We developed two-photon (TP) probes for DNA (ABI-Nu), cytoplasm (Pyr-CT), and mitochondria (BF-MT). We found that ABI-Nu binds to AT in the minor groove, while ABI-Nu and BF-MT are effective for tracking in the cytoplasm and mitochondria, respectively. These probes showed very large effective two-photon action cross section values of 2230, 1555, and 790 Göppert-Mayer units (1 GM  =  10(-50) cm(4) s photon(-1) molecule(-1)) at 740 nm with emission maxima at 473, 561, and 560 nm, respectively, in each organelle. Using these probes, we quantitatively estimated the mean nuclear area and the ratios of nuclei to cytoplasm and mitochondria to nuclei in human colon tissues by dual-colour two-photon microscopy imaging within 2  h after biopsy. The mean nuclear area and the nuclei to cytoplasm and mitochondria to cytoplasm ratios increased in the following order: normal colon mucosa

The safety of polyethylene glycol plus ascorbic acid has not been fully investigated in patients with renal insufficiency. High-dose ascorbic acid could induce hyperoxaluria, thereby causing tubule-interstitial nephritis and renal failure. This study aims to evaluate the safety and efficacy of polyethylene glycol plus ascorbic acid in patients with chronic kidney disease.We retrospectively reviewed prospectively collected data on colonoscopy in patients with impaired renal function. Patients were divided into 2 groups: 2 L polyethylene glycol plus ascorbic acid (n = 61) and 4 L polyethylene glycol (n = 80). The safety of the 2 groups was compared by assessing the differences in laboratory findings before and after bowel cleansing.The laboratory findings were not significantly different before and after the administration of 2 L polyethylene glycol plus ascorbic acid or 4 L polyethylene glycol. In both groups, the estimated glomerular filtration rate was not influenced by the administration of the bowel-cleansing agent. Patients' reports on tolerance and acceptability were better in the 2 L polyethylene glycol plus ascorbic acid group than in the 4 L polyethylene glycol group.The 2 L polyethylene glycol plus ascorbic acid solution is a safe choice for bowel preparation before colonoscopy in patients with impaired renal function.

Chronic inflammation is a driving force for the development of metabolic disease including diabetes and obesity. However, the functional characteristics of T-cell senescence in the abnormal glucose homeostasis are not fully understood. We studied the patients visiting a hospital for routine health check-ups, who were divided into two groups: normal controls and people with prediabetes. Gene expression profiling of peripheral blood mononuclear cells from normal controls and patients with type 2 diabetes was undertaken using microarray analysis. We also investigated the immunometabolic characteristics of peripheral and hepatic senescent T cells in the normal subjects and patients with prediabetes. Moreover, murine senescent T cells were tested functionally in the liver of normal or mice with metabolic deterioration caused by diet-induced obesity. Human senescent (CD28-CD57+) CD8+ T cells are increased in the development of diabetes and proinflammatory cytokines and cytotoxic molecules are highly expressed in senescent T cells from patients with prediabetes. Moreover, we demonstrate that patients with prediabetes have higher concentrations of reactive oxygen species (ROS) in their senescent CD8+ T cells via enhancing capacity to use glycolysis. These functional properties of senescent CD8+ T cells contribute to the impairment of hepatic insulin sensitivity in humans. Furthermore, we found an increase of hepatic senescent T cells in mouse models of aging and diet-induced obesity. Adoptive transfer of senescent CD8+ T cells also led to a significant deterioration in systemic abnormal glucose homeostasis, which is improved by ROS scavengers in mice. This study defines a new clinically relevant concept of T-cell senescence-mediated inflammatory responses in the pathophysiology of abnormal glucose homeostasis. We also found that T-cell senescence is associated with systemic inflammation and alters hepatic glucose homeostasis. The rational modulation of T-cell senescence would be a promising avenue for the treatment or prevention of diabetes.

It is currently unclear which antiviral agent, entecavir (ETV) or tenofovir disoproxil fumarate (TDF), is superior for improving prognosis in patients with chronic hepatitis B (CHB). Here, we assessed the ability of these 2 antivirals to prevent liver-disease progression in treatment-naïve patients with CHB.

The seroclearance of hepatitis B virus (HBV) surface antigen (HBsAg) is regarded as a functional cure of chronic hepatitis B (CHB) although it occurs rarely. Recently, several genome-wide association studies (GWASs) revealed various genetic alterations related to the clinical course of HBV infection. However, all of these studies focused on the progression of HBV infection to chronicity and had limited application because of the heterogeneity of HBV genotypes. In the present study, we aimed to determine susceptibility genetic markers for seroclearance of HBsAg in CHB patients with a homogenous viral genotype.

