Transcriptome Sequencing Analysis Research Articles

Treponema pallidum protein Tp0136 promotes angiogenesis to facilitate the dissemination of Treponema pallidum

The incompletely eliminated Treponema pallidum (T. pallidum) during primary syphilis chancre infection can result in the progression of secondary, tertiary, or latent syphilis in individuals, suggesting that T. pallidum has successfully evaded the immune response and spread to distant sites. The mechanism underlying the dissemination of T. pallidum is unclear. Here, a syphilitic rabbit model dorsal-injected with recombinant Tp0136 protein or Tp0136 antibody subcutaneously was used to demonstrate the role of Tp0136 protein in promoting the dissemination of T. pallidum to the testis and angiogenesis in vivo; vascular endothelial cell line HMEC-1 was employed to display that Tp0136 protein enhances the angiogenesis. Furthermore, the three-dimensional microfluidic angiogenesis system showed that the angiogenesis would heighten vascular permeability. Then transcriptome sequencing analysis, in conjunction with cell-level validation, elucidated the critical role of the PI3K-AKT signaling pathway in the promotion of angiogenesis by Tp0136 protein, resulting in heightened permeability. These findings elucidate the strategy employed by T. pallidum in evading immune clearance.

LAIR1 promotes hepatocellular carcinoma cell metastasis and induces M2-macrophage infiltration through activating AKT-IKKβ-p65 axis.

LAIR1, a receptor found on immune cells, is capable of binding to collagen and is involved in immune-related diseases. However, the precise contribution of LAIR1 expressed on hepatocellular carcinoma (HCC)cells to tumor microenvironment is still unclear. In our study, bioinformatics analysis and immunofluorescence were employed to study the correlation between LAIR1 levels and clinical indicators. Transwell and scratch tests were used to evaluate how LAIR1 affected the migration and invasion of HCC cells. The chemotactic capacity and alternative activation of macrophages were investigated using RT-qPCR,transwell, and immunofluorescence. To investigate the molecular mechanisms, transcriptome sequencing analysis, Western blot, nucleus/cytoplasm fractionation, ELISA, and cytokine microarray were employed. We revealed a significant correlation between the presence of LAIR1 and an unfavorable outcome in HCC. We indicated that LAIR1 promoted migration and invasion of HCC cells through the AKT-IKKβ-p65axis. Additionally, the alternative activation and infiltration of tumor-associated macrophages induced by LAIR1 were reliant on the upregulation of IL6 and CCL5 within this axis, respectively. In conclusion, blocking LAIR1 was found to be an effective approach in combating the cancerous advancement of HCC.

Hydrolyzed Chicken Meat Extract and Its Bioactive Cyclopeptides Protect Neural Function by Attenuating Inflammation and Apoptosis via PI3K/AKT and AMPK Pathways.

Cognitive decline is inevitable with age, and due to the lack of well-established pharmacotherapies for neurodegenerative disorders, dietary supplements have become important alternatives to ameliorate brain deterioration. Hydrolyzed chicken meat extract (HCE) and its bioactive components were previously found to improve neuroinflammation and cognitive decline by regulating microglia polarization. However, the effects and mechanisms of these bioactives on neurons remain unclear. Here, the most potent bioactive component on neural function in HCE was screened out, and the detailed mechanism was clarified through in vivo and in vitro experiments. We found that HCE, cyclo(Val-Pro), cyclo(Phe-Phe), cyclo(His-Pro), cyclo(Leu-Lys), and arginine exerted stronger anti-inflammatory and antioxidant effects among the 12 bioactives in amyloid β (Aβ)-treated HT-22 cells. Further transcriptome sequencing and polymerase chain reaction (PCR) array analysis showed that these bioactives participated in different signaling pathways, and cyclo(Val-Pro) was identified as the most potent cyclic dipeptide. In addition, the antiapoptotic and neuroprotective effect of cyclo(Val-Pro) was partly regulated by the activation of PI3K/AKT and AMPK pathways, and the inhibition of these pathways abolished the effect of cyclo(Val-Pro). Moreover, cyclo(Val-Pro) enhanced cognitive function and neurogenesis and alleviated neuroinflammation and oxidative stress in middle-aged mice, with an effect similar to HCE. Hippocampal transcriptome analysis further revealed that HCE and cyclo(Val-Pro) significantly enriched the neuroactive ligand-receptor interaction pathway, verified by enhanced neurotransmitter levels and upregulated neurotransmitter receptor-related gene expression. Therefore, the mechanism of cyclo(Val-Pro) on neural function might be associated with PI3K/AKT and AMPK pathway-mediated antiapoptotic effect and neurogenesis and the activation of the neurotransmitter-receptor pathway.

