Activation Pathway Research Articles

The Hippo pathway promotes platinum-based chemotherapy by inhibiting MTF1-dependent heavy metal response

The platinum-based compounds are widely used in treating various types of cancer through their heavy metal component platinum. However, the development of chemoresistance often limits their clinical effectiveness. In this study, we report the roles of heavy metal response and its associated Hippo pathway in regulating platinum-based chemotherapy. Our data show that the MTF1-dependent heavy metal response induces cancer cell resistance to platinum-based compounds both in vitro and in vivo. This resistance is mitigated by Hippo pathway-mediated phosphorylation of MTF1. Moreover, pharmacological activation of the Hippo pathway sensitizes cancer cells to platinum-based compounds. Clinically, lung adenocarcinoma (LUAD) patients with high MTF1 activity exhibit poor overall survival rates, and Hippo pathway inactivation is positively correlated with elevated MTF1 transcriptional activity in platinum-treated LUAD patients. Collectively, our findings not only unveil a critical role of the Hippo-MTF1 pathway in regulating the response to platinum-based chemotherapy, but also suggest new strategies to enhance its efficacy by targeting the heavy metal response.

Anti-Markovnikov hydroallylation reaction of alkenes via scandium-catalyzed allylic C‒H activation

Compared with rare-earth (RE)…heteroatom interaction, RE…π interaction, frequently used in facilitating regio- and stereoselectivity of olefin polymerizations, is seldomly used to trigger catalytic C − H functionalization. Here, we describe a direct anti-Markovnikov hydroallylation reaction of styrene derivatives with 1-aryl-2-alkyl alkenes and α-alkenes by use of RE…π interaction. This protocol provides a straightforward and atom-efficient route for the synthesis of valuable chain elongated internal alkenes (65 examples, up to 99% yield, > 19:1 E/Z ratio). The reaction proceeds via an allylic Csp3‒H activation pathway initiated by site-selective deprotonation with the assistance of cationic imidazolin-2-iminato scandium alkyl species followed by alkene insertion into the resulting scandium-allyl bond. A catalytic amount of Lewis base additives, such as amine and tetrahydrofuran (THF) show significant effects on the reactivity and E/Z selectivity. The reaction mechanism is elucidated by experimental studies and theoretical calculations.

Structural pathway for PI3-kinase regulation by VPS15 in autophagy.

The class III phosphatidylinositol-3 kinase complexes I and II (PI3KC3-C1 and -C2) have vital roles in macroautophagy and endosomal maturation, respectively. We elucidated a structural pathway of enzyme activation through cryo-EM analysis of PI3KC3-C1. The inactive conformation of the VPS15 pseudokinase stabilizes the inactive conformation, sequestering its N-myristate in the N-lobe of the pseudokinase. Upon activation, the myristate is liberated such that the VPS34 lipid kinase catalyzes PI3P production on membranes. The VPS15 pseudokinase domain binds tightly to guanosine triphosphate (GTP), and stabilizes a web of interactions to autoinhibit the cytosolic complex and to promote the activation upon membrane binding. These findings show in atomistic detail how the VPS34 lipid kinase is activated in the context of a complete PI3K complex.

GULP1 as a Novel Diagnostic and Predictive Biomarker in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is characterized by high recurrence and mortality, necessitating the identification of reliable biomarkers. In this study, we aimed to identify the predictive gene signatures for HCC recurrence and evaluate the efficiency of GULP PTB domain-containing engulfment adaptor 1 (GULP1) as a predictive and diagnostic marker and therapeutic target for HCC. We analyzed genomic datasets from The Cancer Genome Atlas and Gene Expression Omnibus databases via least absolute shrinkage and selection operator Cox regression and 10-fold cross-validation, leading to the development of a 15-gene risk score model, which was validated using three independent datasets. Serum GULP1 and α-fetoprotein levels were assessed to determine the diagnostic accuracy of the model. Using clinical cohorts and patient sera, GULP1 roles were examined, and functional assays in vitro and in vivo were used to evaluate its effects on cell growth, epithelial-mesenchymal transition (EMT), ADP-ribosylation factor 6 activation, and β-catenin signaling. Our newly developed risk-score model accurately predicted recurrent HCC in all datasets. Among the 15 genes in the risk score model, GULP1 was overexpressed in patients with HCC and independently predicted HCC recurrence. Its expression modulation influenced cell growth and EMT, with observed effects on ADP-ribosylation factor 6 activation and β-catenin signaling pathways. GULP1 is a crucial biomarker for HCC, serving as a non-invasive diagnostic and predictive tool. It also plays key roles in HCC progression. Our findings highlight the potential use of GULP1 in treatment strategies targeting EMT and HCC recurrence to improve the personalized care and patient outcomes.