Mitochondrial trans-2-enoyl-CoA reductase (MECR) is involved in mitochondrial synthesis of fatty acids and is highly expressed in mitochondria. MECR is also known as nuclear receptor binding factor-1, which was originally reported with yeast two-hybrid screening as a binding protein of the nuclear hormone receptor peroxisome proliferator-activated receptor α (PPARα). However, MECR and PPARα are localized at different compartment, mitochondria, and the nucleus, respectively. Therefore, the presence of a cytosolic or nuclear isoform of MECR is necessary for functional interaction between MECR and PPARα.

Although early recognition and treatment with effective antibiotics have lead to improvements in the prognosis of patients with spontaneous bacterial peritonitis (SBP), it remains to be a serious complication in cirrhotic patients. This study was designed to evaluate the clinical manifestations and prognosis of patients with liver cirrhosis and SBP in Korea.

Copine1 (CPNE1), known as a calcium-dependent membrane-binding protein, has tandem C2 domains and an A domain. We previously demonstrated that CPNE1 directly induces neuronal differentiation via Protein kinase B (AKT) phosphorylation in the hippocampal progenitor cell line, HiB5. To better understand its cellular function, we carried out a yeast two-hybrid screening to find CPNE1 binding partners. Among the identified proteins, 14-3-3γ appears to directly interact with CPNE1. Between CPNE1 and 14-3-3γ, the physical interaction as well as the specific binding regions of CPNE1 was confirmed in vitro and in vivo. Furthermore, among the seven 14-3-3 isotypes, only 14-3-3γ directly interacts with CPNE1. Our results also demonstrate that AKT phosphorylation, neurite outgrowth and expression of the neuronal marker protein are increased when 14-3-3γ is overexpressed in CPNE1 high expressed HiB5 cells. Furthermore, the neighboring Ser54 amino acids residue of C2A domain in CPNE1 has an important role in binding with 14-3-3γ, and in differentiation-related function of CPNE1. Moreover, mutation of Ser54 amino acids residue in CPNE1 effectively decreased association with 14-3-3γ and neuronal differentiation of HiB5 cells. Collectively, our findings indicate that 14-3-3γ regulates the differentiation ability of CPNE1 through the binding with C2A domain of CPNE1 in HiB5 cells.

Previously, we noted that SLIT3, slit guidance ligand 3, had an osteoprotective role with bone formation stimulation and bone resorption suppression. Additionally, we found that global Slit3 KO mice had smaller long bone. Skeletal staining showed short mineralized length in the newborn KO mice and wide hypertrophic chondrocyte area in the embryo KO mice, suggesting delayed chondrocyte maturation. The recombinant SLIT3 did not cause any change in proliferation of ATDC5 cells, but stimulated expressions of chondrocyte differentiation markers, such as COL2A1, SOX9, COL10A1, VEGF, and MMP13 in the cells. SLIT3 suppressed β-catenin activity in the cells, and activation of Wnt/β-catenin signaling by lithium chloride attenuated the SLIT3-stimulated differentiation markers. ATDC5 cells expressed only ROBO2 among their 4 isotypes, and the Robo2 knock-down with its siRNA reversed the SLIT3-stimulated differentiated markers in chondrocytes. Taken together, these indicate that SLIT3/ROBO2 promotes chondrocyte maturation via the inhibition of β-catenin signaling.

High-flow nasal cannula (HFNC) oxygen therapy has been widely used in critically ill patients. Despite the effectiveness of HFNC as a treatment, optimal methods to withdraw HFNC after recovery from preexisting conditions have not been investigated to date. In this study, we will evaluate the safety and efficacy of simultaneous reduction of flow and fraction of inspired oxygen (FiO2) compared with sequential reduction of either flow first or FiO2 reduction first in patients with HFNC.

Knee osteoarthritis (OA) is characterized by knee cartilage degeneration and secondary bone hyperplasia, resulting in pain, stiffness, and gait disturbance. The relationship between knee OA and neurodegenerative diseases is still unclear. This study used an Alzheimer's disease (AD) mouse model to observe whether osteoarthritis accelerates dementia progression by analyzing brain histology and neuroinflammation. Knee OA was induced by destabilizing the medial meniscus (DMM) in control (WT) and AD (5xFAD) mice before pathological symptoms. Mouse knee joints were scanned with a micro-CT scanner. A sham operation was used as control. Motor and cognitive abilities were tested after OA induction. Neurodegeneration, β-amyloid plaque formation, and neuroinflammation were analyzed by immunostaining, Western blotting, and RT-PCR in brain tissues. Compared with sham controls, OA in AD mice increased inflammatory cytokine levels in brain tissues. Furthermore, OA significantly increased β-amyloid deposition and neuronal loss in AD mice compared to sham controls. In conclusion, knee OA accelerated amyloid plaque deposition and neurodegeneration in AD-OA mice, suggesting that OA is a risk factor for AD.

