Molecular landscape and multi-omic measurements of heterogeneity in fetal adenocarcinoma of the lung.

Fetal adenocarcinoma of the lung (FLAC) is a rare form of lung adenocarcinoma and was divided into high-grade (H-FLAC) and low-grade (L-FLAC) subtypes. Despite the existence of some small case series studies, a comprehensive multi-omics study of FLAC has yet to be undertaken. In this study, we depicted the multi-omics landscapes of this rare lung cancer type by performing multi-regional sampling on 20 FLAC cases. A comparison of multi-omics profiles revealed significant differences between H-FLAC and L-FLAC in a multi-omic landscape. Two subtypes also showed distinct relationships between multi-layer intratumor heterogeneity (ITH). We discovered that a lower genetic ITH was significantly associated with worse recurrence-free survival and overall survival in FLAC patients, whereas higher methylation ITH in H-FLAC patients suggested a short survival. Our findings highlight the complex interplay between genetic and transcriptional heterogeneity in FLAC and suggest that different types of ITH may have distinct implications for patient prognosis.

Multi-dimensional cell-free DNA-based liquid biopsy for sensitive early detection of gastric cancer.

BACKGROUND: Gastric cancer is the fifth most common cancer type. Most patients are diagnosed at advanced stages with poor prognosis. A non-invasive assay for the detection of early-stage gastric cancer is highly desirable for reducing associated mortality. METHODS: We collected a prospective study cohort of 110 stage I-II gastric cancer patients and 139 non-cancer individuals. We performed whole-genome sequencing with plasma samples and profiled four types of cell-free DNA (cfDNA) characteristics, fragment size pattern, copy number variation, nucleosome coverage pattern, and single nucleotide substitution. With these differential profiles, we developed an ensemble model to detect gastric cancer signals. Further, we validated the assay in an in-house first validation cohort of 73 gastric cancer patients and 94 non-cancer individuals and an independent second validation cohort of 47 gastric cancer patients and 49 non-cancer individuals. Additionally, we evaluated the assay in a hypothetical 100,000 screening population by Monte Carlo simulation. RESULTS: Our cfDNA-based assay could distinguish early-stage gastric cancer from non-cancer at an AUROC of 0.962 (95% CI: 0.942-0.982) in the study cohort, 0.972 (95% CI: 0.953-0.992) in the first validation cohort and 0.937 (95% CI: 0.890-0.983) in the second validation cohort. The model reached a specificity of 92.1% (128/139) and a sensitivity of 88.2% (97/110) in the study cohort. In the first validation cohort, 91.5% (86/94) of non-cancer individuals and 91.8% (67/73) of gastric cancer patients were correctly identified. In the second validation cohort, 89.8% (44/49) of non-cancer individuals and 87.2% (41/47) of gastric cancer patients were accurately classified. CONCLUSIONS: We introduced a liquid biopsy assay using multiple dimensions of cfDNA characteristics that could accurately identify early-stage gastric cancer from non-cancerous conditions. As a cost-effective non-invasive approach, it may provide population-wide benefits for the early detection of gastric cancer. TRIAL REGISTRATION: This study was registered on ClinicalTrials.gov under the identifier NCT05269056 on March 7, 2022.

YTHDF1 promotes the osteolytic bone metastasis of breast cancer via inducing EZH2 and CDH11 translation.

Bone metastasis is common in breast cancer and more effective therapies are required, however, its molecular mechanism is poorly understood. Additionally, the role of the m6A reader YTHDF1 in bone metastasis of breast cancer has not been reported. Here, we reveal that the increased expression of YTHDF1 is clinically correlated with breast cancer bone metastases. YTHDF1 promotes migration, invasion, and osteoblast adhesion and induces osteoclast differentiation of cancer cells in vitro and vivo. Mechanically, RNA-seq, MeRIP-seq and RIP-seq analysis, and molecular biology experiments demonstrate that YTHDF1 translationally enhances EZH2 and CDH11 expression by reading m6A-enriched sites of their transcripts. Moreover, adeno-associated virus (AAV) was used to deliver shYTHDF1 (shYTHDF1-AAV) in intratibial injection models, eliciting a significant suppressive effect on breast cancer bone metastatic formation and osteolytic destruction. Overall, we uncovered that YTHDF1 promotes osteolytic bone metastases of breast cancer by inducing EZH2 and CDH11 translation.

