Rheumatoid arthritis belongs to arthralgia in the theory of traditional Chinese medicine, with cold dampness obstruction pattern (CDO) and damp heat obstruction pattern (DHO) as the main pattern types. Fine therapeutic effects have been obtained in clinical practice following the differentiation of CDO and DHO. However, suitable animal models are not available currently. For adapting to the clinical diagnosis, as well as carrying out basic research of integrated Chinese and Western medicine and preclinical study on new Chinese medicine in a better way, the “Guidelines for Establishing Animal Models of Rheumatoid Arthritis with Cold Dampness Obstruction Pattern and Damp Heat Obstruction Pattern” (referred to as “Guidelines”) were compiled by our research group in cooperation with renowned experts in clinical, pharmaceutical, zoological, and methodological research fields. According to the theory of disease and syndrome integration, experts standardized the establishment methods for animal models of rheumatoid arthritis with CDO and DHO. The Guidelines were compiled via the nominal group method according to the principle of evidence coming in main place, consensus as the auxiliary, and experience as the references. Contents such as animal species, arthritis induction methods, external stimulation conditions, and assessment indicators were specified in the Guidelines based on the comprehensive evaluation of pathogenesis homology, behavioral phenotypic consistency, and drug therapy predictability between animal models and human diseases. The Guidelines will be applicable to the research on syndromes in rheumatoid arthritis, elucidation of acting mechanism, and development of new medicine, and provide reference for standardizing other types of animal models with the integration of disease and syndrome, to promote the modernization of Chinese medicine research.

Objective: To improve the efficiency of patient treatment in routine, emergency, and mass casualty scenarios (both peacetime and wartime), eliminate reliance on blood type compatibility, and promote the productization of platelet-rich plasma (PRP), this study investigated the optimal mixing ratio for developing a universal PRP formulation. Methods: PRP was prepared from donors with blood types O, A, B, and AB. Samples were mixed in varying proportions, and the optimal ratio was determined based on the trend of anti-A and anti-B antibody titers in the mixed PRP. Concentrations of the growth factors PDGF-AA, PDGF-BB, EGF, TGF-β1, and VEGF were measured by enzyme-linked immunosorbent assay (ELISA) before and after mixing. Results: A mixing ratio of A∶B∶AB=6∶2.5∶24 resulted in no detectable agglutination with either A or B cells in the microcolumn gel system, indicating the absence of immunological reactivity and confirming universal compatibility. Importantly, the levels of all measured growth factors remained unchanged following mixing, suggesting preservation of biological activity. Conclusion: These findings suggest that universal PRP can be prepared by neutralizing blood type antibodies through antigen-antibody reactions.

The Guidelines for Establishing Animal Models of Rheumatoid Arthritis with Cold-dampness Obstruction Syndrome and Dampness-heat Obstruction Syndrome （hereinafter referred to as the Guidelines） （No. T/CACM1567-2024） was published by Chinese Association of Chinese Medicine on January 11， 2024. To assist researchers and medical workers in understanding and applying the Guidelines more accurately， and also to provide reference and assistance for the establishment of guidelines on other types of diseases and syndromes combined with animal models， this paper made a declaration of the workflow， technological links， development references， promotion of its application and after-effect evaluation of the Guidelines that has been made according to the requirements of "Draft Group Standard of the Standardization Office of the Chinese Association of Traditional Chinese Medicine".

Hepatic encephalopathy is a difficult and critical disease with rapid progression and limited treatment methods in the field of liver disease， and it is urgently needed to make breakthroughs in its pathogenesis. Selection of appropriate prevention and treatment strategies is of great importance in delaying disease progression and reducing the incidence and mortality rates. This article reviews the theory of “turbid toxin pathogenesis” and related prevention and treatment strategies for hepatic encephalopathy in traditional Chinese medicine/Zhuang medicine， proposes a new theory of “turbid toxin pathogenesis”， analyzes the scientific connotations of “turbid”， “toxin”， and the theory of “turbid toxin pathogenesis”， and constructs the “four-step” prevention and treatment strategies for hepatic encephalopathy， thereby establishing the new clinical prevention and treatment regimen for hepatic encephalopathy represented by “four prescriptions and two techniques” and clarifying the effect mechanism and biological basis of core prescriptions and techniques in the prevention and treatment of hepatic encephalopathy， in order to provide a reference for the prevention and treatment of hepatic encephalopathy.

