Molecular Mechanism Of Action Research Articles

Chemopreventive potential of goniothalamin in diethylnitrosamine-induced hepatocellular carcinoma through the suppression of P13K/AKT signalling pathway.

Liver cancer is the most lethal form of cancer and carries a high risk of death around the world. Goniothalamin (GTN) is a styryl-lactone that possesses antiproliferative and apoptotic activity. The molecular action of GTN is not yet fully evaluated. Thus, our research has been intended to assess the chemopreventive and apoptotic activities of diethylnitrosamine (DEN)-induced hepatocellular carcinoma (HCC) in rats. Rats were separated into 4 groups: control, DEN only, DEN + GTN (30 mg/kg bw), and GTN (30 mg/kg bw) alone. We evaluated body weight, liver weight, tumor incidence, hepatic toxic markers, antioxidants, inflammatory cytokines, histopathology, immunohistochemistry, and Western blot studies. DEN lessened body weight, antioxidants, and apoptosis, whereas it elevated tumor incidence, toxic markers, cytokines, and Bcl-2 expression. GTN treatment maintains body weight, liver weight, and antioxidant levels, and it also prevents tumor incidence, oxidative stress, toxic markers, pro-inflammatory cytokines, and histological changes. It triggers apoptosis by constraining Bcl-2 and elevating caspase-3 levels. GTN also attenuated the P13K/ AKT signaling which enhanced apoptosis. These findings revealed that GTN subdues the P13K/AKT pathway and has auspicious chemopreventive and apoptotic actions in DEN-induced HCC. Therefore, GTN would be suggested as a new medicine in natural remedies for liver cancer.

An epigenetic editor targeting human PCSK9 efficiently and durably lowers Low Density Lipoprotein Cholesterol (LDL-C) in non-human primates

Abstract Background Reducing the risk of atherosclerotic cardiovascular disease is dependent on both the magnitude and cumulative duration of LDL-C lowering. However, in clinical practice achieving treatment goals is often hampered by low adherence to standard of care. Although PCSK9 inhibitors represent an effective option for LDL-C lowering, they require chronic life-long treatment. Epigenetic editing is a new therapeutic approach designed to durably silence genes by leveraging nature’s endogenous cellular mechanism for gene regulation via CpG methylation without the inherent genotoxic risks of genome editing approaches that nick or cut the DNA. We are developing a PCSK9 epigenetic editor (PCSK9-EE) to durably silence PCSK9, with the promise of lifelong reduction in LDL-C. Methods Here, we describe the development of an epigenetic editor targeting human PCSK9. This epigenetic editor consists of a DNA targeting component fused to a transcriptional repressor domain and a DNA methyltransferase domain. PCSK9-EE was first screened and evaluated for specificity and efficacy in primary human hepatocytes (PHH). In vivo efficacy and durability were then evaluated in a transgenic mouse carrying the human PCSK9 genomic locus (hPCSK9-Tg mouse) and non-human primates (NHP). Results Our initial screen of PCSK9-EEs identified several hits that efficiently suppressed secreted PCSK9 levels in immortalized liver cells. We confirmed that our top PCSK9-EE candidates robustly decreased PCSK9 secretion in PHH and were found to be highly specific at targeting PCSK9 as assessed by RNA-seq, methylation array, and whole-genome methylation sequencing. To examine in vivo activity, we treated hPCSK9-Tg mice with a single administration of a lipid nanoparticle delivering mRNA encoding our PCSK9-EE and observed >97% reduction in liver PCSK9 mRNA, and >98% reduction in plasma PCSK9 for the duration of the study (one year). We then delivered PCSK9-EE in NHP and observed a robust reduction in plasma PCSK9 with a concomitant reduction in LDL-C. These effects were found to be durable for at least 6 months following a single administration of PCSK9-EE in NHP. To establish a mechanistic relationship between PCSK9-EE’s molecular action and durable PCSK9 silencing, we performed serial liver biopsies in each NHP approximately 2 months apart. Robust CpG methylation at the PCSK9 genomic locus was observed in the first liver biopsy and was comparable to that observed in the second liver biopsy suggesting that PCSK9-EE induced stable CpG methylation in NHP livers. Conclusions We believe PCSK9-EE may offer an effective, safe, and durable approach to suppress hepatic PCSK9 expression and achieve long-lasting reduction in LDL-C. Furthermore, epigenetic editing may hold promise for one-and-done approaches for the treatment of atherosclerotic cardiovascular disease without cutting, nicking, or altering the DNA sequence.

