Cancer stem cells (CSCs) represent a distinct subpopulation of cells characterized by self-renewal capacity, differentiation potential, and critical roles in driving tumor progression, therapeutic resistance, recurrence, and maintenance of the tumor microenvironment. Targeting CSCs has emerged as a pivotal direction in cancer research, offering novel strategies to overcome drug resistance and prevent metastasis and relapse. Lysosomes, traditionally recognized as central organelles for intracellular degradation and recycling, are indispensable for cellular homeostasis. Dysregulation of lysosomal function is intimately linked to various diseases, including cancer. In tumors, aberrant lysosomal activity can promote malignant progression through mechanisms such as altering metabolic pathways, enhancing lysosomal exocytosis, modulating drug resistance, and interfering with autophagy-lysosomal pathways. Recent studies have underscored the involvement of lysosomes in regulating CSC properties. This review synthesizes findings on lysosomal regulation of CSCs through the following aspects. (1) Lysosomes exert complex and critical bidirectional control over CSC stemness maintenance through three degradation pathways that are dependent on their degradative function. (i) The lysophagy pathway. This pathway exhibits dual roles. Activation can sustain CSC functions; for instance, in glioblastoma, hypoxia upregulates Gal-8 via the STAT3/HIF1α signaling axis to induce autophagy, supporting stem cell survival. In head and neck squamous cell carcinoma, degradation of GSK3β activates the Wnt pathway, enhancing stemness. Conversely, this pathway can suppress stemness by degrading stemness-related proteins such as BMI-1 and OCT4A, thereby impairing CSC self-renewal capacity. (ii) Mitophagy pathway. In non-small cell lung cancer stem cells, mitophagy-related mechanisms, such as the accumulation of mitochondrial DNA (mtDNA) activating the TLR9-Notch1-AMPK signaling axis, have been shown to promote CSC proliferation. (iii) Autophagosome-dependent lysosomal degradation pathway. This pathway directly regulates stemness-related proteins in a bidirectional manner. Enhanced degradative function can promote CSC properties, exemplified by the degradation of NUMB to activate Notch signaling. Conversely, attenuated degradative function can also enhance stemness by stabilizing oncoproteins (e.g., protecting Frizzled-1 from degradation to sustain Wnt signaling) or preventing the degradation of tumor suppressors (e.g., inhibiting Notch degradation). (2) Constituent proteins of lysosomes, including membrane proteins and luminal acid hydrolases, participate in regulating CSC stemness. Regarding membrane proteins, LAMP2A facilitates chaperone-mediated autophagy to maintain stemness in glioblastoma and ovarian cancer. V-ATPase, by maintaining an acidic luminal environment, promotes proliferation and drug resistance in glioma stem cells. Among hydrolases, cathepsins B and L are highly expressed in pancreatic and ovarian cancers and correlate with poor prognosis. Furthermore, targeting lysosomes to induce lysosomal membrane permeabilization (LMP) triggers lysosome-mediated cell death, presenting a potential therapeutic strategy for eradicating CSCs.(3) The acidic luminal environment, single-membrane structure, and the presence of transmembrane transporters (e.g., ABCA3) enable lysosomes to passively trap or actively uptake and sequester chemotherapeutic drugs. Subsequent drug extrusion via exocytosis confers drug resistance. In CSCs, this lysosome-mediated drug sequestration, often cooperating with autophagy, establishes multimodal drug resistance. Therefore, targeting lysosomal function represents a potential strategy to overcome therapy resistance. The central role of lysosomes in regulating CSC stemness and resistance positions them as highly promising therapeutic targets. Strategies aimed at disrupting lysosomal function to selectively eliminate CSCs include: inhibiting the lysosome-autophagy system using agents like IITZ or lovastatin; inducing lysosomal membrane permeabilization (LMP) with compounds such as hexamethylene amiloride to compromise membrane stability; and disrupting the acidic luminal environment using drugs like siramesine or the K/H transport compound 2. In conclusion, lysosomes critically regulate CSC stemness maintenance and drug resistance through degradative pathways, membrane protein functions, luminal hydrolase activities, and drug sequestration mechanisms. This redefines the lysosome from a traditional “waste disposal unit” to a “signal integration center” in CSCs. The duality and context-dependency of lysosomal function in CSCs offer novel insights into the heterogeneity observed across different tumors. Targeting lysosomal vulnerabilities—such as inducing LMP, disrupting acidity, or blocking autophagic flux—provides a strategy to bypass canonical CSC resistance mechanisms and directly trigger cell death. This establishes the lysosome as a key target to overcome CSC-mediated therapy resistance, paving the way for developing diverse candidate drugs and innovative combination therapies in oncology.

