ObjectiveTo investigate the optimal ratio of goat horn replacing antelope horn in Fufang Lingjiao Jiangya pills and the blood pressure-lowering mechanism of this medicine. MethodsThe blood pressure-lowering efficacy of Fufang Lingjiao Jiangya pills with varying proportions of goat horn replacing antelope horn was evaluated on spontaneous hypertensive rats （SHR）. In this experiment， 50 SHR rats were randomly grouped as follows： model （n=8）， captopril （0.01 g·kg^-1） （n=6）， low-dose blank Fufang Lingjiao Jiangya pills （0.342 g·kg^-1） （n=6）， high-dose blank Fufang Lingjiao Jiangya pills （0.684 g·kg^-1） （n=6）， low-dose antelope horn-containing Fufang Lingjiao Jiangya pills （0.378 g·kg^-1） （n=6）， high-dose antelope horn-containing Fufang Lingjiao Jiangya pills （0.756 g·kg^-1） （n=6）， low-dose goat horn-containing Fufang Lingjiao Jiangya pills （0.378 g·kg^-1） （n=6）， and high-dose goat horn-containing Fufang Lingjiao Jiangya pills （0.756 g·kg^-1） （n=6）. Additionally， 8 WKY rats were used as the normal group. Drugs were administered by gavage for 4 weeks while an equal volume of distilled water was administered for the normal and model groups. Blood pressure was measured before administration， 3 h post administration， and biweekly thereafter. In the experiment for Fufang Lingjiao Jiangya pills with goat horn replacing antelope horn in different proportions， 48 SHR rats were randomly grouped as follows： model， blank Fufang Lingjiao Jiangya pills （0.684 g·kg^-1）， antelope horn-containing Fufang Lingjiao Jiangya pills （0.756 g·kg^-1）， 2× goat horn-containing Fufang Lingjiao Jiangya pills （0.824 g·kg^-1）， 4× goat horn Fufang Lingjiao Jiangya pills （0.969 g·kg^-1）， and 6× goat horn Fufang Lingjiao Jiangya pills （1.112 g·kg^-1）. The normal group included 8 WKY rats， and the normal group and model group received an equal volume of distilled water. The treatment lasted for 2 weeks， and blood pressure was recorded at various time points （pre-administration， 3 h post administration， and on days 4， 7， 10， and 14 of administration）. Serum levels of angiotensin-converting enzyme （ACE）， angiotensin Ⅱ（Ang Ⅱ）， renin， and interleukin-6 （IL-6） were measured by enzyme-linked immunosorbent assay. Histopathological changes in the heart， kidney， and thoracic aorta were observed by hematoxylin-eosin staining. The protein levels of ACE2， angiotensin Ⅱ type 1 receptor （AT1R）， and angiotensinogen （AGT） in the kidney tissue were determined by Western blot， while the expression of nuclear factor （NF）-κB p65 and Toll-like receptor 4 （TLR4） in the thoracic aorta tissue was assessed by immunohistochemistry. ResultsCompared with the model group， all treatment groups showed lowered blood pressure （P<0.05， P<0.01）， and the 6× goat horn-containing Fufang Lingjiao Jiangya pills group showed consistent blood pressure-lowering effect with the antelope horn-containing Fufang Lingjiao Jiangya pills group. Compared with the normal group， the model group showed elevated serum levels of ACE， Ang Ⅱ， renin， and IL-6， while the elevations were declined in the Fufang Lingjiao Jiangya pills groups （P<0.05， P<0.01）. Pathological changes in the heart， kidney， and thoracic aorta were alleviated in all the treatment groups， with the 6× goat horn- and antelope horn-containing Fufang Lingjiao Jiangya pills groups exhibited the best effect. Western blot and immunohistochemistry results showed that all the treatment groups exhibited down-regulated protein levels of AT1R， AGT， NF-κB p65， and TLR4 and up-regulated protein levels of ACE2 （P<0.05， P<0.01） compared with model group， with the 6×goat horn- and antelope horn-containing Fufang Lingjiao Jiangya pills groups showcasing the best effect. ConclusionReplacing antelope horn with 6×goat horn in Fufang Lingjiao Jiangya pills can achieve consistent blood pressure-lowering effect with the original prescription. The prescription may exert the effect by inhibiting the renin-angiotensin-aldosterone system （RAAS） and TLR4/NF-κB signaling pathways.

