In-Depth Analysis of Molecular Network Based on Liquid Chromatography Coupled with Tandem Mass Spectrometry in Natural Products: Importance of Redundant Nodes Discovery.

The identification of molecules within complex mixtures is a major bottleneck in natural products (NPs) research. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) has emerged as the main tool for the high-throughput characterization of NPs. The large amount of data sets by LC-MS/MS presents a challenge for data processing and interpretation, and the LC-MS/MS molecular network (MN) is one of the most prominent tools for analyzing large MS/MS data sets, widely used for rapid classification, identification, and structural speculation of unknown compounds. However, the existence of a large number of redundant nodes leads to false-positive results. To solve this problem, we proposed the in-depth analysis of MN. In this study, in-depth analysis of MN of five NPs representing the common structures of saponin, steroid, flavonoid, alkaloid, and phenolic acid revealed the presence of redundant nodes (including other adducts, isotope, and in-source fragmentation) in addition to the normal nodes, which can lead to false-positive identification results. Additionally, the reasons for different redundant nodes are discussed and experimentally verified, and it was found that the impact of redundant nodes can be mitigated by optimizing the experimental conditions and employing Feature-Based Molecular Networking. Furthermore, Ion Identity Molecular Networking can rapidly discover and screen redundant nodes, simplifying the in-depth analysis of MN and improving the network connectivity of structurally related molecules. Finally, a combination formulation of 7 NPs is used as an example to provide a guide for in-depth analysis of MN for comprehensive characterization of complex systems. This study highlights the importance of an in-depth analysis of MN for better understanding and utilization of MS/MS data in complex systems to reduce the false-positive rate of identification by screening and filtering redundant nodes.

Zdhhc1- and Zdhhc2-mediated Gpm6a palmitoylation is essential for maintenance of mammary stem cell activity.

Adult mammary stem cells (aMaSCs) are vital to tissue expansion and remodeling during the process of postnatal mammary development. The protein C receptor (Procr) is one of the well-identified surface markers of multipotent aMaSCs. However, an understanding of the regulatory mechanisms governing Procr's protein stability remains incomplete. In this study, we identified Glycoprotein m6a (Gpm6a) as a critical protein for aMaSC activity modulation by using the Gpm6a knockout mouse model. Interestingly, we determined that Gpm6a depletion results in a reduction of Procr protein stability. Mechanistically, Gpm6a regulates Procr protein stability by mediating the formation of lipid rafts, a process requiring Zdhhc1 and Zdhhc2 to palmitate Gpm6a at Cys17,18 and Cys246 sites. Our findings highlight an important mechanism involving Zdhhc1- and Zdhhc2-mediated Gpm6a palmitoylation for the regulation of Procr stability, aMaSC activity, and postnatal mammary development.

Programmable Lanthanide Metal-Organic Framework for Ultra-Efficient Nucleic Acids Extraction and Interaction Analysis.

Efficient, mild, and reversible adsorption of nucleic acids onto nanomaterials represents a promising analytical approach for medical diagnosis. However, there is a scarcity of efficient and reversible nucleic acid adsorption nanomaterials. Additionally, the lack of comprehension of the molecular mechanisms governing their interactions poses significant challenges. These issues hinder the rational design and analytical applications of the nanomaterials. Herein, we propose an ultra-efficient nucleic acid affinity nanomaterial based on programmable lanthanide metal-organic frameworks (Ln-MOFs). Through experiments and density functional theory calculations, a rational design guideline for nucleic acid affinity of Ln-MOF was proposed, and a modular and flexible preparation scheme was provided. Then, Er-TPA (terephthalic acid) MOF emerged as the optimal candidate due to its pore size-independent adsorption and desorption capabilities for nucleic acids, enabling ultra-efficient adsorption (about 150% mass ratio) within 1 min. Furthermore, we elucidate the molecular-level mechanisms underlying the Ln-MOF adsorption of single- and double-stranded DNA and G4 structures. The affinity nanomaterial based on Ln-MOF exhibits robust nucleic acid extraction capability (4-fold higher than commercial reagent kits) and enables mild and reversible CRISPR/Cas9 functional regulation. This method holds significant promise for broad application in DNA/RNA liquid biopsy and gene editing, facilitating breakthroughs in analytical chemistry, pharmacy, and medical research.

