ObjectiveProtein-protein interactions (PPIs) are fundamental to the execution of biological functions within living cells. However, traditional biochemical methods, such as co-immunoprecipitation (Co-IP), often fail to capture transient, weak, or membrane-associated interactions due to the stringent detergent requirements for cell lysis. Proximity labeling (PL) has emerged in recent years as a transformative technology for mapping the proteomes of specific subcellular compartments and identifying dynamic interactomes in situ. Golgi protein 73 (GP73, also known as GOLPH2), a resident type II Golgi transmembrane protein, is a well-recognized clinical biomarker for liver diseases, including hepatocellular carcinoma (HCC). Despite its clinical significance, the comprehensive physiological and pathological functions of GP73 remain partially understood. This study aims to establish an APEX2-mediated proximity labeling system specifically targeting GP73 to map its interactome in a living cellular environment, thereby providing new insights into its molecular roles and regulatory mechanisms. MethodsTo achieve spatial specificity, we first constructed a stable cell line expressing a fusion protein consisting of GP73 and the engineered soybean peroxidase APEX2. The localization of the GP73-APEX2 fusion protein was validated to ensure it correctly targeted the Golgi apparatus. The proximity labeling reaction was initiated by incubating the cells with biotin-phenol (BP) for 30 min, followed by a brief (1 min) treatment with1 mmol/L hydrogen peroxide (H₂O₂). This catalytic reaction converts BP into highly reactive, short-lived biotin-phenoxyl radicals that covalently attach to endogenous proteins within a small labeling radius of the GP73-APEX2 enzyme. Subsequently, the cells were quenched, and biotinylated proteins were enriched using high-affinity streptavidin-coated magnetic beads. The captured “neighbor” proteins were subjected to on-bead digestion and analyzed via liquid chromatography-tandem mass spectrometry (LC-MS/MS) for high-throughput identification. Rigorous bioinformatics analysis, including Gene Ontology (GO) enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and protein-protein interaction network mapping, was performed to interpret the biological significance of the identified candidates. ResultsOur results demonstrate the successful establishment of a robust and sensitive APEX2-based proximity labeling system for GP73. We identified a total of 95 high-confidence interacting proteins that were significantly enriched in the GP73 proximity proteome compared to control groups. Bioinformatics analysis revealed that these interactors were predominantly associated with biological processes such as vesicular transport, protein localization, and, most notably, molecular functions related to “ribosome binding” and “translation regulation”. This suggested an unexpected role for the Golgi-resident GP73 in the cellular translation machinery. To validate these findings, we performed targeted biochemical assays which confirmed a direct interaction between GP73 and the subunits of the eukaryotic translation initiation factor 3 (eIF3) complex, specifically EIF3G and EIF3I. Furthermore, functional validation using the surface sensing of translation (SUnSET) assay—a non-radioactive method to monitor protein synthesis—revealed that the overexpression of GP73 significantly promoted global protein translation levels in the cell, whereas its depletion or inhibition resulted in reduced translation efficiency. ConclusionThis study successfully utilized APEX2-mediated proximity labeling to provide the first systematic map of GP73 interactome in living cells. Our findings uncover a novel, unconventional function of GP73 as a regulator of cellular protein translation, likely mediated through its interaction with the eIF3 complex. This discovery significantly broadens our understanding of the biological roles of GP73 beyond its traditional function in the Golgi apparatus and suggests that it may act as a bridge between Golgi-related trafficking and the protein synthesis machinery. Furthermore, the technical framework established in this study provides a valuable template for investigating other complex organelle-associated protein networks and resolving transient macromolecular interactions in various physiological and pathological contexts.