Clinical implications of CSF-ctDNA positivity in newly diagnosed diffuse large B cell lymphoma.

The clinical implications of CSF-ctDNA positivity in newly diagnosed diffuse large B cell lymphoma (ND-DLBCL) remains largely unexplored. One hundred ND-DLBCL patients were consecutively enrolled as training cohort and another 26 ND-DLBCL patients were prospectively enrolled in validation cohort. CSF-ctDNA positivity (CSF(+)) was identified in 25 patients (25.0%) in the training cohort and 7 patients (26.9%) in the validation cohort, extremely higher than CNS involvement rate detected by conventional methods. Patients with mutations of CARD11, JAK2, ID3, and PLCG2 were more predominant with CSF(+) while FAT4 mutations were negatively correlated with CSF(+). The downregulation of PI3K-AKT signaling, focal adhesion, actin cytoskeleton, and tight junction pathways were enriched in CSF(+) ND-DLBCL. Furthermore, pretreatment CSF(+) was significantly associated with poor outcomes. Three risk factors, including high CSF protein level, high plasma ctDNA burden, and involvement of high-risk sites were used to predict the risk of CSF(+) in ND-DLBCL. The sensitivity and specificity of pretreatment CSF-ctDNA to predict CNS relapse were 100% and 77.3%. Taken together, we firstly present the prevalence and the genomic and transcriptomic landscape for CSF-ctDNA(+) DLBCL and highlight the importance of CSF-ctDNA as a noninvasive biomarker in detecting and monitoring of CSF infiltration and predicting CNS relapse in DLBCL.

Enrichment of oligodendrocyte precursor phenotypes in subsets of low-grade glioneuronal tumours.

Current histological classification of low-grade glioneuronal tumours does not adequately represent their underlying biology. The neural lineage(s) and differentiation stage(s) involved and the cell state(s) affected by the recurrent genomic alterations are unclear. Here, we describe dysregulated oligodendrocyte lineage developmental programmes in three low-grade glioneuronal tumour subtypes. Ten dysembryoplastic neuroepithelial tumours, four myxoid glioneuronal tumours and five rosette-forming glioneuronal tumours were collected. Besides a comprehensive characterization of clinical features, known diagnostic markers and genomic alterations, we used comprehensive immunohistochemical stainings to characterize activation of rat sarcoma/mitogen-activated protein kinase pathway, involvement of neuronal component, resemblance to glial lineages and differentiation blockage along the stages of oligodendrocyte lineage. The findings were further complemented by gene set enrichment analysis with transcriptome data of dysembryoplastic neuroepithelial tumours from the literature. Dysembryoplastic neuroepithelial tumours, myxoid glioneuronal tumours and rosette-forming glioneuronal tumours occur at different ages, with symptoms closely related to tumour location. Dysembryoplastic neuroepithelial tumours and myxoid glioneuronal tumours contain oligodendrocyte-like cells and neuronal component. Rosette-forming glioneuronal tumours contained regions of rosette-forming neurocytic and astrocytic features. Scattered neurons, identified by neuronal nuclei antigen and microtubule-associated protein-2 staining, were consistently observed in all dysembryoplastic neuroepithelial tumours and myxoid glioneuronal tumours examined, but only in one rosette-forming glioneuronal tumour. Pervasive neurofilament-positive axons were observed only in dysembryoplastic neuroepithelial tumour and myxoid glioneuronal tumour samples. Alterations in B-Raf proto-oncogene, serine/threonine kinase, fibroblast growth factor receptor 1, fibroblast growth factor receptor 3 and platelet-derived growth factor receptor alpha occurred in a mutually exclusive manner, coinciding with strong staining of phospho-p44/42 mitogen-activated protein kinase and low apoptotic signal. All dysembryoplastic neuroepithelial tumours, myxoid glioneuronal tumours and the neurocytic regions of rosette-forming glioneuronal tumours showed strong expression of neuron-glia antigen 2, platelet-derived growth factor receptor alpha (markers of oligodendrocyte precursor cells) and neurite outgrowth inhibitor-A (a marker of developing oligodendrocytes), but lacked the expression of oligodendrocyte markers ectonucleotide pyrophosphatase/phosphodiesterase family member 6 and myelin basic protein. Notably, transcriptomes of dysembryoplastic neuroepithelial tumours were enriched in oligodendrocyte precursor cell signature, but not in signatures of neural stem cells, myelinating oligodendrocytes and astrocytes. Dysembryoplastic neuroepithelial tumour, myxoid glioneuronal tumour and rosette-forming glioneuronal tumour resemble oligodendrocyte precursor cells, and their enrichment of oligodendrocyte precursor cell phenotypes is closely associated with the recurrent mutations in rat sarcoma/mitogen-activated protein kinase pathway.