Multi-omics investigation of high-transglutaminase production mechanisms in Streptomyces mobaraensis and co-culture-enhanced fermentation strategies

Transglutaminase (TGase) has been widely applied in the food industry. However, achieving high-yield TGase production remains a challenge, limiting its broader industrial application. In this study, a high-yield strain with stable genetic traits was obtained through UV-ARTP combined mutagenesis, achieving a maximum TGase activity of 13.77 U/mL, representing a 92.43% increase. Using this strain as a forward mutation gene pool, comparative genomic research identified 95 mutated genes, which were mostly due to base substitutions that led to changes in codon usage preference. Transcriptomic analysis revealed significant expression changes in 470 genes, with 232 upregulated and 238 downregulated genes. By investigating potential key regulatory factors, comprehensive analysis indicated that changes in codon usage preference, amino acid metabolism, carbon metabolism, protein export processes, TGase activation, and spore production pathways collectively contributed to the enhancement of TGase activity. Subsequently, the in vitro activation efficiency of TGase was further improved using co-cultivation techniques with neutral proteases secreted by Bacillus amyloliquefaciens CICC10888, and a TGase activity of 16.91 U/mL was achieved, accounting for a 22.71% increase. This study provides a comprehensive understanding of the mechanisms underlying high-yield TGase production and valuable insights and data references for future research.

EGFR inhibition augments the therapeutic efficacy of the NAT10 inhibitor Remodelin in Colorectal cancer

BackgroundColorectal cancer (CRC) is the second leading cause of cancer-related death worldwide, and treatment options for advanced CRC are limited. The regulatory mechanisms of aberrant NAT10-mediated N4-acetylcytidine (ac4C) modifications in cancer progression remains poorly understood. Consequently, an integrated transcriptomic analysis is necessary to fully elucidate the role of NAT10-mediated ac4C modifications in CRC progression.MethodsNAT10 expression levels were analyzed in CRC samples and compared with those in corresponding normal tissues. The potential mechanisms of NAT10 in CRC were investigated using RNA sequencing, RNA immunoprecipitation sequencing, and acetylated RNA immunoprecipitation sequencing. Additional in vivo and in vitro experiments, including CCK-8 assays, colony formation and mouse xenograft models, were conducted to explore the biological role of NAT10-mediated ac4C modifications. We also evaluated and optimized a potential treatment strategy targeting NAT10.ResultsWe found that NAT10 is highly expressed in CRC samples and plays a pro-oncogenic role. NAT10 knockdown led to PI3K-AKT pathway inactivation, thereby inhibiting CRC progression. However, treatment with the NAT10 inhibitor Remodelin induced only a limited and reversible growth arrest in CRC cells. Further epigenetic and transcriptomic analysis revealed that NAT10 enhances the stability of ERRFI1 mRNA by binding to its coding sequence region in an ac4C-dependent manner. NAT10 knockdown decreased ERRFI1 expression, which subsequently activated the EGFR pathway and counteracted the inhibitory effects on CRC. Based on these findings, we demonstrated that dual inhibition of NAT10 and EGFR using Remodelin and the EGFR-specific monoclonal antibody cetuximab resulted in improved therapeutic efficacy compared to either drug alone. Moreover, we observed that 5-Fluorouracil promoted the interaction between NAT10 and UBR5, which increased the ubiquitin-mediated degradation of NAT10, leading to ERRFI1 downregulation and EGFR reactivation. Triple therapy with Remodelin, cetuximab, and 5-Fluorouracil enhanced tumor regression in xenograft mouse models of CRC with wild-type KRAS, NRAS and BRAF.ConclusionsOur study elucidated the mechanism underlying 5-Fu-induced NAT10 downregulation, revealing that NAT10 inhibition destabilizes ERRFI1 mRNA through ac4C modifications, subsequently resulting in EGFR reactivation. A triple therapy regimen of Remodelin, cetuximab, and 5-Fu showed potential as a treatment strategy for CRC with wild-type KRAS, NRAS and BRAF.

