Antimicrobial resistance profile in Salmonella enterica serovar Choleraesuis isolates from diseased pigs in Taiwan.

This study investigated antimicrobial resistance in Salmonella enterica serovar Choleraesuis (S. Choleraesuis) isolates from diseased pigs in Taiwan (2015-2020). Among 272 isolates, florfenicol (96.7%), enrofloxacin (96.3%), doxycycline (91.2%), gentamicin (84.6%), and tiamulin (80.5%) exhibited high resistance. 99.3% of the isolates were resistant to at least one antibiotic, and 97.8% of the isolates were multidrug resistant. This study illustrated that S. Choleraesuis isolates exhibited high resistance to antimicrobials currently used in the Taiwanese swine industry.

Implementation of point-of-care platforms for rapid detection of porcine circovirus type 2.

BACKGROUND: Porcine circovirus type 2 (PCV2) infection is ubiquitous around the world. Diagnosis of the porcine circovirus-associated disease requires clinic-pathological elements together with the quantification of viral loads. Furthermore, given pig farms in regions lacking access to sufficient laboratory equipment, developing diagnostic devices with high accuracy, accessibility, and affordability is a necessity. OBJECTIVES: This study aims to investigate two newly developed diagnostic tools that may satisfy these criteria. METHODS: We collected 250 specimens, including 170 PCV2-positive and 80 PCV2-negative samples. The standard diagnosis and cycle threshold (Ct) values were determined by quantitative polymerase chain reaction (qPCR). Then, two point-of-care (POC) diagnostic platforms, convective polymerase chain reaction (cPCR, qualitative assay: positive or negative results are shown) and EZtargex (quantitative assay: Ct values are shown), were examined and analyzed. RESULTS: The sensitivity and specificity of cPCR were 88.23% and 100%, respectively; the sensitivity and specificity of EZtargex were 87.65% and 100%, respectively. These assays also showed excellent concordance compared with the qPCR assay (κ = 0.828 for cPCR and κ = 0.820 for EZtargex). The statistical analysis showed a great diagnostic power of the EZtargex assay to discriminate between samples with different levels of positivity. CONCLUSIONS: The two point-of-care diagnostic platforms are accurate, rapid, convenient and require little training for PCV2 diagnosis. These POC platforms can discriminate viral loads to predict the clinical status of the animals. The current study provided evidence that these diagnostics were applicable with high sensitivity and specificity in the diagnosis of PCV2 infection in the field.

Antimicrobial susceptibility and resistome of Actinobacillus pleuropneumoniae in Taiwan: a next-generation sequencing analysis.

Actinobacillus pleuropneumoniae infection causes a high mortality rate in porcine animals. Antimicrobial resistance poses global threats to public health. The current study aimed to determine the antimicrobial susceptibilities and probe the resistome of A. pleuropneumoniae in Taiwan. Herein, 133 isolates were retrospectively collected; upon initial screening, 38 samples were subjected to next-generation sequencing (NGS). Over the period 2017-2022, the lowest frequencies of resistant isolates were found for ceftiofur, cephalexin, cephalothin, and enrofloxacin, while the highest frequencies of resistant isolates were found for oxytetracycline, streptomycin, doxycycline, ampicillin, amoxicillin, kanamycin, and florfenicol. Furthermore, most isolates (71.4%) showed multiple drug resistance. NGS-based resistome analysis revealed aminoglycoside- and tetracycline-related genes at the highest prevalence, followed by genes related to beta-lactam, sulfamethoxazole, florphenicol, and macrolide. A plasmid replicon (repUS47) and insertion sequences (IS10R and ISVAp11) were identified in resistant isolates. Notably, the multiple resistance roles of the insertion sequence IS10R were widely proposed in human medicine; however, this is the first time IS10R has been reported in veterinary medicine. Concordance analysis revealed a high consistency of phenotypic and genotypic susceptibility to florphenicol, tilmicosin, doxycycline, and oxytetracycline. The current study reports the antimicrobial characterization of A. pleuropneumoniae for the first time in Taiwan using NGS.

Experimental study of modified Tavis-Cummings model with directly-coupled superconducting artificial atoms.

