OBJECTIVE To analyze the risk factors associated with serum digoxin concentration （SDC） exceeding the warning threshold and to construct a risk prediction model. METHODS Clinical data were retrospectively collected from hospitalized patients who received regular oral digoxin and completed therapeutic drug monitoring at Guangzhou First People’s Hospital and Nansha Branch of Guangzhou First People’s Hospital between September 2020 and March 2025. Patients with SDC＞2.0 ng/mL were classified as exceeding the warning threshold group， while those with SDC≤2.0 ng/mL were classified as the non-exceeding the warning threshold group. Based on univariate factor analysis， multivariate Logistic regression analysis was used to identify independent risk factors for SDC exceeding the warning threshold. A prediction model was developed and a nomogram was plotted accordingly. The discriminative ability of the model was evaluated by receiver operating characteristic （ROC） curve analysis， and the calibration curve were plotted to assess the calibration of the model. The Hosmer-Lemeshow test was employed to evaluate the goodness of fit of the model， and clinical utility was evaluated by decision curve analysis （DCA）. RESULTS A total of 254 patients were included， among whom 49 patients （19.29%） had SDC exceeding the warning threshold. Univariate factor analysis and multivariate Logistic regression analysis showed that increased daily dose per kilogram of body weight， advanced age， concomitant coronary heart disease， elevated serum creatinine levels， concomitant use of amiodarone， and concomitant use of deslanoside wer e independent risk factors for SDC exceeding the warning threshold （ P ＜0.05）. The area under the ROC curve of the model was 0.869 （95% confidence interval： 0.818-0.920）， with a sensitivity of 0.796 and a specificity of 0.842. The Hosmer-Lemeshow test showed good calibration （ P ＝0.570）. The calibration curve was closely aligned with the ideal curve， with a mean absolute error of 0.012. The model provided a higher net benefit across a threshold probability range of 6% to 82%. CONCLUSIONS The increased daily dose per kilogram of body weight， advanced age， concomitant coronary heart disease， elevated serum creatinine levels， concomitant use of amiodarone， and concomitant use of deslanoside are independent risk factors for SDC exceeding the warning threshold. The nomogram prediction model developed based on the aforementioned factors can be used to predict the risk of SDC exceeding the warning threshold.

Objective To investigate the epidemiological characteristics of cross-county imported dengue fever cases within Yunnan province in 2023, so as to provide insights into formulation of preventive and control measures for intra-provincial spread of dengue fever. Methods All data pertaining cross-county imported dengue fever cases within Yunnan Province in 2023 were collected, and the temporal, regional and population distributions of the cases were descriptively analyzed. Results A total of 1 664 intra-provincial cross-county imported dengue fever cases were reported in 95 counties (cities, districts) cross 16 profectures (cities) in Yunnan Province in 2023, accounting for 12.34% of total cases in the province. Cross-county imported dengue fever cases were predominantly reported during the period between August and October (1 516 cases, 91.11% of total cases), and peaked in September (659 cases), with a single-day peak on October 8 (36 cases). During the period from September 4 to 10, five counties (cities) with local dengue fever epidemics, including Jinghong City of Xishuangbanna Dai Autonomous Prefecture, Gengma Dai and Wa Autonomous County of Lincang City, Ruili City of Dehong Dai and Jingpo Autonomous Prefecture, Mengla Coun ty of Xishuangbanna Dai Autonomous Prefecture, and Zhenkang County of Lincang City, exported 165 cross-county imported dengue fever cases to the rest of the province. Among the 1 644 intra-provincial cross-county imported dengue fever cases, the male to female ratio was 1.40∶1.00, and 1 329 cases were at ages of 15 to 55 years (79.87%), with farmers as the predominant occupation (886 cases, 53.25%). The top 5 counties (cities/districts) reporting the highest number of intra-provincial cross-county imported dengue fever cases included Simao District (266 cases) and Lancang Lahu Autonomous County (118 cases) of Pu’er City, Mengla County (91 cases) and Menghai County (91 cases) of Xishuangbanna Dai Autonomous Prefecture, and Mangshi City (73 cases) of Dehong Dai and Jingpo Autonomous Prefecture, which accounting for 38.40% of total imported cases. These intra-provincial cross-county imported dengue fever cases originated from 7 counties (cities/districts) in 4 prefectures (cities), including 1 261 cases (76.70%) from Jinghong City of Xishuangbanna Dai Autonomous Prefecture, 224 cases (13.63%) from Ruili City of Dehong Dai and Jingpo Autonomous Prefecture, 103 cases (6.27%) from Gengma Dai and Wa Autonomous County of Lincang City, 31 cases (1.89%) from Mengla County of Xishuangbanna Dai Autonomous Prefecture, 30 cases (1.82%) from Zhenkang County of Lincang City, 10 cases (0.61%) from Cangyuan Wa Autonomous County of Lincang City, and 5 cases (0.30%) from Mohan-Boten Economic Cooperation Zone of Kunming City. In addition, local dengue fever epidemics following intra-provincial cross-county importation of dengue fevers cases in Simao District, Jinggu Dai and Yi Autonomous County, Mangshi City, Longchuan County, and Cangyuan Wa Autonomous County. Conclusions Farmers and students are high-risk populations for intra-provincial cross-county imported dengue fever cases in Yunnan Province, and health education pertaining personal protection against dengue fever should be strengthened among these high-risk populations by governments at all levels. There is a high risk of local out-break of dengue fever following continuous introduction of intra-provincial cross-county imported cases. Standardized management of intra-provincial cross-county imported dengue fever cases should be reinforced to reduce the risk of local epidemics.

