Genotoxicity and safety pharmacology of the rVSVInd(GML)-mspSGtc vaccine against SARS-CoV-2 in Sprague-Dawley rats and Beagle dogs.

The emergence of coronavirus disease (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) led to a pandemic, prompting rapid vaccine development. Although vaccines are effective, the occurrence of rare adverse events following vaccination highlights the necessity of determining whether the benefits outweigh the risks posed by the infection itself. The recombinant Vesicular Stomatitis Virus (rVSV) platform is a promising vector for vaccines against emerging viruses. However, limited studies have evaluated the genotoxicity and safety pharmacology of this viral vector vaccine, which is crucial to ensure the safety of vaccines developed using this platform. Hence, the present study aimed to assess the genotoxicity and safety pharmacology of the rVSVInd(GML)-mspSGtc COVID-19 vaccine using micronucleus and comet assays, as well as neurobehavioral, body temperature, respiratory, and cardiovascular assessments in Sprague-Dawley rats and beagle dogs. The intramuscular administration of rVSVInd(GML)-mspSGtc at doses up to 1.5 × 109 PFU/animal did not increase the number of bone marrow micronucleated polychromatic erythrocytes or cause liver DNA damage. Additionally, it had no significant impact on neurobehavioral functions in rats and showed marginal temporary changes in body temperature, respiratory rate, heart rate, and electrocardiogram parameters in rats and dogs, all of which resolved within 24 h. Overall, following genotoxicity and pharmacological safety assessments, rVSVInd(GML)-mspSGtc displayed no notable systemic adverse effects in rats and dogs, suggesting its potential as a vaccine candidate for human clinical trials.

Prevalence and characterization of non-typhoidal Salmonella in egg from grading and packing plants in Korea.

Egg washing guidelines vary across countries; however, since 2020, Korea has required that all eggs produced from farms with more than 10,000 laying hens must be washed through egg grading and packing (GP) plant. This study investigated the prevalence and characterization of non-typhoidal Salmonella in eggs after washing at GP plants. In total, 16,800 eggs were collected from 60 egg GP plants located inside commercial layer farms, and 840 pooled eggshell and egg contents were tested for Salmonella, respectively. Of the 60 GP plants tested, 11 (18.3%) and 12 (20.0%) plants were positive for Salmonella spp. In the eggshells and egg contents, respectively. In particular, High Salmonella prevalence in the eggshells and egg contents occurred most often in farms with laying hens older than 80 weeks (33.3% and 40.0%, respectively). However, among 840 pooled eggshells and egg content samples, only 19 (2.3%) of each sample type were positive only for non-typhoidal Salmonella spp. The most common Salmonella serovar in both eggshells and egg contents was S. Infantis, which was found in five (8.3%) of 60 GP plants for both samples types. The other Salmonella serovars detected in eggshells were S. Bareilly (5.0%), S. Agona (3.3%), S. Enteritidis (1.7%), and S. Montevideo (1.7%), whereas those detected in egg contents were S. Enteritidis (5.0%), S. Agona (3.3%), S. Newport (3.3%), S. Senftenberg (3.3%), and S. Derby (1.7%). Of the 19 virulence genes tested, 14 genes were detected in all Salmonella. Interestingly, the spvB gene was detected only in S. Enteritidis, and the sefC gene was detected only in S. Enteritidis and S. Senftenberg. Moreover, all S. Infantis isolates showed multidrug resistance (MDR) against five or more classes, and the other serovars only showed MDR against three to four classes or no MDR. These results suggest that comprehensive surveillance and advanced management approaches for egg GP plants are required to minimize egg contamination with non-typhoidal Salmonella.

Emergence of Salmonella Infantis carrying the pESI-like plasmid from eggs in egg grading and packing plants in Korea.

