Transcriptome Sequencing Analysis Research Articles

Study on the inactivation effect and mechanism of EGCG disinfectant on Bacillus subtilis

The widespread use of chlorine-based disinfectants in drinking water treatment has led to the proliferation of chlorine-resistant bacteria and the risk of disinfection byproducts (DBPs), posing a serious threat to public health. This study aims to explore the effectiveness and potential applications of epigallocatechin gallate (EGCG) against chlorine-resistant Bacillus and its spores in water, providing new insights for the control of chlorine-resistant bacteria and improving the biological stability of distribution systems. The inactivation effects of EGCG on Bacillus subtilis (B. subtilis) and its spores were investigated using transmission electron microscopy, ATP measurement, and transcriptome sequencing analysis to determine changes in surface structure, energy metabolism, and gene expression levels, thereby elucidating the inactivation mechanism. The results demonstrate the potential application of EGCG in continuously inhibiting chlorine-resistant B. subtilis in water, effectively improving the biological stabilit of the distribution system. However, EGCG is not suitable for treating raw water with high spore content and is more suitable as a supplementary disinfectant for processes with strong spore removal capabilities, such as ozone, ultraviolet, or ultrafiltration. EGCG exhibits a disruptive effect on the morphological structure and energy metabolism of B. subtilis and suppresses the synthesis of substances, energy metabolism, and normal operation of the antioxidant system by inhibiting the expression of multiple genes, thereby achieving the inactivation of B. subtilis.

Transcriptome-based protein-protein interaction analysis reveals immune gene network elucidating white body immunity mechanisms in response to LPS stimulation in Amphioctopus fangsiao

One of the major challenges in the aquaculture of marine organisms is the susceptibility to pathogens. Immune cells defend bacterial invasion by recognizing pathogen-associated molecular patterns (PAMPs) such as Lipopolysaccharides (LPS) in Gram-negative bacteria. The white body are key organs for immune response and hematopoiesis, and play an important role in understanding the antimicrobial biological response in gold-ringed octopus (Amphioctopus fangsiao). In the present research, transcriptome sequencing and bioinformatics analysis were conducted on A. fangsiao white body tissue after 6 h and 24 h of LPS stimulation. The results showed 2,029 and 73 differentially expressed genes (DEGs) at the 6- and 24-hour time points, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were beneficial in determining immune-related terms and signaling pathways. Subsequently, a protein-protein interaction (PPI) network was constructed and 20 hub genes, such as AKT3, MAPK14, and PIK3CA, were identified. These genes are involved in a variety of signaling pathways, including the Hippo signaling pathway, chemokine signaling pathway, and leukocyte trans endothelial migration. Using qRT-PCR, the accuracy of the 20 hub genes was verified. The present study provides a substantial theoretical foundation for the research on A. fangsiao white body immunity, enhancing the current understanding of cephalopods' innate immunity.

Multiomics Analysis of Interleukin-21 as a Potential Immunologic and Biomarker in Hepatocellular Carcinoma.

Interleukin-21 (IL-21) is an important antitumor cytokine that contributes to the proliferation and differentiation of CD8+ T cells. It has been proven to enhance the response to immune checkpoint inhibitors (ICIs) in various solid tumors. However, its role in hepatocellular carcinoma (HCC) has not yet been clarified. In this research, we aimed to investigate the antitumor effect of IL-21 in HCC and its effect on ICI treatment. Through transcriptome sequencing analysis and immunohistochemistry validation, we found that patients with high IL-21 expression had a better prognosis. HCCs with high expression of IL-21 had higher infiltration of CD8+ T cells, increased expression of immune checkpoints, and an improved response to ICI treatment. In conclusion, IL-21 can enhance the efficacy of ICI treatment and improve the prognosis of patients by promoting the infiltration of CD8+ T cells and the expression of immune checkpoint-related genes.

Identification of apoptosis-immune-related gene signature and construction of diagnostic model for sepsis based on single-cell sequencing and bulk transcriptome analysis.

