Real-time monitoring of geosmin based on an aptamer-conjugated graphene field-effect transistor

In this paper, we propose a novel field-effect transistor (FET) using graphene, which is a two-dimensional (2D) nanomaterial, capable of evaluating water quality, and immobilizing the surface of a graphene micropatterned transistor with a highly responsive bioprobe for a water contamination indicator, geosmin, with high selectivity. A high-quality bioprobe-immobilized graphene FET (GFET) was fabricated for the real-time monitoring of geosmin using a liquid-gate measurement configuration. Immobilization was confirmed by measuring the change in the electrical characteristics of the platform (slope of the current-voltage (I？V) curve) and fluorescence images. In addition, a selectivity test showed remarkable implementation of the highly sensitive sensing platform with an insignificant signal when a nontarget was added. Using the fabricated device, the linear range for geosmin detection was determined to be from 0.01 nM - 1 μM with a detection limit of 0.01 nM. In addition, geosmin concentrations as low as 10 nM could be determined from river water samples with the sensor platform. This sensor can be utilized to immediately determine the presence of odorous substances by analyzing a water supply source without additional pretreatment. Another advantage is that the sensor device is a promising tool that does not have special equipment that requirs careful maintenance. In addition, the device provides a new platform for detecting harmful substances in various water sources by varying the bioprobes that are empolyed.

Characteristics of genetic variations associated with Lennox-Gastaut syndrome in Korean families

Lennox-Gastaut syndrome (LGS) is a severe type of childhood-onset epilepsy characterized by multiple types of seizures, specific discharges on electroencephalography, and intellectual disability. Most patients with LGS do not respond well to drug treatment and show poor long-term prognosis. Approximately 30% of patients without brain abnormalities have unidentifiable causes. Therefore, accurate diagnosis and treatment of LGS remain challenging. To identify causative mutations of LGS, we analyzed the whole-exome sequencing data of 17 unrelated Korean families, including patients with LGS and LGS-like epilepsy without brain abnormalities, using the Genome Analysis Toolkit. We identified 14 mutations in 14 genes as causes of LGS or LGS-like epilepsy. 64 percent of the identified genes were reported as LGS or epilepsy-related genes. Many of these variations were novel and considered as pathogenic or likely pathogenic. Network analysis was performed to classify the identified genes into two network clusters: neuronal signal transmission or neuronal development. Additionally, knockdown of two candidate genes with insufficient evidence of neuronal functions, SLC25A39 and TBC1D8, decreased neurite outgrowth and the expression level of MAP2, a neuronal marker. These results expand the spectrum of genetic variations and may aid the diagnosis and management of individuals with LGS.

Molecular profile of African swine fever virus (ASFV) circulating in Vietnam during 2019-2020 outbreaks

African swine fever (ASF) is a highly infectious disease of pigs caused by African swine fever virus (ASFV). In order to identify potential genetic variations among ASFV strains circulating in Vietnam, 26 ASFV isolates from organs and blood samples collected from domestic pigs from 23 different provinces of northern, central and southern Vietnam during 2019-2020 ASF outbreaks were genetically characterized. Nucleotide sequences were determined for a portion of the B646L (p72) gene, the complete E183L (p54) gene, the variable region of EP402R (CD2v), the central variable region (CVR) of pB602L, and a tandem repeat sequence (TRS) between the I73R and I329L genes. Analysis of the partial B646L (p72) and EP402R (CD2v) gene sequences and the full-length E183L (p54) gene sequence showed that all 26 ASFV isolates belonged to genotype II and serotype VIII and that they were identical to the strain Georgia/2007/1 and all ASFV strains sequenced in China. The TRS between the I73R and I329L genes contained a 10-nucleotide insertion that was observed in the Chinese ASFV strain CN201801 isolated from domestic pigs in 2018, but not in the Georgia/2007/1 and China/Jilin/2018/boar strains isolated from wild boar in China. This is the first intra-epidemic genome analysis reported for the ASFV strains circulating in Vietnam.

Identifying genome-wide off-target sites of CRISPR RNA-guided nucleases and deaminases with Digenome-seq

Digested genome sequencing (Digenome-seq) is a highly sensitive, easy-to-carry-out, cell-free method for experimentally identifying genome-wide off-target sites of programmable nucleases and deaminases (also known as base editors). Genomic DNA is digested in vitro using clustered regularly interspaced short palindromic repeats ribonucleoproteins (RNPs; plus DNA-modifying enzymes to cleave both strands of DNA at sites containing deaminated base products, in the case of base editors) and subjected to whole-genome sequencing (WGS) with a typical sequencing depth of 30×. A web-based program is available to map in vitro cleavage sites corresponding to on- and off-target sites. Chromatin DNA, in parallel with histone-free genomic DNA, can also be used to account for the effects of chromatin structure on off-target nuclease activity. Digenome-seq is more sensitive and comprehensive than cell-based methods for identifying off-target sites. Unlike other cell-free methods, Digenome-seq does not involve enrichment of DNA ends through PCR amplification. The entire process other than WGS, which takes ~1-2 weeks, including purification and preparation of RNPs, digestion of genomic DNA and bioinformatic analysis after WGS, takes about several weeks.

