A nomogram model for predicting the risk of axillary lymph node metastasis in patients with early breast cancer and cN0 status.

Axillary staging is commonly performed via sentinel lymph node biopsy for patients with early breast cancer (EBC) presenting with clinically negative axillary lymph nodes (cN0). The present study aimed to investigate the association between axillary lymph node metastasis (ALNM), clinicopathological characteristics of tumors and results from axillary ultrasound (US) scanning. Moreover, a nomogram model was developed to predict the risk for ALNM based on relevant factors. Data from 998 patients who met the inclusion criteria were retrospectively reviewed. These patients were then randomly divided into a training and validation group in a 7:3 ratio. In the training group, receiver operating characteristic curve analysis was used to identify the cutoff values for continuous measurement data. R software was used to identify independent ALNM risk variables in the training group using univariate and multivariate logistic regression analysis. The selected independent risk factors were incorporated into a nomogram. The model differentiation was assessed using the area under the curve (AUC), while calibration was evaluated through calibration charts and the Hosmer-Lemeshow test. To assess clinical applicability, a decision curve analysis (DCA) was conducted. Internal verification was performed via 1000 rounds of bootstrap resampling. Among the 998 patients with EBC, 228 (22.84%) developed ALNM. Multivariate logistic analysis identified lymphovascular invasion, axillary US findings, maximum diameter and molecular subtype as independent risk factors for ALNM. The Akaike Information Criterion served as the basis for both nomogram development and model selection. Robust differentiation was shown by the AUC values of 0.855 (95% CI, 0.817-0.892) and 0.793 (95% CI, 0.725-0.857) for the training and validation groups, respectively. The Hosmer-Lemeshow test yielded P-values of 0.869 and 0.847 for the training and validation groups, respectively, and the calibration chart aligned closely with the ideal curve, affirming excellent calibration. DCA showed that the net benefit from the nomogram significantly outweighed both the 'no intervention' and the 'full intervention' approaches, falling within the threshold probability interval of 12-97% for the training group and 17-82% for the validation group. This underscores the robust clinical utility of the model. A nomogram model was successfully constructed and validated to predict the risk of ALNM in patients with EBC and cN0 status. The model demonstrated favorable differentiation, calibration and clinical applicability, offering valuable guidance for assessing axillary lymph node status in this population.

Effect of exogenous melatonin on antioxidant properties and fruit softening of 'Fengtang' plum fruit (Prunus salicina Lindl.) during storage at room temperature.

'Fengtang' plums soften quickly and lose flavor after harvest. This study comprehensively evaluated the effect of exogenous melatonin on the fruit quality of 'Fengtang' plums. According to our findings, exogenous melatonin prevented plum fruit from losing water, delayed the decline in firmness, and preserved a high TSS/TA level. Additionally, exogenous melatonin also enhanced the activity of antioxidant enzymes and increased the non-enzymatic antioxidants, thereby further increasing the antioxidant capacity of plum fruit. Notably, exogenous melatonin delayed the degradation of covalent soluble pectin (CSP), cellulose, and hemicellulose, as well as the rise in water-soluble pectin (WSP) concentration and the activity of cell wall degrading enzymes. Further investigation using atomic force microscopy (AFM) revealed that the chain-like structure of ionic-soluble pectin (ISP) and the self-assembly network structures of CSP were depolymerized, and melatonin treatment retarded the depolymerization of pectin structures. Our results showed that exogenous melatonin preserved the postharvest quality of plum fruits by controlling fruit softness and antioxidant capacity during storage.

Physiological Mechanisms of Citrus Fruit Cracking: Study on Cell Wall Components, Osmoregulatory Substances, and Antioxidant Enzyme Activities.