The Tavis-Cummings model is intensively investigated in quantum optics and has important applications in generation of multi-atom entanglement. Here, we employ a superconducting circuit quantum electrodynamic system to study a modified Tavis-Cummings model with directly-coupled atoms. In our device, three superconducting artificial atoms are arranged in a chain with direct coupling through fixed capacitors and strongly coupled to a transmission line resonator. By performing transmission spectrum measurements, we observe different anticrossing structures when one or two qubits are resonantly coupled to the resonator. In the case of the two-qubit Tavis-Cummings model without qubit-qubit interaction, we observe two dips at the resonance point of the anticrossing. The splitting of these dips is determined by Δ λ=2g12+g32, where g1 and g3 are the coupling strengths between Qubit 1 and the resonator, and Qubit 3 and the resonator, respectively. The direct coupling J12 between the two qubits results in three dressed states in the two-qubit Tavis-Cummings model at the frequency resonance point, leading to three dips in the transmission spectrum. In this case, the distance between the two farthest and asymmetrical dips, arising from the energy level splitting, is larger than in the previous case. The frequency interval between these two dips is determined by the difference in eigenvalues (Δ λ=ε 1+-ε 1-), obtained through numerical calculations. What we believe as novel and intriguing experimental results may potentially advance quantum optics experiments, providing valuable insights for future research.

Clinical treatment of cryptococcal meningitis: an evidence-based review on the emerging clinical data.

Cryptococcal meningitis (CM) is a fatal fungal central nervous system (CNS) infection caused by Cryptococcus infecting the meninges and/or brain parenchyma, with fever, headache, neck stiffness, and visual disturbances as the primary clinical manifestations. Immunocompromised individuals with human immunodeficiency virus (HIV) infection or who have undergone organ transplantation, as well as immunocompetent people can both be susceptible to CM. Without treatment, patients with CM may have a mortality rate of up to 100% after hospital admission. Even after receiving therapy, CM patients may still suffer from problems such as difficulty to cure, poor prognosis, and high mortality. Therefore, timely and effective treatment is essential to improve the mortality and prognosis of CM patients. Currently, the clinical outcomes of CM are frequently unsatisfactory due to limited drug choices, severe adverse reactions, drug resistance, etc. Here, we review the research progress of CM treatment strategies and discuss the suitable options for managing CM, hoping to provide a reference for physicians to select the most appropriate treatment regimens for CM patients.

Study on injectable chitosan hydrogel with tendon-derived stem cells for enhancing rotator cuff tendon-to-bone healing / 中国修复重建外科杂志

OBJECTIVE@#To explore the effect of chitosan (CS) hydrogel loaded with tendon-derived stem cells (TDSCs; hereinafter referred to as TDSCs/CS hydrogel) on tendon-to-bone healing after rotator cuff repair in rabbits.@*METHODS@#TDSCs were isolated from the rotator cuff tissue of 3 adult New Zealand white rabbits by Henderson step-by-step enzymatic digestion method and identified by multidirectional differentiation and flow cytometry. The 3rd generation TDSCs were encapsulated in CS to construct TDSCs/CS hydrogel. The cell counting kit 8 (CCK-8) assay was used to detect the proliferation of TDSCs in the hydrogel after 1-5 days of culture in vitro, and cell compatibility of TDSCs/CS hydrogel was evaluated by using TDSCs alone as control. Another 36 adult New Zealand white rabbits were randomly divided into 3 groups ( n=12): rotator cuff repair group (control group), rotator cuff repair+CS hydrogel injection group (CS group), and rotator cuff repair+TDSCs/CS hydrogel injection group (TDSCs/CS group). After establishing the rotator cuff repair models, the corresponding hydrogel was injected into the tendon-to-bone interface in the CS group and TDSCs/CS group, and no other treatment was performed in the control group. The general condition of the animals was observed after operation. At 4 and 8 weeks, real-time quantitative PCR (qPCR) was used to detect the relative expressions of tendon forming related genes (tenomodulin, scleraxis), chondrogenesis related genes (aggrecan, sex determining region Y-related high mobility group-box gene 9), and osteogenesis related genes (alkaline phosphatase, Runt-related transcription factor 2) at the tendon-to-bone interface. At 8 weeks, HE and Masson staining were used to observe the histological changes, and the biomechanical test was used to evaluate the ultimate load and the failure site of the repaired rotator cuff to evaluate the tendon-to-bone healing and biomechanical properties.@*RESULTS@#CCK-8 assay showed that the CS hydrogel could promote the proliferation of TDSCs ( P<0.05). qPCR results showed that the expressions of tendon-to-bone interface related genes were significantly higher in the TDSCs/CS group than in the CS group and control group at 4 and 8 weeks after operation ( P<0.05). Moreover, the expressions of tendon-to-bone interface related genes at 8 weeks after operation were significantly higher than those at 4 weeks after operation in the TDSCs/CS group ( P<0.05). Histological staining showed the clear cartilage tissue and dense and orderly collagen formation at the tendon-to-bone interface in the TDSCs/CS group. The results of semi-quantitative analysis showed that compared with the control group, the number of cells, the proportion of collagen fiber orientation, and the histological score in the TDSCs/CS group increased, the vascularity decreased, showing significant differences ( P<0.05); compared with the CS group, the proportion of collagen fiber orientation and the histological score in the TDSCs/CS group significantly increased ( P<0.05), while there was no significant difference in the number of cells and vascularity ( P>0.05). All samples in biomechanical testing failed at the repair site during the testing process. The ultimate load of the TDSCs/CS group was significantly higher than that of the control group ( P<0.05), but there was no significant difference compared to the CS group ( P>0.05).@*CONCLUSION@#TDSCs/CS hydrogel can induce cartilage regeneration to promote rotator cuff tendon-to-bone healing.

