Inhibition of voltage-dependent K+ channels in rabbit coronary arterial smooth muscle cells by the atypical antipsychotic agent sertindole.

The regulation of membrane potential and the contractility of vascular smooth muscle cells (VSMCs) by voltage-dependent K+ (Kv) potassium channels are well-established. In this study, native VSMCs from rabbit coronary arteries were used to investigate the inhibitory effect of sertindole, an atypical antipsychotic agent, on Kv channels. Sertindole induced dose-dependent inhibition of Kv channels, with an IC50 of 3.13 ± 0.72 µM. Although sertindole did not cause a change in the steady-state activation curve, it did lead to a negative shift in the steady-state inactivation curve. The application of 1- or 2-Hz train pulses failed to alter the sertindole-induced inhibition of Kv channels, suggesting use-independent effects of the drug. The inhibitory response to sertindole was significantly diminished by pretreatment with a Kv1.5 inhibitor but not by Kv2.1 and Kv7 subtype inhibitors. These findings demonstrate the sertindole dose-dependent and use-independent inhibition of vascular Kv channels (mainly the Kv1.5 subtype) through a mechanism that involves altering steady-state inactivation curves. Therefore, the use of sertindole as an antipsychotic drug may have adverse effects on the cardiovascular system.

Second-generation antipsychotic quetiapine blocks voltage-dependent potassium channels in coronary arterial smooth muscle cells.

Voltage-dependent K+ (Kv) channels play an important role in restoring the membrane potential to its resting state, thereby maintaining vascular tone. In this study, native smooth muscle cells from rabbit coronary arteries were used to investigate the inhibitory effect of quetiapine, an atypical antipsychotic agent, on Kv channels. Quetiapine showed a concentration-dependent inhibition of Kv channels, with an IC50 of 47.98 ± 9.46 µM. Although quetiapine (50 µM) did not alter the steady-state activation curve, it caused a negative shift in the steady-state inactivation curve. The application of 1 and 2 Hz train steps in the presence of quetiapine significantly increased the inhibition of Kv current. Moreover, the recovery time constants from inactivation were prolonged in the presence of quetiapine, suggesting that its inhibitory action on Kv channels is use (state)-dependent. The inhibitory effects of quetiapine were not significantly affected by pretreatment with Kv1.5, Kv2.1, and Kv7 subtype inhibitors. Based on these findings, we conclude that quetiapine inhibits Kv channels in both a concentration- and use (state)-dependent manner. Given the physiological significance of Kv channels, caution is advised in the use of quetiapine as an antipsychotic due to its potential side effects on cardiovascular Kv channels.

Prevalence and Genetic Characterization of Porcine Respiratory Coronavirus in Korean Pig Farms.

Porcine respiratory coronavirus (PRCV) is a member of the species Alphacoronavirus 1 within the genus Alphacoronavirus of the family Coronaviridae. A few studies have been conducted on the prevalence of PRCV since its first identification in 1997, but there have been no recent studies on the prevalence and genetic characterization of the virus in Korea. In this study, the seroprevalence of PRCV was determined in Korean pig farms using a commercially available TGEV/PRCV differential enzyme-linked immunosorbent assay kit. The farm-level seroprevalence of PRCV was determined to be 68.6% (48/70), similar to previous reports in Korea, suggesting that PRCV is still circulating in Korean pig herds nationwide. Among the 20 PRCV-seropositive farms tested in this study, PRCV RNAs were detected in 17 oral fluid samples (28.3%) from nine farms (45.0%), while TGEV RNAs were not detected in any sample. To investigate the genetic characteristics of Korean PRCV strains, genetic and phylogenetic analyses were conducted on PRCV spike gene sequences obtained in this study. The three Korean PRCV strains (KPRCV2401, KPRCV2402, and KPRCV2403) shared 98.5-100% homology with each other and 96.2-96.6% and 91.6-94.5% homology with European and American strains, respectively. A 224-amino acid deletion was found in the S gene of both Korean and European PRCVs but not in that of American PRCVs, suggesting a European origin for Korean PRCVs. Phylogenetic analysis showed that Korean PRCVs are more closely related to European PRCVs than American PRCVs but clustered apart from both, suggesting that Korean PRCV has evolved independently since its emergence in Korean PRCVs. The results of this study will help expand knowledge on the epidemiology and molecular biology of PRCV currently circulating in Korea.