Modulation of Albumin Esterase Activity by Warfarin and Diazepam

Data are accumulating on the hydrolytic activity of serum albumin towards esters and organophosphates. Previously, with the help of the technology of proton nuclear magnetic resonance (1H NMR) spectroscopy, we observed the yield of acetate in the solution of bovine serum albumin and p-nitrophenyl acetate (NPA). Thus, we showed that albumin possesses true esterase activity towards NPA. Then, using the methods of molecular docking and molecular dynamics, we established site Sudlow I as the catalytic center of true esterase activity of albumin. In the present work, to expand our understanding of the molecular mechanisms of albumin pseudoesterase and true esterase activity, we investigated—in experiments in vitro and in silico—the interaction of anticoagulant warfarin (WRF, specific ligand of site Sudlow I) and benzodiazepine diazepam (DIA, specific ligand of site Sudlow II) with albumins of different species, and determined how the binding of WRF and DIA affects the hydrolysis of NPA by albumin. It was found that the characteristics of the binding modes of WRF in site Sudlow I and DIA in site Sudlow II of human (HSA), bovine (BSA), and rat (RSA) albumins have species differences, which are more pronounced for site Sudlow I compared to site Sudlow II, and less pronounced between HSA and RSA compared to BSA. WRF competitively inhibits true esterase activity of site Sudlow I towards NPA and does not affect the functioning of site Sudlow II. Diazepam can slow down true esterase activity of site Sudlow I in noncompetitive manner. It was concluded that site Sudlow I is more receptive to allosteric modulation compared to site Sudlow II.

Applications, impacts and consequences of botulinum toxin usage in medicine

Introduction Botulinum toxin is a neurotoxin produced by the gram-positive bacteria Clostridium botulinum. Botulinum toxin has found great interest in the field of aesthetic medicine, but it has many other applications in other areas of medicine. Aim of the study The aim of this comprehensive review is to provide an overview of the usage of botulinum toxin in medicine, with a focus on specific areas of medicine. Attention will be paid to the safety of the use of this substance, taking into account its molecular structure and mechanism of action. Materials and methods An analysis of the scientific articles available in the Pubmed, Google Scholar and web of science databases was performed. Recent publications which most appropriately encompassed the issues under discussion have been selected for the purpose of this study. The process of search was carried out with the following keywords: neurotoxin, botulinum toxins, botulism, muscle paralysis. Summary Despite numerous studies, there is ample room for further research into the safety of using botulinum toxin in patients as well as new therapies with this substance. Botulinum toxin is a neurotoxin that is widely used in medicine, but has many side effects.

A Comprehensive Review of Pedunculagin: Sources, Chemistry, Biological and Pharmacological Insights

Pedunculagin is a widely abundant ellagitannin found in the plant kingdom, with a chemical structure featuring two hexahydroxydiphenoyl units linked to a glucose core. It has demonstrated various biological activities, including anti-cancer, anti-inflammatory, and anti-bacterial effects. This review aims to summarize the bioactivities, chemistry, and health-promoting properties of pedunculagin and plant preparations containing it. It is the first comprehensive summary covering pedunculagin’s chemistry, sources, metabolism, and other relevant research. The search databases were Google Scholar, EBSCO Discovery Service, REAXYS Database, SCILIT, SCOPUS, PubMed, MEDLINE, Web of Science, Wiley Online Library, Science Direct/ELSEVIER, WordCat, and Taylor and Francis Online. All the databases were methodically searched for data published from 1911 until 2024. Various biological effects were proven in vitro for pedunculagin; however, due to the limited availability of the isolated compound, they have not been so far directly confirmed on more advanced in vivo and clinical models. However, its bioactivity can be deduced from studies conducted for plant preparations containing this ellagitannin as a dominant constituent, consequently indicating beneficial health effects. Further studies are needed to determine the molecular mechanism of action following topical application as well as the contribution of gut microbiota postbiotic metabolites– urolithins–being formed following the oral ingestion of preparations containing pedunculagin.

