The theory of constitution in traditional Chinese medicine （TCM） has emerged as a new discipline in recent years. Constitution plays a vital role in the onset，progression，transformation，and prognosis of diseases. At present，some clinical scholars have adopted a novel diagnostic and treatment model of "constitution differentiation-disease identification-syndrome differentiation"，in which constitution is regarded as a core element throughout the diagnostic and therapeutic process. Constitution is closely associated with etiology，onset，pathogenesis，syndrome differentiation，and treatment. Against this background，the construction of animal models based on constitution holds far-reaching significance for advancing clinical research. This paper focuses on the construction and evaluation of rodent models with Qi-deficiency constitution，aiming to explore how to further induce Qi-deficiency syndromes and related disease states on the basis of Qi-deficiency constitution models，thereby developing an integrated animal model that embodies the trinity of "constitution-disease-syndrome". The establishment of this model not only provides a solid experimental foundation for the development of new therapies and drugs in TCM targeting specific constitutions，diseases，and syndromes，but also greatly promotes the modernization and scientific advancement of TCM theory. By comprehensively applying multidisciplinary technologies and methods，the study evaluates the model's validity，reliability，and practicality，with the aim of opening new avenues for future research in TCM and promoting the development of the field.

The theory of constitution in traditional Chinese medicine （TCM） has emerged as a new discipline in recent years. Constitution plays a vital role in the onset，progression，transformation，and prognosis of diseases. At present，some clinical scholars have adopted a novel diagnostic and treatment model of "constitution differentiation-disease identification-syndrome differentiation"，in which constitution is regarded as a core element throughout the diagnostic and therapeutic process. Constitution is closely associated with etiology，onset，pathogenesis，syndrome differentiation，and treatment. Against this background，the construction of animal models based on constitution holds far-reaching significance for advancing clinical research. This paper focuses on the construction and evaluation of rodent models with Qi-deficiency constitution，aiming to explore how to further induce Qi-deficiency syndromes and related disease states on the basis of Qi-deficiency constitution models，thereby developing an integrated animal model that embodies the trinity of "constitution-disease-syndrome". The establishment of this model not only provides a solid experimental foundation for the development of new therapies and drugs in TCM targeting specific constitutions，diseases，and syndromes，but also greatly promotes the modernization and scientific advancement of TCM theory. By comprehensively applying multidisciplinary technologies and methods，the study evaluates the model's validity，reliability，and practicality，with the aim of opening new avenues for future research in TCM and promoting the development of the field.

Objective: To analyze the prevalence status and clustering patterns of risk factors for chronic diseases among adult residents in Rui' an City, Zhejiang Province, so as to provide a basis for formulating strategies for the prevention and control of chronic diseases and implementing risk factor interventions. Methods: A multi-stage random cluster sampling method was used to select residents aged ≥18 years in 5 townships (sub-districts) of Rui' an City as survey subjects from December 2023 to March 2024. Data on basic information, history of major chronic diseases, lifestyle behaviors, height, weight, and blood biochemical indicators were collected through questionnaire surveys, physical examinations, and laboratory tests. Descriptive analysis was used to analyze the prevalence status of 5 risk factors for chronic diseases. K-means clustering analysis was used to analyze clustering patterns. Results: A total of 3 060 people were surveyed, including 1 476 males, accounting for 48.24%, and 1 584 females, accounting for 51.76%. The median age was 49.00 (interquartile range, 25.00) years. There were 1 275 cases (41.67%) of hypertension, 462 cases (15.10%) of diabetes, and 1 460 cases (47.71%) of dyslipidemia. Insufficient fruit and vegetable intake, excessive salt intake, overweight and obesity, excessive red meat intake, and smoking involved 2 201, 1 878, 1 541, 1 337, and 571 people, with prevalences of 71.93%, 61.37%, 50.36%, 43.69%, and 18.66%, respectively. K-means clustering analysis identified 4 clustering patterns: smoking-insufficient fruit and vegetable intake type (249 people, accounting for 8.20%), low intake type (1 421 people, accounting for 46.75%). high red meat type (245 people, accounting for 8.07%), and high BMI-high salt type (1 118 people, accounting for 36.96%). The prevalences of hypertension and diabetes were higher in adult residents of the high BMI-high salt type, at 56.53% and 20.30%, respectively. The prevalence of dyslipidemia was higher in adult residents of the smoking-insufficient fruit and vegetable intake type, at 59.44%. Conclusions The prevalence of insufficient fruit and vegetable intake among adult residents in Rui' an City is relatively high. The clustering pattern of chronic disease risk factors is dominated by the low intake type. The prevalence of chronic diseases is higher in adult residents of the high BMI-high salt type and smoking-insufficient fruit and vegetable intake type. It is suggested to carry out health education and collaborative intervention of risk factors for different risk groups.

