-
International Journal of Molecular... May 2024Pulmonary arterial hypertension (PAH) is a progressive cardiopulmonary disease characterized by pathologic vascular remodeling of small pulmonary arteries. Endothelial...
Pulmonary arterial hypertension (PAH) is a progressive cardiopulmonary disease characterized by pathologic vascular remodeling of small pulmonary arteries. Endothelial dysfunction in advanced PAH is associated with proliferation, apoptosis resistance, and endothelial to mesenchymal transition (EndoMT) due to aberrant signaling. DLL4, a cell membrane associated NOTCH ligand, plays a pivotal role maintaining vascular integrity. Inhibition of DLL4 has been associated with the development of pulmonary hypertension, but the mechanism is incompletely understood. Here we report that silencing in pulmonary artery endothelial cells (PAECs) activated AKT and suppressed the expression of DLL4. Consistent with these in vitro findings, increased AKT activation and reduced DLL4 expression was found in the small pulmonary arteries of patients with PAH. Increased NOTCH1 activation through exogenous DLL4 blocked AKT activation, decreased proliferation and reversed EndoMT. Exogenous and overexpression of DLL4 induced and PPRE promoter activity, and and mRNA in idiopathic PAH (IPAH) ECs. PPARγ, a nuclear receptor associated with EC homeostasis, suppressed by BMPR2 loss was induced and activated by DLL4/NOTCH1 signaling in both -silenced and IPAH ECs, reversing aberrant phenotypic changes, in part through AKT inhibition. Directly blocking AKT or restoring DLL4/NOTCH1/PPARγ signaling may be beneficial in preventing or reversing the pathologic vascular remodeling of PAH.
Topics: Humans; Proto-Oncogene Proteins c-akt; Signal Transduction; Bone Morphogenetic Protein Receptors, Type II; PPAR gamma; Receptor, Notch1; Pulmonary Artery; Endothelial Cells; Adaptor Proteins, Signal Transducing; Calcium-Binding Proteins; Pulmonary Arterial Hypertension; Male; Cell Proliferation; Hypertension, Pulmonary; Female; Cells, Cultured
PubMed: 38791441
DOI: 10.3390/ijms25105403 -
International Journal of Molecular... May 2024The pathogenicity of many bacteria, including and , depends on pore-forming toxins (PFTs), which cause the lysis of host cells by forming pores in the membranes of...
The pathogenicity of many bacteria, including and , depends on pore-forming toxins (PFTs), which cause the lysis of host cells by forming pores in the membranes of eukaryotic cells. Bioinformatic analysis revealed a region homologous to the Lys171-Gly250 sequence in hemolysin II (HlyII) from in over 600 PFTs, which we designated as a "homologous peptide". Three β-barrel PFTs were used for a detailed comparative analysis. Two of them-HlyII and cytotoxin K2 (CytK2)-are synthesized in ; the third, α-toxin (Hla), is the most investigated representative of the family. Protein modeling showed certain amino acids of the homologous peptide to be located on the surface of the monomeric forms of these β-barrel PFTs. We obtained monoclonal antibodies against both a cloned homologous peptide and a 14-membered synthetic peptide, DSFNTFYGNQLFMK, as part of the homologous peptide. The HlyII, CytK2, and Hla regions recognized by the obtained antibodies, as well as an antibody capable of suppressing the hemolytic activity of CytK2, were identified in the course of this work. Antibodies capable of recognizing PFTs of various origins can be useful tools for both identification and suppression of the cytolytic activity of PFTs.
Topics: Bacterial Toxins; Bacillus cereus; Hemolysin Proteins; Staphylococcus aureus; Amino Acid Sequence; Hemolysis; Pore Forming Cytotoxic Proteins; Models, Molecular; Animals; Antibodies, Monoclonal; Humans; Bacterial Proteins
PubMed: 38791367
DOI: 10.3390/ijms25105327 -
International Journal of Molecular... May 2024Drug induced fatty liver disease (DIFLD) is a form of drug-induced liver injury (DILI), which can also be included in the more general metabolic dysfunction-associated... (Review)
Review
Drug-Induced Fatty Liver Disease (DIFLD): A Comprehensive Analysis of Clinical, Biochemical, and Histopathological Data for Mechanisms Identification and Consistency with Current Adverse Outcome Pathways.
