Objective To investigate the role and regulatory mechanism of LINC00657 in the progression of cervical cancer. Methods Bioinformatics analysis predicted potential binding sites between LINC00657 and miR-30a-5p and between miR-30a-5p and Skp2. These sites were verified by using RNA immunoprecipitation and dual-luciferase reporter experiments. LINC00657, miR-30a-5p, and Skp2 mRNA expression levels in cervical cancer tissues and cell lines were assessed by utilizing RT-qPCR. Western blot analysis was employed to examine the protein levels of Skp2 in cells and subcutaneous xenograft tumor models in nude mice. Immunohistochemistry was applied to analyze Skp2 expression in animal tissues. The cellular processes of cervical cancer cell lines were evaluated through CCK-8, scratch, and Transwell assays. Results LINC00657 and Skp2 presented binding sites for miR-30a-5p. In cervical cancer, LINC00657 and Skp2 showed high expression levels (P<0.05), whereas miR-30a-5p displayed low expression (P<0.05). Functional experiments demonstrated that linc00657 upregulates Skp2 expression, a process that is dependent on its sequestration of miR-30a-5p. Conclusion LINC00657 promoted the malignant progression of cervical cancer by upregulating Skp2 expression through specifically sequestering miR-30a-5p, thereby relieving its inhibitory effect on the target gene Skp2.

Objective To investigate the causes and management strategies for lower limb ischemic necrosis following xenogeneic heterotopic heart transplantation from a multigene-edited pig to a rhesus monkey. Methods A xenogeneic heterotopic heart transplantation was performed on December 16, 2023, at the Institute of Experimental Animals of Sichuan Academy of Medical Sciences & Sichuan Provincial People's Hospital, using a quintuple-gene-edited pig as the donor and a rhesus monkey as the recipient. On postoperative day (POD) 9, the recipient monkey underwent left lower limb amputation due to ischemic necrosis. Blood samples were collected at various time points after transplantation for analysis of hematologic parameters, liver and renal function, myocardial enzymes, and coagulation profiles. Ultrasound and computed tomography (CT) were used to evaluate anastomotic patency and cardiac structure. Immunological assays, including complement-dependent cytotoxicity (CDC) and IgG/IgM antibody detection, combined with clinical observations, were employed to assess rejection type and therapeutic response. Results The recipient monkey survived for 46 days after transplantation. Echocardiography demonstrated preserved biventricular systolic function in the recipient’s native heart, with left ventricular ejection fraction (LVEF) consistently exceeding 50%. In the donor pig heart, left ventricular endocardial thickening was noted on POD 9, followed by right ventricular endocardial thickening on POD 24, while LVEF remained around 35%. No hyperacute or acute rejection was detected immunologically. CDC positivity ranged between 3.4% and 5.1%, with IgG/IgM antibody binding trends consistent with CDC results. Following amputation, the recipient exhibited elevated inflammatory markers, coagulopathy, and reactive thrombocytosis, which later normalized. Immunohistochemical staining of the necrotic limb revealed arterial and venous thrombosis; however, no T-cell or B-cell infiltration was observed in vascular structures, thrombi, nerves, muscles, fascia, or skin tissues, with CD3 and CD20 staining both negative. Conclusion Limb ischemia after xenogeneic heart transplantation may be associated with lower extremity vascular thrombosis triggered by local trauma in the context of transplantation-induced inflammatory activation and coagulation dysfunction. While no clear lymphocyte-mediated rejection was observed, further studies are needed to explore the potential role of non-lymphocyte-mediated immune mechanisms.

