Discovery of Kebanmycins with Antibacterial and Cytotoxic Activities from the Mangrove-Derived Streptomyces sp. SCSIO 40068.

Mangrove derived actinomycetes are a rich reservoir of bioactive natural products and play important roles in pharmaceutical chemistry. In a screen of actinomycetes from mangrove rhizosphere sedimental environments, the isolated strain Streptomyces sp. SCSIO 40068 displayed strong antibacterial activity. Further fractionation of the extract yielded four new compounds kebanmycins A-D (1-4) and two known analogues FD-594 (5) and the aglycon (6). The structures of 1-6 were determined based on extensive spectroscopic data and single-crystal X-ray diffraction analysis. 1-3 featured a fused pyranonaphthaxanthene as an integral part of a 6/6/6/6/6/6 polycyclic motif, and showed bioactivity against a series of Gram-positive bacteria and cytotoxicity to several human tumor cells. In addition, the kebanmycins biosynthetic gene cluster (keb) was identified in Streptomyces sp. SCSIO 40068, and KebMT2 was biochemically characterized as a tailoring sugar-O-methyltransferase, leading to a proposed biosynthetic route to 1-6. This study paves the way to further investigate 1 as a potential lead compound.

Helicobacter pylori disrupts gastric mucosal homeostasis by stimulating macrophages to secrete CCL3.

BACKGROUND: Helicobacter pylori (H. pylori) is the predominant etiological agent of gastritis and disrupts the integrity of the gastric mucosal barrier through various pathogenic mechanisms. After H. pylori invades the gastric mucosa, it interacts with immune cells in the lamina propria. Macrophages are central players in the inflammatory response, and H. pylori stimulates them to secrete a variety of inflammatory factors, leading to the chronic damage of the gastric mucosa. Therefore, the study aims to explore the mechanism of gastric mucosal injury caused by inflammatory factors secreted by macrophages, which may provide a new mechanism for the development of H. pylori-related gastritis. METHODS: The expression and secretion of CCL3 from H. pylori infected macrophages were detected by RT-qPCR, Western blot and ELISA. The effect of H. pylori-infected macrophage culture medium and CCL3 on gastric epithelial cells tight junctions were analyzed by Western blot, immunofluorescence and transepithelial electrical resistance. EdU and apoptotic flow cytometry assays were used to detect cell proliferation and apoptosis levels. Dual-luciferase reporter assays and chromatin immunoprecipitation assays were used to study CCL3 transcription factors. Finally, gastric mucosal tissue inflammation and CCL3 expression were analyzed by hematoxylin and eosin staining and immunohistochemistry. RESULTS: After H. pylori infection, CCL3 expressed and secreted from macrophages were increased. H. pylori-infected macrophage culture medium and CCL3 disrupted gastric epithelial cells tight junctions, while CCL3 neutralizing antibody and receptor inhibitor of CCL3 improved the disruption of tight junctions between cells. In addition, H. pylori-infected macrophage culture medium and CCL3 recombinant proteins stimulated P38 phosphorylation, and P38 phosphorylation inhibitor improved the disruption of tight junctions between cells. Besides, it was identified that STAT1 was a transcription factor of CCL3 and H. pylori stimulated macrophage to secret CCL3 through the JAK1-STAT1 pathway. Finally, after mice were injected with murine CCL3 recombinant protein, the gastric mucosal injury and inflammation were aggravated, and the phosphorylation level of P38 was increased. CONCLUSIONS: In summary, our findings demonstrate that H. pylori infection stimulates macrophages to secrete CCL3 via the JAK1-STAT1 pathway. Subsequently, CCL3 damages gastric epithelial tight junctions through the phosphorylation of P38. This may be a novel mechanism of gastric mucosal injury in H. pylori-associated gastritis.

Accuracy of cone-beam breast computed tomography for assessing breast cancer tumor size-comparison with breast magnetic resonance imaging.