OBJECTIVES: To investigate the effect of Scleromitrion diffusum on gastric mucosal epithelial-mesenchymal transition (EMT) in rats with gastric precancerous lesion. METHODS: Fifty SD rats were randomly divided into blank control group (n=11), model control group (n=13), Scleromitrion diffusum (SD) group (n=13) and vitase group (n=13). Gastric precancerous lesion animal model was prepared by 1-methyl-3-nitro-1-nitrosoguanidine complex polyfactor method, and the drugs were administrated by gavage once a day for 6 weeks. The pathological changes of gastric mucosa were observed with hematoxylin and eosin staining, the expression of EMT marker proteins were detected with immunohistochemical staining and Western blotting. RESULTS: Compared with the model control group, the gastric mucosal injury was significantly attenuated in the Scleromitrion diffusum group, the mucosal tissue structure gradually recovered, the saccular expansion area was reduced, and the inflammatory infiltration was ameliorated. The expression of epithelial cadherin was higher, and the expression of neural cadherin and vimentin in the Scleromitrion diffusum group were lower than those of model control group (all P<0.05). CONCLUSIONS Scleromitrion diffusum can ameliorate gastric mucosal injury in rats with gastric precancerous lesion by reversing the EMT.
Animals ; Rats ; Rats, Sprague-Dawley ; Epithelial-Mesenchymal Transition/drug effects* ; Precancerous Conditions/metabolism* ; Gastric Mucosa/metabolism* ; Stomach Neoplasms/drug therapy* ; Male ; Cadherins/metabolism*

Sjögren's syndrome (SS) is an autoimmune disease characterized primarily by oral and periocular dryness. Astragalus-Salvia (AS) and Ophiopogon-Dendrobium (OD) represent two frequently utilized herb pairs in SS treatment. While the combination of AS-OD herb pairs demonstrates clinical efficacy in alleviating SS symptoms, its underlying mechanism remains unclear. This investigation sought to assess the therapeutic effects and elucidate the potential mechanisms of AS-OD in non-obese diabetic (NOD)/Ltj mice with SS. The study utilized NOD/Ltj mice as SS models, administering AS-OD treatment for 10 weeks at doses of 113.1, 226.2, and 339.3 mg·d^-1·20 g^-1. Results demonstrated that AS-OD improved SS symptoms, evidenced by enhanced salivary flow rate, decreased anti-SSA/Ro and anti-SSB/La antibody levels, increased swimming duration, and reduced lactate (LA) and blood urea nitrogen (BUN) levels in NOD/Ltj mice. AS-OD reduced lymphocyte infiltration, enhanced Aquaporin-5 (AQP5) expression in the submandibular gland, decreased inflammatory cytokine levels in the submandibular gland, and reduced the T helper type 17/regulatory T lymphocyte (Th17/Treg) cell ratio in the spleen. Transcriptomic and proteomic analyses indicated AS-OD's involvement in regulating phosphatidylinositol-3-kinase/protein kinase B (PI3K/AKT) and Janus kinase 3/signal transducer and activator of transcription 3 (JAK1/STAT3) pathways, with inhibitory effects validated in both NOD/Ltj mice submandibular gland and A-253 cells. Furthermore, AS-OD enhanced cell viability and reduced A-253 cell apoptosis through the PI3K/AKT pathway. In A-253 cells, AS-OD reduced inflammatory cytokine levels, CXC chemokine ligand 9/10 (CXCL9/10), and T-cell chemotaxis by inhibiting the JAK1/STAT3 pathway. AS-OD mitigates SS by suppressing inflammation and immune responses through the PI3K/AKT and JAK1/STAT3 pathways.
Animals ; STAT3 Transcription Factor/genetics* ; Sjogren's Syndrome/immunology* ; Mice, Inbred NOD ; Proto-Oncogene Proteins c-akt/genetics* ; Phosphatidylinositol 3-Kinases/genetics* ; Mice ; Drugs, Chinese Herbal/administration & dosage* ; Signal Transduction/drug effects* ; Janus Kinase 1/genetics* ; Humans ; Female ; Astragalus Plant/chemistry* ; Male