Allicin Ameliorated High-glucose Peritoneal Dialysis Solution-induced Peritoneal Fibrosis in Rats via the JAK2/STAT3 Signaling Pathway.

Peritoneal fibrosis (PF) is one of the most serious complications of peritoneal dialysis (PD) and is the greatest obstacle to the clinical application of PD. Chinese herbal monomers have been effective in the prevention and treatment of PF. The aim of this study was to observe the effect of allicin on PF in rats induced by high glucose and to investigate its molecular mechanism of action. A rat model of PF was established by using a 4.25% glucose-based standard peritoneal dialysis solution. The degree of peritoneal pathological damage was assessed by Hematoxylin and eosin (H&E) staining. Peritoneal collagen deposition was detected by Masson's trichrome staining. The levels of Interleukin-6 (IL-6), Tumor necrosis factor-α (TNF-α), Interleukin-1β (IL-1β) and monocyte chemoattractant protein-1 (MCP-1) in the serum were measured by Enzyme Linked Immunosorbent Assay (ELISA). The expression levels of TGF-β, α-smooth muscle actin (α-SMA) and collagen I were examined by western blotting and immunohistochemistry. The protein expression levels and mRNA levels of E-cadherin, N-cadherin, vimentin, janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) in peritoneal tissue were determined by western blotting and qRT-PCR. TGF-β1 stimulated human peritoneal mesothelial cells (HPMCs), and the cells were treated with allicin and the JAK2/STAT3 pathway activator colivelin alone or in combination. A cell counting kit-8 (CCK-8) assay was used to measure cell viability. The role of JAK2/STAT3 in the effects of allicin was confirmed via in vitro mechanistic research by western blotting, wound healing assays and Transwell assays. Allicin relieves the inflammatory response by downregulating the levels of IL-1β, IL-6, MCP-1 and TNF-α. Furthermore, allicin decreased the expression of TGF-β, α-SMA and collagen I. Allicin also alleviated epithelial-to-mesenchymal transition (EMT), as specifically manifested by increased E-cadherin and reduced N-cadherin and vimentin. Further studies revealed that allicin reduced the protein levels of JAK2, STAT3, p-JAK2, and p-STAT3. The results of the cellular experiments verified the above results. The ability of allicin to inhibit fibrosis and the EMT process was significantly attenuated after HPMCs were treated with colivelin. Taken together, these findings suggest that allicin inhibits inflammation and EMT, thereby improving PF, and this protective effect may be achieved by inhibiting the JAK2/STAT3 signaling pathway.

Insight into structural properties of viral G protein-coupled receptors and their role in the viral infection: IUPHAR Review 41.

G protein-coupled receptors (GPCRs) are pivotal in cellular signalling and drug targeting. Herpesviruses encode GPCRs (vGPCRs) to manipulate cellular signalling, thereby regulating various aspects of the virus life cycle, such as viral spreading and immune evasion.vGPCRs mimic host chemokine receptors, often with broader signalling and high constitutive activity. This review focuses on the recent advancements in structural knowledge about vGPCRs, with an emphasis on molecular mechanisms of action and ligand binding. The structures of US27 and US28 from human cytomegalovirus (HCMV) are compared to their closest human homologue, CX3CR1. Contrasting US27 and US28, the homotrimeric UL78 structure (HCMV) reveals more distance to chemokine receptors. Open reading frame 74 (ORF74; Kaposi's sarcoma-associated herpesvirus) is compared to CXCRs, whereas BILF1 (Epstein-Barr virus) is discussed as a putative lipid receptor. Furthermore, the rolesof vGPCRs in latency and lytic replication, reactivation, dissemination and immune evasion arereviewed, together with their potential as drug targets for virus infections and virus-related diseases.

