Osteoarthritis （OA） and stroke are common clinical diseases that reduce patients' quality of life and place a burden on families and society. Ruyi Zhenbaowan， a classic prescription in Tibetan medicine， have the functions of clearing heat， awakening the brain and opening orifices， relaxing tendons and promoting meridian circulation， and eliminating yellow water. Clinically， they are used to treat osteoarthritis， post-stroke sequelae， neuropathic pain， and other related conditions. Modern pharmacological studies have demonstrated their anti-inflammatory， analgesic， and nerve-repairing effects. However， current research remains insufficient regarding the appropriate indications， timing， and efficacy of this medicine in treating relevant diseases. To enhance clinicians' understanding of this medicine and promote its standardized and rational clinical use， a panel of national experts， including clinical specialists， Tibetan medicine practitioners， pharmacologists， and methodologists， formulated this consensus based on clinical experience and evidence-based practice. The Cochrane systematic review framework， the Grading of Recommendations Assessment， Development and Evaluation （GRADE） system， and the nominal group method were employed to generate seven graded recommendations and 19 consensus-based suggestions. These recommendations clearly define the key points in the clinical application of Ruyi Zhenbaowan， including therapeutic indications， dosage and administration， treatment duration， and medication safety. The consensus specifically addresses the clinical efficacy， appropriate timing of administration， dosage strategies， treatment cycles， and combination medication strategies for treating osteoarthritis and stroke and provides an overview of safety considerations. The aim is to provide standardized guidance for hospitals and healthcare institutions nationwide to ensure the rational application of Ruyi Zhenbaowan in the treatment of osteoarthritis and stroke， reduce medication-related risks， and further leverage its clinical advantages. This consensus has been approved and issued by the China Association of Chinese Medicine， with the standard number GS/CACM 369-2024.

OBJECTIVE: To investigate the correlation between platelet alloantibodies and CD8⁺ T cell with platelet desialylation and apoptosis in platelet transfusion refractoriness(PTR). METHODS: The expression of RCA-1, CD62P and Neu1 on platelets were detected in 135 PTR patients and 260 healthy controls. The ability of PTR patients' sera with anti-HLA antibody, anti-CD36 antibody and antibody-negative groups to induce platelet desialylation and apoptosis, and the potential effect of FcγR inhibitors on desialylation and apoptosis were evaluated. Additionally, the association between CD8⁺ T cells and platelet desialylation in patients was analyzed. RESULTS: The expression of RCA-1 and Neu1 on platelets in PTR patients were significantly higher than those in healthy donors（P < 0.05）, but were not related to platelet alloantibody (P >0.05). The sera of PTR patients generally induced platelet desialylation in vitro （P < 0.05）, with no significant differences among the groups（P >0.05）. However, the sera with anti-CD36 antibodies could induce platelet apoptosis significantly higher than that in the anti-HLA antibody group and antibody-negative group in vitro (P < 0.05). In PTR patients with anti-CD36 antibodies, platelet apoptosis was dependent on FcγR signaling, while desialylation is not. Moreover, CD8⁺ T cells in PTR patients were significantly associated with platelet desialylation (P < 0.05). CONCLUSION Platelet desialylation is a common pathological phenomenon in PTR patients， which involves the participation of CD8⁺ T cell, but isn't associated with platelet alloantibody; while anti-CD36 antibodies have potential clinical significance in predicting platelet apoptosis in PTR patients.
Humans ; Apoptosis ; CD8-Positive T-Lymphocytes/immunology* ; Blood Platelets/metabolism* ; Platelet Transfusion ; Isoantibodies ; Male ; Female ; Middle Aged

Ischemic stroke (IS) is a globally life-threatening disease. Presently, few therapeutic medicines are available for treating IS, and rt-PA is the only drug approved by the US Food and Drug Administration (FDA) in the US. In fact, many agents showing excellent neuroprotection but no blood flow-improving activity in animals have not achieved ideal clinical efficacy, while thrombolytic drugs only improving blood flow without neuroprotection have limited their wider application. To address these challenges and meet the huge unmet clinical need, we have designed and identified a novel compound AAPB with dual effects of neuroprotection and cerebral blood flow improvement. AAPB significantly reduced cerebral infarction and neural function deficit in tMCAO rats, pMCAO rats, and IS rhesus monkeys, as well as displayed exceptional safety profiles and excellent pharmacokinetic properties in rats and dogs. AAPB has now entered phase I of clinical trials fighting IS in China.