The antidiabetic drug ipragliflozin induces vasorelaxation of rabbit femoral artery by activating a Kv channel, the SERCA pump, and the PKA signaling pathway.

We explored the vasorelaxant effects of ipragliflozin, a sodium-glucose cotransporter-2 inhibitor, on rabbit femoral arterial rings. Ipragliflozin relaxed phenylephrine-induced pre-contracted rings in a dose-dependent manner. Pre-treatment with the ATP-sensitive K+ channel inhibitor glibenclamide (10 µM), the inwardly rectifying K+ channel inhibitor Ba2+ (50 µM), or the Ca2+-sensitive K+ channel inhibitor paxilline (10 µM) did not influence the vasorelaxant effect. However, the voltage-dependent K+ (Kv) channel inhibitor 4-aminopyridine (3 mM) reduced the vasorelaxant effect. Specifically, the vasorelaxant response to ipragliflozin was significantly attenuated by pretreatment with the Kv7.X channel inhibitors linopirdine (10 µM) and XE991 (10 µM), the sarcoplasmic/endoplasmic reticulum Ca2+-ATPase (SERCA) pump inhibitors thapsigargin (1 µM) and cyclopiazonic acid (10 µM), and the cAMP/protein kinase A (PKA)-associated signaling pathway inhibitors SQ22536 (50 µM) and KT5720 (1 µM). Neither the cGMP/protein kinase G (PKG)-associated signaling pathway nor the endothelium was involved in ipragliflozin-induced vasorelaxation. We conclude that ipragliflozin induced vasorelaxation of rabbit femoral arteries by activating Kv channels (principally the Kv7.X channel), the SERCA pump, and the cAMP/PKA-associated signaling pathway independent of other K+ (ATP-sensitive K+, inwardly rectifying K+, and Ca2+-sensitive K+) channels, cGMP/PKG-associated signaling, and the endothelium.

Molecular Detection of Porcine Parainfluenza Viruses 1 and 5 Using a Newly Developed Duplex Real-Time RT-PCR in South Korea.

Two species of porcine parainfluenza viruses (PPIV), PPIV1 and PPIV5, are globally distributed in pig herds and associated with porcine respiratory diseases, and a diagnostic tool for the simultaneous detection of the two viruses is required. In this study, a TaqMan probe-based duplex real-time reverse transcription polymerase chain reaction (dqRT-PCR) assay was first developed for the differential detection of PPIV1 and PPIV5 nucleocapsid protein (NP) genes in porcine clinical samples. The dqRT-PCR assay was highly sensitive, its limit of detection was approximately 10 RNA copies/reaction, it specifically amplified the targeted NP genes of PPIV1 and PPIV5 without cross-reacting with other porcine pathogens, and their clinical detection rates were 15.2% and 0.7%, respectively. The results from 441 clinical samples taken from 278 Korean domestic pig farms showed that the prevalence of PPIV1 and PPIV5 was 11.2% and 1.1%, respectively, and co-infection of both viruses was confirmed in a farm, suggesting that PPIV1 and PPIV5 are co-circulating in current Korean pig herds. Phylogenetic analysis based on the partial NP genes suggested that genetically diverse PPIV1 strains are circulating in Korean pig herds. The developed dqRT-PCR assay was found to be an accurate, reliable, and quantitative detection tool for PPIV1 and PPIV5 RNA in clinical pig samples and will be useful for etiological and epidemiological studies and the control of viral infections in the field.

An Improved Duplex Real-Time Quantitative RT-PCR Assay with a Canine Endogenous Internal Positive Control for More Sensitive and Reliable Detection of Canine Parainfluenza Virus 5.