Overexpression of Nrf2 in bone marrow mesenchymal stem cells promotes B-cell acute lymphoblastic leukemia cells invasion and extramedullary organ infiltration through stimulation of the SDF-1/CXCR4 axis

BackgroundTumor microenvironment (TME) represents the key factor inducing leukemia development. As stromal cells within the leukemia microenvironment, Bone Marrow Mesenchymal Stem Cells (BM-MSCs) can trigger leukemia progression under certain conditions. As a critical transcription factor, nuclear factor erythroid related factor 2 (Nrf2) can modulate antioxidant response and antioxidant enzyme gene expression, and prevent various oxidative changes. We previously identified a novel mechanism by which Nrf2 promotes leukemia resistance, providing a potential therapeutic target for the treatment of drug-resistant/refractory leukemias. However, the role of Nrf2 in BM-MSCs from B-cell acute lymphoblastic leukemia (B-ALL) patients has not been clearly reported. The present work focused on investigating the effect of Nrf2 overexpression within MSCs on leukemia cell invasion, extramedullary infiltration and proliferation as well as its downstream pathway.MethodsThrough clinical sample detection, in vitro cell experiments and in vivo animal experiments, the role of Nrf2 within MSCs within adult B-ALL cell migration and invasion and its potential molecular mechanism was explored through transcriptome sequencing analysis, RT-PCR, Western blot, cell migration, cell invasion, lentivirus transfection and other experiments.ResultsNrf2 was highly expressed in BM-MSCs from patients with B-ALL as well as in BM-MSCs co-cultured with leukemia cells. Overexpression of Nrf2 within MSCs significantly promoted leukemia cell migration, invasion and proliferation. The extramedullary organ infiltration rate in B-ALL model mice receiving the combined infusion of both cell types dramatically increased relative to that of leukemia cells alone, accompanied by the significantly shortened survival time. Mechanism study found that Nrf2 overexpression within MSCs promoted PI3K-AKT/ERK1/2 phosphorylation in the downstream pathway by activating SDF-1/CXCR4 axis, ultimately leading to extramedullary infiltration of leukemia cells.ConclusionHigh Nrf2 expression with in MSCs enhances leukemia cell invasion and migration, which then accelerates infiltration in leukemic extramedullary organs. Targeting Nrf2 or inhibiting its downstream signal molecules may be the effective interventions for B-ALL patients treatment.

Insights into the Early Steps of the Symbiotic Interaction between Soybean (Glycine max) and Sinorhizobium fredii Symbiosis Using Transcriptome, Small RNA, and Degradome Sequencing.

Symbiotic nitrogen fixation carried out by the soybean-rhizobia symbiosis increases soybean yield and reduces the amount of nitrogen fertilizer that has been applied. MicroRNAs (miRNAs) are crucial in plant growth and development, prompting an investigation into their role in the symbiotic interaction of soybean with partner rhizobia. Through integrated small RNA, transcriptome, and degradome sequencing analysis, 1215 known miRNAs, 314 of them conserved, and 187 novel miRNAs were identified, with 44 differentially expressed miRNAs in soybean roots inoculated with Sinorhizobium fredii HH103 and a ttsI mutant. The study unveiled that the known miRNA gma-MIR398a-p5 was downregulated in the presence of the ttsI mutation, while the target gene of gma-MIR398a-p5, Glyma.06G007500, associated with nitrogen metabolism, was upregulated. The results of this study offer insights for breeding high-efficiency nitrogen-fixing soybean varieties, enhancing crop yield and quality.

