Constructing an efficient and easy-to-operate multigene expression system is currently a crucial part of plant genetic engineering. In this study, a fragment carrying three independent gene expression cassettes and the expression unit of the gene-silencing suppressor protein(RNA silencing suppressor 19 kDa protein, P19) simultaneously was designed and constructed. This fragment was cloned into the commonly used plant expression vector pCAMBIA300, and the plasmid pC1300-TP2-P19 was obtained. Each gene expression cassette consists of different promoters, fusion tags, and terminators. The target gene can be flexibly inserted into the corresponding site through enzymatic digestion and ligation or recombination and fused with different protein tags, which provides great convenience for subsequent detection. The enhanced green fluorescent protein(eGFP) reporter gene was individually constructed into each expression cassette to verify the feasibility of this vector system. The results of tobacco transient expression and laser-confocal microscopy showed that each expression cassette presented independent and normal expression. Meanwhile, the three key enzyme genes in the betanin synthesis pathway, BvCYP76AD, BvDODA1, and DbDOPA5GT, were constructed into the three expression cassettes. The results of tobacco transient expression phenotype, protein immunoblotting(Western blot), and chemical detection of product demonstrated that the three exogenous genes were highly expressed, and the target compound betanin was successfully produced. The above results indicated that the constructed multigene expression system for plants in this study was efficient and reliable and can achieve the co-transformation of multiple plant genes. It can provide a reliable vector platform for the analysis of plant natural product synthesis pathways, functional verification, and plant metabolic engineering.
Nicotiana/metabolism* ; Genetic Vectors/metabolism* ; Gene Expression Regulation, Plant ; Plant Proteins/metabolism* ; Plants, Genetically Modified/metabolism* ; Genetic Engineering/methods* ; Green Fluorescent Proteins/metabolism* ; Gene Expression

OBJECTIVE: This study aimed to explore the association between body mass index (BMI) and mortality based on the 10-year population-based multicenter prospective study. METHODS: A general population-based multicenter prospective study was conducted at four sites in rural China between 2013 and 2023. Multivariate Cox proportional hazards models and restricted cubic spline analyses were used to assess the association between BMI and mortality. Stratified analyses were performed based on the individual characteristics of the participants. RESULTS: Overall, 19,107 participants with a sum of 163,095 person-years were included and 1,910 participants died. The underweight (< 18.5 kg/m ²) presented an increase in all-cause mortality (adjusted hazards ratio [ aHR] = 2.00, 95% confidence interval [ CI]: 1.66-2.41), while overweight (≥ 24.0 to < 28.0 kg/m ²) and obesity (≥ 28.0 kg/m ²) presented a decrease with an aHR of 0.61 (95% CI: 0.52-0.73) and 0.51 (95% CI: 0.37-0.70), respectively. Overweight ( aHR = 0.76, 95% CI: 0.67-0.86) and mild obesity ( aHR = 0.72, 95% CI: 0.59-0.87) had a positive impact on mortality in people older than 60 years. All-cause mortality decreased rapidly until reaching a BMI of 25.7 kg/m ² ( aHR = 0.95, 95% CI: 0.92-0.98) and increased slightly above that value, indicating a U-shaped association. The beneficial impact of being overweight on mortality was robust in most subgroups and sensitivity analyses. CONCLUSION This study provides additional evidence that overweight and mild obesity may be inversely related to the risk of death in individuals older than 60 years. Therefore, it is essential to consider age differences when formulating health and weight management strategies.
Humans ; Body Mass Index ; China/epidemiology* ; Male ; Female ; Middle Aged ; Prospective Studies ; Rural Population/statistics & numerical data* ; Aged ; Follow-Up Studies ; Adult ; Mortality ; Cause of Death ; Obesity/mortality* ; Overweight/mortality*

