Objective To report the diagnosis of a canine distemper virus outbreak among a colony of cynomolgus monkeys at an experimental monkey farm in 2019. MethodsA total of 46 samples were collected from 21 diseased cynomolgus monkeys (exhibiting symptoms such as facial rash, skin scurf, runny nose, and diarrhea) and from one deceased monkey at an experimental monkey breeding farm in South China in late 2019, including serum, skin rash swabs, and anticoagulated whole blood, liver, lung, and skin tissues were submitted for testing. All submitted samples were tested for canine distemper virus gene fragments using real-time quantitative PCR, while immunohistochemical staining was performed to detect canine distemper virus nucleoprotein in lung tissues. The skin tissue of the deceased monkey was ground and sieved. The filtrate was inoculated into a monolayer MDCK cell line for virus isolation. Then, whole-genome sequencing was performed to identify the isolated virus. The Clustal Omega tool was used to align and analyze the homology of different Asian canine distemper virus isolates. A phylogenetic tree was constructed, followed by genetic evolutionary analysis. ResultsClinical retrospective analysis revealed that the diseased cynomolgus monkeys exhibited symptoms similar to those observed in cynomolgus monkeys infected with measles virus. Necropsy findings showed red lesions in the lungs and significant hemorrhage in the colonic mucosa. Real-time quantitative PCR detected canine distemper virus nucleic acid in the serum, skin rash swabs of the infected monkeys, and various tissue samples of the deceased monkey, all of which tested positive. Calculation based on the standard curve formula indicated the viral load was highest in the skin tissue. Immunohistochemical staining of the deceased monkey's lung tissue demonstrated aggregation of CDV nucleoprotein in alveolar epithelial cells, bronchi, and bronchioles. A CDV strain was isolated from the skin tissue of the deceased monkey. Phylogenetic analysis indicated that this strain shares the closest relationship (98.86%) with the Asian-1 type canine distemper virus strain CDV/dog/HCM/33/140816, previously identified in dogs in Vietnam. ConclusionBased on comprehensive analysis of clinical symptoms, nucleic acid detection, viral protein immunohistochemistry, and whole-genome sequencing results, the diagnosis confirms that the cynomolgus monkeys in this facility are infected with canine distemper virus. It is recommended to include canine distemper virus as a routine surveillance target in captive monkey populations. Additionally, this study provides a foundation for further research on the molecular biological characteristics of canine distemper virus.

Objective To report the diagnosis of a canine distemper virus outbreak among a colony of cynomolgus monkeys at an experimental monkey farm in 2019. MethodsA total of 46 samples were collected from 21 diseased cynomolgus monkeys (exhibiting symptoms such as facial rash, skin scurf, runny nose, and diarrhea) and from one deceased monkey at an experimental monkey breeding farm in South China in late 2019, including serum, skin rash swabs, and anticoagulated whole blood, liver, lung, and skin tissues were submitted for testing. All submitted samples were tested for canine distemper virus gene fragments using real-time quantitative PCR, while immunohistochemical staining was performed to detect canine distemper virus nucleoprotein in lung tissues. The skin tissue of the deceased monkey was ground and sieved. The filtrate was inoculated into a monolayer MDCK cell line for virus isolation. Then, whole-genome sequencing was performed to identify the isolated virus. The Clustal Omega tool was used to align and analyze the homology of different Asian canine distemper virus isolates. A phylogenetic tree was constructed, followed by genetic evolutionary analysis. ResultsClinical retrospective analysis revealed that the diseased cynomolgus monkeys exhibited symptoms similar to those observed in cynomolgus monkeys infected with measles virus. Necropsy findings showed red lesions in the lungs and significant hemorrhage in the colonic mucosa. Real-time quantitative PCR detected canine distemper virus nucleic acid in the serum, skin rash swabs of the infected monkeys, and various tissue samples of the deceased monkey, all of which tested positive. Calculation based on the standard curve formula indicated the viral load was highest in the skin tissue. Immunohistochemical staining of the deceased monkey's lung tissue demonstrated aggregation of CDV nucleoprotein in alveolar epithelial cells, bronchi, and bronchioles. A CDV strain was isolated from the skin tissue of the deceased monkey. Phylogenetic analysis indicated that this strain shares the closest relationship (98.86%) with the Asian-1 type canine distemper virus strain CDV/dog/HCM/33/140816, previously identified in dogs in Vietnam. ConclusionBased on comprehensive analysis of clinical symptoms, nucleic acid detection, viral protein immunohistochemistry, and whole-genome sequencing results, the diagnosis confirms that the cynomolgus monkeys in this facility are infected with canine distemper virus. It is recommended to include canine distemper virus as a routine surveillance target in captive monkey populations. Additionally, this study provides a foundation for further research on the molecular biological characteristics of canine distemper virus.