Knee osteoarthritis (KOA) is a common chronic degenerative disease that not only causes pain and reduces the quality of life for patients but also imposes a significant societal burden. Clinical pathways can be developed by referencing recommendations from clinical practice guidelines to localize guidelines within the context of integrated traditional Chinese and western medical systems. However, existing clinical pathways suffer from shortcomings such as deficiencies in integrated traditional Chinese and western medical diagnosis and treatment, inadequate shared decision-making between healthcare providers and patients, and suboptimal visualization of clinical pathways. This study aimed to address and optimize the clinical pathway of KOA by comprehensively organizing and localizing the recommended guidelines. The concept of integrated traditional Chinese and western medicine was reflected through the construction of a path of joint decision-making between doctors and patients, emphasizing the coexistence of diagnosis and screening, the combination of clinical and imaging staging, joint decision-making between doctors and patients, and treatment stages. This pathway emphasizes patient-centered approach, with pain relief and functional rehabilitation running parallel, achieving the implementation of evidence-based concepts in practical medical practice. It provides a concrete basis for joint decision-making between doctors and patients in the integrated treatment of KOA with traditional Chinese and western medicine, which helps to improve diagnosis and treatment efficiency and patient quality of life.

Objective:To evaluate the dynamic changes in fibrinolytic states after initial resuscitation in severe trauma patients, and to analyze the relationship between the changes and clinical outcomes.Methods:A prospective cohort study was conducted on severe trauma patients admitted to the trauma center in Taicang Hospital, affiliated with Soochow University, from July 2021 to December 2022. Resuscitation treatments, including tranexamic acid (TXA), were administered. Thromboelastography was performed at three intervals: upon admission, 1 hour and 8 hours after initial resuscitation. Fibrinolytic states were categorized into three phenotypes based on clot lysis at 30 minutes: fibrinolysis shutdown (SD), physiologic fibrinolysis (PY), and hyperfibrinolysis (HF). The primary outcomes included all-cause mortality at 24 hours and 28 days. Multivariate logistic regression was used to analyze the association between early fibrinolytic changes and clinical outcomes.Results:A total of 132 patients with severe trauma were included. Upon admission, fibrinolytic phenotypes were distributed as follows: SD in 61 patients (46.2%), PY in 59 patients (44.7%), and HF in 12 patients (9.1%). After resuscitation with TXA and other interventions, SD and PY remained predominant, whereas HF further decreased. Compared with the SD and PY groups, the HF group had significantly higher 24-hour mortality (25.0% vs. 3.3% vs. 3.4%, P<0.05) and 28-day mortality (58.3% vs. 32.8% vs. 11.9%, P<0.05), with massive hemorrhage being the primary cause of death. Among the non-HF groups, 28-day mortality was significantly higher in the SD group than in the PY group (32.8% vs. 11.9%, P<0.05), with traumatic brain injury as the leading cause of death. After the exclusion of 12 HF patients, multivariate logistic regression showed that after adjusting for age, Glasgow Coma Scale score ≤ 8, prothrombin time, and 24-hour crystalloid infusion volume, identified persistent SD was a risk factor for 28-day mortality in severe trauma patients, compared with sustained PY status ( OR=7.009, 95% CI: 1.141-43.079, P=0.036). Conclusions:In patients with severe trauma, SD and PY are the predominant fibrinolysis phenotypes after initial and early resuscitation. Persistent SD following resuscitation is significantly associated with an increased risk of 28-day mortality.