Sepsis leads to multi-organ dysfunction due to disorders of the host response to infections, which makes diagnosis and prognosis challenging. Apoptosis, a classic programmed cell death, contributes to the pathogenesis of various diseases. However, there is much uncertainty about its mechanism in sepsis. Three sepsis gene expression profiles (GSE65682, GSE13904, and GSE26378) were downloaded from the Gene Expression Omnibus database. Apoptosis-related genes were obtained from the Kyoto Encyclopedia of Genes and Genomes Pathway database. We utilized LASSO regression and SVM-RFE algorithms to identify characteristic genes associated with sepsis. CIBERSORT and single cell sequencing analysis were employed to explore the potential relationship between hub genes and immune cell infiltration. The diagnostic capability of hub genes was validated across multiple external datasets. Subsequently, the animal sepsis model was established to assess the expression levels of hub genes in distinct target organs through RT-qPCR and Immunohistochemistry analysis. We identified 11 apoptosis-related genes as characteristic diagnostic markers for sepsis: CASP8, VDAC2, CHMP1A, CHMP5, FASLG, IFNAR1, JAK1, JAK3, STAT4, IRF9, and BCL2. Subsequently, a prognostic model was constructed using LASSO regression with BCL2, FASLG, IRF9 and JAK3 identified as hub genes. Apoptosis-related genes were closely associated with the immune response during the sepsis process. Furthermore, in the validation datasets, aside from IRF9, other hub genes demonstrated similar expression patterns and diagnostic abilities as observed in GSE65682 dataset. In the mouse model, the expression differences of hub genes between sepsis and control group revealed the potential impacts on sepsis-induced organ injury. The current findings indicated the participant of apoptosis in sepsis, and apoptosis-related differentially expressed genes could be used for diagnosis biomarkers. BCL2, FASLG, IRF9 and JAK3 might be key regulatory genes affecting apoptosis in sepsis. Our findings provided a novel aspect for further exploration of the pathological mechanisms in sepsis.

Transcriptome and metabolome sequencing identifies glutamate and LPAR1 as potential factors of anlotinib resistance in thyroid cancer.

To explore the mechanism of anlotinib resistance in thyroid carcinoma. We constructed an anlotinib-resistant thyroid carcinoma cell line and observed the effect of drug resistance on the functional activity of these cell lines. Transcriptome sequencing and metabolomic sequencing combined with biosynthesis analysis were used to explore and screen possible drug resistance regulatory pathways. Through transcriptomic sequencing analysis of drug-resistant cell lines, it was found that the differentially expressed genes of drug-resistant strains were enriched mainly in the interleukin 17, transforming growth factor-β, calcium, peroxisome proliferator activated receptor, and other key signaling pathways. A total of 354 differentially expressed metabolic ions were screened using liquid chromatography-mass spectrometry/mass spectrometry to determine the number of metabolic ions in the drug-resistant strains. The results of the Venn diagram correlation analysis showed that glutamate is closely related to multiple pathways and may be an important regulatory factor of anlotinib resistance in thyroid carcinoma. In addition, eight common differentially expressed genes were screened by comparing the gene expression profiling interactive analysis database and sequencing results. Further quantitative real time polymerase chain reaction verification, combined with reports in the literature, showed that LPAR1 may be an important potential target. This is the first study in which the drug resistance of thyroid cancer to anlotinib was preliminarily discussed. We confirmed that anlotinib resistance in thyroid cancer promotes the progression of malignant biological behavior. We conclude that glutamate may be a potential factor for anlotinib resistance in thyroid cancer and that LPAR1 is also a potentially important target.

SIRT1 mediates the excitability of spinal CaMKIIα-positive neurons and participates in neuropathic pain by controlling Nav1.3.

Neuropathic pain is a common chronic pain disorder, which is largely attributed to spinal central sensitization. Calcium/calmodulin-dependent protein kinase II alpha (CaMKIIα) activation in the spinal dorsal horn (SDH) is a major contributor to spinal sensitization. However, the exact way that CaMKIIα-positive (CaMKIIα+) neurons in the SDH induce neuropathic pain is still unclear. This study aimed to explore the role of spinal CaMKIIα+ neurons in neuropathic pain caused by chronic constriction injury (CCI) and investigate the potential epigenetic mechanisms involved in CaMKIIα+ neuron activation. CCI-induced neuropathic pain mice model, Sirt1loxP/loxP mice, and chemogenetic virus were used to investigate whether the activation of spinal CaMKIIα+ neurons is involved in neuropathic pain and its involved mechanism. Transcriptome sequence, western blotting, qRT-PCR, and immunofluorescence analysis were performed to assay the expression of related molecules and activation of neurons. Co-immunoprecipitation was used to observe the binding relationship of protein. Chromatin immunoprecipitation (ChIP)-PCR was applied to analyze the acetylation of histone H3 in the Scn3a promoter region. The expression of sodium channel Nav1.3 was increased and the expression of SIRT1 was decreased in the spinal CaMKIIα+ neurons of CCI mice. CaMKIIα neurons became overactive after CCI, and inhibiting their activation relieved CCI-induced pain. Overexpression of SIRT1 reversed the increase of Nav1.3 and alleviated pain, while knockdown of SIRT1 or overexpression of Nav1.3 promoted CaMKIIα+ neuron activation and induced pain. By knocking down spinal SIRT1, the acetylation of histone H3 in the Scn3a (encoding Nav1.3) promoter region was increased, leading to an increased expression of Nav1.3. The findings suggest that an aberrant reduction of spinal SIRT1 after nerve injury epigenetically increases Nav1.3, subsequently activating CaMKIIα+ neurons and causing neuropathic pain.