Myelodysplastic syndrome (MDS), a myeloid tumor derived from the malignant clones of hematopoietic stem cells, has an annually increasing incidence. The contemporary research direction has shifted to analyzing the synergistic effect of immune surveillance collapse and abnormal bone marrow microenvironment in the pathological process of MDS. Against this backdrop, the immune checkpoint molecule TIM3 has emerged as a key target because of its persistently high expression on the surface of important immune cells such as T and NK cells. The abnormal activation of the TIM3 pathway is the mechanism by which solid tumors and hematological malignancies achieve immune escape and is a key hub in the formation of immune exhaustion phenotypes. This work integrates the original discoveries of our team with the latest international progress, systematically demonstrating the bidirectional regulatory network of TIM3 between the malignant clone proliferation of MDS and the immunosuppressive microenvironment. Integrating the evidence from emerging clinical trials allows us to consider the clinical significance of TIM3-targeted blocking for MDS, providing a transformative path to overcome the resistance of traditional treatments and marking a new chapter in the active immune reconstitution of MDS treatment.

OBJECTIVE To standardize the main processes and related technical links of the clinical comprehensive evaluation of drugs， and provide guidance and reference for improving the quality of comprehensive evaluation evidence and its transformation and application value. METHODS The construction of Guideline for the Workflow of Clinical Comprehensive Evaluation of Drugs was based on the standard guideline formulation method of the World Health Organization （WHO）， strictly followed the latest definition of guidelines by the Institute of Medicine of the National Academy of Sciences of the United States， and conformed to the six major areas of the Guideline Research and Evaluation Tool Ⅱ. Delphi method was adopted to construct the research questions； research evidence was established by applying the research methods of evidence-based medicine. The evidence quality classification system of the Chinese Evidence-Based Medicine Center was adopted for evidence classification and evaluation. The recommendation strength was determined by the recommendation strength classification standard formulated by the Oxford University Evidence-Based Medicine Center， and the recommendation opinions were formed through the expert consensus method. RESULTS & CONCLUSIONS The Guideline for the Workflow of Clinical Comprehensive Evaluation of Drugs covers 4 major categories of research questions， including topic selection， evaluation implementation， evidence evaluation， and application and transformation of results. The formulation of this guideline has standardized the technical links of the entire process of clinical comprehensive evaluation of drugs， which can effectively guide the high-quality and high-efficient development of this work， enhance the standardized output and transformation application value of evaluation evidence， and provide high-quality evidence support for the scientific decision-making of health and the rationalization of clinical medication.

The superficial zone (SFZ) of articular cartilage is an important interface that isolates deeper zones from the microenvironment of the articular cavity and is directly exposed to various biological and mechanical stimuli. The SFZ is not only a crucial structure for maintaining the normal physiological function of articular cartilage but also the earliest site of osteoarthritis (OA) cartilage degeneration and a major site of cartilage progenitor cells, suggesting that the SFZ might represent a key target for the early diagnosis and treatment of OA. However, to date, SFZ research has not received sufficient attention, accounting for only about 0.58% of cartilage tissue research. The structure, biological composition, function, and related mechanisms of the SFZ in the physiological and pathological processes of articular cartilage remain unclear. This article reviews the key role of the SFZ in articular cartilage physiology and pathology and focuses on the characteristics of SFZ in articular cartilage degeneration and regeneration in OA, aiming to provide researchers with a systematic understanding of the current research status of the SFZ of articular cartilage, hoping that scholars will give more attention to the SFZ of articular cartilage in the future.
Cartilage, Articular/pathology* ; Humans ; Regeneration/physiology* ; Animals ; Osteoarthritis/physiopathology*