A combination of Olea europaea leaf extract and Spirodela polyrhiza extract alleviates atopic dermatitis by modulating immune balance and skin barrier function in a 1-chloro-2,4-dinitrobenzene-induced murine model

Background: Atopic dermatitis is a chronic inflammatory skin disease in humans. Although Olea europaea leaf extract (OLE) and Spirodela polyrhiza extract (SPE) have been used to protect against skin damage, the effects of their combined administration on atopic dermatitis have yet to studied. Purpose: In this study, we evaluated the potential therapeutic effects of an OLE and SPE combination on the progression of atopic dermatitis and the possible mechanisms underlying these effects in 1-chloro-2,4-dinitrobenzene (DNCB)-treated NC/Nga mice. Methods: Atopic dermatitis was induced by topical application of 0.2% w/v DNCB prepared in an olive oil:acetone solution (1:3), and thereafter OLE, SPE and OLE + SPE were administered orally for 5 weeks. We determined atopic dermatitis symptoms, serum IgE levels, and levels of cytokine- and gene expression in the dorsal skin and splenocytes, and performed histological and immune cell subtype analyses. The expression of skin barrier-related proteins (filaggrin, sirtuin 1, and claudin 1) was also evaluated. Results: The OLE + SPE combination significantly ameliorated atopic dermatitis symptoms, including dermatitis scores, and reduced epidermal thickness and infiltration of different inflammatory cells in mice with DNCB-induced atopic dermatitis. It also significantly reduced the number of CD4+, CD8+, and CD4+/CD69+ T cells; immunoglobulin E-producing B cells (CD23+/B220+) in the axillary lymph nodes; CD3+ T-cell eosinophils (chemokine-chemokine receptor 3+/CD11b+) in the skin; and CD3+ T cells, immunoglobulin E-producing B cells (CD23+/B220+), and eosinophils in peripheral blood mononuclear cells. Additionally, the experimental combination lowered levels of serum immunoglobulin E and histamine, as well as Th2-mediated cytokines, and interleukin-4, -5, and -13, whereas it increased the levels of Th1-mediated cytokine interferon-γ in splenocytes. Furthermore, the preparation significantly restored expression of the skin barrier-related proteins filaggrin, sirtuin 1, and claudin 1, and also reduced the expression of the inflammatory cytokine interleukin-6 and chemokine-chemokine receptor 3, as well as the pruritus-related cytokine interleukin-31 and interleukin-31 receptor, in atopic dermatitis skin lesions. Conclusion: Taken together, our findings indicate that administration of a combination of OLE and SPE can alleviate atopic dermatitis symptoms by regulating immune balance and skin barrier function and may be an effective therapeutic option for the treatment of atopic dermatitis.

Effect of microbial short-chain fatty acids on CYP3A4-mediated metabolic activation of human pluripotent stem cell-derived liver organoids

The early and accurate prediction of the hepatotoxicity of new drug targets during nonclinical drug development is important to avoid postmarketing drug withdrawals and late-stage failures. We previously established long-term expandable and functional human-induced pluripotent stem cell (iPSC)-derived liver organoids as an alternative source for primary human hepatocytes. However, PSC-derived organoids are known to present immature fetal characteristics. Here, we treated these liver organoids with microbial short-chain fatty acids (SCFAs) to improve metabolic maturation based on microenvironmental changes in the liver during postnatal development. The effects of the three main SCFA components (acetate, propionate, and butyrate) and their mixture on liver organoids were determined. Propionate (1 μM) significantly promoted the CYP3A4/CYP3A7 expression ratio, and acetate (1 μM), propionate (1 μM), and butyrate (1 μM) combination treatment, compared to no treatment (control), substantially increased CYP3A4 activity and albumin secretion, as well as gene expression. More importantly, mixed SCFA treatment accurately revealed troglitazone-induced hepatotoxicity, which was redeemed on a potent CYP3A4 inhibitor ketoconazole treatment. Overall, we determined, for the first time, that SCFA mixture treatment might contribute to the accurate evaluation of the CYP3A4-dependent drug toxicity by improving metabolic activation, including CYP3A4 expression, of liver organoids.

