Exploring the potential of the TCR repertoire as a tumor biomarker (Review).

T cells play an important role in adaptive immunity. Mature T cells specifically recognize antigens on major histocompatibility complex molecules through T-cell receptors (TCRs). As the TCR repertoire is highly diverse, its analysis is vital in the assessment of T cells. Advances in sequencing technology have provided convenient methods for further investigation of the TCR repertoire. In the present review, the TCR structure and the mechanisms by which TCRs function in tumor recognition are described. In addition, the potential value of the TCR repertoire in tumor diagnosis is reviewed. Furthermore, the role of the TCR repertoire in tumor immunotherapy is introduced, and the relationships between the TCR repertoire and the effects of different tumor immunotherapies are discussed. Based on the reviewed literature, it may be concluded that the TCR repertoire has the potential to serve as a biomarker for tumor prognosis. However, a wider range of cancer types and more diverse subjects require evaluation in future research to establish the TCR repertoire as a biomarker of tumor immunity.

Simulation-based caregiving skills training for family members of high-grade glioma patients.

Background: Because family caregivers of patients with a high-grade glioma experience high levels of distress and feel unprepared to perform the complex caregiving tasks associated with the disease and its treatment, we pilot-tested a caregiving skills intervention that integrates hands-on caregiving with coping skill training. Methods: In this single-arm trial, caregivers participated in a 4-session research nurse-led intervention involving simulation-based caregiving skills training at the hospital and psychoeducation delivered via videoconference. We collected measures of patients' and caregivers' psychological symptoms; caregivers' caregiving self-efficacy and role adjustment; and patients' cancer-related symptoms (MDASI) at baseline and again postintervention. We tracked feasibility data. Results: We approached 29 dyads of which 10 dyads (34%) consented. All patients (mean age: 60 years, 89% male) and caregivers (mean age: 58 years, 80% female, 80% spouses) completed the baseline and 7 dyads completed the follow-up assessments (attrition was related to patient's hospice transfer). Seven caregivers completed all 4 sessions and rated the program as beneficial. Paired t-tests revealed a significant improvement in caregiving self-efficacy at 6 weeks postintervention (t = -3.06, P = .02). Although improvements in caregiver role adjustment and patient and caregiver symptoms were not observed, no decreases in symptom burden or role adjustment were found during the follow-up period. Conclusions: This novel supportive care program appears to be safe, feasible, acceptable, and perceived as useful for caregivers of patients with high-grade glioma. Based on feasibility indicators and a signal of intervention efficacy, a randomized controlled trial is warranted.

The causal associations of inflammatory cytokines with obesity and systemic lupus erythematosus: A Mendelian randomization study.

OBJECTIVE: Previous studies have partly discussed the roles of inflammatory cytokines in obesity and systemic lupus erythematosus (SLE), but the causal relationship among inflammatory cytokines, obesity, and SLE is unclear. It is challenging to comprehensively evaluate the causal relationship between these variables. This study aimed to investigate the role of cytokines as intermediates between obesity and SLE. METHODS: The inverse-variance weighted method (IVW) of mendelian randomization (MR) is mainly used to explore the causal relationship between exposure and outcome by using the genetic variation of the open large genome-wide association studies (GWAS), namely single-nucleotide polymorphisms (SNPs) related to obesity (more than 600 000 participants), inflammatory cytokines (8293 healthy participants) and SLE (7219 cases). Methods such as weighted median, MR-Egger are used to evaluate the reliability of causality. Reverse analysis is performed for each MR analysis to avoid reverse causality. Cochran's Q statistic and funnel chart are used to detect heterogeneity, MR-Egger intercept test and leave-one-out sensitivity analyses evaluated pleiotropy. RESULTS: Obesity was associated with 25 cytokines, and 3 cytokines were associated with SLE, including CTACK (OR = 1.19, 95% CI: 1.06, 1.33, p = .002), IL-18 (OR = 1.13, 95% CI: 1.01, 1.26, p = .027), SCGFb (OR = 0.89, 95% CI: 0.79, 0.99, p = .044). In the opposite direction, SLE was associated with 18 cytokines, and 2 cytokines were associated with obesity, including IP-10 (ßIVW = -.03, 95% CI: -0.05, -0.01, p = .002), MIP-1B (ßIVW = -.03, 95% CI: -0.05, -0.01, p = .004). CONCLUSION: Our MR study suggested that CTACK, IL-18 and SCGFb may play an intermediary role in obesity to SLE, while IP-10 and MIP-1B may play an intermediary role in SLE to obesity.