Hydrogen Nanobubbles and Oxidative Windows: Investigating How Varying Hydrogen Concentrations Influence Seed Germination and Stress Defense

The hydrogen molecule can effectively regulate plant growth and development, improving plant resistance to abiotic stresses. However, studies regarding the optimal concentration of hydrogen and the associated mechanisms of action in organisms are lacking. This study showed that the maximum germination rate of radish seeds decreased from 90% to 50% under the stress of cadmium ions (Cd2+), and hydrogen nanobubble (NB) water significantly alleviated the stress effect of Cd2+ on radish seed germination. A hydrogen concentration of 0.8 ppm had the best effect, reducing Cd2+ accumulation in radish seeds by 63.23% and increasing the maximum germination rate from 50% to 65%. At concentrations exceeding 1.2 ppm, the beneficial effect of hydrogen was weakened or even reversed. Consequently, we integrated the concept of the oxidative window into a REDOX balance model and demonstrated that an appropriate hydrogen concentration can effectively maintain the REDOX state within organisms. Transcriptome sequencing analysis revealed that hydrogen NB water modulated Cd2+ absorption and accumulation in seeds by regulating cell wall components, alleviating oxidative stress through oxidoreductase activity, and enhancing nutrient synthesis and metabolism. This collectively alleviated the inhibitory effect of Cd2+ on seed germination. This study is helpful for further understanding the effect of hydrogen concentration on the REDOX balance of seed germination, providing a theoretical basis for selecting hydrogen concentration to improve its effectiveness in agricultural fields.Environmental ImplicationCadmium is one of the most common heavy metal pollutants. It is more easily absorbed by plants and has great biological toxicity. If the cadmium concentration in soil is too high, it will cause significant inhibitory effects or serious damage to plant cell growth and division, photosynthesis, plant enzyme activity and nucleic acid metabolism, thereby inhibiting plant growth and development. Research indicates that hydrogen nanobubble water effectively alleviates the toxic effects of heavy metal stress on plants and aquatic organisms, offering new insights into addressing heavy metal pollution. This discovery holds significant importance for environmental restoration and maintaining ecological balance.

Novel non-invasive molecular signatures for oral cavity cancer, by whole transcriptome and small non-coding RNA sequencing analyses: Predicted association with PI3K/AKT/mTOR pathway.

Identification of molecular biomarkers in the saliva and serum of oral cavity cancer patients represents a first step in the development of essential and efficient clinical tools for early detection and post-treatment monitoring. We hypothesized that molecular analyses of paired saliva and serum samples from an individual would likely yield better results than analyses of either serum or saliva alone. We performed whole-transcriptome and small non-coding RNA sequencing analyses on 32 samples of saliva and serum collected from the same patients with oral squamous cell carcinoma (OSCC) and healthy controls (HC). We identified 12 novel saliva and serum miRNAs and a panel of unique miRNA and mRNA signatures, significantly differentially expressed in OSCC patients relative to HC (log2 fold change: 2.6-26.8; DE: 0.02-0.000001). We utilized a combined panel of the 10 top-deregulated miRNAs and mRNAs and evaluated their putative diagnostic potential (>87% sensitivity; 100% specificity), recommending seven of them for further validation. We also identified unique saliva and serum miRNAs associated with OSCC and smoking history (OSCC smokers vs. never-smokers or HC: log2 fold change: 22-23; DE: 0.00003-0.000000001). Functional and pathway analyses indicated interactions between the discovered OSCC-related non-invasive miRNAs and mRNAs and their targets, through PI3K/AKT/mTOR signaling. Our data support our hypothesis that using paired saliva and serum from the same individuals and deep sequencing analyses can provide unique combined mRNA and miRNA signatures associated with canonical pathways that may have a diagnostic advantage relative to saliva or serum alone and may be useful for clinical testing. We believe this data will contribute to effective preventive care by post-treatment monitoring of patients, as well as suggesting potential targets for therapeutic approaches.