ObjectiveTo evaluate the efficacy and safety of Janus kinase （JAK） inhibitors combined with Chinese herbal medicine （CHM） in treating rheumatoid arthritis （RA）. MethodsClinical data from 169 RA patients were retrospectively collected. Among them， 71 cases received JAK inhibitors as the control group， while 98 cases received JAK inhibitors plus CHM as the observation group， both treated for 24 weeks. The rheumatoid factor （RF）， cyclic citic peptide antibody （anti-CCP）， erythrocyte sedimentation rate （ESR）， C-reactive protein （CRP）， and white blood cell count （WBC） were recorded before and after treatment. Databases including CNKI， Wanfang， VIP， PubMed and Web of Science were searched from inception till August 31st， 2025 for randomized controlled trials （RCTs） on the combined use of JAK inhibitors and CHM for RA. The methodological quality of the included studies was evaluated using the risk of bias assessment tool. Meta-analyses were performed for RF， anti-CCP， ESR， CRP， 28-joint disease activity score （DAS28）， overall clinical effective rate， and incidence of adverse events. Sensitivity analysis were also performed. ResultsThe retrospective study demonstrated that after treatment， ESR， CRP， and anti-CCP levels decreased in the observation group， while ESR and CRP levels decreased in the control group （P＜0.05）. Moreover， ESR and RF levels in the observation group were lower than those in the control group （P＜0.05）. A total of 9 RCTs involving 770 patients were included in the meta-analysis. The results indicated that the JAK inhibitors plus CHM group was superior to the JAK inhibitors group in reducing RF （MD=-8.97， 95%CI -15.01 to -2.94， P=0.004）， CRP （MD=-3.34， 95%CI -3.82 to -2.86， P＜0.001）， ESR （MD=-5.33， 95%CI -7.98 to -2.69， P＜0.001）， and DAS28 score （MD=-0.54， 95%CI -0.74 to -0.34， P＜0.001）， as well as in improving the overall clinical effective rate （OR=4.53， 95%CI 2.55 to 8.03， P＜0.001）. No statistically significant differences were observed between groups in anti-CCP levels （SMD=-2.08， 95%CI -4.41 to 0.24， P=0.080） or incidence of adverse events （OR=0.93， 95%CI 0.55 to 1.57， P=0.790）. ConclusionThe combination of JAK inhibitors and CHM demonstrates remarkable efficacy in treating RA， contributing to improved disease activity and reduced inflammatory markers with a favorable safety profile.

BACKGROUND: While emotional distress, encompassing anxiety and depression, has been associated with negative clinical outcomes, its impact across various clinical departments and general hospitals has been less explored. Previous studies with limited sample sizes have examined the effectiveness of specific treatments (e.g., antidepressants) rather than a systemic management strategy for outcome improvement in non-psychiatric inpatients. To enhance the understanding of the importance of addressing mental health care needs among non-psychiatric patients in general hospitals, this study retrospectively investigated the impacts of emotional distress and the effects of early detection and management of depression and anxiety on hospital length of stay (LOS) and rate of long LOS (LLOS, i.e., LOS >30 days) in a large sample of non-psychiatric inpatients. METHODS: This retrospective cohort study included 487,871 inpatients from 20 non-psychiatric departments of a general hospital. They were divided, according to whether they underwent a novel strategy to manage emotional distress which deployed the Huaxi Emotional Distress Index (HEI) for brief screening with grading psychological services (BS-GPS), into BS-GPS ( n = 178,883) and non-BS-GPS ( n = 308,988) cohorts. The LOS and rate of LLOS between the BS-GPS and non-BS-GPS cohorts and between subcohorts with and without clinically significant anxiety and/or depression (CSAD, i.e., HEI score ≥11 on admission to the hospital) in the BS-GPS cohort were compared using univariable analyses, multilevel analyses, and/or propensity score-matched analyses, respectively. RESULTS: The detection rate of CSAD in the BS-GPS cohort varied from 2.64% (95% confidence interval [CI]: 2.49%-2.81%) to 20.50% (95% CI: 19.43%-21.62%) across the 20 departments, with a average rate of 5.36%. Significant differences were observed in both the LOS and LLOS rates between the subcohorts with CSAD (12.7 days, 535/9590) and without CSAD (9.5 days, 3800/169,293) and between the BS-GPS (9.6 days, 4335/178,883) and non-BS-GPS (10.8 days, 11,483/308,988) cohorts. These differences remained significant after controlling for confounders using propensity score-matched comparisons. A multilevel analysis indicated that BS-GPS was negatively associated with both LOS and LLOS after controlling for sociodemographics and the departments of patient discharge and remained negatively associated with LLOS after controlling additionally for the year of patient discharge. CONCLUSION Emotional distress significantly prolonged the LOS and increased the LLOS of non-psychiatric inpatients across most departments and general hospitals. These impacts were moderated by the implementation of BS-GPS. Thus, BS-GPS has the potential as an effective, resource-saving strategy for enhancing mental health care and optimizing medical resources in general hospitals.
