Liver-specific Coxsackievirus and adenovirus receptor deletion develop metabolic dysfunction-associated fatty liver disease.

Metabolic dysfunction-associated fatty liver disease (MAFLD) is a common liver disease associated with obesity and is caused by the accumulation of ectopic fat without alcohol consumption. Coxsackievirus and adenovirus receptor (CAR) are vital for cardiac myocyte-intercalated discs and endothelial cell-to-cell tight junctions. CAR has also been reported to be associated with obesity and high blood pressure. However, its function in the liver is still not well understood. The liver of obese mice exhibit elevated CAR mRNA and protein levels. Furthermore, in the liver of patients with non-alcoholic steatohepatitis, CAR is reduced in hepatocyte cell-cell junctions compared to normal levels. We generated liver-specific CAR knockout (KO) mice to investigate the role of CAR in the liver. Body and liver weights were not different between wild-type (WT) and KO mice fed a paired or high-fat diet (HFD). However, HFD induced significant liver damage and lipid accumulation in CAR KO mice compared with WT mice. Additionally, inflammatory cytokines transcription, hepatic permeability, and macrophage recruitment considerably increased in CAR KO mice. We identified a new interaction partner of CAR using a protein pull-down assay and mass spectrometry. Apolipoprotein B mRNA editing enzyme catalytic polypeptide-like 3C (APOBEC3C) demonstrated a complex relationship with CAR, and hepatic CAR expression tightly regulated its level. Moreover, Apolipoprotein B (ApoB) and Low-density lipoprotein receptor (LDLR) levels correlated with APOBEC3C expression in the liver of CAR KO mice, suggesting that CAR may regulate lipid accumulation by controlling APOBEC3C activity. In this study, we showed that hepatic CAR deficiency increased cell-to-cell permeability. In addition, CAR deletion significantly increased hepatic lipid accumulation by inducing ApoB and LDLR expression. Although the underlying mechanism is unclear, CARs may be a target for the development of novel therapies for MAFLD.

Transcription factors BZR1 and PAP1 cooperate to promote anthocyanin biosynthesis in Arabidopsis shoots.

Anthocyanins play critical roles in protecting plant tissues against diverse stresses. The complicated regulatory networks induced by various environmental factors modulate the homeostatic level of anthocyanins. Here, we show that anthocyanin accumulation is induced by brassinosteroids (BRs) in Arabidopsis (Arabidopsis thaliana) shoots and shed light on the underlying regulatory mechanism. We observed that anthocyanin levels are altered considerably in BR-related mutants, and BRs induce anthocyanin accumulation by upregulating the expression of anthocyanin biosynthetic genes. Our genetic analysis indicated that BRASSINAZOLE RESISTANT 1 (BZR1) and PRODUCTION OF ANTHOCYANIN PIGMENT 1 (PAP1) are essential for BR-induced anthocyanin accumulation. The BR-responsive transcription factor BZR1 directly binds to the PAP1 promoter, regulating its expression. In addition, we found that intense anthocyanin accumulation caused by the pap1-D-dominant mutation is significantly reduced in BR mutants, implying that BR activity is required for PAP1 function after PAP1 transcription. Moreover, we demonstrated that BZR1 physically interacts with PAP1 to cooperatively regulate the expression of PAP1-target genes, such as TRANSPARENT TESTA 8, DIHYDROFLAVONOL 4-REDUCTASE, and LEUKOANTHOCYANIDIN DIOXYGENASE. Our findings indicate that BZR1 functions as an integral component of the PAP1-containing transcription factor complex, contributing to increased anthocyanin biosynthesis. Notably, we also show that functional interaction of BZR1 with PAP1 is required for anthocyanin accumulation induced by low nitrogen stress. Taken together, our results demonstrate that BR-regulated BZR1 promotes anthocyanin biosynthesis through cooperative interaction with PAP1 of the MBW complex.