Shexiang Baoxin Pill enriches Lactobacillus to regulate purine metabolism in patients with stable coronary artery disease.

BACKGROUND: It has been clinically confirmed that the Shexiang Baoxin Pill (SBP) dramatically reduces the frequency of angina in patients with stable coronary artery disease (SCAD). However, potential therapeutic mechanism of SBP has not been fully explored. PURPOSE: The study explored the therapeutic mechanism of SBP in the treatment of SCAD patients. METHODS: We examined the serum metabolic profiles of patients with SCAD following SBP treatment. A rat model of acute myocardial infarction (AMI) was established, and the potential therapeutic mechanism of SBP was explored using metabolomics, transcriptomics, and 16S rRNA sequencing. RESULTS: SBP decreased inosine production and improved purine metabolic disorders in patients with SCAD and in animal models of AMI. Inosine was implicated as a potential biomarker for SBP efficacy. Furthermore, SBP inhibited the expression of genes involved in purine metabolism, which are closely associated with thrombosis, inflammation, and platelet function. The regulation of purine metabolism by SBP was associated with the enrichment of Lactobacillus. Finally, the effects of SBP on inosine production and vascular function could be transmitted through the transplantation of fecal microbiota. CONCLUSION: Our study reveals a novel mechanism by which SBP regulates purine metabolism by enriching Lactobacillus to exert cardioprotective effects in patients with SCAD. The data also provide previously undocumented evidence indicating that inosine is a potential biomarker for evaluating the efficacy of SBP in the treatment of SCAD.

Efficacy and safety of Qushi Huayu, a traditional Chinese medicine, in patients with nonalcoholic fatty liver disease in a randomized controlled trial.

BACKGROUND: The effective treatment of non-alcoholic fatty liver disease (NAFLD) is an unmet medical need. Qushi Huayu (QSHY) is an empirical herbal formula with promising effects in NAFLD rodent models and a connection to gut microbiota regulation. HYPOTHESIS/PURPOSE: This study aimed to evaluate the effects of QSHY in patients with NAFLD through a multicenter, randomized, double-blind, double-dummy clinical trial. STUDY DESIGN: A total of 246 eligible patients with NAFLD and liver dysfunction were evenly divided to receive either QSHY and Dangfei Liganning capsule (DFLG) simulant or QSHY simulant and DFLG (an approved proprietary Chinese medicine for NAFLD in China) for 24 weeks. The primary outcomes were changes in liver fat content, assessed using vibration-controlled transient elastography, and serum alanine aminotransferase (ALT) levels from baseline to Week 24. RESULTS: Both QSHY and DFLG led to reductions in liver fat content and liver enzyme levels post-intervention (p < 0.05). Compared to DFLG, QSHY treatment improved ALT (ß, -0.128 [95 % CI, -0.25, -0.005], p = 0.041), aspartate transaminase (ß, -0.134 [95 % CI, -0.256 to -0.012], p = 0.032), and fibrosis-4 score (ß, -0.129 [95 % CI, -0.254 to -0.003], p = 0.044) levels. QSHY markedly improved gut dysbiosis compared to DFLG, with changes in Escherichia-Shigella and Bacteroides abundance linked to its therapeutic effect on reducing ALT. Patients with a high ALT response after QSHY treatment showed superior reductions in peripheral levels of phenylalanine and tyrosine, along with an elevation in the related microbial metabolite p-Hydroxyphenylacetic acid. CONCLUSION: Our results demonstrate favorable clinical potential for QSHY in the treatment of NAFLD.

Qijiao Shengbai Capsule alleviated leukopenia by interfering leukotriene pathway: Integrated network study of multi-omics.