Exploring the molecular pathways of the activation process in PPARγ recurrent bladder cancer mutants.

The intricate involvement of Peroxisome Proliferator-Activated Receptor Gamma (PPARγ) in glucose homeostasis and adipogenesis is well-established. However, its role in cancer, particularly luminal bladder cancer, remains debated. The overexpression and activation of PPARγ are implicated in tumorigenesis. Specific gain-of-function mutations (M280I, I290M, and T475M) within the ligand-binding domain of PPARγ are associated with bladder cancer and receptor activation. The underlying molecular pathways prompted by these mutations remain unclear. We employed a dual-basin structure-based model (db-SBM) to explore the conformational dynamics between the inactive and active states of PPARγ and examined the effects of the M280I, I290M, and T475M mutations. Our findings, consistent with the existing literature, reveal heightened ligand-independent transcriptional activity in the I290M and T475M mutants. Both mutants showed enhanced stabilization of the active state compared to the wild-type receptor, with the I290M mutation promoting a specific transition route, making it a prime candidate for further study. Electrostatic analysis identified residues K303 and E488 as pivotal in the I290M activation cascade. Biophysical assays confirmed that disrupting the K303-E488 interaction reduced the thermal stabilization characteristic of the I290M mutation. Our study demonstrates the predictive capabilities of combining simulation and cheminformatics methods, validated by biochemical experiments, to gain insights into molecular activation mechanisms and identify target residues for protein modulation.

Molecular mechanism of IgE-mediated FcεRI activation.

Allergic diseases, affecting over a quarter of individuals in industrialized countries, have become significant public health concerns1,2. The high-affinity Fc receptor for IgE (FcεRI), mainly present on mast cells and basophils, plays a crucial role in allergic diseases3-5. Monomeric IgE binding to FcεRI regulates mast cell survival, differentiation, and maturation6-8. However, the underlying molecular mechanism remains unclear. Here we demonstrate that, prior to IgE binding, FcεRI mostly exists as a homo-dimer on human mast cell membrane. The structure of human FcεRI confirms the dimeric organization, with each promoter comprising one α subunit, one β subunit, and two γ subunits. The transmembrane helices of the α subunits form a layered arrangement with those of the γ and β subunits. The dimeric interface is mediated by a four-helix bundle of the α and γ subunits at the intracellular juxtamembrane region. Cholesterol-like molecules embedded within the transmembrane domain may stabilize the dimeric assembly. Upon IgE binding, the dimeric FcεRI dissociates into two protomers, each binding to an IgE molecule. Importantly, this process elicits transcriptional activation of Egr1/3 and Ccl2 in rat basophils, which can be attenuated by inhibiting the FcεRI dimer-to-monomer transition. Collectively, our study unveils the mechanism of antigen-independent, IgE-mediated FcεRI activation.

Dimethyl trisulfide reduces postharvest anthracnose and enhances mango quality, and a potential molecular mechanism against Colletotrichum gloeosporioides

Mango anthracnose, mainly caused by Colletotrichum gloeosporioides, is the major destructive postharvest disease of mango during storage and transport. Dimethyl trisulfide (DMTS), an organic volatile found in some microorganisms or plants, inhibited growth of C. gloeosporioides in vitro, but its effects on mango anthracnose and its molecular mechanisms of action have not been well characterized. In this study, the EC50 of DMTS against Colletotrichum spp. from mango mainly ranged from 2.3 to 20.0 μL/L. In vivo, the fumigation rates of 20 μL/L of DMTS for 24 h, or 80 μL/L for 3 h or 6 h could effectively reduce severity of anthracnose (natural inoculum) on postharvest mangoes with inhibitory effects of 61.7 %, 65.7 %, and 69.4 %, respectively, as observed 10 days after treatment. Furthermore, there was no detectable DMTS residue in mango skin or flesh, and an overall improvement in the quality of the fruit with higher soluble solids, total sugars, vitamin c, and β-carotene, and lower titratable acidity than the non-treated control. In addition, DMTS could significantly reduce ergosterol content in mycelia of C. gloeosporioides, and gene expression analysis showed DMTS significantly suppressed expression of ergosterol biosynthesis-related genes Cgerg6 and Cgerg11 after mycelia were exposed to DMTS. Knock-out mutants for each of these two genes showed reduced sensitivity to DMTS. After gene complementation in situ, the sensitivity of complementary transformants to DMTS was restored to that of the parental strain. Therefore, we concluded that the genes Cgerg6 and Cgerg11 are involved in an interaction with the antifungal activity of DMTS. This is the first study to demonstrate a control effect of DMTS on mango postharvest anthracnose resulting in reduced disease severity and enhanced fruit quality. Transformant studies also revealed some potential molecular mechanisms of the antifungal activity of DMTS that may lead to improved management of mango postharvest anthracnose.

