BACKGROUND:In recent years,numerous studies have shown that autophagy and mitophagy play an important role in the regulation of bone metabolism.Under non-physiological conditions,mitophagy breaks the balance of bone metabolism and triggers metabolism disorders,which affect osteoblasts,osteoclasts,osteocytes,chondrocytes,bone marrow mesenchymal stem cells,etc. OBJECTIVE:To summarize the mechanism of mitophagy in regulating bone metabolic diseases and its application in clinical treatment. METHODS:PubMed,Web of Science,CNKI,WanFang and VIP databases were searched by computer using the keywords of"mitophagy,bone metabolism,osteoblasts,osteoclasts,osteocytes,chondrocytes,bone marrow mesenchymal stem cells"in English and Chinese.The search time was from 2008 to 2023.According to the inclusion criteria,90 articles were finally included for review and analysis. RESULTS AND CONCLUSION:Mitophagy promotes the generation of osteoblasts through SIRT1,PINK1/Parkin,FOXO3 and PI3K signaling pathways,while inhibiting osteoclast function through PINK1/Parkin and SIRT1 signaling pathways.Mitophagy leads to bone loss by increasing calcium phosphate particles and tissue protein kinase K in bone tissue.Mitophagy improves the function of chondrocytes through PINK1/Parkin,PI3K/AKT/mTOR and AMPK signaling pathways.Modulation of mitophagy shows great potential in the treatment of bone diseases,but there are still some issues to be further explored,such as different stages of drug-activated mitophagy,and the regulatory mechanisms of different signaling pathways.

BACKGROUND: G protein-coupled receptor kinase 2 (GRK2) could participate in the regulation of diverse cells via interacting with non-G-protein-coupled receptors. In the present work, we explored how paroxetine, a GRK2 inhibitor, modulates the differentiation and activation of immune cells in rheumatoid arthritis (RA). METHODS: The blood samples of healthy individuals and RA patients were collected between July 2021 and March 2022 from the First Affiliated Hospital of Anhui Medical University. C57BL/6 mice were used to induce the collagen-induced arthritis (CIA) model. Flow cytometry analysis was used to characterize the differentiation and function of dendritic cells (DCs)/T cells. Co-immunoprecipitation was used to explore the specific molecular mechanism. RESULTS: In patients with RA, high expression of GRK2 in peripheral blood lymphocytes, accompanied by the increases of phosphatidylinositol 3 kinase (PI3K), protein kinase B (AKT), and mammalian target of rapamycin (mTOR). In animal model, a decrease in regulatory T cells (T regs ), an increase in the cluster of differentiation 8 positive (CD8 + ) T cells, and maturation of DCs were observed. Paroxetine, when used in vitro and in CIA mice, restrained the maturation of DCs and the differentiation of CD8 + T cells, and induced the proportion of T regs . Paroxetine inhibited the secretion of pro-inflammatory cytokines, the expression of C-C motif chemokine receptor 7 in DCs and T cells. Simultaneously, paroxetine upregulated the expression of programmed death ligand 1, and anti-inflammatory cytokines. Additionally, paroxetine inhibited the PI3K-AKT-mTOR metabolic pathway in both DCs and T cells. This was associated with a reduction in mitochondrial membrane potential and changes in the utilization of glucose and lipids, particularly in DCs. Paroxetine reversed PI3K-AKT pathway activation induced by 740 Y-P (a PI3K agonist) through inhibiting the interaction between GRK2 and PI3K in DCs and T cells. CONCLUSION Paroxetine exerts an immunosuppressive effect by targeting GRK2, which subsequently inhibits the metabolism-related PI3K-AKT-mTOR pathway of DCs and T cells in RA.
G-Protein-Coupled Receptor Kinase 2/metabolism* ; Arthritis, Rheumatoid/immunology* ; Animals ; Dendritic Cells/metabolism* ; Paroxetine/therapeutic use* ; Proto-Oncogene Proteins c-akt/metabolism* ; Mice ; Humans ; Mice, Inbred C57BL ; Signal Transduction/drug effects* ; Male ; Phosphatidylinositol 3-Kinases/metabolism* ; Lymphocyte Activation/drug effects* ; Female ; T-Lymphocytes/metabolism* ; Middle Aged