This study aims to investigate the correlations of the appearance traits, total antioxidant capacity, and component content of Forsythiae Fructus processed by different methods, explore the effects of different processing methods on the abovementioned three aspects of Forsythiae Fructus, and screen out the internal and external indicators that have important effects on its quality. It determined the length, diameter, stem length, chroma value L~*, a~*, b~*, and other appearance indexes and antioxidant activity of Forsythiae Fructus processed by different methods. The content of forsythiaside A, rutin, forsythin, pinoresinol, and phillygenin was determined by ultra performance liquid chromatography(UPLC). Correlation analysis, principal component analysis(PCA), orthogonal partial least squares discriminant analysis(OPLS-DA), and independent sample t-test analysis were performed on the appearance indexes and the component content. The correlation analysis showed that there were differences in the appearance traits and the component content. L~* and E~* had highly significant negative correlations with pinoresinol and phillygenin(P<0.01) and significant positive correlations with forsythiaside A(P<0.05). There were a highly significant negative correlation between a~* and forsythiaside A(P<0.01) and highly significant positive correlations of a~* with pinoresinol and phillygenin(P<0.01). There were a highly significant positive correlation between b~* and forsythiaside A(P<0.01) and highly significant negative correlations of b~* with pinoresinol and phillygenin(P<0.01). The total antioxidant capacity had highly significant negative correlations with pinoresinol and phillygenin(P<0.01). The PCA results showed that there were differences among Forsythiae Fructus samples processed by different methods. OPLS-DA marked five important indicators, which were forsythiaside A, stem length, E~*, L~*, and b~*. The results of independent sample t-test showed that the content of forsythiaside A, pinoresinol, and phillygenin, the total antioxidant capacity, and the appearance traits such as L~*, a~*, b~*, and E~* were significantly different between the Forsythiae Fructus samples processed by steaming and boiling(P<0.05). According to content determination and a related biological activity analysis, steaming is a good choice from the perspective of improving the stability of chemical constituents and antioxidant activity of Forsythiae Fructus. From the point of view of improving the stability of chemical constituents and anti-inflammatory and anti-cancer activities of Forsythiae Fructus, it is recommended to use boiling as the processing method. Based on the above analysis methods, the main indexes for the appearance traits of Forsythiae Fructus processed by different methods are powder chroma value(L~*, a~*, b~*, E~*), stem length, and total antioxidant capacity, and those for chemical constituents are the content of forsythiaside A, pinoresinol, and phillygenin. This study provides reference for seeking scientific processing methods of Forsythiae Fructus.
Forsythia/chemistry* ; Drugs, Chinese Herbal/isolation & purification* ; Fruit/chemistry* ; Antioxidants/analysis* ; Chromatography, High Pressure Liquid ; Glycosides/analysis* ; Principal Component Analysis ; Furans ; Lignans

