Over the past two decades, genome-wide association study(GWAS) has identified numerous genetic variants and loci associated with heritable diseases. With the gradual maturation and saturation of GWAS methodologies, transcriptome-wide association study(TWAS) offers a novel perspective by linkinggenetic phenotypes to gene expression levels. By integrating TWAS with other multi-omics analyses, researchers can gain a deeper understanding of heritable diseases. This article provides an overview of recent groundbreaking and representative TWAS methods and tools, analyzes their strengths and limitations, and discusses future trends in TWAS development.

Doxorubicin (DOX)-induced cardiotoxicity and sepsis-induced myocardial injury (SIMI) represent significant clinical challenges in patients undergoing chemotherapy, sharing a common pathological basis of oxidative stress and mitochondrial dysfunction. Ferroptosis, an iron-dependent form of regulated cell death driven by lipid peroxidation, has recently been shown to play a critical role in DOX-induced cardiotoxicity and lipopolysaccharide (LPS)-induced SIMI. This article systematically reviews the mechanisms underlying myocardial injury caused by DOX and sepsis, identifying ferroptosis as a central common pathway. DOX triggers a burst of reactive oxygen species within mitochondria and inhibits glutathione peroxidase 4 (GPX4) activity through redox cycling of its quinone group and high-affinity accumulation in mitochondrial cardiolipin. LPS, by activating pattern recognition receptors and related inflammatory signaling pathways, provokes a cytokine storm and mitochondrial dysfunction. Both can disrupt the core regulatory axis of cysteine-glutathione (GSH)-GPX4, synergistically promoting ferroptosis in cardiomyocytes. Moreover, epigenetic regulation plays a key role in DOX- and LPS-induced cardiomyocyte ferroptosis and may serve as a promising therapeutic target. A deeper understanding of the ferroptosis mechanism and its epigenetic regulatory network in the synergistic injury induced by DOX and sepsis is of great importance for developing novel strategies to mitigate chemotherapy-related cardiotoxicity and improve outcomes in cancer patients with concurrent infections.

Objective: To investigate the distribution characteristics of ABO and Rh blood group antigen phenotypes among blood donors in the Yantai, Shandong. Methods: Blood samples from 310 180 voluntary blood donors in Yantai collected from January 2019 to December 2023 were tested for ABO and Rh blood group antigens using standard serological methods. RhD-negative samples were further typed for C, c, E, and e antigens. Population genetic analysis of blood groups was performed: allele frequencies were inferred from ABO phenotypes, and Rh allele/haplotype frequencies were estimated based on the proportion of RhD-negative donors and CcEe antigen typing, followed by Hardy-Weinberg equilibrium testing. Results: The phenotypic distribution frequency of ABO blood groups was B(32.72%)>O(28.93%)>A(27.65%)>AB(10.70%). The inferred allele frequencies were r(53.74%)>q(24.78%)>p(21.48%), consistent with Hardy-Weinberg equilibrium (P>0.05). A total of 1 872 Rh-negative donors (0.603%) were identified. The most common Rh phenotypes were ccdee (59.56%) and Ccdee (30.18%). The distribution of Rh antigen phenotypes deviated significantly from Hardy-Weinberg equilibrium (χ =37.15, P<0.001), with the cde haplotype showing the highest frequency. There was no statistically significant difference in ABO blood group distribution between RhD-positive and RhD-negative donors (P>0.05). Conclusion: The ABO blood group distribution among voluntary blood donors in Yantai is generally stable and consistent with population genetic equilibrium, whereas the Rh antigen phenotype distribution deviates from equilibrium, indicating potential underlying genetic structural differences.

