Fluorescein, eosin Y, and rose bengal are dyes used in clinical medicine and considered (photo-)chemically stable. Upon extensive irradiation with visible light in aqueous solutions, we found that these compounds release carbon monoxide (CO) - a bioactive gasotransmitter - in 40-100% yields along with the production of low-mass secondary photoproducts, such as phthalic and formic acids, in a multistep degradation process. Such photochemistry should be considered in applications of these dyes, and they could also be utilized as visible-light activatable CO-releasing molecules (photoCORMs) with biological implications.
Fluorescein is a fluorescent dye used as a diagnostic tool in various fields of medicine. Although fluorescein itself possesses low toxicity, after photoactivation, it releases potentially toxic molecules, such as singlet oxygen (1O2) and, as we demonstrate in this work, also carbon monoxide (CO). As both of these molecules can affect physiological processes, the main aim of this study was to explore the potential biological impacts of fluorescein photochemistry. In our in vitro study in a human hepatoblastoma HepG2 cell line, we explored the possible effects on cell viability, cellular energy metabolism, and the cell cycle. We observed markedly lowered cell viability (≈30%, 75-2400 μM) upon irradiation of intracellular fluorescein and proved that this decrease in viability was dependent on the cellular oxygen concentration. We also detected a significantly decreased concentration of Krebs cycle metabolites (lactate and citrate < 30%; 2-hydroxyglutarate and 2-oxoglutarate < 10%) as well as cell cycle arrest (decrease in the G2 phase of 18%). These observations suggest that this photochemical reaction could have important biological consequences and may account for some adverse reactions observed in fluorescein-treated patients. Additionally, the biological activities of both 1O2 and CO might have considerable therapeutic potential, particularly in the treatment of cancer.
- MeSH
- angiografie MeSH
- antitumorózní látky chemie farmakologie MeSH
- buňky Hep G2 MeSH
- citrátový cyklus účinky léků účinky záření MeSH
- fluorescein chemie farmakologie MeSH
- fotochemické procesy MeSH
- kontrolní body buněčného cyklu účinky léků účinky záření MeSH
- lidé MeSH
- oxid uhelnatý analýza MeSH
- plynová chromatografie s hmotnostně spektrometrickou detekcí MeSH
- singletový kyslík analýza MeSH
- světlo MeSH
- viabilita buněk účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
For severe unconjugated hyperbilirubinemia the gold standard treatment is phototherapy with blue-green light, producing more polar photo-oxidation products, believed to be non-toxic. The aim of the present study was to compare the effects of bilirubin (BR) and lumirubin (LR), the major BR photo-oxidation product, on metabolic and oxidative stress markers. The biological activities of these pigments were investigated on several human and murine cell lines, with the focus on mitochondrial respiration, substrate metabolism, reactive oxygen species production, and the overall effects on cell viability. Compared to BR, LR was found to be much less toxic, while still maintaining a similar antioxidant capacity in the serum as well as suppressing activity leading to mitochondrial superoxide production. Nevertheless, due to its lower lipophilicity, LR was less efficient in preventing lipoperoxidation. The cytotoxicity of BR was affected by the cellular glycolytic reserve, most compromised in human hepatoblastoma HepG2 cells. The observed effects were correlated with changes in the production of tricarboxylic acid cycle metabolites. Both BR and LR modulated expression of PPARα downstream effectors involved in lipid and glucose metabolism. Proinflammatory effects of BR, evidenced by increased expression of TNFα upon exposure to bacterial lipopolysaccharide, were observed in murine macrophage-like RAW 264.7 cells. Collectively, these data point to the biological effects of BR and its photo-oxidation products, which might have clinical relevance in phototherapy-treated hyperbilirubinemic neonates and adult patients.
