Factor Activity Research Articles

The KAT module of the SAGA complex maintains the oncogenic gene expression program in MYCN-amplified neuroblastoma.

Pediatric cancers are frequently driven by genomic alterations that result in aberrant transcription factor activity. Here, we used functional genomic screens to identify multiple genes within the transcriptional coactivator Spt-Ada-Gcn5-acetyltransferase (SAGA) complex as selective dependencies for MYCN-amplified neuroblastoma, a disease of dysregulated development driven by an aberrant oncogenic transcriptional program. We characterized the DNA recruitment sites of the SAGA complex in neuroblastoma and the consequences of loss of SAGA complex lysine acetyltransferase (KAT) activity on histone acetylation and gene expression. We demonstrate that loss of SAGA complex KAT activity is associated with reduced MYCN binding on chromatin, suppression of MYC/MYCN gene expression programs, and impaired cell cycle progression. Further, we showed that the SAGA complex is pharmacologically targetable in vitro and in vivo with a KAT2A/KAT2B proteolysis targeting chimeric. Our findings expand our understanding of the histone-modifying complexes that maintain the oncogenic transcriptional state in this disease and suggest therapeutic potential for inhibitors of SAGA KAT activity in MYCN-amplified neuroblastoma.

Characterization of extracellular vesicles from patients with severe aortic stenosis undergoing transcatheter aortic valve implantation and assessment of their coagulation potential

Abstract Funding Acknowledgements None. Background The pathogenesis of aortic stenosis (AS) includes processes of inflammation and calcification. Patients with aortic stenosis have increased risk of stroke. The amount of circulating extracellular vesicles (EVs) is elevated in patients with severe AS and decreases following valve intervention. The function of these EVs in the clinical setting of severe AS is not fully known. Aim With flow cytometry characterize EVs in patients with severe AS before and after Transcatheter Aortic Valve Implantation (TAVI). To investigate whether these EVs have a procoagulant effects by themselves or when incubated with human aortic endothelial cells (hAoECs), human cardiac microvascular endothelial cells (hCMVEC), human cardiac myocytes (hCM) and spheroids of hCMVEC and hCM. Methods 25 patients with severe AS were included in the study and blood samples collected before and 2 months after TAVI. The CytoFLEX flow cytometer (Beckman Coulter) was used for characterization of surface antigens on purified EVs. 2D-cultures of hAoECs, hCMVEC and hCM were incubated with EVs (100 mg/mL) for 6 hours (h) and spheroids of hCM and hCMVEC with EVs for 48h. An inhouse assay determining Tissue Factor (TF) activity by means of FXa generation in cell lysates were utilized. TF activity in purified EVs alone was determined using Tissue Factor Activity Assay kit (Human, colorimetric) from abcam. Results We found an increased level of TF positive EVs, phosphatidylserine (PS) positive EVs as well as double positive PS / TF expressing EVs in AS patients before TAVI compared to healthy controls and reduction of PS+ and TF+/PS+ expressing EVs after TAVI (Fig 1). When analyzing TF activity by means of FXa generation we found no procoagulant effect in the purified EVs alone nor when incubating EVs with hAoEC or hCM. In contrast, we found an increased TF activity in hCMVEC and spheroids of hCM and hCMVEC after incubation with EVs from AS patients compared to healthy EVs. Conclusion Patients with AS have increased amount of EVs expressing typically procoagulant surface antigens. The EVs by themselves have no procoagulant activity. However, upon interaction with hCMVEC and spheroids, consisting of hCMVEC and hCM, EVs appear to induce procoagulant activity in the recipient cells thus suggesting a possible link to thromboembolic events in coronary arteries. Additional studies are warranted to elaborate the mechanism and function of these TF, PS and TF / PS positive EVs.

Immunoinhibitory effects of anti-tuberculosis therapy induce the host vulnerability to tuberculosis recurrence.

The host immune responses play a pivotal role in the establishment of long-term memory responses, which effectively aids in infection clearance. However, the prevailing anti-tuberculosis therapy, while aiming to combat tuberculosis (TB), also debilitates innate and adaptive immune components of the host. In this study, we explored how the front-line anti-TB drugs impact the host immune cells by modulating multiple signaling pathways and subsequently leading to disease relapse. Administration of these drugs led to a reduction in innate immune activation and also the cytokines required to trigger protective T cell responses. Moreover, these drugs led to activation-induced cell death in the mycobacterial-specific T cell leading to a reduced killing capacity. Furthermore, these drugs stalled the T cell differentiation into memory subsets by modulating the activation of STAT3, STAT4, FOXO1, and NFκB transcription factors and hampering the Th1 and Th17-mediated long-term host protective memory responses. These findings suggest the urgent need to augment directly observed treatment, short-course (DOTS) therapy with immunomodulatory agents to mitigate the adverse effects linked to the treatment.IMPORTANCEAs a central component of TB eradication initiatives, directly observed treatment, short-course (DOTS) therapy imparts immune-dampening effects during the course of treatment. This approach undermines the host immune system by delaying the activation process and lowering the immune response. In our investigation, we have unveiled the impact of DOTS on specific immune cell populations. Notably, the signaling pathways involving STAT3 and STAT4 critical for memory responses and NFκβ associated with pro-inflammation were substantially declined due to the therapy. Consequently, these drugs exhibit limited effectiveness in preventing recurrence of the disease. These observations highlight the imperative integration of immunomodulators to manage TB infection.