BACKGROUND: Leukopenia could be induced by chemotherapy, which leads to bone marrow suppression and even affects the therapeutic progression of cancer. Qijiao Shengbai Capsule (QSC) has been used for the treatment of leukopenia in clinic, but its bioactive components and mechanisms have not yet been elucidated clearly. PURPOSE: This study aimed to elucidate the molecular mechanisms of QSC in treating leukopenia. STUDY DESIGN: Serum pharmacochemistry, multi-omics, network pharmacology, and validation experiment were combined to study the effect of QSC in murine leukopenia model. METHODS: First, UPLC-QTOF-MS was used to clarify the absorbed components of QSC. Then, cyclophosphamide (CTX) was used to induce mice model with leukopenia, and the therapeutic efficacy of QSC was assessed by an integrative approach of multi-omics and network pharmacology strategy. Finally, molecular mechanisms and potential therapeutic targets were identified by validated experiments. RESULTS: 121 compounds absorbed in vivo were identified. QSC significantly increase the count of white blood cells (WBCs) in peripheral blood of leukopenia mice with 15 days treatment. Multi-omics and network pharmacology revealed that leukotriene pathway and MAPK signaling pathway played crucial roles during the treatment of leukopenia with QSC. Six targets (ALOX5, LTB4R, CYSLTR1, FOS, JUN, IL-1ß) and 13 prototype compounds were supposed to be the key targets and potential active components, respectively. The validation experiment further confirmed that QSC could effectively inhibit the inflammatory response induced by leukopenia. The inhibitors of ALOX5 activity can significantly increase the number of WBCs in leukopenia mice. Molecular docking of ALOX5 suggested that calycosin, daidzein, and medicarpin were the potentially active compounds of QSC. CONCLUSION: Leukotriene pathway was found for the first time to be a key role in the development of leukopenia, and ALOX5 was conformed as the potential target. QSC may inhibit the inflammatory response and interfere the leukotriene pathway, it is able to improve hematopoiesis and achieve therapeutic effects in the mice with leukopenia.

The pathogenesis and potential therapeutic targets in sepsis.

Sepsis is defined as "a life-threatening organ dysfunction caused by dysregulated host systemic inflammatory and immune response to infection." At present, sepsis continues to pose a grave healthcare concern worldwide. Despite the use of supportive measures in treating traditional sepsis, such as intravenous fluids, vasoactive substances, and oxygen plus antibiotics to eradicate harmful pathogens, there is an ongoing increase in both the morbidity and mortality associated with sepsis during clinical interventions. Therefore, it is urgent to design specific pharmacologic agents for the treatment of sepsis and convert them into a novel targeted treatment strategy. Herein, we provide an overview of the molecular mechanisms that may be involved in sepsis, such as the inflammatory response, immune dysfunction, complement deactivation, mitochondrial damage, and endoplasmic reticulum stress. Additionally, we highlight important targets involved in sepsis-related regulatory mechanisms, including GSDMD, HMGB1, STING, and SQSTM1, among others. We summarize the latest advancements in potential therapeutic drugs that specifically target these signaling pathways and paramount targets, covering both preclinical studies and clinical trials. In addition, this review provides a detailed description of the crosstalk and function between signaling pathways and vital targets, which provides more opportunities for the clinical development of new treatments for sepsis.

Dracorhodin targeting CMPK2 attenuates inflammation: A novel approach to sepsis therapy.