This paper introduces a real-world cohort research model for community-acquired pneumonia （CAP） based on the Jiangsu Traditional Chinese Medicine （TCM） Dominant Diseases Diagnosis and Treatment Data Platform. Firstly， data cleaning is performed by standardizing diagnosis， symptoms， treatment and imaging， intelligently extracting unstructured information， and cleaning and constructing a standardized database. Secondly， for cohort establishment， CAP patients across the province are screened in accordance with CAP diagnostic criteria to build a high-quality disease-specific cohort. Lastly， in terms of protocol design， the characteristics of TCM research and the CAP disease profile are considered to determine appropriate inclusion and exclusion criteria， estimate sample size， define interventions， outcomes and economic evaluations， providing a reference for real-world TCM research on CAP.

This paper introduces a real-world cohort research model for community-acquired pneumonia （CAP） based on the Jiangsu Traditional Chinese Medicine （TCM） Dominant Diseases Diagnosis and Treatment Data Platform. Firstly， data cleaning is performed by standardizing diagnosis， symptoms， treatment and imaging， intelligently extracting unstructured information， and cleaning and constructing a standardized database. Secondly， for cohort establishment， CAP patients across the province are screened in accordance with CAP diagnostic criteria to build a high-quality disease-specific cohort. Lastly， in terms of protocol design， the characteristics of TCM research and the CAP disease profile are considered to determine appropriate inclusion and exclusion criteria， estimate sample size， define interventions， outcomes and economic evaluations， providing a reference for real-world TCM research on CAP.

Objective To explore the effect of management model of clinical pharmacists participating in multidisciplinary collaborative diagnosis and treatment(MDT)for new anti-tumor drugs in the national medical insurance drug negotiation(hereinafter referred to as"national negotiation"),including efficacy,safe-ty,economy and rationality.Methods The medical records of 326 cases using novel anti-tumor drugs by na-tional negotiation and conforming to the including and excluding standards in this hospital from July 2018 to June 2023 were retrospectively analyzed.The patients were divided into the MDT group(n=122)and non-MDT group(n=204).The patients diagnosed as non-small cell lung cancer(NSCLC)in the two groups were extracted and defined as the MDT-NSCLC subgroup(n=41)and non-MDT-NSCLC subgroup(n=77).The progression-free survival(PFS),overall survival(OS),disease control rate(DCR)and the indexes such as survival quality and medical quality control were compared between the groups.Results The median PFS in the two groups was 12.7 months and 8.0 months,the median OS was 75.2 months and 56.3 months,DCR was 96.72％and 81.86％respectively,and the differences were statistically significant(P＜0.05).The COX multivariate regression analysis indicated that the HR value of clinical pharmacists participating in MDT was higher than the other influencing factors.The median PFS time in the MDT-NSCLC subgroup and non-MDT-NSCLC subgroup was 10.5 months and 6.7 months,DCR was 97.30％and 75.64％respectively,and the differences were statistically significant(P＜0.05),the median OS time was 55.1 months and 40.3 months respectively,and the difference was statistically significant(P＞0.05).The COX multivariate regression anal-ysis indicated that the HR value with clinical pharmacists participating in MDT was higher than the other in-fluencing factors;The adverse reaction occurrence rate in the MDT group and non-MDT group was 45.9％and 58.3％respectively,and the difference was statistically significant(P＜0.05).The KPS score after treatment in the MDT group was higher than that in the non-MDT group,and the difference was statistically significant;in the aspect of medical quality control,the average drug proportion in the MDT group and non-MDT group was 63.93％and 64.54％respectively,the rational drug rate of comments on prescription was 98.36％and 88.73％respectively,the patient satisfaction average value was 90.69 points and 87.36 points respectively and the differences were statistically significant(P＜0.05).Conclusion Clinical pharmacists participating in MDT related to novel anti-tumor drugs by national negotiation is beneficial to improve the therapeutic effects,living quality and patient satisfaction,also benefit to management and control of off-label drug use and medical quality control indexes.