Buyang Huanwu Decoction(BYHWD), as one of the classic formulas in traditional Chinese medicine(TCM) for the treatment of cerebral ischemic stroke(CIS), has demonstrated definite effects in clinical practice. However, the material basis and mechanism of treatment have not been systematically elucidated. This study employed network pharmacology and molecular docking to analyze the potential targets and mechanisms of blood-and brain-penetrating active components of BYHWD in reducing cell apoptosis in CIS. Cell experiments were then carried out to validate the prediction results. In the experiments, five active components including hydroxysafflor yellow A( HSYA), tetramethylpyrazine( TMP), astragaloside Ⅳ( AS-Ⅳ), amygdalin( AMY), and paeoniflorin(PF) were selected to explore the pharmacological effects of BYHWD. HT22 cells were treated with BYHWD, and the cell counting kit-8(CCK-8) method was employed to examine the toxic and side effects of BYHWD. A cell model of oxygen-glucose deprivation/reoxygenation( OGD/R) was constructed, with apoptosis and pyroptosis as the main screening indicators. The levels of lactate dehydrogenase(LDH) and glutathione(GSH) were measured to assess the cell membrane integrity. Flow cytometry was employed to detect apoptosis, and the activities of caspase-3 and caspase-1 were measured to clarify the status of apoptosis and pyroptosis. ELISA was employed to determine the levels of interleukin(IL)-1β and IL-18 to confirm pyroptosis. HSYA and AMY were identified in this study as the active components regulating apoptosis and pyroptosis. TUNEL was employed to detect the apoptosis rate, and Western blot was employed to determine the expression levels of apoptosis-related proteins B-cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax), and caspase-3, which confirmed that the anti-apoptotic effect of the combined component group was superior to that of the single component groups. The molecular docking results revealed strong binding affinity of HSYA and AMY with SDF-1α and CXCR4.AMD3100, a selective antagonist of CXCR4, was then used for intervention. The results of Western blot showed alterations in the expression levels of apoptosis-associated proteins, SDF-1α, and CXCR4. In conclusion, HSYA and AMY influence cellular apoptosis by modulating the SDF-1α/CXCR4 signaling cascade.
Drugs, Chinese Herbal/chemistry* ; Apoptosis/drug effects* ; Animals ; Neurons/cytology* ; Mice ; Molecular Docking Simulation ; Cell Line ; Glucose/metabolism* ; Humans ; Neuroprotective Agents/pharmacology*

The ultra-high performance liquid chromatography( UPLC) characteristic fingerprints of Angelica dahurica and A. dahurica var. formosana were established. The compounds corresponding to common peaks were identified by ultra-high performance liquid chromatography with quadrupole time-of-flight mass spectrometry( UPLC-Q-TOF-MS/MS). The results were combined with chemometrics and quantitative analysis of multi-components with a single-marker method(QAMS) to study the quality control of A. dahurica and A. dahurica var. formosana. The separation was performed on a Titank C_(18) column(2. 1 mm × 150 mm, 1. 8 μm)with a mobile phase of acetonitrile-0. 2% formic acid at a flow rate of 0. 3 m L·min~(-1). The column temperature was 35 ℃ and the injection volume was 1. 2 μL. Seven batches of A. dahurica and 11 batches of A. dahurica var. formosana were injected and analyzed. The UPLC characteristic fingerprints of A. dahurica and A. dahurica var. formosana were established according to the Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine( version 2012), and 19 and 20 characteristic peaks were matched respectively. The common peaks were identified by reference substance comparison and UPLC-Q-TOF-MS/MS. Cluster analysis(CA), principal component analysis(PCA), and orthogonal partial least squares-discriminant analysis(OPLS-DA)were performed to analyze the chemical pattern recognition of A. dahurica and A. dahurica var. formosana. The results of CA and PCA could distinguish Angelicae Dahuricae Radix from different producing areas, and the differential quality markers of A. dahurica and A. dahurica var. formosana were obtained by OPLS-DA. With imperatorin as the internal reference, the relative correction factors of oxypeucedanin hydrate, byakangelicin, bergapten, isopimpinellin, oxypeucedanin, and isoimperatorin were 1. 310, 1. 069, 0. 729, 0. 633, 0. 753, and 1. 010, respectively. There was no significant difference between the QAMS and external standard method(ESM)results of each component, indicating that the QAMS established with imperatorin as the internal reference was accurate and reliable. The characteristic fingerprints, chemometrics, and QAMS established in this study can quickly and efficiently control the quality of A. dahurica and A. dahurica var. formosana.
Quality Control ; Chromatography, High Pressure Liquid/methods* ; Drugs, Chinese Herbal/chemistry* ; Angelica/chemistry* ; Chemometrics/methods* ; Tandem Mass Spectrometry/methods* ; Principal Component Analysis

