Effect of cervical pillow in phacoemulsification surgery for age-related cataract patients: a prospective randomized controlled study.

PURPOSE: To explore the application effect of cervical pillow in phacoemulsification surgery for age-related cataract patients. METHODS: 104 cases of age-related cataract patients admitted to our hospital in June 2023 were enrolled and divided into the control group (traditional supine position) and the experimental group (the cervical pillow supine position) by the digital parity method (52 cases per group). The two groups were evaluated for the discomfort score, the satisfaction of patients and doctors, the head displacement rate, the number of displacement, the operation time and the time of body position during the operation and after the operation. RESULTS: There was no significant difference in the gender (P = 0.84), age (P = 0.86), course of disease (P = 0.82) and the time spent on position placement (P = 0.15) of the two groups. The patient in the experimental group had lower discomfort score (P = 0.0001), higher patients satisfaction (P = 0.0001) and higher doctors satisfaction (P = 0.0001) than patients in the control group. There was no significant difference between the experimental group and the control group in the proportion of intraoperative (P = 0.36) and postoperative pain (P = 0.65). Besides, the number of head transfers (P = 0.001), number of head shifts (P = 0.0001), the surgical time (P = 0.0001) and laparoscopic time (P = 0.0001) in the experimental group were significantly lower than those in the control group. CONCLUSION: The additional cervical pillow for age-related cataract patients in the traditional supine position during the operation will not increase the preparation time before the operation, but will help improve the patient satisfaction, improve the comfort and maintain a good position of the operative eye field, bringing obvious comfort and smooth operation for the surgeon in the operation, thus reducing the risk of the operation, shortening the operation time.

Qizhu anticancer prescription enhances immunosurveillance of liver cancer cells by regulating p21-dependent secretory phenotypes.

ETHNOPHARMACOLOGICAL RELEVANCE: Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related death worldwide, largely due to the limitations of available therapeutic strategies. The traditional Chinese medicine Qizhu Anticancer Prescription (QZACP) can improve the quality of life and prolong the survival time of patients with HCC. However, the precise mechanisms underlying the anti-cancer properties of QZACP remain unclear. PURPOSE: This study examined the anti-hepatocarcinogenic properties of QZACP, with a specific focus on its influence on the p21-activated secretory phenotype (PASP)-mediated immune surveillance, to elucidate the underlying molecular pathways involved in HCC. MATERIALS AND METHODS: Cell proliferation was measured using the Cell Counting Kit-8, 5-ethynyl-2'-deoxyuridine, and clonogenic assays. The cell cycle was evaluated using flow cytometry, and senescence was identified by staining with senescence-associated beta-galactosidase (SA-ß-gal). A primary liver cancer model produced by diethylnitrosamine was established in C57 BL/6 mice to assess the tumor-inhibitory effect of QZACP. The liver's pathological characteristics were examined using hematoxylin and eosin staining. PASP screening was performed using GeneCards, DisGeNet, Online Mendelian Inheritance in Man, and The Cancer Genome Atlas databases. Western blot analysis, enzyme-linked immunosorbent assay (ELISA), immunofluorescence staining, and Transwell migration assays were performed. RESULTS: Serum containing QZACP enhanced p21 expression, triggered cell cycle arrest, accelerated cell senescence, and suppressed cell proliferation in Huh7 and MHCC-97H liver cancer cells. QZACP reduced the quantity and dimensions of liver tumor nodules and enhanced p21 protein expression, SA-ß-Gal staining in tumor lesions, and cytotoxic CD8+ T cell infiltration. Bioinformatic analyses indicated that PASP factors, including hepatocyte growth factor, decorin (DCN), dermatopontin, C-X-C motif chemokine ligand 14 (CXCL14), and Wnt family member 2 (WNT2), play an important role in the development of HCC. In addition, these factors are associated with the presence of natural killer cells and CD8+ T cells within tumors. Western blotting and ELISA confirmed that QZACP increased DCN, CXCL14, and WNT2 levels in tumor tissues and peripheral blood. CONCLUSIONS: QZACP's suppression of HCC progression may involve cell senescence mediated via p21 upregulation, DCN, CXCL14, and WNT2 secretion, and reversal of the immunosuppressive microenvironment. This study provides insights that can be used in the development of new treatment strategies for HCC.

Benefit-finding profiles and comparison of caregiving ability among informal caregivers of patients with lung cancer: A latent profile analysis.