Humans ; Retrospective Studies ; Male ; Length of Stay/statistics & numerical data* ; Female ; Middle Aged ; Adult ; Psychological Distress ; Inpatients/psychology* ; Aged ; Anxiety/diagnosis* ; Depression/diagnosis*

The present study aimed to investigate the effects of eccentric treadmill exercise on adaptive hypertrophy of skeletal muscle in rats. Thirty-two 3-month-old Sprague Dawley (SD) rats were selected and randomly assigned to one of the four groups based on their body weights: 2-week quiet control group (2C), 2-week downhill running exercise group (2E), 4-week quiet control group (4C), and 4-week downhill running exercise group (4E). The downhill running protocol for rats in the exercise groups involved slope of -16°, running speed of 16 m/min, training duration of 90 min, and 5 training sessions per week. Twenty-four hours after the final session of training, all the four groups of rats underwent an exhaustion treadmill exercise. After resting for 48 h, all the rats were euthanized and their gastrocnemius muscles were harvested for analysis. HE staining was used to measure the cross-sectional area (CSA) and diameter of muscle fibers. Transmission electron microscope was used to observe the ultrastructural changes in muscle fibers. Purithromycin surface labeling translation method was used to measure protein synthesis rate. Immunofluorescence double labeling was used to detect the colocalization levels of lysosomal-associated membrane protein 2 (Lamp2)-leucyl-tRNA synthetase (LARS) and Lamp2-mammalian target of rapamycin (mTOR). Western blot was used to measure the protein expression levels of myosin heavy chain (MHC) IIb and LARS, as well as the phosphorylation levels of mTOR, p70 ribosomal protein S6 kinase (p70S6K), and eukaryotic translation initiation factor 4E binding protein 1 (4E-BP1). The results showed that, compared with the 2C group rats, the 2E group rats showed significant increases in wet weight of gastrocnemius muscle, wet weight/body weight ratio, running distance, running time, pre- and post-exercise blood lactate levels, myofibrillar protein content, colocalization levels of Lamp2-LARS and Lamp2-mTOR, and LARS protein expression. Besides these above changes, compared with the 4C group, the 4E group further exhibited significantly increased fiber CSA, fiber diameter, protein synthesis rate, and phosphorylation levels of mTOR, p70S6K, and 4E-BP1. Compared with the quiet control groups, the exercise groups exhibited ultrastructural damage of rat gastrocnemius muscle, which was more pronounced in the 4E group. These findings suggest that eccentric treadmill exercise may promote mTOR translocation to lysosomal membrane, activating mTOR signaling via up-regulating LARS expression. This, in turn, increases protein synthesis rate through the mTOR-p70S6K-4E-BP1 signaling pathway, promoting protein deposition and inducing adaptive skeletal muscle hypertrophy. Although the ultrastructural changes of skeletal muscle are more pronounced, the relatively long training cycles during short-term exercise periods have a more significant effect on promoting gastrocnemius muscle protein synthesis and adaptive hypertrophy.