Background: Accurate preoperative assessment of tumor size is important in developing a surgical plan for breast cancer. The purpose of this study was to evaluate the accuracy of cone-beam breast computed tomography (CBBCT) and magnetic resonance imaging (MRI) in the assessment of tumor size and to analyze the factors influencing the discordance. Methods: In this retrospective study, patients with breast cancer who underwent preoperative contrast-enhanced CBBCT (CE-CBBCT) and dynamic contrast-enhanced MRI (DCE-MRI) and received a complete pathologic diagnosis from August 2020 to December 2021 were included, using the pathological result as the gold standard. Two radiologists assessed the CBBCT and MRI features and measured the tumor size with a 2-week washout period. Intraclass correlation coefficient (ICC) and Bland-Altman analyses were used to assess inter-observer reproducibility and agreement based on CBBCT, MRI and pathology. Univariate analyses of differences in clinical, pathological and CBBCT/MRI features between the concordant and discordant groups was performed using the t-test, Mann-Whitney U-test, Chi-squared test and Fisher's exact test. Multivariate analyses were used to identify factors associated with discordance of CBBCT/MRI with pathology. Results: A total of 115 female breast cancer patients (115 lesions) were included. All patients had a single malignant tumor of the unilateral breast. The reproducibility and the agreement ranged from moderate to excellent (ICC =0.607-0.983). Receiver operating characteristic (ROC) analyses showed that the cut-off values of CBBCT-pathology and MRI-pathology discordance were 2.25 and 2.65 cm, respectively. CBBCT/MRI-pathology concordance was significantly associated with the extent of pathology, lesion type, presence of calcification, human epidermal growth factor receptor 2 (HER2) status and fatty infiltration (P<0.05). In lesions containing calcification, the difference of CBBCT-pathology was significantly smaller than MRI-pathology (P=0.021). Non-mass enhancement (NME) was the main predictor of CBBCT- or MRI-pathology discordance [odds ratio (OR) =3.293-6.469, P<0.05], and HER2 positivity was a predictor of CBBCT-pathology discordance (OR =3.514, P=0.019). Conclusions: CBBCT and MRI have comparable accuracy in measurement of tumor size, and CBBCT is advantageous in assessing the size of calcified lesions. NME and HER2 positivity are significant predictors of CBBCT-pathology discordance. This suggests that CBBCT might serve as an alternative imaging technique to assess tumor size when patients do not tolerate MRI.

RIOK3 sustains colorectal cancer cell survival under glucose deprivation via an HSP90α-dependent pathway.

Glucose oxidation via the pentose phosphate pathway serves as the primary cellular mechanism for generating nicotinamide adenine dinucleotide phosphate (NADPH). The central regions of solid tumors typically experience glucose deficiency, emphasizing the need for sustained NADPH production crucial to tumor cell survival. This study highlights the crucial role of RIOK3 in maintaining NADPH production and colorectal cancer (CRC) cell survival during glucose deficiency. Our findings revealed upregulated RIOK3 expression upon glucose deprivation, with RIOK3 knockout significantly reducing cancer cell survival. Mechanistically, RIOK3 interacts with heat shock protein 90α (HSP90α), a chaperone integral to various cellular processes, thereby facilitating HSP90α binding to isocitrate dehydrogenase 1 (IDH1). This interaction further upregulates IDH1 expression, enhancing NADPH production and preserving redox balance. Furthermore, RIOK3 inhibition had no discernible effect on intracellular NADPH levels and cell death rates in HSP90α-knockdown cells. Collectively, our findings suggest that RIOK3 sustains colon cancer cell survival in low-glucose environments through an HSP90α-dependent pathway. This highlights the significance of the RIOK3-HSP90α-IDH1 cascade, providing insights into potential targeted therapeutic strategies for CRC in metabolic stress conditions.

Hinokitiol protects gastric injury from ethanol exposure via its iron sequestration capacity.

Hinokitiol is a natural bioactive tropolone derivative isolated from Chamaecyparis obtusa and Thuja plicata, which exhibits promising potential in terms of antioxidant and anti-inflammatory properties and possesses potent iron-binding capacity. In this study, we aimed to investigate the potential role of hinokitiol in protecting against ethanol-induced gastric injury and elucidate the underlying mechanism. Our results demonstrated that hinokitiol effectively attenuated hemorrhagic gastric lesions, epithelial cell loss, and inflammatory response in mice with ethanol-induced gastric injury. Intriguingly, we found that ethanol exposure affects iron levels both in vivo and in vitro. Moreover, the disturbed iron homeostasis was involved in the development of ethanol-induced injury. Iron depletion was found to enhance defense against ethanol-induced damage, while iron repletion showed the opposite effect. To further explore the role of iron sequestration in the protective effects of hinokitiol, we synthesized methylhinokitiol, a compound that shields the iron binding capacity of hinokitiol with a methyl group. Interestingly, this compound significantly diminishes the protective effect against ethanol-induced injury. These findings collectively demonstrated that hinokitiol could potentially be used to prevent or improve gastric injury induced by ethanol through regulating cellular iron homeostasis.