The bacterial CBASS(cyclic-oligonucleotide-based anti-phage signaling system) represents a novel innate immune defense mechanism mediated by cyclic nucleotides.The system employs cGAS/DncV-like nucleotidyltransferases (CD-NTases) to recognize exogenous nucleic acids, catalyzing the production of second messenger molecules such as cyclic GMP-AMP (cGAMP) to activate effector proteins (e.g., nucleases), thereby triggering "cell suicide" to combat phage infection. The molecular mechanisms and evolutionary features of CBASS not only uncover the diversity of bacterial immune defenses but also provide critical insights into the conservation of prokaryotic-eukaryotic immune pathways. Notably, the bacterial CBASS system exhibits profound homology with the eukaryotic cGAS-STING (Cyclic GMP-AMP Synthase—Stimulator of Interferon Genes) pathway across multiple dimensions, including the catalytic domains of CD-NTase/cGAS and cyclic nucleotide signaling mechanisms. This finding supports the hypothesis that eukaryotic innate immunity may have originated from prokaryotic horizontal gene transfer and highlights an evolutionary shift in defense strategies from bacterial "population lysis" to "individual inflammatory activation" in higher organisms.This review systematically synthesizes the functional architecture and mechanisms of the CBASS system, with a deep exploration of its evolutionary connections to the eukaryotic cGAS-STING pathway. The insights gained herein offer fresh perspectives for understanding the origin and evolution of the innate immune system, and for driving advancements in biotechnological research and development.

Objective:To assess the efficacy of different fusion strategies involving the convolutional block attention module (CBAM) and Unet for automatic pancreas segmentation in enhanced CT images of patients with acute pancreatitis.Methods:A retrospective analysis was conducted on 1 158 patients with acute pancreatitis admitted to the Affiliated Hospital of North Sichuan Medical College between January 1st, 2016 and July 30th, 2021. Among them, 141 patients with first-episode acute pancreatitis were randomly categorized into mild, moderate, and severe cases. The test set comprised 5 mild and 15 severe cases, while the remaining 126 cases were used for training. Within the training set, 20% of the data was randomly allocated as the validation set. Different fusion paths of the CBAM and Unet networks were trained, utilizing the Dice similarity coefficient, Hausdorff distance (HD), and pixel accuracy (PA) as evaluation metrics. The model demonstrating the best performance on the validation set was selected and evaluated on the test set. Additionally, the Unet model was combined with the attention gate attention mechanism (AttentionUnet) in the skip connection, and the ResBlock replaced the original convolution module (ResUnet) in the Unet network. Moreover, the skip connection branch module of feature extraction was integrated with CBAM (ResUnet_CBAM) for comparison.Results:Unet_CBAM achieved better results on the test set with a Dice value of 80.06%, a HD value of 3.765 9 and a PA value of 0.992 3, all surpassing other fusion strategies. The segmentation accuracy of the pancreatic region in CT images of acute pancreatitis patients was notably enhanced compared to Unet and its related variant networks.Conclusions:The Unet network integrated into CBAM after skip connection can better perform pancreatic segmentation on enhanced CT images of patients with acute pancreatitis and can effectively improve the efficiency of relevant personnel in pancreatic segmentation on enhanced CT images of patients with acute pancreatitis.