Objective: This study aimed to explore the benefit finding (BF) profiles among informal caregivers of patients with lung cancer, identify demographic and disease characteristics, and analyze differences in caregiving ability between profiles. Methods: This cross-sectional study utilized convenience sampling to select 272 informal caregivers of patients with lung cancer from a tertiary care hospital in Guangzhou, China. The research instruments used included the Demographic and Disease Characteristics Questionnaire, the revised version of the BF Scale, and the Chinese version of the Family Caregiver Task Inventory. Data analysis was performed using latent profile analysis, chi-square test, Fisher's exact probability test, Kruskal-Wallis test, and multivariate logistic regression. Results: (1) BF can be divided into three profiles: "high benefit-family and personal growth" (Profile 1, 7.7%), "moderate benefit-unclear perception" (Profile 2, 44.9%), and "low benefit-coping ability deficient" (Profile 3, 47.4%). (2) Having a cocaregiver and a disease duration of 6-12 months were more likely to belong to Profile 1; caregivers of patients aged 40-60 years tended to belong to Profile 2; caregivers of older patients with disease duration > 12 months and clinical stage II or III were more likely to belong to Profile 3. (3) There were significant differences in the total score of caregiving ability and the scores of each dimension among the different BF profiles (P < 0.001), and the caregiving abilities of Profile 1 and Profile 2 were higher than those of Profile 3. Conclusions: There was heterogeneity in BF among informal caregivers of patients with lung cancer. Healthcare professionals can identify the key profiles of lung-cancer caregivers based on characteristics such as age, clinical stage, disease duration, and cocaregiver status and enhance their caregiving ability through targeted nursing guidance.

Exosomal ncRNAs: Multifunctional contributors to the immunosuppressive tumor microenvironment of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a malignant liver cancer characterized by aggressive progression, unfavorable prognosis, and an increasing global health burden. Therapies that precisely target immunological checkpoints and immune cells have gained significant attention as possible therapeutics in recent years. In truth, the efficacy of immunotherapy is heavily contingent upon the tumor microenvironment (TME). Recent studies have indicated that exosomes serve as a sophisticated means of communication among biomolecules, executing an essential part in the TME of immune suppression. Exosomal non-coding RNAs (ncRNAs) can induce the activation of tumor cells and immunosuppressive immune cells that suppress the immune system, such as cancer-associated fibroblasts (CAFs), tumor-associated macrophages (TAMs), tumor-associated neutrophils (TANs), CD+8 T cells, regulatory T cells (Tregs), and regulatory B cells (Bregs). This cell-cell crosstalk triggered by exosomal ncRNAs promotes tumor proliferation and metastasis, angiogenesis, malignant phenotype transformation, and drug resistance. Hence, it is imperative to comprehend how exosomal ncRNAs regulate tumor cells or immune cells within the TME to devise more comprehensive and productive immunotherapy programs. This study discusses the features of exosomal ncRNAs in HCC and how the activation of the exosomes redefines the tumor's immunosuppressive microenvironment, hence facilitating the advancement of HCC. Furthermore, we also explored the potential of exosomal ncRNAs as a viable biological target or natural vehicle for HCC therapy.

Action Speaks Louder: The Role of Proactive Behavior between Creative Leadership and Employees' Creativity.

Drawing on social learning theory, this study presents a moderated mediation model to examine the role of proactive behavior and conformity value in the positive relationship between creative leadership and employees' creativity. Two-wave data are collected from 230 employees and their leaders in an automobile manufacturing enterprise in China, in which employees are encouraged to be creative because they need to implement novel designs and proposals to attract consumers. Statistical analysis reveals that proactive behavior partially mediates the influence of creative leadership on employees' creativity. Conformity value moderates not only the relationship between creative leadership and proactive behavior but also the indirect effect of creative leadership on employees' creativity via proactive behavior. The relationship and the mediating mechanism are stronger in the presence of employees' higher conformity value. We discuss the theoretical and practical implications as well as future research directions of the findings.