Activation of plant immunity through conversion of a helper NLR homodimer into a resistosome.

Nucleotide-binding domain and leucine-rich repeat (NLR) proteins can engage in complex interactions to detect pathogens and execute a robust immune response via downstream helper NLRs. However, the biochemical mechanisms of helper NLR activation by upstream sensor NLRs remain poorly understood. Here, we show that the coiled-coil helper NLR NRC2 from Nicotiana benthamiana accumulates in vivo as a homodimer that converts into a higher-order oligomer upon activation by its upstream virus disease resistance protein Rx. The cryo-EM structure of NbNRC2 in its resting state revealed intermolecular interactions that mediate homodimer formation and contribute to immune receptor autoinhibition. These dimerization interfaces have diverged between paralogous NRC proteins to insulate critical network nodes and enable redundant immune pathways, possibly to minimise undesired cross-activation and evade pathogen suppression of immunity. Our results expand the molecular mechanisms of NLR activation pointing to transition from homodimers to higher-order oligomeric resistosomes.

Molecular mechanism of proteolytic cleavage-dependent activation of CadC-mediated response to acid in E. coli

Colonizing in the gastrointestinal tract, Escherichia coli confronts diverse acidic challenges and evolves intricate acid resistance strategies for its survival. The lysine-mediated decarboxylation (Cad) system, featuring lysine decarboxylase CadA, lysine/cadaverine antiporter CadB, and transcriptional activator CadC, plays a crucial role in E. coli’s adaptation to moderate acidic stress. While the activation of the one-component system CadC and subsequent upregulation of cadBA operon in response to acid and lysine presence have been proposed, the molecular mechanisms governing the transition of CadC from an inactive to an active state remain elusive. Under neutral conditions, CadC is inhibited by forming a complex with lysine-specific permease LysP, stabilized in this inactive state by a disulfide bond. Our study unveils that, in an acidic environment, the disulfide bond in CadC is reduced by the disulfide bond isomerase DsbC, exposing R184 to periplasmic proteases, namely DegQ and DegP. Cleavage at R184 by DegQ and DegP generates an active N-terminal DNA-binding domain of CadC, which binds to the cadBA promoter, resulting in the upregulated transcription of the cadA and cadB genes. Upon activation, CadA decarboxylates lysine, producing cadaverine, subsequently transported extracellularly by CadB. We propose that accumulating cadaverine gradually binds to the CadC pH-sensing domain, preventing cleavage and activation of CadC as a feedback mechanism. The identification of DegP, DegQ, and DsbC completes a comprehensive roadmap for the activation and regulation of the Cad system in response to moderate acidic stress in E. coli.

Molecular mechanisms of novel selective glucocorticoid receptor agonist action in breast cancer cells