The plasmid of emerging S. Infantis (pESI) or pESI-like plasmid in Salmonella enterica Infantis are consistently reported in poultry and humans worldwide. However, there has been limited research on these plasmids of S. Infantis isolated from eggs. Therefore, this study aimed to analyze the prevalence and characteristics of S. Infantis carrying the pESI-like plasmid from eggs in egg grading and packing plants. In this study, the pESI-like plasmid was only detected in 18 (78.3%) of 23 S. Infantis isolates, and it was absent in the other 9 Salmonella serovars. In particular, S. Infantis isolates carrying the pESI-like plasmid showed the significantly higher resistance to ß-lactams, phenicols, cephams, aminoglycosides, quinolones, sulfonamides, and tetracyclines than Salmonella isolates without the pESI-like plasmid (p < 0.05). Moreover, all S. Infantis isolates carrying the pESI-like plasmid were identified as extended-spectrum ß-lactamase (ESBL) producer, harboring the blaCTX-M-65 and blaTEM-1 genes, and carried non-ß-lactamase resistance genes (ant(3'')-Ia, aph(4)-Ia, aac(3)-IVa, aph(3')-Ic, sul1, tetA, dfrA14, and floR) against five antimicrobial classes. However, all isolates without the pESI-like plasmid only carried the blaTEM-1 gene among the ß-lactamase genes, and either had no non-ß-lactamase resistance genes or harbored non-ß-lactamase resistance genes against one or two antimicrobial classes. Furthermore, all S. Infantis isolates carrying the pESI-like plasmid carried class 1 and 2 integrons and the aadA1 gene cassette, but none of the other isolates without the pESI-like plasmid harbored integrons. In particular, D87Y substitution in the gyrA gene and IncP replicon type were observed in all the S. Infantis isolates carrying the pESI-like plasmid but not in the S. Infantis isolates without the pESI-like plasmid. The distribution of pulsotypes between pESI-positive and pESI-negative S. Infantis isolates was clearly distinguished, but all S. Infantis isolates were classified as sequence type 32, regardless of whether they carried the pESI-like plasmid. This study is the first to report the characteristics of S. Infantis carrying the pESI-like plasmid isolated from eggs and can provide valuable information for formulating strategies to control the spread of Salmonella in the egg industry worldwide.

Genetic and Phenotypic Diversity of Listeria monocytogenes in Pig Slaughterhouses in Korea.

Listeria monocytogenes is a foodborne pathogen that has variable subtypes associated with human listeriosis and occurs in food and processing environments. This study was conducted to provide the genetic and phenotypic characterization of L. monocytogenes in pig carcasses and environments of slaughterhouses in Korea. A total of 22 L. monocytogenes were isolated from eight of 26 pig slaughterhouses between 2020 and 2022, and the most common serotype was 1/2c (40.9%), followed by serotypes 1/2b (31.8%) and 1/2a (27.3%). The isolates showed a significantly high prevalence of virulence genes located in Listeria pathogenicity island-1 (LIPI-1) and internalins (90.9-100%; p < 0.05). However, the prevalence rates of llsX, ptsA, and stress survival islet-1 (SSI-1) located in LIPI-3, LIPI-4, and SSI were only 9.1%, 22.7%, and 31.8%, respectively. In addition, among the epidemic clones (EC), ECI, ECII, ECIII, and ECV, only one isolate was represented as ECV. Isolates identified from the same slaughterhouses were divided into two or more pulsotypes, except for two slaughterhouses. Furthermore, the seven STs were classified into seven clonal complexes (CCs) (CC8, CC9, CC37, CC87, CC121, CC155, and CC288), and all CCs belonged to lineages I (31.8%) and II (68.1%). Interestingly, the isolates showed a high prevalence of oxacillin resistance (59.1%), and most isolates of the serotypes 1/2a and 1/2b exhibited oxacillin resistance, whereas only one of nine serotype 1/2c isolates exhibited oxacillin resistance. These results provide the genetic diversity of L. monocytogenes in pig carcasses and environments of slaughterhouses, and continuous monitoring will be helpful in predicting food safety risks.

Determination of pepsin in human saliva using pepsin-susceptible peptide reporter and colorimetric dipstick assay: a prospective, cross-sectional study.