BACKGROUND: Despite all modern advances in medicine, an effective drug for treating sepsis has yet to be found. The discovery of CMPK2 spurred hopes for the treatment of sepsis. However, CMPK2-untapped target inhibitors are still an enormous obstacle that has hindered the CMPK2-centric treatment of sepsis. METHODS: Here, we found that the CMPK2 gene is highly expressed in the whole blood of sepsis patients by RNA-Seq. First, recombinant CMPK2 was purified by a eukaryotic expression purification system, and the activity of recombinant CMPK2 was detected by the ADP-GLO assay. Second, we developed an affinity MS strategy combined with quantitative lysine reactivity profiling to discover CMPK2 ligands from the active ingredients of Chinese herbs. In addition, the dissociation constant Kd of the ligand and the target protein CMPK2 was further detected by microscale thermophoresis technology. Third, we used this strategy to identify a naturally sourced small molecule, dracorhodin (DP). Using mass spectrometry-based quantitative lysine reactivity profiling combined with a series of mutant tests, the results show that K265 acts as a bright hotspot of DP inhibition of CMPK2. Fourth, immune-histochemical staining, ELISAs, RT-qPCR, flow cytometry and immunoblotting were used to illustrate the potential function and related mechanism of DP in regulating sepsis injury. RESULTS: Our results suggest that DP exerts powerful anti-inflammatory effects by regulating the NLRP3 inflammasome via the lipopolysaccharide (LPS)-induced CMPK2 pathway. Strikingly, DP significantly attenuated LPS-induced sepsis in a mouse model, but its effect was weakened in mice with myeloid-specific Cmpk2 ablation. CONCLUSION: We provide a new framework that provides more valuable information for new therapeutic approaches to sepsis, including the establishment of screening strategies and the development of target drugs to provide a theoretical basis for ultimately improving clinical outcomes for sepsis patients. Collectively, these findings reveal that DP is a promising CMPK2 inhibitor for the treatment of sepsis.

A mitochondrial EglN1-AMPKα axis drives breast cancer progression by enhancing metabolic adaptation to hypoxic stress.

Mitochondria play essential roles in cancer cell adaptation to hypoxia, but the underlying mechanisms remain elusive. Through mitochondrial proteomic profiling, we here find that the prolyl hydroxylase EglN1 (PHD2) accumulates on mitochondria under hypoxia. EglN1 substrate-binding region in the ß2ß3 loop is responsible for its mitochondrial translocation and contributes to breast tumor growth. Furthermore, we identify AMP-activated protein kinase alpha (AMPKα) as an EglN1 substrate on mitochondria. The EglN1-AMPKα interaction is essential for their mutual mitochondrial translocation. After EglN1 prolyl-hydroxylates AMPKα under normoxia, they rapidly dissociate following prolyl-hydroxylation, leading to their immediate release from mitochondria. In contrast, hypoxia results in constant EglN1-AMPKα interaction and their accumulation on mitochondria, leading to the formation of a Ca2+ /calmodulin-dependent protein kinase 2 (CaMKK2)-EglN1-AMPKα complex to activate AMPKα phosphorylation, ensuring metabolic homeostasis and breast tumor growth. Our findings identify EglN1 as an oxygen-sensitive metabolic checkpoint signaling hypoxic stress to mitochondria through its ß2ß3 loop region, suggesting a potential therapeutic target for breast cancer.

Exploration of epithelial-mesenchymal transition-related lncRNA signature and drug sensitivity in breast cancer.

Background: Breast cancer (BRCA) has become the most diagnosed cancer worldwide for female and seriously endanger female health. The epithelial-mesenchymal transition (EMT) process is associated with metastasis and drug resistance in BRCA patients. However, the prognostic value of EMT-related lncRNA in BRCA still needs to be revealed. The aim of this study is to construct an EMT-related lncRNA (ERL) signature with accuracy predictive ability for the prognosis of BRCA patients. Methods: RNA-seq expression data and Clinical characteristics obtained from the TCGA (The Cancer Genome Atlas) were used in the study. First, we identified the EMT-related lncRNA by the Pearson correlation analysis. An EMT-related lncRNAs prognostic risk signature was constructed using univariate Cox regression and Lasso-penalized Cox regression analyses. The model's performance was validated using Kaplan-Meier (KM) survival analysis, ROC curve and C-index. Finally, a nomogram was constructed for clinical practice in evaluating the patients with BRCA and validated by calibration curve and decision curve analysis (DCA). We also evaluated the drug sensitivity of signature lncRNA and the tumor immune cell infiltration in breast cancer. Results: We constructed a 10-lncRNA risk score signature based on the lncRNAs associated with the EMT process. We could assign BRCA patients to the high- and low-risk group according to the median risk score. The prognostic risk signature showed excellent accuracy and demonstrated sufficient independence from other clinical characteristics. The immune cell infiltration analysis showed that the prognostic risk signature was related to the infiltration of the immune cell subtype. Drug sensitivity analysis proved ERLs signature could effectively predict the sensitivity of patients to common chemotherapy drugs in BRCA and provide guidance for chemotherapy drugs for high-risk and low-risk patients. Conclusion: Our ERL signature and nomogram have excellent prognostic value and could become reliable tools for clinical guidance.

