Single-cell analysis of Crohn's disease: Unveiling heterogeneity and evaluating ustekinumab outcomes.

BACKGROUND: The present study aimed to dissect the cellular complexity of Crohn's disease (CD) using single-cell RNA sequencing, focusing on identifying key cell populations and their transcriptional profiles in inflamed tissue. METHODS: We applied scRNA-sequencing to compare the cellular composition of CD patients with healthy controls, utilizing Seurat for clustering and annotation. Differential gene expression analysis and protein-protein interaction networks were constructed to identify crucial genes and pathways. RESULTS: Our study identified eight distinct cell types in CD, highlighting crucial fibroblast and T cell interactions. The analysis revealed key cellular communications and identified significant genes and pathways involved in the disease's pathology. The role of fibroblasts was underscored by elevated expression in diseased samples, offering insights into disease mechanisms and potential therapeutic targets, including responses to ustekinumab treatment, thus enriching our understanding of CD at a molecular level. CONCLUSIONS: Our findings highlight the complex cellular and molecular interplay in CD, suggesting new biomarkers and therapeutic targets, offering insights into disease mechanisms and treatment implications.

TMPRSS2 and glycan receptors synergistically facilitate coronavirus entry.

The entry of coronaviruses is initiated by spike recognition of host cellular receptors, involving proteinaceous and/or glycan receptors. Recently, TMPRSS2 was identified as the proteinaceous receptor for HCoV-HKU1 alongside sialoglycan as a glycan receptor. However, the underlying mechanisms for viral entry remain unknown. Here, we investigated the HCoV-HKU1C spike in the inactive, glycan-activated, and functionally anchored states, revealing that sialoglycan binding induces a conformational change of the NTD and promotes the neighboring RBD of the spike to open for TMPRSS2 recognition, exhibiting a synergistic mechanism for the entry of HCoV-HKU1. The RBD of HCoV-HKU1 features an insertion subdomain that recognizes TMPRSS2 through three previously undiscovered interfaces. Furthermore, structural investigation of HCoV-HKU1A in combination with mutagenesis and binding assays confirms a conserved receptor recognition pattern adopted by HCoV-HKU1. These studies advance our understanding of the complex viral-host interactions during entry, laying the groundwork for developing new therapeutics against coronavirus-associated diseases.

Automatic soft-tissue analysis on orthodontic frontal and lateral facial photographs based on deep learning.

BACKGROUND: To establish the automatic soft-tissue analysis model based on deep learning that performs landmark detection and measurement calculations on orthodontic facial photographs to achieve a more comprehensive quantitative evaluation of soft tissues. METHODS: A total of 578 frontal photographs and 450 lateral photographs of orthodontic patients were collected to construct datasets. All images were manually annotated by two orthodontists with 43 frontal-image landmarks and 17 lateral-image landmarks. Automatic landmark detection models were established, which consisted of a high-resolution network, a feature fusion module based on depthwise separable convolution, and a prediction model based on pixel shuffle. Ten measurements for frontal images and eight measurements for lateral images were defined. Test sets were used to evaluate the model performance, respectively. The mean radial error of landmarks and measurement error were calculated and statistically analysed to evaluate their reliability. RESULTS: The mean radial error was 14.44 ± 17.20 pixels for the landmarks in the frontal images and 13.48 ± 17.12 pixels for the landmarks in the lateral images. There was no statistically significant difference between the model prediction and manual annotation measurements except for the mid facial-lower facial height index. A total of 14 measurements had a high consistency. CONCLUSION: Based on deep learning, we established automatic soft-tissue analysis models for orthodontic facial photographs that can automatically detect 43 frontal-image landmarks and 17 lateral-image landmarks while performing comprehensive soft-tissue measurements. The models can assist orthodontists in efficient and accurate quantitative soft-tissue evaluation for clinical application.

Tumor cell-intrinsic MELK enhanced CCL2-dependent immunosuppression to exacerbate hepatocarcinogenesis and confer resistance of HCC to radiotherapy.