Objective To investigate the mechanism by which suppressor of cytokine signaling 3(SOCS3)regulates microglial inflammation through nuclear factor-kappaB(NF-κB),providing novel mechanistic insights into microglial involvement in Parkinson's disease(PD)pathogenesis.Methods ① Ten male C57BL/6 mice(12 weeks old,weighing 20～25 g)were subjected to intraperitoneal injection of 15 mg/kg MPTP to establish a PD model.Rotarod test was used to assess motor function.Western blotting was employed to detect the protein expression of tyrosine hydroxylase(TH)and ionized calcium-binding adapter molecule 1(IBA-1)in the substantia nigra.RT-qPCR was utilized to measure the mRNA level of SOCS3 in the substantia nigra.Immunohistochemistry was performed to assess NF-κB p65 subunit expression.The expression of SOCS3,NF-κB and p-NF-κB was measured with Western blotting.② Microglial cell line BV2 was stimulated with 1 000 ng/mL lipopolysaccharide(LPS)for 6 h to establish an inflammatory model.Subsequently,SOCS3 was knocked down.NF-κB inhibitor BAY 11-7082 was used to treat the cells.RT-qPCR and Western blotting were used to measure the expression of SOCS3 at mRNA and protein levels.Western blotting was also applied to detect the expression of NF-κB and p-NF-κB,and ELISA was conducted to measure TNF-α and IL-1β levels in the culture supernatant.Immunofluorescence assay was carried out to localize NF-κB(nuclear vs cytoplasmic).③ A co-culture system of BV2 microglia and N2a neuroblastoma cells was established to investigate the regulatory effects of microglia on neuronal cells.MTT assay and TUNEL staining were used respectively to determine cell viability and apoptosis of N2a cells.Results ① Compared to the control mice,the PD mouse model exhibited reduced rotarod fall latency,down-regulation in TH and SOCS3(P<0.01),up-regulation in IBA-1 and increased p-NF-κB/NF-κB ratio(P<0.01).② In BV2 cells,LPS stimulation increased TNF-α,IL-1β,and p-NF-κB/NF-κB ratio(P<0.01),while down-regulated SOCS3 expression(P<0.01).SOCS3 knockdown in LPS-stimulated BV2 cells further increased the p-NF-κB/NF-κB ratio(P<0.01),increased nuclear localization of NF-κB,and elevated TNF-α and IL-1β levels(P<0.01).BAY 11-7082 treatment in these SOCS3-knockdown,LPS-stimulated cells resulted in reduced p-NF-κB/NF-κB ratio,TNF-α,and IL-1β(P<0.01),and decreased NF-κB nuclear distribution.③ LPS-stimulated BV2 cells reduced cell viability and increased cell apoptosis in N2a cells(P<0.01).SOCS3 knockdown in BV2 cells exacerbated the reduction in N2a cell viability(P<0.01)and the increase in cell apoptosis in N2a cells(P<0.01).BAY 11-7082 treatment of these SOCS3-knockdown BV2 microglia attenuated the reduction in N2a cell viability and decreased apoptosis in N2a cells(P<0.01).Conclusion SOCS3 inhibits microglia inflammatory response through down-regulation of NF-kB activity,and in turn attenuates neuronal cell death and ameliorates PD nerve injury.