Exosome-derived circUPF2 enhances resistance to targeted therapy by redeploying ferroptosis sensitivity in hepatocellular carcinoma

BackgroundAdvanced hepatocellular carcinoma (HCC) can be treated with sorafenib, which is the primary choice for targeted therapy. Nevertheless, the effectiveness of sorafenib is greatly restricted due to resistance. Research has shown that exosomes and circular RNAs play a vital role in the cancer’s malignant advancement. However, the significance of exosomal circular RNAs in the development of resistance to sorafenib in HCC remains uncertain.MethodsUltracentrifugation was utilized to isolate exosomes (Exo-SR) from the sorafenib-resistant HCC cells’ culture medium. Transcriptome sequencing and differential expression gene analysis were used to identify the targets of Exo-SR action in HCC cells. To identify the targets of Exo-SR action in HCC cells, transcriptome sequencing and analysis of differential expression genes were employed. To evaluate the impact of exosomal circUPF2 on resistance to sorafenib in HCC, experiments involving gain-of-function and loss-of-function were conducted. RNA pull-down assays and mass spectrometry analysis were performed to identify the RNA-binding proteins interacting with circUPF2. RNA immunoprecipitation (RIP), RNA pull-down, electrophoretic mobility shift assay (EMSA), immunofluorescence (IF) -fluorescence in situ hybridization (FISH), and rescue assays were used to validate the interactions among circUPF2, IGF2BP2 and SLC7A11. Finally, a tumor xenograft assay was used to examine the biological functions and underlying mechanisms of Exo-SR and circUPF2 in vivo.ResultsA novel exosomal circRNA, circUPF2, was identified and revealed to be significantly enriched in Exo-SR. Exosomes with enriched circUPF2 enhanced sorafenib resistance by promoting SLC7A11 expression and suppressing ferroptosis in HCC cells. Mechanistically, circUPF2 acts as a framework to enhance the creation of the circUPF2-IGF2BP2-SLC7A11 ternary complex contributing to the stabilization of SLC7A11 mRNA. Consequently, exosomal circUPF2 promotes SLC7A11 expression and enhances the function of system Xc- in HCC cells, leading to decreased sensitivity to ferroptosis and resistance to sorafenib.ConclusionsThe resistance to sorafenib in HCC is facilitated by the exosomal circUPF2, which promotes the formation of the circUPF2-IGF2BP2-SLC7A11 ternary complex and increases the stability of SLC7A11 mRNA. Focusing on exosomal circUPF2 could potentially be an innovative approach for HCC treatment.Graphical

Shenqi Fuzheng injection hinders non-small cell lung cancer cell growth by regulating the Bax/Bcl-2 signaling pathway

IntroductionLung cancer (LC) is the most common solid tumor and is currently considered the primary cause of cancer-related deaths worldwide. In clinical efficacy studies, it was not difficult to find that the combination of SFI and chemotherapy could improve the general condition of patients, reduce the side effects of chemotherapy drugs, and have a cooperative antitumor effect in NSCLC patients. However, whether SFI can be used as a novel antitumor drug is still unknown.MethodsFirst, meta-analysis aimed to explore the efficacy of SFI in NSCLC patients, and SFI was identified by ultra-performance liquid chromatography‒mass spectrometry (UPLC‒MS). Cell proliferation, migration, and invasion were explored by Cell Counting Kit-8 (CCK-8), scratch healing, and Transwell assays, respectively. Cell cycle and apoptosis assays were performed by flow cytometry. Transcriptome sequencing analysis was performed in four NSCLC cell lines. Differential expression analysis was used to identify potential targets. Apoptosis-related protein levels were detected by Western blotting assays. The effects of SFI in NSCLC were further investigated by mouse xenografts.ResultsSFI could markedly improve the chemotherapy efficacy of NSCLC patients. The main active ingredients include flavonoids and terpenoids, which can effectively exert antitumor effects. SFI could not only inhibit tumor cell proliferation and cell migration/invasion but also regulate the cell cycle and promote tumor cell apoptosis. In NSCLC, SFI could enhance the transcription level of the CHOP gene, thereby upregulating the expression of the proapoptotic proteins Bax and caspase 3, and inhibiting the expression of the antiapoptotic protein Bcl-2. SFI hindered the growth of mouse NSCLC xenografts in vivo.ConclusionsSFI hindered tumor progression and might promote apoptosis by increasing the expression of Bax, caspase 3 and decreasing the level of Bcl-2 in NSCLC.

Curcuminoid PBPD induces cuproptosis and endoplasmic reticulum stress in cervical cancer via the Notch1/RBP-J/NRF2/FDX1 pathway.