Background: Alzheimer’s disease (AD) is the most prevalent neurodegenerative disorder. Cellular drug screening represents a key approach in the preclinical discovery of AD therapeutics. Moreover, selecting an appropriate cellular model is essential for exploring the mechanisms underlying AD pathogenesis. Objective: This study aimed to establish a progressive strategy integrating cell model construction, monomer evaluation, and effective component group-based drug discovery for the development of novel therapeutics targeting AD. Methods: A novel AD12 cell line stably expressing the double-transgenic genes amyloid precursor protein (APP)695/presenilin 1(PS1)-ΔE9 was first constructed and subjected to lipopolysaccharide (LPS) stimulation. The resulting LPS-induced neuroinflammatory response was characterized using metabolomic analysis. Subsequently, 16 alkaloids derived from traditional Chinese medicine(TCM) were screened with the LPS-AD12 cell model. Molecular docking was then performed to assess the binding affinity of benzoylaconitine (BAC) with APP and PS1 proteins. A targeted metabolomics approach focusing on amino acids and the tricarboxylic acid (TCA) cycle was applied to BAC-treated LPS-AD12 cells. Finally, aconite extract was administered to five-familial-AD-mutations(5xFAD) transgenic mice, and therapeutic efficacy was evaluated through phenotypic and metabolomic analyses. Results: LPS stimulation induced neuroinflammation in AD12 cells, characterized by elevated levels of interleukin-6, amyloid-β (Aβ ), and PS1. The primary metabolic pathways affected in AD12 cells included alanine, aspartate, and glutamate metabolism, as well as arginine biosynthesis. Notably, BAC significantly reduced the levels of Aβ and interleukin-6 in LPS-AD12 cells. Furthermore, molecular docking analyses revealed a strong binding affinity of BAC with APP and PS1 proteins. Targeted metabolomics analysis indicated that the pharmacological activity of BAC was primarily associated with its anti-inflammatory effects, modulation of the cholinergic signaling pathway, and regulation of energy metabolism. Behavioral experiments in 5xFAD mice, together with immunofluorescence assessments of brain tissues (amyloid-β, glial fibrillary acidic protein, and ionized calcium-binding adapter molecule 1), demonstrated that aconite extract treatment significantly alleviated AD-related symptoms. In addition, metabolomics-based mechanistic analyses revealed that serine, acetylcholine, ornithine, and gamma-aminobutyric acid exhibited consistent trends in both brain tissue and serum samples from treated mice. Conclusion: This study established a novel AD12 cell model and an effective, reliable LPS-induced cell-based screening system. Additionally, we proposed a new drug discovery strategy integrating cell model construction, monomer evaluation, and effective component group-based drug discovery. This screening system has the potential to accelerate the identification of anti-AD agents and provides a feasible framework for subsequent in-depth investigations into new drug development from TCM, thereby contributing to the modernization of TCM-derived therapeutics.