Disrupted bone morphogenetic protein type 2 receptor (BMPR2) signaling in endothelial cells drives pulmonary arterial hypertension (PAH). However, targeted recovery of this signaling pathway by lipid nanoparticles (LNPs) has not been explored as a therapy. Here, we employed Design of Experiments to optimize the delivery efficiency of LNPs targeting pulmonary endothelial cells developed by our laboratory, resulting in a remarkable 35-fold increase in a simplified three-component formulation without helper lipids. Administration of BMPR2 mRNA LNPs effectively reversed established PAH in two experimental rat models (monocrotaline or SU5416-hypoxia) by reversing pulmonary vascular remodeling. Specifically, BMPR2 mRNA LNPs replenished the expression of BMPR2 protein and subsequently activated downstream pathways, as confirmed by elevated levels of p-SMAD1/5/9 and ID1 proteins. The relief of pulmonary arterial occlusion was demonstrated by thinned pulmonary arterial media and decreased proportion of full muscularized vessels. Alleviation of right ventricular hypertrophy was indicated by declined Fulton index, the cross-sectional area of right ventricular cardiomyocytes as well as collagen deposition. Effective recovery of right ventricular function was evidenced by increased pulmonary artery flow acceleration time/pulmonary artery flow ejection time ratio. These findings underscore the potential of restoring BMPR2 signaling through pulmonary endothelial cell-specific LNPs for treating PAH.

OBJECTIVES: To investigate the effects of periodontitis induced by Prevotella nigrescens (Pn) combined with ligation on cognitive functions in mice. METHODS: Twenty-four C57BL/6J mice were randomly divided into control group, ligation group, and ligation + Pn treatment (P+Pn) group. Experimental periodontitis was induced by silk ligation of the first molars followed by topical application of Pn for 6 weeks. After modeling, alveolar bone resorption was assessed using micro-CT and histological analysis. Learning and memory abilities of the mice were evaluated using open field test (OFT), novel object recognition test (NORT), and Morris water maze test (MWM). Seven weeks after the start of modeling, the mice were sacrificed for examining histopathological changes in the hippocampus using HE and Nissl staining. RESULTS: After 6 weeks of molar ligation, micro-CT revealed horizontal alveolar bone resorption and furcation exposure in the mice, and histological analysis showed apical migration of the junctional epithelium, epithelial ridge hyperplasia, and lymphocyte infiltration, and these changes were obviously worsened in P+Pn group. Alveolar bone height decreased significantly in both ligation groups compared to the control group. Cognitive tests showed that the mice in both of the ligation groups traveled shorter distances in OFT, showed reduced novel object preference in NORT, and exhibited longer escape latencies in MWM, and the mice in P+Pn group had significantly poorer performances in the tests. Histologically, obvious neuronal cytoplasmic degeneration, necrosis, nuclear pyknosis, vacuolation, and reduced Nissl bodies and viable neurons were observed in the hippocampal regions of the mice in the two ligation groups. CONCLUSIONS Pn infection aggravates alveolar bone destruction, accelerates necrosis and causes morphological abnormalities of neuronal cells in the hippocampus to reduce cognitive functions of mice with periodontitis.
Animals ; Periodontitis/microbiology* ; Mice ; Mice, Inbred C57BL ; Cognition ; Alveolar Bone Loss ; Hippocampus/pathology* ; Male ; Inflammation ; Maze Learning

OBJECTIVE: Patients with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection frequently develop central nervous system damage, yet the mechanisms driving this pathology remain unclear. This study investigated the primary pathways and key factors underlying brain tissue damage induced by the SARS-CoV-2 beta variant (lineage B.1.351). METHODS: K18-hACE2 and C57BL/6 mice were intranasally infected with the SARS-CoV-2 beta variant. Viral replication, pathological phenotypes, and brain transcriptomes were analyzed. Gene Ontology (GO) analysis was performed to identify altered pathways. Expression changes of host genes were verified using reverse transcription-quantitative polymerase chain reaction and Western blot. RESULTS: Pathological alterations were observed in the lungs of both mouse strains. However, only K18-hACE2 mice exhibited elevated viral RNA loads and infectious titers in the brain at 3 days post-infection, accompanied by neuropathological injury and weight loss. GO analysis of infected K18-hACE2 brain tissue revealed significant dysregulation of genes associated with innate immunity and antiviral defense responses, including type I interferons, pro-inflammatory cytokines, Toll-like receptor signaling components, and interferon-stimulated genes. Neuroinflammation was evident, alongside activation of apoptotic and pyroptotic pathways. Furthermore, altered neural cell marker expression suggested viral-induced neuroglial activation, resulting in caspase 4 and lipocalin 2 release and disruption of neuronal molecular networks. CONCLUSION These findings elucidate mechanisms of neuropathogenicity associated with the SARS-CoV-2 beta variant and highlight therapeutic targets to mitigate COVID-19-related neurological dysfunction.
Animals ; COVID-19/genetics* ; Mice ; Brain/metabolism* ; Apoptosis ; Mice, Inbred C57BL ; SARS-CoV-2/physiology* ; Pyroptosis ; Gene Expression Profiling ; Transcriptome ; Male ; Female