Microbiota governs host chenodeoxycholic acid glucuronidation to ameliorate bile acid disorder induced diarrhea

BackgroundDisorder in bile acid (BA) metabolism is known to be an important factor contributing to diarrhea. However, the pathogenesis of BA disorder-induced diarrhea remains unclear.MethodsThe colonic BA pool and microbiota between health piglets and BA disorder-induced diarrheal piglets were compared. Fecal microbiota transplantation and various cell experiments further indicated that chenodeoxycholic acid (CDCA) metabolic disorder produced CDCA-3β-glucuronide, which is the main cause of BA disorder diarrhea. Non-targeted metabolomics uncovered the inhibition of the BA glucuronidation by Lactobacillus reuteri (L. reuteri) is through deriving indole-3-carbinol (I3C). In vitro, important gene involved in the reduction of BA disorder induced-diarrhea were screened by RNA transcriptomics sequencing, and activation pathway of FXR-SIRT1-LKB1 to alleviate BA disorder diarrhea and P53-mediated apoptosis were proposed in vitro by multifarious siRNA interference, CO-IP, immunofluorescence, and so on, which mechanism was also verified in a variety of mouse models.ResultsHere, we reveal for the first time that core microbiota derived I3C represses gut epithelium glucuronidation, particularly 3β-glucuronic CDCA production, which reaction is mediated by host UDP glucuronosyltransferase family 1 member A4 (UGT1A4) and necessary of BA disorder induced diarrhea. Mechanistically, L. reuteri derived I3C activates aryl hydrocarbon receptor to decrease UGT1A4 transcription and CDCA-3β-glucuronide content, thereby upregulating FXR-SIRT1-LKB1 signal. LKB1 binds with P53 based on protein interaction, ultimately resists to apoptosis and diarrhea. Moreover, I3C assists CDCA to attain the ameliorative effects of FXR activation in BA disorder diarrhea, through reversion of abnormal metabolism pathway, improving the outcomes of CDCA supplement.ConclusionThese findings uncover the crucial interplay between gut epithelial cells and microbes, highlighting UGT1A4-mediated conversion of CDCA-3β-glucuronide as a key target for ameliorating BA disorder-induced diarrhea.FSirWKmfeZU8YSVhEvyAL2Video

G protein peptidomimetics reveal allosteric effects and stepwise interactions in ghrelin receptor-G protein coupling.

G protein-coupled receptor (GPCR) signaling is a dynamic process involving various conformational intermediates in addition to those captured in static three-dimensional structures. Here, we used newly developed G protein peptidomimetics to characterize the interactions of the ghrelin receptor (GHSR) with G proteins. Coupling to the G protein peptidomimetic not only affected the conformational features of the cytoplasmic regions of the receptor where the G protein binds but also allosterically affected the extracellular ligand-binding pocket. These conformational and allosteric changes increased the affinity of G protein-coupled GHSR for the endogenous agonist ghrelin. In addition, our data identified different complexes along the G protein activation pathway that differed in the engagement of the Gαq C-terminal helix. Given that this helix is the main link between the activated receptor and the Gα nucleotide-binding pocket, these findings suggested a stepwise process involving distinct states in GPCR-catalyzed G protein activation. Collectively, our results provide evidence for the dynamic behavior of GPCR-G protein signaling complexes, with such dynamics most likely contributing to signaling selectivity and/or efficacy.