UCHL1-dependent control of hypoxia-inducible factor transcriptional activity during liver fibrosis.

Liver fibrosis is the excessive accumulation of extracellular matrix proteins that occurs in most types of chronic liver disease. At the cellular level, liver fibrosis is associated with the activation of hepatic stellate cells (HSCs) which transdifferentiate into a myofibroblast-like phenotype that is contractile, proliferative and profibrogenic. HSC transdifferentiation induces genome-wide changes in gene expression that enable the cell to adopt its profibrogenic functions. We have previously identified that the deubiquitinase ubiquitin C-terminal hydrolase 1 (UCHL1) is highly induced following HSC activation; however, the cellular targets of its deubiquitinating activity are poorly defined. Here, we describe a role for UCHL1 in regulating the levels and activity of hypoxia-inducible factor 1 (HIF1), an oxygen-sensitive transcription factor, during HSC activation and liver fibrosis. HIF1 is elevated during HSC activation and promotes the expression of profibrotic mediator HIF target genes. Increased HIF1α expression correlated with induction of UCHL1 mRNA and protein with HSC activation. Genetic deletion or chemical inhibition of UCHL1 impaired HIF activity through reduction of HIF1α levels. Furthermore, our mechanistic studies have shown that UCHL1 elevates HIF activity through specific cleavage of degradative ubiquitin chains, elevates levels of pro-fibrotic gene expression and increases proliferation rates. As we also show that UCHL1 inhibition blunts fibrogenesis in a pre-clinical 3D human liver slice model of fibrosis, these results demonstrate how small molecule inhibitors of DUBs can exert therapeutic effects through modulation of HIF transcription factors in liver disease. Furthermore, inhibition of HIF activity using UCHL1 inhibitors may represent a therapeutic opportunity with other HIF-related pathologies.

Flexible modeling of regulatory networks improves transcription factor activity estimation

Transcriptional regulation plays a crucial role in determining cell fate and disease, yet inferring the key regulators from gene expression data remains a significant challenge. Existing methods for estimating transcription factor (TF) activity often rely on static TF-gene interaction databases and cannot adapt to changes in regulatory mechanisms across different cell types and disease conditions. Here, we present a new algorithm - Transcriptional Inference using Gene Expression and Regulatory data (TIGER) - that overcomes these limitations by flexibly modeling activation and inhibition events, up-weighting essential edges, shrinking irrelevant edges towards zero through a sparse Bayesian prior, and simultaneously estimating both TF activity levels and changes in the underlying regulatory network. When applied to yeast and cancer TF knock-out datasets, TIGER outperforms comparable methods in terms of prediction accuracy. Moreover, our application of TIGER to tissue- and cell-type-specific RNA-seq data demonstrates its ability to uncover differences in regulatory mechanisms. Collectively, our findings highlight the utility of modeling context-specific regulation when inferring transcription factor activities.

Non-transcriptional regulatory activity of SMAX1 and SMXL2 mediates karrikin-regulated seedling response to red light in Arabidopsis

Karrikins and strigolactones govern plant development and environmental responses through closely related signaling pathways. The transcriptional repressor proteins SUPPRESSOR OF MAX2 1 (SMAX1), SMAX1-like2 (SMXL2), and D53-like SMXLs mediate karrikin and strigolactone signaling by directly binding downstream genes or by inhibiting the activities of transcription factors. In this study, we characterized the non-transcriptional regulatory activities of SMXL proteins in Arabidopsis. We discovered that SMAX1 and SMXL2 with mutations in their ethylene-response factor-associated amphiphilic repression (EAR) motif had undetectable or weak transcriptional repression activities but still partially rescued the hypocotyl elongation defects and fully reversed the cotyledon epinasty defects of the smax1 smxl2 mutant. SMAX1 and SMXL2 directly interact with PHYTOCHROME INTERACTION FACTOR 4 (PIF4) and PIF5 to enhance their protein stability by interacting with phytochrome B (phyB) and suppressing the association of phyB with PIF4 and PIF5. The karrikin-responsive genes were then identified by treatment with GR24ent−5DS, a GR24 analog showing karrikin activity. Interestingly, INDOLE-3-ACETIC ACID INDUCIBLE 29 (IAA29) expression was repressed by GR24ent−5DS treatment in a PIF4- and PIF5-dependent and EAR-independent manner, whereas KARRIKIN UPREGULATED F-BOX 1 (KUF1) expression was induced in a PIF4- and PIF5-independent and EAR-dependent manner. Furthermore, the non-transcriptional regulatory activity of SMAX1, which is independent of the EAR motif, had a global effect on gene expression. Taken together, these results indicate that non-transcriptional regulatory activities of SMAX1 and SMXL2 mediate karrikin-regulated seedling response to red light.