Bone remodeling is essential for maintaining skeletal integrity and mineral homeostasis and depends on the coordinated activities of osteoclasts, osteoblasts, and osteocytes. Osteoclast-mediated bone resorption must remain closely coupled with osteoblast-mediated bone formation, while osteocytes continuously sense mechanical loading and changes in the local and systemic environment. Reactive oxygen species are involved in these physiological processes, particularly in the regulation of osteoclast differentiation and osteoblast function. Once oxidative stress exceeds the capacity of cellular antioxidant systems, however, this balance may be disturbed, leading to enhanced bone resorption, impaired bone formation, and deterioration of bone quality. In recent years, ferroptosis has attracted increasing attention in studies of bone metabolism. Ferroptosis is characterized by iron-dependent accumulation of lipid peroxides, but ferroptosis-related alterations in bone tissue may occur before obvious cell death becomes detectable. Changes in the labile iron pool, membrane lipid composition, antioxidant capacity, and mitochondrial metabolism can already influence the biological behavior of bone cells at an earlier stage. On this basis, ferroptosis susceptibility may provide a useful way to describe a reversible metabolic state in which cells become increasingly vulnerable to lipid peroxidation while widespread membrane damage and terminal cell death have not yet occurred. Such a state should not be identified by a single molecular marker. A more appropriate assessment would combine changes in intracellular iron handling, lipid peroxidation, antioxidant defense, cell viability, bone-cell function, and the response to ferroptosis-related interventions. Measurements of Fe²⁺ or the labile iron pool, C11-BODIPY oxidation, 4-HNE and MDA, GPX4 activity or expression, GSH/GSSG balance, LDH release, and PI staining may therefore provide complementary information when interpreted together with functional changes and rescue experiments. Several metabolic processes may contribute to the development of ferroptosis susceptibility in the bone. Disturbances in iron uptake, ferritin storage, ferritinophagy, and ferroportin-mediated iron export can increase the availability of redox-active iron. At the same time, ACSL4- and LPCAT3-dependent incorporation of polyunsaturated fatty acids into membrane phospholipids increases the pool of oxidizable lipid substrates. Reduced activity of the system Xc^--GSH-GPX4 pathway further weakens the ability of cells to remove lipid hydroperoxides, while FSP1-CoQ and mitochondrial DHODH-CoQ systems provide additional antioxidant protection. Mitochondrial dysfunction, inflammation, and abnormalities in calcium-phosphate and endocrine regulation may further increase this metabolic pressure. Importantly, the consequences are unlikely to be identical among bone-cell populations. In osteoclasts, increased iron availability and oxidative stress may strengthen RANKL-NFATc1-associated differentiation and resorptive activity. In osteoblasts, persistent lipid peroxidation may interfere with RUNX2-related differentiation, collagen synthesis, matrix production, and mineralization before extensive cell loss occurs. Osteocytes, because of their long lifespan and specialized location within the mineralized matrix, may be particularly sensitive to chronic metabolic stress, resulting in impaired mechanosensing and disturbed RANKL- or FGF23-related feedback. Evidence linking these changes with osteoporosis, diabetes-related bone disorders, chronic kidney disease-mineral and bone disorder, inflammatory bone loss, and particle-induced osteolysis is increasing, although the strength of evidence differs among disease models. Changes in ROS, MDA, GPX4, or other individual indicators alone are insufficient to demonstrate ferroptosis-related susceptibility, and stronger conclusions require evidence of iron dependence, lipid peroxidation, temporal sequence, and reversibility. Emerging single-cell, spatial, proteomic, lipidomic, and metabolomic approaches may help identify susceptible cell populations and bone-remodeling regions more precisely, although the mineralized matrix and the effects of sample processing remain important technical limitations. Overall, ferroptosis susceptibility may represent an intermediate metabolic state linking disturbances in iron and redox homeostasis with progressive uncoupling of bone resorption and formation. Clarifying its cell-specific features, temporal development, and reversible window may improve our understanding of how chronic metabolic stress contributes to bone remodeling disorders and may also provide a basis for earlier and more targeted intervention.

AIM To establish a UPLC-MS/MS method for the simultaneous content determination of isofraxidin,saikosaponin c,ginsenoside Re,puerarin,rosmarinic acid,praeruptorin A,daidzein,baicalin and 5-O-methylvisammioside in Chaige Changyuan Mixture.METHODS The analysis was performed on a 35 ℃ Titank C18 column(100 mm×2.1 mm,3 μm),with the mobile phase comprising of acetonitrile-0.1％formic acid flowing at 0.3 mL/min in a gradient elution manner,and electron spray ionization source was adopted in positive and negative ion scanning with multiple reaction monitoring mode.RESULTS Nine constituents showed good linear relationships within their own ranges(r≥0.999 0),whose average recoveries were 84.08％-115.40％with the RSDs of 0.21％-4.47％.CONCLUSION This efficient,simple,sensitive and specific method can be used for the quality control of Chaige Changyuan Mixture.