A simple and effective pepsin detection assay is reported based on a pepsin-susceptible peptide (PSP) reporter degradation strategy. PSP, which can be specifically cleaved by pepsin, was modified with fluorescein isothiocyanate (FITC) and biotin at the N- and C-terminals to be used as a reporter for colorimetric detection of dipsticks. A universal lateral flow dipstick consisting of a streptavidin test line for biotin binding and a sample pad immobilized with a gold-labeled polyclonal (rabbit) anti-FITC antibody was used to verify PSP-based pepsin detection. When the PSP reporter reacts with pepsin in a tube, it cleaves into two fragments, and the cleaved fragments do not display any color on the test line. Therefore, the higher the concentration of pepsin is, the greater is the decrease in test line intensity (IT-line) and the higher is the control line intensity (IC-line). First, the PSP cleavage and dipstick assay conditions for pepsin detection was optimized. The ratio of color intensity (IT-line/IC-line) of PSP-based dipstick assay showed a linear relationship with log concentration of pepsin ranging between 4 and 500 ng/mL (R2 = 0.98, n = 6), with a limit of detection of 1.4 ng/mL. It also exhibited high specificity and good reproducibility. Finally, pepsin levels were quantified in saliva samples from healthy controls (n = 34) and patients with laryngopharyngeal reflux (LPR, n = 61). Salivary pepsin levels were higher in patients with LPR than in healthy controls. The salivary pepsin levels correlated with those measured using a conventional enzyme-linked immunosorbent assay kit. Therefore, this PSP-based dipstick assay is a convenient tool for assessing salivary pepsin levels.

Therapeutic Efficacy of YM155 to Regulate an Epigenetic Enzyme in Major Subtypes of RCC.

Renal cell carcinoma (RCC) is the most common type of kidney cancer and includes more than 10 subtypes. Compared to the intensively investigated clear cell RCC (ccRCC), the underlying mechanisms and treatment options of other subtypes, including papillary RCC (pRCC) and chromogenic RCC (chRCC), are limited. In this study, we analyzed the public databases for ccRCC, pRCC, and chRCC and found that BIRC5 was commonly overexpressed in a large cohort of pRCC and chRCC patients as well as ccRCC and was closely related to the progression of RCCs. We investigated the potential of BIRC5 as a therapeutic target for these three types of RCCs. Loss and gain of function studies showed the critical role of BIRC5 in cancer growth. YM155, a BIRC5 inhibitor, induced a potent tumor-suppressive effect in the three types of RCC cells and xenograft models. To determine the mechanism underlying the anti-tumor effects of YM155, we examined epigenetic modifications in the BIRC5 promoter and found that histone H3 lysine 27 acetylation (H3K27Ac) was highly enriched on the promoter region of BIRC5. Chromatin-immunoprecipitation analysis revealed that H3K27Ac enrichment was significantly decreased by YM155. Immunohistochemistry of xenografted tissue showed that overexpression of BIRC5 plays an important role in malignancy in RCC. Furthermore, high expression of P300 was significantly associated with the progression of RCC. Our findings demonstrate the P300-H3K27Ac-BIRC5 cascade in three types of RCC and provide a therapeutic path for future research on RCC.

ß-Cyclodextrin Nanophotosensitizers for Redox-Sensitive Delivery of Chlorin e6.