A novel combinatorial approach usingsulforaphane- and withaferin A-rich extracts for prevention of estrogen receptor-negative breast cancer through epigenetic and gut microbial mechanisms.

Estrogen receptor-negative [ER(-)] mammary cancer is the most aggressive type of breast cancer (BC) with higher rate of metastasis and recurrence. In recent years, dietary prevention of BC with epigenetically active phytochemicals has received increased attention due to its feasibility, effectiveness, and ease of implementation. In this regard, combinatorial phytochemical intervention enables more efficacious BC inhibition by simultaneously targeting multiple tumorigenic pathways. We, therefore, focused on investigation of the effect of sulforaphane (SFN)-rich broccoli sprouts (BSp) and withaferin A (WA)-rich Ashwagandha (Ash) combination on BC prevention in estrogen receptor-negative [ER(-)] mammary cancer using transgenic mice. Our results indicated that combinatorial BSp + Ash treatment significantly reduced tumor incidence and tumor growth (~ 75%) as well as delayed (~ 21%) tumor latency when compared to the control treatment and combinatorial BSp + Ash treatment was statistically more effective in suppressing BC compared to single BSp or Ash intervention. At the molecular level, the BSp and Ash combination upregulated tumor suppressors (p53, p57) along with apoptosis associated proteins (BAX, PUMA) and BAX:BCL-2 ratio. Furthermore, our result indicated an expressional decline of epigenetic machinery HDAC1 and DNMT3A in mammary tumor tissue because of combinatorial treatment. Interestingly, we have reported multiple synergistic interactions between BSp and Ash that have impacted both tumor phenotype and molecular expression due to combinatorial BSp and Ash treatment. Our RNA-seq analysis results also demonstrated a transcriptome-wide expressional reshuffling of genes associated with multiple cell-signaling pathways, transcription factor activity and epigenetic regulations due to combined BSp and Ash administration. In addition, we discovered an alteration of gut microbial composition change because of combinatorial treatment. Overall, combinatorial BSp and Ash supplementation can prevent ER(-) BC through enhanced tumor suppression, apoptosis induction and transcriptome-wide reshuffling of gene expression possibly influencing multiple cell signaling pathways, epigenetic regulation and reshaping gut microbiota.

Xiaowugui decoction alleviates experimental rheumatoid arthritis by suppressing Rab5a-mediated TLR4 internalization in macrophages

BackgroundRheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by exacerbated synovial inflammation and joint destruction. Recent studies suggest toll-like receptor 4 (TLR4) internalization facilitate inflammatory response of macrophage. The role of TLR4 internalization in the pathogenesis of RA is unknown. PurposeTo investigate the role and mechanism of TLR4 internalization in macrophage inflammatory response of RA and explore whether TLR4 internalization mediates the anti-arthritic effect of Xiaowugui (XWG) decoction, a patented herbal formula used in China. MethodsThe co-expression of TLR4 and the internalization marker, early endosome antigen 1 (EEA1), in the synovial samples of RA patients and joint tissue of collagen-induced arthritis (CIA) mice, were evaluated using immunofluorescence. The effect of Rab5a-mediated early internalization of TLR4 on the activation induced by lipopolysaccharide (LPS) in RAW264.7 cells was investigated using small interfering RNAs that act against Rab5a. CIA was induced in Rab5a−/− mice to evaluate the role of Rab5a in vivo. The disease progression and expression of Rab5a and TLR4 in the joint tissue were evaluated in CIA mice treated with XWG. Inflammatory factors production, TLR4 internalization, and activation of downstream signaling pathways were examined in RAW264.7 cells treated with XWG in vitro. ResultsThe co-expression and co-localization of TLR4 and EEA1 were elevated in the synovial samples of RA patients and joint tissue of CIA mice. Pharmaceutical inhibition of TLR4 internalization reduced macrophages inflammatory responses induced by LPS. The co-expression and co-localization of Rab5a and TLR4 were significantly increased in macrophages treated with LPS. Silencing Rab5a reduced LPS-induced TLR4 internalization, inflammatory factors production, and phosphorylation of Jun N-terminal kinases (JNK) and p65. Genetic deletion of Rab5a inhibited TLR4 internalization and the development of arthritis in vivo. The co-expression of TLR4 and Rab5a was also elevated in the synovial samples of RA patients. XWG treatment of mice with CIA alleviated arthritis and reduced the co-expression of Rab5a and TLR4 in the joint tissue. XWG treatment of macrophage inhibited LPS-induced IL-6 and TNF-α production, co-expression of Rab5a and TLR4, and phosphorylation of JNK and p65. ConclusionsOur findings highlight the pathogenic role of TLR4 internalization in patients with RA and identify a novel Rab5a-dependent internalization pathway that promotes macrophage inflammatory response. XWG treatment demonstrated outstanding therapeutic effects in experimental arthritis, and targeting the Rab5a-mediated internalization of TLR4 may be the main underlying mechanism.