Patterns of lymph node metastasis in level IIB and contralateral level VI for papillary thyroid carcinoma with pN1b and safety of low collar extended incision for neck dissection in level II.

OBJECTIVE: To explore relevant clinical factors of level IIB and contralateral level VI lymph node metastasis and evaluate the safety of low-collar extended incision (LCEI) for lymph node dissection in level II for papillary thyroid carcinoma (PTC) with pN1b. METHOD: A retrospective analysis was performed on 218 patients with PTC with pN1b who were treated surgically in the Head and Neck Surgery Center of Sichuan Cancer Hospital from September 2021 to May 2022. Data on age, sex, body mass index (BMI), tumor location, maximum tumor diameter, multifocality, Braf gene, T staging, surgical incision style, and lymph node metastasis in each cervical subregion were collected. The chi-square test was used for comparative analysis of relevant factors. All statistical analyses were completed by SPSS 24 software. RESULT: Each subgroup on sex, age, BMI, multifocality, tumor location, extrathyroidal extension, Braf gene, and lymphatic metastasis in level III, level IV, and level V had no significant difference in the positive rate of lymph node metastasis in level IIB (P > 0.05). In contrast, patients with bilateral lateral cervical lymphatic metastasis were more likely to have level IIB lymphatic metastasis than those with unilateral lateral cervical lymphatic metastasis, with a statistically significant difference (P = 0.000). In addition, lymph node metastasis in level IIA was significantly associated with lymph node metastasis in level IIB (P = 0.001). After multivariate analysis, lymph node metastasis in level IIA was independently associated with lymph node metastasis in level IIB (P = 0.010). The LCEI group had a similar lymphatic metastasis number and lymphatic metastasis rate in both level IIA and level IIB as the L-shaped incision group (P > 0.05). There were 86 patients with ipsilateral central lymphatic metastasis (78.2%). Patients with contralateral central lymphatic metastasis accounted for 56.4%. The contralateral central lymphatic metastasis rate was not correlated with age, BMI, multifocality, tumor invasion, or ipsilateral central lymphatic metastasis, and there was no significant difference (P > 0.05). The contralateral central lymphatic metastasis in males was slightly higher than that in females, and the difference was statistically significant (68.2% vs. 48.5%, P = 0.041). CONCLUSION: Lymphatic metastasis in level IIA was an independent predictor of lymphatic metastasis in level IIB. When bilateral lateral cervical lymphatic metastasis or lymph node metastasis of level IIA is found, lymph node dissection in level IIB is strongly recommended. When unilateral lateral cervical lymphatic metastasis and lymphatic metastasis in level IIA are negative, lymph node dissection in level IIB may be performed as appropriate on the premise of no damage to the accessory nerve. LCEI is safe and effective for lymph node dissection in level II. When the tumor is located in the unilateral lobe, attention should be given to contralateral central lymph node dissection because of the high lymphatic metastasis rate.

Improving the Data Quality of Untargeted Metabolomics through a Targeted Data-Dependent Acquisition Based on an Inclusion List of Differential and Preidentified Ions.

Metabolomics based on high-resolution mass spectrometry has become a powerful technique in biomedical research. The development of various analytical tools and online libraries has promoted the identification of biomarkers. However, how to make mass spectrometry collect more data information is an important but underestimated research topic. Herein, we combined full-scan and data-dependent acquisition (DDA) modes to develop a new targeted DDA based on the inclusion list of differential and preidentified ions (dpDDA). In this workflow, the MS1 datasets for statistical analysis and metabolite preidentification were first obtained using full-scan, and then, the MS/MS datasets for metabolite identification were obtained using targeted DDA of quality control samples based on the inclusion list. Compared with the current methods (DDA, data-independent acquisition, targeted DDA with time-staggered precursor ion list, and iterative exclusion DDA), dpDDA showed better stability, higher characteristic ion coverage, higher differential metabolites' MS/MS coverage, and higher quality MS/MS spectra. Moreover, the same trend was verified in the analysis of large-scale clinical samples. More surprisingly, dpDDA can distinguish patients with different severities of coronary heart disease (CHD) based on the Canadian Cardiovascular Society angina classification, which we cannot distinguish through conventional metabolomics data collection. Finally, dpDDA was employed to differentiate CHD from healthy control, and targeted metabolomics confirmed that dpDDA could identify a more complete metabolic pathway network. At the same time, four unreported potential CHD biomarkers were identified, and the area under the receiver operating characteristic curve was greater than 0.85. These results showed that dpDDA would expand the discovery of biomarkers based on metabolomics, more comprehensively explore the key metabolites and their association with diseases, and promote the development of precision medicine.