A duplex real-time quantitative reverse transcription-polymerase chain reaction (dqRT-PCR) assay was successfully developed to simultaneously detect canine parainfluenza virus 5 (CPIV5) and a canine endogenous internal positive control (EIPC) in canine clinical samples. Two sets of primers and probes for the CPIV5 L and canine 16S rRNA genes were included in the dqRT-PCR assay to detect CPIV and monitor invalid results throughout the qRT-PCR process. The developed dqRT-PCR assay specifically detected CPIV5 but no other canine pathogens. Furthermore, 16S rRNA was stably amplified by dqRT-PCR assay in all samples containing canine cellular materials. The assay's sensitivity was determined as below ten RNA copies per reaction, with CPIV5 L gene standard RNA and 1 TCID50/mL with the CPIV5 D008 vaccine strain, which was 10-fold higher than that of the previous HN gene-specific qRT-PCR (HN-qRT-PCR) assays and was equivalent to that of the previous N gene-specific qRT-PCR (N-qRT-PCR) assays, respectively. Moreover, the Ct values of the CPIV5-positive samples obtained using the dqRT-PCR assay were lower than those obtained using the previous HN- and N-qRT-PCR assays, indicating that the diagnostic performance of the dqRT-PCR assay was superior to those of previous HN- and N-qRT-PCR assays. The calculated Cohen's kappa coefficient values (95% confidence interval) between dqRT-PCR and the HN- or N-specific qRT-PCR assays were 0.97 (0.90-1.03) or 1.00 (1.00-1.00), respectively. In conclusion, the newly developed dqRT-PCR assay with high sensitivity, specificity, and reliability will be a promising diagnostic tool for the detection of CPIV5 in clinical samples and useful for etiological and epidemiological studies of CPIV5 infection in dogs.

Simple and Rapid Colorimetric Detection of Canine Parainfluenza Virus 5 (Orthorubulavirus mammalis) Using a Reverse-Transcription Loop-Mediated Isothermal Amplification Assay.

Despite its many advantages, a reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay has yet to be developed for canine parainfluenza virus 5 (CPIV5). In this study, a visual RT-LAMP (vRT-LAMP) assay was developed for the rapid detection of CPIV5 in clinical samples. At a constant reaction temperature of 62 °C, the assay was completed within 40 min, and the results could be directly detected with the naked eye using a hydroxynaphthol blue (HNB) metal indicator without any additional detection apparatuses. The assay specifically amplified CPIV5 RNA with a limit of detection of 10 RNA copies/reaction, which was 10-fold more sensitive than the previously reported conventional reverse-transcription polymerase chain reaction (cRT-PCR) assay and was comparable to the previously reported real-time RT-PCR (qRT-PCR) assay. In a clinical evaluation using 267 nasopharyngeal swab samples collected from hospitalized dogs with respiratory symptoms, the CPIV5 detection rate using the vRT-LAMP assay was 5.24% (14/267), which was higher than that of the cRT-PCR assay (4.49%, 12/267) and consistent with that of the qRT-PCR assay, demonstrating 100% concordance with a kappa coefficient value (95% confidence interval) of 1 (1.00-1.00). The discrepancies in the results of the assays were confirmed to be attributed to the low sensitivity of the cRT-PCR assay. Owing to the advantages of a high specificity, rapidity, and simplicity, the developed vRT-LAMP assay using an HNB metal indicator will be a valuable diagnostic tool for the detection of CPIV5 in canine clinical samples, even in resource-limited laboratories.

Genetic Characterization and Evolution of Porcine Deltacoronavirus Isolated in the Republic of Korea in 2022.

Porcine deltacoronavirus (PDCoV) is an emerging coronavirus that causes diarrhea in nursing piglets. Since its first outbreak in the United States in 2014, this novel porcine coronavirus has been detected worldwide, including in Korea. However, no PDCoV case has been reported since the last report in 2016 in Korea. In June 2022, the Korean PDCoV strain KPDCoV-2201 was detected on a farm where sows and piglets had black tarry and watery diarrhea, respectively. We isolated the KPDCoV-2201 strain from the intestinal samples of piglets and sequenced the viral genome. Genetically, the full-length genome and spike gene of KPDCoV-2201 shared 96.9-99.2% and 95.8-98.8% nucleotide identity with other global PDCoV strains, respectively. Phylogenetic analysis suggested that KPDCoV-2201 belongs to G1b. Notably, the molecular evolutionary analysis indicated that KPDCoV-2201 evolved from a clade different from that of previously reported Korean PDCoV strains and is closely related to the emergent Peruvian and Taiwanese PDCoV strains. Furthermore, KPDCoV-2201 had one unique and two Taiwanese strain-like amino acid substitutions in the receptor-binding domain of the S1 region. Our findings suggest the possibility of transboundary transmission of the virus and expand our knowledge about the genetic diversity and evolution of PDCoV in Korea.