Objective:To investigate the differences in clinical outcomes of septic children with varying serum zinc levels,and to analyze the relationship between reduced serum zinc levels and organ dysfunction as well as 28-day mortality in septic children.Methods:This study conducted a retrospective analysis of clinical data from pediatric patients diagnosed with sepsis or septic shock in the Department of critical care medicine of the children's Hospital of Soochow University between January 2017 and December 2022.Clinical characteristics,organ dysfunction,and prognosis were compared between two groups:children with low serum zinc levels and those with normal zinc levels.Results:The serum zinc level of septic children within 24 hours of admission was 9.60(5.52,13.80)μmol/L,with 50.54%(94/186)of the children exhibiting low serum zinc levels(<10.07 μmol/L).Compared to the normal serum zinc group,the low serum zinc group had a significantly lower Pediatric Critical Illness Score(PCIS)[(78.71±9.35)vs.(85.12±8.51),P=0.005]and higher 28-day mortality(46.80%vs.14.13%,P<0.001).The low serum zinc group also had a higher proportion of invasive mechanical ventilation(64.89%vs.47.82%,P=0.019),renal replacement therapy(15.59%vs.3.26%,P=0.003),and use of vasoactive drugs(56.38%vs.30.43%,P<0.001).The rate of underlying conditions in the low serum zinc group was significantly higher than that in the normal serum zinc group(57.44%vs.36.95%,P=0.005).Additionally,the low serum zinc group had a higher incidence of disseminated intravascular coagulation(DIC),respiratory failure,acute kidney injury,shock,and multiple organ dysfunction syndrome(MODS)compared to the normal serum zinc group(P<0.05).Serum zinc levels had predictive value for 28-day mortality in septic children(AUC=0.813;95%CI:0.725～0.902;P<0.001).A serum zinc level of less than 6.950 μmol/L predicted the death of septic children with a sensitivity of 0.618 and a specificity of 0.902.Conclusion:Sepsis in children is commonly associated with low serum zinc levels,especially in those with underlying conditions such as hematologic and oncologic disorders.Sepsis patients hypozincemia with a higher incidence of DIC,respiratory failure,acute kidney injury,shock,and MODS.A serum zinc level below 6.95 μmol/L serves as a significant predictor of 28-day mortality in children with severe sepsis.

Objective To explore the role of X chromosome encoded epigenetic regulator UTX in NK cell-mediated anti-tumor activity.Methods Male Ncr1-iCre mice were crossed with female UTXfl/fl mice to generate F1 Ncr1-iCre+UTXfl/-male mice,which were further crossed with female UTXfl/fl mice to obtain male Ncr1-iCre-UTX fl/-control mice(M-Con)and NK-specific deletion of UTX male mice Ncr1-iCre+UTXfl/-(M-KO),as well as female Ncr1-iCre-UTXfl/fl control mice(F-Con)and UTX-deficient female mice Ncr1-iCre+UTXfl/fl(F-KO).UTX-deficient mice were injected with melanoma cell line B16F10 via tail vein to observe pulmonary metastatic tumor nodules.Moreover,flow cytometry was applied to detect the proportion and quantity of pulmonary NK cells(CD3-CD19-NK1.1+),maturation makers KLRG1 and CD11b,activation receptors NKG2D and CD69,and effector molecules,including perforin,granzyme B,CD107a,and IFN-γ.Then pulmonary NK cells were sorted and co-cultured with B16F10 cells,and the apoptosis of the melanoma cells was measured with flow cytometry.Results Compared with the M-Con mice,the M-KO mice presented less number of pulmonary tumor nodules(P<0.05),increased proportion and quantity of NK cells in the tumor microenvironment(P<0.01),though no obvious changes in the ratio of NK maturation makers KLRG1 to CD11b,enhanced expression level of cytotoxic molecule perforin(P<0.01),but no changes in the expression of effector molecule granzyme B,degranulation marker CD107a and cytokine IFN-γ in NK cells.Co-culture of NK cells and B16F10 cells promoted the apoptosis of tumor cells(P<0.05).Compared with the F-Con mice,the F-KO mice had no statistical difference in the number of pulmonary tumor nodules,but larger proportion and number of NK cells(P<0.05),decreased ratio of KLRG1 to CD11b(P<0.01),elevated level of perforin but decreased levels of granzyme B,CD107a and IFN-γ in NK cells(P<0.01).The co-culture of NK cells and B16F10 cells reduced the apoptosis of tumor cells in F-KO female mice(P<0.05).Conclusion NK-specific deletion of UTX regulates pulmonary metastasis of melanoma in a sex-dependent manner.

High mobility group box 1 (HMGB1), when released extracellularly, plays a pivotal role in the development of spinal cord synapses and exacerbates autoimmune diseases within the central nervous system. In experimental autoimmune encephalomyelitis (EAE), a condition that models multiple sclerosis, the levels of extracellular HMGB1 and interleukin-33 (IL-33) have been found to be inversely correlated. However, the mechanism by which IL-33 deficiency enhances HMGB1 release during EAE remains elusive. Our study elucidates a potential signaling pathway whereby the absence of IL-33 leads to increased binding of P300/CBP-associated factor with HMGB1 in the nuclei of astrocytes, upregulating HMGB1 acetylation and promoting its release from astrocyte nuclei in the spinal cord of EAE mice. Conversely, the addition of IL-33 counteracts the TNF-α-induced increase in HMGB1 and acetylated HMGB1 levels in primary astrocytes. These findings underscore the potential of IL-33-associated signaling pathways as a therapeutic target for EAE treatment.
Animals ; Encephalomyelitis, Autoimmune, Experimental/metabolism* ; Astrocytes/metabolism* ; Interleukin-33/metabolism* ; HMGB1 Protein/metabolism* ; Acetylation ; Mice, Knockout ; Mice, Inbred C57BL ; p300-CBP Transcription Factors/metabolism* ; Mice ; Spinal Cord/metabolism* ; Cells, Cultured ; Female ; Signal Transduction