AIM To study the chemical constituents from Tetrastigma hemsleyanum Diels et Gilg and their antitumor activity in vitro.METHODS Silica gel,ODS,Sephadex LH-20 and semi-preparative HPLC were used for isolation and purification,then the structures of obtained compounds were identified by physicochemical properties and spectral data.The antitumor activity in vitro was determined by MTT mothod.RESULTS Twenty-eight compounds were isolated and identified as triphyllin A(1),eruberin B(2),(2S,4R)-5,7-dihydroxy-4,4'-dimethyl-6,8-dimethyl-flavan-5-O-β-D-6-acetylglucopyranoside-7-O-β-D-glucopyranoside(3),eruberin A(4),abacopterin Ⅰ(5),matteucinol(6),homoerodictyol(7),(2S)-5,3',4'-trihydroxy-7-methoxy-flavanone(8),(2S)-5,2',5'-trihydroxy-7-methoxyflavanone(9),galinsonside B(10),quercetin-3-O-β-D-glucopyranoside(11),kaempferol 3-O-robinobioside(12),rutin(13),geniposide(14),jasminoside A(15),β-sitostenone(16),sitosterol palmitate(17),β-sitosterol(18),ursolic acid(19),hyptadienic acid(20),3,4-dihydroxybenzoic acid(21),3,4-dimethoxybenzoic acid(22),gallic acid(23),dibutylphthalate(24),bis-(2-ethylhexyl)phthalate(25),9-nonadecenoic acid(26),triacylglycerol(27),crocin Ⅰ(28).The IC50 values of compound 1 for human gastric adenocarcinoma cells BGC-823 and human colon cancer cells HCT-116 were(22.07±0.38),(20.67±0.11)μmol/L,respectively.The IC50 value of compound 9 for BGC-823 cells was(21.58±0.05)μmol/L,and the IC50 value of compound 4 for HCT-116 cells was(16.67±0.36)μmol/L.CONCLUSION Compounds 1-10,14-15 and 28 are first isolated from Tetrastigma genus.Compounds 1,4,9 have weak antitumor activity in vitro.

OBJECTIVE: To review the current research status on the diagnosis and treatment of fracture-related infection (FRI). METHODS: The research literature in the field of FRI both domestically and internationally in recent years were widely reviewed, and the research progress of FRI from the aspects of definition and classification, epidemiological characteristics, diagnosis and treatment elaborated, in order to provide reference for clinical practices. RESULTS: In recent years, specific classifications for FRI have gradually emerged. FRI is characterized by high incidence, high recurrence, high disability rates, and significant economic costs. Key diagnostic points include clinical signs and symptoms, imaging tests, serological biomarkers, pathogen identification, and histopathological examination. Treatment principles encompass debridement, management of implants (retention or removal), systemic and local antibiotic use, reconstruction of bone and soft tissue defects, and functional and psychological rehabilitation. CONCLUSION Although FRI is a catastrophic complication following limb bone trauma, early precise diagnosis and standardized treatment are key to improving cure rates, reducing recurrence, and enhancing patients' quality of life.
Humans ; Fractures, Bone/complications* ; Bone Diseases, Infectious/therapy* ; Debridement/methods* ; Anti-Bacterial Agents/therapeutic use* ; Orthopedic Procedures/methods*

Diabetic foot (DF) is one of the most serious chronic complications of diabetes. The incidence rate among global diabetes patients is as high as 15% to 25%, and about 50% of patients will develop contralateral foot ulcers within 5 years after the first unilateral ulcer. As a non-invasive prevention and control solution, the application progress of functional insoles is mainly reflected in the following aspects:(1) Material innovation. The application of new composite materials and smart materials has significantly enhanced the pressure reduction effect and comfort. (2) Structural optimization. The development of multi-layer design and local pressure reduction structure has achieved more precise pressure distribution regulation. (3) Manufacturing process. 3D printing and parametric design have enabled the personalized customization of functional insoles. (4) Intelligent monitoring. It integrates functions such as pressure sensing and temperature monitoring, achieving real-time monitoring and early warning of foot conditions. Clinical research has confirmed that personalized functional insoles could reduce the incidence of foot ulcers and shorten the healing time of ulcers. At present, the research hotspots mainly focus on the development of smart materials, the construction of multi-functional integration and remote monitoring systems. However, in-depth research is still needed in the aspects of biomechanical mechanisms, standardized evaluation systems and long-term efficacy assessment. The development of future functional insoles should focus on the coordinated advancement of "personalization-intelligence-standardization", with the aim of providing more effective solutions for the prevention and treatment of DF.
Humans ; Diabetic Foot/therapy* ; Foot Orthoses