Molecular characteristics of early-onset compared with late-onset colorectal cancer: A case controlled study.

BACKGROUND: Early-Onset Colorectal Cancer (EOCRC) is associated with a poorer prognosis relative to Late-Onset Colorectal Cancer (LOCRC), and its incidence has witnessed a gradual escalation in recent years. This necessitates a comprehensive examination of the underlying pathogenesis and the identification of therapeutic targets specific to EOCRC patients. The present study aimed to delineate the distinct molecular landscape of EOCRC by juxtaposing it with that of LOCRC. METHODS: A total of 11,344 colorectal cancer patients, diagnosed between 2003 and 2022, were enrolled in this study, comprising 578 EOCRC cases and 10,766 LOCRC cases. Next-generation sequencing technology was employed to assess the tumor-related mutation and tumor mutation burden (TMB) in these patients. PD-L1 expression was quantified using immunohistochemistry. Microsatellite instability (MSI) was determined via capillary electrophoresis (2B3D NCI Panel). RESULTS: Upon comparing LOCRC with EOCRC patients, the latter group demonstrated a tendency towards advanced TNM stage, lower tumor differentiation, and less favorable histological types. Among LOCRC patients, those with MSI-H status were found to have an earlier TNM stage compared to those with MSI-L/MSS status. Significantly, the incidence of MSI-H was notably higher in EOCRC (10.2%) compared to LOCRC (2.2%). Mutations in the 7-gene panel (ARID1A, FANCI, CASP8, DGFRA, DPYD, TSHR, and PRKCI) were more prevalent in LOCRC. Within the EOCRC cohort, patients with the MSI-H subtype displayed an earlier TNM stage but concurrently exhibited poorer tissue differentiation and a higher frequency of mucinous adenocarcinoma. Among EOCRC patients, FBXW7, FAT1, ATM, ARID1A, and KMT2B mutations were significantly enriched in the MSI-H subgroup. A comparative analysis of MSI-H patients revealed heightened mutation frequencies of FGFBR2, PBRM1, RNF43, LRP1B, FBXW7, ATM, and ARID1A in the EOCRC group. Furthermore, EOCRC patients demonstrated a higher overall TMB, particularly in the MSI-H subtype. PD-L1 expression was elevated in EOCRC and positively associated with MSI status. CONCLUSIONS: This study revealed a significantly higher MSI-H distribution rate in early-onset colorectal cancer, and EOCRC exhibits a distinct mutational signature coupled with higher PD-L1 expression. These findings hold promise in guiding personalized therapeutic strategies for improved disease management in EOCRC patients.

Prediction model of deep vein thrombosis risk after lower extremity orthopedic surgery.

Purpose: This investigation was conceived to engineer and appraise a pioneering clinical nomogram, crafted to bridge the extant chasm in literature regarding the postoperative risk stratification for deep vein thrombosis (DVT) in the aftermath of lower extremity orthopedic procedures. This novel tool offers a sophisticated and discerning algorithm for risk prediction, heretofore unmet by existing methodologies. Methods: In this retrospective observational study, clinical records of hospitalized patients who underwent lower extremity orthopedic surgery were collected at the Wuxi TCM Hospital Affiliated to the Nanjing University of Chinese Medicine between Jan 2017 and Oct 2019. The univariate and multivariate analysis with the backward stepwise method was applied to select features for the predictive nomogram. The performance of the nomogram was evaluated with respect to its discriminant capability, calibration ability, and clinical utility. Result: A total of 5773 in-hospital patients were eligible for the study, with the incidence of deep vein thrombosis being approximately 1 % in this population. Among 31 variables included, 5 of them were identified to be the predictive features in the nomogram, including age, mean corpuscular hemoglobin concentration (MCHC), D-dimer, platelet distribution width (PDW), and thrombin time (TT). The area under the receiver operating characteristic (ROC) curve in the training and validation cohort was 85.9 % (95%CI: 79.96 %-90.04 %) and 85.7 % (95%CI: 78.96 %-90.69 %), respectively. Both the calibration curves and decision curve analysis demonstrated the overall satisfactory performance of the model. Conclusion: Our groundbreaking nomogram is distinguished by its unparalleled accuracy in discriminative and calibrating functions, complemented by its tangible clinical applicability. This innovative instrument is set to empower clinicians with a robust framework for the accurate forecasting of postoperative DVT, thus facilitating the crafting of bespoke and prompt therapeutic strategies, aligning with the rigorous standards upheld by the most esteemed biomedical journals.