BACKGROUND: The outcome of hepatocellular carcinoma (HCC) is limited by its complex molecular characteristics and changeable tumor microenvironment (TME). Here we focused on elucidating the functional consequences of Maternal embryonic leucine zipper kinase (MELK) in the tumorigenesis, progression and metastasis of HCC, and exploring the effect of MELK on immune cell regulation in the TME, meanwhile clarifying the corresponding signaling networks. METHODS: Bioinformatic analysis was used to validate the prognostic value of MELK for HCC. Murine xenograft assays and HCC lung metastasis mouse model confirmed the role of MELK in tumorigenesis and metastasis in HCC. Luciferase assays, RNA sequencing, immunopurification-mass spectrometry (IP-MS) and coimmunoprecipitation (CoIP) were applied to explore the upstream regulators, downstream essential molecules and corresponding mechanisms of MELK in HCC. RESULTS: We confirmed MELK to be a reliable prognostic factor of HCC and identified MELK as an effective candidate in facilitating the tumorigenesis, progression, and metastasis of HCC; the effects of MELK depended on the targeted regulation of the upstream factor miR-505-3p and interaction with STAT3, which induced STAT3 phosphorylation and increased the expression of its target gene CCL2 in HCC. In addition, we confirmed that tumor cell-intrinsic MELK inhibition is beneficial in stimulating M1 macrophage polarization, hindering M2 macrophage polarization and inducing CD8 + T-cell recruitment, which are dependent on the alteration of CCL2 expression. Importantly, MELK inhibition amplified RT-related immune effects, thereby synergizing with RT to exert substantial antitumor effects. OTS167, an inhibitor of MELK, was also proven to effectively impair the growth and progression of HCC and exert a superior antitumor effect in combination with radiotherapy (RT). CONCLUSIONS: Altogether, our findings highlight the functional role of MELK as a promising target in molecular therapy and in the combination of RT therapy to improve antitumor effect for HCC.

A bispecific antibody exhibits broad neutralization against SARS-CoV-2 Omicron variants XBB.1.16, BQ.1.1 and sarbecoviruses.

The Omicron subvariants BQ.1.1, XBB.1.5, and XBB.1.16 of SARS-CoV-2 are known for their adeptness at evading immune responses. Here, we isolate a neutralizing antibody, 7F3, with the capacity to neutralize all tested SARS-CoV-2 variants, including BQ.1.1, XBB.1.5, and XBB.1.16. 7F3 targets the receptor-binding motif (RBM) region and exhibits broad binding to a panel of 37 RBD mutant proteins. We develop the IgG-like bispecific antibody G7-Fc using 7F3 and the cross-neutralizing antibody GW01. G7-Fc demonstrates robust neutralizing activity against all 28 tested SARS-CoV-2 variants and sarbecoviruses, providing potent prophylaxis and therapeutic efficacy against XBB.1 infection in both K18-ACE and BALB/c female mice. Cryo-EM structure analysis of the G7-Fc in complex with the Omicron XBB spike (S) trimer reveals a trimer-dimer conformation, with G7-Fc synergistically targeting two distinct RBD epitopes and blocking ACE2 binding. Comparative analysis of 7F3 and LY-CoV1404 epitopes highlights a distinct and highly conserved epitope in the RBM region bound by 7F3, facilitating neutralization of the immune-evasive Omicron variant XBB.1.16. G7-Fc holds promise as a potential prophylactic countermeasure against SARS-CoV-2, particularly against circulating and emerging variants.

Effects of myricetin and its derivatives on nonenzymatic glycation: A mechanism study based on proteomic modification and fluorescence spectroscopy analysis.