Fruit cracking affects both the yield and economic efficiency of citrus; however, the underlying mechanism remains unclear. Therefore, this study focused on resistant and susceptible cultivars to identify the mechanisms underlying fruit cracking. The results showed that in 'Mingrijian', pectin morphological transformation and hemicellulose and lignin degradation in the pericarp were important contributing factors. During the critical fruit-cracking period (115-150 days after flowering), the water-soluble pectin, protopectin, and lignin contents in the pericarp of 'Daya' presented inverse changes relative to those in 'Mingrijian', thus enhancing the mechanical properties and resistance of pericarp. From 115 to 150 days after flowering, the soluble sugar content in the pulp of 'Mingrijian' increased rapidly by 97.35%, aiding in pulp water absorption and expansion. Moreover, the soluble protein content in the pericarp of 'Mingrijian' exhibited a declining trend and was lower than that of 'Daya', thus affecting the overall metabolism. The superoxide dismutase (SOD) activity in the pericarp of 'Mingrijian' gradually decreased from 115 to 180 days after flowering, while the peroxidase (POD) activity remained at a low level, resulting in weaker antioxidant capacity and lower environmental resistance. This study provides valuable insights into the mechanisms of citrus fruit cracking, laying the foundation for preventive and control strategies.

Interaction between dietary flavonoid intake and trouble sleeping on non-alcoholic fatty liver disease risk: a cross-sectional study.

OBJECTIVE: The possible interaction of dietary flavonoid intake and sleep on non-alcoholic fatty liver disease (NAFLD) has not been well studied. This study investigated the interaction between dietary flavonoid intake and trouble sleeping on the risk of NAFLD. METHODS: Three discrete National Health and Nutrition Examination Survey data cycles from 2007 to 2010 and 2017 to 2018 were used. NAFLD was diagnosed by a US Fatty Liver Index ≥30. A sleep questionnaire diagnosed trouble sleeping. Univariate and multivariate logistic regression, restricted cubic spline (RCS) and subgroup analyses were used to evaluate the association between dietary flavonoids, trouble sleeping and NAFLD. We employed the relative excess risk due to interaction, attributable proportion of interaction and synergy index to evaluate additive interactions. RESULTS: Ultimately, 5056 participants were enrolled, and higher anthocyanidins and flavanones intake was negatively correlated with NAFLD. Conversely, trouble sleeping was positively associated with NAFLD. These correlations remained stable after adjusting for confounders, and there was a sex difference in this relationship. In the RCS model, anthocyanins were negatively non-linearly related to NAFLD, while flavanones showed a negative linear relationship. Moreover, there was a synergistic interplay between low dietary anthocyanin intake and trouble sleeping on the risk of NAFLD. A similar relationship existed for flavanone intake. CONCLUSION: Anthocyanin and flavanone intake were negatively associated, whereas trouble sleeping was positively associated with NAFLD risk. There was a synergistic effect of low anthocyanin intake and trouble sleeping. The same relationship existed for low flavanone intake.

Circular RNA hsa_circ_0001846 facilitates the malignant behaviors of pancreatic cancer by sponging miR-204-3p and upregulating KRAS expression.

Pancreatic cancer (PC) is mainly derived from the exocrine pancreatic ductal epithelial cells, and it is strongly aggressive malignant tumor. Due to its insidious onset and the lack of effective diagnostic biomarkers, PC currently remains one of the main causes of cancer-related mortality worldwide. Recent studies have found that hsa_circ_0001846 is involved in the progression of multiple cancers and has the potential to become biomarkers, but its function and mechanism in PC remains unclear. We found by qRT-PCR experiments that hsa_circ_0001846 was upregulated in PC cells and tissues, while circBase, Sanger sequencing, agarose gel electrophoresis and FISH experiments identified the splicing site, ring structure and cellular localization of hsa_circ_0001846. Various functional experiments by using the construction of small interfering RNA targeting hsa_circ_0001846 and overexpression plasmid demonstrated that hsa_circ_0001846 promoted the proliferation, migration and invasion of PC cells. Moreover, the tumor weight and volume of nude mice were significantly reduced after the stable knockdown of hsa_circ_0001846. In the mechanism exploration, RNA pull-down experiments and dual-luciferase experiments helped us to determine that hsa_circ_0001846 regulated the KRAS expression by sponging miR-204-3p in PC, thus playing a pro-cancer role. In this study, the effect of miR-204-3p on PC was also explored for the first time, and we found that knockdown of miR-204-3p reversed the tumor suppressive effect caused by silencing hsa_circ_0001846, and silencing KRAS also rescued the pro-cancer effect caused by overexpression of hsa_circ_0001846. In conclusion, our study revealed the pro-cancer role of hsa_circ_0001846 in PC, and for the first time identified the mechanism that hsa_circ_0001846 regulated KRAS by sponging miR-204-3p to promote PC progression and had the potential to become a cancer biomarker.