Radiomics in cone-beam breast CT for the prediction of axillary lymph node metastasis in breast cancer: a multi-center multi-device study.

OBJECTIVES: To develop a radiomics model in contrast-enhanced cone-beam breast CT (CE-CBBCT) for preoperative prediction of axillary lymph node (ALN) status and metastatic burden of breast cancer. METHODS: Two hundred and seventy-four patients who underwent CE-CBBCT examination with two scanners between 2012 and 2021 from two institutions were enrolled. The primary tumor was annotated in each patient image, from which 1781 radiomics features were extracted with PyRadiomics. After feature selection, support vector machine models were developed to predict ALN status and metastatic burden. To avoid overfitting on a specific patient subset, 100 randomly stratified splits were made to assign the patients to either training/fine-tuning or test set. Area under the receiver operating characteristic curve (AUC) of these radiomics models was compared to those obtained when training the models only with clinical features and combined clinical-radiomics descriptors. Ground truth was established by histopathology. RESULTS: One hundred and eighteen patients had ALN metastasis (N + (≥ 1)). Of these, 74 had low burden (N + (1~2)) and 44 high burden (N + (≥ 3)). The remaining 156 patients had none (N0). AUC values across the 100 test repeats in predicting ALN status (N0/N + (≥ 1)) were 0.75 ± 0.05 (0.67~0.93, radiomics model), 0.68 ± 0.07 (0.53~0.85, clinical model), and 0.74 ± 0.05 (0.67~0.88, combined model). For metastatic burden prediction (N + (1~2)/N + (≥ 3)), AUC values were 0.65 ± 0.10 (0.50~0.88, radiomics model), 0.55 ± 0.10 (0.40~0.80, clinical model), and 0.64 ± 0.09 (0.50~0.90, combined model), with all the ranges spanning 0.5. In both cases, the radiomics model was significantly better than the clinical model (both p < 0.01) and comparable with the combined model (p = 0.56 and 0.64). CONCLUSIONS: Radiomics features of primary tumors could have potential in predicting ALN metastasis in CE-CBBCT imaging. CLINICAL RELEVANCE STATEMENT: The findings support potential clinical use of radiomics for predicting axillary lymph node metastasis in breast cancer patients and addressing the limited axilla coverage of cone-beam breast CT. KEY POINTS: • Contrast-enhanced cone-beam breast CT-based radiomics could have potential to predict N0 vs. N + (≥ 1) and, to a limited extent, N + (1~2) vs. N + (≥ 3) from primary tumor, and this could help address the limited axilla coverage, pending future verifications on larger cohorts. • The average AUC of radiomics and combined models was significantly higher than that of clinical models but showed no significant difference between themselves. • Radiomics features descriptive of tumor texture were found informative on axillary lymph node status, highlighting a higher heterogeneity for tumor with positive axillary lymph node.

TRIM28 suppresses cancer stem-like characteristics in gastric cancer cells through Wnt/ß-catenin signaling pathways.

The influences of TRIM28 on the gastric tumorigenesis together with potential molecular mechanisms remain to be studied. We aimed at exploring the important effects of TRIM28 on gastric cancer (GC) and uncovering underling molecular mechanisms. Through immunohistochemistry analysis of 20 pairs of GC and the peritumoral tissues, the expression level of TRIM28 was determined. A variety of assays were applied to explore the important roles of TRIM28 in GC. Western blotting and qRT-PCR analyses were used to analyze the association between TRIM28 and the Wnt/ß-catenin signaling pathway. TRIM28 was highly expressed in GC tissues than peritumoral tissues. And high expression level of TRIM28 in GC was associated with good prognostic effects. In vitro functional assays suggested TRIM28 knockdown enhanced the proliferation and clone formation of GC cell. Moreover, TRIM28 knockdown enhanced the expression level of stemness markers, strengthened sphere-forming and drug-resistance properties of GC cells, suggesting important effect on GC cell stemness. Besides, our analysis showed that the Wnt/ß-catenin signaling was involved in the effect of TRIM28 on GC cell stemness property, and blocking Wnt/ß-catenin signaling pathway obviously rescued the promotion influence of TRIM28 knockdown. Overall, TRIM28 has an important influence on regulating the stem-like property of GC cell via Wnt/ß-catenin signaling, suggesting TRIM28 a promising drug target and a potential predictor of prognosis.