Single cell RNA sequencing of human eosinophils from nasal polyps reveals eosinophil heterogeneity in chronic rhinosinusitis tissue

BackgroundChronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by type 2 inflammation in the US, yet the actual roles of eosinophils in CRSwNP are largely unclear. ObjectiveTo reveal the roles and heterogeneity of eosinophils in nasal polyp (NP) tissue by single cell RNA-Sequencing (scRNA-Seq) analysis of NP tissue. MethodsSinonasal tissues (NP and control sinus tissue) and patient matched peripheral blood (PB) samples were obtained from 5 control patients and 5 patients with CRSwNP. Eosinophils were enriched prior to processing for scRNA-Seq. The gene expression profiles in eosinophils were determined by the microwell-based scRNA-Seq technology (BD Rhapsody platform). We predicted the overall function of NP eosinophils by gene ontology (GO) enrichment and pathway analyses and confirmed expression of selected genes by flow cytometry. ResultsAfter filtering out contaminating cells, we detected 5,542 eosinophils from control PB, 3,883 eosinophils from CRSwNP PB, 101 eosinophils from control sinus tissues (not included in further analyses) and 9,727 eosinophils from NPs by scRNA-Seq. We found that 204 genes were down-regulated, and 354 genes were up-regulated in NP eosinophils, compared to all PB eosinophils (>1.5-fold, Padj<0.05). Up-regulated genes in NP eosinophils were associated with activation, cytokine-mediated signaling, growth factor activity, NF-κB signaling and anti-apoptotic molecules. NP eosinophils displayed 4 clusters revealing potential heterogeneity of eosinophils in NP tissue. ConclusionsElevated eosinophils in NP tissue appear to exist in several subtypes that may play important pathogenic roles in CRSwNP, in part via controlling inflammation and hyperproliferation of other cells.

The use of neuroleptics, sedatives and anesthetics for anxiolytic therapy in animals

An important area for improving antidepressant treatment is the expansion of the indications for neuroleptics, anesthetics and sedatives. All these drugs have pronounced neurotropic effects. The aim of our work is to study the published scientifc evidence on the potential of neuroleptics, anesthetics and sedatives for anxiolytic therapy. A systematic literature review was used to search for publications on the topic of the study. The PubMed database (https://pubmed.ncbi.nlm.nih.gov) was used to search for scientifc articles. The literature data indicates that ketamine is used to treat recurrent anxiety in both humans and animals. It provides rapid and sustained relief of anxiety symptoms in a variety of its clinical presentations. The anxiolytic effect occurs within the frst 12 hours after administration and remains effective for 1 to 2 weeks. The anxiolytic effect of ketamine is due to its effect on hippocampal neurotropic factor activity. Diazepam can signifcantly reduce anxiety and depressive symptoms as well as neuroinflammation in brain-injured mice. It causes a dose-dependent increase in motor activity. In combination with metformin, diazepam is the preferred treatment for type 2 diabetes mellitus in stressed animals. Intermittent use of diazepam is useful to avoid the development of physical dependence when the drug is used for a long period of time. Acepromazine is promising to be used for anxiolytic effect in animals. A combined protocol with acepromazine signifcantly reduced signs of stress, anxiety and aggression during veterinary visits and had an anxiolytic effect in dogs. Acepromazine reduces the negative effects of transport stress in wild ungulates. Dexmedetomidine is used for sedation in both human and veterinary medicine. This drug is a promising candidate for the experimental treatment of stress-related diseases such as anxiety disorders or post-traumatic stress disorder. We believe that further research in this area should be conducted in controlled comparative studies to determine the optimal doses and duration of administration of potential anxiolytics, considering the species, age, sex, physiological state and other relevant clinical parameters of the animals studied. Therefore, systematic and detailed studies will help us not only to understand the effectiveness, but also to provide safe and individualized treatment. This research can improve our understanding of the use of anxiolytics in veterinary practice, which is of key importance for improving the quality of animal welfare. In our opinion, the use of anesthetics, neuroleptics and sedatives, which are now widely used for sedation or general anesthesia, opens new possibilities for the treatment of behavioral and anxiety disorders in animals. Key words: anxiety, veterinary medicine, neuroleptics, sedatives, ketamine, acepromazine, diazepam, medetomidine.

