RESUMEN
The NLRP3 inflammasome plays a key role in responding to pathogens, and endogenous damage and mitochondria are intensively involved in inflammasome activation. The NLRP3 inflammasome forms multiprotein complexes and its sequential assembly is important for its activation. Here, we show that NLRP3 is ubiquitinated by the mitochondria-associated E3 ligase, MARCH5. Myeloid cell-specific March5 conditional knockout (March5 cKO) mice failed to secrete IL-1ß and IL-18 and exhibited an attenuated mortality rate upon LPS or Pseudomonas aeruginosa challenge. Macrophages derived from March5 cKO mice also did not produce IL-1ß and IL-18 after microbial infection. Mechanistically, MARCH5 interacts with the NACHT domain of NLRP3 and promotes K27-linked polyubiquitination on K324 and K430 residues of NLRP3. Ubiquitination-defective NLRP3 mutants on K324 and K430 residues are not able to bind to NEK7, nor form NLRP3 oligomers leading to abortive ASC speck formation and diminished IL-1ß production. Thus, MARCH5-dependent NLRP3 ubiquitination on the mitochondria is required for NLRP3-NEK7 complex formation and NLRP3 oligomerization. We propose that the E3 ligase MARCH5 is a regulator of NLRP3 inflammasome activation on the mitochondria.
Asunto(s)
Inflamasomas , Proteína con Dominio Pirina 3 de la Familia NLR , Animales , Ratones , Inflamasomas/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Interleucina-18/metabolismo , Ubiquitinación , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Caspasa 1/metabolismoRESUMEN
Cells are constantly challenged by genotoxic stresses that can lead to genome instability. The integrity of the nuclear genome is preserved by the DNA damage response (DDR) and repair. Additionally, these stresses can induce mitochondria to transiently hyperfuse; however, it remains unclear whether canonical DDR is linked to these mitochondrial morphological changes. Here, we report that the abolition of mitochondrial fusion causes a substantial defect in the ATM-mediated DDR signaling. This deficiency is overcome by the restoration of mitochondria fusion. In cells with fragmented mitochondria, genotoxic stress-induced activation of JNK and its translocation to DNA lesion are lost. Importantly, the mitochondrial fusion machinery of MFN1/MFN2 associates with Sab (SH3BP5) and JNK, and these interactions are indispensable for the Sab-mediated activation of JNK and the ATM-mediated DDR signaling. Accordingly, the formation of BRCA1 and 53BP1 foci, as well as homology and end-joining repair are impaired in cells with fragmented mitochondria. Together, these data show that mitochondrial fusion-dependent JNK signaling is essential for the DDR, providing vital insight into the integration of nuclear and cytoplasmic stress signals.
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Daño del ADN , Reparación del ADN , Humanos , Reparación del ADN/genética , Inestabilidad Genómica , Mitocondrias/genética , Transducción de Señal/genéticaRESUMEN
Mesenchymal stem cell (MSC)-derived exosomes have emerged as an attractive cell-free tool in tissue engineering and regenerative medicine. The current study aimed to examine the anti-inflammatory, pro-angiogenic, and wound-repair effects of both exosomes and selenium-stimulated exosomes, and check whether the latter had superior wound healing capacity over others. The cellular and molecular network of exosomes, as a paracrine signal, was extensively studied by performing miRNA arrays to explore the key mediators of exosomes in wound healing. Selenium is known to play a critical role in enhancing the proliferation, multi-potency, and anti-inflammatory effects of MSCs. Selenium-stimulated exosomes showed significant effects in inhibiting inflammation and improving pro-angiogenesis in human umbilical vein endothelial cells. Cell growth and the migration of human dermal fibroblasts and wound regeneration were more enhanced in the selenium-stimulated exosome group than in the selenium and exosome groups, thereby further promoting the wound healing in vivo. Taken together, selenium was found to augment the therapeutic effects of adipose MSC-derived exosomes in tissue regeneration. We concluded that selenium may be considered a vital agent for wound healing in stem cell-based cell-free therapies.