Photoinduced Catalyst-Free Deuterodefunctionalization of Aryltriazenes with CDCl3.

A photoinduced deuterodetriazenation of aryltriazenes with CDCl3 under catalyst-free conditions is reported. The reactions featured simple operation, ecofriendly conditions, readily available reagents, inexpensive D sources, precise site selectivity, and a wide range of substrates. Since aryltriazenes could be readily synthesized from arylamine, this protocol can be used as a general method for easily and accurately incorporating deuterium into aromatic systems by using CDCl3 as a D source.

Recombinant human growth hormone promotes wound angiogenesis in burned mice through the ERK signaling pathway.

Burns are the most severe type of trauma, and the resulting ischemia and hypoxia damage can promote the dysfunction and even failure of tissues and organs throughout the body, endangering patients' life safety. Recombinant human growth hormone (rhGH) has the functions of promoting protein synthesis to reverse negative nitrogen balance, accelerating wound healing, and improving immune function, which is widely used in the treatment of burns. However, the exact mechanism and pathway of rhGH's action is not yet fully understood. In this study, we observed the wound repair effect of recombinant human growth hormone (rhGH) on burned mice and further analyzed the mechanism of action, which can provide more comprehensive reference opinions for clinical practice. First, by establishing a burn mouse model and and intervening with different doses of rhGH, we found that the wound healing capacity of mice was significantly enhanced and the inflammatory and oxidative stress responses were obviously alleviated, confirming the excellent promotion of wound repair and anti-inflammatory and antioxidant effects of rhGH. Subsequently, we found that the expression of p-ERK1/2/ERK1/2, EGF, TGF-ß, and VEGF proteins was elevated in the traumatic tissues of mice after rhGH intervention, suggesting that the pathway of action of rhGH might be related to the activation of ERK pathway to promote the regeneration of traumatic capillaries.

Functional and structural dissection of glycosyltransferases underlying the glycodiversity of wolfberry-derived bioactive ingredients lycibarbarspermidines.

Lycibarbarspermidines are unusual phenolamide glycosides characterized by a dicaffeoylspermidine core with multiple glycosyl substitutions, and serve as a major class of bioactive ingredients in the wolfberry. So far, little is known about the enzymatic basis of the glycosylation of phenolamides including dicaffeoylspermidine. Here, we identify five lycibarbarspermidine glycosyltransferases, LbUGT1-5, which are the first phenolamide-type glycosyltransferases and catalyze regioselective glycosylation of dicaffeoylspermidines to form structurally diverse lycibarbarspermidines in wolfberry. Notably, LbUGT3 acts as a distinctive enzyme that catalyzes a tandem sugar transfer to the ortho-dihydroxy group on the caffeoyl moiety to form the unusual ortho-diglucosylated product, while LbUGT1 accurately discriminates caffeoyl and dihydrocaffeoyl groups to catalyze a site-selective sugar transfer. Crystal structure analysis of the complexes of LbUGT1 and LbUGT3 with UDP, combined with molecular dynamics simulations, revealed the structural basis of the difference in glycosylation selectivity between LbUGT1 and LbUGT3. Site-directed mutagenesis illuminates a conserved tyrosine residue (Y389 in LbUGT1 and Y390 in LbUGT3) in PSPG box that plays a crucial role in regulating the regioselectivity of LbUGT1 and LbUGT3. Our study thus sheds light on the enzymatic underpinnings of the chemical diversity of lycibarbarspermidines in wolfberry, and expands the repertoire of glycosyltransferases in nature.

Comparative Epigenetic Profiling Reveals Distinct Features of Mucosal Melanomas Associated with Immune Cell Infiltration and Their Clinical Implications.