Inteins are unique protein insertion sequences capable of self-excision, enabling the covalent ligation of flanking extein peptides via amide bond formation. This process proceeds spontaneously without requiring external enzymes, cofactors, or chemical reagents, granting inteins exceptional biocompatibility and traceless performance in protein engineering applications. Split inteins represent a specialized and versatile subclass whose splicing domains are encoded by two separate gene fragments rather than a single continuous open reading frame. These fragments, known as the N-terminal (IntN) and C-terminal (IntC) split inteins, associate through non-covalent interactions including hydrophobic forces, hydrogen bonds, and van der Waals forces to assemble into an active three-dimensional structure, which then drives efficient extein ligation and enables protein trans-splicing. Protein trans-splicing mediated by split inteins has become a cornerstone for traceless protein ligation owing to its high specificity and irreversibility, fundamentally reshaping strategies for protein modification, assembly, and functional regulation. Compared with traditional chemical ligation methods, split intein systems require no complex chemical derivatization of peptide fragments and can operate efficiently at micromolar concentrations under physiological conditions, thus avoiding structural and functional damage caused by organic reagents. In contrast to enzymatic ligation tools such as sortase, split inteins eliminate the need for additional enzymes or cofactors, simplifying reaction systems, reducing costs, and minimizing non-specific side products. These distinctive advantages render split inteins highly promising for applications in chemical biology, synthetic biology, and biopharmaceutical development. In recent years, deepened mechanistic understanding has established structure-guided rational design as the primary approach to overcoming key limitations of split inteins, including intrinsic aggregation propensity, strict extein sequence dependence, and limited splicing efficiency. Bioinformatic tools have been used to identify aggregation-prone regions in the IntN fragment, and site-directed mutagenesis of hydrophobic residues, relocation of split sites, or removal of misfolding-prone sequences has substantially reduced in vitro aggregation and improved soluble expression and assembly activity. Rational engineering of catalytic residues and adjacent flexible loops has relaxed strict amino acid preferences at extein junctions, enhancing sequence tolerance and reducing the risk of functional impairment in target proteins. Consensus design based on multiple sequence alignments has yielded ultra-fast splicing variants such as Cfa DnaE and Cat-TerL, which exhibit significantly accelerated kinetics and improved tolerance to denaturing conditions. Meanwhile, advances in structural biology have further clarified the conformational dynamics and catalytic mechanisms of splicing, supporting the precise design of high-performance intein modules. On this basis, electrostatic interaction tuning and metagenomic screening have yielded multiple mutually orthogonal split intein pairs, enabling selective multi-fragment protein ligation and providing new routes for the efficient synthesis of large multi-domain functional proteins. With these engineered split inteins offering continuously improved performance and expanded applicability, protein trans-splicing has been widely applied in numerous cutting-edge areas of protein research and biomedicine. In gene delivery, split intein-based systems overcome the packaging limit of adeno-associated viral vectors, enabling the accurate reconstitution of large therapeutic proteins and base editors in target cells, thereby enhancing the efficacy and scope of gene therapy for genetic diseases. In internal protein sequence editing, split inteins mediate precise sequence replacement and modification in flexible regions or loops of target proteins, without the need for complex multi-step ligation and protein refolding involved in traditional protein semisynthesis. In protein-protein interaction studies, intein-mediated splicing covalently captures transient and weak intracellular complexes, enabling sensitive, high-throughput interaction detection and drug screening. In synthetic biology, conditionally controllable splicing systems support the construction of diverse intracellular and cell-surface biological logic gates for the precise regulation of cellular behavior. In mechanistic biochemical research, split inteins enable photocatalytic proximity labeling and