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

Diabetic kidney disease (DKD) is the leading cause of end-stage renal disease. Therefore, understanding the molecular regulatory mechanisms underlying the pathogenesis of DKD is imperative. In this study, we aimed to explore the molecular mechanisms of tubule region endothelial dysfunction in early DKD. Early-stage DKD model was established in 16-week-old female db/db mice for 16 weeks. Body weight, glucose level, and urine albumin-to-creatinine ratio (UACR) were measured. Hematoxylin and eosin (H&E) and periodic acid-Schiff (PAS) staining were performed to evaluate pathological lesions. RNA sequencing data of the kidneys and integrated publicly available single-cell and spatial transcriptome datasets were used to investigate the mechanism of endothelial dysfunction. There was a significant increase in body weight (p = 0.001), glucose levels (p=0.0008), and UACR (p=0.006) in db/db mice compared with db/m mice. H&E and PAS staining showed that vacuolar lesions and protein casts of tubules were the major histopathological changes observed in early-stage DKD mice. The apoptotic pathway in endothelial cells was notably activated in DKD, and Thbs1 was identified as the central gene involved in this apoptotic process. Deconvolution of the cell composition in the RNA sequencing data showed a decrease in the proportion of endothelial cells in the DKD mice. Further analysis of the activity and regulatory network of transcription factors showed that Creb1 was activated in both mouse and human early-stage DKD, suggesting that Creb1 activation may be involved in early kidney injury. The endothelial cell apoptotic pathway is activated in DKD, and the proportion of endothelial cells was reduced in the DKD mice, which is significantly associated with Thbs1. Keywords: Diabetic kidney disease, Endothelial dysfunction, RNA sequencing,Thbs1, Creb1.

• MeSH
• apoptóza MeSH
• diabetické nefropatie * patologie   metabolismus   patofyziologie   genetika   MeSH
• endoteliální buňky metabolismus   patologie   MeSH
• ledvinové kanálky patologie   metabolismus   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• progrese nemoci * MeSH
• protein vázající cAMP responzivní element metabolismus   genetika   MeSH
• thrombospondin 1 metabolismus   genetika   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Liver sinusoidal endothelial cells (LSECs) play a crucial role in regulating the hepatic function. Endoglin (ENG), a transmembrane glycoprotein, was shown to be related to the development of endothelial dysfunction. In this study, we hypothesized the relationship between changes in ENG expression and markers of liver sinusoidal endothelial dysfunction (LSED) during liver impairment. Male C57BL/6J mice aged 9-12 weeks were fed with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet (intrahepatic cholestasis) or choline-deficient l-amino acid defined high-fat diet (CDAA-HFD) (non-alcoholic steatohepatitis (NASH)). Significant increases in liver enzymes, fibrosis, and inflammation biomarkers were observed in both cholestasis and NASH. Decreased p-eNOS/eNOS and VE-cadherin protein expression and a significant increase in VCAM-1 and ICAM-1 expression were detected, indicating LSED in both mouse models of liver damage. A significant reduction of ENG in the DDC-fed mice, while a significant increase of ENG in the CDAA-HFD group was observed. Both DDC and CDAA-HFD-fed mice showed a significant increase in MMP-14 protein expression, which is related to significantly increased levels of soluble endoglin (sENG) in the plasma. In conclusion, we demonstrated that intrahepatic cholestasis and NASH result in an altered ENG expression, predominantly in LSECs, suggesting a critical role of ENG expression for the proper function of liver sinusoids. Both pathologies resulted in elevated sENG levels, cleaved by MMP-14 expressed predominantly from LSECs, indicating sENG as a liver injury biomarker.

• MeSH
• acetamidy * MeSH
• dieta s vysokým obsahem tuků škodlivé účinky   MeSH
• endoglin metabolismus   MeSH
• endoteliální buňky metabolismus   MeSH
• intrahepatální cholestáza * MeSH
• matrixová metaloproteinasa 14 MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• nealkoholová steatóza jater * patologie   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Fuchs endothelial corneal dystrophy (FECD) is an age-related cause of vision loss, and the most common repeat expansion-mediated disease in humans characterised to date. Up to 80% of European FECD cases have been attributed to expansion of a non-coding CTG repeat element (termed CTG18.1) located within the ubiquitously expressed transcription factor encoding gene, TCF4. The non-coding nature of the repeat and the transcriptomic complexity of TCF4 have made it extremely challenging to experimentally decipher the molecular mechanisms underlying this disease. Here we comprehensively describe CTG18.1 expansion-driven molecular components of disease within primary patient-derived corneal endothelial cells (CECs), generated from a large cohort of individuals with CTG18.1-expanded (Exp+) and CTG 18.1-independent (Exp-) FECD. We employ long-read, short-read, and spatial transcriptomic techniques to interrogate expansion-specific transcriptomic biomarkers. Interrogation of long-read sequencing and alternative splicing analysis of short-read transcriptomic data together reveals the global extent of altered splicing occurring within Exp+ FECD, and unique transcripts associated with CTG18.1-expansions. Similarly, differential gene expression analysis highlights the total transcriptomic consequences of Exp+ FECD within CECs. Furthermore, differential exon usage, pathway enrichment and spatial transcriptomics reveal TCF4 isoform ratio skewing solely in Exp+ FECD with potential downstream functional consequences. Lastly, exome data from 134 Exp- FECD cases identified rare (minor allele frequency <0.005) and potentially deleterious (CADD>15) TCF4 variants in 7/134 FECD Exp- cases, suggesting that TCF4 variants independent of CTG18.1 may increase FECD risk. In summary, our study supports the hypothesis that at least two distinct pathogenic mechanisms, RNA toxicity and TCF4 isoform-specific dysregulation, both underpin the pathophysiology of FECD. We anticipate these data will inform and guide the development of translational interventions for this common triplet-repeat mediated disease.