Mucosal melanoma exhibits limited responsiveness to anti-PD-1 therapy. However, a subgroup of mucosal melanomas, particularly those situated at specific anatomic sites like primary malignant melanoma of the esophagus (PMME), display remarkable sensitivity to anti-PD-1 treatment. The underlying mechanisms driving this superior response and the DNA methylation patterns in mucosal melanoma have not been thoroughly investigated. We collected tumor samples from 50 patients with mucosal melanoma, including 31 PMME and 19 non-esophageal mucosal melanoma (NEMM). Targeted bisulfite sequencing was conducted to characterize the DNA methylation landscape of mucosal melanoma and explore the epigenetic profiling differences between PMME and NEMM. Bulk RNA sequencing and multiplex immunofluorescence staining were performed to confirm the impact of methylation on gene expression and immune microenvironment. Our analysis revealed distinct epigenetic signatures that distinguish mucosal melanomas of different origins. Notably, PMME exhibited distinct epigenetic profiling characterized by a global hypermethylation alteration compared with NEMM. The prognostic model based on the methylation scores of a 7-DMR panel could effectively predict the overall survival of patients with PMME and potentially serve as a prognostic factor. PMME displayed a substantial enrichment of immune-activating cells in contrast to NEMM. Furthermore, we observed hypermethylation of the TERT promoter in PMME, which correlated with heightened CD8+ T-cell infiltration, and patients with hypermethylated TERT were likely to have improved responses to immunotherapy. Our results indicated that PMME shows a distinct methylation landscape compared with NEMM, and the epigenetic status of TERT might be used to estimate prognosis and direct anti-PD-1 treatment for mucosal melanoma. SIGNIFICANCE: This study investigated the intricate epigenetic factor of mucosal melanomas contributed to the differential immune checkpoint inhibitor response, and found that PMME exhibited a global hypermethylation pattern and lower gene expression in comparison to NEMM. TERT hypermethylation may contribute to the favorable responses observed in patients with mucosal melanoma undergoing immunotherapy.

Tau Aggregation-Dependent Lipid Peroxide Accumulation Driven by the hsa_circ_0001546/14-3-3/CAMK2D/Tau Complex Inhibits Epithelial Ovarian Cancer Peritoneal Metastasis.

Intraperitoneal dissemination is the main method of epithelial ovarian cancer (EOC) metastasis, which is related to poor prognosis and a high recurrence rate. Circular RNAs (circRNAs) are a novel class of endogenous RNAs with covalently closed loop structures that are implicated in the regulation of tumor development. In this study, hsa_circ_0001546 is downregulated in EOC primary and metastatic tissues vs. control tissues and this phenotype has a favorable effect on EOC OS and DFS. hsa_circ_0001546 can directly bind with 14-3-3 proteins to act as a chaperone molecule and has a limited positive effect on 14-3-3 protein stability. This complex recruits CAMK2D to induce the Ser324 phosphorylation of Tau proteins, changing the phosphorylation status of Tau bound to 14-3-3 and ultimately forming the hsa_circ_0001546/14-3-3/CAMK2D/Tau complex. The existence of this complex stimulates the production of Tau aggregation, which then induces the accumulation of lipid peroxides (LPOs) and causes LPO-dependent ferroptosis. In vivo, treatment with ferrostatin-1 and TRx0237 rescued the inhibitory effect of hsa_circ_0001546 on EOC cell spreading. Therefore, based on this results, ferroptosis caused by Tau aggregation occurs in EOC cells, which is not only in Alzheimer's disease- or Parkinson's disease-related cells and this kind of ferroptosis driven by the hsa_circ_0001546/14-3-3/CAMK2D/Tau complex is LPO-dependent rather than GPX4-dependent is hypothesized.

Integrated analysis of metabolome, transcriptome, and bioclimatic factors of Acer truncatum seeds reveals key candidate genes related to unsaturated fatty acid biosynthesis, and potentially optimal production area.