Investigating the molecular mechanism of Qizhu anticancer prescription in inhibiting hepatocellular carcinoma based on high-resolution mass spectrometry and network pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: Of all primary liver cancer cases, hepatocellular carcinoma (HCC) accounts for about 90%. Most patients with HCC receive a diagnosis in the medium-to-late stages or with chronic liver disease, have lost the opportunity for radical treatment, such as surgical resection, and their 5-year survival rate is low. Qizhu Anticancer Prescription (QZACP) is an empirical formula composed of traditional Chinese herbs that can clinically relieve HCC symptoms, inhibit the progression of HCC, reduce recurrence rate, and prolong survival; however, its exact mode of action remains unknown. AIM OF THE STUDY: This study's purpose was to investigate the mode of action of QZACP in the prevention and treatment of HCC. MATERIALS AND METHODS: Initially, drug components in the QZACP decoction were analyzed using high-resolution mass spectrometry. A subcutaneous tumor xenograft model in nude mice was constructed to further analyze the active components of QZACP that had entered tumor tissues through oral administration. Potential targets of QZACP in the prevention and treatment of HCC were identified and then confirmed in vivo via network pharmacology and molecular docking. In addition, regulatory effects of QZACP on HCC cell proliferation and the cell cycle were detected using a CCK-8 assay and flow cytometry. RESULTS: High-resolution mass spectrometry revealed that the QZACP decoction contained deacetyl asperulosidic acid methyl ester (DAAME), paeoniflorin, calycosin-7-glucoside, liquiritin, glycyrrhizic acid, astragaloside IV, saikosaponin A, curdione, and atractylenolide II. In nude mice, QZACP could effectively inhibit the growth of subcutaneous tumors, where DAAME, paeoniflorin, liquiritin, and glycyrrhizic acid could enter liver cancer tissues after oral administration. Among these, DAAME was the most highly expressed in HCC tissues and may be an important active component of QZACP for inhibiting HCC. Utilizing network pharmacology, the targets of action of these four drug components were identified. After verification using western blotting, STAT3, VEGFA, JUN, FGF2, BCL2L1, AR, TERT, MMP7, MMP1, ABCB1, CA9, and ESR2 were identified as targets of QZACP inhibition in HCC. In vitro experiments revealed that QZACP inhibited the proliferation of HCC cells while inducing G0/G1 phase cell cycle arrest. In vivo experiments demonstrated that DAAME significantly inhibited HCC growth. After intersection of the 24 DAAME targets predicted using network pharmacology with the 435 HCC disease targets, only CA9 was identified as a DAAME-HCC crossover target. Molecular docking results revealed that the binding site of DAAME and CA9 had good stereo-complementarity with a docking score of -8.1 kcal/mol. Western blotting and immunohistochemical results also confirmed that DAAME significantly decreased CA9 protein expression in HCC. CONCLUSIONS: QZACP inhibits HCC by reducing the expression of STAT3, VEGFA, JUN, FGF2, BCL2L1, AR, TERT, MMP7, MMP1, ABCB1, CA9, and ESR2. DAAME may be an important active component of QZACP for the prevention and treatment of HCC, inhibiting it by targeting the expression of CA9.

Prediction of methylation status using WGS data of plasma cfDNA for multi-cancer early detection (MCED).

BACKGROUND: Cell-free DNA (cfDNA) contains a large amount of molecular information that can be used for multi-cancer early detection (MCED), including changes in epigenetic status of cfDNA, such as cfDNA fragmentation profile. The fragmentation of cfDNA is non-random and may be related to cfDNA methylation. This study provides clinical evidence for the feasibility of inferring cfDNA methylation levels based on cfDNA fragmentation patterns. We performed whole-genome bisulfite sequencing and whole-genome sequencing (WGS) on both healthy individuals and cancer patients. Using the information of whole-genome methylation levels, we investigated cytosine-phosphate-guanine (CpG) cleavage profile and validated the method of predicting the methylation level of individual CpG sites using WGS data. RESULTS: We conducted CpG cleavage profile biomarker analysis on data from both healthy individuals and cancer patients. We obtained unique or shared potential biomarkers for each group and built models accordingly. The modeling results proved the feasibility to predict the methylation status of single CpG sites in cfDNA using cleavage profile model from WGS data. CONCLUSION: By combining cfDNA cleavage profile of CpG sites with machine learning algorithms, we have identified specific CpG cleavage profile as biomarkers to predict the methylation status of individual CpG sites. Therefore, methylation profile, a widely used epigenetic biomarker, can be obtained from a single WGS assay for MCED.

Feasibility analysis of arterial CT radiomics model to predict the risk of local and metastatic recurrence after radical cystectomy for bladder cancer.