Tailored Multiplex Real-Time RT-PCR with Species-Specific Internal Positive Controls for Detecting SARS-CoV-2 in Canine and Feline Clinical Samples.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections have been frequently reported in companion dogs and cats worldwide during the ongoing coronavirus disease. However, RT-qPCR methods developed for humans have been used for the diagnosis of SARS-CoV-2 infections in suspected companion dogs and cats owing to the lack of the companion animal-tailored methods. Therefore, we developed a multiplex RT-qPCR (mRT-qPCR) using newly designed primers and probes targeting RdRp and N genes of all currently circulating SARS-CoV-2 variants as well as the canine or feline 16S rRNA gene as an endogenous internal positive control (EIPC) for reliable diagnosis of SARS-CoV-2 infection from suspected dogs and cats. The developed mRT-qPCR assay specifically detected the target genes of SARS-CoV-2 but no other canine or feline pathogens. Furthermore, canine and feline EIPCs were stably amplified by mRT-qPCR in samples containing canine- or feline-origin cellular materials. This assay has high repeatability and reproducibility, with an optimal limit of detection (<10 RNA copies per reaction) and coefficients of variation (<1.0%). The detection rate of SARS-CoV-2 of the developed mRT-qPCR was 6.6% for canine and feline nasopharyngeal samples, which was consistent with that of a commercial mRT-qPCR kit for humans. Collectively, the newly developed mRT-qPCR with canine and feline EIPC can efficiently diagnose and evaluate the viral load in field specimens and will be a valuable tool for etiological diagnosis, epidemiological study, and controlling SARS-CoV-2 infections in canine and feline populations.

An Advanced Multiplex Real-Time Reverse Transcription Loop-Mediated Isothermal Amplification Assay for Rapid and Reliable Detection of Porcine Epidemic Diarrhea Virus and Porcine Internal Positive Control.

For rapid and reliable detection of porcine epidemic diarrhea virus (PEDV) from pig clinical samples, a multiplex, real-time, reverse transcription loop-mediated isothermal amplification (mqRT-LAMP) was developed using two sets of primers and assimilating probes specific to the PEDV N gene and the Sus scrofa ß-actin gene, which was used as an endogenous internal positive control (EIPC) to avoid false-negative results. The assay specifically amplified both target genes of PEDV and EIPC in a single reaction without any interference but did not amplify other porcine viral nucleic acids. The limit of detection was 10 copies/µL, 100-fold lower than that of a reverse transcription-polymerase chain reaction (RT-PCR) and equivalent to that of quantitative/real-time RT-PCR (qRT-PCR). This assay has high repeatability and reproducibility with coefficients of variation < 4.0%. The positive signal of the mqRT-LAMP assay was generated within 25 min, demonstrating advantages in rapid detection of PEDV over RT-PCR or qRT-PCR assay, which require at least 2 h turnaround times. In clinical evaluation, the detection rate of PEDV by mqRT-LAMP assay (77.3%) was higher than that of RT-PCR assay (69.7%), and comparable to qRT-PCR (76.8%) with almost 100% concordance (kappa value 0.98). The developed mqRT-LAMP assay can serve as an advanced alternative method for PEDV diagnosis because it has high sensitivity and specificity, rapidity, and reliability even in resource-limited laboratories.

First Detection and Genetic Characterization of Swine Orthopneumovirus from Domestic Pig Farms in the Republic of Korea.

Novel swine orthopneumovirus (SOV) infections have been identified in pigs in the USA and some European countries but not in Asian countries, including South Korea, to date. The current study reports the first SOV infections in four domestic pig farms located in four provinces across South Korea. The detection rate of SOV in oral fluid samples using qRT-PCR was 4.4% (14/389), indicating the presence of the virus in pigs at commercial farms in Korea. Two complete genome sequences and one glycoprotein (G) gene sequence were obtained from SOV-positive samples. The complete genome analysis of KSOV-2201 and KSOV-2202 strains showed 98.2 and 95.4% homologies with a previously reported SOV, and the phylogenetic tree exhibited a high correlation with a previously reported SOV strain from the US and a canine pneumovirus (CPnV) strain from China. Based on the genetic analysis of the viral G gene, the murine pneumonia virus (MPV)-like orthopneumoviruses (MLOVs) were divided into two genogroups (G1 and G2). Seventeen CPnVs and two feline pneumoviruses were grouped into G1, while the Korean SOV strains identified in this study were grouped into G2 along with one SOV and two CPnVs. These results will contribute to expanding our understanding of the geographical distribution and genetic characteristics of the novel SOV in the global pig population.