Zebrafish myo7aa affects congenital hearing by regulating Rho-GTPase signaling

Introductionmyo7aa, the homolog of the human Usher 1B syndrome pathogenic gene, myo7A, plays an important role in stereociliary development and maintenance, therefore, is critical for hearing and balance. However, the molecular mechanisms that myo7aa regulate hearing and balance still need to be studied.MethodsIn this study, we generated two independent zebrafish myo7aa knockout lines using CRISPR/Cas9 technology. To investigate the effects of myo7aa on hearing, YO-PRO-1 staining and startle response assay were used. To gain insight into the specific molecular mechanisms by which myo7aa affects hearing, transcriptome sequencing and bioinformatics analysis were employed.ResultsOur study showed that hair cells of myo7aa-/- zebrafish can not take up YO-PRO-1 fluorescent dye and are insensitive to acoustic stimulation in myo7aa-/- zebrafish compared to wild type. Genes related to the Rho GTPase signaling pathway, such as arhgap33, dab2ip, and arghef40, are significantly down-regulated in myo7aa-/- zebrafish embryos at 3 dpf. GTP and ATP compensation can partially rescue the hair cell defects in myo7aa knockout zebrafish.DiscussionOur findings suggest that zebrafish myo7aa affects congenital hearing by regulating Rho GTPase signaling, and loss of myo7aa leads to abnormal Rho GTPase signaling and impairs hair cell function. myo7aa, myo7A, arhgap33, dab2ip, arghef40 and myo7aa-/- fonts in the abstract are italicized. -/- is a superscript format.

Integrated Transcriptome and Metabolome to Elucidate the Mechanism of Aluminum-Induced Blue-Turning of Hydrangea Sepals

Hydrangea macrophylla is an ornamental plant with varied calyx colors. Interestingly, from red, to purple, to blue, the colors of all Hydrangea macrophylla are formed by unique delphinidin-3-O-glucoside and aluminum ions (Al3+) and 5-O-p-coumaroylquinic acid. The sepals of ‘Blue Mama’ changed from pink to blue, and the contents of delphinidin-3-O-glucoside and aluminum ions increased under 3 g/L aluminum sulfate treatment. However, the mechanism of the effect of aluminum ions on the synthesis and metabolism of anthocyanins in Hydrangea macrophylla is still unclear. In this project, transcriptome sequencing and anthocyanin metabolome analysis were performed on the sepals of ‘Blue Mama’ during flower development at the bud stage (S1), discoloration stage (S2) and full-bloom stage (S3) under aluminum treatment. It was found that delphinidin, delphinidin-3-O-glucoside and delphinidin-3-O-galactoside were the main differential metabolites. The structural genes CHS, F3H, ANS, DFR and BZI in the anthocyanin synthesis pathway were up-regulated with the deepening in sepal color. There was no significant difference between the aluminum treatment and the non-aluminum treatment groups. However, seven transcription factors were up-regulated and expressed to regulate anthocyanin synthesis genes CHS, F3H, BZI and 4CL, promoting the sepals to turn blue. The KEGG enrichment pathway analysis of differentially expressed genes showed that the glutathione metabolism and the ABC transporter pathway were closely related to anthocyanin synthesis and aluminum-ion transport. GST (Hma1.2p1_0158F.1_g069560.gene) may be involved in the vacuolar transport of anthocyanins. The expression of anthocyanin transporter genes ABCC1 (Hma1.2p1_0021F.1_g014400.gene), ABCC2 (Hma1.2p1_0491F.1_g164450.gene) and aluminum transporter gene ALS3 (Hma1.2p1_0111F.1_g053440.gene) were significantly up-regulated in the aluminum treatment group, which may be an important reason for promoting the transport of anthocyanin and aluminum ions to vacuoles and making the sepals blue. These results preliminarily clarified the mechanism of aluminum ion in the synthesis and transport of anthocyanin in Hydrangea macrophylla, laying a foundation for the further study of the formation mechanism of ‘blue complex’ in Hydrangea macrophylla.

Transcriptomic Expression of T2-Inflammation Genes in Peripheral Blood Mononuclear Cells and Longitudinal Clinical Outcomes in Asthma: Insights from the COREA Study.

Gene expression can provide distinct information compared to clinical biomarkers in the context of longitudinal clinical outcomes in asthma patients. This study examined the association between the gene expression levels of upstream (IL-25, IL-33, and TSLP) and downstream cytokines (IL-5, IL-4, and IL-13) in the T2 inflammatory pathway with a 12-month follow-up of exacerbation, lung function, and steroid use. Transcriptomic sequencing analysis was performed on peripheral blood mononuclear cells from 279 adult asthmatics. Survival analysis and linear mixed-effect models were used to investigate potential differences between the high-level and low-level gene expression groups and the clinical outcomes. Analysis was performed separately for the upstream, downstream, and all 6 cytokines. In general, T2 inflammatory cytokine gene expression showed a weak correlation with blood eosinophil counts (all r < 0.1) and clinical outcomes. Among moderate-to-severe eosinophilic asthma (MSEA) patients, individuals with elevated levels of downstream cytokines were at increased risk of time-to-first exacerbation (p = 0.044) and a greater increase of inhaled corticosteroid use over time (p = 0.002) compared to those with lower gene expression. There was no association between baseline T2 inflammatory cytokine gene expression and the longitudinal changes in lung function over time among MSEA patients. These findings suggest that, among MSEA patients, the gene expression levels of downstream cytokines in the T2 inflammatory pathway may serve as indicators for endotyping asthma.

