Herein, we investigated the anti-amoebic activity of phosphonium-chloride-based deep eutectic solvents against pathogenic Acanthamoeba castellanii of the T4 genotype. Deep eutectic solvents are ionic fluids composed of two or three substances, capable of self-association to form a eutectic mixture with a melting point lower than each substance. In this study, three distinct hydrophobic deep eutectic solvents were formulated, employing trihexyltetradecylphosphonium chloride as the hydrogen bond acceptor and aspirin, dodecanoic acid, and 4-tert-butylbenzoic acid as the hydrogen bond donors. Subsequently, all three deep eutectic solvents, denoted as DES1, DES2, DES3 formulations, underwent investigations comprising amoebicidal, adhesion, excystation, cytotoxicity, and cytopathogenicity assays. The findings revealed that DES2 was the most potent anti-amoebic agent, with a 94% elimination rate against the amoebae within 24 h at 30 °C. Adhesion assays revealed that deep eutectic solvents hindered amoebae adhesion to human brain endothelial cells, with DES2 exhibiting 88% reduction of adhesion. Notably, DES3 exhibited remarkable anti-excystation properties, preventing 94% of cysts from reverting to trophozoites. In cytopathogenicity experiments, deep eutectic solvent formulations and dodecanoic acid alone reduced amoebae-induced human brain endothelial cell death, with DES2 showing the highest effects. Lactate dehydrogenase assays revealed the minimal cytotoxicity of the tested deep eutectic solvents, with the exception of trihexyltetradecylphosphonium chloride, which exhibited 35% endothelial cell damage. These findings underscore the potential of specific deep eutectic solvents in combating pathogenic Acanthamoeba, presenting promising avenues for further research and development against free-living amoebae.
- MeSH
- Acanthamoeba castellanii * účinky léků genetika MeSH
- amébicidy farmakologie chemie MeSH
- buněčná adheze účinky léků MeSH
- endoteliální buňky účinky léků MeSH
- genotyp * MeSH
- lidé MeSH
- organofosforové sloučeniny farmakologie chemie MeSH
- rozpouštědla * chemie farmakologie MeSH
- trofozoiti účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Waterpipe smoking (WPS) has adverse health effects that include endothelial dysfunction with mechanisms involving oxidative stress and inflammation. Nonetheless, there is a scarcity of data on the direct impact of WPS on endothelial function. In this study, we assessed the in vitro effects of waterpipe smoke extract (WPSE) on aortic endothelial cell lines, namely the TeloHAEC. The WPSE markedly caused concentration- and time-dependent decreases in cellular viability. When compared with the control, at a concentration of 20 % and an incubation period of 48 h, the WPSE significantly increased the levels of lactate dehydrogenase, and markers of oxidative stress including thiobarbituric acid reactive substances, superoxide dismutase, catalase, and reduced glutathione. Moreover, the concentrations of proinflammatory cytokine (tumor necrosis factor alpha), and adhesion molecules (E-selectin and intercellular adhesion molecule-1) were also significantly augmented. Likewise, WPSE triggered mitochondrial dysfunction, DNA oxidative damage, as well as apoptosis in TeloHAEC cells. Similarly, cells cultured with WPSE have shown increased expression of phosphorylated nuclear factor-kappaB and hypoxia-inducible factor 1-alpha (HIF-1alpha). In conclusion, our study showed that WPSE triggers endothelial inflammation, oxidative stress, DNA damage, mitochondrial dysfunction, and apoptosis via mechanisms involving the activation of nuclear factor-kappaB and HIF-1alpha. Key words Waterpipe smoking, Aortic endothelial cells, Inflammation, Oxidative Stress.