A machine learning-based radiomics model for prediction of tumor mutation burden in gastric cancer.

Purpose: To evaluate the potential of machine learning (ML)-based radiomics approach for predicting tumor mutation burden (TMB) in gastric cancer (GC). Methods: The contrast enhanced CT (CECT) images with corresponding clinical information of 256 GC patients were retrospectively collected. Patients were separated into training set (n = 180) and validation set (n = 76). A total of 3,390 radiomics features were extracted from three phases images of CECT. The least absolute shrinkage and selection operator (LASSO) model was used for feature screening. Seven machine learning (ML) algorithms were employed to find the optimal classifier. The predictive ability of radiomics model (RM) was evaluated with receiver operating characteristic. The correlation between RM and TMB values was evaluated using Spearman's correlation coefficient. The explainability of RM was assessed by the Shapley Additive explanations (SHAP) method. Results: Logistic regression algorithm was chosen for model construction. The RM showed good predictive ability of TMB status with AUCs of 0.89 [95% confidence interval (CI): 0.85-0.94] and 0.86 (95% CI: 0.74-0.98) in the training and validation sets. The correlation analysis revealed a good correlation between RM and TMB levels (correlation coefficient: 0.62, p < 0.001). The RM also showed favorable and stable predictive accuracy within the cutoff value range 6-16 mut/Mb in both sets. Conclusion: The ML-based RM offered a promising image biomarker for predicting TMB status in GC patients.

Both extracellular vesicles from helicobacter pylori-infected cells and helicobacter pylori outer membrane vesicles are involved in gastric/extragastric diseases.

Bacterial-derived extracellular vesicles (EVs) have emerged as crucial mediators in the cross-talk between hosts and pathogens, playing a significant role in infectious diseases and cancers. Among these pathogens, Helicobacter pylori (H. pylori) is a particularly important bacterium implicated in various gastrointestinal disorders, gastric cancers, and systemic illnesses. H. pylori achieves these effects by stimulating host cells to secrete EVs and generating internal outer membrane vesicles (OMVs). The EVs derived from H. pylori-infected host cells modulate inflammatory signaling pathways, thereby affecting cell proliferation, apoptosis, cytokine release, immune cell modification, and endothelial dysfunction, as well as disrupting cellular junctional structures and inducing cytoskeletal reorganization. In addition, OMVs isolated from H. pylori play a pivotal role in shaping subsequent immunopathological responses. These vesicles incite both inflammatory and immunosuppressive reactions within the host environment, facilitating pathogen evasion of host defenses and invasion of host cells. Despite this growing understanding, research involving H. pylori-derived EVs remains in its early stages across different domains. In this comprehensive review, we present recent advancements elucidating the contributions of EV components, such as non-coding RNAs (ncRNAs) and proteins, to the pathogenesis of gastric and extragastric diseases. Furthermore, we highlight their potential utility as biomarkers, therapeutic targets, and vehicles for targeted delivery.

Feasibility analysis of magnetic resonance imaging-based radiomics features for preoperative prediction of nuclear grading of ductal carcinoma in situ.

Background: The nuclear grading of ductal carcinoma in situ (DCIS) affects its clinical risk. The aim of this study was to investigate the possibility of predicting the nuclear grading of DCIS, by magnetic resonance imaging (MRI)-based radiomics features. And to develop a nomogram combining radiomics features and MRI semantic features to explore the potential role of MRI radiomic features in the assessment of DCIS nuclear grading. Methods: A total of 156 patients (159 lesions) with DCIS and DCIS with microinvasive (DCIS-MI) were enrolled in this retrospective study, with 112 lesions included in the training cohort and 47 lesions included in the validation cohort. Radiomics features were extracted from Dynamic contrast-enhanced MRI (DCE-MRI) phases 1st and 5th. After feature selection, radiomics signature was constructed and radiomics score (Rad-score) was calculated. Multivariate analysis was used to identify MRI semantic features that were significantly associated with DCIS nuclear grading and combined with Rad-score to construct a Nomogram. Receiver operating characteristic curves were used to evaluate the predictive performance of Rad-score and Nomogram, and decision curve analysis (DCA) was used to evaluate the clinical utility. Results: In multivariate analyses of MRI semantic features, larger tumor size and heterogeneous enhancement pattern were significantly associated with high-nuclear grade DCIS (HNG DCIS). In the training cohort, Nomogram had an area under curve (AUC) of 0.879 and Rad-score had an AUC of 0.828. Similarly, in the independent validation cohort, Nomogram had an AUC value of 0.828 and Rad-score had an AUC of 0.772. In both the training and validation cohorts, Nomogram had a significantly higher AUC value than Rad-score (P<0.05). DCA confirmed that Nomogram had a higher net clinical benefit. Conclusions: MRI-based radiomic features can be used as potential biomarkers for assessing nuclear grading of DCIS. The nomogram constructed by radiomic features combined with semantic features is feasible in discriminating non-HNG and HNG DCIS.