Asunto(s)
Exosomas , MicroARNs , Selenio , Movimiento Celular , Células Endoteliales de la Vena Umbilical Humana , Humanos , Inflamación , MicroARNs/farmacología , Selenio/farmacología , Cicatrización de HeridasRESUMEN
The endocytosis-mediating performances of two types of peptide ligands, cell receptor binding peptide (CRBP) and cell membrane penetrating peptide (CMPP), were analyzed and compared using a common carrier of peptide ligands-human ferritin heavy chain (hFTH) nanoparticle. Twenty-four copies of a CMPP(human immunodeficiency virus-derived TAT peptide) and/or a CRBP (peptide ligand with strong and specific affinity for either human integrin(αv ß3 ) or epidermal growth factor receptor I (EGFR) that is overexpressed on various cancer cells) were genetically presented on the surface of each hFTH nanopariticle. The quantitative level of endocytosis and intracellular localization of fluorescence dye-labeled CRBP- and CMPP-presenting nanoparticles were estimated in the in vitro cultures of integrin- and EGFR-overexpressing cancer and human dermal fibroblast cells(control). From the cancer cell cultures treated with the CMPP- and CRBP-presenting nanoparticles, it was notable that CRBPs resulted in quantitatively higher level of endocytosis than CMPP (TAT) and successfully transported the nanoparticles to the cytosol of cancer cells depending on concentration and treatment period of time, whereas TAT-mediated endocytosis localized most of the nanoparticles within endosomal vesicles under the same conditions. These novel findings provide highly useful informations to many researchers both in academia and in industry who are interested in developing anticancer drug delivery systems/carriers.
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Membrana Celular/metabolismo , Endocitosis , Nanopartículas/metabolismo , Péptidos/metabolismo , Receptores de Superficie Celular/metabolismo , Apoferritinas/metabolismo , Células Cultivadas , Receptores ErbB/metabolismo , Humanos , Integrina alfaVbeta3/metabolismo , Nanopartículas/química , Unión Proteica , Propiedades de SuperficieRESUMEN
The local heating of poly(3,4-ethylenedioxythiophene) (PEDOT) by a photothermal effect directed by near-infrared (NIR) light induces unfolding of absorbed collagen triple helices, yielding soluble collagen single-helical structures. This dissociation of collagens allowed the harvesting of a living idiomorphic cell sheet, achieved upon irradiation with NIR light (λ=808â nm). The PEDOT layer was patterned and cells were successfully cultured on the patterned substrate. Cell sheets of various shapes mirroring the PEDOT pattern could be detached after a few minutes of irradiation with NIRâ light. The PEDOT patterns guided not only the entire shape of the cell sheets but also the spreading direction of the cells in the sheets. This photothermally induced dissociation of collagen provided a fast non-invasive harvesting method and tailor-made cell-sheet patterns.
Asunto(s)
Separación Celular/métodos , Colágeno/metabolismo , Colágeno/efectos de la radiación , Rayos Infrarrojos , Procesos Fotoquímicos , Temperatura , Compuestos Bicíclicos Heterocíclicos con Puentes/química , Fibroblastos/citología , Fibroblastos/metabolismo , Fibroblastos/efectos de la radiación , Humanos , Polímeros/químicaRESUMEN
BACKGROUND AIMS: Mesenchymal stromal cells (MSCs) have the ability to self-renew and differentiate into various cell types. Their plasticity and easy availability make them promising candidates for regenerative medicine. However, for successful clinical application, MSCs need to be expanded under a Good Manufacturing Practices-compliant system to obtain a large quantity of these cells. Although the viability and potency of these in vitro-expanded MSCs need to be maintained during preparation and transportation before transplantation, these characteristics have not thoroughly been examined. Our goal in this study was to standardize MSC preparation and storage before their clinical application to ensure reproducible quality and potency for their clinically intended purpose. METHODS: We examined the viability, self-renewal capacity and differentiation capability of MSCs on short-term in vitro storage in saline or dextrose solution at 4°C and room temperature. RESULTS: MSCs harvested and suspended in saline for 1-2 h showed >90% viability regardless of storage temperature. However, when cells were stored for >2 h in saline, their viability decreased gradually over time. The viability of cells in dextrose deteriorated rapidly. MSCs lost colony-forming unit and differentiation capacities rapidly as storage time increased. Collectively, we found that a storage period >2 h resulted in a significant decrease in cell viability, cell proliferation capacity and differentiation potency. CONCLUSIONS: Storage of culture-harvested MSCs for >2 h is likely to result in suboptimal MSC-mediated tissue regeneration because of decreased cell viability and differentiation capacity.