• MeSH
• alternativní sestřih genetika   MeSH
• endoteliální buňky metabolismus   MeSH
• expanze trinukleotidových repetic * genetika   MeSH
• Fuchsova endoteliální dystrofie * genetika   MeSH
• lidé MeSH
• rohovkový endotel metabolismus   patologie   MeSH
• transkripční faktor 4 * genetika   metabolismus   MeSH
• transkriptom genetika   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví 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

Oxidative stress and autophagy are potential mechanisms associated with cerebral ischemia/reperfusion injury (IRI) and is usually linked to inflammatory responses and apoptosis. Curcumin has recently been demonstrated to exhibit anti-inflammatory, anti-oxidant, anti-apoptotic and autophagy regulation properties. However, mechanism of curcumin on IRI-induced oxidative stress and autophagy remains not well understood. We evaluated the protective effects and potential mechanisms of curcumin on cerebral microvascular endothelial cells (bEnd.3) and neuronal cells (HT22) against oxygen glucose deprivation/reoxygenation (OGD/R) in vitro models that mimic in vivo cerebral IRI. The cell counting kit-8 (CCK-8) and lactate dehydrogenase (LDH) activity assays revealed that curcumin attenuated the OGD/R-induced injury in a dose-specific manner. OGD/R induced elevated levels of inflammatory cytokines TNF-alpha, IL-6 as well as IL-1beta, and these effects were notably reduced by curcumin. OGD/R-mediated apoptosis was suppressed by curcumin via upregulating B-cell lymphoma-2 (Bcl-2) and downregulating Bcl-associated X (Bax), cleaved-caspase3 and TUNEL apoptosis marker. Additionally, curcumin increased superoxide dismutase (SOD) and glutathione (GSH), but suppressed malondialdehyde (MDA) and reactive oxygen species (ROS) content. Curcumin inhibited the levels of autophagic biomarkers such as LC3 II/LC3 I and Beclin1. Particularly, curcumin induced p62 accumulation and its interactions with keap1 and promoted NF-E2-related factor 2 (Nrf2) translocation to nucleus, accompanied by increased NADPH quinone dehydrogenase (Nqo1) and heme oxygenase 1 (HO-1). Treatment of curcumin increased phosphorylation-phosphatidylinositol 3 kinase (p-PI3K) and p-protein kinase B (p-AKT). The autophagy inhibitor 3-methyladenine (3-MA) activated the keap-1/Nrf2 and PI3K/AKT pathways. This study highlights the neuroprotective effects of curcumin on cerebral IRI.

• MeSH
• antioxidancia farmakologie   metabolismus   MeSH
• autofagie fyziologie   MeSH
• endoteliální buňky metabolismus   MeSH
• faktor 2 související s NF-E2 metabolismus   MeSH
• fosfatidylinositol-3-kinasy metabolismus   MeSH
• KEAP-1 metabolismus   MeSH
• kurkumin * farmakologie   MeSH
• kyslík metabolismus   MeSH
• neuroprotektivní látky * farmakologie   MeSH
• oxidační stres MeSH
• protoonkogenní proteiny c-akt metabolismus   MeSH
• signální transdukce MeSH
• Publikační typ
• časopisecké články MeSH

Aim: Human induced pluripotent stem cells (iPSCs) are inefficiently derived from somatic cells by overexpression of defined transcription factors. Overexpression of H2A histone variant macroH2A1.1, but not macroH2A1.2, leads to increased iPSC reprogramming by unclear mechanisms. Materials & methods: Cleavage under targets and tagmentation (CUT&Tag) allows robust epigenomic profiling of a low cell number. We performed an integrative CUT&Tag-RNA-Seq analysis of macroH2A1-dependent orchestration of iPSCs reprogramming using human endothelial cells. Results: We demonstrate wider genome occupancy, predicted transcription factors binding, and gene expression regulated by macroH2A1.1 during reprogramming, compared to macroH2A1.2. MacroH2A1.1, previously associated with neurodegenerative pathologies, specifically activated ectoderm/neural processes. Conclusion: CUT&Tag and RNA-Seq data integration is a powerful tool to investigate the epigenetic mechanisms occurring during cell reprogramming.