BACKGROUND: Lipids found in plant seeds are essential for controlling seed dormancy, dispersal, and defenses against biotic and abiotic stress. Additionally, these lipids provide nutrition and energy and are therefore important to the human diet as edible oils. Acer truncatum, which belongs to the Aceaceae family, is widely cultivated around the world for its ornamental value. Further because its seed oil is rich in unsaturated fatty acids (UFAs)- i.e. α-linolenic acid (ALA) and nervonic acid (NA)- and because it has been validated as a new food resource in China, the importance of A. truncatum has greatly risen. However, it remains unknown how UFAs are biosynthesized during the growth season, to what extent environmental factors impact their content, and what areas are potentially optimal for their production. RESULTS: In this study, transcriptome and metabolome of A. truncatum seeds at three representative developmental stages was used to find the accumulation patterns of all major FAs. Cumulatively, 966 metabolites and 87,343 unigenes were detected; the differential expressed unigenes and metabolites were compared between stages as follows: stage 1 vs. 2, stage 1 vs. 3, and stage 2 vs. 3 seeds, respectively. Moreover, 13 fatty acid desaturases (FADs) and 20 ß-ketoacyl-CoA synthases (KCSs) were identified, among which the expression level of FAD3 (Cluster-7222.41455) and KCS20 (Cluster-7222.40643) were consistent with the metabolic results of ALA and NA, respectively. Upon analysis of the geographical origin-affected diversity from 17 various locations, we found significant variation in phenotypes and UFA content. Notably, in this study we found that 7 bioclimatic variables showed considerable influence on FAs contents in A. truncatum seeds oil, suggesting their significance as critical environmental parameters. Ultimately, we developed a model for potentially ecological suitable regions in China. CONCLUSION: This study provides a comprehensive understanding of the relationship between metabolome and transcriptome in A. truncatum at various developmental stages of seeds and a new strategy to enhance seed FA content, especially ALA and NA. This is particularly significant in meeting the increasing demands for high-quality edible oil for human consumption. The study offers a scientific basis for A. truncatum's novel utilization as a woody vegetable oil rather than an ornamental plant, potentially expanding its cultivation worldwide.

Evolutionary patterns of DNA base composition at polymorphic sites highlight the role of the environment in shaping barley and rice genomes.

Insights into changes in genome base composition underlying crop domestication can be gained by using comparative genomics. With this approach, previous studies have reported that crop genomes during domestication accumulate more nucleotides adenine (A) and thymine (T) (termed as [AT]-increase) across polymorphic sites. However, the potential influence of the environment or its factors, for example, solar ultraviolet (UV) radiation and temperature, on the [AT]-increase has not been well elucidated. Here, we investigated the [AT]-increase in barley (Hordeum vulgare L.) and rice (Oryza sativa L.) and the association with natural environments, where accessions are distributed. With 12,798,376 and 2,861,535 single-nucleotide polymorphisms from 368 barley and 1375 rice accessions, respectively, we discovered that [AT] increases from wild accessions to improved cultivars, and genomic regions with larger [AT]-increase tend to have higher UV-related motif frequencies, suggesting solar UV radiation as a potential factor in driving genome variation. To link [AT] change with the geographic distribution, we gathered georeferenced accessions and examined their local environments. Interestingly, negative correlations between [AT] and environmental factors were observed (r = -0.39 ∼ -0.75) and modern accessions with higher [AT] values, as compared with wild relatives, are from the environments with lower solar UV radiation or lower temperature. With [AT] and environmental factors as phenotypes, genome-wide association mapping identified three candidate genes that have the potential to contribute to [AT] variation under the effect of environmental conditions. Our findings provide genomic and environmental insights into evolutionary pattern of DNA base composition and underlying mechanisms.

p-Smad3 differentially regulates the cytological behavior of osteoclasts before and after osteoblasts maturation.

BACKGROUND: A series of previous investigations have revealed that p-Smad3 plays a facilitative role in the differentiation and maturation of osteoblasts, while also regulating the expression of certain intercellular communication factors. However, the effects of p-Smad3 in osteoblasts before and after maturation on the proliferation, migration, differentiation, apoptosis and other cellular behaviors of osteoclasts have not been reported. METHODS: MC3T3-E1 cells were cultured in osteogenic induction medium for varying durations, After that, the corresponding conditioned medium was collected and the osteoclast lineage cells were treated. To elucidate the regulatory role of p-Smad3 within osteoblasts, we applied the activator TGF-ß1 and inhibitor SIS3 to immature and mature osteoblasts and collected corresponding conditioned media for osteoclast intervention. RESULTS: We observed an elevation of p-Smad3 and Smad3 during the early stage of osteoblast differentiation, followed by a decline in the later stage. we discovered that as osteoblasts mature, their conditioned media inhibit osteoclasts differentiation and the osteoclast-coupled osteogenic effect. However, it promotes apoptosis in osteoclasts and the angiogenesis coupled with osteoclasts. p-Smad3 in immature osteoblasts, through paracrine effects, promotes the migration, differentiation, and osteoclast-coupled osteogenic effects of osteoclast lineage cells. For mature osteoblasts, p-Smad3 facilitates osteoclast apoptosis and the angiogenesis coupled with osteoclasts. CONCLUSIONS: As pre-osteoblasts undergo maturation, p-Smad3 mediated a paracrine effect that transitions osteoclast cellular behaviors from inducing differentiation and stimulating bone formation to promoting apoptosis and coupling angiogenesis.