Genome-wide analysis of the SWEET gene family in Hemerocallis citrina and functional characterization of HcSWEET4a in response to salt stress

Sugars will be eventually effluxed transporters (SWEETs) have been confirmed to play diverse physiological roles in plant growth, development and stress response. However, the characteristics and functions of the SWEET genes in Hemerocallis citrina remain unclear and poorly elucidated. In this study, the whole genome of Hemerocallis citrina was utilized to conduct bioinformatics analysis and a total of 19 HcSWEET genes were successfully identified. Analysis of the physicochemical properties indicated dominant differences among these HcSWEETs. A phylogenetic analysis revealed that HcSWEET proteins can be divided into 4 clades ranging from Clade I to IV, where proteins within the same clade exhibited shared conserved motifs and gene structures. Five to six exons were contained in the majority of HcSWEET genes, which were unevenly distributed across 11 chromosomes. The gene duplication analysis showed the presence of 4 gene pairs. Comparative syntenic maps revealed that the HcSWEET gene family might present more closed homology in monocotyledons than dicotyledons. Cis-acting element analysis of HcSWEET genes indicated key responsiveness to various hormones, light, and stresses. Additionally, transcriptome sequencing analysis suggested that most HcSWEET genes had a relatively higher expression in roots, and HcSWEET4a was significantly up-regulated under salt stress. Overexpression further verified the possibility that HcSWEET4a was involved in response to salt stress, which provides novel insights and facilitates in-depth studies of the functional analysis of HcSWEETs in resistance to abiotic stress.

Deficiency of ValRS-m Causes Male Infertility in Drosophila melanogaster.

Drosophila spermatogenesis involves the renewal of germline stem cells, meiosis of spermatocytes, and morphological transformation of spermatids into mature sperm. We previously demonstrated that Ocnus (ocn) plays an essential role in spermatogenesis. The ValRS-m (Valyl-tRNA synthetase, mitochondrial) gene was down-regulated in ocn RNAi testes. Here, we found that ValRS-m-knockdown induced complete sterility in male flies. The depletion of ValRS-m blocked mitochondrial behavior and ATP synthesis, thus inhibiting the transition from spermatogonia to spermatocytes, and eventually, inducing the accumulation of spermatogonia during spermatogenesis. To understand the intrinsic reason for this, we further conducted transcriptome-sequencing analysis for control and ValRS-m-knockdown testes. The differentially expressed genes (DEGs) between these two groups were selected with a fold change of ≥2 or ≤1/2. Compared with the control group, 4725 genes were down-regulated (dDEGs) and 2985 genes were up-regulated (uDEGs) in the ValRS-m RNAi group. The dDEGs were mainly concentrated in the glycolytic pathway and pyruvate metabolic pathway, and the uDEGs were primarily related to ribosomal biogenesis. A total of 28 DEGs associated with mitochondria and 6 meiosis-related genes were verified to be suppressed when ValRS-m was deficient. Overall, these results suggest that ValRS-m plays a wide and vital role in mitochondrial behavior and spermatogonia differentiation in Drosophila.

Single-cell transcriptome analysis reveals two subtypes of tumor cells of sclerosing pneumocytoma with distinct molecular features and clinical implications

Pulmonary sclerosing pneumocytoma (PSP) is a rare, distinctive benign lung adenoma of pneumocyte origin. Despite its rarity, the tumor's unique cellular morphology has sparked ongoing debates regarding the origin of its constituent cells. This study aimed to elucidate the molecular features of PSP tumor cells and enhance our understanding of the cellular processes contributing to PSP formation and biological behavior. Tissue samples from PSP and corresponding normal lung tissues (n = 4) were collected. We employed single-cell RNA sequencing and microarray-based spatial transcriptomic analyses to identify cell types and investigate their transcriptomes, with a focus on transcription factors, enriched gene expression, and single-cell trajectory evaluations. Our analysis identified two types of tumor cells: mesenchymal epithelial dual phenotype cells (MEDPs) and a distinct subpopulation of type II alveolar epithelial cells exhibiting characteristics slightly reminiscent of type I alveolar epithelial cells (AT2Cs), corresponding to histological round stromal cells and surface cuboidal cells, respectively. MEDPs displayed weak alveolar epithelial differentiation but strong collagen production capabilities, as indicated by the expression of both TTF-1 and vimentin. These cells played a pivotal role in forming the solid and sclerotic areas of PSP. Moreover, MEDPs exhibited a pronounced propensity for epithelial-mesenchymal transition, suggesting a greater potential for metastasis compared to AT2Cs. The capillary endothelial cells of PSP displayed notable diversity. Overall, this study provides, for the first time, a comprehensive mapping of the single-cell transcriptome profile of PSP. Our findings delineate two distinct subtypes of tumor cells, MEDPs and AT2Cs, each with its own biological characteristics and spatial distribution. A deeper understanding of these cell types promises insights into the histology and biological behaviors of this rare tumor.