Epidemiological and Molecular Approaches for a Fatal Feline Panleukopenia Virus Infection of Captive Siberian Tigers (Panthera tigris altaica) in the Republic of Korea.

Feline panleukopenia virus (FPV), a member of the species Protoparvovirus carnivoran1, is one of the most fatal pathogens of domestic and wild carnivores. The virus endemically infects domestic carnivores worldwide and its cross-species transmission threatens endangered wild carnivores, including Siberian tigers. In this study, a fatal FPV infection in endangered Siberian tigers was investigated to trace the origin of the virus and elucidate the reason behind FPV's infection of the vaccinated tigers. Our genetic characterization and phylogenetic analysis revealed that the virus detected in the infected tigers, designated as the KTPV-2305 strain, was closely related to FPV strains circulating in Korean cats, suggesting that it might have been transmitted from stray cats wandering around the zoo. Compared with the prototype FPV reference strains, the KTPV-2305 strain carried three distinct amino acid (aa) mutations in the VP2 protein sequence (I101T, I232V, and L562V) in this study. These three mutations are commonly found in most global FPV strains, including Korean strains, indicating that these mutations are common evolutionary characteristics of currently circulating global FPVs. The reason why the vaccinated tigers were infected with FPV was most likely the insufficient protective immunity of the affected tigress or vaccine failure triggered by the interference of maternal-derived antibodies in the affected tiger cubs. These findings suggest that improved vaccination guidelines are urgently needed to save the lives of wild carnivores from this fatal virus.

Cell death is induced by ciglitazone, a peroxisome proliferator-activated receptor γ (PPARγ) agonist, independently of PPARγ in human glioma cells.

Peroxisome proliferator-activated receptor γ (PPARγ) regulates multiple signaling pathways, and its agonists induce apoptosis in various cancer cells. However, their role in cell death is unclear. In this study, the relationship between ciglitazone (CGZ) and PPARγ in CGZ-induced cell death was examined. At concentrations of greater than 30 µM, CGZ, a synthetic PPARγ agonist, activated caspase-3 and induced apoptosis in T98G cells. Treatment of T98G cells with less than 30 µM CGZ effectively induced cell death after pretreatment with 30 µM of the PPARγ antagonist GW9662, although GW9662 alone did not induce cell death. This cell death was also observed when cells were co-treated with CGZ and GW9662, but was not observed when cells were treated with CGZ prior to GW9662. In cells in which PPARγ was down-regulated cells by siRNA, lower concentrations of CGZ (<30 µM) were sufficient to induce cell death, although higher concentrations of CGZ (≥30 µM) were required to induce cell death in control T98G cells, indicating that CGZ effectively induces cell death in T98G cells independently of PPARγ. Treatment with GW9662 followed by CGZ resulted in a down-regulation of Akt activity and the loss of mitochondrial membrane potential (MMP), which was accompanied by a decrease in Bcl-2 expression and an increase in Bid cleavage. These data suggest that CGZ is capable of inducing apoptotic cell death independently of PPARγ in glioma cells, by down-regulating Akt activity and inducing MMP collapse.

Pathogenesis and genetic characteristics of low pathogenic avian influenza H10 viruses isolated from migratory birds in South Korea during 2010-2019.

Human infection by avian-origin subtype H10 influenza viruses has raised concerns about the pandemic potential of these microbes. H10 subtype low pathogenic avian influenza viruses (LPAIVs) have been isolated from wild birds and poultry worldwide. Here, we isolated 36 H10 LPAIVs from wild bird habitats (a mean annual rate of 3.8% of all avian influenza virus isolations) from January 2010 to April 2019 through a nationwide active surveillance program for avian influenza viruses (AIVs). Phylogenetic analysis revealed that the haemagglutinin (HA) gene of H10 isolates formed eight distinct genetic subgroups (HA-A-H). Unlike other Eurasian-origin subgroups, the HA-H subgroup belonged to the North American lineage. Gene-constellation analysis revealed that 24 H10 LPAIVs constituted ≥18 distinct genotypes, representing high levels of genetic diversity. An intravenous pathogenicity index (IVPI) experiment showed that the pathogenicity of representative strains of the HA-B, E and G subgroups possessing an IVPI score >1.2 was associated with replication capacity in the chicken kidney in the absence of trypsin. Intranasal inoculation experiments showed that a representative strain of the HA-D subgroup replicated and transmitted in chickens without clinical signs. Subclinical virus shedding in chickens may contribute to its silent spread among the poultry population. Moreover, six representative viruses replicated in the lungs of mice without prior adaptation and a representative strain of the HA-C subgroup caused 40% mortality, with severe body weight loss. These findings highlight the importance of intensive surveillance of wild bird habitats, poultry farms and the animal-human interface, along with appropriate risk assessment of isolated viruses.