site-specific tagging, allowing the preparation of homogeneous protein samples carrying precise post-translational modifications such as ubiquitination and polyglutamylation for chromatin interactome analysis and epigenetic studies. Moreover, covalent trapping strategies using split inteins stabilize transient enzymatic intermediates, providing unprecedented insights into molecular mechanisms such as nucleosome ubiquitination that are difficult to elucidate using conventional methods. This review systematically summarizes key technological advances in split inteins over the past decade, highlighting engineering strategies, mechanistic insights, and the development of orthogonal components. It comprehensively surveys emerging applications at the frontiers of protein research, analyzes current core challenges, and proposes future directions, particularly emphasizing artificial intelligence-driven de novo design and novel splicing pathways to break existing technical bottlenecks. By enabling traceless, efficient, and versatile protein manipulation, split inteins continue to serve as indispensable tools that drive innovation in protein engineering and fundamental life science research.

ObjectiveTo observe the clinical efficacy and safety of Yishen Jiangtang decoction combined with conventional Western medicine treatment in managing stage G3A3 diabetic kidney disease （DKD） in the patients with the pattern of Qi-Yin deficiency combined with dampness-heat and blood stasis， and to explore its mechanism of inhibiting the endoplasmic reticulum stress （ERS）-apoptosis axis and delaying renal fibrosis （RF）. MethodsA single-center， randomized， parallel-controlled clinical trial was conducted. A total of 78 patients diagnosed with stage G3A3 DKD and traditional Chinese medicine （TCM） pattern of Qi-Yin deficiency combined with dampness-heat and blood stasis and treated in the Department of Nephrology and the Department of Endocrinology of the People's Hospital Affiliated to Fujian University of Traditional Chinese Medicine from March 2025 to February 2026 were enrolled. They were randomly assigned at a 1∶1 ratio into a control group （conventional Western medicine treatment） and an experimental group （Yishen Jiangtang decoction combined with conventional Western medicine treatment）， with 39 patients in each group. The treatment lasted for 12 weeks. Basic demographic data and medical history were collected before and after treatment. The two groups before and after treatment were compared in terms of the following parameters： Blood urea nitrogen （BUN）， serum creatinine （SCr）， estimated glomerular filtration rate （eGFR）， urinary albumin-to-creatinine ratio （UACR）， fasting blood glucose （FBG）， triglycerides （TG）， total cholesterol （TC）， and low-density lipoprotein cholesterol （LDL-C）. Enzyme-linked immunosorbent assay （ELISA） kits were used to measure serum glucose-regulated protein 78 （GRP78）， serum cysteinyl aspartate-specific protease-3 （Caspase-3）， and serum and urinary transforming growth factor-β₁ （TGF-β₁） levels before and after treatment. TCM symptom scores， TCM syndrome efficacy， and overall clinical efficacy were evaluated before and after treatment. Spearman correlation analysis was performed to explore the correlations of serum levels of GRP78 and Caspase-3， as well as serum and urinary levels of TGF-β1， with renal function， proteinuria， and glucose and lipid metabolism indicators in patients with DKD. In addition， safety indicators were also monitored. ResultsA total of 75 patients completed the study （38 in the control group and 37 in the experimental group）. ① Baseline data： The two groups were comparable in terms of sex and age， with no statistically significant difference. ② Efficacy outcomes： Compared with baseline levels， both groups showed reductions in BUN， SCr， UACR， FBG， TG， TC， LDL-C， and TCM symptom scores （P<0.01） and increases in eGFR level （P<0.01）. Moreover， the experimental group outperformed the control group in altering the indicators above （P<0.05）. Regarding TCM syndrome efficacy， the total response rate in the experimental group was 83.78%， which was higher than that （57.89%） in the control group （P<0.05）. Regarding overall clinical efficacy， the total response rate in the experimental group was 78.38%， which was higher than that （50.00%） in the control group （P<0.05）. ③ Mechanism outcomes： Compared with baseline levels， both groups showed declined serum levels of GRP78， Caspase-3， and TGF-β₁ and urinary level of TGF-β₁ after treatment （P<0.01）. Moreover， the experimental group showed greater declines than the control group （P<0.05）. The serum levels of