This study primarily investigates the effect of Liuwei Dihuang Pills on the activation and proliferation of female germline stem cells(FGSCs) in the ovaries and cortex of mice with premature ovarian failure(POF), and how it improves ovarian function. ICR mice were randomly divided into the control group, model group, Liuwei Dihuang Pills group, Liuwei Dihuang Pills double-dose group, and estradiol valerate group. A mouse model of POF was established by intraperitoneal injection of cyclophosphamide. After successful modeling, the mice were treated with Liuwei Dihuang Pills or estradiol valerate for 28 days. Vaginal smears were prepared to observe the estrous cycle and body weight. After the last administration, mice were sacrificed and sampled. Serum levels of estradiol(E_2), follicle-stimulating hormone(FSH), luteinizing hormone(LH), and anti-Müllerian hormone(AMH) were measured by enzyme-linked immunosorbent assay(ELISA). Hematoxylin-eosin(HE) staining was used to observe ovarian morphology and to count follicles at all stages to evaluate ovarian function. Immunohistochemistry was used to detect the expression of mouse vasa homolog(MVH), a marker of ovarian FGSCs. Immunofluorescence staining, using co-labeling of MVH and proliferating cell nuclear antigen(PCNA), was used to detect the expression and localization of specific markers of FGSCs. Western blot was employed to assess the protein expression of MVH, octamer-binding transcription factor 4(Oct4), and PCNA in the ovaries. The results showed that compared with the control group, the model group exhibited disordered estrous cycles, decreased ovarian index, increased atretic follicles, and a reduced number of follicles at all stages. FSH and LH levels were significantly elevated, while AMH and E_2 levels were significantly reduced, indicating the success of the model. After treatment with Liuwei Dihuang Pills or estradiol valerate, hormone levels improved, the number of atretic follicles decreased, and the number of follicles at all stages increased. MVH marker protein and PCNA proliferative protein expression in ovarian tissue also increased. These results suggest that Liuwei Dihuang Pills regulate estrous cycles and hormone disorders in POF mice, promote the proliferation of FGSCs, improve follicular development in POF mice, and enhance ovarian function.
Animals ; Female ; Drugs, Chinese Herbal/administration & dosage* ; Mice ; Cell Proliferation/drug effects* ; Mice, Inbred ICR ; Ovary/cytology* ; Primary Ovarian Insufficiency/genetics* ; Follicle Stimulating Hormone/metabolism* ; Humans ; Anti-Mullerian Hormone/blood* ; Antineoplastic Agents/adverse effects* ; Luteinizing Hormone/metabolism* ; Cyclophosphamide/adverse effects*

Traditional Chinese medicine(TCM), a treasure of the Chinese nation, contains abundant chemical components and demonstrates unique pharmacological activities, showing important values in clinical applications. With profound connotations and broad application prospects, TCM urgently needs us to further explore and conduct systematic research. Puerarin is a small-molecule natural isoflavonoid carbon glycoside extracted from plants of Pueraria. It is also the main active ingredient of Puerariae Lobata Radix, a Chinese herbal medicine with both medicinal and edible values. Puerarin has a variety of pharmacological effects such as blood pressure-lowering, anti-atherosclerosis, anti-ischemia-reperfusion injury, antithrombotic, anti-tumor, anti-inflammatory, liver-protecting, nerve cell-protecting, and intestinal microbiota-regulating effects. It is also an active ingredient that has been widely studied. This article comprehensively reviews the research progress in the pharmacological effects and molecular mechanisms of puerarin over the years, aiming to provide references and theoretical support for the in-depth research and development as well as clinical application of puerarin.
Isoflavones/chemistry* ; Humans ; Animals ; Drugs, Chinese Herbal/chemistry* ; Pueraria/chemistry*