Introduction. Glucocorticoids (GC) are widely used in breast cancer (BC) therapy to reduce the side effects of cytostatic drugs and may exhibit antiproliferative effects on luminal BC cells. The biological action of GC is mediated by glucocorticoid receptor (GR) by two mechanisms: transrepression, which determines the therapeutic effect of GC, and transactivation (associated with the development of side effects, resistance to cytotoxic drugs, cancer progression and metastasis). Selective GR agonists (SEGRA) which may selective activate transrepression are a promising alternative to GC to use in combination cancer therapy. One of the most studied SEGRA is Compound A (CpdA). The instability of CpdA limits its use in clinical practice. So recently we performed synthesis and evaluation of biological activities of the CpdA analogue, CpdA-03.Aim. To compare the effects of SEGRA CpdA-03 and CpdA and dexamethasone on proliferative activity of breast cancer cells, as well as receptor nuclear translocation and activation of GR-dependent genes in breast cancer cells.Materials and methods. Luminal (MCF-7) and triple negative (MDA-MB-231) BC cell lines were used. The effect of CpdA-03 on proliferation was evaluated by direct counting of viable cells with trypan blue staining. The effect of the compound on cell distribution by cell cycle phases was assessed by flow cytofluorimetry with propidium iodide staining. Changes in the expression of GR-dependent genes after incubation with CpdA-03 were evaluated by quantitative polymerase chain reaction. Additionally, the ability of the new SEGRA to induce transactivation-associated translocation of the receptor to the nucleus was evaluated by Western blotting.Results. CpdA-03 was shown to suppress proliferation of luminal and triple negative BC cells. This compound causes changes in the expression of a number of GC-inducible genes, but does not stimulate GR phosphorylation and translocation to the nucleus in BC cells.Conclusion. The observed suppression of cell proliferation, as well as the ability of CpdA-03 to reduce gene expression of proteins regulating intercellular adhesion and cell migration, intracellular signaling, stress response, and transcription in BC cells makes it relevant for further development of the drug for use in the combination therapy of cancers including breast cancer.

Escherichia coli and Micrococcus luteus Activate the CG45045 Gene in Drosophila S2 Cell Line.

The humoral immune system of Drosophila melanogaster, which is the best studied of all eukaryotes, is activated by the canonical IMD and Toll signalling pathways. Recently, long non-coding RNAs (lncRNAs) and genes encoding short polypeptides have been identified as potential regulators of the innate immune response. S2 cells are a macrophage-like cell line. They are used as a model system to study the molecular mechanisms of immune response gene activation. We used this cell line to study the effect of Escherichia coli and Micrococcus luteus bacteria on the transcription of the lncRNA-CR30055 and the CG45045 and CG44404 genes, encoding short polypeptides. We found that pathogens activate only CG45045, while the transcription levels of CR30055 and CG44404 remain unchanged. No activation of Cecropin C and some Bomanin family genes was observed, suggesting differing patterns of immune response gene activation in S2 cells and adult flies. The highest activation of CG45045 was observed between 6 and 12 hours of cell incubation with pathogens. The activation patterns of CG45045 after exposure to E. coli and M. luteus were similar, suggesting common mechanisms of transcriptional activation of this gene. Thus, CG45045 may be a novel gene involved in the humoral immune response of Drosophila.

Exploring the Potential Mechanisms of Danshen for the Treatment of Ulcerative Colitis based on Serum Pharmacochemistry, Gene Expression Profiling, and Network Pharmacology: Regulation of Cell Apoptosis and Inflammatory Response.

As a traditional Chinese medicine, Danshen shows potential efficacy for treating ulcerative colitis (UC). However, the bioactive components and mode of action were unclear. This paper uses a combination of network pharmacology, serum medicinal chemistry, and gene expression profiling to clarify its possible molecular mechanism of action and material basis. Ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) was utilized to analyze the herbal components and metabolites from the serum of Danshen-treated mice. Gene expression profiles were applied to construct a database of Danshen action targets. Then, active ingredient-target-biological functional module networks were constructed to analyze the mechanism of action. Molecular docking has further confirmed the possibility of its components to the targets. As a result, 193 common targets between 1684 Danshen-related DEGs and 1492 UC targets were determined as the potential targets for Danshen in treatment with UC. Serum pharmacochemistry and target prediction showed that 22 components in serum acted on 777 targets. Intersection with common targets yielded 46 core targets, and an active ingredienttarget- biological functional module network was constructed for analysis. Network prediction and molecular docking results showed that the main action modules were inflammatory response and cell apoptosis, which mainly acted on targets SRC, RELA, HSP90AA1, CTNNB1, STAT3, and CASP3. The main components of Danshen intervention in UC were predicted to include Catechol, 3,9-Dimethoxypterocarpan, 8-Prenylnaringenin, Isoferulic acid, Salvianolic acid C, and Danshensu. The present study provides a scientific foundation for further explicating the mechanisms of Danshen against UC.