Nanoporous Crystalline Materials for the Recognition and Applications of Nucleic Acids.

Nucleic acid plays a crucial role in countless biological processes. Hence, there is great interest in its detection and analysis in various fields from chemistry, biology, to medicine. Nanoporous crystalline materials exhibit enormous potential as an effective platform for nucleic acid recognition and application. These materials have highly ordered and uniform pore structures, as well as adjustable surface chemistry and pore size, making them good carriers for nucleic acid extraction, detection, and delivery. In this review, the latest developments in nanoporous crystalline materials, including metal organic frameworks (MOFs), covalent organic frameworks (COFs), and supramolecular organic frameworks (SOFs) for nucleic acid recognition and applications are discussed. Different strategies for functionalizing these materials are explored to specifically identify nucleic acid targets. Their applications in selective separation and detection of nucleic acids are highlighted. They can also be used as DNA/RNA sensors, gene delivery agents, host DNAzymes, and in DNA-based computing. Other applications include catalysis, data storage, and biomimetics. The development of novel nanoporous crystalline materials with enhanced biocompatibility has opened up new avenues in the fields of nucleic acid analysis and therapy, paving the way for the development of sensitive, selective, and cost-effective diagnostic and therapeutic tools with widespread applications.

Prognosis difference between HER2-low and HER2-zero breast cancer patients: a systematic review and meta-analysis.

BACKGROUND: HER2-low breast cancer (BC) is proposed to be a special population of patients with an immunohistochemistry (IHC) score of 1 + or 2 + and non-amplified in situ hybridization (ISH) results. The role and prognostic impact of HER2-low BC is still controversial. This meta-analysis aims to explore the prognostic difference between of HER2-low and HER2-zero characteristic in BC patients. METHODS: A meta-analysis was conducted using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines and eligible studies were search in PubMed, Web of Science and EMBASE databases. Quality assessment of included studies were performed by Quality in Prognostic Studies (QUIPS) tool. Hazard ratios (HRs) and corresponding 95% confidence interval (CI) for overall survival (OS) and disease-free survival (DFS) were pooled in a meta-analysis. Furthermore, subgroup analysis, sensitivity analysis, and analysis for publication bias were conducted. RESULTS: Eighteen studies comprising a total of 93,317 patients were included for meta-analysis. BC patients with HER2-low characteristic have longer OS (HRs 0.87, 95% CI 0.81-0.93, p < 0.0001) and DFS (HRs 0.82, 95% CI 0.73-0.93, p = 0.001) compared to those with HER2-zero characteristic. Subgroup analysis indicate that the source of heterogeneity may come from the hormone receptor (HR) status group. Although, the publication bias was detected, sensitivity analysis and the trim-and-fill method analysis demonstrated the stability and reliability of the results. CONCLUSION: HER2-low BC patients have longer OS and DFS compared to HER2-zero BC patients, and its prognostic value is consistent among different HR status patients. Whether HER2-low breast cancer is an independent subtype of breast cancer is still a subject of ongoing research, and more studies are needed to fully understand the molecular and clinical features of this subtype.

Prophylactic Central Neck Dissection Based on Preoperative Imaging and Intraoperative Surgeon's Palpation Versus Total Thyroidectomy Alone for Papillary Thyroid Cancer.