Objective To demonstrate that neferine(Nef)alleviates Parkinson's disease(PD)by inhibiting microglial pyroptosis mediated through the reactive oxygen species(ROS)/NOD-like receptor protein 3(NLRP3)/Caspase-1 pathway.Methods BV2 microglial cells were divided into:control group,lipopolysaccharides(LPS)-adenosine triphosphate(ATP)group,and LPS-ATP+Nef group.Pyroptosis was induced by 1 μg/mL LPS+5 mmol/L ATP,with 2 mmol/L Nef pretreatment.Eighteen 10-12-week-old male C57BL/6 mice(22～25 g)were randomly assigned to:control(n=6),1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)(n=6),and MPTP+Nef(n=6)groups.Detection methods included:flow cytometry for pyroptosis,Cell Counting Kit-8(CCK-8)for viability,2',7'-dichlorodihydrofluorescein diacetate(DCFH-DA)for ROS,commercial kits for malondialdehyde(MDA),superoxide dismutase(SOD),glutathione(GSH),ELISA/Western blot for interleukin-1β(IL-1β)/IL-18,immunofluorescence/immunohistochemistry for NLRP3/Caspase-1,tyrosine hydroxylase(TH)immunohistochemistry,hematoxylin-eosin staining for neuropathology,and modified neurological severity score(mNSS).Results Versus control,LPS-ATP group showed decreased viability(P=0.002),increased pyroptosis(P<0.001),elevated ROS(P<0.001)/MDA(P<0.001)/IL-1β(P<0.001)/IL-18(P<0.001),upregulated NLRP3(P<0.001)/Caspase-1(P<0.001),and reduced GSH(P<0.001)/SOD(P<0.001).Nef treatment reversed these effects(all P<0.05).According to the results of murine studies,compared with the control group,the MPTP group had increased mNSS(P<0.001)/tissue ROS(P<0.001),downregulated TH(P<0.001),upregulated NLRP3(P<0.001)/Caspase-1(P<0.001).Nef treatment significantly attenuated the MPTP-induced deleterious effects(P<0.05).Histopathological analysis revealed that control group exhibited uniformly distributed hippocampal neurons with distinct nuclear morphology;MPTP group showed neuronal swelling,interstitial edema,and nuclear atrophy;MPTP+Nef group demonstrated ameliorated neuronal damage.Conclusion Nef inhibits microglial pyroptosis via ROS/NLRP3/Caspase-1 axis,ameliorating PD neuroinflammation and pathology.

ObjectiveTo analyze the temporal trend of knockdown resistance (kdr) gene mutations highly correlated with pyrethroid resistance in field populations of Aedes aegypti in Jinghong City of Yunnan Province, and to provide a scientific basis for formulating rational insecticide use strategies. MethodsAdult mosquito samples of Aedes aegypti from 2016 to 2023 and larvae mosquito samples from July 2022 to June 2023 were collected in Jinghong City of Yunnan Province. Allele specific PCR (AS-PCR) was used to measure kdr mutations at amino acid positions 989, 1016 and 1534 of the voltage-gated sodium ion channel (VGSC) gene. Data such as mutation rate and mutation allele frequency were calculated, SPSS software was used to perform trend chi square tests on mutation rate and mutation allele frequency with year and month, as well as comparison of mutation allele frequencies and genotype distributions between the dry and rainy seasons, thereby delineating the temporal trend of kdr gene mutations. ResultsAmong the 173 samples collected from 2016 to 2023, the mutation rates of S989P and V1016G were 100.00% for each year, while the mutation rate of F1534C ranged from 62.50% to 100.00%. The mutation rate and mutation allele frequency of F1534C were increased over the years (χ²=22.079, P<0.001; χ²=42.971, P<0.001). Concurrently, the proportion of the PPGGCC genotype was increased annually (χ²=60.790, P<0.001). Among the 288 samples collected from July 2022 to June 2023, the monthly mutation rates for S989P, V1016G, and F1534C were consistently 100.00%. There was only one type of mutation present, namely S989P+V1016G+F1534C. In the combinations of the three genotypes, the SPGGCC genotype accounted for 1.39% (4/288), the PPGGFC accounted for 2.78% (8/288), and the PPGGCC had the highest proportion at 95.83% (276/288). After tesiting the samples collected in August 2023, the mutation rates of 989, 1016 and 1534 sites of VGSC in females, males, and larvae of the same generation were all 100.00%. ConclusionSince 2016, the gene mutations at S989P and V1016G loci in the VGSC gene of wild Aedes aegypti in Jinghong City have remained consistently at 100.00%, while the mutation rate and mutant allele frequency of F1534C have increased year by year during the testing period. By 2023, the mutation rates at three loci in the VGSC gene of Aedes aegypti in Jinghong City had all reached 100.00%, and neither changes in insect developmental stage nor gender differences during transmission exerted a detectable impact on the mutation rates. In the control of Aedes aegypti in Jinghong City, the use of pyrethroid insecticides should be stopped or reduced, and regular monitoring of kdr genes should be carried out to promptly detect new mutations.