#894 Dynamic multi-omics and mechanistic modelling approach provides deeper insight into kidney fibrosis progression

Abstract Background and Aims Chronic kidney disease (CKD) affects more than 10% of the world's population and causes millions of deaths annually. Organ fibrosis is the main driver of CKD, leading to excessive accumulation of extracellular matrix (ECM). The presence of myofibroblasts correlates with disease progression. The pro-fibrotic responses of these cells, such as ECM production, are driven by the master regulator TGF-β. Despite progress in understanding the disease, there are no treatments available, and current diagnostic indicators neither facilitate early detection of CKD nor correlate with actual renal damage [1–4] Using human kidney-derived PDGFRβ+ mesenchymal cells we aim to provide time-resolved mechanistic insight into CKD progression. Method Our goal is to understand and model how these cells contribute to fibrosis by studying their kinetic response to TGF-β. Therefore, we use multiple omics measurements: transcriptomics, secretomics and phospho-/proteomics coupled with quantitative imaging of the ECM. For each time point, we quantified differential changes in omics and estimated the activities of transcription factors, kinases and phosphatases [5]. Furthermore, we used causal reasoning to draw mechanistic relationships between omics layers within and across time points. Results For validation, we focused on a set of transcription factors that also showed importance in the network context that considers several data modalities. Using knock-down experiments, we show the relevance of these transcription factors for the ECM deposition as well as the fibrotic response to TGF-β. Conclusion Taken together, by applying mechanistic modeling and the integration of multiple datasets, we are contributing to a better understanding of the factors and mechanisms driving kidney fibrosis. Furthermore, we will publish this dataset and thus provide a valuable resource for researchers in the field of CKD.

#2769 The impact of carbon footprint in advanced chronic kidney disease treatments

Abstract Background and Aims Climate change has been classified as “the greatest global threat to health in the 21st century”. The quantification of the carbon footprint (CF) currently stands as the most accepted model for assessing the ecological impact of various businesses and healthcare systems. Renal treatments have a significant impact on the CF of healthcare systems, especially in terms of supple chains and waste management. The choice of the most environmentally friendly treatment for pathologies arising from advanced chronic kidney disease (ACKD) or peritoneal dialysis (PD) can impact the amount of waste generated and energy consumed. The objective was to analyze the carbon footprint (CF) impact resulting from the use of two different drugs in patients with the same medical condition: advanced chronic kidney disease (ACKD) or peritoneal dialysis (PD). One group received cinacalcet and another group received subcutaneous darbepoetin. Method Transversal observational study including 41 patients (19 receiving cinacalcet and 22 received darbepoetin alfa) that collected medication from the hospital pharmacy from February 1 to March 30, 2023. The CF was calculated using analytical techniques from hybrid life cycle models of the analyzed medications. For this analysis, three groups were considered: patient travel (distance traveled, trips/year, pickup, mode of transportation, and type of fuel), energy (energy consumption for refrigeration), and waste disposal and transportation (self-administration or administration by a family member, disposal location, and recycling information received). Quantification was performed using the formula: activity x emission factor (activity = sum of CF of the variables composing each group, emission factor = constant provided by the “Ministerio de transición ecológica” database: https://www.miteco.gob.es/es/cambio-climatico). Results Forty-one patients were included with a median age of 72 years [IQR 63-80]. Twenty-seven patients (65%) were male and 28 (68%) were on PD. Nineteen patients (46%) were on treatment with cinacalcet and 22 (54%) with darbepoetin alfa. There were no differences in calcium, phosphorus, parathormone or hemoglobin within the groups. Overall impact: The overall impact of the CF derived from the subcutaneous treatment process with darbepoetin alfa was 95,512.93 kg of CO2/year compared to that derived from cinacalcet treatment, which was 12,199.85 kg of CO2/year (p&amp;lt; 0.001) (Fig. 1). Patient travel to hospital and transportation of supplies Group-wise analysis did not detect significant differences in consumption derived from patient's travel to collect darbepoetin alfa or cinacalcet in the hospital (15.97 [IQR 5.25-28.66] vs 10.20 [IQR 2.08-24.5] kg CO2/year, p = 0.45). The CF of transporting supplies of darbepoetin alfa was 41,526.15 kg CO2/year compared with 10,361.54 kg CO2/year in cinacalcet group. Waste generation and transportation The waste derived CF of patients treated with darbepoetin alfa was 132 kg CO2/year [IQR 93.5-143] compared to patients treated with cinacalcet 36.4 kg CO2/year [IQR 36.3-36.5] with p-value &amp;lt; 0,001. Moreover, the transportation of waste for darbepoetin alfa resulted in a carbon footprint of 42,224.55 kg CO2/year, mainly attributable to the disposal of sharp waste. Energy: The refrigeration of darbepoetin alfa was calculated with a total emission of 8,286.96 kg CO2. Cinacalcet treatment did not require any refrigeration. Conclusion The CF derived from the subcutaneous administration of darbepoetin alfa was significantly greater than the CF derived from oral medication like cinacalcet. This may represent a negative environmental impact of the administration of medications via the subcutaneous route compared to oral medications in this group of patients. The use of drugs that do not require subcutaneous administration could be more sustainable, reducing the carbon footprint by 85%, as they do not require refrigeration or waste incineration.