Curcumin has been shown to have antitumor properties, but its low potency and bioavailability has limited its clinical application. We designed a novel curcuminoid, [1-propyl-3,5-bis(2-bromobenzylidene)-4-piperidinone] (PBPD), which has higher antitumor strength and improves bioavailability. Cell counting kit-8 was used to detect cell activity. Transwell assay was used to detect cell invasion and migration ability. Western blot and quantitative polymerase chain reaction were used to detect protein levels and their messenger RNA expression. Immunofluorescence was used to detect the protein location. PBPD significantly inhibited the proliferation of cervical cancer cells, with an IC50 value of 4.16 μM for Hela cells and 3.78 μM for SiHa cells, leading to the induction of cuproptosis. Transcriptome sequencing analysis revealed that PBPD significantly inhibited the Notch1/Recombination Signal Binding Protein for Immunoglobulin kappa J Region (RBP-J) and nuclear factor erythroid 2-related factor 2 (NRF2) signaling pathways while upregulating ferredoxin 1 (FDX1) expression. Knockdown of Notch1 or RBP-J significantly inhibited NRF2 expression and upregulated FDX1 expression, leading to the inhibition of nicotinamide adenine dinucleotide phosphate activity and the induction of oxidative stress, which in turn activated endoplasmic reticulum stress and induced cell death. The overexpression of Notch1 or RBP-J resulted in the enrichment of RBP-J within the NRF2 promoter region, thereby stimulating NRF2 transcription. NRF2 knockdown resulted in increase in FDX1 expression, leading to cuproptosis. In addition, PBPD inhibited the acidification of tumor niche and reduced cell metabolism to inhibit cervical cancer cell invasion and migration. In conclusion, PBPD significantly inhibits the proliferation, invasion, and migration of cervical cancer cells and may be a novel potential drug candidate for treatment of cervical cancer.

Altered N-linked glycosylation in depression: A pre-clinical study

BackgroundNeuroimmune plays an important role in major depressive disorders (MDD). N-linked protein glycosylation (NLG) might contribute to depression by regulating the neuroinflammatory response. As microglia is the main executor of neuroimmune function in the central neural system (CNS), targeting the process of N-linked protein glycosylation of microglia in the mice used for studying depression might potentially offer new avenues for the strategy for MDD. MethodsThe chronic unpredictable mild stress (CUMS) mouse model was established for the whole brain microglia isolating. Then, RNA samples of microglia were extracted for transcriptome sequencing and mRNA analysis. Immunofluorescence (IF) was used to identify the expression level of NLG-related enzyme, B4galt1, in microglia. ResultsThe data showed that NLG was positively related to depression. Moreover, the NLG-related gene, B4galt1 increased expression in the microglia of CUMS mice. Then, the inhibition of NLG reversed the depressive behavior in CUMS mice. The expression level of B4galt1 in CUMS mice was upregulating following the NLG-inhibitor treatment. Similar results haven't been observed in neurons. Information obtained from these experiments showed increasing expression of B4galt1 in microglia following depressive-like behaviors. ConclusionsThese findings indicate that NLG in microglia is associated with MDD, and suggest that therapeutically targeting NLG might be an effective strategy for depression. LimitationsHow to modulate the B4galt1 or NLG pathways in microglia efficiently and economically request new technologies.

The prognostic genes model of breast cancer drug resistance based on single-cell sequencing analysis and transcriptome analysis

Breast cancer (BC) represents a multifaceted malignancy, with escalating incidence and mortality rates annually. Chemotherapy stands as an indispensable approach for treating breast cancer, yet drug resistance poses a formidable challenge. Through transcriptome data analysis, we have identified two sets of genes exhibiting differential expression in this context. Furthermore, we have confirmed the overlap between these genes and those associated with exosomes, which were subsequently validated in cell lines. The investigation screened the identified genes to determine prognostic markers for BC and utilized them to formulate a prognostic model. The disparities in prognosis and immunity between the high- and low-risk groups were validated using the test dataset. We have discerned different BC subtypes based on the expression levels of prognostic genes in BC samples. Variations in prognosis, immunity, and drug sensitivity among distinct subtypes were examined. Leveraging data from single-cell sequencing and prognostic gene expression, the AUCell algorithm was employed to score individual cell clusters and analyze the pathways implicated in high-scoring groups. Prognostic genes (CCT4, CXCL13, MTDH, PSMD2, and RAB27A) were subsewoquently validated using RT-qPCR. Consequently, we have established a model for predicting prognosis in breast cancer that hinges on drug resistance and ERGs. Furthermore, we have evaluated the prognostic value of this model. The genes identified as prognostic markers can now serve as a reference for precise treatment of this condition.