Ginsenoside Rc alleviates osteoporosis by the TGF-ß/Smad signaling pathway.

Osteoporosis is a common chronic bone disorder in postmenopausal women. Ginsenosides are primary active components in ginseng and the effects of various ginsenoside variants in osteoporosis treatment have been widely revealed. We planned to explore the impact of ginsenoside Rc on bone resorption in an osteoporosis rat model. We used ovariectomized rats to assess the potential impact of ginsenoside Rc on osteoporosis. µ-CT was implemented for analyzing the microstructure of the distal left femur in rats. H&E staining together with Masson staining were applied for bone histomorphometry evaluation. ELISA kits were implemented to detect serum concentrations of TRACP-5b, OCN, CTX, as well as PINP. Ginsenoside Rc treatment lessened the serum levels of TRACP-5b as well as CTX, while increasing serum levels of OCN, and PINP of OVX rats. Moreover, we found that ginsenoside Rc contributed to the synthesis of type I collagen via increasing Col1a1 and Col1a2 levels in femur tissues of ovariectomized rats. Our findings also revealed that ginsenoside Rc activated the TGF-ß/Smad pathway by increasing TGF-ß as well as phosphorylated Smad2/3 protein levels. Ginsenoside Rc alleviates osteoporosis in rats through promoting the TGF-ß/Smad pathway.

[The Regulatory Effect of RNA m6 A Methylation Modification on KDM4B Gene Expression in t (8;21) AML Cells by MeRIP-qPCR].

OBJECTIVE: To confirm the direct regulatory effect of WTAP-mediated RNA m6A modification on the KDM4B gene in t (8;21) acute myeloid leukemia (AML) cells through MeRIP combined with reverse transcription real-time quantitative PCR (RT-qPCR) technology. METHODS: The lentivirus-mediated shRNA target WTAP or KDM4B gene was used to transfect the t (8;21) AML cell lines: Kasumi-1 and SKNO-1, and cells transfected with randomly shuffled shRNA as the control. Using the Ultrapure RNA Extraction Kit (DNase I) to extract RNA. The Magna MeRIPTM m6A Kit was used to enrich methylated modified fragments, and detect the m6A methylated RNA regions by RT-qPCR, and the protein and mRNA expression levels of WTAP and KDM4B in cells were detected by Western blot and reverse transcription real-time quantitative PCR (RT-qPCR). Colony formation assays were used to detect the colony ability of cells in vitro. RESULTS: Silencing the expression of WTAP in Kasumi-1 cells, the enrichment of m6A methylation modification was significantly decreased in the 3'UTR of KDM4B mRNA(P < 0.01), and the protein(P < 0.001) and mRNA (Kasumi-1:P < 0.001; SKNO-1: P < 0.01) expression levels of KDM4B were also significantly inhibited in Kasumi-1 and SKNO-1 cells upon WTAP knockdown (all P < 0.01), accompanied by a significant decrease in the colony-forming ability of both cell lines (both P < 0.01). CONCLUSION: In t(8;21) AML cell lines, WTAP could regulate the expression of KDM4B by regulating the m6A modification of the 3'UTR of KDM4B mRNA, and silencing the expression of KDM4B could inhibit the cellular proliferation in vitro.

Establishment and characterization of a golden pompano (Trachinotus blochii) fin cell line for applications in marine fish pathogen immunology.