Radiomics nomogram for predicting axillary lymph node metastasis-a potential method to address the limitation of axilla coverage in cone-beam breast CT: a bi-center retrospective study.

PURPOSE: Cone-beam breast CT (CBBCT) has an inherent limitation that the axilla cannot be imaged in its entirety. We aimed to develop and validate a nomogram based on clinical factors and contrast-enhanced (CE) CBBCT radiomics features to predict axillary lymph node (ALN) metastasis and complement limited axilla coverage. MATERIAL AND METHODS: This retrospective study included 312 patients with breast cancer from two hospitals who underwent CE-CBBCT examination in a clinical trial (NCT01792999) during 2012-2020. Patients from TCIH comprised training set (n = 176) and validation set (n = 43), and patients from SYSUCC comprised external test set (n = 93). 3D ROIs were delineated manually and radiomics features were extracted by 3D Slicer software. RadScore was calculated and radiomics model was constructed after feature selection. Clinical model was built on independent predictors. Nomogram was developed with independent clinical predictors and RadScore. Diagnostic performance was compared among three models by ROC curve, and decision curve analysis (DCA) was used to evaluate the clinical utility of nomogram. RESULTS: A total of 139 patients were ALN positive and 173 patients were negative. Twelve radiomics features remained after feature selection. Location and focality were selected as independent predictors for ALN status. The AUC of nomogram in external test set was higher than that of clinical model (0.80 vs. 0.66, p = 0.012). DCA demonstrated that the nomogram had higher overall net benefit than that of clinical model. CONCLUSION: The nomogram combined CE-CBBCT-based radiomics features and clinical factors could have potential in distinguishing ALN positive from negative and addressing the limitation of axilla coverage in CBBCT.

Assessing metal ion transporting activity of ZIPs: Intracellular zinc and iron detection.

Zrt/Irt-like proteins (ZIPs or SLC39A) are a large family of metal ion transporters mainly responsible for zinc uptake. Some ZIPs have been shown to specifically transport zinc, whereas others have broader substrate specificity in divalent metal ion trafficking, notably those of zinc and iron ions. Measuring intracellular zinc and iron levels helps assess their molecular and physiological activities. This chapter presents step-by-step methods for evaluating intracellular metal ion concentrations, including direct measurement using inductively coupled plasma-mass spectrometry (ICP-MS), chemical staining, fluorescent probes, and indirect reporter assays such as activity analysis of enzymes whose activities are dependent on metal ion availability.

Accuracy of Preoperative Contrast-enhanced Cone Beam Breast CT in Assessment of Residual Tumor after Neoadjuvant Chemotherapy: A Comparative Study with Breast MRI.

RATIONALE AND OBJECTIVES: To compare the accuracy of preoperative contrast-enhanced cone beam breast CT (CE-CBBCT) and MRI in assessment of residual tumor after neoadjuvant chemotherapy (NAC). MATERIALS AND METHODS: Residual tumor assessments in 91 female patients were performed on preoperative CE-CBBCT and MRI images after NAC. The agreements of tumor size between imaging and pathology were tested by Intraclass Correlation Coefficient (ICC). Subgroup analyses were set according to ductal carcinoma in situ (DCIS), calcifications and molecular subtypes. Correlated-samples Wilcoxon Signed-rank test was used to analyze the difference between imaging and pathology in total and subgroups. AUC, sensitivity, specificity, PPV, and NPV were calculated to compare the performance of CE-CBBCT and MRI in predicting pathological complete response (pCR). RESULTS: Comparing with pathology, the agreement on CE-CBBCT was good (ICC = 0.64, 95% CI, 0.35-0.78), whereas on MRI was moderate (ICC = 0.59, 95% CI, 0.36-0.77), and overestimation on CE-CBBCT was less than that on MRI (median (interquartile range, IQR): 0.24 [0.00, 1.31] cm vs. 0.67 [0.00, 1.81] cm; p = 0.000). In subgroup analysis, CE-CBBCT showed superior accuracy than MRI when residual DCIS (p = 0.000) and calcifications (p = 0.000) contained, as well as luminal A (p = 0.043) and luminal B (p = 0.009) breast cancer. CE-CBBCT and MRI performed comparable in predicting pCR, AUCs were 0.749 and 0.733 respectively (p > 0.05). CONCLUSION: CE-CBBCT showed superior accuracy in assessment of residual tumor compared with MRI, especially when residual DCIS or calcifications contained and luminal subtype. The performance of preoperative CE-CBBCT in predicting pCR was comparable to MRI. CE-CBBCT could be an alternative method used for preoperative assessment after NAC.