PURPOSE: To construct a radiomics-clinical nomogram model for predicting the risk of local and metastatic recurrence within 3 years after radical cystectomy (RC) of bladder cancer (BCa) based on the radiomics features and important clinical risk factors for arterial computed tomography (CT) images and to evaluate its efficacy. METHODS: Preoperative CT datasets of 134 BCa patients (24 recurrent) who underwent RC were collected and divided into training (n = 93) and validation sets (n = 41). Radiomics features were extracted from a 1.5 mm CT layer thickness image in the arterial phase. A radiomics score (Rad-Score) model was constructed using the feature dimension reduction method and a logistic regression model. Combined with important clinical factors, including gender, age, tumor size, tumor number and grade, pathologic T stage, lymph node stage and histology type of the archived lesion, and CT image signs, a radiomics-clinical nomogram was developed, and its performance was evaluated in the training and validation sets. Decision curve analyses (DCA) the potential clinical usefulness. RESULTS: The radiomics model is finally linear combined by 8 features screened by LASSO regression, and after coefficient weighting, achieved good predictive results. The radiomics nomogram developed by combining two independent predictors, Rad-Score and pathologic T stage, was developed in the training set [AUC, 0.840; 95% confidence interval (CI) 0.743-0.937] and validation set (AUC, 0.883; 95% CI 0.777-0.989). The calibration curve showed good agreement between the predicted probability of the radiomics-clinical model and the actual recurrence rate within 3 years after RC for BCa. DCA show the clinical application value of the radiomics-clinical model. CONCLUSION: The radiomics-clinical nomogram model constructed based on the radiomics features of arterial CT images and important clinical risk factors is potentially feasible for predicting the risk of recurrence within 3 years after RC for BCa.

Qizhu Anti-Cancer Recipe promotes anoikis of hepatocellular carcinoma cells by activating the c-Jun N-terminal kinase pathway.

Background: Qizhu Anti-Cancer Recipe (QACR) is a traditional Chinese medicine widely used in treating several liver diseases. However, its function and the relevant mechanism underlying its effect in treating hepatocellular carcinoma (HCC) remain unknown. The aim of this study was to explore the effect of QACR in HCC, which are expected to be a potential therapeutic scheme for HCC. Materials and methods: The chemical compositions of QACR were determined by liquid chromatography/quadrupole time-of-fight mass spectrometry (LC-QTOF-MS). The anoikis-resistant HCC cell proliferation and angiopoiesis were detected using the cell counting kit 8 (CCK8) assay, trypan blue, calcein AM/EthD-1, flow cytometer, Western blot, and tube formation assays. An orthotopic xenograft mouse model was established to evaluate the in vivo effects of the QACR. The expression of proliferating cell nuclear antigen (PCNA), Bcl-2, CD31, caspase-3, caspase-8, caspase-9, PARP-1, DFF40, phospho-c-Jun NH2-terminal kinase (p-JNK), and JNK was assessed using Western blot and immunohistochemical analysis. Results: QACR reduced the growth and tube formation of anoikis-resistant HCC cells and enhanced cell apoptosis in vitro. In the orthotopic xenograft mouse models, QACR suppressed the tumorigenesis of HCC in vivo. Mechanistically, QACR modulated the JNK pathway. The JNK inhibitor (SP600125) reverses the inhibitory effects of QACR on anoikis-resistant HCC cell proliferation and angiopoiesis. Conclusion: Our study suggests that QACR suppresses the proliferation and angiopoiesis of anoikis-resistant HCC cells by activating the JNK pathway. Therefore, QACR is a promising new therapeutic strategy for treating hepatocellular carcinoma.

Holographic 3D renal segments reconstruction protects renal function by promote choice of selective renal artery clamping during robot-assisted partial nephrectomy.