Detection of a novel porcine circovirus 4 in Korean pig herds using a loop-mediated isothermal amplification assay.

A novel porcine circovirus 4 has been recently identified in China and Korea. A sensitive and specific diagnostic method is urgently required to detect the virus in field samples. We developed a loop-mediated isothermal amplification (LAMP) the assay for the visual detection of PCV4 and evaluated its sensitivity, specificity, and applicability in clinical samples. This assay's results can be directly visualized by the naked eye using hydroxynaphthol blue after incubation for 40 min at 64 °C. The assay specifically amplified PCV4 DNA and no other viral nucleic acids. The sensitivity of the assay was <50 DNA copies/reaction, which was 10 times more sensitive than conventional polymerase chain reaction (cPCR) and comparable to real-time PCR (qPCR). Clinical evaluation revealed that the PCV4 detection rate in individual pig samples and at the farm level was 39.3 % (57/145) and 45.7 % (32/70), respectively, which were higher than cPCR (46 samples, 24 farms) and qPCR (52 samples, 29 farms) results. Cumulatively, owing to the advantages of high sensitivity and specificity, direct visual monitoring of the results, no possibility for cross-contamination, and being a low-cost equipment, the developed LAMP assay will be a valuable tool for the detection of the novel PCV4 in clinical samples, even in resource-limited laboratories.

First report of Porcine respirovirus 1 in South Korea.

Porcine respirovirus 1 (PRV1) is a recently emerging porcine respiratory virus that belongs to the genus Respirovirus of the Paramyxoviridae family. Since its first detection in Hong Kong, China in 2009, PRV1 has been subsequently identified in several American and European countries, suggesting that the emerging virus may have been globally distributed. However, in Asia, the virus has been reported only in China. Here, we report that PRV1 was first detected in pigs from 16 farms located in seven provinces across Korea, with a prevalence of 71.4% based on the tested oral fluid samples, suggesting that the virus is already widespread in Korean pig herds. For further genetic characterization of the Korean PRV1 strains, a complete genome and two F gene sequences were obtained from PRV1-positive samples collected from three different pig farms. Phylogenetic analysis based on the complete genome and F gene sequences showed that all three Korean PRV1 strains were grouped into European lineage 1 and were closely related to strains from Hong Kong (China), Germany and Poland. We could not obtain evidence for the origin of Korean PRV1 because of the limited availability of PRV1 sequences. In conclusion, PRV1 was first identified in Korean pig herds and genetically characterized in the present study. These results contribute to a better understanding of the global geographical distribution and genetic characteristics of PRV1.

Correlation between chemokines released from umbilical cord blood-derived mesenchymal stem cells and engraftment of hematopoietic stem cells in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice.

Umbilical cord blood (UCB)-derived mesenchymal stem cells (MSCs) enhance the engraftment of human hematopoietic stem cells (HSCs) when they are cotransplanted in animal and human studies. However, the type of MSCs that preferentially facilitate the engraftment and homing of HSCs is largely unknown. The authors categorized UCB-MSCs as the least-effective MSCs (A) or most-effective MSCs (B) at enhancing the engraftment of HSCs, and compared the gene expression profiles of various cytokines and growth factors in the UCB-MSC populations. The most-effective UCB-MSCs (B) secreted higher levels of several factors, including chemokine (C-X-C motif) ligand 12 (CXCL12), regulated upon activation, normal T cells expressed and secreted (RANTES), epithelial growth factor (EGF), and stem cell factor (SCF), which are required for the engraftment and homing of HSCs. By contrast, levels of growth-related oncogene (GRO), insulin-like growth factor-binding protein 1 (IGFBP1), and interleukin-8 (IL-8), which are associated with immune inflammation, were secreted at higher levels in UCB-MSCs (A). In addition, there were no differences between the transcripts of the 2 UCB-MSC populations after interferon-gamma (IFN-γ) stimulation, except for cyclooxygenase (COX)-1. Based on these findings, the authors propose that these chemokines may be useful for modulating these cells in a clinical setting and potentially for enhancing the effectiveness of the engraftment and homing of HSCs.