Fraxetin inhibits IKKβ, blocks NF-κB pathway and NLRP3 inflammasome activation, and alleviates spleen injury in sepsis.

Sepsis is a systemic inflammatory condition associated with severe organ failure, particularly splenic injury. Fraxetin (Fra), a natural product isolated from ash bark, exhibits anti-inflammatory and antioxidant properties. This study explores the function and mechanism of Fra in sepsis-induced splenic injury using an in vivo mouse model induced by Cecal Ligation and Puncture and an in vitro sepsis model based on LPS/ATP co-stimulated J774A.1 cells. The experimental groups are as follows: Sham operation or control group, Fra control group, CLP or LPS/ATP group, CLP+Fra group or LPS/ATP+Fra group, with Dexamethasone as a positive control. The results indicated that Fra improved the survival rate, inhibited bacteria burden, and reduced spleen edema. Fra also alleviated spleen necrosis, and restored the structural integrity. Blood results showed that Fra restored platelet count and lymphocyte percentage, reduced neutrophil ratio and C-reactive protein increase, and prevented lymphocyte depletion. Immunohistochemistry demonstrated that Fra inhibited MPO levels. Additionally, Fra downregulated Procalcitonin, inhibited pro-inflammatory cytokines, NO release and Arg-1 expression, illustrating its anti-inflammatory effects. DHE staining revealed that Fra inhibited ROS and MDA, enhanced CAT, GSH-PX, and SOD activities. Furthermore, Fra inhibited NLRP3 inflammasome activation, p-IKKβ expression and NF-κB pathway. Mechanistically, molecular docking studies revealed that Fra could bind to IKKβ, thereby blocking the NF-κB pathway and NLRP3 inflammasome, functioning anti-inflammatory effects. In summary, Fra targets IKKβ to block the NF-κB pathway and NLRP3 inflammasome activation, alleviating sepsis-induced splenic injury, making it a promising therapeutic strategy for treating sepsis-induced splenic injury.

Selective JAK2 pathway inhibition enhances anti-leukemic functionality in CD19 CAR-T cells

The integration of molecular targeted therapeutics with chimeric antigen receptor T (CAR-T) cell therapy represents a novel strategy to amplify the anti-tumor efficacy of immunotherapy. While CD19-targeted CAR-T cells and Janus kinase (JAK) inhibitors have independently shown efficacy against certain B-cell leukemias, such as Philadelphia chromosome-like acute lymphoblastic leukemia, the concurrent use of JAK1/2 inhibitors, such as ruxolitinib, has been implicated in reducing CAR-T cell potency by inhibiting the JAK1-dependent T cell activation pathway. This study explores the combinatorial use of a selective type II JAK2 inhibitor, CHZ868, with CD19 CAR-T cells, revealing a synergistic enhancement of anti-leukemic activity across B-cell tumor models irrespective of JAK2 mutational status. CHZ868-mediated JAK2 inhibition did not induce the exhaustion of CAR-T cells, maintaining efficacy over repeated tumor challenges and significantly extending survival in mouse models engrafted with JAK2 inhibitor-resistant leukemia cells (median survival, CD19 CAR-T + CHZ868 vs. CD19 CAR-T + DMSO: 32 days vs. 26 days, p = 0.0303). Transcriptomic analyses suggest that CHZ868 impedes CAR-T cell differentiation while preserving their proliferative capacity, a crucial factor in maintaining CAR-T cell functionality. Therefore, the selective inhibition of the JAK2 pathway may potentiate CAR-T cell therapy and offer a viable treatment strategy for patients with resistant B-cell leukemias.

Enhancing the Quality of kMI for Complex Motor Skills through Simulated Muscle Activation Color Visualization: Evidence from Time-Frequency and Functional Connectivity Analyses.