Onabotulinum toxin injections for shoulder and chest wall muscle pain in breast cancer survivors: retrospective study - preliminary report.

OBJECTIVES: The primary objective of this retrospective review is to describe patient-reported improvement in muscular pain after initial treatment with onabotulinum toxin. A secondary objective was to determine other physiatry (physical medicine & rehabilitation (PM&R)) interventions ordered. METHODS: Preliminary retrospective review of physiatry interventions for 47 patients referred by breast radiation oncology to PM&R at a tertiary referral-based academic cancer centre clinic from 1 January 2018 to 31 December 2021 for muscular shoulder/chest wall pain. RESULTS: Patients were most commonly diagnosed with muscle spasm 27/47 (58%), lymphedema 21/47 (45%), myalgia/myofascial pain 16/47 (34%), radiation fibrosis 14/47 (30%), fatigue/deconditioning 13/47 (28%), neurological impairment 11/47 (23%) and joint pathology 3/47 (6%). The top three physiatric interventions were home exercise programme education (17/47, 36%), botulinum toxin injection (17/47, 36%) and physical or occupational therapy referral (15/47, 32%). Patients who had muscle spasms documented were more likely to have botulinum toxin recommended by physiatry (24/24) compared with those with questionable spasms (4/7) and those without spasms(0/16) (p=0.0005). 17/28 (60.7%) received botulinum toxin injection, and a total of 35 injections were performed during the study period. 94% (16/17) of patients who received botulinum toxin injection voiced improvement in pain after injection. CONCLUSION: Botulinum toxin injections may play a role in the treatment of muscle spasm-related pain in breast cancer survivors. Additional blinded controlled research on the effectiveness of botulinum toxin injection after breast cancer treatment with spastic muscular shoulder/chest wall pain is needed.

Liposomal nanocarriers of preassembled glycocalyx restore normal venular permeability in sepsis: assessed quantitatively with a novel microchamber system.

BACKGROUND: The endothelial glycocalyx (EG), covering the luminal side of endothelial cells, regulates vascular permeability and senses wall shear stress. In sepsis, EG undergoes degradation leading to increased permeability and edema formation. We hypothesized that restoring EG integrity using liposomal nanocarriers of preassembled glycocalyx (LNPG) will restore normal venular permeability in a lipopolysaccharide (LPS)-induced sepsis model of mice. METHODS: To test this hypothesis, we designed a unique perfusion microchamber in which permeability of isolated venules could be assessed by measuring the concentration of Evans blue dye (EBD) in microliter-samples of extravascular solution (ES). RESULTS: Histamine-induced time- and dose-dependent increases in EBD in the ES could be measured, confirming the sensitivity of the microchamber system. Notably, the histamine-induced increase in permeability was significantly attenuated by histamine receptor (H1) antagonist, triprolidine hydrochloride. Subsequently, mice were treated with LPS, or LPS + LNPG. Compared to control mice, venules from LPS-treated mice showed a significant increased permeability, which was significantly reduced by LNPG administration. Moreover, in the presence of wall shear stress, intraluminal administration of LNPG significantly reduced the permeability in isolated venules from LPS-treated mice. We have found no sex differences. CONCLUSION: Our newly developed microchamber system allows us to quantitatively measure the permeability of isolated mesenteric venules. LPS-induced sepsis increases permeability of venules that is attenuated by in vivo LNPG administration, which is also reestablished endothelial responses to shear stress. Thus, LNPG presents a promising therapeutic potential for restoring EG function and thereby mitigating vasogenic edema due to increased permeability in sepsis.

Assessment of rehabilitation practices during hematopoietic stem cell transplantation in the United States: a survey.