M2 Macrophage-Derived Exosomal Ferritin Heavy Chain Promotes Colon Cancer Cell Proliferation.

Colon cancer is a widespread life-threatening malignancy with complex and multifactorial etiology. Both epidemiological cohort studies and basic research support the substantial role of iron metabolism in colon cancer. Thus, understanding the mechanisms of how essential iron metabolic proteins are dysregulated may provide new treatment strategies for colon cancer. Ferritin is the main iron storage protein that occupies a vital position in iron metabolism. Studies reported that ferritin is differentially highly expressed in tissues from multiple malignancies. However, the source and function of highly expressed ferritin in colon cancer have not been explored. In this study, we found that the protein level but not RNA level of ferritin heavy chain (FTH1) was upregulated in colon cancer using paired clinical samples. Co-culture system was used to mimic the in vivo circumstance and study the cell-cell communication of macrophages and colon cancer cells. Results showed that M2 macrophages could substantially increase the FTH1 levels in colon cancer cells. This effect could be blocked by the exosome biogenesis/ secretion inhibitor GW4869, implying the vital role of exosomes in this biological process. Besides, we found that purified exosomes from M2 macrophages could deliver FTH1 into colon cancer cells and promote cell proliferation. Furtherly, EdU assay and live cell imaging system were performed in FTH1-OE (overexpression) colon cancer cell lines and confirmed the cell proliferation promoting effect of FTH1. Our results unveil the source and function of highly expressed FTH1 in colon cancer and provide a new potential therapeutic target for the treatment of colon cancer.

Nitric Oxide Negatively Regulates the Rapid Formation of Pleurotus ostreatus Primordia by Inhibiting the Mitochondrial aco Gene.

Nitric oxide (NO) is as a signaling molecule that participates in the regulation of plant development and in a number of physiological processes. However, the function and regulatory pathway of NO in the growth and development of edible mushrooms are still unknown. This study found that NO played a negative role in the transformation of Pleurotus ostreatus from vegetative growth to reproductive growth by the exogenous addition of NO donors and scavengers. Further studies showed that NO can inhibit the gene expression and enzyme activity of aconitase (ACO). Moreover, the overexpression (OE) of mitochondrial aco and RNA interference (RNAi) confirmed that ACO participates in the regulation of the primordia formation rate. The effects of aco OE and RNAi on the tricarboxylic acid (TCA) cycle and energy metabolism were further measured. The results showed that RNAi-aco mutant strains can affect the enzyme activities of isocitrate dehydrogenase of mitochondria (ICDHm) and α-ketoglutarate dehydrogenase (α-KGDH) in the TCA cycle, thereby reducing the production of nicotinamide adenine dinucleotide (NADH) in the TCA cycle, decreasing the contents of adenosine triphosphate (ATP) and hydrogen peroxide (H2O2), and negatively regulating the rapid formation of primordia. In addition, H2O2 was significantly increased during the transformation from vegetative growth to reproductive growth of P. ostreatus. Additionally, the exogenous addition of H2O2 and its scavengers further confirmed the positive regulation by H2O2 in primordia formation. This study shows that during the growth and development of P. ostreatus, NO can inhibit the expression of the mitochondrial aco gene and ACO protein in the TCA cycle, reduce the production of ATP and H2O2 in the respiratory chain, and negatively regulate the rate of primordia formation.

Comparison of background parenchymal enhancement (BPE) on contrast-enhanced cone-beam breast CT (CE-CBBCT) and breast MRI.