Lazertinib versus Platinum-Based Chemotherapy with Epidermal Growth Factor Receptor (EGFR)-Positive Non-Small-Cell Lung Cancer after Failing EGFR-Tyrosine Kinase Inhibitor: A Real-World External Comparator Study.

BACKGROUND: Lazertinib is a third-generation tyrosine kinase inhibitor of epidermal growth factor receptor (EGFR-TKI) that selectively inhibit common EGFR mutation and T790M mutation in non-small-cell lung cancer (NSCLC) patients. No previous studies have compared lazertinib to platinum-based chemotherapy. We have compared lazertinib with platinum-based chemotherapy in EGFR-mutated NSCLC patients after previous EGFR-TKI therapy. METHODS: We retrospectively compared 200 patients from LASER201, LASER301, and LASER-PMS studies to 334 patients who were treated with platinum-based chemotherapy after previous EGFR-TKI from the Samsung Medical Center. After propensity score matching (PSM), we selected 156 patients from each group. The primary outcome was progression-free survival (PFS), with overall survival (OS), objective response rate (ORR), and time to treatment discontinuation (TTD) as secondary outcomes. RESULTS: The median follow-up of PFS was 15.61 months in the lazertinib group and 21.67 months in the external control group. The PFS was significantly longer in patients who were treated with lazertinib than those treated with platinum-based chemotherapy (10.97 months vs. 5.10 months; adjusted hazard ratio (HR) 0.40; 95% confidence interval (CI), 0.29-0.55; p < 0.01) after PSM. Lazertinib showed superior OS (32.23 months vs. 18.73 months; adjusted HR 0.45; 95% CI, 0.29-0.69; p < 0.001), ORR (64.1% vs. 47.4%), and TTD (11.66 months vs. 6.73 months; adjusted HR 0.54; 95% CI, 0.39-0.75; p < 0.001) compared to platinum-based chemotherapy. CONCLUSION: Based on this retrospective, external control study, lazertinib has demonstrated significantly better efficacy compared with platinum-based chemotherapy. The external controls provide important context to evaluate efficacy in single-arm studies.

Comparative effectiveness of lazertinib in patients with EGFR T790M-positive non-small-cell lung cancer using a real-world external control.

Lazertinib is a recently developed third-generation epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors used for patients with advanced EGFR T790M-positive non-small-cell lung cancer. We evaluated the effectiveness of lazertinib compared with osimertinib using an external control. We obtained individual patient data for the lazertinib arm from the LASER201 trial and the osimertinib arm from registry data at the Samsung Medical Center. In total, 75 and 110 patients were included in the lazertinib and osimertinib groups, respectively. After propensity score matching, each group had 60 patients and all baseline characteristics were balanced. The median follow-up duration was 22.0 and 29.6 months in the lazertinib and osimertinib group, respectively. The objective response rate (ORR) were 76.7% and 86.7% for lazertinib and osimertinib, respectively (p = 0.08). The median progression-free survival (PFS) was 12.3 months (95% confidence interval [CI] 9.5-19.1) and 14.4 months (95% CI 11.8-18.1) for the lazertinib and osimertinib group, respectively (hazard ratio [HR] 0.97; 95% CI 0.64-1.45, p = 0.86). The median overall survival with lazertinib was not reached and that with osimertinib was 29.8 months (HR 0.44; 95% CI 0.25-0.77, p = 0.005). Our study suggests that lazertinib has an ORR and PFS comparable to those of osimertinib and has the potential for superior survival benefits.

Induction of TNF-α by Filifactor alocis in THP-1 macrophagic cells.