- Publikační typ
- časopisecké články MeSH
Granulocyte colony-stimulating factor (G-CSF) is used in clinical practice to mobilize cells from the bone marrow to the blood; however, it is not always effective. We show that cobalt protoporphyrin IX (CoPP) increases plasma concentrations of G-CSF, IL-6, and MCP-1 in mice, triggering the mobilization of granulocytes and hematopoietic stem and progenitor cells (HSPC). Compared with recombinant G-CSF, CoPP mobilizes higher number of HSPC and mature granulocytes. In contrast to G-CSF, CoPP does not increase the number of circulating T cells. Transplantation of CoPP-mobilized peripheral blood mononuclear cells (PBMC) results in higher chimerism and faster hematopoietic reconstitution than transplantation of PBMC mobilized by G-CSF. Although CoPP is used to activate Nrf2/HO-1 axis, the observed effects are Nrf2/HO-1 independent. Concluding, CoPP increases expression of mobilization-related cytokines and has superior mobilizing efficiency compared with recombinant G-CSF. This observation could lead to the development of new strategies for the treatment of neutropenia and HSPC transplantation.
- MeSH
- faktor stimulující kolonie granulocytů metabolismus MeSH
- granulocyty účinky léků MeSH
- hematopoetické kmenové buňky účinky léků MeSH
- hemoxygenasa-1 nedostatek genetika MeSH
- mobilizace hematopoetických kmenových buněk MeSH
- myši inbrední C57BL MeSH
- myši knockoutované MeSH
- myši MeSH
- protoporfyriny farmakologie MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Oxidative stress and inflammation are predominant features of several chronic diseases. The nuclear factor erythroid 2-related factor 2 (Nrf2) is a major arbiter in counteracting these insults via up-regulation of several defensive proteins, including heme oxygenase-1 (HO-1). HO-1-derived carbon monoxide (CO) exhibits anti-inflammatory actions and can be delivered to tissues by CO-releasing agents. In this study we assessed the pharmacological and anti-inflammatory properties of HYCO-3, a dual activity compound obtained by conjugating analogues of the CO-releasing molecule CORM-401 and dimethyl fumarate (DMF), an immunomodulatory drug known to activate Nrf2. HYCO-3 induced Nrf2-dependent genes and delivered CO to cells in vitro and tissues in vivo, confirming that the two expected pharmacological properties of this agent are achieved. In mice challenged with lipopolysaccharide, orally administered HYCO-3 reduced the mRNA levels of pro-inflammatory markers (TNF-α, IL-1β and IL-6) while increasing the expression of the anti-inflammatory genes ARG1 and IL-10 in brain, liver, lung and heart. In contrast, DMF or CORM-401 alone or their combination decreased the expression of pro-inflammatory genes but had limited influence on anti-inflammatory markers. Furthermore, HYCO-3 diminished TNF-α and IL-1β in brain and liver but not in lung and heart of Nrf2-/- mice, indicating that the CO-releasing part of this hybrid contributes to reduction of pro-inflammation and that this effect is organ-specific. These data demonstrate that the dual activity of HYCO-3 results in enhanced efficacy compared to the parent compounds indicating the potential exploitation of hybrid compounds in the development of effective anti-inflammatory therapies.
- MeSH
- antiflogistika farmakologie MeSH
- antioxidancia metabolismus MeSH
- cytokiny genetika metabolismus MeSH
- exprese genu MeSH
- faktor 2 související s NF-E2 genetika metabolismus MeSH
- hemoxygenasa-1 genetika metabolismus MeSH
- kultivované buňky MeSH
- lipopolysacharidy škodlivé účinky MeSH
- makrofágy účinky léků imunologie metabolismus MeSH
- mediátory zánětu metabolismus MeSH
- mikroglie účinky léků metabolismus MeSH
- modely nemocí na zvířatech MeSH
- myši knockoutované MeSH
- myši MeSH
- oxid uhelnatý metabolismus MeSH
- oxidační stres účinky léků MeSH
- zánět farmakoterapie etiologie metabolismus patologie MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Decreased inflammatory status has been reported in subjects with mild unconjugated hyperbilirubinemia. However, mechanisms of the anti-inflammatory actions of bilirubin (BR) are not fully understood. The aim of this study is to assess the role of BR in systemic inflammation using hyperbilirubinemic Gunn rats as well as their normobilirubinemic littermates and further in primary hepatocytes. The rats were treated with lipopolysaccharide (LPS, 6 mg/kg intraperitoneally) for 12 h, their blood and liver were collected for analyses of inflammatory and hepatic injury markers. Primary hepatocytes were treated with BR and TNF-α. LPS-treated Gunn rats had a significantly decreased inflammatory response, as evidenced by the anti-inflammatory profile of white blood cell subsets, and lower hepatic and systemic expressions of IL-6, TNF-α, IL-1β, and IL-10. Hepatic mRNA expression of LPS-binding protein was upregulated in Gunn rats before and after LPS treatment. In addition, liver injury markers were lower in Gunn rats as compared to in LPS-treated controls. The exposure of primary hepatocytes to TNF-α with BR led to a milder decrease in phosphorylation of the NF-κB p65 subunit compared to in cells without BR. In conclusion, hyperbilirubinemia in Gunn rats is associated with an attenuated systemic inflammatory response and decreased liver damage upon exposure to LPS.