Chromogenic in situ hybridization technique for detecting porcine circovirus 3 in lung and lymphoid tissues.

Background and Aim: Porcine circovirus 3 (PCV3) was recently reported in Malaysian commercial pig population in 2020 by conventional polymerase chain reaction (PCR), revealing a molecular prevalence of 17.02% in the sampled domestic pig population. This study aims to describe a chromogenic in situ hybridization (ISH) technique using digoxigenin (DIG)-labeled cloned PCV3 open reading frame 1 (ORF1) fragment DNA to detect and localize the PCV3 antigen in formalin-fixed, paraffin-embedded lung, and lymphoid tissue specimens. Materials and Methods: Since PCV3 was mainly detected in lung and lymphoid tissues, we obtained tissue specimens from these organs from the previous Malaysian PCV3 study. Digoxigenin-labeled ISH probes were designed to target a 69 bp region of PCV3 ORF1 spanning from the nucleotide positions (282-350). Results: Light microscopy analysis revealed that chromogenic staining of PCV3 antigens was visualized within the cytoplasm of pneumocytes and lymphocytes, indicating positive ISH results. The results of molecular detection of PCV3 using PCR and ISH showed a high agreement of 90.91%, including for the negative PCV3 status for all samples. Conclusion: This study reports a chromogenic ISH technique using DIG-labeled probes targeting PCV3 ORF1 to detect PCV3 antigens in lung and lymphoid tissues. Despite the limited availability of PCV3 antibodies, ISH remains relevant for investigating PCV3 replication and pathogenesis and can be used complementarily with PCR for evaluating the localization of antigens in infected tissues.

Molecular Characteristics and Pathogenicity of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) 1 in Taiwan during 2019-2020.

Two variants of porcine reproductive and respiratory syndrome virus (PRRSV), PRRSV 1 and PRRSV 2, have caused abortion in pregnant sows and respiratory distress in nursery pigs worldwide. PRRSV 2 has been thoroughly researched in Taiwan since 1993; however, the first case of PRRSV 1 was not reported until late 2018. To decipher the genetic characteristics of PRRSV 1 in Taiwan, open reading frame 5 (ORF5) genes of PRRSV 1 strains collected from 11 individual pig farms in 2019-2020 were successfully sequenced. All Taiwanese ORF5 sequences were closely related to Spanish-like PRRSV strains, which are considered to share a common evolutionary origin with the strain used for the PRRSV 1 vaccine. Analyses of amino acid (aa) and non-synonymous substitutions showed that genetic variations resulted in numerously specific codon mutations scattered across the neutralizing epitopes within the ORF5 gene. The PRRSV 1 challenge experiment disclosed the pathogenetic capability of the NPUST2789 isolate in nursery pigs. These findings provide comprehensive knowledge of the molecular diversity of the PRRSV 1 variant in local Taiwanese fields and facilitate the development of suitable immunization programs against this disease.