Since tumor mutational burden (TMB) and gene expression profiling (GEP) have complementary effects, they may have improved predictive power when used in combination. Here, we investigated the ability of TMB and GEP to predict the immunotherapy response in patients with non-small cell lung cancer (NSCLC) and assessed if this combination can improve predictive power compared to that when used individually.

Activation of hepatocyte cannabinoid receptor-1 (CB1R) by hepatic stellate cell (HSC)-derived 2-arachidonoylglycerol (2-AG) drives de novo lipogenesis in alcoholic liver disease (ALD). How alcohol stimulates 2-AG production in HSCs is unknown. Here, we report that chronic alcohol consumption induced hepatic cysteine deficiency and subsequent glutathione depletion by impaired transsulfuration pathway. A compensatory increase in hepatic cystine-glutamate anti-porter xCT boosted extracellular glutamate levels coupled to cystine uptake both in mice and in patients with ALD. Alcohol also induced the selective expression of metabotropic glutamate receptor-5 (mGluR5) in HSCs where mGluR5 activation stimulated 2-AG production. Consistently, genetic or pharmacologic inhibition of mGluR5 or xCT attenuated alcoholic steatosis in mice via the suppression of 2-AG production and subsequent CB1R-mediated de novo lipogenesis. We conclude that a bidirectional signaling operates at a metabolic synapse between hepatocytes and HSCs through xCT-mediated glutamate-mGluR5 signaling to produce 2-AG, which induces CB1R-mediated alcoholic steatosis.

Irreversible electroporation (IRE) is a newly developed non-thermal ablative therapy. During the IRE procedure, the permeability of the cell membrane is irreversibly changed by application of high-energy pulses across the tissue. This induces the breakdown of cell homeostasis, and thereby cell death. Here, we present an in vivo study to demonstrate IRE ablation of gastric tissue and characterize the changes that occur with time therein. No significant complications were observed in the test rats during the experiment. The electroporated tissues exhibited apoptosis at 10, 24 and 48 h after IRE ablation. The apoptosis peaked at 10 h after IRE and then declined, suggesting that the ablated tissue rapidly recovered owing to intense metabolic activity. In addition, the electroporated tissues exhibited morphological changes such as pyknosis and karyorrhexis, while histological analysis showed that the blood vessels were preserved. Interestingly, electroporation greatly affected the mucosa and muscularis propria, but not the submucosa and serosa. This study suggests that IRE could potentially be used as a minimally invasive treatment for early gastric cancer that does not exhibit lymph node metastasis or dysplasia.

Useful biomarkers for metabolic syndrome have been insufficient. We investigated the performance of serum milk fat globule-EGF factor-8 (MFG-E8), the key mediator of inflammatory pathway, in diagnosis of metabolic syndrome.

Besifovir dipivoxil maleate (BSV), an acyclic nucleotide phosphonate, shows potent antiviral activity against hepatitis B virus. Our previous 48-week trial revealed that BSV has comparable antiviral efficacy to tenofovir disoproxil fumarate (TDF) and better safety profiles in terms of improved renal and bone safety. This extension study evaluated the prolonged efficacy and safety of BSV in treatment-naive chronic hepatitis B patients.

Exerkines are soluble factors secreted by exercised muscles, mimicking the effects of exercise in various organs, including the muscle itself. Lumican is reportedly secreted from muscles; however, its roles in skeletal muscle remain unknown. Herein, we found that lumican mRNA expression in the extensor digitorum longus was significantly higher in exercised mice than in unloading mice, and lumican stimulated myogenesis in vitro. Additionally, lumican knockdown significantly decreased muscle mass and cross-sectional area (CSA) of the muscle fiber in the gastrocnemius muscle of exercised mice. Lumican upregulated phosphorylation of p38 mitogen-activated protein kinase (MAPK) and a p38 inhibitor near completely blocked lumican-stimulated myogenesis. Inhibitors for integrin α2β1 and integrin ανβ3 also prevented lumican-stimulated myogenesis. Systemic lumican treatment, administered via the tail vein for 4 weeks, significantly increased relative muscle masses by 36.1% in ovariectomized mice. In addition, intramuscular lumican injection into unloaded muscles for 2 weeks significantly increased muscle mass by 8.5%. Both intravenous and intramuscular lumican treatment significantly increased muscle CSA. Our in vitro and in vivo experiments indicate that lumican is a muscle-secreted exerkine that affords protection against muscle loss by activating p38 MAPK via integrin receptors.