ObjectiveThis paper aims to find the identified and validated clinical biomarker data building upon a clinical study of early-phase phase Ⅱ and investigate the correlation analysis of Huanglian Jiedu Wan on syndrome improvement and clinical biomarkers in the treatment of "excess heat-toxicity" based on a machine learning model. Additionally， the effective prediction of clinical biomarker values for the main symptoms of the "excess heat-toxicity" syndrome was assessed. MethodsA total of 229 patients meeting the inclusion criteria for "excess heat-toxicity" syndrome were randomly divided into the Huanglian Jiedu Wan group and the placebo group. Syndrome score transition matrices were constructed for the Huanglian Jiedu Wan group and the placebo group based on three main symptoms of "excess heat-toxicity" syndrome， such as oral ulcers， sore throat， and gum swelling and pain. Data from the patients with these three syndromes were also integrated for an overall analysis. The corresponding syndrome score transition matrices were further constructed to visualize symptom change trends of the patients in the two groups via heatmaps. Based on the identified and validated clinical biomarkers related to inflammation， oxidative stress， and energy metabolism in the early phase， Spearman correlation analysis was employed to analyze and evaluate the associations between clinical biomarkers and syndrome improvement. Key clinical biomarkers reflecting the effect of Huanglian Jiedu Wan were screened through the comparison of differences between groups. An extreme gradient boosting （XGBoost） algorithm was used to develop a prediction model for main symptom classification， with classification performance evaluated through 10-fold cross-validation. Feature importance analysis was applied to identify variables with the greatest contribution to the prediction result. ResultsThe syndrome transition matrix results indicated that the Huanglian Jiedu Wan group showed a superior effect to the placebo group in improving oral ulcers， sore throat， and overall symptoms， with significant effects observed especially in sore throat and overall symptom analyses （P<0.01）. Spearman correlation analysis revealed that several clinical biomarkers positively correlated with "excess heat-toxicity" syndrome and its main symptom improvement， were also called "heat-related biomarkers"， including succinic acid， α-ketoglutaric acid， glycine， lactic acid， adenosine monophosphate （AMP）， tumor necrosis factor-α （TNF-α）， interferon-γ （IFN-γ）， interleukin-1β （IL-1β）， interleukin-4 （IL-4）， interleukin-6 （IL-6）， interleukin-8 （IL-8）， interleukin-10 （IL-10）， and so on. Conversely， clinical biomarkers negatively correlated with symptom severity， were also called "heat-clearing related biomarkers" after administration of Huanglian Jiedu Wan， including malic acid， fumaric acid， cis-aconitic acid， adrenocorticotropic hormone （ACTH）， IL-1β， IL-4， IL-8， succinic acid， and citric acid. The XGBoost classification model using all 52 biomarkers as variables achieved an average test accuracy of 0.754 and an average F1 score of 0.777. Feature importance analysis identified the scores of glutamic acid in saliva and IL-6 were the highest in all the variables， with importance scores of 0.081 and 0.080， respectively. After screening out 14 key variables and optimizing the parameters， model performance improved to an average accuracy of 0.758 and an F1 score of 0.798. Feature importance analysis further determined that the glutamic acid in saliva and IL-6 showed obvious changes after screening the variables， confirming the good syndrome prediction ability of the model constructed by these key clinical biomarkers. ConclusionThis study systematically elucidates the correlation between syndrome improvement and clinical biomarkers of Huanglian Jiedu Wan in the treatment of "excess heat-toxicity" syndrome. An XGBoost classification model based on key clinical biomarkers is successfully established， achieving effective prediction of the symptoms related to the "excess heat-toxicity" syndrome such as oral ulcers and sore throat and providing a new insight for objective identification of traditional Chinese medicine syndromes.