Asunto(s)
Técnicas de Cultivo de Célula , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/fisiología , Células de la Médula Ósea/citología , Diferenciación Celular , Proliferación Celular , Supervivencia Celular , Células Cultivadas , Regeneración Tisular Dirigida , HumanosRESUMEN
Exposure to particulate matter (PM) causes mitochondrial dysfunction and lung inflammation. The cyclooxygenase-2 (COX-2) pathway is important for inflammation and mitochondrial function. However, the mechanisms by which glucocorticoid receptors (GRs) suppress COX-2 expression during PM exposure have not been elucidated yet. Hence, we examined the mechanisms underlying the dexamethasone-mediated suppression of the PM-induced COX-2/prostaglandin E2 (PGE2) pathway in A549 cells. The PM-induced increase in COX-2 protein, mRNA, and promoter activity was suppressed by glucocorticoids; this effect of glucocorticoids was antagonized by the GR antagonist RU486. COX-2 induction was correlated with the ability of PM to increase reactive oxygen species (ROS) levels. Consistent with this, antioxidant treatment significantly abolished COX-2 induction, suggesting that ROS is involved in PM-mediated COX-2 induction. We also observed a low mitochondrial membrane potential in PM-treated A549 cells, which was reversed by dexamethasone. Moreover, glucocorticoids significantly enhanced Bcl-2/GR complex formation in PM-treated A549 cells. Glucocorticoids regulate the PM-exposed induction of COX-2 expression and mitochondrial dysfunction and increase the interaction between GR and Bcl-2. These findings suggest that the COX-2/PGE2 pathway and the interaction between GR and Bcl-2 are potential key therapeutic targets for the suppression of inflammation under PM exposure.
Asunto(s)
Dexametasona , Glucocorticoides , Humanos , Glucocorticoides/farmacología , Glucocorticoides/metabolismo , Ciclooxigenasa 2/genética , Ciclooxigenasa 2/metabolismo , Dexametasona/farmacología , Células A549 , Material Particulado/toxicidad , Dinoprostona/metabolismo , Especies Reactivas de Oxígeno , InflamaciónRESUMEN
Mitochondrial dysfunction is important in various chronic degenerative disorders, and aberrant immune responses elicited by cytoplasmic mitochondrial DNA (mtDNA) may be related. Here, we developed mtDNA-targeted MTERF1-FokI and TFAM-FokI endonuclease systems to induce mitochondrial DNA double-strand breaks (mtDSBs). In these cells, the mtDNA copy number was significantly reduced upon mtDSB induction. Interestingly, in cGAS knockout cells, synthesis of interferon ß1 and interferon-stimulated gene was increased upon mtDSB induction. We found that mtDSBs activated DNA-PKcs and HSPA8 in a VDAC1-dependent manner. Importantly, the mitochondrial E3 ligase MARCH5 bound active DNA-PKcs in cells with mtDSBs and reduced the type Ð interferon response through the degradation of DNA-PKcs. Likewise, mitochondrial damage caused by LPS treatment in RAW264.7 macrophage cells increased phospho-HSPA8 levels and the synthesis of mIFNB1 mRNA in a DNA-PKcs-dependent manner. Accordingly, in March5 knockout macrophages, phospho-HSPA8 levels and the synthesis of mIFNB1 mRNA were prolonged after LPS stimulation. Together, cytoplasmic mtDNA elicits a cellular immune response through DNA-PKcs, and mitochondrial MARCH5 may be a safeguard to prevent persistent inflammatory reactions.