Psoriasis is a chronic inflammatory skin disease, and its pathogenesis is largely modulated by abnormal angiogenesis. Previous research has indicated that AlkB homolog 5 (ALKBH5), an important demethylase affecting N⁶-methyladenosine (m⁶A) modification, plays a role in regulating angiogenesis in cardiovascular and eye diseases. Our present study found that ALKBH5 was upregulated and co-localized with cluster of differentiation 31 (CD31) in the skin of IMQ group compared with control group. ALKBH5-deficient mice decreased IMQ-induced psoriatic dermatitis and exhibited histological improvements, including decreased epidermal thickness, hyperkeratosis, numbers of dermal capillary vessels and inflammatory cell infiltration. ALKBH5-KO mice alleviated angiogenesis in psoriatic lesions by downregulating the protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway. Additionally, the expression of ALKBH5 was significantly upregulated in IL-17A-induced human umbilical vein endothelial cells (HUVECs), which further promoted the expression of angiogenesis-related cytokines and endothelial cell proliferation. Cell proliferation and angiogenesis were suppressed in ALKBH5 knockdown group, whereas ALKBH5 overexpression promoted these processes. The regulation of angiogenesis in HUVECs by ALKBH5 was facilitated through the AKT-mTOR pathway. Collectively, ALKBH5 plays a pivotal role in psoriatic dermatitis and angiogenesis, which may offer a new potential targets for treating psoriasis.
Animals ; Psoriasis/chemically induced* ; Mice ; Humans ; Neovascularization, Pathologic/genetics* ; Human Umbilical Vein Endothelial Cells/metabolism* ; AlkB Homolog 5, RNA Demethylase/genetics* ; Proto-Oncogene Proteins c-akt/metabolism* ; TOR Serine-Threonine Kinases/metabolism* ; Cell Proliferation ; Mice, Knockout ; Disease Models, Animal ; Signal Transduction ; Male ; Skin/blood supply* ; Mice, Inbred C57BL ; Angiogenesis

BACKGROUND:Previous studies have indicated that cathepsin K can intervene with the occurrence and development of osteoporosis by regulating bone mineral density in middle-aged and older adults. However,whether there is a causal relationship between the cathepsin family and bone mineral density in other populations remains unknown. OBJECTIVE:To investigate the causal relationship between cathepsin and bone mineral density.METHODS:Genetic loci associated with eight cathepins were extracted from the IEU Open GWAS database as instrumental variables,and bone mineral density values in five age groups acted as an outcome. The causal relationship between cathepin and bone mineral density was assessed by two-way Mendelian randomization analysis. Heterogeneity of the genetic instrumental variables was assessed using Cochran's Q test,pleiotropy was assessed using the MR-Egger intercept test,and the sensitivity of single nucleotide polymorphisms used as instrumental variables to the causal effect of exposure and outcome was assessed using the leave-one-out method. RESULTS AND CONCLUSION:The results of the inverse variance weighting method with positive Mendelian randomization showed that cathepin H was negatively associated with bone mineral density in people aged 45-60 years[odds ratio (95％ confidence interval)=0.965(0.94-0.99),P=0.04];cathepin Z was negatively associated with bone mineral density in people aged 30-45 year[odds ratio (95％ confidence interval)=1.06 (1.00-1.11),P=0.03]. The results of sensitivity analysis showed a stable causal relationship,and MR-Egger intercept analysis did not detect potential horizontal pleiotropy. The inverse Mendelian randomization results showed that bone mineral density had no significant inverse effect on cathepin. The above results confirm that cathepin can affect bone mineral density in some age groups,which may increase the risk of osteoporosis and should be given more attention.