Drug induced fatty liver disease (DIFLD) is a form of drug-induced liver injury (DILI), which can also be included in the more general metabolic dysfunction-associated steatotic liver disease (MASLD), which specifically refers to the accumulation of fat in the liver unrelated to alcohol intake. A bi-directional relationship between DILI and MASLD is likely to exist: while certain drugs can cause MASLD by acting as pro-steatogenic factors, MASLD may make hepatocytes more vulnerable to drugs. Having a pre-existing MASLD significantly heightens the likelihood of experiencing DILI from certain medications. Thus, the prevalence of steatosis within DILI may be biased by pre-existing MASLD, and it can be concluded that the genuine true incidence of DIFLD in the general population remains unknown. In certain individuals, drug-induced steatosis is often accompanied by concomitant injury mechanisms such as oxidative stress, cell death, and inflammation, which leads to the development of drug-induced steatohepatitis (DISH). DISH is much more severe from the clinical point of view, has worse prognosis and outcome, and resembles MASH (metabolic-associated steatohepatitis), as it is associated with inflammation and sometimes with fibrosis. A literature review of clinical case reports allowed us to examine and evaluate the clinical features of DIFLD and their association with specific drugs, enabling us to propose a classification of DIFLD drugs based on clinical outcomes and pathological severity: Group 1, drugs with low intrinsic toxicity (e.g., ibuprofen, naproxen, acetaminophen, irinotecan, methotrexate, and tamoxifen), but expected to promote/aggravate steatosis in patients with pre-existing MASLD; Group 2, drugs associated with steatosis and only occasionally with steatohepatitis (e.g., amiodarone, valproic acid, and tetracycline); and Group 3, drugs with a great tendency to transit to steatohepatitis and further to fibrosis. Different mechanisms may be in play when identifying drug mode of action: (1) inhibition of mitochondrial fatty acid β-oxidation; (2) inhibition of fatty acid transport across mitochondrial membranes; (3) increased de novo lipid synthesis; (4) reduction in lipid export by the inhibition of microsomal triglyceride transfer protein; (5) induction of mitochondrial permeability transition pore opening; (6) dissipation of the mitochondrial transmembrane potential; (7) impairment of the mitochondrial respiratory chain/oxidative phosphorylation; (8) mitochondrial DNA damage, degradation and depletion; and (9) nuclear receptors (NRs)/transcriptomic alterations. Currently, the majority of, if not all, adverse outcome pathways (AOPs) for steatosis in AOP-Wiki highlight the interaction with NRs or transcription factors as the key molecular initiating event (MIE). This perspective suggests that chemical-induced steatosis typically results from the interplay between a chemical and a NR or transcription factors, implying that this interaction represents the primary and pivotal MIE. However, upon conducting this exhaustive literature review, it became evident that the current AOPs tend to overly emphasize this interaction as the sole MIE. Some studies indeed support the involvement of NRs in steatosis, but others demonstrate that such NR interactions alone do not necessarily lead to steatosis. This view, ignoring other mitochondrial-related injury mechanisms, falls short in encapsulating the intricate biological mechanisms involved in chemically induced liver steatosis, necessitating their consideration as part of the AOP's map road as well.
Topics: Humans; Fatty Liver; Chemical and Drug Induced Liver Injury; Adverse Outcome Pathways; Liver; Oxidative Stress
PubMed: 38791241
DOI: 10.3390/ijms25105203 -
International Journal of Molecular... May 2024The membrane F factor of ATP synthase is highly sensitive to mutations in the proton half-channel leading to the functional blocking of the entire protein. To identify...
The membrane F factor of ATP synthase is highly sensitive to mutations in the proton half-channel leading to the functional blocking of the entire protein. To identify functionally important amino acids for the proton transport, we performed molecular dynamic simulations on the selected mutants of the membrane part of the bacterial FF-ATP synthase embedded in a native lipid bilayer: there were nine different mutations of -subunit residues (E219, H245, N214, Q252) in the inlet half-channel. The structure proved to be stable to these mutations, although some of them (H245Y and Q252L) resulted in minor conformational changes. H245 and N214 were crucial for proton transport as they directly facilitated H transfer. The substitutions with nonpolar amino acids disrupted the transfer chain and water molecules or neighboring polar side chains could not replace them effectively. E219 and Q252 appeared not to be determinative for proton translocation, since an alternative pathway involving a chain of water molecules could compensate the ability of H transmembrane movement when they were substituted. Thus, mutations of conserved polar residues significantly affected hydration levels, leading to drastic changes in the occupancy and capacity of the structural water molecule clusters (W1-W3), up to their complete disappearance and consequently to the proton transfer chain disruption.