- MeSH
- aorta * účinky léků metabolismus MeSH
- apoptóza účinky léků MeSH
- buněčné linie MeSH
- endoteliální buňky * účinky léků metabolismus MeSH
- kouř * škodlivé účinky MeSH
- kouření vodní dýmky * škodlivé účinky metabolismus MeSH
- lidé MeSH
- oxidační stres * účinky léků MeSH
- poškození DNA účinky léků MeSH
- viabilita buněk účinky léků MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Liver sinusoidal endothelial inflammation/dysfunction and fibrosis are a crucial part of Metabolic Dysfunction Associated Steatohepatitis (MASH) development. TRC105 and M1043 are anti-endoglin (ENG) monoclonal antibodies that bind ENG. In this study, we hypothesized that treatment with anti-ENG antibodies would prevent the progression of LSECs inflammation and fibrosis in vivo and in vitro. MASH was induced in male C57BL/6 mice fed a choline-deficient L-amino acid-defined high-fat diet (CDAA-HFD) for 4 or 8 weeks. In the rescue study, mice were divided into three groups: a control group (chow diet), a MASH group (CDAA-HFD + IgG), and a rescue group (CDAA-HFD + M1043). Later, two groups received rat IgG1 (10 mg/kg) and M1043 (10 mg/kg). In in vitro experiments, inflammation was induced in human LSECs by ox-LDL (50 μg/mL) and treated with TRC105 (300 μg/mL). Liver sinusoidal endothelial inflammation/dysfunction in MASH animals was characterized by endothelial overexpression of ENG, VCAM-1, and ICAM-1 and reduced VE-cadherin and p-eNOS/eNOS expression. M1043 treatment prevented the overexpression of ENG, VCAM-1, and ICAM-1, the progression of liver fibrosis, and the increase of liver-to-body weight ratio. In vitro experiments with TRC105 confirmed the prevention of LSECs inflammation development by reduced ENG and VCAM-1 expression, as well as decreased THP-1 monocytic cell adhesion in ox-LDL activated LSECs. In conclusion, we demonstrate that anti-ENG antibody treatment can prevent LSECs inflammation and fibrosis progression in a MASH animal model and LSECs inflammation in vitro. Thus, we propose directly targeted ENG may represent a promising pharmacological approach for addressing LSECs inflammation and liver fibrosis.
- MeSH
- dieta s vysokým obsahem tuků škodlivé účinky MeSH
- endoglin * metabolismus antagonisté a inhibitory MeSH
- endoteliální buňky účinky léků metabolismus patologie MeSH
- jaterní cirhóza * prevence a kontrola patologie farmakoterapie metabolismus MeSH
- játra * patologie účinky léků metabolismus MeSH
- krysa rodu rattus MeSH
- lidé MeSH
- monoklonální protilátky * farmakologie MeSH
- myši inbrední C57BL * MeSH
- myši MeSH
- nealkoholová steatóza jater farmakoterapie prevence a kontrola patologie metabolismus MeSH
- progrese nemoci MeSH
- zánět * patologie farmakoterapie metabolismus prevence a kontrola MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé MeSH
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Ciel: Zistiť efekt opakovaných intravitreálnych aplikácii afliberceptu na endotel rohovky u pacientov s diabetickým makulárnym edémom (DME) a edémom makuly pri oklúzii retinálnej vény (RVO). Metodika: Do prospektívneho sledovania v období od januára 2021 do novembra 2023 bolo zaradených 87 naivných očí s diagnózou DME a RVO. Exklúzne kritérium pre zaradenie bol operačný alebo laserový zákrok počas sledovacieho obdobia, nosenie kontaktných šošoviek, ope rácia katarakty pred menej ako 6 mesiacmi, dystrofie alebo iné ochorenia rohovky, ktoré môžu spôsobiť zmeny endotelu. Okrem rutinných vyšetrení sme v deň 1., 4. a 8. injekcie vyšetrovali aj endotel rohovky pomocou bezkontaktnej endotelovej mikroskopie. Vyhodnocovali sme 4 parametre: hustotu endotelových buniek (CD), hexagonalitu (HEX), koeficient variability (CV) a centrálnu hrúbku rohovky (CCT). Najskôr sme vyhodnocovali celý súbor očí a následne sme ho rozdelili podľa 2 kritérií; podľa diagnózy na DME/RVO a podľa šošovky na fakické/pseu dofakické oči. Výsledky: Vyhodnotených bolo 87 očí (68 pacientov). Priemerný vek pacientov v čase diagnózy bol 66,8 ±9,3 rokov. Z celkového počtu 87 bolo 51 (59 %) fakických a 36 (41 %) pseudofakických očí. S diagnózou DME bolo liečených 61 (70 %) očí a s diagnózou RVO 26 (30 %). V prie behu sledovania nedošlo vplyvom liečby k štatisticky signifikantnej zmene priemerných hodnôt CD, HEX, CV, CCT či už v súbore všetkých očí alebo v rozdelení do podskupín podľa diagnózy či stavu šošovky. Záver: Zistili sme, že intravitreálne aplikácie afliberceptu u pacientov s DME a RVO neovplyvňujú parametre rohovkového endotelu ako CCT, HEX, CD, CV hodnotené pomocou endotelovej mikroskopie pri sledovaní po 8 injekciu.