Comprehensive analyses of the citrus WRKY gene family involved in the metabolism of fruit sugars and organic acids.

Sugars and organic acids are the main factors determining the flavor of citrus fruit. The WRKY transcription factor family plays a vital role in plant growth and development. However, there are still few studies about the regulation of citrus WRKY transcription factors (CsWRKYs) on sugars and organic acids in citrus fruit. In this work, a genome-wide analysis of CsWRKYs was carried out in the citrus genome, and a total of 81 CsWRKYs were identified, which contained conserved WRKY motifs. Cis-regulatory element analysis revealed that most of the CsWRKY promoters contained several kinds of hormone-responsive and abiotic-responsive cis-elements. Furthermore, gene expression analysis and fruit quality determination showed that multiple CsWRKYs were closely linked to fruit sugars and organic acids with the development of citrus fruit. Notably, transcriptome co-expression network analysis further indicated that three CsWRKYs, namely, CsWRKY3, CsWRKY47, and CsWRKY46, co-expressed with multiple genes involved in various pathways, such as Pyruvate metabolism and Citrate cycle. These CsWRKYs may participate in the metabolism of fruit sugars and organic acids by regulating carbohydrate metabolism genes in citrus fruit. These findings provide comprehensive knowledge of the CsWRKY family on the regulation of fruit quality.

Widely targeted metabolomic profiling combined with transcriptome analysis sheds light on flavonoid biosynthesis in sweet orange 'Newhall' (C. sinensis) under magnesium stress.

Sweet orange 'Newhall' peels (SOPs) are abundant in flavonoids, making them increasingly popular in the realms of nutrition, food, and medicine. However, there is still much unknown about flavonoid components in SOPs and the molecular mechanism of flavonoid biosynthesis when subjected to magnesium stress. The previous experiment conducted by the research group found that the total flavonoid content of Magnesium deficiency (MD) was higher than Magnesium sufficiency (MS) in SOPs. In order to study the metabolic pathway of flavonoids under magnesium stress, an integrative analysis of the metabolome and transcriptome was performed in SOPs at different developmental stages, comparing MS and MD. A comprehensive analysis revealed the identification of 1,533 secondary metabolites in SOPs. Among them, 740 flavonoids were classified into eight categories, with flavones identified as the dominant flavonoid component. The influence of magnesium stress on flavonoid composition was evaluated using a combination of heat map and volcanic map, which indicated significant variations between MS and MD varieties at different growth stages. The transcriptome detected 17,897 differential genes that were significantly enriched in flavonoid pathways. Further analysis was performed using Weighted gene correlation network analysis (WGCNA) in conjunction with flavonoid metabolism profiling and transcriptome analysis to identify six hub structural genes and ten hub transcription factor genes that play a crucial role in regulating flavonoid biosynthesis from yellow and blue modules. The correlation heatmap and Canonical Correspondence Analysis (CCA) results showed that CitCHS had a significant impact on the synthesis of flavones and other flavonoids in SOPs, as it was the backbone gene in the flavonoid biosynthesis pathway. The qPCR results further validated the accuracy of transcriptome data and the reliability of candidate genes. Overall, these results shed light on the composition of flavonoid compounds in SOPs and highlight the changes in flavonoid metabolism that occur under magnesium stress. This research provides valuable insights for improving the cultivation of high-flavonoid plants and enhancing our understanding of the molecular mechanisms underlying flavonoid biosynthesis.

Pathogenicity classification of SOD1 variants of uncertain significance by in vitro aggregation propensity.