Unveiling underlying mechanism of combined He–Ne laser and UV mutagenesis in Bacillus subtilis CICC 21927: A transcriptomic analysis

Conventional random mutation breeding remains the simplest and most cost-effective approach in enhancing strains in fermentation industry. Nonetheless, extensive research on compound mutagenesis are limited to screening target strains, and few studies on the interaction mechanism of compound mutagenesis factors. Herein, transcriptomic was employed to elucidate the molecular mechanism behind helium-neon (He–Ne) laser and ultraviolet (UV) compound mutagenesis in Bacillus subtilis. Transcriptomic sequencing analysis was conducted on five sample groups (control/CK, UV, laser/LA, UV-laser/UL, and laser-UV/LU), facilitating the statistical comparison of differentially expressed genes (DEGs) across 4 sample relationships (CK-vs-UV, CK-vs-LA, CK-vs-LU, and CK-vs-UL). The DEGs revealed that UV irradiation improved the biosynthesis of pyrimidine nucleotides by upregulating the activity of key enzymes (pyrimidine nucleotides synthetase, pyruvate dehydrogenase, phosphotransferase system (PTS) fructose-specific enzyme), while laser irradiation stimulated metabolism through the regulation of transmembrane transporter activity. Further KEGG enrichment analysis of 1891 DEGs in UV-vs-LU sample relationship unveiled 89 genes significantly enriched in the two-component system, which potentially linked to phytochrome. Moreover, analysis of 784 DEGs in UV-vs-UL sample relationship indicated up-regulation of genes associated with translation, replication, and genetic repair pathways, affirming the laser's induction of UV damage repair mechanisms. Subsequently, 9 pivotal genes (liaS, yvfT, yxdK, cheA, ykoW, uvrC, pcrA, recA, and ruvB) were selected for quantitative reverse transcriptional PCR (qRT-PCR) validation, demonstrating consistent trends with transcriptomic data. Conclusively, our findings unveiled interplay mechanism between UV and laser compound mutagenesis at molecular level, offered valuable insights for mutagenesis mode selection and further elucidation of mutagenesis mechanisms.

Transcriptome-wide profiling for melanocytes derived from newborn and adult human epidermis with enhanced proliferation.

The senescence-associated protein p16INK4A acts as a limiter element in cell-cycle progression. The loss of p16INK4A function is causally related to cellular immortalization. The increase in p16INK4A levels with advancing age was demonstrated in melanocytes. However, the characteristic difference between young and senescent melanocytes affecting immortalization of melanocytes remains unclear. In this study, we generated 10 different cell lines in total from newborn (NB) and adult (AD) primary normal human epidermal melanocytes (NHEM) using four different methods, transduction of CDK4R24C and cyclin D1 (K4D), K4D with TERT (K4DT), SV40 T-antigen (SV40T), and HPV16 E6 and E7 (E6/E7) and performed whole transcriptome sequencing analysis (RNA-Seq) to elucidate the differences of genome-wide expression profiles among cell lines. The analysis data revealed distinct differences in expression pattern between cell lines from NB and AD although no distinct biological differences were detected in analyses such as comparison of cell morphology, evaluation of cell proliferation, and cell cycle profiles. This study may provide useful in vitro models to benefit the understanding of skin-related diseases.

The correlation between TRIM28 expression and immune checkpoints in CRPC.