Noninvasive techniques for evaluating the severity of nonalcoholic fatty liver disease (NAFLD) have shown limited diagnostic performance. MicroRNAs (miRNAs) are useful biomarkers for diagnosing and monitoring the progression and treatment response to several diseases. Here, we evaluated whether serum exosomal miRNAs could be used for the diagnosis and prognosis of NAFLD severity. Exosomal miRNAs were isolated from the sera of 41 patients with NAFLD (diagnosed using liver biopsy) for microarray profiling. The degree of NAFLD severity was determined using inflammation, steatosis, and ballooning scores and the NAFLD activity score (NAS). Correlations between miRNA expression, clinical and biochemical parameters, and mRNA expression were analyzed. Overall, 25, 11, 13, and 14 miRNAs correlated with the inflammation score, steatosis score, ballooning score, and NAS, respectively, with 33 significant correlations observed between 27 miRNAs and six clinical variables. Eight miRNAs (let-7b-5p, miR-378h, -1184, -3613-3p, -877-5p, -602, -133b, and 509-3p) showed anticorrelated patterns with the corresponding mRNA expression. In fibrosis, 52 and 30 interactions corresponding to high miRNA-low mRNA and low miRNA-high mRNA expression, respectively, were observed. The present results therefore suggest that serum exosomal miRNAs can be used to evaluate NAFLD severity and identify potential targets for NAFLD treatment.

Background: Crosstalk between brown adipose tissue (BAT) and the liver is receiving increasing attention. This study investigated the effect of BAT dysfunction by thermoneutral (TN) housing on liver fibrosis in mice and examined the effect of secreted factors from brown adipocytes on the activation of hepatic stellate cells (HSCs). Methods: The carbon tetrachloride (CCl4)-induced liver fibrosis mouse model was used to evaluate fibrotic changes in the livers of mice housed under standard and TN conditions. The effect of BAT on the activation of HSCs was examined using cultured cells treated with conditioned media from brown adipocytes. Results: Under TN conditions, mice with CCl4-induced liver fibrosis exhibited increased liver injury, collagen deposition, and alpha smooth muscle actin (α-SMA) expression in the liver compared with mice maintained at room temperature. The numbers of liver-infiltrating immune cells and T cells producing IL-17A and IFN-γ were also significantly increased in the livers of mice housed under TN conditions. Treatment of HSCs with conditioned media from brown adipocytes markedly attenuated HSC activation, as shown by down-regulated α-SMA expression at day 4, day 7 and day 10 of culture. At thermoneutrality, with CCl4 administration, IL-10-deficient mice exhibited more severe liver fibrosis than wild-type mice. Interestingly, conditioned media from IL-10-deficient brown adipocytes could up-regulate the expression of α-SMA and induce HSCs activation. Conclusions: BAT inactivation by thermoneutrality contributes to the activation of pro-inflammatory and pro-fibrotic pathways in mice with CCl4-induced liver fibrosis. Normal brown adipocytes secreted factors that impair the activation of HSCs, while this protective effect was lost in IL-10-deficient brown adipocytes. Thus, the BAT-liver axis may serve as a potential therapeutic target for liver fibrosis, and IL-10 may be a key factor regulating the activation of HSCs by BAT.

Alcoholic liver disease (ALD) and nonalcoholic fatty liver disease (NAFLD) are typically asymptomatic and slow-progressing but potentially fatal diseases that are common causes of liver cirrhosis and related complications. Exosomes are nano-sized extracellular vesicles that have been linked to various intercellular communication processes and can carry biological materials reflecting the state and severity of disease. In this study, shotgun proteomic analysis of the protein expression profiles of extracellular vesicles, including exosomes and microvesicles, enriched from human serum samples of 24 patients diagnosed with various fatty liver diseases was performed using liquid chromatography tandem mass spectrometry (LC-MS/MS) followed by protein identification and label-free quantification using the MaxQuant platform. A total of 329 proteins, including 190 previously reported exosome-specific proteins, were identified from four types of liver disease, where significant differences in protein expression were found in apolipoproteins, immunoglobulins, and other previously reported markers of liver disease. Principal component analysis of 61 proteins identified from MaxQuant analysis of the LC-MS/MS data provided a confident differentiation between ALD and NAFLD. SIGNIFICANCE: The current investigation revealed the difference among various types of liver disease using LC-MS/MS of exosomes enriched from human serum samples of 24 patients where the most significantly up-regulation proteins were alpha-2-macroglobulin for alcoholic hepatitis and apolipoprotein C3 for nonalcoholic fatty liver disease.