INTRODUCTION: To compare the overall morbidity and recurrence of papillary thyroid cancer (PTC) after total thyroidectomy (TT) with or without prophylactic central compartment neck dissection (CCND) in cases of both preoperative and intraoperative nonsuspicious central lymph nodes (CLNs). METHODS: A total of 570 PTC patients who harbored no preoperative and intraoperative suspicious CLNs at two institutions were enrolled. They were randomly assigned to TT alone or TT with prophylactic CCND (pCCND) after intraoperative assessment of CLNs during the surgery. Lymph nodes that were hard or large enough to be palpated were regarded as suspicious metastatic lymph nodes during the surgery. The characteristics, postoperative complications, and locoregional recurrence of the two groups were recorded and compared. RESULTS: With a median follow-up of 5 y, the rates of lymph node recurrence in the TT alone and TT with pCCND groups were similar (7.3% versus 4.6%, P = 0.247), but there were significantly higher rates of overall morbidity (6.6% versus 19.1%, P < 0.001) when pCCND was performed. CONCLUSIONS: pCCND is not recommended for patients with clinically node-negative PTC preoperatively and intraoperatively because of the high complication rate and lack of benefit of reducing recurrence.

Optimizing ex vivo culture conditions to study human gut microbiome.

The inter-individual variations of gut microbiome contribute to the different responses toward drug therapy among populations, developing a reliable ex vivo culture method for mixed bacteria is the urgent need for predicting personal reaction to drug therapy. Unfortunately, very few attentions have been paid to the bias that could be introduced during the culture process for mixed bacteria. Here we systemically evaluated the factors that may affect the outcomes of cultured bacteria from human feces. We demonstrated that inter-individual difference of host gut microbiome was the main factor affecting the outcomes of cultured bacteria, followed by the culture medium and time point. We further optimized a new medium termed GB based on our established multi-dimensional evaluation method, which could mimic the status of in situ host gut microbiome to the highest extent. Finally, we assessed the inter-individual metabolism by host gut microbiome from 10 donors on three frequently used clinical drugs (aspirin, levodopa and doxifluridine) based on the optimized GB medium. Our results revealed obvious variation in drug metabolism by microbiome from different donors, especially levodopa and doxifluridine. This work suggested the optimized culture medium had the potential for exploring the inter-individual impacts of host gut microbiome on drug metabolism.

Microbiome-metabolome reveals that the Suxiao Jiuxin pill attenuates acute myocardial infarction associated with fatty acid metabolism.

ETHNOPHARMACOLOGICAL RELEVANCE: The Suxiao Jiuxin pill (SJP) is a Chinese medical patent drug on the national essential drug list of China, with well-established cardiovascular protective effects in the clinic. However, the mechanisms underlying the protective effects of SJP on cardiovascular disease have not yet been elucidated clearly, especially its relationship with the gut microbiota. AIM OF THE STUDY: This study aimed to investigate the cardioprotective effect of SJP against isoproterenol-induced acute myocardial infarction (AMI) by integrating the gut microbiome and metabolome. METHODS: A rat model of AMI was generated using isoproterenol. Firstly, the effect of antibiotic (ABX) treatment on the blood absorption and excretion of the main components of SJP were studied. Secondly, 16S rRNA sequencing and untargeted metabolomics were used to discover the improvement of SJP treatment on gut microbiota and host metabolism in AMI rats. Finally, targeted metabolomics was used to verify the effects of SJP treatment on host metabolism in AMI rats. RESULT: The results showed that ABX treatment could affect the blood absorption and fecal excretion of the main active components of SJP. At the same time, SJP can restore the richness and diversity of gut microbiota, and multiple gut microbiota (including Jeotgalicoccus, Lachnospiraceae, and Blautia) are significantly associated with fatty acids. Untargeted metabolomics also found that SJP could restore the levels of various fatty acid metabolites in serum and cecal contents (p < 0.01, FC > 1.5 and VIP >1). Targeted metabolomics further confirmed that 41, 21, and 39 fatty acids were significantly altered in serum, cecal contents, and heart samples, respectively. Interestingly, these fatty acids belong to the class of eicosanoids, and SJP can significantly downregulate these eicosanoids in AMI rats. CONCLUSION: The results of this study suggest that SJP may exert its cardioprotective effects by remodeling the gut microbiota and host fatty acid metabolism.