Background: Rehabilitation therapy is important to treat physical and functional impairments that may occur in individuals receiving physically taxing, yet potentially curative hematopoietic stem cell transplants (HSCT). However, there is scarce data on how rehabilitation is delivered during HSCT in real-life setting. Our objective is to assess the rehabilitation practices for adult patients hospitalized for HSCT in the United States. Methods: A 48-question online survey with cancer centers with the top 10% HSCT volumes (per American registries). We obtained data on patient characteristics, rehabilitation therapy details (timing, indication, administering providers), physical function objective and subjective outcome measures, and therapy activity precautions. Results: Fourteen (out of 21) institutions were included. Rehabilitation therapy referrals occurred at admission for all patients at 35.7% of the centers for: functional decline (92.9%), fall risk (71.4%), and discharge planning (71.4%). Participating institutions had physical therapists (92.9%), occupational therapists (85.7%), speech language pathologists (64.3%) and therapy aides (35.7%) in their rehabilitation team. Approximately 71% of centers used objective functional measures including sit-to-stand tests (50.0%), balance measures (42.9%), and six-minute walk/gait speed (both 35.7%). Monitoring of blood counts to determine therapy modalities frequently occurred and therapies held for low platelet or hemoglobin values; but absolute neutrophil values were not a barrier to participate in resistance or aerobic therapies (42.9%). Discussion: Rehabilitation practices during HSCT varied among the largest volume cancer centers in the United States, but most centers provided skilled therapy, utilized objective, clinician and patient reported outcomes, and monitored blood counts for safety of therapy administration.

Clinical Validation of a Deep Learning-Based Software for Lumbar Bone Mineral Density and T-Score Prediction from Chest X-ray Images.

Screening for osteoporosis is crucial for early detection and prevention, yet it faces challenges due to the low accuracy of calcaneal quantitative ultrasound (QUS) and limited access to dual-energy X-ray absorptiometry (DXA) scans. Recent advances in AI offer a promising solution through opportunistic screening using existing medical images. This study aims to utilize deep learning techniques to develop a model that analyzes chest X-ray (CXR) images for osteoporosis screening. This study included the AI model development stage and the clinical validation stage. In the AI model development stage, the combined dataset of 5122 paired CXR images and DXA reports from the patients aged 20 to 98 years at a medical center was collected. The images were enhanced and filtered for hardware retention such as pedicle screws, bone cement, artificial intervertebral discs or severe deformity in target level of T12 and L1. The dataset was then separated into training, validating, and testing datasets for model training and performance validation. In the clinical validation stage, we collected 440 paired CXR images and DXA reports from both the TCVGH and Joy Clinic, including 304 pared data from TCVGH and 136 paired data from Joy Clinic. The pre-clinical test yielded an area under the curve (AUC) of 0.940, while the clinical validation showed an AUC of 0.946. Pearson's correlation coefficient was 0.88. The model demonstrated an overall accuracy, sensitivity, and specificity of 89.0%, 88.7%, and 89.4%, respectively. This study proposes an AI model for opportunistic osteoporosis screening through CXR, demonstrating good performance and suggesting its potential for broad adoption in preliminary screening among high-risk populations.

Novel hypoxia- and lactate metabolism-related molecular subtyping and prognostic signature for colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is a serious global health burden because of its high morbidity and mortality rates. Hypoxia and massive lactate production are hallmarks of the CRC microenvironment. However, the effects of hypoxia and lactate metabolism on CRC have not been fully elucidated. This study aimed to develop a novel molecular subtyping based on hypoxia-related genes (HRGs) and lactate metabolism-related genes (LMRGs) and construct a signature to predict the prognosis of patients with CRC and treatment efficacy. METHODS: Bulk and single-cell RNA-sequencing and clinical data of CRC were downloaded from the TCGA and GEO databases. HRGs and LMRGs were obtained from the Molecular Signatures Database. The R software package DESeq2 was used to perform differential expression analysis. Molecular subtyping was performed using unsupervised clustering. A predictive signature was developed using univariate Cox regression, random forest model, LASSO, and multivariate Cox regression analyses. Finally, the sensitivity of tumor cells to chemotherapeutic agents before and after hypoxia was verified using in vitro experiments. RESULTS: We classified 575 patients with CRC into three molecular subtypes and were able to distinguish their prognoses clearly. The C1 subtype, which exhibits high levels of hypoxia, has a low proportion of CD8 + T cells and a high proportion of macrophages. The expression of immune checkpoint genes is generally elevated in C1 patients with severe immune dysfunction. Subsequently, we constructed a predictive model, the HLM score, which effectively predicts the prognosis of patients with CRC and the efficacy of immunotherapy. The HLM score was validated in GSE39582, GSE106584, GSE17536, and IMvigor210 datasets. Patients with high HLM scores exhibit high infiltration of CD8 + exhausted T cells (Tex), especially terminal Tex, and oxidative phosphorylation (OXPHOS)-Tex in the immune microenvironment. Finally, in vitro experiments confirmed that CRC cell lines were less sensitive to 5-fluorouracil, oxaliplatin, and irinotecan under hypoxic conditions. CONCLUSION: We constructed novel hypoxia- and lactate metabolism-related molecular subtypes and revealed their immunological and genetic characteristics. We also developed an HLM scoring system that could be used to predict the prognosis and efficacy of immunotherapy in patients with CRC.