OBJECTIVES: Filifactor alocis is an emerging periodontal pathogen, and macrophage-produced tumor necrosis factor-α (TNF-α) plays important roles in periodontal pathogenesis. In this study, we investigated F. alocis-stimulated TNF-α production in THP-1 macrophagic cells. DESIGN: Phorbol 12-myristate 13-acetate-differentiated THP-1 macrophagic cells were challenged with F. alocis ATCC 35896 for various durations. TNF-α mRNA expression and protein secretion were determined using RT-PCR and ELISA, respectively. Activation of protein kinases and transcription factor proteins was evaluated by Western blot analysis. RESULTS: Live F. alocis stimulated THP-1 cells to produce TNF-α in a dose-dependent manner. However, glutaraldehyde-killed or heat-killed F. alocis showed no effectiveness for TNF-α induction. In contrast, both live and killed Porphyromonas gingivalis robustly increased TNF-α expression. Furthermore, F. alocis was unable to stimulate TNF-α expression in Toll-like receptor 2 (TLR2) knockout THP-1 cells. F. alocis activated all three mitogen-activated protein kinases: extracellular signal-regulated kinase (ERK), p38, and c-Jun N-terminal kinase (JNK). Pharmacological inhibition of ERK and JNK, but not p38, significantly reduced F. alocis-induced TNF-α production. Finally, increased levels of phospho-c-Jun were detected in F. alocis-stimulated THP-1 cells. CONCLUSIONS: These results suggest that F. alocis induces TNF-α production in THP-1 macrophagic cells primarily by activating the TLR2, JNK, and c-Jun pathways.

Fatal outcome of severe fever with thrombocytopenia syndrome (SFTS) and severe and critical COVID-19 is associated with the hyperproduction of IL-10 and IL-6 and the low production of TGF-ß.

Severe fever with thrombocytopenia syndrome virus (SFTSV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can cause the hyperproduction of inflammatory cytokines, which have pathological effects in patient including severe or fatal cytokine storms. To characterize the effect of SFTSV and SARS-CoV-2 infection on the production of cytokines in severe fever with thrombocytopenia syndrome (SFTS) and COVID-19 patients, we performed an analysis of cytokines in SFTS and COVID-19 patients and also investigated the role of interleukin-10 (IL-10) in vitro studies: lipopolysaccharide-induced THP-1-derived macrophages, SFTSV infection of THP-1 cells, and SARS-CoV-2 infection of THP-1 cells. In this study, we found that levels of both IL-10 and IL-6 were significantly elevated, the level of transforming growth factor-ß (TGF-ß) was significantly decreased and IL-10 was elevated earlier than IL-6 in severe and critical COVID-19 and fatal SFTS patients, and inhibition of IL-10 signaling decreased the production of IL-6 and elevated that of TGF-ß. Therefore, the hyperproduction of IL-10 and IL-6 and the low production of TGF-ß have been linked to cytokine storm-induced mortality in fatal SFTS and severe and critically ill COVID-19 patients and that IL-10 can play an important role in the host immune response to severe and critical SARS-CoV-2 and fatal SFTSV infection.

Heterologous vaccination utilizing viral vector and protein platforms confers complete protection against SFTSV.

Severe fever with thrombocytopenia syndrome virus was first discovered in 2009 as the causative agent of severe fever with thrombocytopenia syndrome. Despite its potential threat to public health, no prophylactic vaccine is yet available. This study developed a heterologous prime-boost strategy comprising priming with recombinant replication-deficient human adenovirus type 5 (rAd5) expressing the surface glycoprotein, Gn, and boosting with Gn protein. This vaccination regimen induced balanced Th1/Th2 immune responses and resulted in potent humoral and T cell-mediated responses in mice. It elicited high neutralizing antibody titers in both mice and non-human primates. Transcriptome analysis revealed that rAd5 and Gn proteins induced adaptive and innate immune pathways, respectively. This study provides immunological and mechanistic insight into this heterologous regimen and paves the way for future strategies against emerging infectious diseases.

Antioxidant effect of ergothioneine on in vitro maturation of porcine oocytes.