The ambiguity of etiology makes temporomandibular joint osteoarthritis (TMJOA) "difficult-to-treat". Emerging evidence underscores the therapeutic promise of exosomes in osteoarthritis management. Nonetheless, challenges such as low yields and insignificant efficacy of current exosome therapies necessitate significant advances. Addressing lower strontium (Sr) levels in arthritic synovial microenvironment, we studied the effect of Sr element on exosomes and miRNA selectively loading in synovial mesenchymal stem cells (SMSCs). Here, we developed an optimized system that boosts the yield of SMSC-derived exosomes (SMSC-EXOs) and improves their miRNA profiles with an elevated proportion of beneficial miRNAs, while reducing harmful ones by pretreating SMSCs with Sr. Compared to untreated SMSC-EXOs, Sr-pretreated SMSC-derived exosomes (Sr-SMSC-EXOs) demonstrated superior therapeutic efficacy by mitigating chondrocyte ferroptosis and reducing osteoclast-mediated joint pain in TMJOA. Our results illustrate Alix's crucial role in Sr-triggered miRNA loading, identifying miR-143-3p as a key anti-TMJOA exosomal component. Interestingly, this system is specifically oriented towards synovium-derived stem cells. The insight into trace element-driven, site-specific miRNA selectively loading in SMSC-EXOs proposes a promising therapeutic enhancement strategy for TMJOA.
MicroRNAs/metabolism* ; Mesenchymal Stem Cells/drug effects* ; Osteoarthritis/drug therapy* ; Exosomes/drug effects* ; Strontium/pharmacology* ; Synovial Membrane/cytology* ; Humans ; Animals ; Temporomandibular Joint Disorders/therapy* ; Temporomandibular Joint

Gene regulation relies on the precise binding of transcription factors (TFs) at regulatory elements, but simultaneously detecting hundreds of TFs on chromatin is challenging. We developed cFOOT-seq, a cytosine deaminase-based TF footprinting assay, for high-resolution, quantitative genome-wide assessment of TF binding in both open and closed chromatin regions, even with small cell numbers. By utilizing the dsDNA deaminase SsdAtox, cFOOT-seq converts accessible cytosines to uracil while preserving genomic integrity, making it compatible with techniques like ATAC-seq for sensitive and cost-effective detection of TF occupancy at the single-molecule and single-cell level. Our approach enables the delineation of TF footprints, quantification of occupancy, and examination of chromatin influences on TF binding. Notably, cFOOT-seq, combined with FootTrack analysis, enables de novo prediction of TF binding sites and tracking of TF occupancy dynamics. We demonstrate its application in capturing cell type-specific TFs, analyzing TF dynamics during reprogramming, and revealing TF dependencies on chromatin remodelers. Overall, cFOOT-seq represents a robust approach for investigating the genome-wide dynamics of TF occupancy and elucidating the cis-regulatory architecture underlying gene regulation.
Transcription Factors/genetics* ; Humans ; Chromatin/genetics* ; Animals ; Binding Sites ; Mice ; DNA Footprinting/methods*

Objective:This study empirically analyzes the relationship between outpatient and inpatient services under the impact of healthcare payment reform,and evaluates the effects of the reform.Methods:Data from healthcare services and basic medical insurance payments in eight districts of Jinhua City from 2020 to 2022 were used.A fixed-effects model for outpatient and inpatient services was constructed to analyze the impact of healthcare payment reforms and outpatient services on inpatient services.Results:The DRG-based payment had a significant positive effect on inpatient visits and a significant negative effect on employee basic medical insurance inpatient costs.The"capitation+APG"outpatient payment policy had a significant negative effect on inpatient visits and a significant negative effect on residents'basic medical insurance inpatient costs.The interaction between outpatient payment and outpatient visits had a significant negative effect on employee basic medical insurance inpatient visits,while the interaction between outpatient payment and outpatient costs had a significant negative effect on both overall and employee inpatient costs.Conclusions:The DRG payment reform led to an increase in inpatient visits and a reduction in employee basic medical insurance inpatient costs.The outpatient"capitation+APG"payment reform reduced inpatient visits and lowered residents'basic medical insurance inpatient costs,thereby slowing down the complementary effect between outpatient and inpatient services.