Asunto(s)
Lipopolisacáridos , Ubiquitina-Proteína Ligasas , Humanos , Ubiquitina-Proteína Ligasas/metabolismo , Lipopolisacáridos/metabolismo , Proteínas de la Membrana/metabolismo , Mitocondrias/metabolismo , ADN Mitocondrial/genética , ADN Mitocondrial/metabolismo , Interferones/metabolismo , ARN Mensajero/metabolismoRESUMEN
Stem cell-derived exosomes are efficient and safe therapeutic tools for transferring endogenous biological cargo or functional biomolecules for regenerative medicine. The regulation of inflammation and angiogenesis plays a pivotal role in wound healing and tissue regeneration. The purpose of this study was to investigate the anti-inflammatory and pro-angiogenic roles of human adipose mesenchymal stem cell-derived exosomes, focusing on the underlying mechanisms. Exosomes inhibited LPS-induced inflammation by activating ROCK1 and PTEN expression. Moreover, microRNAs (miR-132 and miR-146a) released from exosomes upregulated the expression of pro-angiogenic genes and promoted proliferation activity and tube formation in human umbilical vein endothelial cells. Exosomal effects were verified using ROCK1/PTEN inhibitors for anti-inflammation and miR-132/miR-146a inhibitors for pro-angiogenesis. Our findings suggest that exosomes exert anti-inflammatory effects by targeting the ROCK1/PTEN pathway and exhibit pro-angiogenic effects via delivery of miR-132 and miR-146a. Taken together, these results suggest that exosomes may be promising therapeutic candidates for curing diseases involved in inflammation and angiogenesis.
Asunto(s)
Tejido Adiposo/metabolismo , Exosomas/metabolismo , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Madre Mesenquimatosas/metabolismo , Neovascularización Fisiológica , Humanos , Inflamación/metabolismoRESUMEN
Interactions of gelatin and albumin with a photo-reactive diphenylamino-s-triazine bridged p-phenylene vinylene polymer (DTOPV) were examined by using surface plasmon resonance (SPR) spectroscopy to explore the effect of the polymer structure on protein coverage of DTOPV nanofilms. The SPR data revealed a significant increase of gelatin adsorption on UV-DTOPV nanofilms, while the adsorption of albumin was decreased by UV exposure in the time frame of the experiment. We also found that the selective adsorption of these proteins was highly dependent on the protein concentration; the highest selectivity of protein adsorption was obtained at the lowest concentration (3.5 µg ml(-1)), while no selective adsorption was confirmed at high concentrations (350 and 1000 µg ml(-1)). The selective attachment of mesenchymal stem cells (MSCs) was directly correlated with the selective adsorption of these proteins onto DTOPV nanofilms. The MSCs attachment onto UV-DTOPV films was promoted with only small mass coverage of gelatin, which led to MSC patterning onto the patterned DTOPV nanofilms successfully. The role of cell adhesion proteins that we found in this study will be a clue to elucidate the complex response of biomolecules on functional polymer nanolayers, and contribute to build up biocompatible surfaces on various advanced materials for the sake of cell engineering and medical implants.
Asunto(s)
Albúminas/química , Gelatina/química , Células Madre Mesenquimatosas/química , Polivinilos/química , Adsorción , Humanos , Resonancia por Plasmón de Superficie , Propiedades de SuperficieRESUMEN
The present work describes the design and biological applications of a novel colorimetric and fluorescence turn-on probe for hydrosulfide detection. The probe was designed to introduce hemicyanine as the fluorescent skeleton and 7-nitro-1,2,3-benzoxadiazole as the recognition site. The optical properties and responses of the probe towards HS-, anions and some biothiols indicate an impressively high selectivity of the probe towards HS- such that it can be effectively used as an indicator for monitoring the level of HS- in living cells. In biological experiments using the probe, the H2S levels are found to be higher in cancer cells than in normal cells. In addition, the probe is shown to specifically and rapidly detect endogenous H2S, which is produced primarily in the mitochondria of cancer cells, as demonstrated by a co-localization experiment using specific trackers for the detection of cellular organelles in pharmacological inhibition or stimulation studies, without any significant cytotoxic effects. Thus, the results of the chemical and biological experiments described herein demonstrate the potential of this novel probe to specifically, safely, and rapidly detect H2S to distinguish cancer cells from normal cells by targeting it specifically in mitochondria.