Native high-density lipoproteins (HDL) with higher paraoxonase exerts a potent antiviral effect against SARS-CoV-2 (COVID-19), while glycated HDL lost the antiviral activity

Human high-density lipoproteins (HDL) show a broad spectrum of antiviral activity in terms of anti-infection. Although many reports have pointed out a correlation between a lower serum HDL-C and a higher risk of COVID-19 infection and progression, the in vitro antiviral activity of HDL against SARS-CoV-2 has not been reported. HDL functionality, such as antioxidant and anti-infection, can be impaired by oxidation and glycation and a change to pro-inflammatory properties. This study compared the antiviral activity of native HDL with glycated HDL via fructosylation and native low-density lipoproteins (LDL). After 72 h of fructosylation, glycated HDL showed a typical multimerized protein pattern with an elevation of yellowish fluorescence. Glycated HDL showed a smaller particle size with an ambiguous shape and a loss of paraoxonase activity up to 51% compared to native HDL. The phagocytosis of acetylated LDL was accelerated 1.3-fold by glycated HDL than native HDL. Native HDL showed 1.7 times higher cell viability and 3.6 times higher cytopathic effect (CPE) inhibition activity against SARS-CoV-2 than that of glycated HDL under 60 μg/mL (approximately final 2.2 μM) in a Vero E6 cell. Native HDL showed EC50 = 52.1 ± 1.1 μg/mL (approximately final 1.8 μM) for the CPE and CC50 = 79.4 ± 1.5 μg/mL (around 2.8 μM). The selective index (SI) of native HDL was calculated to be 1.52. In conclusion, native HDL shows potent antiviral activity against SARS-CoV-2 without cytotoxicity, while the glycation of HDL impairs its antiviral activity. These results may explain why patients with diabetes mellitus or hypertension are more sensitive to a COVID-19 infection and have a higher risk of mortality.

Regulatory function of peroxiredoxin I on 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced lung cancer development

Smoking is a major cause of lung cancer, and 4？(methylnitrosamino)？1？(3？pyridyl)？1？butanone (NNK) is one of the most important carcinogens in cigarette smoke. NNK modulates the expression of peroxiredoxin (Prdx) I in lung cancer. Prdx1 is upregulated in lung squamous cell carcinoma and lung adenocarcinoma, and considered a potential biomarker for lung cancer. The current article reviewed the role and regulatory mechanisms of Prdx1 in NNK？induced lung cancer cells. Prdx1 protects erythrocytes and DNA from NNK？induced oxidative damage, prevents malignant transformation of cells and promotes cytotoxicity of natural killer cells, hence suppressing tumor formation. In addition, Prdx1 has the ability to prevent NNK？induced lung tumor metabolic activity and generation of large amount of reactive oxygen species (ROS) and ROS？induced apoptosis, thus promoting tumor cell survival. In contrast to this, Prdx1, together with NNK, can promote the epithelial？mesenchymal transition and migration of lung tumor cells. The signaling pathways associated with NNK and Prdx1 in lung cancer cells have been discussed in present review; however, numerous potential pathways are yet to be studied. To develop novel methods for treating NNK？induced lung cancer, and improve the survival rate of patients with lung cancer, further research is needed to understand the complete mechanism associated with NNK.

Safety pharmacology of self-assembled-micelle inhibitory RNA-targeting amphiregulin (SAMiRNA-AREG), a novel siRNA nanoparticle platform

The present safety pharmacology core battery studies (neurobehavior, respiratory, cardiovascular system, and human ether a-go-go (hERG) channel current) investigated the potential harmful effects of self-assembled-micelle inhibitory RNA-targeting amphiregulin (SAMiRNA-AREG). The SAMiRNA-AREG was administered by single intravenous injection at up to 300？mg/kg and 100？mg/kg in mice and monkeys, respectively. The hERG assay was performed in Chinese hamster ovary (CHO) cells at SAMiRNA-AREG concentrations of up to 200？μg/mL. In the evaluation on neurobehavior, a transient decrease in body temperature was found at 0.5？h (30？min) post-dose at both sexes in mice, with a single 300？mg/kg dose of SAMiRNA-AREG. However, these effects had returned to normal at 1？h post-dose. In the evaluation on hERG channel current, there were statistically significant differences in the inhibition of peak hERG potassium channel current between the 20, 100, and 200？μg/mL SAMiRNA-AREG treatment groups and the vehicle control group. However, these effects were less potent than that of E-4031, a positive control article. For the respiratory and cardiovascular systems, no treatment-related changes were observed in mice or monkeys. Thus, under these experimental conditions, these studies suggest that SAMiRNA-AREG showed no adverse effects on the neurobehavior, respiratory, and cardiovascular function.

Tryptophanyl-tRNA synthetase as a potential therapeutic target

Tryptophanyl-tRNA synthetase (WRS) is an essential enzyme that catalyzes the ligation of tryptophan (Trp) to its cognate tRNAtrp during translation via aminoacylation. Interestingly, WRS also plays physiopathological roles in diseases including sepsis, cancer, and autoimmune and brain diseases and has potential as a pharmacological target and therapeutic. However, WRS is still generally regarded simply as an enzyme that produces Trp in polypeptides; therefore, studies of the pharmacological effects, therapeutic targets, and mechanisms of action of WRS are still at an emerging stage. This review summarizes the involvement of WRS in human diseases. We hope that this will encourage further investigation into WRS as a potential target for drug development in various pathological states including infection, tumorigenesis, and autoimmune and brain diseases.