The aim of this study is to prepare redox-sensitive nanophotosensitizers for the targeted delivery of chlorin e6 (Ce6) against cervical cancer. For this purpose, Ce6 was conjugated with ß-cyclodextrin (bCD) via a disulfide bond, creating nanophotosensitizers that were fabricated for the redox-sensitive delivery of Ce6 against cancer cells. bCD was treated with succinic anhydride to synthesize succinylated bCD (bCDsu). After that, cystamine was attached to the carboxylic end of bCDsu (bCDsu-ss), and the amine end group of bCDsu-ss was conjugated with Ce6 (bCDsu-ss-Ce6). The chemical composition of bCDsu-ss-Ce6 was confirmed with 1H and 13C NMR spectra. bCDsu-ss-Ce6 nanophotosensitizers were fabricated by a dialysis procedure. They formed small particles with an average particle size of 152.0 ± 23.2 nm. The Ce6 release rate from the bCDsu-ss-Ce6 nanophotosensitizers was accelerated by the addition of glutathione (GSH), indicating that the bCDsu-ss-Ce6 nanophotosensitizers have a redox-sensitive photosensitizer delivery capacity. The bCDsu-ss-Ce6 nanophotosensitizers have a low intrinsic cytotoxicity against CCD986Sk human skin fibroblast cells as well as Ce6 alone. However, the bCDsu-ss-Ce6 nanophotosensitizers showed an improved Ce6 uptake ratio, higher reactive oxygen species (ROS) production, and phototoxicity compared to those of Ce6 alone. GSH addition resulted in a higher Ce6 uptake ratio, ROS generation, and phototoxicity than Ce6 alone, indicating that the bCDsu-ss-Ce6 nanophotosensitizers have a redox-sensitive biological activity in vitro against HeLa human cervical cancer cells. In a tumor xenograft model using HeLa cells, the bCDsu-ss-Ce6 nanophotosensitizers efficiently accumulated in the tumor rather than in normal organs. In other words, the fluorescence intensity in tumor tissues was significantly higher than that of other organs, while Ce6 alone did not specifically target tumor tissue. These results indicated a higher anticancer activity of bCDsu-ss-Ce6 nanophotosensitizers, as demonstrated by their efficient inhibition of the growth of tumors in an in vivo animal tumor xenograft study.

Next-Generation Sequencing Results Vary Between Cultured and Uncultured Microbes.

Next-generation sequencing (NGS) technology has led to innovations in environmental metagenomics and investigations involving humans and microbes. However, it is necessary to analyze the components that will affect analysis of the method upon processing a large amount of information. In particular, the processing method after sample collection affects the NGS results, and it is necessary to check for inaccurate results. Here, we show that the microbial communities obtained from fingertip samples differ from those obtained from fingertips remaining on mobile phones and desks, when cultured or not for 24 h. We also confirmed changes in microbial communities in fingertip samples from desks incubated for 2, 4, 8, 16, and 24 h. Samples of prints from mobile phones that are considerably vulnerable to external factors were not analyzed. Ratios of Firmicutes and Bacillus were, respectively, increased in cultures at the phylum and species levels. Collectively, we identified bacterial species that can aid in determining whether a sample has been cultured. In addition, although microbial communities differed depending on sample types, we confirmed changes after culture for 4 and 8 h. However, since this study is a sample limited to three types, it is necessary to analyze other types of samples in the same way and check whether they are applicable to all types. This strategy can verify the suitability of samples for deriving informative results from cultured or uncultured bacterial communities.

Simple and Sensitive Detection of Bacterial Hydrogen Sulfide Production Using a Paper-Based Colorimetric Assay.

Hydrogen sulfide (H2S) is known to participate in bacteria-induced inflammatory response in periodontal diseases. Therefore, it is necessary to quantify H2S produced by oral bacteria for diagnosis and treatment of oral diseases including halitosis and periodontal disease. In this study, we introduce a paper-based colorimetric assay for detecting bacterial H2S utilizing silver/Nafion/polyvinylpyrrolidone membrane and a 96-well microplate. This H2S-sensing paper showed a good sensitivity (8.27 blue channel intensity/µM H2S, R2 = 0.9996), which was higher than that of lead acetate paper (6.05 blue channel intensity/µM H2S, R2 = 0.9959). We analyzed the difference in H2S concentration released from four kinds of oral bacteria (Eikenella corrodens, Streptococcus sobrinus, Streptococcus mutans, and Lactobacillus casei). Finally, the H2S level in Eikenella corrodens while varying the concentration of cysteine and treatment time was quantified. This paper-based colorimetric assay can be utilized as a simple and effective tool for in vitro screening of H2S-producing ability of many bacteria as well as salivary H2S analysis.

Phenethyl Isothiocyanate-Conjugated Chitosan Oligosaccharide Nanophotosensitizers for Photodynamic Treatment of Human Cancer Cells.