Myricetin and its derivatives, myricitrin and dihydromyricetin, are flavonoids widely presented in foods and phytomedicine that possess tremendous health potential. In this study, we compared the antiglycation activity of myricetin and its derivatives, then investigated the underlying mechanism using proteomic modification and fluorescence spectroscopy analysis. All three compounds exhibited thorough inhibition on nonenzymatic glycation process, with the inhibitory effects on AGEs reaching 85% at 40 µmol/L. They effectively protected bovine serum albumin (BSA) structure by inhibiting protein oxidation, preventing the conversion from α-helix to ß-sheet, and reducing amyloid-like cross-ß structure formation. Among the three compounds, myricetin showed a predominant antiglycation activity. Proteomic analysis identified the early glycated sites that were protected by myricetin, including lysine K235, 256, 336, 421, 420, 489, etc. Additionally, fluorescence spectroscopy revealed spontaneous interactions between BSA and myricetin. Overall, myricetin holds promise as an antiglycation agent in both the food and drug industries.

Exosomal circSIPA1L3-mediated intercellular communication contributes to glucose metabolic reprogramming and progression of triple negative breast cancer.

BACKGROUND: Breast cancer is the most common malignant tumor, and metastasis remains the major cause of poor prognosis. Glucose metabolic reprogramming is one of the prominent hallmarks in cancer, providing nutrients and energy to support dramatically elevated tumor growth and metastasis. Nevertheless, the potential mechanistic links between glycolysis and breast cancer progression have not been thoroughly elucidated. METHODS: RNA-seq analysis was used to identify glucose metabolism-related circRNAs. The expression of circSIPA1L3 in breast cancer tissues and serum was examined by qRT-PCR, and further assessed its diagnostic value. We also evaluated the prognostic potential of circSIPA1L3 by analyzing a cohort of 238 breast cancer patients. Gain- and loss-of-function experiments, transcriptomic analysis, and molecular biology experiments were conducted to explore the biological function and regulatory mechanism of circSIPA1L3. RESULTS: Using RNA-seq analysis, circSIPA1L3 was identified as the critical mediator responsible for metabolic adaption upon energy stress. Gain- and loss-of-function experiments revealed that circSIPA1L3 exerted a stimulative effect on breast cancer progression and glycolysis, which could also be transported by exosomes and facilitated malignant behaviors among breast cancer cells. Significantly, the elevated lactate secretion caused by circSIPA1L3-mediated glycolysis enhancement promoted the recruitment of tumor associated macrophage and their tumor-promoting roles. Mechanistically, EIF4A3 induced the cyclization and cytoplasmic export of circSIPA1L3, which inhibited ubiquitin-mediated IGF2BP3 degradation through enhancing the UPS7-IGF2BP3 interaction. Furthermore, circSIPA1L3 increased mRNA stability of the lactate export carrier SLC16A1 and the glucose intake enhancer RAB11A through either strengthening their interaction with IGF2BP3 or sponging miR-665, leading to enhanced glycolytic metabolism. Clinically, elevated circSIPA1L3 expression indicated unfavorable prognosis base on the cohort of 238 breast cancer patients. Moreover, circSIPA1L3 was highly expressed in the serum of breast cancer patients and exhibited high diagnostic value for breast cancer patients. CONCLUSIONS: Our study highlights the oncogenic role of circSIPA1L3 through mediating glucose metabolism, which might serve as a promising diagnostic and prognostic biomarker and potential therapeutic target for breast cancer.

Steam explosion-assisted grinding improves the functional properties and antioxidant activity of Java tea-leaf powders (Clerodendranthus spicatus).

BACKGROUND: Java tea is widely consumed and has multiple health effects. This study established a steam explosion (SE) pretreatment method to prepare Java tea-leaf powders. The physicochemical, functional properties, phenolic extraction, and antioxidant activity of Java tea-leaf powders produced by simple and SE-assisted milling methods were investigated. RESULTS: In comparison with simple milling, SE pretreatment broke the cell wall effectively and reduced the particle size of Java tea-leaf powders. Steam explosion-treated powders showed higher values for sensory signals, bulk and tap density, and for the water solubility index. After SE treatment, the adsorption capacities to glucose, soybean oil, and cholesterol of leaf powders were increased by up to 55, 95, and 80% respectively. The extracts from SE-treated powders also showed higher total polyphenol content and antioxidant activity. CONCLUSION: Steam explosion treatment is helpful for the improvement of functional properties and antioxidant activity, which can benefit the development and application of Java tea-leaf powders. © 2024 Society of Chemical Industry.