Aim: To determine the effect of repeated intravitreal injections of aflibercept on the corneal endothelium in patients with diabetic macular edema (DME) and macular edema due to retinal vein occlusion (RVO). Methods: In a prospective study conducted between January 2021 and November 2023, a total of 87 treatment-naive eyes with DME and RVO were evaluated. The exclusion criteria were surgery or laser intervention during the follow-up period, contact lens wear, cataract surgery in the last 6 months, dystrophy, or other corneal condition that may cause endothelial damage. In addition to routine examinations on the day of application, we also measured the corneal endothelium using specular microscopy on the 1st, 4th and 8th day of injection. We evaluated 4 parameters: endothelial cell density (CD), hexagonality (HEX), coefficient of variability (CV) and central corneal thickness (CCT). First of all, we evaluated the entire cohort of eyes, and then divided it according to 2 criteria; the diagnosis into DME/RVO and according to the lens status into phakic/pseudophakic eyes. Results: A total of 87 eyes of 68 patients were evaluated. The average age of the patients at the time of diagnosis was 66.8 ±9.3 years. Within the cohort 51 (59%) eyes were phakic and 36 (41%) pseudophakic. A total of 61 (70%) eyes with a diagnosis of DME were treated, and 26 (30%) with RVO. During the follow-up, there were no significant changes in the average values of CD, HEX, CV, CCT due to aflibercept treatment, either in the whole group or in subgroups according to diagnosis or lens condition. Conclusions: The results of this study suggest that intravitreal administration of aflibercept in patients with DME and RVO did not have an impact on corneal endothelial parameters, including CCT, HEX, CD and CV. These parameters were measured using endothelial microscopy during an 8-injection observation period.
- Klíčová slova
- aflibercept,
- MeSH
- bevacizumab aplikace a dávkování farmakologie terapeutické užití MeSH
- endoteliální buňky účinky léků MeSH
- humanizované monoklonální protilátky farmakologie klasifikace terapeutické užití MeSH
- inhibitory angiogeneze * aplikace a dávkování farmakologie klasifikace terapeutické užití MeSH
- injekce intravitreální * klasifikace metody MeSH
- lidé MeSH
- makulární edém chemicky indukované etiologie MeSH
- mikroskopie klasifikace metody MeSH
- počet buněk MeSH
- prospektivní studie MeSH
- rohovkový endotel diagnostické zobrazování účinky léků MeSH
- vaskulární endoteliální růstový faktor A antagonisté a inhibitory MeSH
- Check Tag
- lidé MeSH
The global shortage of corneal endothelial graft tissue necessitates the exploration of alternative therapeutic strategies. Rho-associated protein kinase inhibitors (ROCKi), recognized for their regenerative potential in cardiology, oncology, and neurology, have shown promise in corneal endothelial regeneration. This study investigates the repurposing potential of additional ROCKi compounds. Through screening a self-assembled library of ROCKi on B4G12 corneal endothelial cells, we evaluated their dose-dependent effects on proliferation, migration, and toxicity using live-cell imaging. Nine ROCKi candidates significantly enhanced B4G12 proliferation compared to the basal growth rate. These candidates were further assessed for their potential to accelerate wound closure as another indicator for tissue regeneration capacity, with most demonstrating notable efficacy. To assess the potential impact of candidate ROCKi on key corneal endothelial cell markers related to cell proliferation, leaky tight junctions and ion efflux capacity, we analyzed the protein expression of cyclin E1, CDK2, p16, ZO-1 and Na+/K+-ATPase, respectively. Immunocytochemistry and western blot analysis confirmed the preservation of corneal endothelial markers post-treatment with ROCKi hits. However, notable cytoplasm enlargement and nuclear fragmentation were detected after the treatment with SR-3677 and Thiazovivin, indicating possible cellular stress. In compared parameters, Chroman-1 at a concentration of 10 nM outperformed other ROCKi, requiring significantly 1000-fold lower effective concentration than established ROCKi Y-27632 and Fasudil. Altogether, this study underscores the potential of repurposing ROCKi for treating corneal endothelial dysfunctions, offering a viable alternative to conventional grafting methods, and highlights Chroman-1 as a promising candidate structure for hit-to-lead development.