Objective To evaluate the bioequivalence and safety of ezetimibe tablets in healthy Chinese subjects.Methods The study was designed as a single-center,randomized,open-label,two-period,two-way crossover,single-dose trail.Subjects who met the enrollment criteria were randomized into fasting administration group and postprandial administration group and received a single oral dose of 10 mg of the subject presparation of ezetimibe tablets or the reference presparation per cycle.The blood concentrations of ezetimibe and ezetimibe-glucuronide conjugate were measured by high-performance liquid chromatography-tandem mass spectrometry(HPLC-MS/MS),and the bioequivalence of the 2 preparations was evaluated using the WinNonlin 7.0 software.Pharmacokinetic parameters were calculated to evaluate the bioequivalence of the 2 preparations.The occurrence of all adverse events was also recorded to evaluate the safety.Results The main pharmacokinetic parameters of total ezetimibe in the plasma of the test and the reference after a single fasted administration:Cmax were(118.79±35.30)and(180.79±51.78)nmol·mL-1;tmax were 1.40 and 1.04 h;t1/2 were(15.33±5.57)and(17.38±7.24)h;AUC0-t were(1 523.90±371.21)and(1 690.99±553.40)nmol·mL-1·h;AUC0-∞ were(1 608.70±441.28),(1 807.15±630.00)nmol·mL-1·h.The main pharmacokinetic parameters of total ezetimibe in plasma of test and reference after a single meal:Cmax were(269.18±82.94)and(273.93±87.78)nmol·mL-1;Tmax were 1.15 and 1.08 h;t1/2 were(22.53±16.33)and(16.02±5.84)h;AUC0_twere(1 463.37±366.03),(1 263.96±271.01)nmol·mL-1·h;AUC0-∞ were(1 639.01±466.53),(1 349.97±281.39)nmol·mL-1·h.The main pharmacokinetic parameters Cmax,AUC0-tand AUC0-∞ of the two preparations were analyzed by variance analysis after logarithmic transformation.In the fasting administration group,the 90％CI of the log-transformed geometric mean ratios were within the bioequivalent range for the remaining parameters in the fasting dosing group,except for the Cmax of ezetimibe and total ezetimibe,which were below the lower bioequivalent range.The Cmax of ezetimibe,ezetimibe-glucuronide,and total ezetimibe in the postprandial dosing group was within the equivalence range,and the 90％CI of the remaining parameters were not within the equivalence range for bioequivalence.Conclusion This test can not determine whether the test preparation and the reference preparation of ezetimibe tablets have bioequivalence,and further clinical trials are needed to verify it.

Cognitive impairment refers to the abnormality of the hippocampus， cortex and other parts of the brain， which is manifested by the decline of cognitive abilities such as learning， memory and attention. With the increase in people's work pressure and bad living habits， the incidence of cognitive impairment is getting higher and higher， which seriously affects people's normal life. However， there are adverse reactions such as gastrointestinal reactions and extrapyramidal reactions in Western drug treatment for cognitive impairment. Therefore， the development of a drug with relatively minimal adverse reactions is of great significance. Traditional Chinese medicine （TCM） has the characteristics of "multi-component， multi-pathway and multi-target"， and the incidence of adverse reactions is relatively low. Studies have shown that the pathogenesis of cognitive impairment is closely related to oxidative stress， inflammation， apoptosis， autophagy and other processes of neurons in the cerebral cortex and hippocampus. Phosphatidylinositol 3-kinase （PI3K）-protein kinase B （Akt） signal pathway plays an important role in the transmission of intracellular and intracellular signals， and in the regulation of cellular inflammation， apoptosis， autophagy， etc. TCM monomers， TCM extracts， and TCM compounds exert anti-inflammatory， antioxidant， anti-apoptotic and autophagy regulation effects by regulating the PI3K/Akt signaling pathway to improve cognitive impairment. This review first summarized the composition and regulatory process of the PI3K/Akt signaling pathway， and then discussed the research progress on the improvement of cognitive impairment through the improvement of oxidative stress， inflammation， apoptosis and autophagy of neurons. Finally， the recent research status of the regulation of this signaling pathway by TCM extracts， TCM monomers and TCM compounds to improve cognitive impairment was summarized. This study provides a theoretical basis for the future study of new TCM related to cognitive impairment.