The aim of this study is to synthesize phenethyl-conjugated chitosan oligosaccharide (COS) (abbreviated as ChitoPEITC) conjugates and then fabricate chlorin E6 (Ce6)-incorporated nanophotosensitizers for photodynamic therapy (PDT) of HCT-116 colon carcinoma cells. PEITC was conjugated with the amine group of COS. Ce6-incorporated nanophotosensitizers using ChitoPEITC (ChitoPEITC nanophotosensitizers) were fabricated by dialysis method. 1H nuclear magnetic resonance (NMR) spectra showed that specific peaks of COS and PEITC were observed at ChitoPEITC conjugates. Transmission electron microscope (TEM) confirmed that ChitoPEITC nanophotosensitizers have spherical shapes with small hydrodynamic diameters less than 200 nm. The higher PEITC contents in the ChitoPEITC copolymer resulted in a slower release rate of Ce6 from nanophotosensitizers. Furthermore, the higher Ce6 contents resulted in a slower release rate of Ce6. In cell culture study, ChitoPEITC nanophotosensitizers showed low toxicity against normal CCD986Sk human skin fibroblast cells and HCT-116 human colon carcinoma cells in the absence of light irradiation. ChitoPEITC nanophotosensitizers showed a significantly higher Ce6 uptake ratio than that of free Ce6. Under light irradiation, cellular reactive oxygen species (ROS) production of nanophotosensitizers was significantly higher than that of free Ce6. Especially, PEITC and/or ChitoPEITC themselves contributed to the production of cellular ROS regardless of light irradiation. ChitoPEITC nanophotosensitizers showed significantly higher PDT efficacy against HCT-116 cells than that of free Ce6. These results indicate that ChitoPEITC nanophotosensitizers have superior potential in Ce6 uptake, ROS production and PDT efficacy. In the HCT-116 cell-bearing mice tumor-xenograft model, ChitoPEITC nanophotosensitizers efficiently inhibited growth of tumor volume rather than free Ce6. In the animal imaging study, ChitoPEITC nanophotosensitizers were concentrated in the tumor tissue, i.e., fluorescence intensity in the tumor tissue was stronger than that of other tissues. We suggest that ChitoPEITC nanophotosensitizers are a promising candidate for the treatment of human colon cancer cells.

Status of Helminthic Infections in Residents around River Basins in the Republic of Korea for 10 Years (2011-2020).

The positive rate of Clonorchis sinensis is the highest among intestinal parasites in the Republic of Korea (Korea). More than 1.2 million people were at risk of C. sinensis infection in Korea in 2012. An intensive control program is being implemented for residents of the 5 major river basins to reduce helminthic infections, including C. sinensis infection. This study evaluated the continuous intensive control program for parasitic diseases including clonorchiasis in areas near the 5 major river basins in Korea over the past 10 years (2011-2020). A total of 335,020 fecal samples (one sample per resident) prepared by the modified sedimentation technic were microscopically examined. Those who expelled helminth eggs were treated with anthelmintics through local health centers and re-examined 3 months later. The overall positive rate of helminths egg was 7.1%. The annual positive rates were dramatically decreased from 14.4% (2011) to 5.9% (2020). The egg positive rate was highest in C. sinensis (5.3%), followed by heterophyid flukes (1.5%) and Trichuris trichiura (0.2%). The prevalence of C. sinensis was significantly higher in males (7.6%) than in females (3.7%), and the highest in the 50-59 years (7.0%) age group. Our results are beneficial to establish prevention and control policies against helminthiases including clonorchiasis in endemic areas in this country.

Treatment with Extracellular Vesicles from Giardia lamblia Alleviates Dextran Sulfate Sodium-Induced Colitis in C57BL/6 Mice.

Inflammatory bowel disease (IBD) is a chronic and recurrent illness of the gastrointestinal tract. Treatment of IBD traditionally involves the use of aminosalicylic acid and steroids, while these drugs has been associated with untoward effects and refractoriness. The absence of effective treatment regimen against IBD has led to the exploration of new targets. Parasites are promising as an alternative therapy for IBD. Recent studies have highlighted the use of parasite-derived substances, such as excretory secretory products, extracellular vesicles (EVs), and exosomes, for the treatment of IBD. In this report, we examined whether EVs secreted by Giardia lamblia could prevent colitis in a mouse model. G. lamblia EVs (GlEVs) were prepared from in vitro cultures of Giardia trophozoites. Clinical signs, microscopic colon tissue inflammation, and cytokine expression levels were detected to assess the effect of GlEV treatment on dextran sulfate sodium (DSS)-induced experimental murine colitis. The administration of GlEVs prior to DSS challenge reduced the expression levels of pro-inflammatory cytokines, including tumor necrosis factor alpha, interleukin 1 beta, and interferon gamma. Our results indicate that GlEV can exert preventive effects and possess therapeutic properties against DSS-induced colitis.