• MeSH
• endoteliální buňky metabolismus   MeSH
• histony * metabolismus   MeSH
• indukované pluripotentní kmenové buňky * metabolismus   MeSH
• lidé MeSH
• přeprogramování buněk genetika   MeSH
• sekvenování transkriptomu MeSH
• transkripční faktory genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

OBJECTIVE: An increase in the number of neutrophils (NEUs) has long been associated with infections in the knee joints; however, their impact on knee osteoarthritis (KOA) pathophysiology remains largely unexplored. DESIGN: This study compared the phenotypic and functional characteristics of synovial fluid (SF)-derived NEUs in KOA and knee infection (INF). RESULTS: KOA NEUs were characterised by a lower expression of CD11b, CD54, and CD64 and higher expression of CD62L, TLR2, and TLR4 compared with INF NEUs. Except for CCL2, lower levels of inflammatory mediators and proteases were detected in KOA SF than in INF SF. Functionally, KOA NEUs displayed increased reactive oxygen species production and phagocytic activity compared with INF NEUs. Moreover, KOA and INF NEUs differed in cell sizes, histological characteristics of the surrounding synovial tissues, and their effects on the endothelial cells assessed by human umbilical vein endothelial cells. When KOA patients were subdivided based on the SF NEU abundance, patients with high NEUs (10%-60%) were characterised by i) elevated SF protein levels of TNF-α, IL-1RA, MMP-9, sTREM-1, VILIP-1 and ii) lower CD54, CD64, TLR2 and TLR4 expression compared to patients with low NEUs (<10%). Analysis of paired SF samples suggests that low or high NEU percentages, respectively, persist throughout the course of disease. CONCLUSIONS: Our findings suggest that NEU may play a significant role in KOA pathophysiology. Further studies should explore the mechanisms that contribute to the increased number of NEUs in SF and the clinical consequences of neutrophilic phenotype in KOA.

• MeSH
• artróza kolenních kloubů * MeSH
• endoteliální buňky metabolismus   MeSH
• fenotyp MeSH
• kolenní kloub patologie   MeSH
• lidé MeSH
• neutrofily MeSH
• synoviální tekutina * metabolismus   MeSH
• toll-like receptor 2 metabolismus   MeSH
• toll-like receptor 4 metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKIs) show potent efficacy in several ALK-driven tumors, but the development of resistance limits their long-term clinical impact. Although resistance mechanisms have been studied extensively in ALK-driven non-small cell lung cancer, they are poorly understood in ALK-driven anaplastic large cell lymphoma (ALCL). Here, we identify a survival pathway supported by the tumor microenvironment that activates phosphatidylinositol 3-kinase γ (PI3K-γ) signaling through the C-C motif chemokine receptor 7 (CCR7). We found increased PI3K signaling in patients and ALCL cell lines resistant to ALK TKIs. PI3Kγ expression was predictive of a lack of response to ALK TKI in patients with ALCL. Expression of CCR7, PI3Kγ, and PI3Kδ were up-regulated during ALK or STAT3 inhibition or degradation and a constitutively active PI3Kγ isoform cooperated with oncogenic ALK to accelerate lymphomagenesis in mice. In a three-dimensional microfluidic chip, endothelial cells that produce the CCR7 ligands CCL19/CCL21 protected ALCL cells from apoptosis induced by crizotinib. The PI3Kγ/δ inhibitor duvelisib potentiated crizotinib activity against ALCL lines and patient-derived xenografts. Furthermore, genetic deletion of CCR7 blocked the central nervous system dissemination and perivascular growth of ALCL in mice treated with crizotinib. Thus, blockade of PI3Kγ or CCR7 signaling together with ALK TKI treatment reduces primary resistance and the survival of persister lymphoma cells in ALCL.

• MeSH
• anaplastická lymfomová kináza MeSH
• anaplastický velkobuněčný lymfom * farmakoterapie   genetika   patologie   MeSH
• endoteliální buňky metabolismus   MeSH
• fosfatidylinositol-3-kinasy MeSH
• inhibitory proteinkinas farmakologie   terapeutické užití   MeSH
• inhibitory tyrosinkinasy MeSH
• krizotinib farmakologie   terapeutické užití   MeSH
• lidé MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nádorové mikroprostředí MeSH
• nádory plic * farmakoterapie   MeSH
• nemalobuněčný karcinom plic * farmakoterapie   MeSH
• receptory CCR7 genetika   MeSH
• tyrosinkinasové receptory metabolismus   MeSH
• tyrosinkinasy 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