ObjectiveTo observe the effect of Zuoguiwan on ovarian reserve in the female offspring rat model of prenatal stress （PS） and explore the mechanism based on Toll-like receptor 4/nuclear factor-κB p65 （TLR4/NF-κB p65） signaling pathway. MethodThirty-two pregnant rats were prepared and randomized into four groups （n=8）： control， model， Zuoguiwan （18.9 mg·kg^-1）， and vitamin E （1.44 mg·kg^-1）. Except the control group， the other three groups were subjected to chronic unpredictable mild stress （CUMS） from day 11 of pregnancy， and the modeling was accompanied by gavage with corresponding drugs until delivery. The PS model was evaluated by the sucrose preference test， open field test， and serum corticosterone （CORT） level. The estrous cycle was monitored and the morphological changes in the ovarian tissue were observed. The serum levels of estradiol （E₂）， luteinizing hormone （LH）， follicle-stimulating hormone （FSH）， and anti-Mullerian hormone （AMH） in the 75-day-old offspring rats were measured by enzyme-linked immunosorbent assay （ELISA） to evaluate the ovarian reserve. The ovary and uterus indices were calculated. The serum levels of interleukin-1β （IL-1β） and tumor necrosis factor-α （TNF-α） were measured by enzyme-linked immunosorbent assay （ELISA）. The morphology of the ovarian tissue in the offspring on the day of birth and day 75 after birth was observed by hematoxylin-eosin staining. The transport of NF-κB p65 to the nucleus in the ovaries of the 75-day-old offspring was detected by the immunofluorescence （IF） assay. The expression of TLR4， NF-κB p65 and other related proteins in the ovarian tissue was determined by Western blot. ResultCompared with the control group， the model group showed reduced primordial follicles in the offspring on the day of birth （P<0.01） as well as disturbed estrous cycle， decreased ovary index and uterus index （P<0.01）， reduced corpus luteum， increased atretic follicles （P<0.01）， lowered serum levels of AMH and E₂ （P<0.01）， elevated serum levels of LH， FSH， IL-1β， and TNF-α （P<0.05， P<0.01）， and up-regulated protein levels of TLR4， NF-κB p65， recombinant myeloid differentiation factor 88 （MyD88）， and phosphorylated NF-κB inhibitor （p-IκBα） （P<0.01） in the 75-day-old offspring rats. Compared with the model group， Zuoguiwan and vitamin E increased the primordial follicles in the offspring on the day of birth （P<0.01）. Moreover， they resumed the estrous cycle， increased the ovary and uterine indices （P<0.05， P<0.01） and corpus luteum （P<0.01）， reduced atretic follicles （P<0.01）， elevated the serum levels of AMH and E₂ （P<0.05， P<0.01）， lowered the serum levels of LH， FSH， IL-1β， and TNF-α （P<0.05， P<0.01）， and down-regulated the expression of TLR4， NF-κB p65， MyD88， and p-IκB-α （P<0.05， P<0.01） in the 75-day-old offspring. ConclusionZuoguiwan can improve the ovarian reserve in the offspring rat model of congenital kidney deficiency by regulating the TLR4/NF-κB p65 signaling pathway.

In the era of evidence-based medicine, it is necessary to explore the unique advantages of traditional Chinese medicine (TCM) based on standardized technical methods and operating procedures in order to achieve the modernization and internationalization of TCM and benefit all humanity. The proposal of a three-pronged evidence system combining TCM theory, human experience and experimental evidence marks an important progress in the thinking method of the TCM evaluation system. The multi-evidence body integrated through appropriate methods provides a strong support for the clinical guideline recommendations and evidence-based health decision-making in TCM. Based on the current methodological progress of international evidence synthesis and grading, this paper proposes a novel approach for integrating multi-evidence in TCM: the MERGE framework. The aim is to establish a solid foundation for the development of this methodology and provide guidance for the advancement of evidence-based medicine framework in TCM.

Objective:To analyze the consistency and repeatability of ultrasonography in diaphragm function evaluation. Method:Sixty healthy subjects participated in the present study,using a PA12A portable color doppler untra-sound diagnostic system.The diaphragm data were recorded during calm breathing and deep breathing,includ-ing diaphragm muscle mobility during calm breathing and deep breathing,diaphragmatic thickness during calm exhalation and calm inspiration,and diaphragmatic thickness during deep exhalation and deep inspiration.The in-tra-test reliability,inter-tester reliability and retest reliability of the above indexes were analyzed respectively. Result:For intra-test reliability,the intra-class correlation coefficient(ICC)values for diaphragm muscle mo-bility during calm breathing,diaphragm muscle mobility during deep breathing,diaphragmatic thickness during calm exhalation,diaphragmatic thickness during calm inspiration,diaphragmatic thickness during maximal exha-lation and diaphragmatic thickness during maximal inspiration were 0.99,0.99,0.99,0.99,0.99,0.99 and 0.99,respectively.The ICC values for the six indicators were 0.84,0.80,0.82,0.82,0.83,and 0.77 for in-ter-tester reliability.For retest reliability,the ICC values for the six indicators were 0.98,0.86,0.98,0.97,0.94,and 0.86 for relative reliability.The standard error of measurement(SEM)for the absolute reliability of the six indicators were 0.09,0.39,0.01,0.02,0.02,0.03,SEM％were 5.23％,8.40％,5.38％,5.7％,8.17％,8.77％,and the minimal detectable change(MDC)were 0.25,1.08,0.03,0.04,0.04,0.09,and MDC％were 14.49％,23.28％,14.9％,15.8％,22.64％,24.31％,respectively.Bland-Altman graphical analy-sis showed no systematic errors. Conclusion:Ultrasound has good reliability in diaphragm function evaluation and can provide objective evi-dence in clinical application.