Asunto(s)
Colorantes Fluorescentes/farmacología , Sulfuro de Hidrógeno/metabolismo , Mitocondrias/metabolismo , Oxadiazoles/farmacología , Línea Celular , Supervivencia Celular/efectos de los fármacos , Colorimetría , Fluorescencia , HumanosRESUMEN
BACKGROUND: Human adipose-derived mesenchymal stem cells (AMSCs) are an attractive resource for wound healing because their regenerative capacity improves injury repair. Recently, stem cell-derived exosomes have been shown to play a positive role in stem cell-based therapies. However, the effects of exosomes derived from AMSCs (AEXOs) on wound healing are unclear. In this study, we aimed to examine the role of AEXOs in attenuating inflammation and explore their effects in normal wound healing. METHODS: We isolated exosomes from AMSCs and established a cellular model of inflammation by treatment with the inflammatory cytokines, interferon gamma and tumor necrosis factor alpha, to determine whether AEXOs can inhibit inflammation. We examined the wound healing effects of AEXOs in in vitro wound healing models and performed a miRNA array to understand the role of AEXOs in inflammation and wound healing. RESULTS: A significant difference was observed in wound closure and the expression of anti-inflammatory and wound-healing-related factors between control and AEXO-treated cells. CONCLUSION: Our results showed that besides alleviating the inflammation response, AEXOs also promote wound healing. Thus, AEXOs represent a novel, stem-cell-based, therapeutic strategy for wound healing.
Asunto(s)
Exosomas , Células Madre Mesenquimatosas , MicroARNs , Tejido Adiposo , Humanos , MicroARNs/genética , Cicatrización de HeridasRESUMEN
The chromatin remodeler RSF1 enriched at mitotic centromeres is essential for proper chromosome alignment and segregation and underlying mechanisms remain to be disclosed. We here show that PLK1 recruitment by RSF1 at centromeres creates an activating phosphorylation on Thr236 in the activation loop of Aurora B and this is indispensable for the Aurora B activation. In structural modeling the phosphorylated Thr236 enhances the base catalysis by Asp200 nearby, facilitating the Thr232 autophosphorylation. Accordingly, RSF1-PLK1 is central for Aurora B-mediated microtubule destabilization in error correction. However, under full microtubule-kinetochore attachment RSF1-PLK1 positions at kinetochores, halts activating Aurora B and phosphorylates BubR1, regardless of tension. Spatial movement of RSF1-PLK1 to kinetochores is triggered by Aurora B-mediated phosphorylation of centromeric histone H3 on Ser28. We propose a regulatory RSF1-PLK1 axis that spatiotemporally controls on/off switch on Aurora B. This feedback circuit among RSF1-PLK1-Aurora B may coordinate dynamic microtubule-kinetochore attachment in early mitosis when full tension yet to be generated.
Asunto(s)
Aurora Quinasa B/genética , Proteínas de Ciclo Celular/genética , Segregación Cromosómica , Mitosis , Proteínas Nucleares/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Proto-Oncogénicas/genética , Transducción de Señal/genética , Transactivadores/genética , Ácido Aspártico/metabolismo , Aurora Quinasa B/metabolismo , Proteínas de Ciclo Celular/metabolismo , Cromatina/química , Cromatina/metabolismo , Retroalimentación Fisiológica , Regulación de la Expresión Génica , Células HeLa , Histonas/genética , Histonas/metabolismo , Humanos , Cinetocoros/metabolismo , Cinetocoros/ultraestructura , Microtúbulos/metabolismo , Microtúbulos/ultraestructura , Proteínas Nucleares/deficiencia , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Serina/metabolismo , Transactivadores/deficiencia , Quinasa Tipo Polo 1RESUMEN
The positive effects of mesenchymal stem cells (MSCs) are primarily activated through molecular secretions known as paracrine activity, which regulates the function of various cell types including immune cells. Accumulating evidence shows that exosomes of soluble factors released from MSCs are potential alternative agents for stem cell-based therapy, although the exact underlying mechanism has not been elucidated. The purpose of this study was to evaluate the potential effects of exosomes produced by adipose-derived MSCs and to examine the changes in anti-inflammatory genes in concurrence with the polarization of M2 macrophages in cellular models ex vivo. Isolated exosomes were used to investigate the inflammatory modulation in pro-inflammatory cytokine-treated fibroblasts and THP-1 cells. The anti-inflammatory mRNA expression associated with M2 macrophages was significantly upregulated after exosome treatment in an interferon gamma and tumor necrosis factor alpha-treated inflammatory environment. Furthermore, melatonin-stimulated exosomes exerted superior anti-inflammatory modulation via exosomal miRNAs miR-34a, miR-124, and miR-135b, compared with exosomes. Our results indicate that melatonin-stimulated exosomes originating from adipose-derived MSCs are safe and efficient tools for regenerative medicine to treat inflammatory diseases.