Fascia lata autograft bridging combined with long head of biceps tendon transposition for irreparable massive rotator cuff tear / 中华骨科杂志

Objective:To investigate the early clinical effect of fascia lata autograft bridging combined with the long head of biceps tendon transposition for treatment of irreparable massive rotator cuff tear.Methods:All of 31 cases of massive irreparable rotator cuff tear treated in our hospital from March 2016 to March 2020 were analyzed retrospectively. Among them, 17 cases (10 males, 7 females) were repaired with fascia lata autograft bridging under arthroscopy (patch group), the average age was 61.47±6.63 (ranging from 51 to 72) and 14 cases (4 males, 10 females) were repaired with fascia lata autograft bridging combined with the long head of biceps tendon transposition (combined group), the average age was 62.57±6.11 (ranging from 53 to 71). The operation time, intraoperative blood loss, postoperative complications, visual analogue scale (VAS) of pain before operation, at 1 week and 12 months after operation, Constant-Murley score of shoulder joint and American Association of shoulder and elbow Surgeons (ASES) score before operation, at 6 months and 12 months after operation were compared between the two groups. The outcome of rotator cuff healing was evaluated by MRI 1 year after operation.Results:All patients were followed up for 12-27 months (mean 18.33 ±6.8 months). There was no perioperative complication, and there was no significant difference in operation time between the two groups ( P>0.05) . The VAS score in the patch group was significantly higher than the combined group 1 week after operation ( t=2.09, P=0.048) , and there was no significant difference in VAS score 12 months after operation between the two groups. Constant-Murley score and ASES score in the combined group were significantly higher than the patch group at 6 months after operation ( t=5.23, P<0.001; t=4.45, P<0.001) , and there was no significant difference in Constant score and ASES score between the two groups at 12 months after operation. Constant score and ASES score in the two groups were significantly higher than those before operation. One year after operation, the MRI of the affected shoulder showed that the incidence of autograft patch thinning (Sugaya grade III) was 52.94%, the autograft patch structure failure rate (Sugaya grade IV and V) was 17.65% in the patch group, the autograft patch thinning rate (Sugaya grade III) was 35.71%, and the structural failure rate (Sugaya grade IV and V) was 7.14% in the combined group. The difference was statistically significant (χ 2=7.12, P=0.028) . Conclusion:Fascia lata autograft patch bridging combined with long head of biceps tendon transposition technique for treatment of irreparable massive rotator cuff tear has less pain 1 week after operation and better recovery of shoulder function half a year after operation. MRI showed better patch healing 1 year after operation.

Clinical Efficacy of Huanglian Jiedutang in Adjuvant Treatment of Acute Cerebral Infarction Complicated with Gastric Motility Disorder / 中国实验方剂学杂志

ObjectiveTo explore the clinical efficacy of Huanglian Jiedutang as an adjunctive treatment for acute cerebral infarction complicated with gastric motility disorder. MethodSixty patients with acute cerebral infarction complicated with gastric motility disorder with fire toxin syndrome were randomly divided into a western medicine control group （control group） and a traditional Chinese medicine （TCM） combined treatment group （observation group）， with 30 cases in each group. The control group received basic treatment for cerebral infarction and relevant western medical symptomatic treatment based on the patients' gastrointestinal symptoms. The observation group received Huanglian Jiedutang in addition to the treatment provided to the control group. The treatment course was 7 days. Neurological deficit scores and gastrointestinal dysfunction scores were assessed in both groups before treatment and on the 4th and 7th days of treatment. Gastrointestinal electrographic parameters， serum citrulline （CIT）， and motilin （MTL） levels were measured in both groups before treatment and on the 7th day of treatment. Clinical efficacy was compared between the two groups. ResultCompared with the baseline in both groups， the neurological deficit scores and gastrointestinal dysfunction scores were significantly reduced on the 4th and 7th days of treatment （P<0.05）. The reductions in these scores were more significant on the 7th day compared with those on the 4th day of treatment （P<0.05）. On the 4th and 7th days of treatment， the observation group showed a significantly greater reduction in neurological deficit scores and gastrointestinal dysfunction scores compared with the control group （P<0.05）. On the 7th day of treatment， compared with the baseline， both groups showed a significant increase in gastric antral and gastric body electric wave amplitudes as well as serum CIT and MTL levels （P<0.05）， and there were no statistically significant differences in the frequency of gastric antral and gastric body electric waves. On the 7th day of treatment， compared with the control group， the observation group had a significant increase in gastric antral and gastric body electric wave amplitudes as well as serum CIT and MTL levels （P<0.05）， and there were no statistically significant differences in the frequency of gastric antral and gastric body electric waves. After 7 days of treatment， the total effective rate in the observation group was 90.00% （27/30）， higher than 76.67% （23/30） in the control group， but the difference was not statistically significant. ConclusionAdjunctive treatment with Huanglian Jiedutang can effectively improve the symptoms of neurological function impairment and gastrointestinal dysfunction in patients with acute cerebral infarction complicated with gastric motility disorder， increase gastric antral and gastric body electric wave amplitudes， improve gastric motility disorder， and increase serum CIT and MTL levels， thereby improving the imbalanced secretion function of the gastrointestinal tract.