Objective:To compare the effects of different test meals on postprandial triglycerides and to optimize the standard meal composition and the blood sampling protocol for the oral fat tolerance test.Methods:This study is a prospective, open-label, randomized, cross-over trial. In March 2023, 36 volunteers were recruited in Hebei General Hospital. They underwent a health examination and oral glucose tolerance test. Twenty-six healthy volunteers(11 males and 15 females) were included in this study, with an average age of(39.08±4.56) years. Each volunteer received 75 g protein meal, 75 g fat meal, 700 kcal fixed-calorie high-fat mixed meal, and a high-fat mixed meal with energy adjusted based on 10 kcal/kg body weight. A one-week washout period of regular diet was applied before each trial. Blood was collected at fasting status and 1, 2, 3, 4, 5, and 6 hours after a meal to detect serum triglycerides, total cholesterol, low density lipoprotein-cholesterol(LDL-C), high density lipoprotein-cholesterol(HDL-C), glucose, and insulin. The variations of postprandial metabolic indicators over time following the consumption of different test meals were analyzed. The disparities in postprandial metabolic responses between the two types of mixed meals were compared.Results:The protein meal, fat meal, fixed-calorie high-fat mixed meal, and adjusted-calorie high-fat mixed meal resulted in postprandial triglyceride increases of 22.45%, 115.40%, 77.14%, and 63.63%, and insulin increase of 560.43%, 85.69%, 554.18%, and 598.97%, respectively, and with reductions in total cholesterol, LDL-C, and HDL-C ranging from 5.64%-21.81%, respectively. The blood glucose changed slightly. Changes in metabolic indicators mainly occured within 4 hours. The comparison of the characteristics of postprandial triglycerides between the two high-fat mixed meals showed no statistically significant differences( P>0.05). Conclusion:A standardize protocol with a 700 kcal fixed-calorie high-fat mixed meal as test meal, and blood lipid levels measured at fasting and at 1, 2, 3, and 4 hours after consumption, can serve as an optimized approach for oral fat tolerance test.

AIM:To identify transcriptional differences between the ocular surface ectoderm(OSE)and surface ectoderm(SE)using RNA-seq, and elucidate the OSE transcriptome landscape and the regulatory networks involved in its development.METHODS:OSE and SE cells were differentiated from human embryonic stem(hES)cells. Differentially expressed genes(DEGs)between OSE and SE were analyzed using RNA-seq. Based on the DEGs, we performed gene ontology(GO)analysis, Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analysis, and protein-protein interaction(PPI)network analysis. Transcription factors(TFs)and hub genes were screened. Subsequently, TF-gene and TF-miRNA regulatory networks were constructed using the NetworkAnalyst platform.RESULTS:A total of 4 182 DEGs were detected between OSE and SE cells, with 2 771 up-regulated and 1 411 down-regulated genes in OSE cells. GO-BP analysis revealed that up-regulated genes in OSE were enriched in the regulation of ion transmembrane transport, axon development, and modulation of chemical synaptic transmission. Down-regulated genes were primarily involved in nuclear division, chromosome segregation, and regulation of cell cycle phase transition. KEGG analysis indicated that up-regulated genes in OSE cells were enriched in signaling pathways such as cocaine addiction, axon guidance, and amphetamine addiction, while down-regulated genes were enriched in proteoglycans in cancer, ECM-receptor interaction, protein digestion and absorption, and cytokine-cytokine receptor interaction. Additionally, compared with SE, 204 TFs(including FOS, EGR1, POU5F1, SOX2, and PAX6)were up-regulated, and 80 TFs(including HAND2, HOXB6, HOXB5, HOXA5, and HOXB8)were down-regulated in OSE cells. Furthermore, we identified 6 up-regulated and 9 down-regulated hub genes in OSE cells, and constructed TF-gene and TF-miRNA regulatory networks based on these hub genes.CONCLUSIONS:The transcriptome characteristics of OSE and SE cells were elucidated through RNA-seq analysis. These findings may provide a novel insight for studies on the development and in vitro directed induction of OSE and corneal epithelial cells.

Rheumatoid arthritis (RA) is an autoimmune disease with a complex etiology. Monocyte-derived macrophages (MDMs) infiltration are associated with RA severity. We have reported the deletion of G-protein-coupled receptor kinase 2 (GRK2) reprograms macrophages toward an anti-inflammatory phenotype by recovering G-protein-coupled receptor signaling. However, as more GRK2-interacting proteins were discovered, the GRK2 interactome mechanisms in RA have been understudied. Thus, in the collagen-induced arthritis mouse model, we performed genetic GRK2 deletion using GRK2^f/fLyz2-Cre^+/- mice. Synovial inflammation and M1 polarization were improved in GRK2^f/fLyz2-Cre^+/- mice. Supporting experiments with RNA-seq and dual-luciferase reporter assays identified peroxisome proliferator-activated receptor γ (PPARγ) as a new GRK2-interacting protein. We further confirmed that fms-related tyrosine kinase 1 (Flt-1), which promoted macrophage migration to induce angiogenesis, was inhibited by GRK2-PPARγ signaling. Mechanistically, excess GRK2 membrane recruitment in CIA MDMs reduced the activation of PPARγ ligand-binding domain and enhanced Flt-1 transcription. Furthermore, the treatment of mice with GRK2 activity inhibitor resulted in significantly diminished CIA pathology, Flt-1⁺ macrophages induced-synovial inflammation, and angiogenesis. Altogether, we anticipate to facilitate the elucidation of previously unappreciated details of GRK2-specific intracellular signaling. Targeting GRK2 activity is a viable strategy to inhibit MDMs infiltration, affording a distinct way to control joint inflammation and angiogenesis of RA.