Transcriptome sequencing and metabolome analysis reveal the molecular mechanism of Salvia miltiorrhiza in response to drought stress

Salvia miltiorrhiza is commonly used as a Chinese herbal medicine to treat different cardiovascular and cerebrovascular illnesses due to its active ingredients. Environmental conditions, especially drought stress, can affect the yield and quality of S. miltiorrhiza. However, moderate drought stress could improve the quality of S. miltiorrhiza without significantly reducing the yield, and the mechanism of this initial drought resistance is still unclear. In our study, transcriptome and metabolome analyses of S. miltiorrhiza under different drought treatment groups (CK, A, B, and C groups) were conducted to reveal the basis for its drought tolerance. We discovered that the leaves of S. miltiorrhiza under different drought treatment groups had no obvious shrinkage, and the malondialdehyde (MDA) contents as well as superoxide dismutase (SOD) and peroxidase (POD) activities dramatically increased, indicating that our drought treatment methods were moderate, and the leaves of S. miltiorrhiza began to initiate drought resistance. The morphology of root tissue had no significant change under different drought treatment groups, and the contents of four tanshinones significantly enhanced. In all, 5213, 6611, and 5241 differentially expressed genes (DEGs) were shared in the A, B, and C groups compared with the CK group, respectively. The results of KEGG and co-expression analysis showed that the DEGs involved in plant-pathogen interactions, the MAPK signaling pathway, phenylpropanoid biosynthesis, flavonoid biosynthesis, and plant hormone signal transduction responded to drought stress and were strongly correlated with tanshinone biosynthesis. Furthermore, the results of metabolism analysis indicated that 67, 72, and 92 differentially accumulated metabolites (DAMs), including fumarate, ferulic acid, xanthohumol, and phytocassanes, which were primarily involved in phenylpropanoid biosynthesis, flavonoid biosynthesis, and diterpenoid biosynthesis pathways, were detected in these groups. These discoveries provide valuable information on the molecular mechanisms by which S. miltiorrhiza responds to drought stress and will facilitate the development of drought-resistant and high-quality S. miltiorrhiza production.

#1948 Transcriptome analysis of the effect of roxadustat on osteoclasts

Abstract Background and Aims In recent years, hypoxia-inducing factor prolyl hydroxylase inhibitors (HIF-PHIs) have been widely used in the treatment of renal anemia, bringing a revolutionary breakthrough in the treatment of anemia in CKD patients. However, HIF-PHIs acts on a variety of cells and regulates the expression of multiple target genes. Some studies have reported that HIF-PHIs plays an important role in regulating bone metabolism and promoting the differentiation and calcification of osteoblasts, but its influence on osteoclasts remains controversial. This study intends to explore the effect of roxadustat, a representative drug of HIF-PHIs, on osteoclasts. Method Bone marrow mononuclear cells (BMMCs) from 6-8 weeks old male C57BL/6 wild mice were extracted and induced to differentiate into osteoclasts using macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). Meanwhile, lipopolysaccharide (LPS) was used to simulate the inflammatory environment in CKD patients. According to the experimental group, different concentrations of roxadustat (0 μM, 2.5 μM, 5 μM, 10μM) were added simultaneously. When osteoclast induction reached day 6, mRNA was extracted for transcriptome sequencing and bioinformatics analysis. Results As shown in Table 1, different concentrations of roxadustat were given to primary bone marrow monocytes under both conditions, and there were 3 biological replicates for each sample. The reproducibility scatter plot showed that the data in each group had good repeatability (Fig. 1), and the heat map for basic difference analysis of each sample was shown in Fig. 2. The trend analysis results of the two groups were shown in Fig. 3. BMMCs group and LPS stimulated group showed two upward trends, respectively. KEGG enrichment analysis of these two upward trends showed that HIF-1, MAPK, PI3K-Akt, NF-κB and osteoclast differentiation pathways were significantly enriched in both groups. It is suggested that roxadustat can promote osteoclast differentiation and increase inflammatory response with the increase of roxadustat concentration. The enrichment analysis of the two groups showed that there was no significant enrichment pathway. Conclusion We explored the differentiation of osteoclasts under normal conditions and osteoclast differentiation under simulated inflammation in CKD patients respectively. Transcriptomic analysis results showed that HIF-1, MAPK, PI3K-Akt, NF-κB and osteoclast differentiation pathways were significantly enriched with the increase of roxadustat concentration. It is suggested that the use of roxadustat may promote bone destruction and increase the risk of inflammatory response.