This study delves into the unexplored realm of castration-resistant prostate cancer (CRPC) by investigating the role of TRIM28 and its intricate molecular mechanisms using high-throughput single-cell transcriptome sequencing and advanced bioinformatics analysis. Our comprehensive examination unveiled dynamic TRIM28 expression changes, particularly in immune cells such as macrophages and CD8+ T cells within CRPC. Correlation analyses with TCGA data highlighted the connection between TRIM28 and immune checkpoint expression and emphasized its pivotal influence on the quantity and functionality of immune cells. Using TRIM28 knockout mouse models, we identified differentially expressed genes and enriched pathways, unraveling the potential regulatory involvement of TRIM28 in the cGAS-STING pathway. Invitro, experiments further illuminated that TRIM28 knockout in prostate cancer cells induced a notable anti-tumor immune effect by inhibiting M2 macrophage polarization and enhancing CD8+ T cell activity. This impactful discovery was validated in an insitu transplant tumor model, where TRIM28 knockout exhibited a deceleration in tumor growth, reduced proportions of M2 macrophages, and enhanced infiltration of CD8+ T cells. In summary, this study elucidates the hitherto unknown anti-tumor immune role of TRIM28 in CRPC and unravels its potential regulatory mechanism via the cGAS-STING signaling pathway. These findings provide novel insights into the immune landscape of CRPC, offering promising directions for developing innovative therapeutic strategies.

P-682 Malic enzyme 1 in granulosa cells promotes ovarian aging through inhibiting the AMPK-FoXO1-IDH2 signaling pathway

Abstract Study question Reduced levels of oxidative phosphorylation in granulosa cells are a key factor in regulating ovarian aging, but the upstream regulatory mechanisms are unknown. Summary answer Our study revealed that elevated malic enzyme 1 in granulosa cells promotes ovarian aging by inhibiting oxidative phosphorylation levels through the AMPK-FoXO1-IDH2 signaling pathway. What is known already With ovarian aging, the ovarian granulosa cells appear hallmarks of high apoptosis activity and low-proliferative activity, which lead to a reduction in ovarian reserve and a decrease in hormone synthesis, resulting in female infertility and reduced quality of life. And the decline of isocitrate dehydrogenase, IDH, in non-human primate ovarian granulosa cells is a key causative factor for the decline in the level of oxidative phosphorylation in granulosa cells. Recently, researchers reported that malic enzyme 1, which encodes a cytosolic, NADP-dependent Enzyme that generates NADPH for fatty acid biosynthesis inhibits IDH2 expression in tumor cells. Study design, size, duration Utilizing clinical samples, cells and animal models, with the help of research tools such as biochemical experiments, gene editing and single-cell transcriptome sequencing, at different levels, including molecular, cellular, animal and human samples, we systematically analyzed the characteristic changes and key molecules in granulosa cells along with ovarian aging and reveal the intracellular molecular pathways through which ME1 and IDH2 regulate the process of ovarian aging for 1 year. Participants/materials, setting, methods We first apply single-cell transcriptome sequencing analysis of young and old mouse ovaries to reveal cellular subpopulation changes and transcriptomic hallmarks of ovarian granulosa cell with aging. And then we validate the biological function and revealing the molecular mechanism to regulate ovarian aging by using physiological ovarian aging mouse model, clinical ovarian granulosa cells from young and old female and human ovarian granulosa cell line KGN utilizing technologies of protein immunology, qPCR and CRISPR/Cas9. Main results and the role of chance Ovarian single-cell transcriptome sequencing data from young and old mice showed that ovarian aging was accompanied by a dramatic decrease in the number of granulosa cells and drastic changes in the proportions of different subpopulations. The proportion of mitotically active granulosa cells was significantly lower in aging ovaries than in young mouse ovaries, and GO enrichment analysis showed that granulosa cell cycle, antioxidant and hormone synthesis-related signaling pathways were significantly down-regulated in aging ovaries. Using immunoblotting, immunofluorescence and qPCR, we demonstrated that the expression of ME1 in granulosa cells of aging mice and human ovaries was significantly elevated, while the expression of IDH2 was significantly decreased. Performing the overexpression and knockdown of ME1 in human ovarian granulosa cell line KGN, which further demonstrated that ME1 could inhibit the transcriptionally regulated IDH2 expression of downstream genes, such as FoXO1, by suppressing the phosphorylation of AMPK. The decrease of IDH2 in mitochondria reduces the level of oxidative phosphorylation in granulosa cells, resulting in the increase of intracellular reactive oxygen species, activation of the intracellular p53-p21 signaling pathway, inhibition of the cell cycle, and promotion of apoptosis, which in turn impairs the function of granulosa cells to synthesize hormones and promotes ovarian aging. Limitations, reasons for caution Malic enzyme 1, which links the glycolytic cycle to the citric acid cycle, expressing in other ovarian cells including oocytes, and the regulation of its expression is complex. Future studies will focus on its biological functions in other ovarian cells and its molecular mechanism in regulating ovarian aging. Wider implications of the findings Our study suggests that granulosa cell ME1 and IDH2 co-regulate oxidative phosphorylation levels to regulate ovarian aging, and that ME1 and IDH2 may serve as potential targets for the finding and treatment of premature ovarian aging and delaying ovarian aging in women. Trial registration number not applicable