OBJECTIVE: To investigate the impact of selective artery clamping (SAC) and main artery clamping (MAC) during robot-assisted partial nephrectomy (RAPN) on renal function and the influence of holographic three-dimensional (3D) reconstruction of renal segments on the selection between SAC and MAC. METHODS: This retrospective observational study included patients who underwent RAPN at First Hospital Affiliated to the Army Medical University between December 2016 and July 2022. According to the clamping methods, the patients were divided into the SAC group and the MAC group. The primary outcome was renal function. RESULTS: A total of 422 patients (194 in the SAC group and 228 in the MAC group) were included. The RAPN procedures were all completed successfully. The patients in SAC group had less glomerular filtration rate (GFR) decline in the affected kidney (8.6 ± 7.0 ml/min vs. 18.7 ± 10.9 ml/min, P < 0.001) and minor estimated glomerular filtration rate (eGFR) decrease (4.3 ± 10.5 ml/min vs. 12.6 ± 12.1 ml/min, P < 0.001) than those in MAC group. Among 37 patients with baseline renal insufficiency, the GFR decline of the affected kidney in the SAC subgroup was significantly lower than in the MAC subgroup (5.5 ± 6.5 ml/min vs. 14.3 ± 9.2 ml/min, P = 0.002). The proportion of patients who underwent 3D reconstruction was significant higher in the SAC group than in the MAC group. (65.46% vs. 28.07%, P < 0.001). CONCLUSION: The SAC technique during RAPN may serve as a protective measure for renal function, while the implementation of holographic 3D renal segment reconstruction technique may facilitate optimal selection of SAC.

FAT4 expression in peripheral blood mononuclear cells is associated with prognosis and immune cell infiltration in hepatocellular carcinoma.

Peripheral blood mononuclear cell (PBMC) genes reflect the host immune status and could be suitable for evaluating the prognosis of patients with hepatocellular carcinoma (HCC), for which a reliable biomarker is unavailable and the host immune responses to cancer cells. This study aimed to investigate prognostically relevant genes in HCC PBMCs and assessed whether their expression represents tumor immune infiltration. Gene expression in PBMCs from patients with advanced or terminal HCC who had survived or died was examined. Correlations among FAT atypical cadherin 4 (FAT4) expression, cancer immune characteristics, and infiltrated immune cell gene marker sets were analyzed. FAT4 expression was lower in the PBMCs of patients with advanced or terminal HCC who had died than that in patients who survived. Kaplan-Meier analysis indicated that FAT4 downregulation was associated with a relatively poor prognosis while overexpression was positively correlated with immune cell infiltration, several immune cell markers, and immune checkpoint expression. Hsa-miR-93-5p represented the most probable upstream microRNA of FAT4. Thus, upregulated FAT4 in PBMCs and HCC tissues might indicate a favorable prognosis and increased immune cell infiltration, while miRNA-93-5p could be a modulator of FAT4 expression. Collectively, these findings suggest novel immunotherapy targets for HCC.

Association of mitochondrial DNA copy number with chronic kidney disease in older adults.

BACKGROUND: Mitochondrial dysfunction in kidney cells has been implicated in the pathogenesis of chronic kidney disease (CKD). Estimation of mitochondrial DNA copy number (mtDNA-CN) is considered a convenient method for representing mitochondrial function in large samples. However, no study has investigated the association between mtDNA-CN and CKD in older adults with the highest prevalence. The objective is to examine cross-sectional and prospective associations between mtDNA-CN values and CKD risk in older adults to determine whether mtDNA-CN represents a novel potential biomarker for the recognition of CKD risk. PATIENTS AND METHODS: In a Chinese community-based cohort of over 65-year-olds, we included 14,467 participants (52.6% females). CKD was defined by eGFR < 60 mL/min/1.73 m2 or ICD-10 codes (patients = 3831 (26.5%)). Participants had peripheral blood levels of mtDNA-CN calculated from probe intensities of the Axiom CAS Array. RESULTS: The risk of CKD prevalence decreased with mtDNA-CN per 1-SD increment, independent of established risk factors for older CKD (odds ratio [OR] per SD 0.90, 95% confidence interval [CI] 0.86, 0.93, P < 0.001), and has comparable strength of association with these established risk factors. Furthermore, the progression of kidney function was stratified according to the worsening of eGFR categories. The risk of kidney function progression to a more severe stage gradually decreased as the mtDNA-CN increased (P trend < 0.001). Non-CKD participants in the highest quartile of mtDNA-CN had a lower risk of developing CKD compared to the lowest quartile within 2 years of follow-up, reducing the risk of CKD by 36% (95% CI 0.42, 0.97; P = 0.037). CONCLUSIONS: Based on the analysis of the largest sample to date investigating the association between mtDNA-CN and CKD in older adults, higher levels of mtDNA-CN were found to be associated with a lower risk of CKD, suggesting that a reduced level of mtDNA-CN is a potential risk factor for CKD.

State of CD8+ T cells in progression from nonalcoholic steatohepatitis to hepatocellular carcinoma: From pathogenesis to immunotherapy.