A simple colorimetric detection of porcine epidemic diarrhea virus by reverse transcription loop-mediated isothermal amplification assay using hydroxynaphthol blue metal indicator.

A simple reverse transcription loop-mediated isothermal amplification combined with visual detection method (vRT-LAMP) assay was developed for rapid and specific detection of porcine epidemic diarrhea virus (PEDV) in this study, which overcomes the shortcomings of previously described RT-LAMP assays that require additional detection steps or pose a risk of cross-contamination. The assay results can be directly detected by the naked eye using hydroxynaphthol blue after incubating for 40 min at 62 °C. The assay specifically amplified PEDV RNA and no other viral nucleic acids. The limit of detection of the assay was less than 50 RNA copies per reaction, which was 100 times more sensitive than conventional reverse transcription polymerase chain reaction (RT-PCR) and comparable to real-time RT-PCR (RRT-PCR). In the clinical evaluation, the PEDV detection rate of vRT-LAMP was higher than that of RRT-PCR, showing 99 % concordance, with a kappa value (95 % confidence interval) of 0.97 (0.93-1.01). Considering the advantages of high sensitivity and specificity, simple and direct visual monitoring of the results, no possibility for cross-contamination, and being able to be used as low-cost equipment, the developed vRT-LAMP assay will be a valuable tool for detecting PEDV from clinical samples, even in resource-limited laboratories.

A nonthiazolidinedione peroxisome proliferator-activated receptor α/γ dual agonist CG301360 alleviates insulin resistance and lipid dysregulation in db/db mice.

Activation of peroxisome proliferator-activated receptors (PPARs) have been implicated in the treatment of metabolic disorders with different mechanisms; PPARα agonists promote fatty acid oxidation and reduce hyperlipidemia, whereas PPARγ agonists regulate lipid redistribution from visceral fat to subcutaneous fat and enhance insulin sensitivity. To achieve combined benefits from activated PPARs on lipid metabolism and insulin sensitivity, a number of PPARα/γ dual agonists have been developed. However, several adverse effects such as weight gain and organ failure of PPARα/γ dual agonists have been reported. By use of virtual ligand screening, we identified and characterized a novel PPARα/γ dual agonist, (R)-1-(4-(2-(5-methyl-2-p-tolyloxazol-4-yl)ethoxy)benzyl)piperidine-2-carboxylic acid (CG301360), exhibiting the improvement in insulin sensitivity and lipid metabolism. CG301360 selectively stimulated transcriptional activities of PPARα and PPARγ and induced expression of their target genes in a PPARα- and PPARγ-dependent manner. In cultured cells, CG301360 enhanced fatty acid oxidation and glucose uptake and it reduced pro-inflammatory gene expression. In db/db mice, CG301360 also restored insulin sensitivity and lipid homeostasis. Collectively, these data suggest that CG301360 would be a novel PPARα/γ agonist, which might be a potential lead compound to develop against insulin resistance and hyperlipidemia.

An advanced loop-mediated isothermal amplification assay for the rapid detection of beak and feather disease virus in psittacine birds.

Swarm primer-applied loop-mediated isothermal amplification (sLAMP) assay was developed for the rapid and specific detection of the ORF V1 gene of beak and feather disease virus (BFDV). The amplification can be completed in 40 min at 62 °C, and the results can be visually detected by the naked eye. The assay specifically amplified BFDV DNA and not amplified other viral nucleic acids. The limit of detection of the assay was 5 × 102 DNA copies/reaction, which was lower than that of the previously described LAMP (preLAMP) assay and comparable to that of a previously reported real time quantitative PCR (qPCR) assay. The detection rates of BFDV from psittacine clinical samples by the sLAMP, qPCR and preLAMP assays were 36.0 %, 36.0 % and 25.6 %, respectively, and the sLAMP results showed 100.0 % concordance with the qPCR results with a kappa value of 1.0. On the other hand, the preLAMP assay did not detect nine out of the 31 samples that were positive by sLAMP and qPCR assays, probably due to low sensitivity of the assay. These data suggest that the newly developed sLAMP assay will be a valuable tool for the rapid, sensitive, specific and reliable detection of BFDV in suspected psittacine birds, even in resource-limited laboratories.