- MeSH
- buněčné linie MeSH
- endoteliální buňky účinky léků MeSH
- inhibitory proteinkinas * farmakologie MeSH
- kinázy asociované s rho * antagonisté a inhibitory metabolismus MeSH
- lidé MeSH
- pohyb buněk účinky léků MeSH
- přehodnocení terapeutických indikací léčivého přípravku MeSH
- proliferace buněk * účinky léků MeSH
- regenerace * účinky léků MeSH
- rohovkový endotel * účinky léků MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Drug delivery to central nervous pathologies is compromised by the blood-brain barrier (BBB). A clinically explored strategy to promote drug delivery across the BBB is sonopermeation, which relies on the combined use of ultrasound (US) and microbubbles (MB) to induce temporally and spatially controlled opening of the BBB. We developed an advanced in vitro BBB model to study the impact of sonopermeation on the delivery of the prototypic polymeric drug carrier pHPMA as a larger molecule and the small molecule antiviral drug ribavirin. This was done under standard and under inflammatory conditions, employing both untargeted and RGD peptide-coated MB. The BBB model is based on human cerebral capillary endothelial cells and human placental pericytes, which are co-cultivated in transwell inserts and which present with proper transendothelial electrical resistance (TEER). Sonopermeation induced a significant decrease in TEER values and facilitated the trans-BBB delivery of fluorescently labeled pHPMA (Atto488-pHPMA). To study drug delivery under inflamed endothelial conditions, which are typical for e.g. tumors, neurodegenerative diseases and CNS infections, tumor necrosis factor (TNF) was employed to induce inflammation in the BBB model. RGD-coated MB bound to and permeabilized the inflamed endothelium-pericyte co-culture model, and potently improved Atto488-pHPMA and ribavirin delivery. Taken together, our work combines in vitro BBB bioengineering with MB-mediated drug delivery enhancement, thereby providing a framework for future studies on optimization of US-mediated drug delivery to the brain.