Animals ; Rats, Sprague-Dawley ; Physical Conditioning, Animal/physiology* ; Rats ; Muscle, Skeletal/metabolism* ; TOR Serine-Threonine Kinases/metabolism* ; Male ; Hypertrophy ; Adaptation, Physiological/physiology* ; Adaptor Proteins, Signal Transducing/metabolism* ; Ribosomal Protein S6 Kinases, 70-kDa/metabolism* ; Intracellular Signaling Peptides and Proteins

The circadian clock plays a critical role in regulating various physiological processes, including gene expression, metabolic regulation, immune response, and the sleep-wake cycle in living organisms. Post-translational modifications (PTMs) are crucial regulatory mechanisms to maintain the precise oscillation of the circadian clock. By modulating the stability, activity, cell localization and protein-protein interactions of core clock proteins, PTMs enable these proteins to respond dynamically to environmental and intracellular changes, thereby sustaining the periodic oscillations of the circadian clock. Different types of PTMs exert their effects through distincting molecular mechanisms, collectively ensuring the proper function of the circadian system. This review systematically summarized several major types of PTMs, including phosphorylation, acetylation, ubiquitination, SUMOylation and oxidative modification, and overviewed their roles in regulating the core clock proteins and the associated pathways, with the goals of providing a theoretical foundation for the deeper understanding of clock mechanisms and the treatment of diseases associated with circadian disruption.
Protein Processing, Post-Translational/physiology* ; Circadian Rhythm/physiology* ; Humans ; Animals ; CLOCK Proteins/physiology* ; Circadian Clocks/physiology* ; Phosphorylation ; Acetylation ; Ubiquitination ; Sumoylation

The aim of this study is to investigate the regulation of Kaixin San-medicated serum(KXS-MS) on autophagy induced by Aβ_(25-35) in SH-SY5Y cells. The SH-SY5Y cell model of Aβ_(25-35)(25 μmol·L~(-1))-induced injury was established, and different concentrations of KXS-MS were added into the culture media of cells, which were then incubated for 24 h. Cell viability was measured by the methyl thiazolyl tetrazolium(MTT) assay. The protein levels of microtubule-associated protein 1 light chain 3(LC3)Ⅰ, LC3Ⅱ, protein kinase B(Akt), p-Akt, mammalian target of rapamycin(mTOR), and p-mTOR were assessed by Western blot. Furthermore, the combination of rapamycin(Rapa)/3-methyladenine(3-MA) and low concentration of KXS-MS was added to the culture medium of SH-SY5Y cells injured by Aβ_(25-35), and the cell viability and the expression levels of the above proteins were determined. The results showed that Aβ_(25-35) decreased the cell viability, up-regulated the expression levels of LC3Ⅱ and LC3Ⅱ/LC3Ⅰ, and down-regulated the expression levels of p-Akt, p-mTOR, p-Akt/Akt, and p-mTOR/mTOR. Compared with the Aβ_(25-35) model group, KXS-MS treatment attenuated Aβ_(25-35)-induced injury and enhanced the survival of SH-SY5Y cells. Meanwhile, KXS-MS down-regulated the LC3Ⅱ/LC3Ⅰ level and up-regulated the p-Akt/Akt and p-mTOR/mTOR levels. Compared with the low-concentration KXS-MS group, Rapa did not affect the cell survival and the levels of p-Akt and p-Akt/Akt, while it up-regulated the levels of LC3Ⅱ and LC3Ⅱ/LC3Ⅰ and down-regulated the levels of p-mTOR and p-mTOR/mTOR. 3-MA significantly reduced the cell survival rate and p-Akt, p-Akt/Akt level in the KXS-MS group, while it had no significant effect on the levels of LC3Ⅱ, LC3Ⅱ/LC3Ⅰ, p-mTOR, and p-mTOR/mTOR. The above results indicate that KXS-MS exhibits protective effects against Aβ_(25-35)-induced damage in SH-SY5Y cells by up-regulating Akt/mTOR activity to inhibit autophagy.