Astrocytic LRP1 enables mitochondria transfer to neurons and mitigates brain ischemic stroke by suppressing ARF1 lactylation.

Low-density lipoprotein receptor-related protein-1 (LRP1) is an endocytic/signaling cell-surface receptor that regulates diverse cellular functions, including cell survival, differentiation, and proliferation. LRP1 has been previously implicated in the pathogenesis of neurodegenerative disorders, but there are inconsistencies in its functions. Therefore, whether and how LRP1 maintains brain homeostasis remains to be clarified. Here, we report that astrocytic LRP1 promotes astrocyte-to-neuron mitochondria transfer by reducing lactate production and ADP-ribosylation factor 1 (ARF1) lactylation. In astrocytes, LRP1 suppressed glucose uptake, glycolysis, and lactate production, leading to reduced lactylation of ARF1. Suppression of astrocytic LRP1 reduced mitochondria transfer into damaged neurons and worsened ischemia-reperfusion injury in a mouse model of ischemic stroke. Furthermore, we examined lactate levels in human patients with stroke. Cerebrospinal fluid (CSF) lactate was elevated in stroke patients and inversely correlated with astrocytic mitochondria. These findings reveal a protective role of LRP1 in brain ischemic stroke by enabling mitochondria-mediated astrocyte-neuron crosstalk.

Anti-inflammatory lindenane sesquiterpenoid dimers from the roots of Chloranthus holostegius var. trichoneurus.

Two new lindenane-type sesquiterpenoid dimers, chlotrichenes C and D (1 and 2) together with five known lindenane-type sesquiterpenoid dimers (3-7) were isolated from the roots of Chloranthus holostegius var. trichoneurus, a famous natural medicine named as "Sikuaiwa" for subduing swellings and relieving pain. The structures including absolute configuration were elucidated by their 1D and 2D NMR, HRESIMS, and ECD data. Compounds 1 and 2 were classical [4 + 2] lindenane-type sesquiterpenoid dimers that differed from known analogs in oxidation profile, side chain profile, and double bond position. The new isolates and compound 3 exhibited significant inhibitory activity on IL-1ß production (IC50: 1-15 µM) in LPS-induced THP-1 cells and other compounds exhibited inhibitory activity on NO production in LPS-induced RAW 264.7 cells (IC50: 24-33 µM).

A comprehensive benchmarking with interpretation and operational guidance for the hierarchy of topologically associating domains.

Topologically associating domains (TADs), megabase-scale features of chromatin spatial architecture, are organized in a domain-within-domain TAD hierarchy. Within TADs, the inner and smaller subTADs not only manifest cell-to-cell variability, but also precisely regulate transcription and differentiation. Although over 20 TAD callers are able to detect TAD, their usability in biomedicine is confined by a disagreement of outputs and a limit in understanding TAD hierarchy. We compare 13 computational tools across various conditions and develop a metric to evaluate the similarity of TAD hierarchy. Although outputs of TAD hierarchy at each level vary among callers, data resolutions, sequencing depths, and matrices normalization, they are more consistent when they have a higher similarity of larger TADs. We present comprehensive benchmarking of TAD hierarchy callers and operational guidance to researchers of life science researchers. Moreover, by simulating the mixing of different types of cells, we confirm that TAD hierarchy is generated not simply from stacking Hi-C heatmaps of heterogeneous cells. Finally, we propose an air conditioner model to decipher the role of TAD hierarchy in transcription.

Patterns of distant metastasis and survival outcomes in de novo metastatic breast cancer according to age groups.