Effects of UVA on Flavonol Accumulation in Ginkgo biloba

Ginkgo is an economic tree species with high medicinal value, and flavonols are its main medicinal components. This research was conducted to investigate the molecular mechanism underlying the influence of Ultraviolet A (UVA) treatment on the synthesis of ginkgo flavonols with the aim of increasing their content. Ginkgo full-sib hybrid offspring were used as test materials. The phenylalanine ammonialyase (PAL), cinnamate 4-hydroxylase (C4H), and 4-coumarate: CoA ligase (4CL) enzyme activities, as well as flavonol contents, were measured under the same intensity of white light (300 μmol·m−2·s−1) with the addition of 20, 40, and 60 μmol·m−2·s−1 UVA separately after 20 days of treatment. The control check (CK) and treatment with the highest flavonol content were chosen for transcriptome sequencing analysis. The results showed that the PAL, C4H, and 4CL enzyme activities, as well as the flavonol and totalflavonol glycoside contents, of ginkgo hybrid progeny differed significantly under different UVA treatments. They showed a tendency to increase and then decrease, reaching a maximum value under UVA-4 (40 μmol·m−2·s−1 ultraviolet UVA light intensity) treatment. Ribonucleic acid (RNA) sequencing revealed the presence of 4165 genes with differential expression, and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed that the metabolic pathways commonly enriched across all four comparison groups included ‘phenylpropanoid biosynthesis’, while the pathways commonly enriched in green-leaf ginkgo UVA-4 treatment (TL), yellow-leaf ginkgo mutant CK treatment (CKY), and green-leaf ginkgo CK treatment (CKL) were related to ‘flavonoid biosynthesis’. Treatment with UVA light led to the increased expression of PAL and 4CL enzymes in the phenylpropanoid biosynthesis pathway, as well as increased expression of chalcone synthase (CHS), Flavanone 3-hydroxylase (F3H), and flavonol synthase (FLS) enzymes in the flavonoid biosynthesis pathway, thereby promoting the synthesis of ginkgo flavonols. In summary, the use of 40 μmol·m−2·s−1 UVA treatment for 20 days significantly increased the flavonol content and the expression of related enzyme genes in ginkgo hybrid offspring, enhancing ginkgo flavonoids and increasing the medicinal value of ginkgo.

ADCY3: the pivotal gene in classical ketogenic diet for the treatment of epilepsy.

Epilepsy is a common neurological disorder characterized by recurrent epilepsy episodes. As a non-pharmacological treatment, the ketogenic diet has been widely applied in treating epilepsy. However, the exact therapeutic mechanism of the ketogenic diet for epilepsy remains unclear. This study investigates the molecular mechanisms of the ketogenic diet in regulating fatty acid metabolism and activating the ADCY3-initiated cAMP signaling pathway to enhance neuronal inhibition and thereby treat epilepsy. Meta-analysis reveals that the ketogenic diet is superior to the conventional diet in treating epilepsy. Animal experiments demonstrate that the ketogenic diet is more effective than the conventional diet in treating epilepsy, with the best results achieved using the classic ketogenic diet. Transcriptome sequencing analysis identifies six essential genes, among which ADCY3 shows increased expression in the ketogenic diet. In vivo experiments confirm that the activation of the cAMP-PKA signaling pathway by ADCY3 enhances neuronal inhibition and improves epilepsy control. Clinical observations indicate that the ketogenic diet improves patient epilepsy episodes by regulating the ADCY3-initiated cAMP signaling pathway.

Whole transcriptome RNA sequencing provides novel insights into the molecular dynamics of ovarian development in mud crab, Scylla paramamosain after mating