Activation of NF-κB by TOPK upregulates Snail/Slug expression in TGF-ß1 signaling to induce epithelial-mesenchymal transition and invasion of breast cancer cells.

TGF-ß1 is known to induce epithelial-mesenchymal transition (EMT), which is a prerequisite for cancer cell invasion. Here we reveal that TOPK upregulates EMT and invasion of human breast cancer MDA-MB-231 or Hs578T cells via NF-κB-dependent Snail/Slug in TGF-ß1 signaling. Endogenous TOPK expression was significantly increased in response to TGF-ß1 and TOPK knockdown mitigated TGF-ß1-induced breast cancer cell invasion. Interestingly, TOPK knockdown restored TGF-ß1 suppression of E-cadherin expression and markedly reduced N-cadherin induced by TGF-ß1. Also, NF-κB activity or expression of EMT markers Snail and Slug induced by TGF-ß1 was decreased by TOPK knockdown. Meanwhile, knockdown of Snail or TOPK attenuated TGF-ß1-induced breast cancer cell invasion. Taken, we conclude that TOPK mediates TGF-ß1-induced EMT and invasion in breast cancer cells via NF-κB/Snail signaling, suggesting novel role of TOPK as therapeutic target in TGF-ß1-mediated breast cancer development.

TOPK promotes epithelial-mesenchymal transition and invasion of breast cancer cells through upregulation of TBX3 in TGF-ß1/Smad signaling.

TOPK has been suggested to contribute to invasion of lung, prostate, gastric, pancreatic or breast cancer cells. However, how TOPK mediates TGF-ß1/Smad signaling leading to epithelial-mesenchymal transition (EMT) and invasion of breast cancer cells remains unknown. Here we report that TOPK upregulates T-box transcription factor TBX3 to enhance TGF-ß1-induced EMT and invasion of MDA-MB-231 breast cancer cells. Expression of endogenous TOPK was promoted by TGF-ß1 treatment of MDA-MB-231 cells time-dependently. In addition, knockdown of TOPK attenuated TGF-ß1-induced phosphorylation or transcriptional activity of Smad3. Meanwhile, levels of both mRNA and protein of TBX3 induced by TGF-ß1 were abolished by TOPK depletion. Also, knockdown of TBX3 inhibited TGF-ß1 induction of EMT-related genes Snail, Slug or Fibronectin. Furthermore, ablation of TOPK or TBX3 suppressed TGF-ß1-induced MDA-MB-231 cell invasion. Collectively, we conclude that TOPK positively regulates TBX3 in TGF-ß1/Smad signaling pathway, thereby enhancing EMT and invasion of breast cancer cells, implying a mechanistic role of TOPK in TGF-ß1/Smad signaling.

Relationship between Breastfeeding, Birth History, and Acute Pyelonephritis in Infants.