Background: Is de novo metastatic breast cancer (dnMBC) the same disease in the elderly as in younger breast cancer remains unclear. This study aimed to determine the metastatic patterns and survival outcomes in dnMBC according to age groups. Methods: We included patients from the Surveillance Epidemiology and End Results program. Chi-square test, multivariate logistic regression analyses, and multivariate Cox regression models were used for statistical analyses. Results: A total of 17719 patients were included. There were 3.6% (n=638), 18.6% (n=3290), 38.0% (n=6725), and 39.9% (n=7066) of patients aged <35, 35-49, 50-64, and ≥65 years, respectively. Older patients had a significantly higher risk of lung metastasis and a significantly lower risk of liver metastasis. There were 19.1%, 25.6%, 30.9%, and 35.7% of patients with lung metastasis in those aged <35, 35-49, 50-64, and ≥65 years, respectively. Moreover, the proportion of liver metastasis was 37.6%, 29.5%, 26.3%, and 19.2%, respectively. Age was the independent prognostic factor associated with breast cancer-specific survival (BCSS) and overall survival (OS). Those aged 50-64 years had significantly inferior BCSS (P<0.001) and OS (P<0.001) than those aged <35 years. Patients aged ≥65 years also had significantly lower BCSS (P<0.001) and OS (P<0.001) than those aged <35 years. However, similar outcomes were found between those aged 35-49 and <35 years. Conclusion: Our study suggests that different age groups may affect the metastatic patterns among patients with dnMBC and the survival of younger patients is more favorable than those of older patients.

Prediction model for developing neuropsychiatric systemic lupus erythematosus in lupus patients.

OBJECTIVE: This study aimed to construct a predictive model for assessing the risk of development of neuropsychiatric systemic lupus erythematosus (NPSLE) among patients with SLE based on clinical, laboratory, and meteorological data. METHODS: A total of 2232 SLE patients were included and were randomly assigned into training and validation sets. Variables such as clinical and laboratory data and local meteorological data were screened by univariate and least absolute shrinkage and selection operator (LASSO) logistic regression modelling. After 10-fold cross-validation, the predictive model was built by multivariate logistic regression, and a nomogram was constructed to visualize the risk of NPSLE. The efficacy and accuracy of the model were assessed by receiver operating characteristic (ROC) curve and calibration curve analysis. Net clinical benefit was assessed by decision curve analysis. RESULTS: Variables that were included in the predictive model were anti-dsDNA, anti-SSA, lymphocyte count, hematocrit, erythrocyte sedimentation rate, pre-albumin, retinol binding protein, creatine kinase isoenzyme MB, Nterminal brain natriuretic peptide precursor, creatinine, indirect bilirubin, fibrinogen, hypersensitive C-reactive protein, CO, and mild contamination. The nomogram showed a broad prediction spectrum; the area under the curve (AUC) was 0.895 (0.858-0.931) for the training set and 0.849 (0.783-0.916) for the validation set. CONCLUSION: The model exhibits good predictive performance and will confer clinical benefit in NPSLE risk calculation. Key Points • Clinical, laboratory, and meteorological data were incorporated into a predictive model for neuropsychiatric systemic lupus erythematosus (NPSLE) in SLE patients. • Anti-dsDNA, anti-SSA, LYM, HCT, ESR, hsCRP, IBIL, PA, RBP, CO, Fib, NT-proBNP, Crea, CO, and mild contamination are predictors of the development of NPSLE and may have potential for research. • The nomogram has good predictive performance and clinical value and can be used to guide clinical diagnosis and treatment.

An updated advancement of bifunctional IL-27 in inflammatory autoimmune diseases.

Interleukin-27 (IL-27) is a member of the IL-12 family. The gene encoding IL-27 is located at chromosome 16p11. IL-27 is considered as a heterodimeric cytokine, which consists of Epstein-Barr virus (EBV)-induced gene 3 (Ebi3) and IL-27p28. Based on the function of IL-27, it binds to receptor IL-27rα or gp130 and then regulates downstream cascade. To date, findings show that the expression of IL-27 is abnormal in different inflammatory autoimmune diseases (including systemic lupus erythematosus, rheumatoid arthritis, Sjogren syndrome, Behcet's disease, inflammatory bowel disease, multiple sclerosis, systemic sclerosis, type 1 diabetes, Vogt-Koyanagi-Harada, and ankylosing spondylitis). Moreover, in vivo and in vitro studies demonstrated that IL-27 is significantly in3volved in the development of these diseases by regulating innate and adaptive immune responses, playing either an anti-inflammatory or a pro-inflammatory role. In this review, we comprehensively summarized information about IL-27 and autoimmunity based on available evidence. It is hoped that targeting IL-27 will hold great promise in the treatment of inflammatory autoimmune disorders in the future.