With the obesity epidemic, nonalcoholic steatohepatitis (NASH) is emerging as the fastest growing potential cause of hepatocellular carcinoma (HCC). NASH has been demonstrated to establish a tumor-prone liver microenvironment where both innate and adaptive immune systems are involved. As the most typical anti-tumor effector, the cell function of CD8+ T cells is remodeled by chronic inflammation, metabolic alteration, lipid toxicity and oxidative stress in the liver microenvironment along the NASH to HCC transition. Unexpectedly, NASH may blunt the effect of immune checkpoint inhibitor therapy against HCC due to the dysregulated CD8+ T cells. Growing evidence has supported that NASH is likely to facilitate the state transition of CD8+ T cells with changes in cell motility, effector function, metabolic reprogramming and gene transcription according to single-cell sequencing. However, the mechanistic insight of CD8+ T cell states in the NASH-driven HCC is not comprehensive. Herein, we focus on the characterization of state phenotypes of CD8+ T cells with both functional and metabolic signatures in NASH-driven fibrosis and HCC. The NASH-specific CD8+ T cells are speculated to mainly have a dualist effect, where its aberrant activated phenotype sustains chronic inflammation in NASH but subsequently triggers its exhaustion in HCC. As the exploration of CD8+ T cells on the distribution and phenotypic shifts will provide a new direction for the intervention strategies against HCC, we also discuss the implications for targeting different phenotypes of CD8+ T cells, shedding light on the personalized immunotherapy for NASH-driven HCC.

Gene mutation profiling and clinical significances in patients with renal cell carcinoma.

OBJECTIVES: The pathological mechanisms of patients with Renal Cell Carcinoma (RCC) remain defined. This study aimed to evaluate relationships between the landscape of gene mutations and their clinical significance in RCC patients. METHODS: Tissue and peripheral blood samples of 42 patients with RCC were collected and performed for the Next Generation Sequencing (NGS) with Geneseeq PrimeTM 425-gene panel probes. Their landscapes of gene mutation were analyzed. We also carried out an evaluation of Tumor-Node-Metastasis (TNM) staging, RENAL nephelometry score, surgery, and targeted drug treatment of patients. Then we compared the correlations of landscape in gene mutations and the prognosis. RESULTS: The most common gene alternations, including BAP1, PBRM1, SETD2, CSF1R, NPM1, EGFR, POLE, RB1, and VHL genes, were identified in tissue and blood samples of 75% of patients. EGFR, POLE, and RB1 gene mutations frequently occurred in relapsed and metastatic patients. BAP1, CCND2, KRAS, PTPN11, ERBB2/3, JAK2, and POLE were presented in the patients with > 9 RENAL nephelometry score. Univariable analysis indicated that SETD2, BAP1, and PBRM1 genes were key factors for Disease-Free Survival (DFS). Multivariable analysis confirmed that mutated SETD1, NPM1, and CSF1R were critical factors for the Progression Free Survival (PFS) of RCC patients with target therapy. CONCLUSIONS: Wild-type PBRM1 and mutated BAP1 in patients with RCC were strongly associated with the outcomes of the patient. The PFS of the patients with SETD2, NPM1, and CSF1R mutations were significantly shorter than those patients without variants.

Identification of immune-related target and prognostic biomarkers in PBMC of hepatocellular carcinoma.

BACKGROUND:  Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related deaths worldwide, and is characterized by insidious onset, rapid progression, and poor prognosis. Immunotherapy is a first-line treatment for advanced HCC. The identification of immune-related prognostic markers may be an effective strategy to predict and improve clinical response rate of immunotherapy. METHODS:  The DESeq2, edgeR, and limma R packages were used to compare the transcriptomes of HCC with different prognoses. Cancer-related databases such as UALCAN, TNMplot, GEPIA, muttarget and Human Protein Atlas (HPA), and the Kaplan-Meier Plotter platform were used to analyze the relationship between CLDN18 and the clinical characteristics, as well as prognosis of HCC. The co-expressed genes of CLDN18 were obtained from LinkedOmics platform, and GO functional enrichment and KEGG pathway analysis were performed. The CIBERSORT, TIMER, Timer 2.0 and TISIDB algorithms were used to analyze immune infiltration. RESULTS:  CLDN18 was differentially expressed in HCC patients with different prognoses, and its expression level in PBMC was positively correlated with the stage of BCLC. In addition, CLDN18 was significantly overexpressed in HCC tumor tissues compared to adjacent non-tumor tissues, which was consistent with PBMC sequencing results and immunohistochemical data from human protein profiles. CLDN18 was also positively correlated with HCC staging and grading, and high expression levels of CLDN18 predicted shorter overall survival. Functional annotation of CLDN18 in HCC revealed enrichment of the cellular senescence and protein activation cascade, along with biological processes such as cell cycle, inflammatory response, and cellular ketone metabolism. In addition, CLDN18 was also associated with tumor infiltrating immune cells, suppressive immune cell markers, T lymphocyte depletion and activation of HCC, and low expression of CLDN18 was associated with higher CD8 + T cell infiltration and better survival rates. CONCLUSIONS: CLDN18 is a potential prognostic marker and immunotherapeutic target for HCC.

