

Bacterial type III effector protein HopQ inhibits melanoma motility through autophagic degradation of vimentin.

Malignant melanoma is a fatal disease that rapidly spreads to the whole body. Treatments have limited efficiency owing to drug resistance and various side effects. Pseudomonas syringae pv. tomato (Pto) is a model bacterial pathogen capable of systemic infection in plants. Pto injects the effector protein HopQ into the plant cytosol via a type III secretion machinery and suppresses the host immunity. Intriguingly, host plant proteins regulated by HopQ are conserved even in humans and conferred in tumor metastasis. Nevertheless, the potential for HopQ to regulate human cancer metastasis was unknown. In this study, we addressed the suitability of HopQ as a possible drug against melanoma metastasis. In melanoma cells, overexpressed HopQ is phosphorylated and bound to 14-3-3 through its N-terminal domain, resulting in stronger interaction between HopQ and vimentin. The binding of HopQ to vimentin allowed for degradation of vimentin via p62-dependent selective autophagy. Attenuation of vimentin expression by HopQ inhibited melanoma motility and in vivo metastasis. These findings demonstrated that HopQ directly degraded vimentin in melanoma cells and could be applied to an inhibitor of melanoma metastasis.