Evaluation of coagulation and platelet activation state and function in heartworm-infected dogs.

Enhanced platelet responses have been demonstrated in heartworm-infected (HWI) dogs; however, the cause and clinical implications of altered platelet function have not been fully elucidated. This study evaluated platelet function in HWI dogs. Anticoagulated whole blood collected from eight HWI and eight uninfected dogs was evaluated using turbidometric platelet aggregometry, a platelet function analyzer (PFA-100), a total thrombus analysis system (T-TAS), tissue factor-activated and tissue plasminogen activator modified thromboelastography (TF- and tPA-TEG), CBC, von Willebrand Factor activity, and fibrinogen concentrations. Platelet activation state and the presence of reticulated platelets were assessed via flow cytometric expression of P-selection (CD-62P) and thiazole orange staining. Platelet aggregation responses to adenosine diphosphate (ADP, 10 μM) or collagen (20 μg/mL), PFA-100 closure times, and T-TAS occlusion times did not differ between groups. TEG values TF-R, tPA-R, TF-K, and TF-LY60 were decreased (P = .025, P = .047, P = .038, P = .025) and TF-MA, tPA-MA, TF-G, tPA-G and TF-alpha angle were increased (P < .04) in HWI dogs. HWI dogs had higher fibrinogen concentrations (465.6 ± 161 mg/dL vs 284.5 ± 38 mg/dL, P = .008) and eosinophil counts (0.686 ± 0.27 × 103/μL vs 0.267 ± 0.20 × 103/μL, P = .003). There was no difference in hematocrit, activation state, or percent of reticulated platelets. Non-activated reticulated platelets exhibited higher CD62P expression compared with mature platelets. Chronic canine heartworm disease was accompanied by hypercoagulability, hyperfibrinogenemia, and decreased fibrinolysis. Enhanced platelet activation was not identified in this group of HWI dogs.

Transcriptional Response of Wolfberry to Infestation with the Endophytic Fusarium nematophilum Strain NQ8GII4.

Fusarium nematophilum NQ8GII4 is an endophytic fungus isolated from the root of healthy wolfberry (Lycium barbarum). Previous studies have reported that NQ8GII4 could dwell in wolfberry roots and enhance the defense responses in wolfberry against root rot, which is caused by F. oxysporum. To further elucidate the molecular mechanism of wolfberry disease resistance induced by NQ8GII4, in the present study, we adopted RNA sequencing analysis to profile the transcriptome of wolfberry response to NQ8GII4 infestation over a time course of 3 and 7 days postinoculation. Gene ontology enrichment analysis revealed that differentially expressed genes (DEGs) were enriched in biological regulation, response to stimulus, signaling, detoxification, immune system process, transporter activity, electron carrier activity, transcription factor activity, nucleic acid binding transcription factor, and antioxidant activity. Through Kyoto Encyclopedia of Genes and Genomes analysis, it was found that many of these DEGs were enriched in pathways related to plant-pathogen interactions, hormone signal transduction, and the phenylpropanoid biosynthesis pathway in wolfberry. This result suggested that innate immunity, phytohormone signaling, and numerous phenylpropanoid compounds comprise a complex defense network in wolfberry. Chloroplast 50S ribosomal proteins were consistently located at the core position of the response in wolfberry following infestation with NQ8GII4 analyzed by the protein-protein interaction network. This study elucidated the molecular mechanism underlying the interaction between NQ8GII4 and wolfberry, clarified the wolfberry immune response network to endophytic fungi infestation, identified candidate resistance genes in wolfberry, and provided a fundamental date for subsequent work.

Human cytomegalovirus infection coopts chromatin organization to diminish TEAD1 transcription factor activity.