Role of liensinine in sensitivity of activated macrophages to ferroptosis and in acute liver injury.

Acute liver injury (ALI) is an acute inflammatory liver disease with a high mortality rate. Alternatively, activated macrophages (AAMs) have been linked to the inflammation and recovery of ALI. However, the mechanism underlying AAM death in ALI has not been studied sufficiently. We used liensinine (Lie) as a drug of choice after screening a library of small-molecule monomers with 1488 compounds from traditional Chinese remedies. In ALI, we evaluated the potential therapeutic effects and underlying mechanisms of action of the drug in ALI and found that it effectively inhibited RSL3-induced ferroptosis in AAM. Lie significantly reduced lipid peroxidation in RSL3-generated AAM. It also improved the survival rate of LPS/D-GalN-treated mice, reduced serum transaminase activity, suppressed inflammatory factor production, and may have lowered AAM ferroptosis in ALI. Lie also inhibited ferritinophagy and blocked Fe2+ synthesis. Following combined treatment with RSL3 and Lie, super-resolution microscopy revealed a close correlation between ferritin and LC3-positive vesicles in the AAM. The co-localization of ferritin and LC3 with LAMP1 was significantly reduced. These findings suggest that Lie may ameliorate ALI by inhibiting ferritinophagy and enhancing AMM resistance to ferroptosis by inhibiting autophagosome-lysosome fusion. Therefore, Lie may be used as a potential therapeutic agent for patients with ALI.

High­throughput screening identification of a small­molecule compound that induces ferroptosis and attenuates the invasion and migration of hepatocellular carcinoma cells by targeting the STAT3/GPX4 axis.

Hepatocellular carcinoma (HCC) is a lethal malignancy. Although considerable efforts have been made in recent years regarding treatments, effective therapeutic drugs for HCC remain insufficient. In the present study, polyphyllin VI was identified as a potential therapeutic drug for HCC by screening natural herbal compounds. The therapeutic effects of polyphyllin VI were assessed using Cell Counting Kit­8, lactate dehydrogenase release and colony formation assays. The occurrence of ferroptosis was determined by assessing lipid peroxidation by reactive oxygen species, malondialdehyde levels, intracellular ferrous iron levels, and the mRNA and protein levels of glutathione peroxidase 4 (GPX4). The migratory and invasive abilities of HCC cells were examined using wound healing and Transwell assays. The results revealed that polyphyllin VI inhibited the proliferation, invasion and metastasis of HCC cells (HCCLM3 and Huh7 cells) by inducing ferroptosis. In addition, through a network pharmacology­based approach and molecular docking analyses, it was found that polyphyllin VI may target the signal transducer and activator of transcription 3 (STAT3). HCC cells were treated with polyphyllin VI or a STAT3 inhibitor (Stattic), both of which exerted similar inhibitory effects on protein expression. Furthermore, immunofluorescence staining revealed that polyphyllin VI significantly inhibited the nuclear translocation of p­STAT3 in HCC cells. Mechanistically, by the overexpression of STAT3, it was confirmed that STAT3 binds to GPX4 and promotes its protein expression and transcription, whereas polyphyllin VI induces ferroptosis by inhibiting the STAT3/GPX4 axis. Subsequently, in vivo experiments revealed that polyphyllin VI inhibited the growth of subcutaneously transplanted tumors. On the whole, findings of the present study suggest that polyphyllin VI inhibits STAT3 phosphorylation, which inhibits GPX4 expression and induces the ferroptosis of HCC cells, eventually inhibiting their invasion and metastasis. These data suggest that polyphyllin VI may be a candidate for the prevention and treatment of HCC.