GRP78， Caspase-3，and TGF-β₁ and the urinary level of TGF-β₁ in DKD patients showed positive correlations with BUN， SCr， UACR， and FBG and negative correlations with eGFR （P<0.05）. ④ Safety outcomes： Neither group showed drug-related adverse reactions， and laboratory safety parameters remained within normal ranges. ConclusionYishen Jiangtang decoction combined with conventional Western medicine treatment may safely and effectively alleviate clinical symptoms， correct glucose and lipid metabolism disorders， and protect the renal function in patients with stage G3A3 DKD and the pattern of Qi-Yin deficiency combined with dampness-heat and blood stasis. The mechanism may be associated with inhibition of the ERS-apoptosis axis and delay of the RF process.

ObjectiveTo observe the clinical efficacy and safety of Yishen Jiangtang decoction combined with conventional Western medicine treatment in managing stage G3A3 diabetic kidney disease （DKD） in the patients with the pattern of Qi-Yin deficiency combined with dampness-heat and blood stasis， and to explore its mechanism of inhibiting the endoplasmic reticulum stress （ERS）-apoptosis axis and delaying renal fibrosis （RF）. MethodsA single-center， randomized， parallel-controlled clinical trial was conducted. A total of 78 patients diagnosed with stage G3A3 DKD and traditional Chinese medicine （TCM） pattern of Qi-Yin deficiency combined with dampness-heat and blood stasis and treated in the Department of Nephrology and the Department of Endocrinology of the People's Hospital Affiliated to Fujian University of Traditional Chinese Medicine from March 2025 to February 2026 were enrolled. They were randomly assigned at a 1∶1 ratio into a control group （conventional Western medicine treatment） and an experimental group （Yishen Jiangtang decoction combined with conventional Western medicine treatment）， with 39 patients in each group. The treatment lasted for 12 weeks. Basic demographic data and medical history were collected before and after treatment. The two groups before and after treatment were compared in terms of the following parameters： Blood urea nitrogen （BUN）， serum creatinine （SCr）， estimated glomerular filtration rate （eGFR）， urinary albumin-to-creatinine ratio （UACR）， fasting blood glucose （FBG）， triglycerides （TG）， total cholesterol （TC）， and low-density lipoprotein cholesterol （LDL-C）. Enzyme-linked immunosorbent assay （ELISA） kits were used to measure serum glucose-regulated protein 78 （GRP78）， serum cysteinyl aspartate-specific protease-3 （Caspase-3）， and serum and urinary transforming growth factor-β₁ （TGF-β₁） levels before and after treatment. TCM symptom scores， TCM syndrome efficacy， and overall clinical efficacy were evaluated before and after treatment. Spearman correlation analysis was performed to explore the correlations of serum levels of GRP78 and Caspase-3， as well as serum and urinary levels of TGF-β1， with renal function， proteinuria， and glucose and lipid metabolism indicators in patients with DKD. In addition， safety indicators were also monitored. ResultsA total of 75 patients completed the study （38 in the control group and 37 in the experimental group）. ① Baseline data： The two groups were comparable in terms of sex and age， with no statistically significant difference. ② Efficacy outcomes： Compared with baseline levels， both groups showed reductions in BUN， SCr， UACR， FBG， TG， TC， LDL-C， and TCM symptom scores （P<0.01） and increases in eGFR level （P<0.01）. Moreover， the experimental group outperformed the control group in altering the indicators above （P<0.05）. Regarding TCM syndrome efficacy， the total response rate in the experimental group was 83.78%， which was higher than that （57.89%） in the control group （P<0.05）. Regarding overall clinical efficacy， the total response rate in the experimental group was 78.38%， which was higher than that （50.00%） in the control group （P<0.05）. ③ Mechanism outcomes： Compared with baseline levels， both groups showed declined serum levels of GRP78， Caspase-3， and TGF-β₁ and urinary level of TGF-β₁ after treatment （P<0.01）. Moreover， the experimental group showed greater declines than the control group （P<0.05）. The serum levels of GRP78， Caspase-3，and TGF-β₁ and the urinary level of TGF-β₁ in DKD patients showed positive correlations with BUN， SCr， UACR， and FBG and negative correlations with eGFR （P<0.05）. ④ Safety outcomes： Neither group showed drug-related adverse reactions， and laboratory safety parameters remained within normal ranges. ConclusionYishen Jiangtang decoction combined with conventional Western medicine treatment may safely and effectively alleviate clinical symptoms， correct glucose and lipid metabolism disorders， and protect the renal function in patients with stage G3A3 DKD and the pattern of Qi-Yin deficiency combined with dampness-heat and blood stasis. The mechanism may be associated with inhibition of the ERS-apoptosis axis and delay of the RF process.