Carbon Chain Length Determines Inhibitory Potency of Perfluoroalkyl Sulfonic Acids on Human Placental 3β-Hydroxysteroid Dehydrogenase 1: Screening, Structure-Activity Relationship, and In Silico Analysis / 生物医学与环境科学（英文）

OBJECTIVE@#This study aimed to compare 9 perfluoroalkyl sulfonic acids (PFSA) with carbon chain lengths (C4-C12) to inhibit human placental 3β-hydroxysteroid dehydrogenase 1 (3β-HSD1), aromatase, and rat 3β-HSD4 activities.@*METHODS@#Human and rat placental 3β-HSDs activities were determined by converting pregnenolone to progesterone and progesterone secretion in JEG-3 cells was determined using HPLC/MS-MS, and human aromatase activity was determined by radioimmunoassay.@*RESULTS@#PFSA inhibited human 3β-HSD1 structure-dependently in the order: perfluorooctanesulfonic acid (PFOS, half-maximum inhibitory concentration, IC 50: 9.03 ± 4.83 μmol/L) > perfluorodecanesulfonic acid (PFDS, 42.52 ± 8.99 μmol/L) > perfluoroheptanesulfonic acid (PFHpS, 112.6 ± 29.39 μmol/L) > perfluorobutanesulfonic acid (PFBS) = perfluoropentanesulfonic acid (PFPS) = perfluorohexanesulfonic acid (PFHxS) = perfluorododecanesulfonic acid (PFDoS) (ineffective at 100 μmol/L). 6:2FTS (1H, 1H, 2H, 2H-perfluorooctanesulfonic acid) and 8:2FTS (1H, 1H, 2H, 2H-perfluorodecanesulfonic acid) did not inhibit human 3β-HSD1. PFOS and PFHpS are mixed inhibitors, whereas PFDS is a competitive inhibitor. Moreover, 1-10 μmol/L PFOS and PFDS significantly reduced progesterone biosynthesis in JEG-3 cells. Docking analysis revealed that PFSA binds to the steroid-binding site of human 3β-HSD1 in a carbon chain length-dependent manner. All 100 μmol/L PFSA solutions did not affect rat 3β-HSD4 and human placental aromatase activity.@*CONCLUSION@#Carbon chain length determines inhibitory potency of PFSA on human placental 3β-HSD1 in a V-shaped transition at PFOS (C8), with inhibitory potency of PFOS > PFDS > PFHpS > PFBS = PFPS = PFHxS = PFDoS = 6:2FTS = 8:2FTS.

Gastrin attenuates sepsis-induced myocardial dysfunction by down-regulation of TLR4 expression in macrophages

Myocardial dysfunction is the most serious complication of sepsis. Sepsis-induced myocardial dysfunction (SMD) is often associated with gastrointestinal dysfunction, but its pathophysiological significance remains unclear. The present study found that patients with SMD had higher plasma gastrin concentrations than those without SMD. In mice, knockdown of the gastrin receptor, cholecystokinin B receptor (Cckbr), aggravated lipopolysaccharide (LPS)-induced cardiac dysfunction and increased inflammation in the heart, whereas the intravenous administration of gastrin ameliorated SMD and cardiac injury. Macrophage infiltration plays a significant role in SMD because depletion of macrophages by the intravenous injection of clodronate liposomes, 48 h prior to LPS administration, alleviated LPS-induced cardiac injury in Cckbr-deficient mice. The intravenous injection of bone marrow macrophages (BMMs) overexpressing Cckbr reduced LPS-induced myocardial dysfunction. Furthermore, gastrin treatment inhibited toll-like receptor 4 (TLR4) expression through the peroxisome proliferator-activated receptor α (PPAR-α) signaling pathway in BMMs. Thus, our findings provide insights into the mechanism of the protective role of gastrin/CCKBR in SMD, which could be used to develop new treatment modalities for SMD.

Genome-wide analysis of transmembrane 9 superfamily genes in wheat (Triticum aestivum) and their expression in the roots under nitrogen limitation and Bacillus amyloliquefaciens PDR1 treatment conditions.