OBJECTIVES: To compare the background parenchymal enhancement (BPE) levels on contrast-enhanced cone-beam breast CT (CE-CBBCT) and MRI, evaluate inter-reader reliability, and analyze the relationship between clinical factors and BPE level on CE-CBBCT. METHODS: In this retrospective study, patients who underwent both CE-CBBCT and MRI were analyzed. BPE levels on CE-CBBCT and MRI were assessed by five specialists independently in random fashion, with a wash-out period of 4 weeks. Weighted kappa was used to analyze the agreement between CE-CBBCT and MRI, and intraclass correlation coefficient (ICC) was used to evaluate the inter-reader reliability for each modality. The association between BPE level on CE-CBBCT and clinical factors was evaluated by univariate and multivariate logistic regression. RESULTS: A total of 221 patients from January 2017 to April 2021 were enrolled. CE-CBBCT showed substantial agreement (weighted kappa = 0.690) with MRI for BPE evaluation, with good degree of inter-reader reliability on both CE-CBBCT (ICC = 0.712) and MRI (ICC = 0.757). Based on majority reports, BPE levels on CE-CBBCT were lower than MRI (p < 0.001). BPE level on CE-CBBCT was significantly associated with menstrual status (odds ratio, OR = 0.125), breast density (OR = 2.308), and previously treated breast cancer (OR = 0.052) (all p < 0.05). BPE level for premenopausal patients was associated with menstrual cycle, with lower BPE level for the 2nd week of menstrual cycle (OR = 0.246). CONCLUSIONS: CE-CBBCT showed substantial agreement and comparable inter-reader reliability with MRI for BPE evaluation, indicating that the corresponding BI-RADS lexicons could be used to describe BPE level on CE-CBBCT. The 2nd week of menstrual cycle timing is suggested as the optimal examination period for CE-CBBCT. KEY POINTS: • CE-CBBCT showed substantial agreement and comparable inter-reader reliability with MRI for BPE evaluation. • Menstrual status, breast density, and previously treated breast cancer were associated with the BPE level on CE-CBBCT images. • The 2ndweek of the menstrual cycle is suggested as the optimal examination period for CE-CBBCT.

Zinc antagonizes iron-regulation of tyrosine hydroxylase activity and dopamine production in Drosophila melanogaster.

BACKGROUND: Dopamine (DA) is a neurotransmitter that plays roles in movement, cognition, attention, and reward responses, and deficient DA signaling is associated with the progression of a number of neurological diseases, such as Parkinson's disease. Due to its critical functions, DA expression levels in the brain are tightly controlled, with one important and rate-limiting step in its biosynthetic pathway being catalyzed by tyrosine hydroxylase (TH), an enzyme that uses iron ion (Fe2+) as a cofactor. A role for metal ions has additionally been associated with the etiology of Parkinson's disease. However, the way dopamine synthesis is regulated in vivo or whether regulation of metal ion levels is a component of DA synthesis is not fully understood. Here, we analyze the role of Catsup, the Drosophila ortholog of the mammalian zinc transporter SLC39A7 (ZIP7), in regulating dopamine levels. RESULTS: We found that Catsup is a functional zinc transporter that regulates intracellular zinc distribution between the ER/Golgi and the cytosol. Loss-of-function of Catsup leads to increased DA levels, and we showed that the increased dopamine production is due to a reduction in zinc levels in the cytosol. Zinc ion (Zn2+) negatively regulates dopamine synthesis through direct inhibition of TH activity, by antagonizing Fe2+ binding to TH, thus rendering the enzyme ineffective or non-functional. CONCLUSIONS: Our findings uncovered a previously unknown mechanism underlying the control of cellular dopamine expression, with normal levels of dopamine synthesis being maintained through a balance between Fe2+ and Zn2+ ions. The findings also provide support for metal modulation as a possible therapeutic strategy in the treatment of Parkinson's disease and other dopamine-related diseases.

Alternative oxidase gene induced by nitric oxide is involved in the regulation of ROS and enhances the resistance of Pleurotus ostreatus to heat stress.