Objective To quantitatively measure the density of hydroxyapatite(HAP)(water)within lumbar vertebral bodies using the gemstone spectral imaging(GSI)material decomposition technique and to compare and analyze the clinical significance of density variations of HAP(water)in different regions of L1-L3 vertebral bodies.Methods A total of 242 patients who underwent lumbar quantitative computed tomography(QCT)scans via utilizing the GSI technique were selected.Following the scans,the HAP(water)density values in four regions(anterosuperior,posterosuperior,anteroinferior,and posteroinferior)of each L1-L3 vertebral bodies were quantitatively measured using the material decomposition technique.Based on the QCT results,all cases were divided into three groups of normal bone mineral density,osteopenia,and osteoporosis.The distributions of HAP(water)density values in the four regions within each vertebral body were compared and analyzed among the groups.Results In all three groups of patients,the highest HAP(water)density values in the L1-L3 vertebral bodies were all located in the posteroinferior region,followed by the posterosuperior region.In the normal bone mineral density group,the lowest HAP(water)density values was found in the anterosuperior region of the L1 vertebral body.In the osteopenia and osteoporosis groups,the lowest HAP(water)density values was found in the anteroinferior region of the L1-L3 vertebral bodies.Conclusion Significant differences in HAP(water)density are present across different regions within lumbar vertebral bodies,which may be related to the development of vertebral osteoporosis and the location of fractures.

Objective This study aims to investigate controversial cases of forensic clinical re-identification of fractures,exploring the characteristics,causes,and countermeasures related to identification errors in primary bone injuries,complications,and subsequent changes.The goal is to provide identification strategies for similar cases regarding the collection of identification materials,timing,and examination method selection,ultimately establishing a paradigm for such identifications.Methods A total of 103 cases of clinical re-identification of fractures accepted by the West China Forensic Identification Center from 2020 to 2024 were collected,and the data from initial identifications and re-identifications were retrospectively analyzed.Results Male cases accounted for 69.90%of the re-identifications,with disability grade(67.96%)and injury degree(30.10%)being the primary concerns.Individual requests represented a high proportion(92.86%)in the initial assessment of disability levels,while unit or joint requests dominated the re-assessment(92.86%).The agreement rates for disability grade and injury degree were 55.26%and 59.38%,respectively.The reassessment of disability grades primarily involved fractures of limb long bones,spine,and ribs,with 75.53%of opinions resulting in downgraded disability levels.Rib,orbital,and nasal bone fractures were the main focus in injury degree reassessments,with 84.62%of opinions indicating aggravated injuries.The consistency rates for fracture identification in disability grade assessments was 92.21%,while rates for injury degree and sequelae were 65.63%and 48.94%,respectively.Inconsistencies in identifying damage facts—including the presence of fractures,distinguishing between fresh and old fractures,and determining the nature of fractures and sequelae—were primarily noted in rib,orbital,and nasal bone fractures.The utilization rate of CT metadata in initial evaluations(25.00%)was significantly lower than in re-evaluations(95.00%).The identification time for joint mobility dysfunction after fracture in re-identifications was significantly longer than in initial identifications(P=0.0002),and the identification time for cases with agreement was significantly shorter than for cases with disagreement(P=0.036).Conclusion Image data type and identification timing are critical factors that may influence the accuracy and consistency of forensic clinical identification of bone injuries.When necessary,dynamic CT metadata in conjunction with image post-processing technology can be routinely employed to identify fractures of the ribs,orbital bones,or nasal bones,thereby reducing the risk of misidentification.