BACKGROUND:Remyelination in the central nervous system is a basic repair process triggered by demyelinating events,mainly through the proliferation,migration,and differentiation of oligodendrocyte precursor cells into oligodendrocytes.The process of remyelination is affected by many factors such as astrocytes,myelin debris,microglia,macrophages,endothelial cells,pericytes,T cells,and age.OBJECTIVE:Astrocytes play an important role in regulating synaptic activity,nutritional support,and tissue repair in the central nervous system.This review aims to provide potential therapeutic targets for demyelinating diseases of central nervous system by reviewing the role of astrocytes in remyelination.METHODS:A search was conducted on relevant literature collected from CNKI,PubMed,and Web of Science from 2014 tO 2024.The search terms were"astrocytes,oligodendrocyte precursor cells,remyelination"in both Chinese and English.Finally,66 articles were included after screening and summarized.RESULTS AND CONCLUSION:(1)The treatment of demyelinating diseases,such as multiple sclerosis,is limited to disease-modifying therapies,and there is no available method to overcome the failure of remyelination.Therefore,it is necessary to explore targets related to remyelination to promote myelin repair.(2)Remyelination is a process in which oligodendrocyte precursor cells proliferate,migrate,differentiate,and mature into oligodendrocytes,and the latter produce myelin to wrap axons to form myelin sheath.(3)Astrocytes regulate remyelination by phagocytosis of myelin debris,participating in inflammatory response,transforming into oligodendrocyte lineage cells,providing energy supply for oligodendrocyte lineage cells,releasing neurotrophic factors,and secreting extracellular matrix components.(4)The drugs screened in this paper use astrocytes and their derived factors as intervention targets to regulate the remyelination.Some drugs have satisfactory effects,but their effectiveness and safety still need more basic research and clinical trials to verify.(5)The mechanism of action of astrocytes in remyelination has not been fully elucidated,and the related molecular targets and signaling pathways can be further studied.

Objective:To explore the clinical significance of chromosome microarray analysis (CMA) in the prenatal detection of DMD gene variants in fetuses without family history during prenatal diagnosis.Methods:A retrospective analysis was conducted on amniotic fluid samples from 12 629 pregnant women who underwent CMA prenatal diagnosis due to high-risk factors at Women and Children′s Hospital Affiliated to Qingdao University between January 2019 and December 2024. Samples with CMA-indicated DMD gene variants were further verified by multiplex ligation-dependent probe amplification (MLPA).Results:(1) Among 12 629 amniotic fluid samples, CMA detected 11 samples with DMD gene deletions or duplications (6 male and 5 female fetuses), which were confirmed by MLPA. (2) All 11 samples with DMD gene variants had no family history of genetic diseases, including 5 deletions and 6 duplications. All of the 5 DMD gene deletions occurred in male fetuses and were all pathogenic, and the pregnant women chose to terminate the pregnancies. Among the 6 DMD gene duplications cases, 1 male fetus was diagnosed as pathogenic and had pregnancy termination; the other 5 duplication cases were female fetuses, in which 1 were pathogenic and 4 were likely pathogenic. They continued pregnancy until delivery, and follow-up showed no DMD-related symptoms. (3) Pedigree analysis revealed that among the 11 samples with DMD gene variants, 3 were de novo mutations, 7 were inherited from mothers, and 1 had an unknown origin.Conclusions:For fetuses without pseudohypertrophic muscalar dystrophy family history but requiring invasive prenatal diagnosis for other reasons, CMA helps to increase the detection of DMD gene variants in fetuses. Testing pregnant women for DMD pathogenic gene carriers could effectively prevent the birth of pseudohypertrophic muscalar dystrophy children.

Fracture-related infection (FRI) is defined as a bone infection occurring at fracture sites or implant interfaces following open fractures or internal fixation, primarily caused by pathogenic bacterial colonization or compromised host immunity, with variable involvement of adjacent soft tissues. As a catastrophic complication of extremity trauma, FRI continues to pose substantial clinical challenges. Diagnostic difficulties stem from multiple factors: imaging confounded by traumatic and postoperative changes, limited specificity of conventional inflammatory biomarkers, high false-negative rates in standard microbial cultures, and lack of standardized histopathological criteria. Therapeutically, challenges persist, including uncertainty regarding optimal debridement extent, lack of evidence-based criteria for implant retention during early infection, ongoing controversy over contentious administration routes and duration of systemic antibiotics, and elevated complication rates in major bone defect reconstruction. To this end, the authors explored challenges and corresponding management strategies in the clinical diagnosis and treatment of FRI, aiming to enhance orthopedic surgeons′ clinical recognition of it.