The neural-muscular activation pathways of quasi-sensory, quasi-perceptual and low "enslavement" kinesthetic motor imagery contribute to the construction of internal cognitive representations of complex motor skills. Based on the perspective that the relative duration and force of muscle activation in the generalized motor program theory facilitate the long-term perceptual memory of motor skills learning, there exists a potential possibility of enhancing the quality of Kinesthetic motor imagery by providing muscle force information in the demonstration models of complex motor skills. Therefore, this study used motion simulation technology (AnyBody Modeling System) to create complex motor skill demonstration materials and compared the EEG rhythms and functional connectivity differences between the key interest brain regions during the kinesthetic motor imagery period with and without muscle force information visual cues. In addition, the results of the VMIQ-2 measurement of the vividness of motor imagery were also measured. The research findings indicated that, under the baseline condition where there was no difference in the kinesthetic motor imagery ability before the experiment, the vividness of kinesthetic motor imagery in the experimental group during the experiment was significantly higher than that in the control group. The ERSP results revealed that both groups exhibited the ERS phenomenon in the frontal, central, and temporal brain regions, and the ERD phenomenon in the parieto-occipital brain regions. However, the ERS value of the experimental group was lower than that of the control group, while the ERD value was higher than that of the control group. The functional connectivity results indicated that the experimental group enhanced the connectivity of the occipital-right parietal (V1-R PPC) and right frontal-right parietal (R DLPFC-R PPC) brain regions. Conclusion: The muscle force information enhanced the vividness of kinesthetic motor imagery, which was associated with the weakened selective inhibition in the frontal-central-temporal brain regions and the enhanced activation in the occipital-parietal brain regions in the brain rhythm information, and it also strengthened the information circulation between the occipital-parietal and frontal-parietal brain regions.

Platelet related gene IQGAP1 contributes to the onset and abnormal immune landscape of ischemic stroke patients.

Ischemic stroke (IS) is a complex illness resulting from a combination of numerous environmental and genetic risk factors. Recent reports have shed light on the vital role that platelets play in the pathophysiology of IS. Here, we aimed to explore the potential platelet-related genes in IS and investigate the effect of platelet-related genes in the immune microenvironment of IS. The data of IS were retrieved from the Gene Expression Omnibus database. Firstly, we screened the platelet-related genes that were correlated with the onset of IS using differential expression analysis, enrichment analyses, and protein-protein interaction (PPI) network. Moreover, we analyzed the clinical value and functional information of platelet-related genes in IS. Finally, we explored the correlation between platelet-related genes and immune cells' infiltration. Ten platelet-related genes that were correlated with the onset of IS were identified, among which IQGAP1 was located at the core of the PPI network. IQGAP1 was found to be expressed in the normal brain tissue and its expression was significantly elevated in IS samples. The area under the curve (AUC) values for IQGAP1 in both the GSE16561 and GSE58294 datasets were close to 1. IQGAP1 knockdown might increase OGD/R‑induced HT22 cell viability. Additionally, FoxO signaling pathway, NOD-like receptor signaling pathway, Phagosome and Platelet activation pathways were significantly activated in IS patients with high IQGAP1 expression compared to those with low IQGAP1 expression. The IS patients in the IQGAP1high and IQGAP1low groups showed dramatically different proportions of immune cells and immune-related functions, and the IQGAP1 expression was correlated with the immune cell' infiltration in IS. In this study, we identified the IQGAP1 might serve as a potential diagnostic marker for IS, and the IQGAP1 expression was very relevant in determining the immune cell' infiltration in IS patients.

Modulating the complement system through epitope-specific inhibition by complement C3 inhibitors.