ObjectiveThis paper aims to find the identified and validated clinical biomarker data building upon a clinical study of early-phase phase Ⅱ and investigate the correlation analysis of Huanglian Jiedu Wan on syndrome improvement and clinical biomarkers in the treatment of "excess heat-toxicity" based on a machine learning model. Additionally， the effective prediction of clinical biomarker values for the main symptoms of the "excess heat-toxicity" syndrome was assessed. MethodsA total of 229 patients meeting the inclusion criteria for "excess heat-toxicity" syndrome were randomly divided into the Huanglian Jiedu Wan group and the placebo group. Syndrome score transition matrices were constructed for the Huanglian Jiedu Wan group and the placebo group based on three main symptoms of "excess heat-toxicity" syndrome， such as oral ulcers， sore throat， and gum swelling and pain. Data from the patients with these three syndromes were also integrated for an overall analysis. The corresponding syndrome score transition matrices were further constructed to visualize symptom change trends of the patients in the two groups via heatmaps. Based on the identified and validated clinical biomarkers related to inflammation， oxidative stress， and energy metabolism in the early phase， Spearman correlation analysis was employed to analyze and evaluate the associations between clinical biomarkers and syndrome improvement. Key clinical biomarkers reflecting the effect of Huanglian Jiedu Wan were screened through the comparison of differences between groups. An extreme gradient boosting （XGBoost） algorithm was used to develop a prediction model for main symptom classification， with classification performance evaluated through 10-fold cross-validation. Feature importance analysis was applied to identify variables with the greatest contribution to the prediction result. ResultsThe syndrome transition matrix results indicated that the Huanglian Jiedu Wan group showed a superior effect to the placebo group in improving oral ulcers， sore throat， and overall symptoms， with significant effects observed especially in sore throat and overall symptom analyses （P<0.01）. Spearman correlation analysis revealed that several clinical biomarkers positively correlated with "excess heat-toxicity" syndrome and its main symptom improvement， were also called "heat-related biomarkers"， including succinic acid， α-ketoglutaric acid， glycine， lactic acid， adenosine monophosphate （AMP）， tumor necrosis factor-α （TNF-α）， interferon-γ （IFN-γ）， interleukin-1β （IL-1β）， interleukin-4 （IL-4）， interleukin-6 （IL-6）， interleukin-8 （IL-8）， interleukin-10 （IL-10）， and so on. Conversely， clinical biomarkers negatively correlated with symptom severity， were also called "heat-clearing related biomarkers" after administration of Huanglian Jiedu Wan， including malic acid， fumaric acid， cis-aconitic acid， adrenocorticotropic hormone （ACTH）， IL-1β， IL-4， IL-8， succinic acid， and citric acid. The XGBoost classification model using all 52 biomarkers as variables achieved an average test accuracy of 0.754 and an average F1 score of 0.777. Feature importance analysis identified the scores of glutamic acid in saliva and IL-6 were the highest in all the variables， with importance scores of 0.081 and 0.080， respectively. After screening out 14 key variables and optimizing the parameters， model performance improved to an average accuracy of 0.758 and an F1 score of 0.798. Feature importance analysis further determined that the glutamic acid in saliva and IL-6 showed obvious changes after screening the variables， confirming the good syndrome prediction ability of the model constructed by these key clinical biomarkers. ConclusionThis study systematically elucidates the correlation between syndrome improvement and clinical biomarkers of Huanglian Jiedu Wan in the treatment of "excess heat-toxicity" syndrome. An XGBoost classification model based on key clinical biomarkers is successfully established， achieving effective prediction of the symptoms related to the "excess heat-toxicity" syndrome such as oral ulcers and sore throat and providing a new insight for objective identification of traditional Chinese medicine syndromes.