Ovarian development in animals is a complicated biological process, requiring the simultaneous coordination among various genes and pathways. To understand the dynamic changes and molecular regulatory mechanisms of ovarian development in mud crab (Scylla paramamosain), both histological observation and whole transcriptome sequencing of ovarian tissues at different mating stages were implemented in this study. The histological results revealed that ovarian development was delayed in unmated females (60 days after courtship behavior but not mating), who exhibited an oocyte diameter of 56.38 ± 15.17 μm. Conversely, mated females exhibited accelerated the ovarian maturation process, with females reaching ovarian stage III (proliferative stage) 23 days after mating and attained an average oocyte diameter of 132.19 ± 15.07 μm. Thus, mating process is essential in promoting the rapid ovarian development in mud crab. Based on the whole transcriptome sequencing analysis, a total of 518 mRNAs, 1502 lncRNAs, 18 circRNAs and 151 miRNAs were identified to be differentially expressed between ovarian tissues at different mating stages. Notably, six differentially expressed genes (DEGs) associated with ovarian development were identified, including ovary development-related protein, red pigment concentrating hormone receptor, G2/mitotic-specific cyclin-B3-like, lutropin-chorio gonadotropic hormone receptor, renin receptor, and SoxB2. More importantly, both DEGs and targets of differentially expressed non-coding RNAs (DEncRNAs) were enriched in renin-angiotensin system, TGF-β signaling, cell adhesion molecules, MAPK signaling pathway, and ECM-receptor interaction, suggesting that these pathways may play significant roles in the ovarian development of mud crabs. Moreover, competition endogenous RNA (ceRNA) networks were constructed while mRNAs were differentially expressed between mating stages were involved in Gene Ontology (GO) biological processes such as developmental process, reproduction, and growth. These findings could provide solid foundations for the future development of female mud crab maturation enhancement strategy, and improve the understanding of the ovarian maturation process in crustaceans.

PPARγ Antagonists Exhibit Antitumor Effects by Regulating Ferroptosis and Disulfidptosis.

Oral squamous cell carcinoma (OSCC) stands as a prevalent subtype of head and neck squamous cell carcinoma, leading to disease recurrence and low survival rates. PPARγ, a ligand-dependent nuclear transcription factor, holds significance in tumor development. However, the role of PPARγ in the development of OSCC has not been fully elucidated. Through transcriptome sequencing analysis, we discovered a notable enrichment of ferroptosis-related molecules upon treatment with PPARγ antagonist. We subsequently confirmed the occurrence of ferroptosis through transmission electron microscopy, iron detection, etc. Notably, ferroptosis inhibitors could not completely rescue the cell death caused by PPARγ inhibitors, and the rescue effect was the greatest when disulfidptosis and ferroptosis inhibitors coexisted. We confirmed that the disulfidptosis phenotype indeed existed. Mechanistically, through qPCR and Western blotting, we observed that the inhibition of PPARγ resulted in the upregulation of heme oxygenase 1 (HMOX1), thereby promoting ferroptosis, while solute carrier family 7 member 11 (SLC7A11) was also upregulated to promote disulfidptosis in OSCC. Finally, a flow cytometry analysis of flight and multiplex immunohistochemical staining was used to characterize the immune status of PPARγ antagonist-treated OSCC tissues in a mouse tongue orthotopic transplantation tumor model, and the results showed that the inhibition of PPARγ led to ferroptosis and disulfidptosis, promoted the aggregation of cDCs and CD8+ T cells, and inhibited the progression of OSCC. Overall, our findings reveal that PPARγ plays a key role in regulating cell death in OSCC and that targeting PPARγ may be a potential therapeutic approach for OSCC.

Edaravone Dexborneol ameliorates the cognitive deficits of APP/PS1 mice by inhibiting TLR4/MAPK signaling pathway via upregulating TREM2

Currently, there are no effective therapeutic agents available to treat Alzheimer's disease (AD). However, edaravone dexborneol (EDB), a novel composite agent used to treat acute ischemic stroke, has recently been shown to exert efficacious neuroprotective effects. However, whether EDB can ameliorate cognitive deficits in AD currently remains unclear. To this end, we explored the effects of EDB on AD and its potential mechanisms using an AD animal model (male APP/PS1 mice) treated with EDB for 10 weeks starting at 6 months of age. Subsequent analyses revealed that EDB-treated APP/PS1 mice exhibited improved cognitive abilities compared to untreated APP/PS1 mice. Administration of EDB in APP/PS1 mice further alleviated neuropathological alterations of the hippocampus, including Aβ deposition, pyramidal cell karyopyknosis, and oxidative damage, and significantly decreased the levels of inflammatory cytokines (IL-1β, IL-6 and TNF-α) and COX-2 in the hippocampus of APP/PS1 mice. Transcriptome sequencing analysis demonstrated the critical role of the inflammatory reaction in EDB treatment in APP/PS1 mice, indicating that the alleviation of the inflammatory reaction by EDB in the hippocampus of APP/PS1 mice was linked to the action of the TREM2/TLR4/MAPK signaling pathway. Further in vitro investigations showed that EDB suppressed neuroinflammation in LPS-stimulated BV2 cells by inhibiting the TLR4/MAPK signaling pathway and upregulating TREM2 expression. Thus, the findings of the present study demonstrate that EDB is a promising therapeutic agent for AD-related cognitive dysfunction.