As an integral part of the innate immune system, the complement system is a tightly regulated proteolytic cascade, playing a critical role in microbial defense, inflammation activation, and dying host cell clearance. Complement proteins are now emerging as subjects of intense research and drug development, since dysregulation of the complement system plays a critical role in several diseases and disorders, such as Paroxysmal Nocturnal Hemoglobinuria (PNH) and Geographic Atrophy (GA). Within the complement cascade, complement C3 is the central component, situated at the convergence of all complement activation pathways, rendering it an attractive target for complement-related diseases. However, due to the complicated Structure-Activity Relationship (SAR) of C3, elucidating the mechanisms of C3 inhibition on diverse epitopes is the basis for the rational design of C3-targeted therapeutics. Here, we have developed a set of comprehensive biochemical assays that are tailored to the specific steps within the complement cascade, allowing for a thorough understanding of the pharmacological consequences of different C3 inhibitors at each stage. Utilizing three Model Inhibitors (MIs) with different epitopes, we found that inhibition of MG4/MG5 domains has potent inhibition efficacy across all the complement activation pathways by interrupting C3-C3 convertase interaction, while inhibition of C345C domain displays a bias over the Alternative Pathway (AP) inhibition by impairing AP C3 proconvertase formation. This study elucidates the intricate impact of C3 inhibition by targeting different epitopes, offering valuable insights into understanding the mechanism and facilitating the rational design of C3-targeted therapeutics.

Chemotaxonomy and Bioactive Potential of High-Mountain Plantago atrata.

Plantago atrata Hoppe is a high-altitude mountain plant exposed to harsh environmental factors. This study aims to elucidate the ecological, phytochemical and pharmacological characteristics of this lesser-known plantain. Despite nutrient-poor peat soil, the leaves of P. atrata contained increased levels of phosphorus, potassium and calcium, and the anatomy revealed an isobilateral mesophyll. Molecular taxonomic identification and phylogenetic analyses confirmed the classification of the studied plant as P. atrata and its clustering with narrow-leaved plantains growing in extreme conditions. Detailed phytochemical profiles revealed primary and secondary metabolites that can be used as taxonomic identifiers and for bioactivity studies. The methanol fraction enriched in phenolic compounds appeared biologically active - by displaying limited antimicrobial activity, however, it possessed significant radical-scavenging and anti-inflammatory capacity. The data demonstrate that P. atrata's leaf parameters and phytochemical arsenal assure plant survival and reveal the plantain's pharmaceutical potential in inhibiting inflammation via the classical pathway of Complement activation.

Tofacitinib Treatment for Active Dermatomyositis and Anti-synthetase Syndrome: A Prospective Cohort Pilot Study.

The objective of this study was to evaluate the efficacy and safety of tofacitinib in the treatment of active dermatomyositis (DM) and anti-synthetase syndrome (ASS). Tofacitinib was administered at a dose of 5 mg twice daily to patients who exhibited inadequate response to conventional treatments. The primary end point was the reduction of T follicular helper (Tfh) cells at week 24. Key secondary endpoints included clinical scores. Moreover, we analyzed the immunological profiles and conducted RNA sequencing (RNAseq) on peripheral blood samples from four patients. A total of twenty-six patients were enrolled, with twenty-one completing the study. Both DM and ASS patients demonstrated significant improvements in disease activity. Among these, the percentage of Tfh cells in peripheral blood decreased by 81.0% (17/21) of patients (p = 0.003). Significant reductions in Th17 cells were observed in vivo in the PBMCs of these patients (p = 0.017). In vitro, Tfh cells (2.88 ± 1.13 vs 2.28 ± 0.92, p < 0.001), Th17 cells (1.42 ± 0.92 vs 1.01 ± 0.74, p = 0.016), Treg cells (2.06 ± 1.26 vs 0.98 ± 0.65, p = 0.019) and Tfh17 cells (33.38 ± 15.14 vs 30.28 ± 4.89, p = 0.014) were inhibited. RNAseq analysis revealed significant downregulation of genes associated with the 'herpes simplex virus 1 infection' and 'IL-17 signalling' pathway. MDAAT scores improved in 21 out of 24 patients. Fifteen (62.5%) patients met the criteria for IMACS DOI. Importantly, no severe adverse events necessitated treatment discontinuation. Tofacitinib demonstrated significant immunologic and clinical effectiveness in DM and ASS patients, reducing key immune cell populations and downregulating immune activation pathways.

From Teeth to Body: The Complex Role of Streptococcus mutans in Systemic Diseases.