BACKGROUND: Ergothioneine (EGT) is a natural amino acid derivative in various animal organs and is a bioactive compound recognized as a food and medicine. OBJECTIVES: This study examined the effects of EGT supplementation during the in vitro maturation (IVM) period on porcine oocyte maturation and subsequent embryonic development competence after in vitro fertilization (IVF). METHODS: Each EGT concentration (0, 10, 50, and 100 µM) was supplemented in the maturation medium during IVM. After IVM, nuclear maturation, intracellular glutathione (GSH), and reactive oxygen species (ROS) levels of oocytes were investigated. In addition, the genes related to cumulus function and antioxidant pathways in oocytes or cumulus cells were investigated. Finally, this study examined whether EGT could affect embryonic development after IVF. RESULTS: After IVM, the EGT supplementation group showed significantly higher intracellular GSH levels and significantly lower intracellular ROS levels than the control group. Moreover, the expression levels of hyaluronan synthase 2 and Connexin 43 were significantly higher in the 10 µM EGT group than in the control group. The expression levels of nuclear factor erythroid 2-related factor 2 (Nrf2) and NAD(P)H quinone dehydrogenase 1 (NQO1) were significantly higher in the oocytes of the 10 µM EGT group than in the control group. In the assessment of subsequent embryonic development after IVF, the 10 µM EGT treatment group improved the cleavage and blastocyst rate significantly than the control group. CONCLUSIONS: Supplementation of EGT improved oocyte maturation and embryonic development by reducing oxidative stress in IVM oocytes.

Structural changes in the retina after implantation of subretinal three-dimensional implants in mini pigs.

The retinal structural changes after subretinal implantation of three-dimensional (3D) microelectrodes were investigated in a mini pig. Three types of electrode were implanted into the subretinal spaces of nine mini pigs: 75-µm-high 3D electrodes on a 200-µm-thick right-angled polydimethylsiloxane (PDMS) substrate (group 1); a 140-µm-thick sloped PDMS substrate without electrodes (group 2); and a 140-µm-thick sloped PDMS substrate with 20-µm-high 3D electrodes (group 3). One mini pig was used as a control. Spectral domain-optical coherence tomography (SD-OCT) images were obtained at baseline and 2, 6, and 12 weeks post-surgery. Retinal specimens were immunostained using a tissue-clearing method 3 months post-implantation. The 75-µm-high 3D electrodes progressively penetrated the inner nuclear layer (INL) and touched the inner plexiform layer (IPL) 2 weeks post-surgery. At 6 weeks post-operatively, the electrodes were in contact with the nerve-fiber layer, accompanied by a severe fibrous reaction. In the other groups, the implants remained in place without subretinal migration. Immunostaining showed that retinal ganglion and bipolar cells were preserved without fibrosis over the retinal implants in groups 2 and 3 during the 12-week implantation period. In summary, SD-OCT and immunohistology results showed differences in the extent of reactions, such as fibrosis over the implants and penetration of the electrodes into the inner retinal layer depending on different types of electrodes. A sloped substrate performed better than a right-angled substrate in terms of retinal preservation over the implanted electrodes. The 20-µm-high electrodes showed better structural compatibility than the 75-µm-high 3D electrodes. There was no significant difference between the results of sloped implants without electrodes and 20-µm-high 3D electrodes, indicating that the latter had no adverse effects on retinal tissue.

Ovarian tumor deubiquitinase 6A regulates cell proliferation via deubiquitination of nucleolin and caspase­7.