Humans ; Autophagy/drug effects* ; TOR Serine-Threonine Kinases/genetics* ; Amyloid beta-Peptides/toxicity* ; Proto-Oncogene Proteins c-akt/genetics* ; Drugs, Chinese Herbal/pharmacology* ; Cell Line, Tumor ; Cell Survival/drug effects* ; Peptide Fragments/toxicity* ; Microtubule-Associated Proteins/genetics*

This study identified 8 members including NjBIN2 of the GSK3 family in Nardostachys jatamansi by bioinformatics analysis. Moreover, the phylogenetic tree revealed that the GKS3 family members of N. jatamansi had a close relationship with those of Arabidopsis. RT-qPCR results showed that NjBIN2 presented a tissue-specific expression pattern with the highest expression in roots, suggesting that NjBIN2 played a role in root growth and development. In addition, the application of epibrassinolide or the brassinosteroid(BR) synthesis inhibitor(brassinazole) altered the expression pattern of NjBIN2 and influenced the photomorphogenesis(cotyledon opening) and root development of N. jatamansi, which provided direct evidence about the functions of NjBIN2. In conclusion, this study highlights the roles of BIN2 in regulating the growth and development of N. jatamansi by analyzing the expression pattern and biological function of NjBIN2. It not only enriches the understanding about the regulatory mechanism of the growth and development of N. jatamansi but also provides a theoretical basis and potential gene targets for molecular breeding of N. jatamansi with improved quality in the future.
Brassinosteroids/metabolism* ; Steroids, Heterocyclic/metabolism* ; Gene Expression Regulation, Plant/drug effects* ; Plant Proteins/metabolism* ; Phylogeny ; Nardostachys/metabolism* ; Plant Growth Regulators/pharmacology* ; Plant Roots/drug effects*

Ecological cultivation is an important way for the sustainable production of traditional Chinese medicine in the context of the carbon peaking and carbon neutrality goals. Facility cultivation and simulative habitat cultivation modes have been developed and applied to develop the endangered Dendrobium huoshanense on the basis of protection. However, the differences in the greenhouse gas emissions and global warming potential of these cultivation modes remain unexplored, which limits the accurate assessment of carbon-friendly ecological cultivation modes of D. huoshanense. Greenhouse gas emission flux monitoring based on the static chamber method provides an effective way to solve this problem. Therefore, this study conducted a field experiment in the facility cultivation and simulative habitat cultivation modes at a D. huoshanense cultivation base in Dabie Mountains, Anhui Province. From April 2023 to March 2024, samples of greenhouse gases were collected every month, and the concentrations of CO_2, CH_4, and N_2O of the samples were then detected by gas chromatography. The greenhouse gas emission fluxes, cumulative emissions, and global warming potential were further calculated, and the following results were obtained.(1)The two cultivation modes of D. huoshanense showed significant differences in greenhouse gas emission fluxes, especially the CO_2 emission flux, with a pattern of facility cultivation>simulative habitat cultivation [(35.60±11.70)mg·m~(-2)·h~(-1) vs(2.10±4.59)mg·m~(-2)·h~(-1)].(2) The annual cumulative CO_2 emission flux in the case of facility cultivation was significantly higher than that of simulative habitat cultivation[(3 077.00±842.00)kg·hm~(-2) vs(221.00±332.00)kg·hm~(-2)], while no significant difference was found in annual cumulative CH_4 and N_2O emission fluxes.(3) The facility cultivation mode had a significantly higher global warming potential than the simulative habitat cultivation mode [(3 053.00±847.00)kg·hm~(-2) vs(196.00±362.00)kg·hm~(-2)]. Overall, the simulative habitat cultivation of D. huoshanense has obvious carbon-friendly characteristics compared with facility cultivation, which is in line with the concept of ecological cultivation of medicinal plants. This study is of great reference significance for the implementation and promotion of the ecological cultivation mode of D. huoshanense under carbon peaking and carbon neutrality goals.