Long-term efficacy of liposomal nanocarriers of preassembled glycocalyx in restoring cerebral endothelial glycocalyx in sepsis.

The endothelial glycocalyx (EG) undergoes early degradation in sepsis. Our recent work introduced a novel therapeutic approach involving liposomal nanocarriers of preassembled glycocalyx (LNPG) to restore EG in lipopolysaccharide (LPS)-induced sepsis model of mice. While short-term effects were promising, this study focuses on the long-term impact of LNPG on mouse cerebral microcirculation. Utilizing cranial window, we assessed the stability of vascular density (VD) and perfused boundary region (PBR), an index of EG thickness, over a five-day period in normal control mice. In septic groups (LPS, LPS + 1-dose LNPG, and LPS + 2-dose LNPG), the exposure of mice to LPS significantly reduced VD and increased PBR within 3 h. Without LNPG treatment, PBR returned to the normal control level by endogenous processes at 48 h, associated with the recovery of VD to the baseline level at 72 h. However, mice receiving LNPG treatment significantly reduced the increment of PBR at 3 h. The therapeutic effect of 1-dose LNPG persisted for 6 h while the 2-dose LNPG treatment further reduced PBR and significantly increased VD at 12 h compared to LPS group. This study provides valuable insights into the potential therapeutic benefits of LNPG in mitigating EG degradation in sepsis.

Deep-potential enabled multiscale simulation of gallium nitride devices on boron arsenide cooling substrates.

High-efficient heat dissipation plays critical role for high-power-density electronics. Experimental synthesis of ultrahigh thermal conductivity boron arsenide (BAs, 1300 W m-1K-1) cooling substrates into the wide-bandgap semiconductor of gallium nitride (GaN) devices has been realized. However, the lack of systematic analysis on the heat transfer across the GaN-BAs interface hampers the practical applications. In this study, by constructing the accurate and high-efficient machine learning interatomic potentials, we perform multiscale simulations of the GaN-BAs heterostructures. Ultrahigh interfacial thermal conductance of 260 MW m-2K-1 is achieved, which lies in the well-matched lattice vibrations of BAs and GaN. The strong temperature dependence of interfacial thermal conductance is found between 300 to 450 K. Moreover, the competition between grain size and boundary resistance is revealed with size increasing from 1 nm to 1000 µm. Such deep-potential equipped multiscale simulations not only promote the practical applications of BAs cooling substrates in electronics, but also offer approach for designing advanced thermal management systems.

Integrated serum pharmacochemistry and network pharmacology to explore the mechanism of Yi-Shan-Hong formula in alleviating chronic liver injury.

BACKGROUND: Chronic liver injury (CLI) is a complex condition that requires effective therapeutic interventions. The Yi-Shan-Hong (YSH) formula is an empirically derived remedy that has shown effectiveness and safety in the management of chronic liver damage. However, the bioactive components and multifaceted mechanisms of YSH remain inadequately understood. PURPOSE: To examine the bioactive compounds and functional processes that contribute to the therapeutic benefits of YSH against CLI. METHODS: Serum pharmacochemistry and network pharmacology were employed to identify active compounds and possible targets of YSH in CLI. In addition, YSH was also given in three doses to d-(+)-galactosamine hydrochloride (D-GalN) -induced CLI rats to test its therapeutic efficacy. RESULTS: The analysis of serum samples successfully detected 25 compounds from YSH. Searches on the databases resulted in 277 genes as being correlated with chemicals in YSH, and 397 genes associated with CLI. In vivo experiments revealed that YSH displayed a notable therapeutic impact on liver injury caused by d-GalN. This was evidenced by enhanced liver function and histopathological improvements, reduced oxidative stress response, proinflammatory factors, and fibrosis levels. Importantly, no discernible adverse effects were observed. Furthermore, the administration of YSH treatment reversed the activation of AKT phosphorylation caused by d-GalN, aligning with the findings of the network pharmacology study. CONCLUSION: These findings provide preclinical evidence of YSH's therapeutic value in CLI and highlight its hepatoprotective action via the PI3K/AKT signaling pathway.