Objective:To develop a recombinant protein vaccine based on KPC-2, a drug resistance target in Klebsiella pneumoniae, and evaluate its immunogenicity, protective efficacy and mechanism in a mouse model of pneumonia. Methods:KPC-2 was expressed in Escherichia coli and purified using GST affinity chromatography. A recombinant protein vaccine was prepared with KPC-2 and used to immunize New Zealand rabbits through subcutaneous injection. Serum samples were isolated from cardiac blood and Protein G chromatography was used to purify polyclonal antibodies against KPC-2. Opsonophagocytic killing assay was used to assess the bactericidal activity of the polyclonal antibodies in vitro. Female BALB/c mice were immunized three times with the recombinant protein vaccine, and the titers of specific IgG antibodies in serum were measured by indirect ELISA. One week after the last vaccination, the mice were infected with Klebsiella pneumoniae strain SRT through tracheal intubation, and received a single intravenous dose of meropenem (0.1 mg) 1 h later. The protective efficacy of the KPC-2 recombinant protein vaccine was evaluated by comparing the survival rates, bacterial colonization and histopathological changes between vaccine group and adjuvant group as well as the survival rates between meropenem group and normal saline group. Moreover, the protective efficacy of polyclonal antibodies against KPC-2 was evaluated through passive immunization. Results:The level of specific IgG antibodies in serum was significantly higher in the vaccine group than in the adjuvant group ( t=4.325, P<0.05). The survival rate in the vaccine group was also higher than that of the adjuvant group [70% (7/10) vs 10% (1/10), P<0.05]. Furthermore, lung inflammation was less severe and bacterial burden was reduced in the vaccine group as compared with those of the control group ( t=3.127, P<0.05). Both active and passive vaccination strategies demonstrated strong protective efficacy against Klebsiella pneumoniae infection, and had a synergistic effect when used in combination with antibiotic therapy. The polyclonal antibodies against KPC-2 had bactericidal activity in vitro ( t=5.427, P<0.05). Conclusions:The prepared KPC-2 vaccine has better immunogenicity and protective efficacy. It can induce strong humoral immune responses. This study suggest that drug resistance target may be used as a candidate antigen for future vaccine development.

Objective:To evaluate the impact of three small compounds, namely sodium diethyldithiocarbamate (DTC), levamisole (LMS) and imiquimod (Imi), on the immunogenicity and protective efficacy of the candidate antigen PA0833 from Pseudomonas aeruginosa ( Pa) and analyze the underlying mechanisms. Methods:PA0833 was formulated with aluminum adjuvant and the above small compounds, respectively. BALB/c mice were immunized with these vaccines intramuscularly on days 0, 14 and 21. Serum samples were collected and the levels of PA0833-specific IgG were measured by ELISA. The protective efficacy of these vaccines was evaluated by assessment of survival rates, body weights, clinical scores, inflammatory factors, and histopathological changes after infecting the immunized mice with Pa PAO1 strains. Besides, the mice were injected with DTC intramuscularly for seven consecutive days to analyze the mechanism of DTC in enhancing immune response using transcriptome sequencing and flow cytometry. Results:All these small compounds were capable of effectively enhancing the immunogenicity of PA0833 formulated with aluminum adjuvant, reducing bacterial loads in lung tissues, inhibiting the secretion of TNF-α, IL-6 and IL-1β, and improving mouse survival rates upon Pa infection. DTC was more effective than the other two compounds. Transcriptome sequencing identified 121 up-regulated genes and 18 down-regulated genes in DTC-treated group as compared with PBS control group. These differentially expressed genes were significantly enriched in immune pathways, with a strong activation of the IL-17 pathway. Flow cytometry analysis demonstrated significant activation of dendritic cells and proliferation of Th17 cells in splenocytes in DTC-treated group as compared with PBS control group. Conclusions:All three small compounds are able of effectively enhance antigen immunogenicity with DTC being the most effective, indicating that DTC can be used as a novel adjuvant in vaccine development.