Gene mutation profiling and clinical significances in patients with renal cell carcinoma

Abstract Objectives The pathological mechanisms of patients with Renal Cell Carcinoma (RCC) remain defined. This study aimed to evaluate relationships between the landscape of gene mutations and their clinical significance in RCC patients. Methods Tissue and peripheral blood samples of 42 patients with RCC were collected and performed for the Next Generation Sequencing (NGS) with Geneseeq PrimeTM 425-gene panel probes. Their landscapes of gene mutation were analyzed. We also carried out an evaluation of Tumor-Node-Metastasis (TNM) staging, RENAL nephelometry score, surgery, and targeted drug treatment of patients. Then we compared the correlations of landscape in gene mutations and the prognosis. Results The most common gene alternations, including BAP1, PBRM1, SETD2, CSF1R, NPM1, EGFR, POLE, RB1, and VHL genes, were identified in tissue and blood samples of 75% of patients. EGFR, POLE, and RB1 gene mutations frequently occurred in relapsed and metastatic patients. BAP1, CCND2, KRAS, PTPN11, ERBB2/3, JAK2, and POLE were presented in the patients with > 9 RENAL nephelometry score. Univariable analysis indicated that SETD2, BAP1, and PBRM1 genes were key factors for Disease-Free Survival (DFS). Multivariable analysis confirmed that mutated SETD1, NPM1, and CSF1R were critical factors for the Progression Free Survival (PFS) of RCC patients with target therapy. Conclusions Wild-type PBRM1 and mutated BAP1 in patients with RCC were strongly associated with the outcomes of the patient. The PFS of the patients with SETD2, NPM1, and CSF1R mutations were significantly shorter than those patients without variants.

Detection and monitoring of HBV-related hepatocellular carcinoma from plasma cfDNA fragmentation profiles.

Most hepatocellular carcinomas (HCCs) are associated with hepatitis B virus infection (HBV) in China. Early detection of HCC can significantly improve prognosis but is not yet fully clinically feasible. This study aims to develop methods for detecting HCC and studying the carcinogenesis of HBV using plasma cell-free DNA (cfDNA) whole-genome sequencing (WGS) data. Low coverage WGS was performed for 452 participants, including healthy individuals, hepatitis B patients, cirrhosis patients, and HCC patients. Then the sequencing data were processed using various machine learning models based on cfDNA fragmentation profiles for cancer detection. Our best model achieved a sensitivity of 87.10% and a specificity of 88.37%, and it showed an increased sensitivity with higher BCLC stages of HCC. Overall, this study proves the potential of a non-invasive assay based on cfDNA fragmentation profiles for the detection and prognosis of HCC and provides preliminary data on the carcinogenic mechanism of HBV.

Germacrone induces caspase-3/GSDME activation and enhances ROS production, causing HepG2 pyroptosis.

Liver cancer is a highly lethal malignancy. Despite considerable efforts made in recent years, the prognosis of patients with liver cancer remains poor. Curcuma zedoaria (known as Ezhu in Chinese) is widely prescribed in traditional Chinese medicine. Germacrone (GM) is a sesquiterpene constituent derived from the essential oil of Ezhu, and exerts anti-carcinogenic effects by inducing apoptosis in various cancer cells. The present study investigated the potential mechanism of GM in HepG2 cells. Cell Counting Kit-8, colony-formation and lactate dehydrogenase-release assays, as well as cell death assays using flow cytometry, were performed to evaluate HepG2 cell proliferation following GM treatment. HepG2 cells were transfected with caspase-3 small interfering RNA and then treated with GM. Caspase-3 expression levels were determined by reverse transcription-quantitative PCR and western blotting. The present study showed that GM inhibited the growth of HepG2 cells and induced the proteolytic cleavage of caspase 3, with concomitant cleavage of gasdermin E (GSDME), by markedly increasing the production of reactive oxygen species (ROS). This led to caspase 3-dependent cleavage of GSDME, thereby promoting pyroptosis in HepG2 cells. However, these changes were rescued by ROS scavengers, such as N-acetylcysteine. Furthermore, GM inhibited tumor growth by promoting the cleavage of caspase 3 and GSDME in HepG2 cell xenograft models. These results indicated that GM induced GSDME-dependent pyroptosis through caspase 3 activation, at least in part, by damaging the mitochondria and enhancing ROS production, thereby supporting the possible development of GM as a candidate for the prevention and treatment of liver cancer.