Topics: Lipid Bilayers; Molecular Dynamics Simulation; Mutation; Protein Conformation; Protein Subunits; Bacterial Proton-Translocating ATPases
PubMed: 38791189
DOI: 10.3390/ijms25105143 -
International Journal of Molecular... May 2024Identifying alterations caused by aging could be an important tool for improving the diagnosis of cardiovascular diseases. Changes in vascular tone regulation involve... (Review)
Review
Identifying alterations caused by aging could be an important tool for improving the diagnosis of cardiovascular diseases. Changes in vascular tone regulation involve various mechanisms, like NO synthase activity, activity of the sympathetic nervous system, production of prostaglandin, endothelium-dependent relaxing, and contracting factors, etc. Surprisingly, Ca-dependent Cl channels (CaCCs) are involved in all alterations of the vascular tone regulation mentioned above. Furthermore, we discuss these mechanisms in the context of ontogenetic development and aging. The molecular and electrophysiological mechanisms of CaCCs activation on the cell membrane of the vascular smooth muscle cells (VSMC) and endothelium are explained, as well as the age-dependent changes that imply the activation or inhibition of CaCCs. In conclusion, due to the diverse intracellular concentration of chloride in VSMC and endothelial cells, the activation of CaCCs depends, in part, on intracellular Ca concentration, and, in part, on voltage, leading to fine adjustments of vascular tone. The activation of CaCCs declines during ontogenetic development and aging. This decline in the activation of CaCCs involves a decrease in protein level, the impairment of Ca influx, and probably other alterations in vascular tone regulation.
Topics: Humans; Aging; Animals; Calcium; Muscle, Smooth, Vascular; Chloride Channels; Endothelium, Vascular; Myocytes, Smooth Muscle
PubMed: 38791133
DOI: 10.3390/ijms25105093 -
Antioxidants (Basel, Switzerland) May 2024Dimethyl sulfoxide (DMSO), an organosulfur compound, is widely used as the gold standard solvent in biological research. It is used in cell culture experiments and as a...
Dimethyl sulfoxide (DMSO), an organosulfur compound, is widely used as the gold standard solvent in biological research. It is used in cell culture experiments and as a component of formulations in in vivo studies. Unfortunately, parameters related to sulfur metabolism are often not taken into account when using DMSO. Therefore, in this work we aim to show that the addition of DMSO to the culture medium (even in amounts commonly considered acceptable) alters some parameters of sulfur metabolism. For this study, we used three cell lines: a commercially available Caco-2 line (HTB-37, ATCC) and two lines created as part of our early studies (likewise previously described in the literature) to investigate the anomalies of sulfur metabolism in mucopolysaccharidosis. As the negative effects of DMSO on the cell membrane are well known, additional experiments with the partial loading of DMSO into polymerosomes (poly(ethylene glycol) methyl ether-block-poly(lactide-co-glycolide), PEG-PLGA) were performed to eliminate these potentially disruptive effects. The results show that DMSO is a source of interference in studies related to sulfur metabolism and that there are not just simple effects that can be corrected in the final result by subtracting control values, since complex synergisms are also observed.
PubMed: 38790687
DOI: 10.3390/antiox13050582 -
Current Opinion in Cell Biology Jun 2024Caveolae are atypical plasma membrane invaginations that take part in lipid sorting and regulation of oxidative and mechanical plasma membrane stress. Caveola formation... (Review)
Review
Caveolae are atypical plasma membrane invaginations that take part in lipid sorting and regulation of oxidative and mechanical plasma membrane stress. Caveola formation requires caveolin, cavin, and specific lipid types. The recent advances in understanding the structure and assembly of caveolin and cavin complexes within the membrane context have clarified the fundamental processes underlying caveola biogenesis. In addition, the curvature of the caveola membrane is controlled by the regulatory proteins EHD2, pacsin2, and dynamin2, which also function to restrain the scission of caveolae from the plasma membrane (PM). Here, this is integrated with novel insights on caveolae as lipid and mechanosensing complexes that can dynamically flatten or disassemble to counteract mechanical, and oxidative stress.
Topics: Humans; Caveolae; Cell Membrane; Animals; Caveolins
PubMed: 38788266
DOI: 10.1016/j.ceb.2024.102371 -
Microbial Genomics May 2024Lyme disease (LD), caused by spirochete bacteria of the genus , remains the most common vector-borne disease in the northern hemisphere. outer surface protein A (OspA)...
Lyme disease (LD), caused by spirochete bacteria of the genus , remains the most common vector-borne disease in the northern hemisphere. outer surface protein A (OspA) is an integral surface protein expressed during the tick cycle, and a validated vaccine target. There are at least 20 recognized genospecies, that vary in OspA serotype. This study presents a new sequence-based method for OspA typing using next-generation sequence data. Using a compiled database of over 400 genomes encompassing the 4 most common disease-causing genospecies, we characterized OspA diversity in a manner that can accommodate existing and new OspA types and then defined boundaries for classification and assignment of OspA types based on the sequence similarity. To accommodate potential novel OspA types, we have developed a new nomenclature: OspA type (IST). Beyond the ISTs that corresponded to existing OspA serotypes 1-8, we identified nine additional ISTs that cover new OspA variants in (IST9-10), (IST11-12), and other genospecies (IST13-17). The IST typing scheme and associated OspA variants are available as part of the PubMLST spp. database. Compared to traditional OspA serotyping methods, this new computational pipeline provides a more comprehensive and broadly applicable approach for characterization of OspA type and genospecies to support vaccine development.