Most proteins maintain protein homeostasis via post­translational modifications, including the ubiquitin­proteasome system. Deubiquitinating enzymes (DUBs) have essential intercellular roles, such as responses to DNA damage, proteolysis and apoptosis. Therefore, it is important to understand DUB­related diseases to identify DUBs that target abnormally regulated proteins in cells. Ovarian tumor deubiquitinase 6A (OTUD6A) was previously reported as a downregulated DUB in HCT116 cells with p53 knockdown. Therefore, it was expected that the relationship between OTUD6A and p53 would affect cell proliferation. In the present study, putative substrates of OTUD6A related to the p53 signaling pathway were identified. Application of liquid chromatography­tandem mass spectrometry and proteomic analysis led to the identification of nucleolin (known to bind p53) as a binding protein. In addition, immunoprecipitation studies determined that caspase­7, an apoptotic protein, is associated with p53 signaling and is regulated by OTUD6A. It was further identified that OTUD6A regulates the protein stability of nucleolin, but not caspase­7. It was also demonstrated that OTUD6A acts as a respective DUB through the deubiquitination of K48­linked polyubiquitin chain of nucleolin and the K63­linked polyubiquitin chain of caspase­7. Furthermore, overexpression of OTUD6A induced cell proliferation via enhancing cell cycle progression of MCF7 cells. Taken together, OTUD6A may be proposed as a target for anticancer therapy.

Combined effects of cadmium and ochratoxin A on intestinal barrier dysfunction in human Caco-2 cells and pig small intestinal epithelial cells.

Hazardous chemicals are commonly found in cereals and cereal-based products. However, most studies focus on the individual effects of these mycotoxins or metals, rather than their combined toxicity. The main objective of this study was to evaluate the combined effects of cadmium (Cd) and ochratoxin A (OTA) on intestinal barrier integrity using Caco-2 cells and pig small intestinal epithelial (PSI) cells as models of intestinal epithelial cells and to measure alterations in cell survival and barrier integrity. The combined effects on cell viability were assessed in terms of a combination of index values. These findings showed that co-exposure to Cd + OTA had synergistic effects on Caco-2 and PSI cells at 25%, 50%, and 75% inhibitory concentrations (IC25, IC50, and IC75, respectively) against cell viability. Individual Cd and OTA treatments had no effect, but combined Cd + OTA exposure resulted in synergistic down-regulation of paracellular apical junction complex proteins, such as claudin-1, occludin, and E-cadherin. The current findings indicate that the combined effects of OTA + Cd may have consequences at the gut level, which should not be underestimated when considering their risk to human health.

Enhancement of TRP Gene Expression and UV Absorption by Bioconverted Chestnut Inner Shell Extracts Using Lactiplantibacillus plantarum.

In this work, the suppression of tyrosinase-related genes, including an improvement in UV absorption effects of bioconverted CS extracts (BCS), was investigated to improve the skin-whitening effect. Total polyphenols and total flavonoids, which are bioactive components, increased 2.6- and 5.4-times in bioconversion using Lactiplantibacillus plantarum SM4, respectively, as compared to ultrasound-assisted extracts (UCS). The effect of BCS on radical scavenging activity, UV-A absorption, and tyrosinase activity inhibition, contributing to skin-whitening, were 1.3-, 1.2-, and 1.2-times higher than those of UCS, respectively. The main component identified in high-performance liquid chromatography (HPLC) was gallic acid in both UCS and BCS, which increased by 2.9-times following bioconversion. The gene expression of tyrosinase-related proteins, including TRP-1 and TRP-2 genes, was studied to confirm the suppression of melanin synthesis by BCS in order to identify the skin-whitening mechanism, and BCS decreased both genes' expression by 1.7- and 1.6-times, demonstrating that BCS effectively suppressed melanin synthesis. These findings imply that the chestnut inner shell can be employed as a cosmetic material by simultaneously inhibiting melanogenesis and enhancing UV-A absorption through bioconversion using L. plantarum SM4.

Protein Kinase B2 (PKB2/AKT2) Is Essential for Host Protection in CVB3-Induced Acute Viral Myocarditis.