OBJECTIVE: To investigate the inhibitory effect of Tanreqing Injection (TRQ) on the activation of nucleotide-binding oligomerization domain-like receptor pyrin domain containing 3 (NLRP3) inflammasome in macrophages infected with influenza A virus and the underlying mechanism based on mitophagy pathway. METHODS: The inflammatory model of murine macrophage J774A.1 induced by influenza A virus [strain A/Puerto Rico/8/1934 (H1N1), PR8] was constructed and treated by TRQ, while the mitochondria-targeted antioxidant Mito-TEMPO and autophagy specific inhibitor 3-methyladenine (3-MA) were used as controls to intensively study the anti-inflammatory mechanism of TRQ based on mitophagy-mitochondrial reactive oxygen species (mtROS)-NLRP3 inflammasome pathway. The levels of NLRP3, Caspase-1 p20, microtubule-associated protein 1 light chain 3 II (LC3II) and P62 proteins were measured by Western blot. The release of interleukin-1β (IL-1β) was tested by enzyme linked immunosorbent assay, the mtROS level was detected by flow cytometry, and the immunofluorescence and co-localization of LC3 and mitochondria were observed under confocal laser scanning microscopy. RESULTS: Similar to the effect of Mito-TEMPO and contrary to the results of 3-MA treatment, TRQ could significantly reduce the expressions of NLRP3, Caspase-1 p20, and autophagy adaptor P62, promote the expression of autophagy marker LC3II, enhance the mitochondrial fluorescence intensity, and inhibit the release of mtROS and IL-1β (all P<0.01). Moreover, LC3 was co-localized with mitochondria, confirming the type of mitophagy. CONCLUSION TRQ could reduce the level of mtROS by promoting mitophagy in macrophages infected with influenza A virus, thus inhibiting the activation of NLRP3 inflammasome and the release of IL-1β, and attenuating the inflammatory response.
Mitophagy/drug effects* ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Animals ; Macrophages/virology* ; Inflammasomes/drug effects* ; Drugs, Chinese Herbal/pharmacology* ; Mice ; Mitochondria/metabolism* ; Reactive Oxygen Species/metabolism* ; Influenza A virus/physiology* ; Interleukin-1beta/metabolism* ; Cell Line ; Injections

OBJECTIVE: To investigate the regulatory effects of two traditional mineral medicines (TMMs), Gypsum Fibrosum (Shigao, GF) and Terra Flava Usta (Zaoxintu, TFU), on gut-beneficial bacteria in mice, and preliminarily explore their mechanisms of action. METHODS: Mice were randomly divided into 3 groups (n=10 per group): the control group (standard diet), the GF group (diet supplemented with 2% GF), and the TFU group (diet supplemented with 2% TFU). After 4-week intervention, 16S rRNA gene sequencing was used to analyze the changes in the gut microbiota (GM). Scanning electron microscopy, in combination with coumarin A tetramethyl rhodamine conjugate and Hoechst stainings, was used to observe the bacteria and biofilm formation. RESULTS: Principal coordinate analysis revealed that GF and TFU significantly altered the GM composition in mice. Further analysis revealed that GF and TFU affected different types of gut bacteria, suggesting that different TMMs may selectively modulate specific bacterial populations. For certain bacteria, such as Faecalibaculum and Ileibacterium, both GF and TFU exhibited growth-promoting effects, implying that they may be sensitive to TMMs and that different TMMs can increase their abundance through their respective mechanisms. Notably, Lactobacillus reuteri, a widely recognized and used probiotic, was significantly enriched in the GF group. Random forest analysis identified Ileibacterium valens as a potential indicator bacterium for TMMs' impact on GM. Further mechanistic studies showed that gut bacteria formed biofilm structures on the TFU surface. CONCLUSIONS This study provides new insights into the interaction between TMMs and GM. As safe and effective natural clays, GF and TFU hold promise as potential candidates for prebiotic development.
Animals ; Gastrointestinal Microbiome/drug effects* ; Bacteria/growth & development* ; Mice ; Biofilms/drug effects* ; Male ; RNA, Ribosomal, 16S/genetics*