Topics: Bacterial Outer Membrane Proteins; Lyme Disease; Lipoproteins; Antigens, Surface; Borrelia burgdorferi; Computer Simulation; Humans; Genome, Bacterial; Borrelia burgdorferi Group; High-Throughput Nucleotide Sequencing; Serogroup; Phylogeny; Bacterial Vaccines
PubMed: 38787376
DOI: 10.1099/mgen.0.001252 -
Current Issues in Molecular Biology May 2024Serine/arginine-rich splicing factors (SRSFs), part of the serine/arginine-rich (SR) protein family, play a crucial role in precursor RNA splicing. Abnormal expression...
Serine/arginine-rich splicing factors (SRSFs), part of the serine/arginine-rich (SR) protein family, play a crucial role in precursor RNA splicing. Abnormal expression of SRSFs in tumors can disrupt normal RNA splicing, contributing to tumor progression. Notably, SRSF7 has been found to be upregulated in hepatocellular carcinoma (HCC), yet its specific role and molecular mechanisms in HCC pathogenesis are not fully understood. We investigated the expression and prognostic significance of SRSF7 in HCC using bioinformatics database analysis. In HepG2 cells, the expressions of SRSF7 and glycolytic enzymes were analyzed using qRT-PCR, and Western blot. Glucose uptake and lactate production were quantified using relevant reagent kits. Additionally, cell proliferation, clonogenicity, invasion, and apoptosis were evaluated using MTS assay, clonal formation assay, Transwell assay, and mitochondrial membrane potential assay, respectively. This study demonstrated significant overexpression of SRSF7 in HCC tissue, correlating with poor prognosis. Knockdown of SRSF7 in HepG2 cells resulted in inhibited proliferation, clonogenicity, and invasion, while apoptosis was enhanced. This knockdown also decreased glucose uptake and lactate production, along with a reduction in the expression of glucose transporter 1 (GLUT1) and lactate dehydrogenase A (LDHA). Furthermore, SRSF7 downregulation increased the pyruvate kinase muscle 1 (PKM1)/PKM2 ratio. The glycolytic boost due to PKM2 overexpression partially counteracted the effects of SRSF7 silencing on HepG2 cell growth. The knockdown of SRSF7 impairs aerobic glycolysis and growth in HepG2 cells by downregulating PKM2 expression.
PubMed: 38785569
DOI: 10.3390/cimb46050301 -
Frontiers in Cellular Neuroscience 2024Engineered 3D neural tissues made of neurons and glial cells derived from human induced pluripotent stem cells (hiPSC) are among the most promising tools in drug...
Engineered 3D neural tissues made of neurons and glial cells derived from human induced pluripotent stem cells (hiPSC) are among the most promising tools in drug discovery and neurotoxicology. They represent a cheaper, faster, and more ethical alternative to animal testing that will likely close the gap between animal models and human clinical trials. Micro-Electrode Array (MEA) technology is known to provide an assessment of compound effects on neural 2D cell cultures and acute tissue preparations by real-time, non-invasive, and long-lasting electrophysiological monitoring of spontaneous and evoked neuronal activity. Nevertheless, the use of engineered 3D neural tissues in combination with MEA biochips still involves series of constraints, such as drastically limited diffusion of oxygen and nutrients within tissues mainly due to the lack of vascularization. Therefore, 3D neural tissues are extremely sensitive to experimental conditions and require an adequately designed interface that provides optimal tissue survival conditions. A well-suited technique to overcome this issue is the combination of the Air-Liquid Interface (ALI) tissue culture method with the MEA technology. We have developed a full 3D neural tissue culture process and a data acquisition system composed of high-end electronics and novel MEA biochips based on porous, flexible, thin-film membranes integrating recording electrodes, named as "Strip-MEA," to allow the maintenance of an ALI around the 3D neural tissues. The main motivation of the porous MEA biochips development was the possibility to monitor and to study the electrical activity of 3D neural tissues under different recording configurations, (i) the Strip-MEA can be placed below a tissue, (ii) or by taking advantage of the ALI, be directly placed on top of the tissue, or finally, (iii) it can be embedded into a larger neural tissue generated by the fusion of two (or more) tissues placed on both sides of the Strip-MEA allowing the recording from its inner part. This paper presents the recording and analyses of spontaneous activity from the three positioning configurations of the Strip-MEAs. Obtained results are discussed with the perspective of developing models of brain diseases and/or impairment of neural network functioning.
PubMed: 38784710
DOI: 10.3389/fncel.2024.1389580