Protein kinase B2 (AKT2) is involved in various cardiomyocyte signaling processes, including those important for survival and metabolism. Coxsackievirus B3 (CVB3) is one of the most common pathogens that cause myocarditis in humans. The role of AKT2 in CVB3 infection is not yet well understood. We used a cardiac-specific AKT2 knockout (KO) mouse to determine the role of AKT2 in CVB3-mediated myocarditis. CVB3 was injected intraperitoneally into wild-type (WT) and KO mice. The mice's survival rate was recorded: survival in KO mice was significantly decreased compared with WT mice (WT vs. KO: 73.3 vs. 27.1%). Myocardial damage and inflammation were significantly increased in the hearts of KO mice compared with those of WT mice. Moreover, from surface ECG, AKT2 KO mice showed a prolonged atria and ventricle conduction time (PR interval, WT vs. KO: 47.27 ± 1.17 vs. 64.79 ± 7.17 ms). AKT2 deletion induced severe myocarditis and cardiac dysfunction due to CVB3 infection. According to real-time PCR, the mRNA level of IL-1, IL-6, and TNF-α decreased significantly in KO mice compared with WT mice on Days 5 after infection. In addition, innate immune response antiviral effectors, Type I interferon (interferon-α and ß), and p62, were dramatically suppressed in the heart of KO mice. In particular, the adult cardiac myocytes isolated from the heart showed high induction of TLR4 protein in KO mice in comparison with WT. AKT2 deletion suppressed the activation of Type I interferon and p62 transcription in CVB3 infection. In cardiac myocytes, AKT2 is a key signaling molecule for the heart from damage through the activation of innate immunity during acute myocarditis.

Pseudoalteromone A, a Ubiquinone Derivative from Marine Pseudoalteromonas spp., Suppresses Melanogenesis.

An ubiquinone derivative, pseudoalteromone A (1), has been isolated from two marine-derived Pseudoalteromonas spp., APmarine002 and ROA-050, and its anti-melanogenesis activity was investigated. The anti-melanogenic capacity of pseudoalteromone A was demonstrated by assessing the intracellular and extracellular melanin content and cellular tyrosinase activity in the B16 cell line, Melan-a mouse melanocyte cell line, and MNT-1 human malignant melanoma cell line. Treatment with pseudoalteromone A (40 µg/mL) for 72 h reduced α-melanocyte-stimulating hormone (α-MSH)-induced intracellular melanin production by up to 44.68% in B16 cells and 38.24% in MNT-1 cells. Notably, pseudoalteromone A induced a concentration-dependent reduction in cellular tyrosinase activity in B16 cell, and Western blot analyses showed that this inhibitory activity was associated with a significant decrease in protein levels of tyrosinase and tyrosinase-related protein 1 (Tyrp-1), suggesting that pseudoalteromone A exerts its anti-melanogenesis activity through effects on melanogenic genes. We further evaluated the skin-whitening effect of pseudoalteromone A in the three-dimensional (3D) pigmented-epidermis model, MelanoDerm, and visualized the 3D distribution of melanin by two-photon excited fluorescence imaging in this human skin equivalent. Collectively, our findings suggest that pseudoalteromone A inhibits tyrosinase activity and expression and that this accounts for its anti-melanogenic effects in melanocytes.

Regulation of Cancer Metabolism by Deubiquitinating Enzymes: The Warburg Effect.

Cancer is a disorder of cell growth and proliferation, characterized by different metabolic pathways within normal cells. The Warburg effect is a major metabolic process in cancer cells that affects the cellular responses, such as proliferation and apoptosis. Various signaling factors down/upregulate factors of the glycolysis pathway in cancer cells, and these signaling factors are ubiquitinated/deubiquitinated via the ubiquitin-proteasome system (UPS). Depending on the target protein, DUBs act as both an oncoprotein and a tumor suppressor. Since the degradation of tumor suppressors and stabilization of oncoproteins by either negative regulation by E3 ligases or positive regulation of DUBs, respectively, promote tumorigenesis, it is necessary to suppress these DUBs by applying appropriate inhibitors or small molecules. Therefore, we propose that the DUBs and their inhibitors related to the Warburg effect are potential anticancer targets.