Introduction: Transmembrane 9 superfamily (TM9SF) proteins play significant roles in plant physiology. However, these proteins are poorly characterized in wheat (Triticum aestivum). The present study aimed at the genome-wide analysis of putative wheat TM9SF (TraesTM9SF) proteins and their potential involvement in response to nitrogen limitation and Bacillus amyloliquefaciens PDR1 treatments. Methods: TraesTM9SF genes were retrieved from the wheat genome, and their physiochemical properties, alignment, phylogenetic, motif structure, cis-regulatory element, synteny, protein-protein interaction (PPI), and transcription factor (TF) prediction analyses were performed. Transcriptome sequencing and quantitative real-time polymerase reaction (qRT-PCR) were performed to detect gene expression in roots under single or combined treatments with nitrogen limitation and B. amyloliquefaciens PDR1. Results and discussion: Forty-seven TraesTM9SF genes were identified in the wheat genome, highlighting the significance of these genes in wheat. TraesTM9SF genes were absent on some wheat chromosomes and were unevenly distributed on the other chromosomes, indicating that potential regulatory functions and evolutionary events may have shaped the TraesTM9SF gene family. Fifty-four cis-regulatory elements, including light-response, hormone response, biotic/abiotic stress, and development cis-regulatory elements, were present in the TraesTM9SF promoter regions. No duplication of TraesTM9SF genes in the wheat genome was recorded, and 177 TFs were predicted to target the 47 TraesTM9SF genes in a complex regulatory network. These findings offer valued data for predicting the putative functions of uncharacterized TM9SF genes. Moreover, transcriptome analysis and validation by qRT-PCR indicated that the TraesTM9SF genes are expressed in the root system of wheat and are potentially involved in the response of this plant to single or combined treatments with nitrogen limitation and B. amyloliquefaciens PDR1, suggesting their functional roles in plant growth, development, and stress responses. Conclusion: These findings may be vital in further investigation of the function and biological applications of TM9SF genes in wheat.

Analysis on prevalence and epidemic risk of animal plague in different ecological plague foci in Inner Mongolia Autonomous Region / 中华预防医学杂志

The risk of plague epidemics and relapse of various types of plague foci persists in Inner Mongolia Autonomous Region. For Marmota sibirica plague foci, the animal plague has not been found but antibody has been detected positive. Nowadays, Marmota sibirica has been increasing in population and distribution in China. In bordering countries Mongolia and Russia, the animal plague has been continuously prevalent. For Spermophilus dauricus plague foci, the animal plague has been taken place now and then. Compared to the above foci, the animal plague is most prevalent in Meriones unguiculatus plague foci and frequently spread to humans. Due to higher strain virulence and historical disaster in Marmota sibirica plague foci and Spermophilus dauricus plague foci, plague prevention and control should be strengthened on these foci. In addition to routine surveillance, epidemic dynamics need to be further monitored in these two foci, in order to prevent their relapse and spread to humans.

Study on Liver Injury of Different Components of Evodia Fructus in Vivo and in Vitro / 中国实验方剂学杂志

ObjectiveTo evaluate the in vivo and in vitro toxicity of Evodia Fructus water extraction and its index components， and provide a basis for basic research on the toxic substances of Evodia Fructus. MethodInstitute of Cancer Research（ICR） mice were divided into high， medium and low dose groups of water extraction of Evodia Fructus and a blank control group. The administration groups were respectively given 80，60，40 g·kg-1 water extraction of Evodia Fructus， the blank control group was given distilled water in equal volume， blood was taken 24 hours later to determine the serum alanine aminotransferase（ALT）and aspartate aminotransferase（AST）values， the liver was weighed and histopathological examination was performed. Evodia Fructus water extract， evodiamine， rutaecarpine and limonin were respectively acted on HepG2 cells for 24 h， and cell counting kit-8（CCK-8） method was used to investigate the cytotoxicity. The ICR Mice were divided into two groups， one group was given by oral gavage and the other group was given intraperitoneal injection. The two routes of administration were separately given 3 index components of Evodia Fructus， and the dosage was 200 mg·kg-1. Take blood 24 hours after administration to determine the activity of ALT and AST in serum， and take liver to calculate liver index. ResultCompared with the blank group， the high and medium dose groups of Evodia Fructus water extract were depressed 24 hours after administration， and the behavior of the low dose group was not significantly abnormal. The serum biochemical results showed that the activities of serum ALT and AST in the high and medium dose groups were significantly increased （P<0.01）， the activities of serum ALT and AST in the low dose group were significantly increased， and the histopathological results showed that the high and medium dose groups were significantly increased Punctate necrosis and vacuolar degeneration appeared in the liver of the medium dose group， and there was no obvious abnormality in the low dose group. Compared with the blank group， evodiamine and rutaecarpine had a certain inhibitory effect on the proliferation of HepG2 cells， but the inhibitory effect was not strong. Limonin had no significant inhibitory effect on the proliferation of HepG2 cells. Compared with the control group， the 3 index components of Evodia Fructus have no effect after oral administration. There was no significant difference in the activity of ALT and AST in serum of mice， and there was no significant difference in liver index. Intraperitoneal injection of evodiamine and rutaecarpine can cause the activity of serum ALT and AST to increase， and limonin can cause ALT activity was significantly increased （P<0.01）， and the liver index was significantly increased （P<0.05）. ConclusionEvodia Fructus water extract can cause acute liver injury in mice， Oral administration of evodiamine， rutaecarpine and limonin had no damage to the liver of mice. Intraperitoneal administration of evodiamine and rutaecarpine had no effect on liver injury in mice， and intraperitoneal administration of limonin could cause acute liver injury in mice.

