Nitro-oleic acid (NO2-OA), pluripotent cell-signaling mediator, was recently described as a modulator of the signal transducer and activator of transcription 3 (STAT3) activity. In our study, we discovered new aspects of NO2-OA involvement in the regulation of stem cell pluripotency and differentiation. Murine embryonic stem cells (mESC) or mESC-derived embryoid bodies (EBs) were exposed to NO2-OA or oleic acid (OA) for selected time periods. Our results showed that NO2-OA but not OA caused the loss of pluripotency of mESC cultivated in leukemia inhibitory factor (LIF) rich medium via the decrease of pluripotency markers (NANOG, sex-determining region Y-box 1 transcription factor (SOX2), and octamer-binding transcription factor 4 (OCT4)). The effects of NO2-OA on mESC correlated with reduced phosphorylation of STAT3. Subsequent differentiation led to an increase of the ectodermal marker orthodenticle homolog 2 (Otx2). Similarly, treatment of mESC-derived EBs by NO2-OA resulted in the up-regulation of both neural markers Nestin and β-Tubulin class III (Tubb3). Interestingly, the expression of cardiac-specific genes and beating of EBs were significantly decreased. In conclusion, NO2-OA is able to modulate pluripotency of mESC via the regulation of STAT3 phosphorylation. Further, it attenuates cardiac differentiation on the one hand, and on the other hand, it directs mESC into neural fate.

• Klíčová slova
• STAT3, cardiomyogenesis, mouse embryonic stem cells, neurogenesis, nitro-oleic acid, pluripotency,
• MeSH
• biologické markery metabolismus   MeSH
• buněčná diferenciace * účinky léků   MeSH
• dusíkaté sloučeniny farmakologie   MeSH
• embryoidní tělíska účinky léků   metabolismus   MeSH
• kardiomyocyty účinky léků   metabolismus   MeSH
• kyseliny olejové farmakologie   MeSH
• myší embryonální kmenové buňky cytologie   účinky léků   metabolismus   MeSH
• myši MeSH
• neurony cytologie   účinky léků   metabolismus   MeSH
• organogeneze účinky léků   MeSH
• pluripotentní kmenové buňky účinky léků   metabolismus   MeSH
• signální transdukce účinky léků   MeSH
• transkripční faktor STAT3 metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• biologické markery MeSH
• CXA-10 MeSH Prohlížeč
• dusíkaté sloučeniny MeSH
• kyseliny olejové MeSH
• transkripční faktor STAT3 MeSH

Many diseases accompanied by chronic inflammation are connected with dysregulated activation of macrophage subpopulations. Recently, we reported that nitro-fatty acids (NO2-FAs), products of metabolic and inflammatory reactions of nitric oxide and nitrite, modulate macrophage and other immune cell functions. Bone marrow cell suspensions were isolated from mice and supplemented with macrophage colony-stimulating factor (M-CSF) or granulocyte-macrophage colony-stimulating factor (GM-CSF) in combination with NO2-OA for different times. RAW 264.7 macrophages were used for short-term (1-5min) experiments. We discovered that NO2-OA reduces cell numbers, cell colony formation, and proliferation of macrophages differentiated with colony-stimulating factors (CSFs), all in the absence of toxicity. In a case of GM-CSF-induced bone marrow-derived macrophages (BMMs), NO2-OA acts via downregulation of signal transducer and activator of transcription 5 and extracellular signal-regulated kinase (ERK) activation. In the case of M-CSF-induced BMMs, NO2-OA decreases activation of M-CSFR and activation of related PI3K and ERK. Additionally, NO2-OA also attenuates activation of BMMs. In aggregate, we demonstrate that NO2-OA regulates the process of macrophage differentiation and that NO2-FAs represent a promising therapeutic tool in the treatment of inflammatory pathologies linked with increased accumulation of macrophages in inflamed tissues.