Cross-Linked Composite Gel Polymer Electrolyte Based on an H-Shaped Poly(ethylene oxide)-Poly(propylene oxide) Tetrablock Copolymer with SiO2 Nanoparticles for Solid-State Supercapacitor Applications.

Achieving high ionic conductivity, wide voltage window, and good mechanical strength in a single material remains a key challenge for polymer-based electrolytes for use in solid-state supercapacitors (SCs). Herein, we report cross-linked composite gel polymer electrolytes (CGPEs) based on multi-cross-linkable H-shaped poly(ethylene oxide)-poly(propylene oxide) (PEO-PPO) tetrablock copolymer precursors, SiO2 nanoparticles, and 1-ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide, an ionic liquid (IL). Self-standing CGPE membranes with a high IL content were prepared using in situ cross-linking reactions between the silane groups present in the precursor and the SiO2 surface. The incorporation of an optimal amount of SiO2 increased the cross-linking density of the resulting CGPE while reducing polymer-chain ordering and, consequently, increasing both ionic conductivity and mechanical strength. As a result, the CGPE with 0.1 wt % SiO2 exhibited a high ionic conductivity (2.22 × 10-3 S cm-1 at 25 °C), good tensile strength (453 kPa), and high thermal stability up to 330 °C. Finally, an all-solid-state SC assembled with the prepared CGPE showed a high operating voltage (3 V), a large specific capacitance (103.9 F g-1 at 1 A g-1), and excellent durability (94% capacitance retention over 10,000 charge/discharge cycles), which highlights its strong potential as a solid-state electrolyte for SCs.

Vascular Endothelial Integrity Affects the Severity of Enterovirus-Mediated Cardiomyopathy.

Coxsackievirus and adenovirus receptor (CAR) is present in epithelial and vascular endothelial cell junctions. We have previously shown a hemorrhagic phenotype in germ-line CAR knock-out mouse embryos; we have also found that CAR interacts with ZO-1 and ß-catenin. However, the role of CAR in vascular endothelial junction permeability has not been proven. To understand the roles of CAR in the vascular endothelial junctions, we generated endothelium-specific CAR knockout (CAR-eKO) mice. In the absence of CAR, the endothelial cell layer showed an increase in transmembrane electrical resistance (TER, Ω) and coxsackievirus permeability. Evans blue dye and 70 kDa dextran-FITC were delivered by tail vein injection. We observed increased vascular permeability in the hearts of adult CAR-eKO mice compare with wild-type (WT) mice. There was a marked increase in monocyte and macrophage penetration into the peritoneal cavity caused by thioglycolate-induced peritonitis. We found that CAR ablation in endothelial cells was not significantly increased coxsackievirus B3 (CVB3) induced myocarditis in murine model. However, tissue virus titers were significantly higher in CAR-eKO mice compared with WT. Moreover, CVB3 was detected in the brain of CAR-eKO mice. Endothelial CAR deletion affects the expression of major endothelial junction proteins, such as cadherin and platelet endothelial cell adhesion molecule-1 (PECAM-1) in the cultured endothelial cells as well as liver vessel. We suggest that CAR expression is required for normal vascular permeability and endothelial tight junction homeostasis. Furthermore, CVB3 organ penetration and myocarditis severities were dependent on the endothelial CAR level.

The antioxidant dieckol reduces damage of oxidative stress-exposed porcine oocytes and enhances subsequent parthenotes embryo development.