Cervical spinal cord injury without fracture-dislocation (CSCIWFD) is referred to as a special type of cervical spinal cord injury characterized by traumatic spinal cord dysfunction and no significant bony structural abnormalities on imagines. Duo to the high risk of missed diagnosis during the initial consultation, CSCIWFD may lead to progressive neurological deterioration or even complete paralysis, severely impacting patients′ prognosis. Currently, there are no established consensuses over the diagnosis and treatment of CSCIWFD, such as the lack of evidence-based standards for indications of non-surgical treatment and risk of secondary neurological injury, as well as debates over the optimal timing for surgical intervention and indications for different surgical approaches. To address these issues, the Spine Trauma Group of the Orthopedic Branch of the Chinese Medical Doctor Association organized experts in the relevant fields to formulate Diagnosis and treatment guideline for acute cervical spinal cord injury without fracture- dislocation in adults ( version 2025) . Based on evidence-based medicine and the principles of scientific rigor and clinical applicability, the guidelines proposed 11 recommendations covering terminology, diagnosis, evaluation treatment, and rehabilitation, etc., aiming to standardize the management of CSCIWFD.

Vertebral refracture following percutaneous vertebral augmentation (PVA) is commonly seen in elderly patients with osteoporotic thoracolumbar compression fractures (OTLCF). It can lead to recurrent pain, loss of vertebral height, progression of kyphosis, and even neurological dysfunction, significantly impairing patients′ quality of life. Current diagnosis and treatment face multiple challenges, including high misdiagnosis rate, difficulty in choosing between surgical and non-surgical treatment options, lack of standardized surgical protocols, interference from intralesional bone cement during procedures, inadequate stability of internal fixation in osteoporotic bone, and suboptimal compliance of anti-osteoporotic therapy. Establishing a standardized diagnostic and therapeutic framework is urgently needed. To standardize the management process and improve outcomes for vertebral refractures after PVA in elderly OTLCF patients, Spinal Trauma Group of the Orthopedic Branch of Chinese Medical Doctor Association organized experts in the field to develop Guideline for the diagnosis and treatment of vertebral refracture after percutaneous vertebral augmentation in elderly patients with osteoporotic thoracolumbar compression fractures ( version 2025), based on current literature and clinical experience, and adhering to principles of scientific rigor and clinical applicability. A total of 11 recommendations were proposed, encompassing diagnosis, treatment, and rehabilitation of vertebral refracture after PVA in elderly patients with OTLCF, aiming to provide a foundation for a standardized management.

Objective:To construct a VP8-mRNA vaccine using human rotavirus spike protein VP8 domain as the immunogen and analyze its immunogenicity in mice.Methods:The VP8-mRNA sequence was designed, optimized, and synthesized. The VP8 gene of rotavirus G1P[8] type was used to construct the plasmid pUC57-VP8-Kan-SapⅠ, which was then sequenced. The plasmid confirmed by sequencing was subjected to large-scale amplification and extraction, followed by linearization, in vitro transcription, and capping. The purified capped products were encapsulated with lipid nanoparticles using a microfluidic control apparatus. The encapsulated VP8-mRNA vaccine was administered intramuscularly to mice at 10, 15, and 20 μg. Serum samples were collected for antibody detection by ELISA. Cellular immune responses were detected by flow cytometry and ELISPOT. Statistical analysis was performed using one-way or two-way analysis of variance and Tukey-Kramer test. Results:The encapsulated VP8-mRNA vaccine was rounded and spherical, with a particle size of about 100 nm, a polymer dispersion index of 0.088, and an encapsulation rate of 92.3%. Two doses of VP8-mRNA vaccine immunization could induce a good immune response in mice. The level of IgG antibody induced after immunization in the 15 μg group was comparable to that of the 20 μg group, and there was no statistical difference ( P>0.05), but the antibody levels in the two groups were significantly higher than that in the 10 μg group ( P<0.000 1). VP8-mRNA vaccine could induce neutralizing antibodies against rotavirus G1 and G9 types. The highest level of neutralizing antibodies against rotavirus type G1 was observed in the 15 μg group, which was significantly higher than that in the 10 μg group ( P<0.05). All immunization groups exhibited good neutralizing ability against rotavirus G9 type. The results of ELISPOT showed that lymphocytes from mice in each vaccine group were able to secrete IFN-γ when stimulated with VP8 peptide. Flow cytometry showed that the proportions of CD8 + T cell subsets in the vaccine groups were higher than that in the control group. Conclusion:The VP8-mRNA vaccine has good immunogenicity in mice and can induce good humoral and T-cell immune responses.