Objective To explore the predictive value of inflammatory burden index(IBI)combined with pregnancy-associated plasma protein-A(PAPP-A)for asymptomatic missed abortion.Methods A total of 108 asymptomatic women with missed abortions admitted to Northwest Women's and Children's Hospital from January 2020 to January 2023 were selected as the study group,and another 100 healthy pregnant women who were examined in the hospital during the same period were selected as the control group.The levels of C-reac-tive protein,neutrophils,lymphocytes,and PAPP-A of all research subjects were detected,and IBI was calcu-lated.The receiver operating characteristic curve was plotted to analyze the predictive value of IBI combined with PAPP-A for asymptomatic isolated abortion,and to explore the influencing factors of asymptomatic iso-lated abortion.Results The IBI of the study group was higher than that of the control group,and the PAPP-A was lower than that of the control group,the difference was statistically significant(P<0.05).The area under the curve(AUC)of the combined prediction of asymptomatic missed abortion by IBI and PAPP-A was higher than AUC predicted by IBI and PAPP-A alone(Z=9.251,10.030,P<0.001).There were statistically significant differences in the proportions of smoking history,drinking history,intrauterine operation history and reproductive tract infection between the study group and the control group(P<0.05).The results of multivariate Logistic regression analysis showed that smoking history,drinking history,intrauterine operation history,reproductive tract infection,and IBI were risk factors for asymptomatic missed abortion,while PAPP-A was a protective factor for asymptomatic missed abortion(P<0.05).Conclusion IBI combined with PAPP-A has certain clinical value in predicting asymptomatic missed abortion.

BACKGROUND: Regulatory dendritic cell (DCreg) subset exhibits a unique capacity for inducing immune tolerance among the variety subsets of dendritic cells (DCs) within gut-associated lymphoid tissues (GALTs). Fms-like tyrosine kinase 3 ligand (FLT3L) is involved in the differentiation of DCregs and the subsequent expansion of regulatory T-cells (Tregs) mediated by DCregs, though the precise mechanism remains poorly understood. This study aimed to explore the expansion mechanism of Treg induced by DCreg and the role of FLT3L in this process. METHODS: DCregs were distinguished from other DC subsets isolated from GALTs of BALB/c mice through a mixed lymphocyte reaction assay. The functions and mechanisms by which FLT3L promoted Treg expansion via DCregs were investigated in vitro through co-culture experiments involving DCregs and either CD4 + CD25 - T-cells or CD4 + CD25 + T-cells. Additionally, an in vivo experiment was conducted using a dextran sulfate sodium (DSS)-induced colitis model in mice. RESULTS: CD103 + CD11b + DC exhibited DCreg-like functionality and was identified as DCreg for subsequent investigation. Analysis of Foxp3 + Treg percentages within a co-culture system of CD4 + CD25 - T-cells and DCregs, with or without FLT3L, demonstrated the involvement of the FLT3/FLT3L axis in driving the differentiation of precursor T-cells into Foxp3 + Tregs induced by DCregs. Cell migration and co-culture assays revealed that the FLT3/FLT3L axis enhanced DCreg migration toward Tregs via the Rho pathway. Additionally, it was observed that DCregs could promote Treg proliferation through the Notch pathway, as inhibition of Notch signaling by DAPT (N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester) suppressed Treg expansion within the co-culture system of DCregs and CD4 + T-cells or CD4 + CD25 + T-cells. Furthermore, the FLT3/FLT3L axis influenced JAG1 expression in DCregs, indirectly modulating Treg expansion. In vivo experiments further established that FLT3L promoted DCreg expansion and restored Treg balance in DSS-induced colitis models, thereby ameliorating colitis symptoms in mice. CONCLUSION The FLT3/FLT3L axis is integral to the maintenance of DCreg function in Treg expansion.
Animals ; T-Lymphocytes, Regulatory/immunology* ; Dendritic Cells/immunology* ; Mice ; Mice, Inbred BALB C ; Membrane Proteins/metabolism* ; Receptors, Notch/metabolism* ; Lymphoid Tissue/metabolism* ; Signal Transduction/physiology* ; Coculture Techniques ; Flow Cytometry