Dendrobium/chemistry* ; Greenhouse Gases/metabolism* ; Carbon/analysis* ; Ecosystem ; Carbon Dioxide/metabolism* ; China ; Global Warming

To determine the optimal cultivation duration and harvest period for cultivated Notopterygium incisum and promote its industrial development, this study established a characteristic chromatographic profile of cultivated N. incisum and employed chemometrics combined with entropy-weighted grey correlation analysis to assess differences in agronomic traits and quality indicators across different cultivation years and harvest periods. By comparing with reference substances, ten common peaks were identified, including chlorogenic acid, p-coumaric acid, ferulic acid, marmesinin, nodakenin, isochlorogenic acid B, notopterol, phenethyl ferulate, isoimperatorin, and falcarindiol. The similarity between the characteristic chromatographic profiles of N. incisum at different cultivation years and the reference profile was all above 0.932. Principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA) revealed that the quality of 1-to 3-year-old cultivated N. incisum was highly dispersed and unstable, whereas the quality of 4-year-old cultivated N. incisum remained relatively stable across different harvest periods. This suggests that the accumulation of relevant compounds in the medicinal material had reached a plateau, confirming that the optimal cultivation period for N. incisum is four years. Entropy-weighted grey correlation analysis indicated that the quality of 4-year-old cultivated N. incisum across different harvest periods ranked from highest to lowest as follows: November, December, October, August, July, and September, demonstrating that November is the optimal harvest time. The findings of this study establish the suitable cultivation duration and optimal harvest period for N. incisum, providing a scientific basis for cultivation guidance and quality standardization.
Chromatography, High Pressure Liquid/methods* ; Apiaceae/chemistry* ; Entropy ; Chemometrics/methods* ; Drugs, Chinese Herbal/chemistry* ; Principal Component Analysis ; Quality Control

OBJECTIVE: Varied acupoint selections represent a potential cause of the uncertainty surrounding the efficacy of acupuncture for knee osteoarthritis (OA). Skin temperature, a guiding factor for acupoint selection, may help to address this issue. This study explored thermal sensitization of acupoints used for the treatment of knee OA. METHODS: This cross-sectional case-control study enrolled cases aged 45-75 years with symptomatic knee OA and age- and gender-matched non-knee OA controls in a 1:1 ratio. All participants underwent infrared thermographic imaging. The primary outcome was the relative skin temperature of acupoint (STA), and the secondary outcome was the absolute STA of 11 acupoints. The Z test was used to compare the relative and absolute STAs between the groups. Principal component analysis was used to extract the common factors (CFs, acupoint cluster) in the STAs. A general linear model was used to identify factors affecting the STA in the knee OA cases. For the group comparisons of relative STA, P < 0.0045 (adjusted for 11 acupoints through Bonferroni correction) was considered to indicate statistical significance. For other analyses, P < 0.05 was used as the threshold for statistical significance. RESULTS: The analysis included 308 participants, consisting of 151 cases (mean age: [64.58 ± 6.67] years; male: 25.83%; mean body mass index: [25.70 ± 3.16] kg/m²) and 157 controls (mean age: [63.37 ± 5.96] years; male: 26.11%; mean body mass index: [24.47 ± 2.84] kg/m²). The relative STAs of ST34 (P = 0.0001), EX-LE2 (P < 0.0001), EX-LE5 (P = 0.0006), SP10 (P < 0.0001), BL40 (P = 0.0012) and GB39 (P = 0.0037) were higher in the knee OA group. No difference was found in the STAs of ST35, ST36, SP9, GB33 and GB34. Four CFs were identified for relative STA in both groups. The acupoints within each CF were consistent between the groups. The mean values of the relative STAs across each CF were higher in the knee OA group. In the knee OA cases, no factors were observed to affect the relative STA, while age and gender were found to affect the absolute STA. CONCLUSION Among patients with knee OA, thermal sensitization occurs in the acupoints of the lower extremity, exhibiting localized and regional thermal consistencies. The thermally sensitized acupoints that we identified in this study, ST34, SP10, EX-LE2, EX-LE5, GB39 and BL40, may be good choices for the acupuncture treatment of knee OA. Please cite this article as: Tu JF, Wang XZ, Yan SY, Wang YR, Yang JW, Shi GX, Zhang WZ, Jing LN, Yang LS, Liu DH, Wang LQ, Mi BH. Thermal sensitization of acupoints in patients with knee osteoarthritis: A cross-sectional case-control study. J Integr Med. 2025; 23(3): 289-296.
Humans ; Osteoarthritis, Knee/physiopathology* ; Male ; Cross-Sectional Studies ; Middle Aged ; Female ; Acupuncture Points ; Case-Control Studies ; Aged ; Skin Temperature ; Acupuncture Therapy