BACKGROUND: Although the clinical importance of the immunological benefits of breastfeeding has been emphasized for decades, their direct relationship with acute pyelonephritis (APN) is still not clear. Our goal was to determine whether breastfeeding truly provides protection against APNs, while investigating the effects of other factors such as sex, age, mode of delivery, and birth weight on APN. METHODS: A total of 62 infants under 6 months of age who had both microbiologically and radiologically-confirmed APN were enrolled in the case group. Healthy infants (n = 178) who visited the hospital for scheduled vaccinations were enrolled in the control group. The following participant characteristics were compared between the case and control groups: age, sex, birth order among siblings, feeding methods, weight percentile by month, birth weight percentile by gestational age, gestational age at birth, and mode of delivery. RESULTS: Babies exclusively fed with manufactured infant formulae before 6 months of age had significantly higher risk for APN than breastfed or mixed-fed infants (odds ratio [OR], 3.4; 95% confidence interval [CI], 1.687-7.031; P = 0.001). Firstborn babies had lower risk for APN than 2nd- or 3rd-born babies (OR, 0.43; 95% CI, 0.210-0.919). Other factors that increased the risk for APN were low birth weight percentiles (OR, 8.33; 95% CI, 2.300-30.166) and birth via caesarean section (OR, 2.32; 95% CI, 1.097-4.887). There were more preterm births in the case group (10.9% vs. 1.7%; P = 0.002), but this did not increase the risk for APN (OR, 4.47; P = 0.063). CONCLUSION: Feeding exclusively with formula before 6 months of age was related to higher risk for APN, which demonstrates that breastfeeding has a protective effect against APN. The other risk factors for APN were birth order (≥ 2nd-born), low birth weight, and birth via caesarean section.

Optimization of Saliva Collection and Immunochromatographic Detection of Salivary Pepsin for Point-of-Care Testing of Laryngopharyngeal Reflux.

Salivary pepsin is a promising marker for the non-invasive diagnosis of laryngopharyngeal reflux (LPR). For reliable results regarding pepsin in saliva, it is critical to standardize the collection, storage, and pre-processing methods. In this study, we optimized the saliva collection protocols, including storage conditions, i.e., solution, temperature, and time, and the pre-processing filter for pepsin. Moreover, we prepared a simple immunochromatographic strip for the rapid detection of pepsin and evaluated its sensing performance. As a result, we selected a polypropylene (PP) filter as the pre-processing filter for salivary pepsin in low resource settings, such as those where point of care testing (POCT) is conducted. This filter showed a similar efficiency to the centrifuge (standard method). Finally, we detected the pepsin using gold nanoparticles conjugated with monoclonal pepsin antibody. Under optimized conditions, the lower limit of detection for pepsin test strips was determined as 0.01 µg/mL. Furthermore, we successfully detected the salivary pepsin in real saliva samples of LPR patients, which were pre-processed by the PP filter. Therefore, we expect that our saliva collection protocol and pepsin immunochromatographic strip can be utilized as useful tools for a non-invasive diagnosis/screening of LPR in POCT.

2-Phenylethylamine (PEA) Ameliorates Corticosterone-Induced Depression-Like Phenotype via the BDNF/TrkB/CREB Signaling Pathway.

Depression is a serious medical illness that is one of the most prevalent psychiatric disorders. Corticosterone (CORT) increases depression-like behavior, with some effects on anxiety-like behavior. 2-Phenethylamine (PEA) is a monoamine alkaloid that acts as a central nervous system stimulant in humans. Here, we show that PEA exerts antidepressant effects by modulating the Brain-derived neurotrophic factor (BDNF)/tropomyosin receptor kinase B (TrkB)/cAMP response element binding protein (CREB) signaling pathway in CORT-induced depression. To investigate the potential effects of PEA on CORT-induced depression, we first treated CORT (50 µM)-induced hippocampal neurons with 100 µM PEA for 24 h. We found that treatment with CORT altered dendritic spine architecture; however, treatment with PEA rescued dendritic spine formation via regulation of BDNF/TrkB/CREB signaling. Next, we used a mouse model of CORT-induced depression. Mice were treated with CORT (20 mg/kg) for 21 days, followed by assessments of a battery of depression-like behaviors. During the final four days of CORT exposure, the mice were treated with PEA (50 mg/kg). We found that CORT injection promoted depression-like behavior and significantly decreased BDNF and TrkB expression in the hippocampus. However, treatment with PEA significantly ameliorated the behavioral and biochemical changes induced by CORT. Our findings reveal that PEA exerts antidepressant effects by modulating the BDNF/TrkB/CREB signaling pathway in a mouse model of CORT-induced depression.

The Abuse Potential of Novel Synthetic Phencyclidine Derivative 1-(1-(4-Fluorophenyl)Cyclohexyl)Piperidine (4'-F-PCP) in Rodents.