Convergent effects of different anesthetics on changes in phase alignment of cortical oscillations.

Many anesthetics cause loss of responsiveness despite having diverse underlying molecular and circuit actions. To explore the convergent effects of these drugs, we examined how anesthetic doses of ketamine and dexmedetomidine affected oscillations in the prefrontal cortex of nonhuman primates. Both anesthetics caused increases in phase locking in the ventrolateral and dorsolateral prefrontal cortex, within and across hemispheres. However, the nature of the phase locking varied. Activity in different subregions within a hemisphere became more anti-phase with both drugs. Local analyses within a region suggested that this finding could be explained by broad cortical distance-based effects, such as large traveling waves. By contrast, homologous areas across hemispheres became more in-phase. Our results suggest that both anesthetics induce strong patterns of cortical phase alignment that are markedly different from those in the awake state, and that these patterns may be a common feature driving loss of responsiveness from different anesthetic drugs.

Dihydrotanshinone I promotes LDL uptake by HepG2 cells through increasing LDLR level

ABSTRACT Dihydrotanshinone I (DHT), a lipophilic compound extracted from Salvia miltiorrhiza, has cardiovascular protective effects, but the underlying molecular mechanisms of action have rarely been reported. Based on this, whether DHT affects cholesterol metabolism by regulating LDLR is investigated in this work. The results revealed that DHT can increase LDLR expression and promote LDL uptake by HepG2 cells. Meanwhile, DHT stabilizes LDLR mRNA expression by activating the epidermal growth factor receptor/extracellular signal-regulated kinase 1/2 (EGFR/erk1/2) signaling pathway. In addition, DHT inhibits the expression of the proprotein convertase subtilisin/kexin type 9 (PCSK9) by down-regulating the liver nuclear transcription factor 1 A (HNF1A) and up-regulating the forkhead box O3 (FOXO3). All of these indicate that DHT increases LDLR expression at the post-transcriptional and post-translational levels thereby promoting LDL-C metabolism. The potential mechanism of DHT to improve lipid metabolism has been revealed as a promising drug against AS.

Study of mRNA expression of thirteen genes of Trypanosoma evansi in response to diminazene aceturate and isometamidium chloride

The monomorphic, non-cyclic, extracellular haemoprotozoan parasite, Trypanosoma evansi leads to Surra disease in domesticated animals. Currently, diminazene aceturate (DA) and isometamidium chloride (ISM) are the most used chemotherapeutic agents for the treatment of Surra in animals. There is still little knowledge on the anti- trypanosomal mechanism of action of DA and ISM. The work addresses a significant gap in the understanding of the anti-typanosomal mechanism of DA and ISM by investigating their effects on mRNA expression profiles of 13 genes of T. evansi. The half maximal inhibitory concentration (IC50) of DA and ISM for a pony isolate of T. evansi was estimated as 335.3 nM and 308.6 nM, respectively. Transcript analysis of DA and ISM exposed T. evansi population showed its effects on the metabolic machinery of T. evansi by down-regulating the mRNA expression of all the 13 targeted genes. However, ISM exposure did not affect mRNA expression of Expression site-associated genes 8 (ESAG8), oligopeptidase B and ornithine decarboxylase genes. The finding provides valuable insights into the molecular action of these drugs, which is crucial for developing more effective treatment of Surra disease. Further, comprehensive transcriptome and proteomic analysis could provide a deeper insight into precise molecular pathway of these medications against T. evansi.

Modulating JAK2/STAT3 signaling by quercetin in Qiling Baitouweng Tang: a potential therapeutic approach for diffuse large B-cell lymphoma.