ObjectiveTaking the cuproptosis/oxidative stress pathway as the entry point， this study investigated the effect and mechanism of Liangyi Paste on hepatic lipid deposition in naturally aged mice fed with a high-fat diet. MethodsAfter adaptive feeding， 80 ten-week-old male C57BL/6 mice were used. Thirty of them were randomly divided into three groups （10 mice per group）： The 12-month-old control group （12MCON）， the 15-month-old control group （15MCON）， and the 15-month-old group with a high-fat diet （15MHFD）. The 12MCON and 15MCON groups were continuously fed a standard diet， while the 15MHFD group started receiving a high-fat diet at 12 months of age. Tissue samples were collected at the corresponding time points for each group. The remaining 50 mice were randomly divided into five groups （10 mice per group）： the 20-month-old control group （20MCON）， the model group， and the low-， medium-， and high-dose Liangyi Paste groups （2.91 ， 5.82 ， 11.64 g·kg^-1·d^-1， respectively）. The 20MCON group was continuously fed a standard diet， while the other groups started receiving a high-fat diet at 15 months of age. At 18 months of age， the Liangyi Paste groups were administered the corresponding doses of Liangyi Paste by gavage， while the 20MCON and model groups were given an equal volume of saline by gavage. After 8 weeks of continuous gavage （when the mice reached 20 months of age）， tissue samples were collected. Hepatic TG levels were measured using assay kits； liver histology and lipid deposition were observed via hematoxylin-eosin （HE） and oil red O staining； reactive oxygen species （ROS） were detected by enzyme-linked immunosorbent assay （ELISA）； Cu²⁺， superoxide dismutase （SOD）， and malondialdehyde （MDA） levels were measured by colorimetry； mRNA and protein expression of genes related to cuproptosis and oxidative stress pathways were analyzed by Real-time polymerase chain reaction（Real-time PCR） and Wes automated protein expression system. ResultsCompared with 12MCON， the 15MCON group showed significantly increased hepatic TG， Cu²⁺， ROS， and MDA levels （P<0.01）， decreased SOD （P<0.01）， hepatocyte swelling， and disordered arrangement. The mRNA and protein levels of ferredoxin 1 （FDX1）， dihydrolipoamide S-acetyltransferase （DLAT）， heat shock protein 70 （HSP70）， dihydrolipoamide dehydrogenase （DLD）， pyruvate dehydrogenase E1 subunit-β （PDHB）， nuclear factor erythroid 2-related factor 2 （Nrf2）， and peroxisome proliferator-activated receptor γ （PPARγ） were significantly elevated （P<0.05， P<0.01）. Compared with 15MCON group， the 15MHFD and 20MCON groups exhibited further increases in TG， Cu²⁺， ROS， and MDA （P<0.01）， reduced SOD （P<0.01）， and aggravated hepatocyte swelling and disorder. There were increased lipid droplets with mild vacuolization in the 15MHFD group， and no significant lipid deposition was observed in the 20MCON group. FDX1， DLAT， HSP70， DLD， PDHB， Nrf2， and PPARγ mRNA and protein levels were significantly increased （P<0.05， P<0.01）. Compared with 20MCON group， the model group demonstrated markedly elevated TG， Cu²⁺， ROS， and MDA （P<0.01）， reduced SOD （P<0.01）， severe hepatic steatosis， and upregulated expression of FDX1， DLAT， HSP70， DLD， PDHB， Nrf2， and PPARγ mRNA and proteins （P<0.05， P<0.01）. All abnormalities were significantly reversed after Liangyi Paste treatment. ConclusionLiangyi paste can ameliorate hepatic lipid deposition in naturally aged mice with a high-fat diet by modulating the cuproptosis/oxidative stress pathway.

OBJECTIVE To analyze the prototype components absorbed into blood and brain of Lithocarpus litseifolius leaves， so as to provide a reference for clarifying the pharmacological material basis of its prevention and treatment of central nervous system dis eases. METHODS The ethanol extract of L. litseifolius leaves， as well as the gastric lavage fluid and perfusion solution were prepared. Using rats as subjects， plasma samples of intestinal wall metabolism， intestinal flora metabolism and hepatic metabolism were prepared via in situ intestinal perfusion and closed intestinal loop method； while comprehensive metabolic plasma samples， brain tissue samples， and cerebrospinal fluid samples were collected after intragastric administration. UPLC-HRMS technology was utilized to analyze and identify chemical components and prototype components absorbed into blood and brain of L. litseifolius leaves. RESULTS A total of 66 chemical constituents were identified in L. litseifolius leaves， primarily consisting of flavonoids， organic acids， and others. A total of 16， 13， 11， and 5 prototype components were identified in intestinal wall metabolism， intestinal flora metabolism， hepatic metabolism， and comprehensive metabolic plasma samples， respectively. Additionally， 4 prototype components were detected in brain tissue and 9 in cerebrospinal fluid. Phloridzin， trilobatin， phloretin-2- O -malonyl hexoside， and phloretin were identified as common components across all sample types. CONCLUSIONS Prototype components absorbed into blood and brain of L. litseifolius leaves， such as phloridzin， trilobatin， phloretin， and other components may serve as the pharmacological material basis for their therapeutic effects on central nervous system diseases.