Functional Characterization of the MeSSIII-1 Gene and Its Promoter from Cassava.

Soluble starch synthases (SSs) play important roles in the synthesis of cassava starch. However, the expression characteristics of the cassava SSs genes have not been elucidated. In this study, the MeSSIII-1 gene and its promoter, from SC8 cassava cultivars, were respectively isolated by PCR amplification. MeSSIII-1 protein was localized to the chloroplasts. qRT-PCR analysis revealed that the MeSSIII-1 gene was expressed in almost all tissues tested, and the expression in mature leaves was 18.9 times more than that in tuber roots. MeSSIII-1 expression was induced by methyljasmonate (MeJA), abscisic acid (ABA), and ethylene (ET) hormones in cassava. MeSSIII-1 expression patterns were further confirmed in proMeSSIII-1 transgenic cassava. The promoter deletion analysis showed that the -264 bp to -1 bp MeSSIII-1 promoter has basal activity. The range from -1228 bp to -987 bp and -488 bp to -264 bp significantly enhance promoter activity. The regions from -987 bp to -747 bp and -747 bp to -488 bp have repressive activity. These findings will provide an important reference for research on the potential function and transcriptional regulation mechanisms of the MeSSIII-1 gene and for further in-depth exploration of the regulatory network of its internal functional elements.

[Stellera chamaejasme against multidrug resistance of triple-negative breast cancer MDA-MB-231 cell through Nrf2].

This study aims to investigate the effect and mechanism of Stellera chamaejasme extract(SCL) on multidrug resistance(MDR) in breast cancer. Human triple-negative breast cancer cell line MDA-MB-231 and its adriamycin-resistant cell line MDA-MB-231/ADR were used in the experiment. Cell viability was detected by methyl thiazolyl tetrazolium(MTT) assay, and cell apoptosis was detected by DAPI staining and Annexin-V/Pi double staining. Western blot(WB) was used to detect the expression levels of Keap1, Nrf2, HO-1, Bcl-2, Bax, caspase-9, and caspase-3. Immunofluorescence staining was used to observe the distribution of Nrf2 in the cell, and flow cytometry was used to detect the level of reactive oxygen species(ROS) in the cell. The results showed that the resis-tance factor of SCL was 0.69, and that of adriamycin and paclitaxel was 8.40 and 16.36, respectively. DAPI staining showed that SCL could cause nuclear shrinkage and fragmentation of breast cancer cells. Annexin-V/Pi double staining showed that the average apoptosis rate of the drug-resistant cells was 32.64% and 50.29%, respectively under medium and high doses of SCL. WB results showed that SCL could significantly reduce the expression levels of anti-apoptotic proteins Bcl-2, caspase-9, and caspase-3 and significantly increase the expression level of pro-apoptotic protein Bax. Further studies showed that SCL could significantly promote the expression of Keap1, significantly inhibit the expression of Nrf2 and HO-1, and significantly reduce the expression level of Nrf2 in the nucleus. Correspondingly, flow cytometry showed that the intracellular ROS level was significantly increased. In conclusion, SCL can significantly inhibit the proliferation of MDA-MB-231 multidrug-resistant cells of triple-negative breast cancer and cause cell apoptosis, and the mechanism is related to inhibiting Keap1/Nrf2 signaling pathway, leading to ROS accumulation in drug-resistant cells and increasing the expression of apoptosis-related proteins.

MeGT2.6 increases cellulose synthesis and active gibberellin content to promote cell enlargement in cassava.