- MeSH
- antivirové látky aplikace a dávkování chemie farmakologie farmakokinetika MeSH
- endoteliální buňky * účinky léků metabolismus MeSH
- hematoencefalická bariéra * metabolismus MeSH
- kokultivační techniky * MeSH
- lékové transportní systémy metody MeSH
- lidé MeSH
- mikrobubliny * MeSH
- oligopeptidy * chemie aplikace a dávkování farmakokinetika MeSH
- pericyty * metabolismus účinky léků MeSH
- polymery chemie aplikace a dávkování MeSH
- ribavirin aplikace a dávkování chemie farmakokinetika MeSH
- ultrazvukové vlny MeSH
- zánět farmakoterapie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Cíl: Analyzovat souvislost mezi podáním reversinu a zvýšenou plasticitou buněk DFAT schopných dělení na různé typy buněk. Metoda: Vykultivované buňky DFAT byly rozděleny do čtyř skupin podle dávky reversinu: na kontrolní (bez reversinu) skupinu a na skupiny s aplikací reversinu v dávkách 10 nM, 20 nM a 40 nM. Každá skupina prochází několika stadii vývoje před další diferenciací na kardiomyocyty (identifikované expresí cTnT), buňky hladké svaloviny (vascular smooth muscle cells, VSMC) (označené expresí afta-SMA) a buňky cévního endotelu (identifikované expresí CD31). Výsledek: V každé skupině buněk DFAT s aplikací reversinu byly nalezeny statisticky významné rozdíly v expresi cTnT, alfa-SMA a CD31 (p = 0,003; resp. < 0,001 a < 0,001). Post hoc analýza s použitím Tukeyova testu prokázala, že pouze ve skupině s reversinem v dávce 10 nM došlo ke statisticky významnému rozdílu oproti kontrolní skupině (p = 0,002) v expresi cTnT a ve skupinách s reversinem v dávkách 10 nM a 20 nM k rozdílu v expresi alfa-SMA a CD31 (p = 0,028, resp. p < 0,001). Závěry: Tato studie prokázala vztah mezi dávkou reversinu a zvýšenou plasticitou buněk DFAT schopných diferenciace na kardiomyocyty (cTnT), VSMC (alfa-SMA) a buňky cévního endotelu (CD31).
Aim: To analyze the association between reversine and increased plasticity of DFAT into cardiac derivative cells. Method: The cultured DFAT cells were divided into four groups based on reversine dose: control (no reversine), 10 nM, 20 nM, and 40 nM reversine. Each group will go through several stages of passage before further differentiation into cardiomyocytes (marked by cTnT expression), VSCMs (marked by alpha-SMA expression), and vascular endothelial cells (marked by alpha-SMA expression) (marked by CD31 expression). Result: There were significant differences in the expression of cTnT, alpha-SMA, and CD31 (p = 0.003, p <0.001, and p <0.001, respectively) in each group of DFAT cells that received reversine. From post-hoc analysis with Tukey test, it was found that only the 10 nM reversine group produced a significant difference compared to the control group (p = 0.002) for cTnT expression and reversine 10 nM and 20 nM group for α-SMA expression and CD31 expression (p = 0.028 and p <0.001, respectively). Conclusions: This study proves that there is a relationship between reversine and increased plasticity of DFAT cells into cardiac derived cells in the form of cardiomyocytes (cTnT), VSMCs (alpha-SMA), and vascular endothelial cells (CD31).
Monocyte homing to the liver and adhesion to the liver sinusoidal endothelial cells (LSECs) are key elements in nonalcoholic steatohepatitis (NASH) pathogenesis. We reported previously that VCAM-1 mediates monocyte adhesion to LSECs. However, the pathogenic role of VCAM-1 in NASH is unclear. Herein, we report that VCAM-1 was a top upregulated adhesion molecule in the NASH mouse liver transcriptome. Open chromatin landscape profiling combined with genome-wide transcriptome analysis showed robust transcriptional upregulation of LSEC VCAM-1 in murine NASH. Moreover, LSEC VCAM-1 expression was significantly increased in human NASH. LSEC VCAM-1 expression was upregulated by palmitate treatment in vitro and reduced with inhibition of the mitogen-activated protein 3 kinase (MAP3K) mixed lineage kinase 3 (MLK3). Likewise, LSEC VCAM-1 expression was reduced in the Mlk3-/- mice with diet-induced NASH. Furthermore, VCAM-1 neutralizing Ab or pharmacological inhibition attenuated diet-induced NASH in mice, mainly via reducing the proinflammatory monocyte hepatic population as examined by mass cytometry by time of flight (CyTOF). Moreover, endothelium-specific Vcam1 knockout mice were also protected against NASH. In summary, lipotoxic stress enhances the expression of LSEC VCAM-1, in part, through MLK3 signaling. Inhibition of VCAM-1 was salutary in murine NASH and might serve as a potential therapeutic strategy for human NASH.