Objective To generate minichromosome maintenance protein 2(MCM2)gene knockout cervi-cal cancer HeLa cell lines using inducible CRISPR/Cas9 technology,and to explore the effect of MCM2 on DNA replication and replication stress.Methods The inducible CRISPR/Cas9 system,TLCV2,was used to construct MCM2 knockout HeLa cell lines.And the cell lines were divided into control group(Control),knockout group 1(KO1),and knockout group 2(KO2).Western blot,Edu incorporation experiment,real-time quantitative PCR(qPCR),immunofluorescence and MTT assay were used to analyze the effects of MCM2 knockout on DNA replica-tion and replication stress induced by hydroxyurea.Results The CRISPR/Cas9 system successfully knocked out the MCM2 gene after induction,and MCM2 knockout affected the stability of MCM2-7 complex.Compared with the control cells,MCM2 knockout cells had a dramatic decrease in the capacity of DNA replication,and the mRNA levels of Cyclin A1,Cyclin E1 and CDK4.Under DNA replication stress,MCM2 knockout cells decreased cell viability,DNA damage repair capacity,and increased genomic instability compared with control cells.Conclusion Knockout of MCM2 gene reduces the DNA replication capacity of HeLa cells under normal conditions and cell viability under replication stress.This study successfully generates MCM2 gene inducible knockout HeLa cell lines,laying the foundation for further research on the role and biological function of MCM2 gene in the occurrence and progression of cervical cancer.

BACKGROUND:Mesenchymal stem cell-derived exosomes may play a crucial role in tissue damage repair,and miRNA is an important component of exosomes for therapeutic effects.Among them,miR-29b-3p has the effect of reducing cell apoptosis,promoting axonal regeneration,and angiogenesis. OBJECTIVE:To study the protective effect of adipose-derived mesenchymal stem cell-derived exosome via miR-29b-3p on a neural cell injury model simulated by H2O2-treated PC12 cells,and explore the relevant mechanisms. METHODS:(1)First,the collagenase digestion method was used to extract rat adipose-derived mesenchymal stem cells.Adipose-derived mesenchymal stem cells were transfected with miR-29b-3p mimics and inhibitors.Exosomes were extracted from the culture supernatant by ultracentrifugation and identified so as to construct adipose-derived mesenchymal stem cell-derived exosomes with high expression and knockdown miR-29b-3p.(2)By constructing a neural cell injury model simulated by PC12 cells treated with H2O2,the relevant mechanisms of the protective effect of adipose-derived mesenchymal stem cell-derived exosome via miR-29b-3p on the simulated neuronal cell injury model were studied. RESULTS AND CONCLUSION:(1)Adipose-derived mesenchymal stem cell-derived exosome had a typical cup-shaped shape and a diameter distribution in the range of 50-140 nm,expressed membrane proteins Alix,CD63,and TSG101,which were specific markers on the surface of exosomes,and could be successfully ingested by PC12 cells.(2)Adipose-derived mesenchymal stem cell-derived exosome pretreatment could reduce cell apoptosis induced by H2O2 treatment in PC12 cells,and this protective effect was enhanced with the increase of miR-29b-3p expression in the exosomes and weakened with the decrease of miR-29b-3p expression in the exosomes.The mechanism of its effect was related to adipose-derived mesenchymal stem cell-derived exosome via miR-29b-3p promoting the expression of anti-apoptotic protein Bcl-2 and inhibiting the expression of apoptotic protein Bax.