BACKGROUND: In China, during the cultivation process of Pleurotus ostreatus, the yield and quality of fruiting bodies are easily affected by high temperatures in summer. Nitric oxide (NO) plays an important regulatory role in the response to abiotic stress, and previous studies have found that NO can induce alternative oxidase (aox) experssion in response to heat stress (HS) by regulating aconitase. However, the regulatory pathway of NO is complex, and the function and regulation of the aox gene in the response to HS remain unclear. RESULTS: In this study, we found that NO affected nicotinamide adenine dinucleotide (NADH) and adenosine triphosphate (ATP) levels, reduced hydrogen peroxide (H2O2) and superoxide anion (O2-) contents, and slowed O2- production. Further RNA-Seq results showed that NO regulated the oxidation-reduction process and oxidoreductase activity, affected the cellular respiration pathway and activated aox gene expression. The function of aox was determined by constructing overexpression (OE) and RNA interference (RNAi) strains. The results showed that the OE-aox strains exhibited obviously improved growth recovery after exposure to HS. During exposure to HS, the OE-aox strains exhibited reduced levels of NADH, the product of the tricarboxylic acid (TCA) cycle, and decreased synthesis of ATP, which reduced the production and accumulation of reactive oxygen species (ROS), whereas the RNAi-aox strains exhibited the opposite result. In addition, aox mediated the expression of antioxidant enzyme genes in the mycelia of P. ostreatus under HS through the retrograde signaling pathway. CONCLUSIONS: This study shows that the expression of the aox gene in P. ostreatus mycelia can be induced by NO under HS, that it regulates the TCA cycle and cell respiration to reduce the production of ROS, and that it can mediate the retrograde signaling pathway involved in the mycelial response to HS.

Lactic acid accumulation under heat stress related to accelerated glycolysis and mitochondrial dysfunction inhibits the mycelial growth of Pleurotus ostreatus.

High temperature is a major threat to Pleurotus ostreatus cultivation. In this study, a potential mechanism by which P. ostreatus mycelia growth is inhibited under heat stress was explored. Lactate, as a microbial fermentation product, was found unexpectedly in the mycelia of P. ostreatus under heat stress, and the time-dependent accumulation and corresponding inhibitory effect of lactate on mycelial growth was further confirmed. The addition of a glycolysis inhibitor, 2-deoxy-D-glucose (2DG), reduced the lactate content in mycelia and slightly restored mycelial growth under high-temperature conditions, which indicated the accumulation of lactate can be inhibited by glycolysis inhibition. Further data revealed mitochondrial dysfunction under high-temperature conditions, with evidence of decreased oxygen consumption and adenosine triphosphate (ATP) synthesis and increased reactive oxygen species (ROS). The removal of ROS with ascorbic acid decreased the lactate content, and mycelial growth recovered to a certain extent, indicating lactate accumulation could be affected by the mitochondrial ROS. Moreover, metabolic data showed that glycolysis and the tricarboxylic acid cycle were enhanced. This study reported the accumulation of lactate in P. ostreatus mycelia under heat stress and the inhibitory effect of lactate on the growth of mycelia, which might provide further insights into the stress response mechanism of edible fungi. Key Points • Lactate can accumulate in Pleurotus ostreatus mycelia under heat stress and inhibit its growth. • The accumulation of lactate may be due to the acceleration of glycolysis and the dysfunction of mitochondria of P. ostreatus mycelia under high-temperature stress. • The glycolysis and tricarboxylic acid cycle of P. ostreatus mycelia were accelerated under high-temperature stress.

A distinctive sequence motif in the fourth transmembrane domain confers ZIP13 iron function in Drosophila melanogaster.

The zinc/iron permease (ZIP/SLC39A) family plays an important role in metal ion transport and is essential for diverse physiological processes. Members of the ZIP family function primarily in the influx of transition metal ions zinc and iron, into cytoplasm from extracellular space or intracellular organelles. The molecular determinants defining metal ion selectivity among ZIP family members remain unclear. Specifically, we reported before that the Drosophila ZIP family member ZIP13 (dZIP13), functions as an iron exporter and was responsible for pumping iron into the secretory pathway. ZIP13 protein is unique in that it differs from the other LIV-1 subfamily members at transmembrane domain IV (TM4), wherein relative positions of the conserved H and D residues in the HNXXD sequence motif are switched, generating a DNXXH motif. In this study, we undertook an in vivo approach to explore the significance of this D/H exchange. Comparative functional analysis of mutants revealed that the relative positions of D and H are critical for the physiological roles of dZIP13 and its close homologue dZIP7. Swapping D/H position of this DNXXH sequence in dZIP13 resulted in loss of iron activity; normal dZIP13 could not complement dZIP7 loss, but swapping the two relative amino acid positions D and H in dZIP13 was sufficient to make it functionally analogous to its close homologue dZIP7. This work provides the first in vivo functional analysis of a structural motif required to differentiate different transporting functions of ZIPs.