Human cytomegalovirus (HCMV) infects up to 80% of the world's population. Here, we show that HCMV infection leads to widespread changes in human chromatin accessibility and chromatin looping, with hundreds of thousands of genomic regions affected 48 hours after infection. Integrative analyses reveal HCMV-induced perturbation of Hippo signaling through drastic reduction of TEAD1 transcription factor activity. We confirm extensive concordant loss of TEAD1 binding, active H3K27ac histone marks, and chromatin looping interactions upon infection. Our data position TEAD1 at the top of a hierarchy involving multiple altered important developmental pathways. HCMV infection reduces TEAD1 activity through four distinct mechanisms: closing of TEAD1-bound chromatin, reduction of YAP1 and phosphorylated YAP1 levels, reduction of TEAD1 transcript and protein levels, and alteration of TEAD1 exon-6 usage. Altered TEAD1-based mechanisms are highly enriched at genetic risk loci associated with eye and ear development, providing mechanistic insight into HCMV's established roles in these processes.

Essential growth factor receptors for fibroblast homeostasis and activation: Fibroblast Growth Factor Receptor (FGFR), Platelet Derived Growth Factor Receptor (PDGFR), and Transforming Growth Factor β Receptor (TGFβR).

Fibroblasts are cells of mesenchymal origin that are found throughout the body. While these cells have several functions, their integral roles include maintaining tissue architecture through the production of key extracellular matrix components, and participation in wound healing after injury. Fibroblasts are also key mediators in disease progression during fibrosis, cancer, and other inflammatory diseases. Under these perturbed states, fibroblasts can activate into inflammatory fibroblasts or contractile myofibroblasts. Fibroblasts require various growth factors and mitogenic molecules for survival, proliferation, and differentiation. While the activity of mitogenic growth factors on fibroblasts in vitro was characterized as early as the 1970s, the proliferation and differentiation effects of growth factors on these cells in vivo are unclear. Recent work exploring the heterogeneity of fibroblasts raises questions as to whether all fibroblast cell states exhibit the same growth factor requirements. Here, we will examine and review existing studies on the influence of fibroblast growth factor receptors (FGFRs), platelet-derived growth factor receptors (PDGFRs), and transforming growth factor β receptor (TGFβR) on fibroblast cell states.

The reverse transcriptase inhibitor 3TC modulates hippocampal transcriptome signatures of inflammation in tauopathy model mice

Reducing neuroinflammation, a key contributor to brain aging and neurodegenerative diseases, is a promising strategy for improving cognitive function in these settings. The FDA-approved nucleoside reverse transcriptase inhibitor 3TC (Lamivudine) has been reported to improve cognitive function in old wild-type mice and multiple mouse models of neurodegenerative disease, but its effects on the brain have not been comprehensively investigated. In the current study, we used transcriptomics to broadly characterize the effects of long-term supplementation with a human-equivalent therapeutic dose of 3TC on the hippocampal transcriptome in male and female rTg4510 mice (a commonly studied model of tauopathy-associated neurodegeneration). We found that tauopathy increased hippocampal transcriptomic signatures of neuroinflammation/immune activation, but 3TC treatment reversed some of these effects. We also found that 3TC mitigated tauopathy-associated activation of key transcription factors that contribute to neuroinflammation and immune activation, and these changes were related to improved recognition memory performance. Collectively, our findings suggest that 3TC exerts protective effects against tauopathy in the hippocampus by modulating inflammation and immune activation, and they may provide helpful insight for ongoing clinical efforts to determine if 3TC and/or related therapeutics hold promise for treating neurodegeneration.

Impacts of Climate Change on the Habitat Suitability and Natural Product Accumulation of the Medicinal Plant Sophora alopecuroides L. Based on the MaxEnt Model.

Sophora alopecuroides L., a perennial herb in the arid and semi-arid regions of northwest China, has the ecological functions of windbreaking and sand fixation and high medicinal value. In recent years, global warming and human activities have led to changes in suitable habitats for S. alopecuroides, which may affect the accumulation of natural products. In this study, MaxEnt 3.4 and ArcGIS 10.4 software were used to predict the distribution of potentially suitable habitats for S. alopecuroides in China under climate change. Furthermore, the geographical distribution of S. alopecuroides as affected by human activities, the differences in the content of natural products of S. alopecuroides between different suitable habitats, and the correlation between natural products and environmental factors were analyzed. The results showed that suitable habitats for S. alopecuroides were projected to expand in the future, and the major environmental factors were temperature (Bio1), rainfall (Bio18), and soil pH (pH). When Bio1, Bio18, and pH were 8.4283 °C, 7.1968 mm, and 9.9331, respectively, the distribution probability (P) of S. alopecuroides was the highest. After adding a human activity factor, the accuracy of the model prediction results was improved, and the area of suitable habitats was greatly reduced, showing a fragmented pattern. Meanwhile, habitat suitability had a specific effect on the content of natural products in S. alopecuroides. Specifically, the content of natural products in S. alopecuroides in wild habitats was higher than that in artificial cultivation, and highly suitable habitats showed higher contents than those in non-highly suitable habitats. The contents of total alkaloids and total flavonoids were positively correlated with human activities and negatively correlated with land use types. Among them, total alkaloids were negatively correlated with aspect, and total flavonoids were positively correlated with aspect. In addition, it is suggested that Xinjiang should be the priority planting area for S. alopecuroides in China, and priority should be given to protection measures in the Alashan area. Overall, this study provides an important foundation for the determination of priority planting areas and resource protection for S. alopecuroides.