Qiling Baitouweng Tang (QLBTWT) is a traditional clinical formula for treating diffuse large B-cell lymphoma (DLBCL), but its molecular action is not fully understood. This research is utilized in silico analysis and liquid chromatography tandem mass spectrometry (LC‒MS/MS) to identify the active constituents of QLBTWT with anti-DLBCL properties and their targets. The study identified 14 compounds, including quercetin, naringenin, and astilbin, as potentially effective against DLBCL. Molecular modeling highlighted the favorable interaction of quercetin with the JAK2 protein. In vitro studies confirmed the ability of quercetin to inhibit DLBCL cell growth and migration while inducing apoptosis and causing G2/M phase cell cycle arrest. Molecular dynamics simulations revealed that quercetin binds to JAK2 as a type II inhibitor. In vivo studies in U2932 xenograft models demonstrated that QLBTWT inhibited tumor growth in a dose-dependent manner, which was associated with the JAK2/STAT3 signaling pathway. Overall, this study elucidates the therapeutic effect of QLBTWT on DLBCL through quercetin-mediated suppression of the JAK2/STAT3 pathway, offering novel therapeutic insights for DLBCL.

9194 Generation of a Human-Induced Pluripotent Stem Cell Line From a Patient With Hypopituitarism and a Novel Nonsense Variant in FOXA2

Abstract Disclosure: M.A. Camilletti: None. G.T. Chirino Felker: None. G. Amin: None. S. Castañeda: None. F. Zabalegui: None. C. Romano Florit: None. A. Waisman: None. M. Ciaccio: None. M.I. Di Palma: None. E. Vaiani: None. A. Belgorosky: None. S.G. Miriuka: None. L.N. Moro: None. M.I. Perez-Millan: None. Congenital Hypopituitarism (CH) is a genetically complex disease characterized by one or more pituitary hormone deficiencies. Forkhead Box A2 (FOXA2, 600288) is a pioneer transcription factor associated with development of multiple tissues in humans and mice. Heterozygous loss of function mutations in FOXA2 were recently shown to cause CH with hyperinsulinemic hypoglycemia, but little is known about the molecular mechanism of FOXA2 action during pituitary development. We identified a novel, heterozygous nonsense variant in FOXA2 (c.686C<A; p.S229X) in a patient diagnosed with GH deficiency at 1.7 years of age, with anterior lobe hypoplasia and absent pituitary stalk, asymptomatic hypoglycemia, and craniofacial malformations. To discover the mechanism of FOXA2 regulation of pituitary development we generated an induced pluripotent stem cell line (FOXA2mut-iPSC) from the patient’s polymorphonuclear blood cells (PBMCs) using an EF1a-hSTEMCCA-loxP vector containing the pluripotency genes OCT-4, SOX-2, c-MYC and KLF4. FOXA2mut-iPSC clone R1 had a normal karyotype and embryonic stem cell-like morphology. These cells express endogenous pluripotency-associated markers NANOG, SOX2, and OCT4, have alkaline phosphatase activity, and can differentiate into all three developmental layers, indicating that these cells are pluripotent. Short tandem repeat (STR) analysis confirmed that FOXA2mut-iPSC R1 clone profiles matched those of the donor PBMCs. Next, we compared pituitary differentiation of FOXA2mut-iPSC vs. control-iPSCs in vitro. Cells were cultured in a differentiation medium containing BMP4, the smoothened agonist SAG, and fibroblast growth factor (FGF), as described in Zimmer et al., 2016 (1). Gene expression analysis (by qRT-PCR) of samples collected on days 0, 4, 7, and 15 of the differentiation protocol showed an increased expression of genes associated with anterior pituitary development including OTX2, PITX1/2, and SIX1 in both cell lines (n=2). These preliminary data suggest that pituitary cell fate can be induced in the heterozygous FOXA2mut-iPSC clone. Ongoing studies will assess pituitary hormone-producing cell differentiation after longer periods of FOXA2mut-iPSC cell culture. In sum, patient iPSC differentiation is a promising tool for assessment of normal and variant FOXA2 function in human pituitary development, and for enlightening the mechanisms of FOXA2 action in human pituitary development. (1) Zimmer et al., Stem Cell Reports. 2016 Jun 14;6(6):858-872. Presentation: 6/1/2024