- MeSH
- apoptóza účinky léků MeSH
- bilirubin farmakologie MeSH
- biologické markery krev MeSH
- cytokiny krev genetika metabolismus MeSH
- cytoprotekce účinky léků MeSH
- fosforylace účinky léků MeSH
- hepatocyty metabolismus MeSH
- hyperbilirubinemie krev komplikace MeSH
- játra metabolismus MeSH
- kultivované buňky MeSH
- leukocyty metabolismus MeSH
- lipopolysacharidy MeSH
- messenger RNA genetika metabolismus MeSH
- NF-kappa B metabolismus MeSH
- potkani Gunn MeSH
- signální transdukce MeSH
- zánět komplikace MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Bilirubin is considered to be one of the most potent endogenous antioxidants in humans. Its serum concentrations are predominantly affected by the activity of hepatic bilirubin UDP-glucuronosyl transferase (UGT1A1). Our objective was to analyze the potential bilirubin-modulating effects of natural polyphenols from milk thistle (Silybum marianum), a hepatoprotective herb. Human hepatoblastoma HepG2 cells were exposed to major polyphenolic compounds isolated from milk thistle. Based on in vitro studies, 2,3-dehydrosilybins A and B were selected as the most efficient compounds and applied either intraperitoneally or orally for seven days to C57BL/6 mice. After, UGT1A1 mRNA expression, serum, intrahepatic bilirubin concentrations, and lipoperoxidation in the liver tissue were analyzed. All natural polyphenols used increased intracellular concentration of bilirubin in HepG2 cells to a similar extent as atazanavir, a known bilirubinemia-enhancing agent. Intraperitoneal application of 2,3-dehydrosilybins A and B (the most efficient flavonoids from in vitro studies) to mice (50 mg/kg) led to a significant downregulation of UGT1A1 mRNA expression (46 ± 3% of controls, p < 0.005) in the liver and also to a significant increase of the intracellular bilirubin concentration (0.98 ± 0.03vs.1.21 ± 0.02 nmol/mg, p < 0.05). Simultaneously, a significant decrease of lipoperoxidation (61 ± 2% of controls, p < 0.005) was detected in the liver tissue of treated animals, and similar results were also observed after oral treatment. Importantly, both application routes also led to a significant elevation of serum bilirubin concentrations (125 ± 3% and 160 ± 22% of the controls after intraperitoneal and oral administration, respectively, p < 0.005 in both cases). In conclusion, polyphenolic compounds contained in silymarin, in particular 2,3-dehydrosilybins A and B, affect hepatic and serum bilirubin concentrations, as well as lipoperoxidation in the liver. This phenomenon might contribute to the hepatoprotective effects of silymarin.
