In Vitro Evaluation of Antipseudomonal Activity and Safety Profile of Peptidomimetic Furin Inhibitors.

Inhibitors of the serine protease furin have been widely studied as antimicrobial agents due to their ability to block the cleavage and activation of certain viral surface proteins and bacterial toxins. In this study, the antipseudomonal effects and safety profiles of the furin inhibitors MI-1851 and MI-2415 were assessed. Fluorescence quenching studies suggested no relevant binding of the compounds to human serum albumin and α-acid glycoprotein.

PK/PD investigation of antiviral host matriptase/TMPRSS2 inhibitors in cell models.

Certain corona- and influenza viruses utilize type II transmembrane serine proteases for cell entry, making these enzymes potential drug targets for the treatment of viral respiratory infections. In this study, the cytotoxicity and inhibitory effects of seven matriptase/TMPRSS2 inhibitors (MI-21, MI-463, MI-472, MI-485, MI-1900, MI-1903, and MI-1904) on cytochrome P450 enzymes were evaluated using fluorometric assays. Additionally, their antiviral activity against influenza A virus subtypes H1N1 and H9N2 was assessed.

Antiviral Drug Candidate Repositioning for Infection in Non-Tumorigenic Cell Models.

The increasing prevalence of antimicrobial resistance against zoonotic bacteria, including , highlights the need for new therapeutical strategies, including the repositioning of drugs. In this study, susceptibilities of bacterial isolates were tested toward ten different 3-amidinophenyalanine (Phe(3-Am)) derivatives via determination of minimum inhibitory concentration (MIC) values. Some of these protease inhibitors, like compounds MI-432, MI-471, and MI-476, showed excellent antibacterial effects against .

In vitro testing of host-targeting small molecule antiviral matriptase/TMPRSS2 inhibitors in 2D and 3D cell-based assays.

The outbreak of coronavirus disease 2019 (COVID-19) pandemic strongly stimulated the development of small molecule antivirals selectively targeting type II transmembrane serine proteases (TTSP), required for the host-cell entry of numerous viruses. A set of 3-amidinophenylalanine derivatives (MI-21, MI-472, MI-477, MI-485, MI-1903 and MI-1904), which inhibit the cleavage of certain viral glycoproteins was characterized in 2D and 3D primary human hepatocyte models on collagen- and Matrigel-coating using a CCK-8 assay to evaluate their cytotoxicity, a resorufin-based method to detect redox imbalances, fluorescence and ultrafiltration experiments to evaluate their interactions with human serum albumin (HSA) and α-acidic glycoprotein (AGP), and luminescence measurement to assess CYP3A4 modulation. For elucidation of selectivity of the applied compounds towards matriptase, transmembrane serine protease 2 (TMPRRS2), thrombin and factor Xa (FXa) K values were determined.

Reductive Treatment of Pt Supported on TiSnO-C Composite: A Route for Modulating the Sn-Pt Interactions.

The composites of transition metal-doped titania and carbon have emerged as promising supports for Pt electrocatalysts in PEM fuel cells. In these multifunctional supports, the oxide component stabilizes the Pt particles, while the dopant provides a co-catalytic function. Among other elements, Sn is a valuable additive.

In Vitro Pharmacokinetic Behavior of Antiviral 3-Amidinophenylalanine Derivatives in Rat, Dog and Monkey Hepatocytes.

Type II transmembrane serine proteases represent pharmacological targets for blocking entry and spread of influenza or coronaviruses. In this study, the depletion rates of the 3-amidinophenylalanine (3-APhA)-derived matriptase/TMPRSS2 inhibitors MI-463, MI-472, MI-485 or MI-1900 were determined by LC-MS/MS measurements over a period of 300 min using suspensions of rat, dog and cynomolgus monkey primary hepatocytes. From these in vitro pharmacokinetic (PK) experiments, intrinsic clearance values (Cl) were evaluated, and in vivo pharmacokinetic parameters (hepatic clearance, hepatic extraction ratio and bioavailability) were predicted.

Luteolin and Chrysin Could Prevent Lipopolysaccharide-Ochratoxin A Combination-Caused Inflammation and Oxidative Stress in In Vitro Porcine Intestinal Model.

Ochratoxin A (OTA) and lipopolysaccharide (LPS) intake can cause gastrointestinal disorders. Polyphenolic chrysin (CHR) and luteolin (LUT) display anti-inflammatory and antioxidant properties. Porcine intestinal epithelial (jejunal) IPEC-J2 cells were treated with OTA (1 µM, 5 µM and 20 µM), E.

In vitro characterization of the furin inhibitor MI-1851: Albumin binding, interaction with cytochrome P450 enzymes and cytotoxicity.

The substrate-analog furin inhibitor MI-1851 can suppress the cleavage of SARS-CoV-2 spike protein and consequently produces significant antiviral effect on infected human airway epithelial cells. In this study, the interaction of inhibitor MI-1851 was examined with human serum albumin using fluorescence spectroscopy and ultrafiltration techniques. Furthermore, the impacts of MI-1851 on human microsomal hepatic cytochrome P450 (CYP) 1A2, 2C9, 2C19, 2D6 and 3A4 activities were assessed based on fluorometric assays.