• Klíčová slova
• Differentiation, Growth factors, Inflammation, Macrophages, Nitro-fatty acids, Nitro-oleic acid, Signaling pathways,
• MeSH
• buněčná diferenciace účinky léků   MeSH
• buňky kostní dřeně účinky léků   MeSH
• extracelulárním signálem regulované MAP kinasy genetika   MeSH
• faktory stimulující kolonie genetika   MeSH
• fosfatidylinositol-3-kinasy genetika   MeSH
• kyselina olejová aplikace a dávkování   chemie   MeSH
• makrofágy účinky léků   MeSH
• MAP kinasový signální systém účinky léků   MeSH
• myši MeSH
• oxid dusnatý aplikace a dávkování   chemie   MeSH
• RAW 264.7 buňky MeSH
• regulace genové exprese u nádorů účinky léků   MeSH
• transkripční faktor STAT5 genetika   MeSH
• zánět farmakoterapie   genetika   patologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• extracelulárním signálem regulované MAP kinasy MeSH
• faktory stimulující kolonie MeSH
• fosfatidylinositol-3-kinasy MeSH
• kyselina olejová MeSH
• oxid dusnatý MeSH
• transkripční faktor STAT5 MeSH

RATIONALE: Pulmonary hypertension (PH) represents a serious health complication accompanied with hypoxic conditions, elevated levels of asymmetric dimethylarginine (ADMA), and overall dysfunction of pulmonary vascular endothelium. Since the prevention strategies for treatment of PH remain largely unknown, our study aimed to explore the effect of nitro-oleic acid (OA-NO2), an exemplary nitro-fatty acid (NO2-FA), in human pulmonary artery endothelial cells (HPAEC) under the influence of hypoxia or ADMA. METHODS: HPAEC were treated with OA-NO2 in the absence or presence of hypoxia and ADMA. The production of nitric oxide (NO) and interleukin-6 (IL-6) was monitored using the Griess method and ELISA, respectively. The expression or activation of different proteins (signal transducer and activator of transcription 3, STAT3; hypoxia inducible factor 1α, HIF-1α; endothelial nitric oxide synthase, eNOS; intercellular adhesion molecule-1, ICAM-1) was assessed by the Western blot technique. RESULTS: We discovered that OA-NO2 prevents development of endothelial dysfunction induced by either hypoxia or ADMA. OA-NO2 preserves normal cellular functions in HPAEC by increasing NO production and eNOS expression. Additionally, OA-NO2 inhibits IL-6 production as well as ICAM-1 expression, elevated by hypoxia and ADMA. Importantly, the effect of OA-NO2 is accompanied by prevention of STAT3 activation and HIF-1α stabilization. CONCLUSION: In summary, OA-NO2 eliminates the manifestation of hypoxia- and ADMA-mediated endothelial dysfunction in HPAEC via the STAT3/HIF-1α cascade. Importantly, our study is bringing a new perspective on molecular mechanisms of NO2-FAs action in pulmonary endothelial dysfunction, which represents a causal link in progression of PH. Graphical Abstract ᅟ.

• Klíčová slova
• Asymmetric dimethylarginine, Human pulmonary artery endothelial cell, Hypoxia, Nitro-oleic acid, Pulmonary hypertension,
• MeSH
• arginin analogy a deriváty   farmakologie   MeSH
• arteria pulmonalis cytologie   MeSH
• buněčná adheze účinky léků   MeSH
• endoteliální buňky účinky léků   metabolismus   fyziologie   MeSH
• faktor 1 indukovatelný hypoxií - podjednotka alfa metabolismus   MeSH
• hypoxie buňky účinky léků   MeSH
• interleukin-6 metabolismus   MeSH
• kultivované buňky MeSH
• kyseliny olejové farmakologie   MeSH
• lidé MeSH
• mezibuněčná adhezivní molekula-1 metabolismus   MeSH
• oxid dusnatý metabolismus   MeSH
• pohyb buněk účinky léků   MeSH
• synthasa oxidu dusnatého, typ III metabolismus   MeSH
• transkripční faktor STAT3 antagonisté a inhibitory   metabolismus   MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• arginin MeSH
• faktor 1 indukovatelný hypoxií - podjednotka alfa MeSH
• HIF1A protein, human MeSH Prohlížeč
• IL6 protein, human MeSH Prohlížeč
• interleukin-6 MeSH
• kyseliny olejové MeSH
• mezibuněčná adhezivní molekula-1 MeSH
• N,N-dimethylarginine MeSH Prohlížeč
• NOS3 protein, human MeSH Prohlížeč
• oxid dusnatý MeSH
• STAT3 protein, human MeSH Prohlížeč
• synthasa oxidu dusnatého, typ III MeSH
• transkripční faktor STAT3 MeSH