Streptococcus mutans, the principal pathogen associated with dental caries, impacts individuals across all age groups and geographic regions. Beyond its role in compromising oral health, a growing body of research has established a link between S. mutans and various systemic diseases, including immunoglobulin A nephropathy (IgAN), nonalcoholic steatohepatitis (NASH), infective endocarditis (IE), ulcerative colitis (UC), cerebral hemorrhage, and tumors. The pathogenic mechanisms associated with S. mutans frequently involve collagen-binding proteins (CBPs) and protein antigens (PA) present on the bacterial surface. These components facilitate intricate interactions with the host immune system, thereby potentially contributing to various pathological processes. Specifically, CBP is implicated in the deposition of IgA and complement component C3, which exhibits characteristics reminiscent of IgAN-like lesions through animal models, recent clinical studies suggest a potential involvement of S. mutans in IgAN. In addition, CBP binds to complement component C1q, effectively inhibiting the classical activation pathway of the complement system. In addition, CBP promotes the induction of host cells to produce interferon-gamma (IFN-γ). Furthermore, CBP leads to direct inhibitory effects on platelets and the activation of matrix metalloproteinase-9 (MMP-9) at sites of vascular injury.Moreover, PA enhances the ability of S. mutans to invade hepatic tissue. Through utilization of its PAc, S. mutans excessively produces kynurenine (KYNA), which promotes the development and progression of oral squamous cell carcinoma (OSCC). This article synthesizes the latest advancements in understanding the mechanisms of intricate interactions between S. mutans and various systemic conditions in humans, expanding our perspective beyond the traditional focus on dental caries.

Unveiling the functional connectivity of astrocytic networks with AstroNet, a graph reconstruction algorithm coupled to image processing

Astrocytes form extensive networks with diverse calcium activity, yet the organization and connectivity of these networks across brain regions remain largely unknown. To address this, we developed AstroNet, a data-driven algorithm that uses two-photon calcium imaging to map temporal correlations in astrocyte activation. By organizing individual astrocyte activation events chronologically, our method reconstructs functional networks and extracts local astrocyte correlations. We create a graph of the astrocyte network by tallying direct co-activations between pairs of cells along these activation pathways. Applied to the CA1 hippocampus and motor cortex, AstroNet reveals notable differences: astrocytes in the hippocampus display stronger connectivity, while cortical astrocytes form sparser networks. In both regions, smaller, tightly connected sub-networks are embedded within a larger, loosely connected structure. This method not only identifies astrocyte activation paths and connectivity but also reveals distinct, region-specific network patterns, providing new insights into the functional organization of astrocytic networks in the brain.

Single-cell sequencing of human Langerhans cells identifies altered gene expression profiles in patients with atopic dermatitis.

Atopic dermatitis (AD) is characterized by dysregulated T cell immunity and skin microbiome dysbiosis with predominance of Staphylococcus aureus, which is associated with exacerbating AD skin inflammation. Specific glycosylation patterns of S. aureus cell wall structures amplify skin inflammation through interaction with Langerhans cells (LCs). Nevertheless, the role of LCs in AD remains poorly characterized. Here, we performed single cell RNA sequencing of primary epidermal LCs and dermal T cells, isolated from skin biopsies of AD patients and healthy control subjects, alongside specific glycoanalysis of S. aureus strains isolated from the AD lesions. Our findings revealed 4 LC subpopulations ie, 2 steady-state clusters [LC1 and LC1H] and 2 proinflammatory/matured subsets [LC2 and migratory LCs]. The latter 2 subsets were enriched in AD skin. AD LCs showed enhanced expression of C-type lectin receptors, the high-affinity IgE receptor, and activation of prostaglandin and leukotriene biosynthesis pathways, upregulated transcriptional signatures related to T cell activation pathways, and increased expression of CCL17 compared with healthy LCs. Correspondingly, T helper 2 and T regulatory cell populations were increased in AD lesions. Complementary, we performed bulk RNA sequencing of primary LCs stimulated with the S. aureus strains isolated from the AD lesions, which showed upregulation of T helper 2-related pathways. Our study provides proof-of-concept for a role of LCs in connecting the S. aureus-T cell axis in the AD inflammatory cycle.