The MADS-box gene family is a very important transcriptional regulator gene, which plays a role in the whole growth and development process of plants. The APETALA1 (AP1) gene is considered to play an important regulatory role in the transformation of plant flowering, but also to control the characteristic development of floral organs. Lonicera macranthoides is used as medicine with dry buds and early flowers. Therefore, studying the potential mechanism of AP1 gene in regulating flower organ development can provide a basis for improving its medicinal value by molecular means. To explore the potential mechanism of the AP1 gene in the regulation of floral organ development in L. macranthoides, the full-length cDNA of the AP1 was cloned by reverse transcription PCR (RT-PCR) and named LmMADS4. The results show that the CDS of the LmMADS4 gene is 729 bp and encodes 242 amino acids, and the LmMADS4 protein contains no signal peptide and no transmembrane structure, which is an unstable hydrophilic protein. Through homologous sequence alignment and phylogenetic analysis, LmMADS4 and L. japonica MADS27 protein cluster into one class and are closely related. Finally, the expression pattern and protein interaction pattern of LmMADS4 were analyzed by real-time reverse transcription-PCR (qRT-PCR) and yeast two-hybrid technology. The qRT-PCR showed that LmMADS4 gene was differentially expressed in the stems, leaves and flower bud at different developmental stages, including bud type variety Longhua and common variety Baiyun; and LmMADS4 gene was highly expressed in the flower buds, and with the development of flower buds, LmMADS4 gene was continuously up-regulated in the flower bud variety Longhua, however, the expression level of LmMADS4 in the Baiyun terminal flower bud was lower than that in the late flower bud, but the difference was not significant. The yeast two-hybrid results showed that the bait vector pGBKT7-LmMADS4 was not toxic to yeast strains and had no self-activating activity. LmMADS4 protein interacted with LmSVP1, LmSVP3 and LmSOC1s proteins. This study can provide a theoretical basis for exploring the mechanism of long flower bud stage and corolla non-unfolding at the molecular level and variety improvement of L. macranthoides.

MADS-box protein family are important transcriptional regulatory factors in plant growth and development. The AGAMOUS 12 (AGL12) subfamily is believed to play an important regulatory role in the process of plant flowering transition. To explore the potential mechanism of AGL12 subfamily involved in regulating the flower development of Lonicera macranthoides, quantitative real-time polymerase chain reaction (qRT-PCR), prokaryotic expression and yeast two-hybrid techniques were used to analyze the expression pattern, protein expression, and protein-protein interaction pattern of LmAGL12 based on transcriptome data. The results showed that the LmAGL12 gene contains a 603 bp open reading frame (ORF), encoding 200 amino acids, and the encoded protein was stable and hydrophilic without a transmembrane region and signal peptide. Through homologous sequence alignment and phylogenetic analysis, it was confirmed that LmAGL12 protein belongs to the MADS-box protein family and is closely related to the AGL12 protein of Heracleum sosnowskyi and Daucus carota subsp. sativus. The LmAGL12 gene was cloned into prokaryotic expression vector pET-28a and the recombinant constructs were transformed into Escherichia coli BL21 (DE3), which inducing the target protein successfully. The yeast two-hybrid results showed that LmAGL12 protein interacts with LmSVP protein, LmSOC1 protein and LmAP1 protein, respectively. The qRT-PCR results showed that LmAGL12 gene were differentially expressed in different development stages of flower bud, stem, and leaves of 'Longhua' and 'Baiyun' in L. macranthoides. The LmAGL12 gene showed the highest expression level in the middle stage of the 'Longhua' floral bud; For the 'Baiyun' variety, the relative expression level of LmAGL12 gene is the highest in the stem. This study cloned the LmAGL12 gene in L. macranthoides and analyzed it expression for the first time, enriching the research on flower organ development and providing a research basis for further exploring the molecular mechanisms of long bud stage and non-unfolded corolla in L. macranthoides, as well as for variety improvement.