The investigation of potential mechanism of Fuzhengkangfu Decoction against Diabetic myocardial injury based on a combined strategy of network pharmacology, transcriptomics, and experimental verification

Background and ObjectivesDiabetic cardiomyopathy (DCM) is a cardiac condition resulting from myocardial damage caused by diabetes mellitus (DM), currently lacking specific therapeutic interventions. Fuzhengkangfu decoction (FZK) plays an important role in the prevention and treatment of various cardiovascular diseases. However, the efficacy and potential mechanisms of FZK are not fully understood. This study aims to investigate the protective effect and mechanisms of FZK against DCM. MethodologiesRats were given a high-calorie diet along with a low dosage of streptozotocin (STZ) to establish a rat model of DCM. The diabetic rats received FZK or normal saline subcutaneously for 12 weeks. Echocardiography was conducted to evaluate their heart function characteristics. Rat heart morphologies were assessed using Sirius Red staining and H&E staining. Transcriptome sequencing analysis and network pharmacology were used to reveal possible targets and mechanisms. Molecular docking was conducted to validate the association between the primary components of FZK and the essential target molecules. Finally, both in vitro and in vivo studies were conducted on the cardioprotective properties and mechanism of FZK. ResultsAccording to the results of network pharmacology, FZK may prevent DCM by reducing oxidative stress and preventing apoptosis. Transcriptomics confirmed that FZK protected against DCM-induced myocardial fibrosis and remodelling, as predicted by network pharmacology, and suggested that FZK regulated the expression of oxidative stress and apoptosis-related proteins. Integrating network pharmacology and transcriptome analysis results revealed that the AGE-RAGE signalling pathway-associated MMP2, SLC2A1, NOX4, CCND1, and CYP1A1 might be key targets. Molecular docking showed that Poricoic acid A and 5-O-Methylvisammioside had the highest docking activities with these targets. We further conducted in vivo experiments, and the results showed that FZK significantly attenuated left ventricular remodelling, reduced myocardial fibrosis, and improved cardiac contractile function. And, our study demonstrated that FZK effectively reduced oxidative stress and apoptosis of cardiomyocytes. The data showed that Erk, NF-κB, and Caspase 3 phosphorylation was significantly inhibited, and Bcl-2/Bax was significantly increased after FZK treatment. In vitro, FZK significantly reduced AGEs-induced ROS increase and apoptosis in cardiomyocytes. Furthermore, FZK significantly inhibited the phosphorylation of Erk and NF-κB proteins and decreased the expression of MMP2. All the results confirmed that FZK inhibited the activation of the Erk/NF-κB pathway in AGE-RAGE signalling and alleviated oxidative stress and apoptosis of cardiomyocytes. In summary, we verified that FZK protects against DCM by inhibiting myocardial apoptotic remodelling through the suppression of the AGE-RAGE signalling pathway. ConclusionIn conclusion, our research indicates that FZK demonstrates anti-cardiac dysfunction properties by reducing oxidative stress and cardiomyocyte apoptosis through the AGE-RAGE pathway in DCM, showing potential for therapeutic use.

GL-V9 induce apoptosis of CML cells via MAPK signaling pathway

GL-V9, a derivative of wogonin, has shown potent antitumor effects in various cancers, yet its impact on chronic myeloid leukemia (CML) remains unexplored. In this study, we found that GL-V9 significantly decreased the viability of CML cells. Annexin V/PI staining demonstrated that GL-V9 induced apoptosis in a concentration-dependent manner. The JC-1 assay indicated a significant reduction in mitochondrial membrane potential (ΔΨm) in cells treated with GL-V9. Additionally, GL-V9 altered reactive oxygen species (ROS) levels in CML cells. Through transcriptomic sequencing and Western blot analysis, we further revealed that GL-V9 activated the MAPK pathway. These results suggest that GL-V9 is a promising therapeutic candidate for CML.

Mitotic catastrophe heterogeneity: implications for prognosis and immunotherapy in hepatocellular carcinoma.