Cassava, a pivotal tropical crop, exhibits rapid growth and possesses a substantial biomass. Its stem is rich in cellulose and serves as a crucial carbohydrate storage organ. The height and strength of stems restrict the mechanised operation and propagation of cassava. In this study, the triple helix transcription factor MeGT2.6 was identified through yeast one-hybrid assay using MeCesA1pro as bait, which is critical for cellulose synthesis. Over-expression and loss-of-function lines were generated, and results revealed that MeGT2.6 could promote a significant increase in the plant height, stem diameter, cell size and thickness of SCW of cassava plant. Specifically, MeGT2.6 upregulated the transcription activity of MeGA20ox1 and downregulated the expression level of MeGA2ox1, thereby enhancing the content of active GA3, resulting in a large cell size, high plant height and long stem diameter in cassava. Moreover, MeGT2.6 upregulated the transcription activity of MeCesA1, which promoted the synthesis of cellulose and hemicellulose and produced a thick secondary cell wall. Finally, MeGT2.6 could help supply additional substrates for the synthesis of cellulose and hemicellulose by upregulating the invertase genes (MeNINV1/6). Thus, MeGT2.6 was found to be a multiple regulator; it was involved in GA metabolism and sucrose decomposition and the synthesis of cellulose and hemicellulose.

Plasticity in resource allocation of the invasive Phytolacca americana: Balancing growth, reproduction, and defense along urban-rural gradients.

In response to varying environments along urban and rural gradients, invasive plants may strategically allocate resources to enhance their invasiveness. However, how invasive plants balance their resources for growth, reproduction, and defense as responses to biotic and abiotic factors across these gradients remain unclear. We conducted field surveys on the growth, reproduction, and herbivory of the invasive species Phytolacca americana across diverse urban and rural habitats. Leaf samples were collected to analyze the nutritional content, primary and secondary metabolites. We found that plant growth rates, specific leaf area, leaf nitrogen content, and concentrations of flavonoids and saponins were higher in urban habitats, while reproduction, herbivory, and carbon-to­nitrogen ratios were lower than those in rural habitats. We also found a trade-off between growth rate and herbivory, as well as trade-offs among defense traits associated with herbivory (e.g., leaf mass per area, the inverse of leaf nitrogen content, and carbon­nitrogen ratio) and the production of metabolites associated with abiotic stress tolerance (e.g., soluble sugars, flavonoids, and saponins). As earlier studies showed low levels of genetic diversity within and between populations, our findings suggest that the urban-rural gradient patterns of resource allocation are primarily phenotypic plasticity in response to herbivory in rural areas and abiotic factors in urban areas. Our study sheds light on the mechanisms by which urbanization affects plant invasions and offers insights for the implementation of their management strategies.

Hepatic kynurenic acid mediates phosphorylation of Nogo-A in the medial prefrontal cortex to regulate chronic stress-induced anxiety-like behaviors in mice.

Exercise training effectively relieves anxiety disorders via modulating specific brain networks. The role of post-translational modification of proteins in this process, however, has been underappreciated. Here we performed a mouse study in which chronic restraint stress-induced anxiety-like behaviors can be attenuated by 14-day persistent treadmill exercise, in association with dramatic changes of protein phosphorylation patterns in the medial prefrontal cortex (mPFC). In particular, exercise was proposed to modulate the phosphorylation of Nogo-A protein, which drives the ras homolog family member A (RhoA)/ Rho-associated coiled-coil-containing protein kinases 1(ROCK1) signaling cascade. Further mechanistic studies found that liver-derived kynurenic acid (KYNA) can affect the kynurenine metabolism within the mPFC, to modulate this RhoA/ROCK1 pathway for conferring stress resilience. In sum, we proposed that circulating KYNA might mediate stress-induced anxiety-like behaviors via protein phosphorylation modification within the mPFC, and these findings shed more insights for the liver-brain communications in responding to both stress and physical exercise.