Objective:To investigate the impact of peripheral blood prolymphocyte percentage on the prognosis of patients with chronic lymphocytic leukemia (CLL) .Methods:This study included 300 patients diagnosed with CLL at the Department of Hematology of Jiangsu Provincial People’s Hospital from October 2011 to December 2020. The association between prolymphocyte percentage and other parameters was analyzed, and the optimal cutoff prolymphocyte percentage was determined by X-tile analysis. Further survival analysis and prognostic model construction were used to validate the predictive value of prolymphocyte percentage.Results:Of the 300 eligible patients with CLL who were enrolled, 50 received Bruton tyrosine kinase inhibitors (BTKi) as first-line treatment. The group with higher prolymphocyte percentage comprised more patients in the advanced stages ( P=0.010) and had higher β 2-microglobulin ( P<0.001), unmutated immunoglobulin heavy-chain variable region gene ( P<0.001), and TP53 aberration ( P=0.004). The optimal cutoff percentage of prolymphocytes was 1%. Patients with a prolymphocyte percentage >1% had significantly shorter treatment-free survival (TFS) ( P<0.001) and overall survival time ( P=0.007) than patients with a prolymphocyte percentage ≤1%. On multivariate analysis, prolymphocyte percentage >1% tended to have an independent prognostic value for TFS [ HR=1.405 (95% CI 0.971~2.032), P=0.071]. Compared with the nomogram of CLL international prognostic index (CLL-IPI) alone, the nomogram of CLL-IPI combined with prolymphocyte percentage showed better discrimination (area under the curve: 0.778 vs. 0.637; P=0.006). In addition, patients with a prolymphocyte percentage >1% were more likely to progress after BTKi treatment ( P=0.038) . Conclusion:Peripheral blood prolymphocyte percentage was associated with various clinical and biological parameters and prognosis among patients with treatment-naive CLL.

Objective:To investigate the role of the IgG subtypes (IgG1 and IgG2a) in anti-glycoprotein (GP) Ⅰbα antibody-induced platelet clearance.Methods:Venous blood was collected from healthy volunteers, and platelets were separated. The phagocytosis of human platelets by human acute monocytic leukemia cells (THP-1 cells) induced by different anti-GPⅠbα antibodies (AN51, AK2, HIP1, TM60, VM16d, WM23, and SZ2) was detected by flow cytometry. The effects of the AN51 full-length antibody, F (ab') 2, and Fab fragments on platelet phagocytosis by THP-1 cells were detected by flow cytometry. Then, the Fc blocking antibody 2.4G2 and normal rat IgG2a or IgG1 were injected into C57BL/6J mice via the posterior ocular vein, and their effects on platelet reduction induced by R300 were detected by a hematology analyzer. Results:Compared with IgG1, the IgG2a subtype of anti-GPⅠbα antibodies induced the phagocytosis of platelets by THP-1 cells in vitro ( P<0.05). In contrast to the AN51 full-length antibody, neither AN51 F (ab') 2 nor the Fab fragment could induce THP-1 cells to phagocytose platelets ( P<0.05). Compared with the control group, anti-mouse GPⅠbα R300-induced thrombocytopenia in mice was reduced at 2, 4, and 6 h after the injection of Fc blocking antibody 2.4G2 ( P<0.05). Similarly, R300-induced thrombocytopenia in mice was reduced at 2, 4, and 6 h after the injection of rat IgG2a ( P<0.05) . Conclusion:IgG2a plays an important role in anti-GPⅠbα-induced clearance.