The cyclic GMP-AMP synthase(cGAS)-stimulator of interferon genes signaling pathway plays a central role in cells'ability to sense the presence of abnormal double-stranded DNA within the cytoplasm.Modulation of the function of the cytoplasmic DNA recognition receptor cGAS allows regulation of this signaling pathway.In recent years,research has identified protein post-translational modifications(PTMs),including phosphorylation,acetylation,ubiquitination,methylation,and SUMOylation,as having significant roles in regulating innate immune signaling pathways.This paper reviews how PTMs influence the function of cGAS and its downstream signaling pathways through various mechanisms.

BACKGROUND:Toll-like receptors are an important class of pattern recognition receptors that have important functions in pathogen immunity and cytokine synthesis by recognizing specific molecular patterns.Previous studies have found that different types of bone tissue cells also express Toll-like receptors.Activation or inhibition of Toll-like receptors can have significant effects on osteoblast and osteoclast function through multiple pathways. OBJECTIVE:To summarize the expression and action pathways of Toll-like receptors in osteoblasts and osteoclasts,in order to further elucidate the biological mechanisms involved in the regulation of Toll-like receptors under physiological and pathological conditions. METHODS:Relevant literature was retrieved from databases such as PubMed and CNKI as of December 2022.The Chinese and English search terms included"Toll-like receptor,osteoblast,osteoclast,mesenchymal stem cells,macrophage,cytokine,signaling pathway".According to the research needs,the corresponding criteria were established to screen the final literature. RESULTS AND CONCLUSION:(1)Toll-like receptors could directly regulate osteoblast and osteoclast differentiation through the activation of related signaling pathways.(2)Toll-like receptor activation induces cytokine production and exerts regulatory effects.(3)Toll-like receptor activation can affect the survival and migration ability of osteoblasts and osteoclasts.(4)Toll-like receptors in osteoblasts and osteoclasts are activated in certain diseases and pathological settings and are involved in intercellular interactions.

BACKGROUND:Tumor necrosis factor-α is a broadly acting inflammatory cytokine that plays an important role in the immune inflammatory response of the body.The current study concluded that tumor necrosis factor-α has significant biological effects on a variety of bone tissue cells. OBJECTIVE:To summarize the expression and action pathways of tumor necrosis factor-α in osteoblastic and osteoclastic cells to further elucidate the regulatory role of tumor necrosis factor-α on bone tissue cells. METHODS:PubMed and CNKI were searched until March 2023,and the Chinese search terms included"tumor necrosis factor α,osteoblast,osteoclast,osteoclast,osteoprogenitor";the English search terms included"TNF-α,osteoblast,osteoclast,osteocyte,osteoprogenitor cell".The corresponding criteria were established according to the research needs,and the final literature was screened.Finally,77 articles were included for review. RESULTS AND CONCLUSION:(1)Tumor necrosis factor-α is participating in regulating the recruitment,appreciation,and differentiation of osteoprogenitor cells,but leads to osteoprogenitor cell stripping and death under specific environments.It also participates in bone resorption directly or indirectly through secreted enzymes.(2)Tumor necrosis factor-α can increase the level of inflammatory factors in the environment by activating relevant signaling pathways in osteoclast lineage cells or directly induce the generation of osteoclasts in specific environments.(3)Tumor necrosis factor-α can inhibit osteogenic differentiation by activating nuclear factor-κB signaling pathway,inhibiting the expression of transcription factors such as RUNX2 and Osterix,and inducing apoptosis and necrotizing apoptosis in osteoblasts.(4)Tumor necrosis factor-α inhibits osteogenesis and promotes osteoclastogenesis by activating the nuclear factor-κB signaling pathway in osteocytes and inducing cytokines such as RANKL,SOST,and DKK1,while enhancing apoptosis of the osteocytes,as well as bone resorption around the apoptotic bone tissue.(5)Taken together,the effect of tumor necrosis factor-α in bone tissue is mainly to inhibit osteogenesis and promote osteoclastosis.The biological effect of tumor necrosis factor-α in bone tissue cells is usually dependent on the activation of tumor necrosis factor receptor and nuclear factor-κB signaling pathways.(6)The interaction of tumor necrosis factor-α with other tissue cell types surrounding bone tissue and its role in bone immune regulation still deserve attention in future studies.