- MeSH
- cévní buněčněadhezivní molekula-1 antagonisté a inhibitory genetika metabolismus MeSH
- endoteliální buňky účinky léků metabolismus MeSH
- játra účinky léků metabolismus patologie MeSH
- lidé MeSH
- MAP kinasový signální systém účinky léků MeSH
- messenger RNA genetika MeSH
- modely nemocí na zvířatech MeSH
- myši inbrední C57BL MeSH
- myši knockoutované MeSH
- myši MeSH
- nealkoholová steatóza jater etiologie genetika metabolismus MeSH
- neutralizující protilátky aplikace a dávkování MeSH
- palmitany toxicita MeSH
- stanovení celkové genové exprese MeSH
- upregulace účinky léků MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Silicon dioxide, in the form of nanoparticles, possesses unique physicochemical properties (size, shape, and a large surface to volume ratio). Therefore, it is one of the most promising materials used in biomedicine. In this paper, we compare the biological effects of both mesoporous silica nanoparticles extracted from Urtica dioica L. and pyrogenic material. Both SEM and TEM investigations confirmed the size range of tested nanoparticles was between 6 and 20 nanometers and their amorphous structure. The cytotoxic activity of the compounds and intracellular ROS were determined in relation to cells HMEC-1 and erythrocytes. The cytotoxic effects of SiO2 NPs were determined after exposure to different concentrations and three periods of incubation. The same effects for endothelial cells were tested under the same range of concentrations but after 2 and 24 h of exposure to erythrocytes. The cell viability was measured using spectrophotometric and fluorimetric assays, and the impact of the nanoparticles on the level of intracellular ROS. The obtained results indicated that bioSiO2 NPs, present higher toxicity than pyrogenic NPs and have a higher influence on ROS production. Mesoporous silica nanoparticles show good hemocompatibility but after a 24 h incubation of erythrocytes with silica, the increase in hemolysis process, the decrease in osmotic resistance of red blood cells, and shape of erythrocytes changed were observed.
- MeSH
- endoteliální buňky účinky léků MeSH
- erytrocyty účinky léků MeSH
- hemolýza účinky léků MeSH
- lidé MeSH
- nanočástice aplikace a dávkování chemie MeSH
- oxid křemičitý aplikace a dávkování chemie MeSH
- oxidační stres účinky léků MeSH
- poréznost MeSH
- povrchové vlastnosti MeSH
- reaktivní formy kyslíku metabolismus MeSH
- viabilita buněk MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: The aim of our study was to assess the impact of different thawing protocols on morphological changes arising in cryopreserved human saphenous vein grafts. METHODS: The study was performed in 12 saphenous vein grafts harvested in brain death donors. Storage in the vapor phase of liquid nitrogen for 3 or 5 years followed. Two thawing protocols were tested: slow thawing in a refrigerator at temperature +4°C for 2 hr and rapid thawing-in a water bath at +37°C. Grafts were processed for scanning electron microscopy. Comparisons of continuous parameters under study between experimental groups were performed using the t-test (age, cold ischemia time, exposure to cryoprotectant, time of storage, total thawing time, mean thawing rate, morphology scoring of thawed HSVG) and the median test (HSVG length). Categorical parameters (sex and blood group) were formally tested using the chi-square test. RESULTS: All samples were evaluated according to morphological changes and scored in terms of morphologically intact endothelium, confluent endothelium with structural inhomogeneity, disruption of the intercellular contacts, separation of the endothelial cells, complete loss of the endothelium, and damage of the subendothelial layers. There is no statistically significant difference between the sample sets at the significance level of 0.05. There was no association with donors' age, sex, and time of storage. CONCLUSIONS: Human cryopreserved saphenous vein grafts in our experimental work showed no difference in terms of structural deterioration of the endothelial surface and basal membrane depending on different thawing protocols used.
- MeSH
- časové faktory MeSH
- dospělí MeSH
- endoteliální buňky účinky léků transplantace ultrastruktura MeSH
- kryoprezervace * MeSH
- kryoprotektivní látky farmakologie MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- odběr tkání a orgánů MeSH
- přežití tkáně MeSH
- vena saphena účinky léků transplantace ultrastruktura MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- srovnávací studie MeSH