A self-monitoring microneedle patch for light-controlled synergistic treatment of melanoma.

Melanoma is the most aggressive and malignant form of skin cancer. Current melanoma treatment methods generally suffer from frequent drug administration as well as difficulty in direct monitoring of drug release. Here, a self-monitoring microneedle (MN)-based drug delivery system, which integrates a dissolving MN patch with aggregation-induced emission (AIE)-active PATC microparticles, is designed to achieve light-controlled pulsatile chemo-photothermal synergistic therapy of melanoma. The PATC polymeric particles, termed D/I@PATC, encapsulate both of chemotherapeutic drug doxorubicin (DOX) and the photothermal agent indocyanine green (ICG). Upon light illumination, PATC gradually dissociates into smaller particles, causing the release of encapsulated DOX and subsequent fluorescence intensity change of PATC particles, thereby not only enabling direct observation of the drug release process under light stimuli, but also facilitating verification of drug release by fluorescence recovery after light trigger. Moreover, encapsulation of ICG in PATC particles displays significant improvement of its photothermal stability both in vitro and in vivo. In a tumor-bearing mouse, the application of one D/I@PATC MN patch combining with two cycles of light irradiation showed excellent controllable chemo-photothermal efficacy and exhibited ∼97% melanoma inhibition rate without inducing any evident systemic toxicity, suggesting a great potential for skin cancer treatment in clinics.

Nephrotoxicity assessment of Esculentoside A using human-induced pluripotent stem cell-derived organoids.

Drug-induced nephrotoxicity is a leading cause of acute kidney injury (AKI). A major obstacle in predicting AKI is the lack of a comprehensive experimental model that mimics stable and physiologically relevant kidney functions and accurately reflects the changes a drug induces. Organoids derived from human-induced pluripotent stem cells (iPSCs) are promising models because of their reproducibility and similarity to the in vivo conditions. In this study, Esculentoside A, the triterpene saponin with the highest concentration isolated from the root of Phytolacca acinose Roxb., was used to induce kidney injury models in vivo and kidney organoids. Esculentoside A induced AKI in mice, together with pathological changes and enhanced apoptosis. Moreover, Esculentoside A damaged podocytes and proximal tubular endothelial cells in kidney organoids in a similar way as in vivo. We also found that treatment with 60 µM Esculentoside A induced the known biomarkers of kidney damage and inflammatory cytokines (such as kidney injury molecule (KIM-1), ß2-microglobulin (ß2-M), and cystatin C (CysC)) in the organoids, in which activation of Cleaved Caspase-3 was involved, possibly due to lowered mitochondrial membrane potential. In summary, this study strongly suggests using kidney organoids as a reliable platform to assess Chinese medicine-induced nephrotoxicity.

The deubiquitinating enzyme UCHL3 promotes anaplastic thyroid cancer progression and metastasis through Hippo signaling pathway.

Yes-associated protein (YAP) is one of major key effectors of the Hippo pathway and the mechanism supporting abnormal YAP expression in Anaplastic thyroid carcinoma (ATC) remains to be characterized. Here, we identified ubiquitin carboxyl terminal hydrolase L3 (UCHL3) as a bona fide deubiquitylase of YAP in ATC. UCHL3 stabilized YAP in a deubiquitylation activity-dependent manner. UCHL3 depletion significantly decreased ATC progression, stem-like and metastasis, and increased cell sensitivity to chemotherapy. Depletion of UCHL3 decreased the YAP protein level and the expression of YAP/TEAD target genes in ATC. UCHL3 promoter analysis revealed that TEAD4, through which YAP bind to DNA, activated UCHL3 transcription by binding to the promoter of UCHL3. In general, our results demonstrated that UCHL3 plays a pivotal role in stabilizing YAP, which in turn facilitates tumorigenesis in ATC, suggesting that UCHL3 may prove to be a potential target for the treatment of ATC.