Deposition of insoluble SOD1 aggregates in motor neurons is the hallmark of SOD1-associated ALS. Mutant SOD1 protein promotes structural instability that leads to misfolded SOD1 protein aggregates, which can be recapitulated in vitro. Therefore, aggregation propensity in cell lines can be a reliable indicator for the pathogenicity classification of SOD1 variants. Herein, we performed in vitro experiment to classify the pathogenicity of 34 SOD1 variants of uncertain significance (VUS) from 215 variants reported previously. The clinical features of 234 ALS patients with 31 SOD1 likely pathogenic (LP) variants were summarized. 31 VUS variants formed aggregates spontaneously, indicating LP variants. Missense variants were mainly located in the C-terminal of SOD1. Among patients with 31 SOD1 LP variants, 75% of patients had lower limb onset. The onset of familial ALS patients (45.7±14.0 years) is earlier than sporadic ALS patients (50.6±13.1 years). Our results expand the spectrum of SOD1 mutations and highlight the natural history of SOD1-positive ALS patients for further clinical trials in SOD1-related ALS.

Effects of Pollen Sources on Fruit Set and Fruit Characteristics of 'Fengtangli' Plum (Prunus salicina Lindl.) Based on Microscopic and Transcriptomic Analysis.

Adequate yield and fruit quality are required in commercial plum production. The pollen source has been shown to influence fruit set and fruit characteristics. In this study, 'Siyueli', 'Fenghuangli' and 'Yinhongli' were used as pollinizers of 'Fengtangli' plum. Additionally, self-pollination, mixed pollination, and open pollination were performed. We characterized the differences in pollen tube growth, fruit set and fruit quality among pollination combinations. 'Fengtangli' flowers pollinated by 'Fenghuangli' had more pistils with pollen tubes penetrating the ovary and the highest fruit set rate, while the lowest fruit set rate was obtained from self-pollination. In self-pollinated flowers, 33% of pistils had at least one pollen tube reaching the ovary, implying that 'Fengtangli' is partially self-compatible. Pollen sources affected 'Fengtangli' fruit size, weight, pulp thickness, soluble solids, and sugar content. Transcriptome analysis of 'Siyueli'-pollinated and 'Yinhongli'-pollinated fruits revealed 2762 and 1018 differentially expressed genes (DEGs) involved in the response to different pollen sources. DEGs were enriched in plant hormone signal transduction, starch and sucrose metabolism, and MAPK signaling pathways. Our findings provide a reference for the selection of suitable pollinizers for 'Fengtangli' plum and promote future research on the metaxenia effect at the molecular level.

Circular RNA hsa_circ_0007367 promotes the progression of pancreatic ductal adenocarcinoma by sponging miR-6820-3p and upregulating YAP1 expression.

Circular RNAs (circRNAs) play critical regulatory roles in cancer biological processes. Nevertheless, the contributions and underlying mechanisms of circRNAs to pancreatic ductal adenocarcinoma (PDAC) remain largely unexplored. Dysregulated circRNAs between cancerous tissues and matched adjacent normal tissues were identified by circRNA microarray in PDAC. The biological effect of hsa_circ_007367 both in vitro and in vivo was demonstrated by gain- and loss-of-function experiments. Further, dual-luciferase reporter and RNA pull-down assays were performed to confirm the interaction among hsa_circ_007367, miR-6820-3p, and Yes-associated protein 1 (YAP1). The expression of hsa_circ_007367 and YAP1 were detected by in situ hybridization (ISH) and immunohistochemistry (IHC) using tissue microarray (TMA) in 128 PDAC samples. We first identified that a novel circRNA, hsa_circ_0007367, was markedly upregulated in PDAC tissues and cells. Functionally, in vivo and in vitro data indicated that hsa_circ_0007367 promotes the proliferation and metastasis of PDAC. Mechanistically, we confirmed that hsa_circ_0007367 could facilitate the expression of YAP1, a well-known oncogene, by sponging miR-6820-3p, which function as a tumor suppresser in PDAC cells. The results of ISH and IHC demonstrated that hsa_circ_0007367 and YAP1 were upregulated in PDAC tissues. Furthermore, clinical data showed that higher hsa_circ_0007367 expression was correlated with advanced histological grade and lymph node metastasis in PDAC patients. In conclusion, our findings reveal that hsa_circ_0007367 acts as an oncogene via modulating miR-6820-3p/YAP1 axis to promote the progression of PDAC, and suggest that hsa_circ_0007367 may serve as a potential therapeutic target for treatment of PDAC.