Youguiwan Regulates CXCL8/CXCR1/2 Signaling Pathway and Ang-1 and Ang-2 Expression to Promote Ovarian Angiogenesis in Rats / 中国实验方剂学杂志

ObjectiveTo study the effects of Chinese herbal compound Youguiwan on angiogenesis of rats with ovarian dysfunction caused by natural aging and its relationship with chemokine interleukin 8 （CXCL8）/CXC chemokine receptor 1/2 （CXCR1/2） signaling pathway， angiopoietin 1 （Ang-1）， and angiopoietin 2 （Ang-2）， so as to explore its mechanism in improving the ovarian function. MethodFifty six female SD rats were randomly divided into the young control group （n=8） and modeling group （n=48， ovarian dysfunction caused by natural aging）. Rats in both the young control and modeling groups were routinely fed， during which the ones in the modeling group underwent exfoliative cytology of vaginal smears for five to seven days. The ones presented with prolonged estrous cycle， followed by continuous estrus and repeated pseudopregnancy revealed by vaginal cytology during four consecutive estrous cycles indicated early aging， and the young rats with keratinocyte proliferation index higher than 50% for 10 consecutive days were classified into the young control group. The successfully modeled rats were randomly divided into the early-aged group， estrogen （65 μg·kg-1·d-1） group， Zuoguiwan （33 g·kg-1·d-1） group， as well as the low-， medium-， and high-dose （1.2， 2.4， 4.8 g·kg-1·d-1） Youguiwan groups. Rats in the young control group and the early-aged group were gavaged with the same volume of normal saline for 30 days. After the experiment， the morphological changes in rat ovary were observed by hematoxylin-eosin （HE） staining. The protein expression levels of chemokines CXCL8， CXCR1， CXCR2， Ang-1， and Ang-2 in rat ovary were detected by Western blotting and immunohistochemistry， and the mRNA expression levels of CXCL8， CXCR1， CXCR2， Ang-1， and Ang-2 by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. ResultCompared with the young control group， the early-aged group exhibited reduced number of growing follicles， corpus luteum， and blood vessels at all levels， elevated atretic follicles （P<0.01）， up-regulated protein and mRNA expression of CXCL8， CXCR1， and CXCR2 in the ovarian tissue （P<0.01）， and down-regulated Ang-1 and Ang-2 protein and mRNA expression （P<0.05）. Compared with the early-aged group， each medication remarkably increased the number of growing follicles， corpus luteum， and blood vessels （P<0.05）， lowered the number of atretic follicles （P<0.05）， down-regulated the protein and mRNA expression levels of CXCL8， CXCR1， and CXCR2 in the ovarian tissue （P<0.05）， and up-regulated the protein and mRNA expression levels of Ang-1 and Ang-2 （P<0.05）. ConclusionYouguiwan down-regulates the levels of CXCL8， CXCR1， and CXCR2 in rat ovary and up-regulates the levels of Ang-1 and Ang-2 to promote ovarian angiogenesis and improve rat ovarian function.

Study on pharmacodynamics and pharmacokinetics of Phytolacca acinosa Roxb.And its mechanism / 中国药理学通报