The mitotic catastrophe (MC) pathway plays an important role in hepatocellular carcinoma (HCC) progression and tumor microenvironment (TME) regulation. However, the mechanisms linking MC heterogeneity to immune evasion and treatment response remain unclear. Based on 94 previously published highly correlated genes for MC, HCC patients' data from the Cancer Genome Atlas (TCGA) and changes in immune signatures and prognostic stratification were studied. Time and spatial-specific differences for MCGs were assessed by single-cell RNA sequencing and spatial transcriptome (ST) analysis. Multiple external databases (GEO, ICGC) were employed to construct an MC-related riskscore model. Identification of two MC-related subtypes in HCC patients from TCGA, with clear differences in immune signatures and prognostic risk stratification. Spatial mapping further associates low MC tumor regions with significant immune escape-related signaling. Nomogram combining MC riskscore and traditional indicators was validated great effect for early prediction of HCC patient outcomes. MC heterogeneity enables immune escape and therapy resistance in HCC. The MC gene signature serves as a reliable prognostic indicator for liver cancer. By revealing clear immune and spatial heterogeneity of HCC, our integrated approach provides contextual therapeutic strategies for optimal clinical decision-making.

Targeting MS4A4A: A novel pathway to improve immunotherapy responses in glioblastoma.

Glioblastoma (GBM) remains a challenging brain tumor to treat, with limited response to PD-1 immunotherapy due to tumor-associated macrophages (TAMs), specifically the M2 phenotype. This study explores the potential of MS4A4A (membrane spanning four domains, subfamily A, member 4A) inhibition in driving M2 macrophage polarization toward the M1 phenotype via the ferroptosis pathway to enhance the effectiveness of immunotherapy in GBM. Single-cell RNA sequencing and spatial transcriptomic analyses were employed to characterize M2 macrophages and MS4A4A expression in GBM. Invitro studies utilizing TAM cultures, flow cytometry, and western blot validations were conducted to assess the impact of MS4A4A on the tumor immune microenvironment and M2 macrophage polarization. Invivo models, including subcutaneous and orthotopic transplantation in mice, were utilized to evaluate the effects of MS4A4A knockout and combined immune checkpoint blockade (ICB) therapy on tumor growth and response to PD-1 immunotherapy. Distinct subsets of GBM-associated macrophages were identified, with spatial distribution in tumor tissue elucidated. Invivo experiments demonstrated that inhibiting MS4A4A and combining ICB therapy effectively inhibited tumor growth, reshaped the tumor immune microenvironment by reducing M2 TAM infiltration and enhancing CD8+ T-cell infiltration, ultimately leading to complete tumor eradication. MS4A4A inhibition shows promise in converting M2 macrophages to M1 phenotype via ferroptosis, decreasing M2-TAM infiltration, and enhancing GBM response to PD-1 immunotherapy. These findings offer a novel approach to developing more effective immunotherapeutic strategies for GBM.

Dihydroartemisinin regulates the apoptosis and growth of colorectal cancer by suppressing DPYSL2 and increasing TF and ACHE

Colorectal cancer (CRC) is considered a tumor of the digestive tract with high incidence and great danger. The complex pathogenesis of CRC has led to a lack of effective therapies and drugs in clinical practice. Study found that dihydroartemisinin (DHA) may selectively kill tumor cells by increasing the expression of transferrin receptors on tumor cell membranes. The intersection targets of DHA and CRC were analyzed by bioinformatics. Cell experiments were used to validate the results of bioinformatics. Flow cytometry was uesd to examine effect of DHA on apoptosis of SW480 cancer cell, the total RNA from cancer cell was used for transcriptome sequencing analysis. The binding stability of DHA to protein targets was verified through molecular docking. Binding free energy, hydrogen bonds and root-mean-square fluctuations were analysed by molecular dynamics. Core interaction protein targets were screened and analyzed by PPI, GO and KEGG. Cell experiments showed that DHA promoted apoptosis in SW480 cancer cell, results of transcriptome sequencing showed that DHA significantly up-regulated gene expression of TF and ACHE, while a down-regulated gene expression of DPYSL2. Molecular docking showed that DHA bound tightly to TF, ACHE and DPYSL2. Binding stability of dihydroartemisinin to protein targets was demonstrated by molecular dynamics. This study found that DHA may not only affect the apoptosis of CRC by regulating TF, but also hinder the growth and migration of CRC through ACHE and DPYSL2. The mechanism of DHA treating CRC was investigated by bioinformatics analysis, cellular experiments, transcriptome sequencing and supercomputer simulation.