Pompano fishes have been widely farmed worldwide. As a representative commercial marine species of the Carangidae family, the golden pompano (Trachinotus blochii) has gained significant popularity in China and worldwide. However, because of rapid growth and high-density aquaculture, the golden pompano has become seriously threatened by various diseases. Cell lines are the most cost-effective resource for in vitro studies and are widely used for physiological and pathological research owing to their accessibility and convenience. In this study, we established a novel immortal cell line, GPF (Golden pompano fin cells). GPF has been passaged over 69 generations for 10 months. The morphology, adhesion and extension processes of GPF were evaluated using light and electron microscopy. GPF cells were passaged every 3 days with L-15 containing 20 % fetal bovine serum (FBS) at 1:3. The optimum conditions for GPF growth were 28 °C and a 20 % FBS concentration. DNA sequencing of 18S rRNA and mitochondrial 16S rRNA confirmed that GPF was derived from the golden pompano. Chromosomal analysis revealed that the number pattern of GPF was 48 chromosomes. Transfection experiments demonstrated that GPF could be utilized to express foreign genes. Furthermore, heavy metals (Cd, Cu, and Fe) exhibited dose-dependent cytotoxicity against GPF. After polyinosinic-polycytidylic acid (poly I:C) treatment, transcription of the retinoic acid-inducible gene I-like receptor (RLR) pathway genes, including mda5, mita, tbk1, irf3, and irf7 increased, inducing the expression of interferon (IFN) and anti-viral proteins in GPF cells. In addition, lipopolysaccharide (LPS) stimulation up-regulated the expression of inflammation-related factors, including myd88, irak1, nfκb, il1ß, il6, and cxcl10 expression. To the best of our knowledge, this is the first study on the immune response signaling pathways of the golden pompano using an established fin cell line. In this study, we describe a preliminary investigation of the GPF cell line immune response to poly I:C and LPS, and provide a more rapid and efficient experimental material for research on marine fish immunology.

Enhancing Built-in Electric Fields via Molecular Symmetry Modulation in Supramolecular Photocatalysts for Highly Efficient Photocatalytic Hydrogen Evolution.

Nature-inspired supramolecular self-assemblies are attractive photocatalysts, but their quantum yields are limited by poor charge separation and transportation. A promising strategy for efficient charge transfer is to enhance the built-in electric field by symmetry breaking. Herein, an unsymmetric protonation, N-heterocyclic π-conjugated anthrazoline-based supramolecular photocatalyst SA-DADK-H+ was developed. The unsymmetric protonation breaks the initial structural symmetry of DADK, resulting in ca. 50-fold increase in the molecular dipole, and facilitates efficient charge separation and transfer within SA-DADK-H+. The protonation process also creates numerous active sites for H2O adsorption, and serves as crucial proton relays, significantly improving the photocatalytic efficiency. Remarkably, SA-DADK-H+ exhibits an outstanding hydrogen evolution rate of 278.2 mmol g-1 h-1 and a remarkable apparent quantum efficiency of 25.1 % at 450 nm, placing it among the state-of-the-art performances in organic semiconductor photocatalysts. Furthermore, the versatility of the unsymmetric protonation approach has been successfully applied to four other photocatalysts, enhancing their photocatalytic performance by 39 to 533 times. These findings highlight the considerable potential of unsymmetric protonation induced symmetry breaking strategy in tailoring supramolecular photocatalysts for efficient solar-to-fuel production.

TMBstable: a variant caller controls performance variation across heterogeneous sequencing samples.

In cancer genomics, variant calling has advanced, but traditional mean accuracy evaluations are inadequate for biomarkers like tumor mutation burden, which vary significantly across samples, affecting immunotherapy patient selection and threshold settings. In this study, we introduce TMBstable, an innovative method that dynamically selects optimal variant calling strategies for specific genomic regions using a meta-learning framework, distinguishing it from traditional callers with uniform sample-wide strategies. The process begins with segmenting the sample into windows and extracting meta-features for clustering, followed by using a pre-trained meta-model to select suitable algorithms for each cluster, thereby addressing strategy-sample mismatches, reducing performance fluctuations and ensuring consistent performance across various samples. We evaluated TMBstable using both simulated and real non-small cell lung cancer and nasopharyngeal carcinoma samples, comparing it with advanced callers. The assessment, focusing on stability measures, such as the variance and coefficient of variation in false positive rate, false negative rate, precision and recall, involved 300 simulated and 106 real tumor samples. Benchmark results showed TMBstable's superior stability with the lowest variance and coefficient of variation across performance metrics, highlighting its effectiveness in analyzing the counting-based biomarker. The TMBstable algorithm can be accessed at https://github.com/hello-json/TMBstable for academic usage only.