The dissociative anesthetic phencyclidine (PCP) and PCP derivatives, including 4'-F-PCP, are illegally sold and abused worldwide for recreational and non-medical uses. The psychopharmacological properties and abuse potential of 4'-F-PCP have not been fully characterized. In this study, we evaluated the psychomotor, rewarding, and reinforcing properties of 4'-F-PCP using the open-field test, conditioned place preference (CPP), and self-administration paradigms in rodents. Using Western immunoblotting, we also investigated the expression of dopamine (DA)-related proteins and DA-receptor-mediated downstream signaling cascades in the nucleus accumbens (NAc) of 4'-F-PCP-self-administering rats. Intraperitoneal administration of 10 mg/kg 4'-F-PCP significantly increased locomotor and rearing activities and increased CPP in mice. Intravenous administration of 1.0 mg/kg/infusion of 4'-F-PCP significantly enhanced self-administration during a 2 h session under fixed ratio schedules, showed a higher breakpoint during a 6 h session under progressive ratio schedules of reinforcement, and significantly altered the expression of DA transporter and DA D1 receptor in the NAc of rats self-administering 1.0 mg/kg 4'-F-PCP. Additionally, the expression of phosphorylated (p) ERK, pCREB, c-Fos, and FosB/ΔFosB in the NAc was significantly enhanced by 1.0 mg/kg 4'-F-PCP self-administration. Taken together, these findings suggest that 4'-F-PCP has a high potential for abuse, given its robust psychomotor, rewarding, and reinforcing properties via activation of DAergic neurotransmission and the downstream signaling pathways in the NAc.

Development of an effective sample transfer device for biomarker detection in nasal secretions.

Nasal secretions (NS) reflect inflammatory activity of the nasal mucosa and thus can be utilized for disease diagnosis and determining treatment effects in Allergic rhinitis (AR). However, non-standardized collection of samples can affect the measured concentration of inflammatory biomarker in NS. In this study, we aimed to develop and evaluate new devices capable of standardizing the collection, storage, and preprocessing methods of NS samples. First, we chose the best swab as polyester (PE) and selected a stimulation method, twirling for 10 s at 1 Hz, to efficiently release AR biomarkers from a PE swab. Storage of sample solutions at -20 °C was optimal for the stability of biomarkers for the detection of AR. The new swab sample transfer device showed excellent concentration recovery efficiency (90-100%) for tryptase (Trp) and eosinophil cationic protein (ECP) without crosstalk between the two biomarkers. Finally, we compared the concentration of Trp in human NS samples of AR patients (n = 6) pre-processed by the new device with that by centrifuge as a standard method. As a result, the concentrations of Trp in NS were very similar in both groups. Therefore, this device can be utilized as an effective sample transfer and pre-processing device for point-of-care testing of AR.

Characterization of H2S releasing properties of various H2S donors utilizing microplate cover-based colorimetric assay.

In this study, we characterized the potential H2S-releasing properties of seven different H2S donors, including sodium sulfide (Na2S), sodium hydrosulfide (NaHS), diallyl disulfide (DADS), diallyl trisulfide (DATS), sodium thiosulfate (Na2S2O3), morpholin-4-ium 4-methoxyphenyl-morpholino-phosphinodithioate (GYY4137), and Lawesson's reagent, in three assay solutions, phosphate buffered saline (PBS, pH 7.4), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) buffered saline (HBS, pH 7.6), and cell growth media (GM), utilizing our microplate cover-based colorimetric assay. For quantitative analyses of H2S-releasing characteristics of the various donors, we evaluated four parameters, maximum concentration of H2S at the steady state (Cmax), the time required to reach half of Cmax (t1/2), maximum releasing rate of H2S (Rmax), and time at H2S (tr-max). The results showed that the H2S-releasing kinetics of each H2S donor were dependent on the type of assay solution. In particular, the addition of GSH to DATS in GM released the fastest and highest amounts of H2S among the four H2S donors in the following order: DATS > DADS > Na2S ~ NaHS. The H2S-releasing characteristics of the H2S donors were well-matched with cell viability results of human prostate cancer PC-3 cells. Therefore, the microplate cover-based colorimetric assay will be a useful tool for accurate and efficient measurements of H2S-releasing dynamics.