In vitro interaction of potential antiviral TMPRSS2 inhibitors with human serum albumin and cytochrome P 450 isoenzymes.

The interactions of four sulfonylated Phe(3-Am)-derived inhibitors (MI-432, MI-463, MI-482 and MI-1900) of type II transmembrane serine proteases (TTSP) such as transmembrane protease serine 2 (TMPRSS2) were examined with serum albumin and cytochrome P450 (CYP) isoenzymes. Complex formation with albumin was investigated using fluorescence spectroscopy. Furthermore, microsomal hepatic CYP1A2, 2C9, 2C19 and 3A4 activities in presence of these inhibitors were determined using fluorometric assays.

Investigation of the inflammatory and oxidative stress-inducing effects of deoxynivalenol and T-2 toxin exposure in non-tumorigenic human intestinal cell model.

Fungi in the Fusarium genus produce trichothecene mycotoxins including deoxynivalenol (DON) and T-2 toxin which may elicit their damaging effects on the gastrointestinal tract following the consumption of contaminated cereal-based foods. The aim of our study was to evaluate the effects of these commonly occurring fusarotoxins alone and in combination using the human, non-cancerous intestinal epithelial cell line HIEC-6. Based on our experimental data, 24 h after treatment with fusarotoxins, hydrogen peroxide levels, intracellular oxidative stress and the amounts of inflammatory interleukins IL-6 and IL-8 significantly increased.

The Effects of Matriptase Inhibition on the Inflammatory and Redox Homeostasis of Chicken Hepatic Cell Culture Models.

The function of the transmembrane serine protease matriptase is well described in mammals, but it has not been elucidated in avian species yet. Hence, the aim of the present study was to assess the effects of the 3-amidinophenylalanine (3-AphA)-type matriptase inhibitors MI432 and MI460 on the inflammatory and oxidative state of chicken primary hepatocyte mono-cultures and hepatocyte-nonparenchymal cell co-cultures, the latter serving as a proper model of hepatic inflammation in birds. Cell cultures were exposed to MI432 and MI460 for 4 and 24 h at 10, 25, and 50 µM concentrations, and thereafter the cellular metabolic activity, extracellular interleukin (IL-)6, IL-8, HO and malondialdehyde concentrations were monitored.

Exposure of human intestinal epithelial cells and primary human hepatocytes to trypsin-like serine protease inhibitors with potential antiviral effect.

Human intestinal epithelial cell line-6 (HIEC-6) cells and primary human hepatocytes (PHHs) were treated with 3-amidinophenylalanine-derived inhibitors of trypsin-like serine proteases for 24 hours. It was proven that treatment with MI-1900 and MI-1907 was tolerated up to 50 μM in HIEC-6. These inhibitors did not cause elevations in extracellular HO levels and in the concentrations of interleukin (IL)-6 and IL-8 and did not alter occludin distribution in HIEC-6.

The impact of quercetin on a porcine intestinal epithelial cell line exposed to deoxynivalenol.

Quercetin (Que) is present in many vegetables and fruits as a secondary antioxidant metabolite. Deoxynivalenol (DON) produced by various Fusarium mould species can induce cytotoxicity and oxidative stress in the gastrointestinal tracts of humans and farm animals. The aim of this study was to investigate the effects of Que on DON-induced oxidative stress in a non-tumourigenic porcine IPEC-J2 cell line.

The Impact of Fermented Wheat Germ Extract on Porcine Epithelial Cell Line Exposed to Deoxynivalenol and T-2 Mycotoxins.

The effect of fermented wheat germ extract (FWGE) () was evaluated with cotreatments with deoxynivalenol (DON) and T-2 toxin (T-2). These mycotoxins are produced by mold species. The effects of FWGE on IPEC-J2 with DON and T-2 have not been studied until now.

3-Amidinophenylalanine-derived matriptase inhibitors can modulate hepcidin production in vitro.

Matriptase-2 (MT-2) is a type II transmembrane serine protease and predominantly attached to the surface of hepatocytes. MT-2 decreases the production of hepcidin, a key regulator of iron homeostasis. In this study, the effects of four 3-amidinophenylalanine-derived combined matriptase-1/matriptase-2 (MT-1/2) inhibitors (MI-432, MI-441, MI-460, and MI-461) on hepcidin production were investigated in hepatocyte mono- and hepatocyte-Kupffer cell co-cultures.

Chlorogenic Acid Combined with Lactobacillus plantarum 2142 Reduced LPS-Induced Intestinal Inflammation and Oxidative Stress in IPEC-J2 Cells.

This study was carried out to investigate protective effect of chlorogenic acid against lipopolysaccharide-induced inflammation and oxidative stress in intestinal epithelial cells. As a marker of inflammatory response, IL-6, IL-8, TNF-α mRNA and protein levels, furthermore, COX-2 mRNA level were followed up. Intracellular redox status and extracellular H2O2 level were also monitored by two fluorescent assays (DCFH-DA, Amplex Red).