This study investigated the effect of the antioxidant dieckol, a component of Ecklonia cava, on maturation and developmental competence of porcine oocytes exposed to oxidative stress in vitro. Oocytes were matured in in vitro maturation (IVM) medium containing various concentrations of dieckol. The blastocyst formation rate was highest in the 0.5 µM dieckol-treated (0.5 DEK) group. The reactive oxygen species level was decreased, and the level of glutathione and expression of antioxidant genes (NFE2L, SOD1, and SOD2) at metaphase II were increased in the 0.5 DEK group. Abnormal spindle organization and chromosome misalignment were prevented in the 0.5 DEK group. Expression of maternal markers (CCNB1 and MOS) and activity of p44/42 mitogen-activated protein kinase were increased in the 0.5 DEK group. After parthenogenetic activation, the total number of cells per blastocyst was increased and the percentage of apoptotic cells was decreased in the 0.5 DEK group. Expression of development-related genes (CX45, CDX2, POU5F1, and NANOG), antiapoptotic genes (BCL2L1 and BIRC5), and a proapoptotic gene (CASP3) were altered in the 0.5 DEK group. These results indicate that the antioxidant dieckol improves IVM and subsequent development of porcine oocytes and can be used to improve the quality of oocytes under peroxidation experimental conditions.

The antioxidant icariin protects porcine oocytes from age-related damage in vitro.

OBJECTIVE: If fertilization does not occur within a specific period, the quality of unfertilized oocytes in the oviduct (in vivo aging) or in culture (in vitro aging) will deteriorate over time. Icariin (ICA), found in all species of Epimedium herbs, has strong antioxidant activity, and is thought to exert anti-aging effects in vitro. We asked whether ICA protects oocytes against age-related changes in vitro. METHODS: We analyzed the reactive oxygen species (ROS) levels and expression of antioxidant, maternal, and estrogen receptor genes, and along with spindle morphology, and the developmental competence and quality of embryos in the presence and absence of ICA. RESULTS: Treatment with 5 µM ICA (ICA-5) led to a significant reduction in ROS activity, but increased mRNA expression of glutathione and antioxidant genes (superoxide dismutase 1 [SOD1], SOD2, peroxiredoxin 5, and nuclear factor erythroid 2-like 2), during aging in vitro. In addition, ICA-5 prevented defects in spindle formation and chromosomal alignment, and increased mRNA expression of cytoplasmic maturation factor genes (bone morphogenetic protein 15, cyclin B1, MOS proto-oncogene, serine/threonine kinase, and growth differentiation factor-9). It also prevented apoptosis, increased mRNA expression of antiapoptotic genes (BCL2-like 1 and baculoviral IAP repeat-containing 5), and reduced mRNA expression of pro-apoptotic genes (BCL2 antagonist/killer 1 and activation of caspase-3). Although the maturation and cleavage rates were similar in all groups, the total cell number per blastocyst and the percentage of apoptotic cells at the blastocyst stage were higher and lower, respectively, in the control and ICA-5 groups than in the aging group. CONCLUSION: ICA protects oocytes against damage during aging in vitro; therefore, it can be used to improve assisted reproductive technologies.

Ab-interno surgical technique for the implantation of a wireless subretinal prosthesis in mini-pigs.

We sought to describe the surgical techniques required in the ab-interno method to implant subretinal prostheses in mini-pigs and suggest tips to facilitate optimal outcomes. During vitrectomy, the use of valved trocar cannulas was essential to stabilize the detached retina and implanted chip. As a first step in retinal detachment, a 23-gauge cannula with very small amount of viscoelastic material was used to establish the retinal hole and promote retinal detachment. Then, balanced salt solution was applied to increase the retinal detachment and diathermy was used to make opening for subretinal prosthesis. For easy positioning of the subretinal prosthesis, a curved laser probe was adopted when handling the subretinal prosthesis under the retina. After surgery, the sclerotomy sites were tightly sutured to prevent silicone oil leakage. Without special equipment, such as a 41-gauge tip, retinal detachment could be induced easily, while the prosthesis was also successfully inserted and manipulated under the retina without an iatrogenic retinal tear. Two weeks after the operation, the oil fully occupied the intraocular volume without leakage. The subretinal prosthesis remained stable without complication. Understanding the principle of the ab-interno method and considering several tips for improving surgical access may help to enhance surgical success rates of subretinal prostheses implantation.