Aim To explore the diuretic effect, diuretic mechanism and pharmacokinetics of Phytolacca acinosa Roxb., and clarify its "quantity-time-effect" relationship.Methods Firstly, qualified rats were modeled by water load model, given different doses of Phytolacca acinosa Roxb.aqueous extract, then the diuretic effect was investigated.Secondly, Western blot was used to detect the protein expression of aquaporins AQP2, AQP4 and the angiotensin II receptors ATGR1, ATGR2, and renin in the RAAS system in kidney tissues.Thirdly, the established LC-MS/MS biological analysis method was used to detect the esculentoside A(EsA)content in the plasma, calculate the pharmacokinetic parameters and analyze the correlation between the blood concentration and the drug effect.Results The water load model was successfully established.Compared with the model group, hydrochlorothiazide had a significant diuretic effect(P<0.01).Low, medium and high dose groups of Phytolacca acinosa Roxb.all had obvious diuretic effects(P<0.01), EsA also had a significant diuretic effect(P<0.05).Phytolacca acinosa Roxb.aqueous extract and EsA significantly down-regulated the expression of AQP2, AQP4, ATGR1 and renin protein.The pharmacokinetic results showed that the Cmax and AUC0-t of EsA in the plasma of rats in the low, medium, and high dose groups of aqueous extract increased with the increase of the dose.Conclusions Phytolacca acinosa Roxb.had a diuretic effect, which is related to inhibiting the expression of aquaporins AQP2 and AQP4 and inhibiting the expression of angiotensin II type 1 receptor and renin, thereby inhibiting the reabsorption of renal tubules and collecting ducts.

Interlaboratory method validation of slope ratio determination for anticoagulant activity of leeches / 中国药理学通报

Aim To investigate the slope ratio method for the determination of anticoagulant activity of leeches. Methods Three batches of leeches, four groups of Japanese medical vermiculite yinpian and fifteen groups of leech preparations were chosen, with contrast medicinal leeches herbs and Philippine cattle leech contrast medicinal materials, and different concentrations of leaching solutions were prepared in parallel. APTT value was determined after anticoagulant activity was determined by slope ratio method for the joint validation of laboratory, intermediate precision and accuracy between the linear range. Results The slope ratio method was accurate and accurate in the determination of anticoagulant activity of leeches, with linearity between 64% and 156% relative titer level. Conclusion Slope ratio method can be used to determine the anticoagulant activity of leeches.

The preliminary exploration of Calu-3 cell air-liquid interface culture and exposure research / 中国药理学通报

Aim To explore the air-liquid interface(ALI)culture conditions of Calu-3 cells and their tolerance to exposure to clean air in the exposure system. Methods Calu-3 cells were cultured in three stages:flask expansion,liquid-liquid culture until full Transwell membrane covered,and ALI culture,and the cell barrier status was determined by cell trans-epithelium electrical resistant(TEER)measurement and expression of tight junction protein ZO-1; Calu-3 cells were exposed to clean air using the air-liquid interface exposure system for 1,2,3,4,5,6 hours,and cells cultured in incubators were used as controls to detect changes in Calu-3 cell activity and TEER after exposure,and to determine the single maximum exposure time of Calu-3 cells in the in vitro exposure system based on ALI culture. Results Calu-3 cells were cultured in liquid-liquid culture for 8±1 days to grow full Transwell membranes and barrier formation,and then were transferred to ALI culture. The barrier was intact and in a stable state for 1 to 18 days of ALI culture,and could be used for exposure experiments. The activity of Calu-3 cells decreased significantly after 6 hours of clean air exposure(P<0.01),and TEER decreased significantly after 4,5,and 6 hours of clean air exposure(P<0.05,P<0.01,P<0.01). Conclusions Calu-3 cells cultured for 1 to 18 days in ALI can be used for air-liquid interface exposure experiments; the combined changes in cell activity and TEER suggest that the maximum single exposure time for Calu-3 cells exposed to clean air in the exposure system is 3 hours.

Phylodynamic analysis and spike protein mutations in porcine deltacoronavirus with a new variant introduction in Taiwan.

Porcine deltacoronavirus (PDCoV) is a highly transmissible intestinal pathogen that causes mild to severe clinical symptoms, such as anorexia, vomiting, and watery diarrhea in pigs. By comparing the genetic sequences of the spike glycoprotein between historical and current Taiwanese PDCoV strains, we identified a novel PDCoV variant that displaced the PDCoV responsible for the 2015 epidemic. This PDCoV variant belongs to a young population within the US lineage, and infected pigs carry high concentrations of the virus. It also has several critical point mutations and an amino acid insertion at position 52 that may enhance the affinity between the B-cell epitopes located in the N-terminal domain with its complementarity regions, consequently facilitating binding or penetration between the fusion peptide and cellular membrane. Furthermore, viral protein structure prediction demonstrated that these amino acid changes may change the ability of the virus to bind to the receptor, which may consequently alter virus infectivity. Our results hence suggest the emergence of new PDCoV strains in Taiwan with the potential for greater transmission and pathogenesis.