BACKGROUND:Enhancing the differentiation of induced pluripotent stem cells into lung stem cells is crucial for repairing lung injuries.NKX2.1 is the earliest marker of lung epithelial differentiation and plays a significant regulatory role in lung development.However,the impact of its expression on the differentiation of induced pluripotent stem cells into lung stem cells remains inadequately understood.OBJECTIVE:To investigate the effect of NKX2.1 on the differentiation of induced pluripotent stem cells into lung stem cells.METHODS:Induced pluripotent stem cells were cultured in vitro.The expression of specific pluripotent stem cell genes was assessed using real-time fluorescence quantitative PCR.NKX2.1 was overexpressed in induced pluripotent stem cells,which were then induced to differentiate into lung stem cells.The expression of FoxA2,SOX9,and P63 was determined via quantitative PCR and immunofluorescence on day 7 of induction of differentiation.The expression of the alveolar marker SPB and SPC was evaluated through immunofluorescence staining on day 7 of induction of differentiation.RESULTS AND CONCLUSION:(1)Induced pluripotent stem cells in vitro were tightly packed and showed typical clonoid growth and significantly expressed stem cell-specific genes OCT-4,SOX2,and NANOG.(2)Compared with the non-transfected control group,the expression of NKX2.1 in human induced pluripotent stem cells was significantly increased in the NKX2.1 overexpression group(P＜0.000 1).(3)Seven days after induction of differentiation,compared with the non-transfected control group,the expression of lung stem cell-related markers FoxA2,SOX9,and P63 was significantly increased in the NKX2.1 overexpression group(P＜0.000 1).(4)Thirteen days after induction of differentiation,compared with the non-transfected control group,the fluorescence intensity of alveolar cell marker molecules SPB and SPC increased significantly in the overexpression NKX2.1 group.The results show that NKX2.1 can promote the differentiation of induced pluripotent stem cells into lung stem cells.

The Janus kinase/signal transducers and activators of transcription (JAK-STAT) control natural killer (NK) cells development and cytotoxic functions, however, whether long non-coding RNAs (lncRNAs) are involved in this pathway remains unknown. We found that miR155HG was elevated in activated NK cells and promoted their proliferation and effector functions in both NK92 and induced-pluripotent stem cells (iPSCs)-derived NK (iPSC-NK) cells, without reliance on its derived miR-155 and micropeptide P155. Mechanistically, miR155HG bound to miR-6756 and relieved its repression of JAK3 expression, thereby promoting the JAK-STAT pathway and enhancing NK cell proliferation and function. Further investigations disclosed that upon cytokine stimulation, STAT3 directly interacts with miR155HG promoter and induces miR155HG transcription. Collectively, we identify a miR155HG-mediated positive feedback loop of the JAK-STAT signaling. Our study will also provide a power target regarding miR155HG for improving NK cell generation and effector function in the field of NK cell adoptive transfer therapy against cancer, especially iPSC-derived NK cells.