OBJECTIVE To investigate the activation of xanthine oxidase(XO)from the human liver by vitamin K3 and the mechanism.METHODS Using human liver S9(0.1 g·L-1)as the source,XO was incubated with substrate xanthine of 0,2,4,8,and 16 μmol·L-1 at 37℃ for 90 min.The Michaelis constant(Km)of the reaction of xanthine oxidation was determined using the liquid chromatography diode array method.At the concentration of Km,the three-point method(1,10 and 100 μmol·L-1)was used to detect the activity of vitamin K3 activators.The multi-point method(vitamin K3 1,2,5,10,20,50,100,200 and 400 μmol·L-1)was adopted to determine the half effective concentration(EC50)of activated XO.Kinetic parameters(Km and Vmax)and the fit of double reciprocal curves were determined via vitamin K3 of 1/2EC50,EC50 and 2EC50.The changes in kinetic behavior at different concentrations of vitamin K3 were observed and their types of activation were analyzed.The interactions between XO and activator vitamin K3 were explored via molecular docking.RESULTS The Km of XO-mediated xanthine oxidation reac-tion was 4.71 μmol·L-1.As an activator of this reaction,vitamin K3 activated XO in a concentration-dependent manner(according to the logistic fitting formula y=A2+(A1-A2)/(1+(x/x0)^p),with an EC50 of 32.0 μmol·L-1.The kinetic parameters also changed after the addition of vitamin K3.The Km value decreased(4.71-1.34 μmol·L-1)with the increase of vitamin K3 concentrations,while the Vmax value increased(0.08-1.31 μmol·min-1·g-1),leading to an increase in Vmax/Km(17.0-977.6 mL·min·g-1).In addition,the double reciprocal curve fitting found that the activation type of vitamin K3 on XO was mixed.The molecular docking results showed that vitamin K3 bound to the molybdopterin domain of XO and maintained hydrogen bonding interactions with Arg599 and Ser605.CONCLUSION Vitamin K3 is an activator of XO,which can form hydrogen bonds with Arg599 and Ser605 in the XO domain,regu-late its affinity with the substrate xanthine,activate XO and increase the uric acid level.

Objective To investigate the inhibitory effects of pyruvate-ferredoxin oxidoreductase(PFOR)by luteolin and its anti-Clostridium difficile effect.Methods The PFOR encoding sequence of Clostridium difficile was cloned into the expression vector pET-2a and transformed into competent Escherichia coli.The crude enzyme was prepared after induction with IPTG(Isopropyl β-D-Thiogalactoside).The inhibitory rate of the test compounds on PFOR was determined after an 8-hour anaerobic reaction between PFOR and 40 μmol·L-1 of test compounds at 25℃.The minimum inhibitory concentration(MIC)of PFOR inhibitors against C.difficile strains(ATCC BAA 1382 and ATCC BAA 1870)was determined by monitoring the OD600 of the bacterial culture.Molecular docking was performed to investigate the possible interaction mechanisms between PFOR and inhibitors.Results Among the tested compounds,the luteolin showed the strongest inhibitory activity against PFOR,with a single-point inhibition rate of approximately 33%,which is comparable to that observed with the positive inhibitor nitazoxanide(40%).Molecular docking revealed that luteolin could form hydrogen bonds with Asp428,Val431,Gly429,Asp456,Lys458,Lys459,and other residues in the PFOR domain.The MIC of luteolin against C.difficile was approximately 32 μg·mL-1.Conclusion Luteolin exhibits good activity against C.difficile,and PFOR may be a target for its antibacterial action.