Integrated analysis of chromatin and transcriptomic profiling of the striatum after cerebral hypoperfusion in mice

BackgroundVascular cognitive impairment (VCI) is a significant contributor to dementia, yet the precise mechanisms underlying the cognitive decline associated with chronic cerebral hypoperfusion (CCH) remain unclear. This study investigated the molecular and epigenetic changes in the striatum, a brain region critical for motor function and cognition, following chronic hypoperfusion using a bilateral common carotid artery stenosis (BCAS) model in mice.MethodsRNA-seq was utilized to identify differentially expressed genes (DEGs) associated with hypoperfusion. In parallel, ATAC-seq was used to assess changes in chromatin accessibility within the striatum, providing insight into the epigenome and potential regulatory mechanisms. The integration of these datasets allowed us to correlate chromatin accessibility with transcriptional activity and to identify key transcription factors driving the observed gene expression changes.ResultsAnalysis of striatum-specific transcriptome revealed significant upregulation of immune response genes, particularly type II interferon signaling, and downregulation of neural activation pathways. Analysis of striatum-specific epigenome showed increased chromatin accessibility at promoters of immune-related genes. Integrated analysis highlighted PU.1 as a key transcription factor in upregulated pathways, while neural pathways lacked epigenetic regulation, revealing distinct molecular responses in the striatum following chronic hypoperfusion.ConclusionsOur findings indicate that upregulated pathways in the striatum following BCAS-induced CCH are driven by epigenetic changes, while downregulated pathways occur independently of these modifications. Additionally, PU.1 plays a critical role in mediating immune responses, offering a potential target for therapeutic intervention.

Salivary proteomic analysis in patients with type 2 diabetes mellitus and periodontitis.

This study aimed to compare the salivary protein profile in individuals with Type 2 Diabetes Mellitus (DM2) and periodontitis and their respective controls. Eighty participants were included in the study. The four groups were formed by individuals with DM2 and periodontitis (DM2 + P, n = 20), DM2 without periodontitis (DM2, n = 20), periodontitis without DM2 (P, n = 20) and individuals without periodontitis and without DM2 (H, n = 20). Periodontal clinical examinations were performed and unstimulated saliva was collected. Proteomic analysis was performed by shotgun mass spectrometry. The results were obtained by searching the Homo sapiens database of the UniProt catalog. A total of 220 proteins were identified in saliva samples. In the comparison between DM2 + P and DM2 groups, 27 proteins were up-regulated [e.g. S100-A8 was 6 times up-regulated (humoral immune response pathway)]. The DM2 + P and P groups had 26 up-regulated proteins [e.g. Immunoglobulin lambda constant 7 more than 2 times up-regulated (complement activation pathway)]. The non-DM2 groups (P and H) presented 22 up-regulated proteins [e.g. Glyceraldehyde-3-phosphate dehydrogenase more than 2 times up-regulated (Peptidyl-cysteine S-nitrosylation pathway)]. The groups without periodontitis (DM2 and H) showed 23 were up-regulated proteins [e.g. Hemoglobin subunit alpha that was more than 10 times up-regulated (cellular oxidant detoxification pathway)]. The presence of DM2 and periodontitis significantly impacts the salivary proteome. Our proteomic analysis demonstrated that changes in the S100 family proteins (S100A8 and S100 A9) are highly related to the presence of DM2 and periodontitis. Diabetes Mellitus (DM) and periodontitis are highly prevalent chronic diseases that present a wide variety of signs and symptoms. They present a bidirectional relationship, where patients with DM have a higher prevalence and severity of periodontitis, and patients with periodontitis have a higher prevalence of DM, worse glycemic control, and more diabetic complications. Diagnosing periodontitis requires specific clinical examinations, which require a highly trained operator. In this study, we used high throughput proteomics in order to evaluate non-invasive biomarkers for periodontitis in type 2 DM subjects. The results can contribute to earlier, more accurate, and less costly diagnosis of periodontitis in diabetic subjects, enabling better diabetes control.