Oral submucosal fibrosis induced by active components in areca nut: a network pharmacology-based analysis and validation of the mechanism

To explore the pharmacologically active components in areca nut that induce oral submucosal fibrosis (OSF) and the possible mechanism. The chemical components in areca nut were analyzed using Thermo QE plus liquid chromatography tandem high-resolution mass spectrometer and Compound discover 3.2 data processing software. The chemical activity of the top 20 compounds was analyzed based on Chinese Pharmacopoeia (2015), PubChem, Chemical book, and SciFinder databases. The potential active components, core targets, biological functions and signaling pathways affecting OSF were analyzed by network pharmacology. The targets of OSF were obtained by integrating Genecards and KEGG databases. The compounds acting on the targets were selected from the Systematic Pharmacology Technology Platform of Traditional Chinese Medicine (TCMSP), and the target-compound, compound-TCM, target-compound-TCM network was constructed. Molecular docking was used to analyze the component-target binding. Immunohistochemistry was used to examine the expressions of key proteins in the PI3K-Akt and MAPK pathways in clinical samples of OSF. The core intersection target genes between the top 10 active ingredients in areca nut extract and OSF involved mainly the PI3K-Akt and MAPK pathways. In the clinical samples, the expressions of PI3K protein decreased and the expressions p-PI3K, AKT1 and PAkt all increased significantly in OSF tissue, where increased JNK protein expression and enhanced activity of c-Jun and c-Fos transcriptional factors were also detected. The OSF patients had significantly elevated plasma levels of IL-6 and IL-8 compared with healthy individuals. The main active ingredients including arecoline, arecaine, and guvacine are capable of activating the PI3K-Akt and MAPK pathways to promote the expressions of inflammatory mediators IL-6 and IL-8 and induce collagen hyperplasia, thus leading to the occurrence of oral submucosal fibrosis.

Differential expressions of endoplasmic reticulum stress-associated genes in aortic dissection and their correlation with immune cell infiltration

To explore differentially expressed endoplasmic reticulum stress-associated genes (ERSAGs) in aortic dissection (AD) and their correlations with immune cell infiltration to identify new therapeutic targets for AD. Two AD mRNA expression datasets (GSE190635 and GSE98770) were downloaded from GEO database for analysis of differentially expressed genes between the aorta of AD patients and normal aorta using R software. ERSAGs dataset was downloaded from GeneCards website, and GeneMANIA database was used to analyze the protein-protein interaction network of the differentially expressed ERSAGs and the proteins interacting with these genes. Based on GSE98770 dataset we analyzed the distributions of 22 immune cells within the aortic wall of AD patients using CIBERSORT package of R software. Surgical aortic wall specimens were obtained from 10 AD patients and 10 non-AD patients for detecting AGER mRNA expression using qRTPCR, and the upstream transcriptional factors, miRNAs, and chemicals targeting AGER were analyzed using the TRRUST database and NetworkAnalyst database. Bioinformatic analysis suggested significant differential expression of AGER in AD, which interacted with 20 proteins involved in pattern recognition receptor signaling pathway, positive regulation of DNA-binding transcription factor activity, myeloid leukocyte migration, leukocyte migration, and regulation of the I-κB kinase/NF-κB signaling. In AD, AGER expression level was positively correlated with Treg cell abundance (r=0.59, P < 0.05). The results of qRT-PCR demonstrated significantly lower expression of AGER mRNA in AD than in non-AD patients (1.00±0.30 vs 1.76±0.68, P < 0.05). ROC curve analysis showed that at the cut-off value of 1.335, AGER had an AUC of 0.86 (95% CI: 0.67-1.00, P= 0.0073) for predicting AD. Three transcriptional factors, 3 miRNAs, and 27 chemicals were predicted in the AGER regulatory network. AGER is lowly expressed in the aorta of AD patients and may influence the occurrence of AD through Treg cells.