The Impact of Acute Matriptase Inhibition in Hepatic Inflammatory Models.

Purpose. Dysfunction of matriptase-2 can be involved in iron regulatory disorder via downregulation of hepcidin expression. In the present study, we investigated the effects of 3-amidinophenylalanine-derived matriptase inhibitors on porcine hepatic inflammatory cell models.

In vitro characterization of TMPRSS2 inhibition in IPEC-J2 cells.

The transmembrane serine protease, TMPRSS2 is an important target in the treatment of seasonal influenza infections and contributes to prostate carcinogenesis and metastasis. In this study, the effect of the synthetic TMPRSS2 inhibitor I-432 on jejunal IPEC-J2 cell monolayers cultured on membrane inserts was characterized. Using a fluorogenic substrate, it was found that the apical addition of I-432 could suppress trypsin-like activity in the supernatants of IPEC-J2 cells.

Polymethoxyflavone Apigenin-Trimethylether Suppresses LPS-Induced Inflammatory Response in Nontransformed Porcine Intestinal Cell Line IPEC-J2.

The in vitro anti-inflammatory effect of apigenin and its trimethylated analogue (apigenin-trimethylether) has been investigated in order to evaluate whether these flavonoids could attenuate LPS-induced inflammation in IPEC-J2 non-transformed intestinal epithelial cells. Levels of IL-6, IL-8, TNF-α, and COX-2 mRNA were measured as a marker of inflammatory response. The extracellular H2O2 level in IPEC-J2 cells was also monitored by Amplex Red assay.

Interaction exists between matriptase inhibitors and intestinal epithelial cells.

The type II trypsin-like transmembrane serine protease matriptase, is mainly expressed in epithelial cells and one of the key regulators in the formation and maintenance of epithelial barrier integrity. Therefore, we have studied the inhibition of matriptase in a non-transformed porcine intestinal IPEC-J2 cell monolayer cultured on polyester membrane inserts by the non-selective 4-(2-aminoethyl)-benzosulphonylfluoride (AEBSF) and four more selective 3-amidinophenylalanine-derived matriptase inhibitors. It was found that suppression of matriptase activity by MI-432 and MI-460 led to decreased transepithelial electrical resistance (TER) of the cell monolayer and to an enhanced transport of fluorescently labelled dextran, a marker for paracellular transport between apical and basolateral compartments.

Gentamicin sulphate permeation through porcine intestinal epithelial cell monolayer.

Gentamicin is an aminoglycoside antibiotic widely used in combination with dimethyl sulphoxide (DMSO) in topical drug formulations. It is not known, however, whether DMSO can enhance the permeation of gentamicin through biological membranes, leading to oto- and nephrotoxic side effects. A simple and reliable high-performance liquid chromatographic (HPLC) method was applied for the quantitative determination of gentamicin collected from the apical and basolateral compartments of the porcine intestinal epithelial cell line IPEC-J2 cell monolayer using fluorometric derivatisation of the analyte with fluorenylmethyloxycarbonyl chloride (FMOC) prior to chromatographic run in the presence and absence of 1% DMSO.

Changes in the distribution of type II transmembrane serine protease, TMPRSS2 and in paracellular permeability in IPEC-J2 cells exposed to oxidative stress.

The effect of oxidative stress on barrier integrity and localization of transmembrane serine proteinase 2 (TMPRSS2) were studied using porcine epithelial IPEC-J2 cells on membrane inserts. Increased paracellular permeability of FITC-dextran 4 kDa (fluorescence intensity 43,508 ± 2,391 versus 3,550 ± 759) and that of gentamicin (3.41 ± 0.

The effects of intestinal LPS exposure on inflammatory responses in a porcine enterohepatic co-culture system.

A porcine enterohepatic co-culture system, with primary hepatocytes as bottom layer and IPEC-J2 epithelial cells as upper layer, was developed to study the effects of lipopolysaccharides (LPS) on the gene expression profile of pro-inflammatory cytokines (interleukin-8 (IL-8) and tumor necrosis factor-α) and CYP enzymes (CYP1A1, CYP1A2, CYP3A29). The barrier integrity of IPEC-J2 cells was investigated by transepithelial electrical resistance measurements and by fluorescein isothiocyanate-dextran-based test. Basolateral IL-8 production was significantly elevated in LPS-treated IPEC-J2 and primary hepatocyte mono-cultures as well as in the co-culture system, in a dose-independent manner.

Protein phosphatases but not reactive oxygen species play functional role in acute amphetamine-mediated dopamine release.

Drug abuse-induced neurodegeneration can be triggered by elevated production of reactive oxygen species (ROS). Involvement of oxidative stress in acute amphetamine (AMPH)-mediated dopamine (DA) release, however, has not been completely understood yet. In order to elucidate the dopaminergic response of PC12 cells to a single dose of 10 μM AMPH, ROS production was measured as related to the extracellular DA level.