SOD1 Mutation Spectrum and Natural History of ALS Patients in a 15-Year Cohort in Southeastern China.

Background: Mutations in superoxide dismutase 1 gene (SOD1) are the most frequent high penetrant genetic cause for amyotrophic lateral sclerosis (ALS) in the Chinese population. A detailed natural history of SOD1-mutated ALS patients will provide key information for ongoing genetic clinical trials. Methods: We screened for SOD1 mutations using whole exome sequencing (WES) in Chinese ALS cases from 2017 to 2021. Functional studies were then performed to confirm the pathogenicity of novel variants. In addition, we enrolled previously reported SOD1 mutations in our centers from 2007 to 2017. The SOD1 mutation spectrum, age at onset (AAO), diagnostic delay, and survival duration were analyzed. Results: We found two novel SOD1 variants (p.G17H and p.E134*) that exerted both gain-of-function and loss-of-function effects in vitro. Combined with our previous SOD1-mutated patients, 32 probands with 21 SOD1 mutations were included with the four most frequently occurring mutations of p.V48A, p.H47R, p.C112Y, and p.G148D. SOD1 mutations account for 58.9% of familial ALS (FALS) cases. The mean (SD) AAO was 46 ± 11.4 years with a significant difference between patients carrying mutations in exon 1 [n = 5, 34.6 (12.4) years] and exon 2 [n = 8, 51.4 (8.2) years] (p = 0.038). The mean of the diagnostic delay of FALS patients is significantly earlier than the sporadic ALS (SALS) patients [9.5 (4.8) vs. 20.3 (9.3) years, p = 0.0026]. In addition, male patients survived longer than female patients (40 vs. 16 months, p = 0.05). Conclusion: Our results expanded the spectrum of SOD1 mutations, highlighted the mutation distribution, and summarized the natural history of SOD1-mutated patients in southeastern China. Male patients were found to have better survival, and FALS patients received an earlier diagnosis. Our findings assist in providing a detailed clinical picture, which is important for ongoing genetic clinical trials.

Transcriptome analysis and functional identification of GmMYB46 in soybean seedlings under salt stress.

Salinity is one of the major abiotic stress that limits crop growth and productivity. We investigated the transcriptomes of salt-treated soybean seedlings versus a control using RNA-seq to better understand the molecular mechanisms of the soybean (Glycine max L.) response to salt stress. Transcriptome analysis revealed 1,235 differentially expressed genes (DEGs) under salt stress. Several important pathways and key candidate genes were identified by KEGG enrichment. A total of 116 differentially expressed transcription factors (TFs) were identified, and 17 TFs were found to belong to MYB families. Phylogenetic analysis revealed that these TFs may be involved in salt stress adaptation. Further analysis revealed that GmMYB46 was up-regulated by salt and mannitol and was localized in the nucleus. The salt tolerance of transgenic Arabidopsis overexpressing GmMYB46 was significantly enhanced compared to wild-type (WT). GmMYB46 activates the expression of salt stress response genes (P5CS1, SOD, POD, NCED3) in Arabidopsis under salt stress, indicating that the GmMYB46 protein mediates the salt stress response through complex regulatory mechanisms. This study provides information with which to better understand the molecular mechanism of salt tolerance in soybeans and to genetically improve the crop.

Clinical and genetic characteristics of Chinese patients with reducing body myopathy.

Reducing body myopathy (RBM) is a rare myopathy characterized by reducing bodies (RBs) in morphological presentation. The clinical manifestations of RBM present a wide clinical spectrum, varying from infantile lethal form through childhood and adult benign forms. FHL1 gene is the causative gene of RBM. To date, only 6 Chinese RBM patients have been reported. Here, we reported the clinical presentations and genetic findings of 3 Chinese RBM patients from two families. Two novel pathogenic variants, c.395G>A and c.401_402insGAC, were identified by whole exome sequencing. Furthermore, by reviewing previous studies, we revealed that most RBM patients manifested with an early onset, symmetric, progressive limb-girdle and axial muscle weakness with joint contractures, rigid spine or scoliosis except familial female patients who exhibited asymmetric benign muscle involvements. Our results provide insightful information to help better diagnose and understand the disease.