Objective:To construct a nanoparticle vaccine displaying the prefusion F (preF) protein of respiratory syncytial virus (RSV) using the I53-50 protein nanoparticle platform, and to systematically evaluate its immunogenicity and protective efficacy.Methods:The RSV preF trimer antigen was genetically fused to I53-50A and assembled in vitro with I53-50B to form preF-I53-50 nanoparticles, theoretically displaying 20 preF antigens per particle. The structure and purity were characterized by size-exclusion chromatography, SDS-PAGE, and negative-stain electron microscopy. BALB/c mice were intramuscularly immunized with varying doses (1 μg or 5 μg) of preF antigen or an equimolar amount of preF-I53-50 nanoparticles. Humoral immunity, B-cell responses, and protective efficacy were assessed following intranasal viral challenge.Results:The preF-I53-50 nanoparticles self-assembled into spherical structures (50-60 nm in diameter) with uniformly arrayed antigens. The nanoparticle vaccine enhanced RSV-specific IgG1 and IgG2a antibody responses, promoting a Th1-biased immune profile. At equimolar preF doses, the neutralizing antibody titers induced by 1 μg and 5 μg nanoparticle formulations were 2.8-fold and 2.3-fold higher, respectively, than those elicited by preF alone ( P<0.05). Notably, even the low-dose nanoparticle group outperformed the high-dose preF group (1.6-fold increase). Viral challenge experiments demonstrated that preF-I53-50 effectively suppressed pulmonary viral replication, mitigated pathological damage, and induced stronger germinal center and memory B-cell responses, suggesting enhanced B-cell affinity maturation and long-term immune memory. Conclusion:The preF-I53-50 vaccine improves the immunogenicity and protective efficacy of RSV preF through multivalent antigen display.

Objective:To observe the tumor control and the degree of radiation-induced lung injury (RILI) between FLASH irradiation and conventional dose rate (CONV) irradiation, and compare the changes in plasma proteomic profiles of mice following whole-lung FLASH and CONV irradiation using proteomics method.Methods:A mouse model with metastatic lung cancer was established. After whole-lung irradiation, changes in normal lung capacity were monitored using CT scans. Then, a RILI model was constructed to examine pathological alterations in lung tissues following whole-lung CONV and FLASH irradiation. Plasma samples were collected from mice receiving whole-lung CONV irradiation ( n = 5) and whole-lung FLASH irradiation ( n = 5), followed by comparison with samples from the control group of healthy mice (also referred to as the healthy control group). These plasma samples were analyzed using isobaric tags for relative and absolute quantification (iTRAQ)-based proteomics, followed by the screening and identification of differentially expressed proteins using high-throughput bioinformatics. Moreover, protein-protein interaction (PPI) network analysis was conducted to identify hub genes using the STRING database and Cytoscape software. Results:Whole-lung FLASH and CONV irradiation produced consistent tumor control, with the former significantly reducing RILI compared to the latter. A total of 609 proteins were identified through proteomic analysis. Among them, 89 differentially expressed proteins were detected in the whole-lung FLASH group. Gene Ontology (GO) enrichment analysis indicated that up-regulated genes were primarily associated with stress and inflammatory responses, whereas down-regulated genes were related to ATP metabolism and angiogenesis regulation. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed that up-regulated genes were predominantly enriched in unfolded protein response pathways, while down-regulated genes were mainly involved in metabolic pathways and oxidative phosphorylation. Integrated PPI analysis and subsequent validation via reverse transcription-polymerase chain reaction (RT-PCR) revealed four key genes.Conclusions:Compared to the whole-lung CONV irradiation, whole-lung FLASH irradiation reduces the RILI of normal lung tissues while maintaining equivalent tumor control in metastatic lung cancer. Proteomic analysis of differentially expressed proteins in plasma after whole-lung FLASH and CONV irradiation provides valuable insights into the molecular mechanisms underlying the FLASH effect.