- MeSH
- bilirubin metabolismus MeSH
- buňky Hep G2 MeSH
- flavonoidy chemie izolace a purifikace farmakologie MeSH
- glukuronosyltransferasa genetika metabolismus MeSH
- hemoxygenasa-1 genetika metabolismus MeSH
- intracelulární prostor metabolismus MeSH
- játra účinky léků metabolismus MeSH
- lidé MeSH
- messenger RNA genetika metabolismus MeSH
- myši inbrední C57BL MeSH
- peroxidace lipidů účinky léků MeSH
- regulace genové exprese enzymů účinky léků MeSH
- silibinin aplikace a dávkování farmakologie MeSH
- silymarin izolace a purifikace farmakologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: The action spectrum for bilirubin photodegradation has been intensively studied. However, questions still remain regarding which light wavelength most efficiently photodegrades bilirubin. In this study, we determined the in vitro effects of different irradiation wavelength ranges on bilirubin photodegradation. METHODS: In our in vitro method, normalized absolute irradiance levels of 4.2 × 1015 photons/cm2/s from light-emitting diodes (ranging from 390-530 nm) and 10-nm band-pass filters were used to irradiate bilirubin solutions (25 mg/dL in 4% human serum albumin). Bilirubin and its major photoisomer concentrations were determined; the half-life time of bilirubin (t1/2) was calculated for each wavelength range, and the spectral characteristics for bilirubin photodegradation products were obtained for key wavelengths. RESULTS: The in vitro photodegradation of bilirubin at 37 °C decreased linearly as the wavelength was increased from 390 to 500 nm with t1/2 decreasing from 63 to 17 min, respectively. At 460 ± 10 nm, a significantly lower rate of photodegradation and thus higher t1/2 (31 min) than that at 500 nm (17 min) was demonstrated. CONCLUSION: In our system, the optimum bilirubin photodegradation and lumirubin production rates occurred between 490 and 500 nm. Spectra shapes were remarkably similar, suggesting that lumirubin production was the major process of bilirubin photodegradation.
- MeSH
- bilirubin analogy a deriváty krev chemie účinky záření MeSH
- fotolýza účinky záření MeSH
- fototerapie metody MeSH
- isomerie MeSH
- lidé MeSH
- lidský sérový albumin chemie účinky záření MeSH
- novorozenec MeSH
- novorozenecká hyperbilirubinemie krev terapie MeSH
- spektrofotometrie MeSH
- světlo * MeSH
- techniky in vitro MeSH
- Check Tag
- lidé MeSH
- novorozenec MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- antioxidancia * izolace a purifikace metabolismus normy MeSH
- bilirubin * izolace a purifikace krev metabolismus MeSH
- cholestáza * etiologie klasifikace metabolismus MeSH
- komorbidita MeSH
- kyselina cholová chemie metabolismus MeSH
- lidé MeSH
- metaanalýza jako téma MeSH
- receptory cytoplazmatické a nukleární metabolismus MeSH
- statistika jako téma MeSH
- žluč metabolismus MeSH
- žlučové kyseliny a soli chemie metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
- přehledy MeSH
Obesity is characterized by accumulation of adipose tissue and is one the most important risk factors in the development of insulin resistance. Carbon monoxide-releasing (CO-releasing) molecules (CO-RMs) have been reported to improve the metabolic profile of obese mice, but the underlying mechanism remains poorly defined. Here, we show that oral administration of CORM-401 to obese mice fed a high-fat diet (HFD) resulted in a significant reduction in body weight gain, accompanied by a marked improvement in glucose homeostasis. We further unmasked an action we believe to be novel, by which CO accumulates in visceral adipose tissue and uncouples mitochondrial respiration in adipocytes, ultimately leading to a concomitant switch toward glycolysis. This was accompanied by enhanced systemic and adipose tissue insulin sensitivity, as indicated by a lower blood glucose and increased Akt phosphorylation. Our findings indicate that the transient uncoupling activity of CO elicited by repetitive administration of CORM-401 is associated with lower weight gain and increased insulin sensitivity during HFD. Thus, prototypic compounds that release CO could be investigated for developing promising insulin-sensitizing agents.
- MeSH
- adenosintrifosfát metabolismus MeSH
- buňky 3T3-L1 MeSH
- dieta s vysokým obsahem tuků MeSH
- hmotnostní přírůstek účinky léků MeSH
- inzulinová rezistence * MeSH
- myši inbrední C57BL MeSH
- myši obézní MeSH
- myši MeSH
- obezita metabolismus MeSH
- organokovové sloučeniny farmakologie MeSH
- oxid uhelnatý metabolismus MeSH
- tukové buňky účinky léků metabolismus MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