OBJECTIVE To study the improvement effects and potential mechanisms of water extract from Chrysanthemum morifolium on skeletal muscle atrophy in rats after ischemic stroke. METHODS Sprague-Dawley rats were randomly divided into sham operation group， model group， ATP group （10 mg/kg）， C. morifolium water extract high-dose and low-dose groups （1.08， 0.54 g/kg）. Except for sham operation group， ischemic stroke models were induced in rats from the other groups using middle cerebral artery occlusion. Starting from the first day after surgery， rats in each group were given corresponding drug/normal saline intragastrically， once a day， for consecutive 7 days. On the 7th day post-surgery， the rats’ body weights were measured， and their motor functions were evaluated， including Longa scores， exercise distance， grip strength； the electrophysiological signals of the skeletal muscles in rats were measured； the pathological morphology of the soleus muscle in rats was observed； the levels of tumor necrosis factor-α （TNF-α） in serum and soleus muscle were measured； the expressions of proteins related to TNF-α/c-Jun N- terminal kinase （JNK）/mitogen-activated protein kinase （MAPK） signaling pathway in the soleus muscle were determined. RESULTS Compared with sham operation group， the body weight， grip strength and exercise distance of rats were decreased/ shortened significantly （P＜0.01）； additionally， there was a notable reduction in the interpeak value of skeletal muscle electrophysiology （P＜0.05 or P＜0.01）. Longa score， as well as the levels of TNF-α in serum and soleus muscle， and the expression levels of TNF-α， phosphorylated JNK， phosphorylated MAPK， muscle ring-finger protein-1， and muscle atrophy Fbox- 1 protein in the soleus muscle， were all significantly elevated （P＜0.01）. The skeletal muscle cells of the soleus muscle in the model group showed significant atrophy， with a markedly decreased cross-sectional area （P＜0.01）. Compared with the model group， the levels of the aforementioned indicators were significantly reversed in C. morifolium water extract groups （P＜0.05 or P＜ 0.01）， and the skeletal muscle cells of the soleus muscle were markedly enlarged. CONCLUSIONS C. morifolium water extract can improve skeletal muscle atrophy in rats after ischemic stroke， the mechanism of which may be associated with suppressing the activation of the TNF- α/JNK/MAPK E-mail：19932015@cdutcm.edu.cn signaling pathway.

Poloxamer， as a non-ionic surfactant， exhibits a unique triblock [polyethylene oxide-poly （propylene oxide）-polyethylene oxide] structure， which endows it with broad application potential in various fields， including solid dispersion technology， nanotechnology， gel technology， biologics， gene engineering and 3D printing. As a carrier， it enhances the solubility and bioavailability of poorly soluble drugs. In the field of nanotechnology， it serves as a stabilizer etc.， enriching preparation methods. In gel technology， its self-assembly behavior and thermosensitive properties facilitate controlled drug release. In biologics， it improves targeting efficiency and reduces side effects. In gene engineering， it enhances delivery efficiency and expression levels. In 3D printing， it provides novel strategies for precise drug release control and the production of high-quality biological products. As a versatile material， poloxamer holds promising prospects in the pharmaceutical field.

Objective This study aims to investigate the effects of varying durations of light exposure on body weight and learning and memory abilities of pubertal NIH mice. Methods Forty pubertal NIH mice, evenly split by gender and with similar initial weights, were subjected to a 12 h light-dark cycle for one week. They were then randomly assigned to groups with daily light exposure durations of 0, 6, 12, 18, and 24 hours, with 8 mice in each group. The experimental period lasted for 7 weeks, with the first 5 weeks as the feeding phase under different light exposure conditions, and the last 2 weeks as the behavioral testing phase. Their body weight was monitored, and learning and memory abilities were assessed using the T-maze, object location test, and eight-arm maze tests. Results During the light exposure period, there were no significant differences in body weight among groups (P>0.05). However, the weight gain of mice in the 24 h group was significantly higher than that of the 0 h group and the 6 h group during the second and third weeks of light exposure (P<0.05). After five weeks of light exposure, in the T-maze test, the latency time of the 0 h light exposure group was significantly longer than that of the 12 h group (P<0.01), and the latency time of the 24 h light exposure group was significantly longer than that of the 12 h group (P<0.05). In the object location test, the mice in 12 h group exhibited a higher discrimination index and spent more time observing the new location compared to the other groups, with significant differences in comparison to the 18 h group (P<0.01) and the 24 h group (P<0.05). In the eight-arm maze test, the time to find food, the reference memory error rate, and the working memory error rate in the 12 h group were all lower than those in the 0 h group, with significant differences (P<0.05). Moreover, the working memory error rate in the 24 h group was higher than that in the 12 h group, with significant differences (P<0.05). Conclusion Continuous 24 h light exposure affects body weight gain, while light exposure durations exceeding 18 h or below 6 h per day weaken the learning and memory abilities of NIH mice.