Asunto(s)
Exosomas/metabolismo , Inflamación/patología , Melatonina/farmacología , MicroARNs/metabolismo , Proliferación Celular/efectos de los fármacos , Exosomas/efectos de los fármacos , Exosomas/ultraestructura , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Mediadores de Inflamación/farmacología , Interferón gamma/farmacología , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , MicroARNs/genética , Células THP-1 , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
The mitochondrial antiviral signaling (MAVS) protein on the mitochondrial outer membrane acts as a central signaling molecule in the RIG-I-like receptor (RLR) signaling pathway by linking upstream viral RNA recognition to downstream signal activation. We previously reported that mitochondrial E3 ubiquitin ligase, MARCH5, degrades the MAVS protein aggregate and prevents persistent downstream signaling. Since the activated RIG-I oligomer interacts and nucleates the MAVS aggregate, MARCH5 might also target this oligomer. Here, we report that MARCH5 targets and degrades RIG-I, but not its inactive phosphomimetic form (RIG-IS8E). The MARCH5-mediated reduction of RIG-I is restored in the presence of MG132, a proteasome inhibitor. Upon poly(I:C) stimulation, RIG-I forms an oligomer and co-expression of MARCH5 reduces the expression of this oligomer. The RING domain of MARCH5 is necessary for binding to the CARD domain of RIG-I. In an in vivo ubiquitination assay, MARCH5 transfers the Lys 48-linked polyubiquitin to Lys 193 and 203 residues of RIG-I. Thus, dual targeting of active RIG-I and MAVS protein oligomers by MARCH5 is an efficient way to switch-off RLR signaling. We propose that modulation of MARCH5 activity might be beneficial for the treatment of chronic immune diseases.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteína 58 DEAD Box/metabolismo , Inmunidad Innata , Proteínas de la Membrana/metabolismo , Mitocondrias/metabolismo , Receptores Inmunológicos/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitina/metabolismo , Animales , Proteína 58 DEAD Box/química , Células HEK293 , Humanos , Lisina/metabolismo , Ratones , Unión Proteica , Dominios Proteicos , Proteolisis , Células RAW 264.7 , Receptores Inmunológicos/química , UbiquitinaciónRESUMEN
Accumulating evidence has shown that the paracrine factors derived from mesenchymal stem cells (MSCs) are capable of regulating the immune system via interaction with various immune cells. In this study, adipose-derived MSCs (AdMSCs) and human peripheral blood monocytes (PBMCs) were isolated and cultured to examine the effects of MSC-induced macrophages (iMΦ) on inflammation and immune modulation. Indirect coculture with MSCs increased the expression of arginase-1 and mannose receptor (CD206), markers of activated M2 macrophages, in the PBMCs demonstrating that MSC-secreted factors promoted M2-MΦ polarization. Additionally, iMΦ exhibited a similar higher inhibitory effect on the growth of activated T cells compared to that in the other groups (AdMSCs only, AdMSCs plus iMΦ), implying that iMΦ can play a sufficient functional role. Interestingly, the population of FoxP3 Treg cells significantly increased when cocultured with iMΦ, suggesting that iMΦ have an immunomodulatory effect on the Treg cells through the modulation of the FoxP3 expression. Notably, iMΦ expressed high levels of immunosuppressive and anti-inflammatory cytokines, namely IL-10 and TSG-6. Furthermore, we confirmed that the AdMSC-derived exosomes modulated macrophage polarization by upregulating the expression of M2 macrophage markers. Conclusively, our results suggest that iMΦ play a significant role in regulating the immunomodulatory- and inflammatory-mediated responses. Thus, iMΦ may be used as a novel stem cell-based cell-free therapy for the treatment of immune-mediated inflammatory disorders.
RESUMEN
The toxic effects of particulate matter have been linked to polycyclic aromatic hydrocarbons (PAHs) such as benzopyrene. PAHs are potent inducers of the aryl hydrocarbon receptor (AhR), which is an expressed nuclear receptor that senses environmental stimuli and modulates gene expression. Even though several studies have shown that the benzopyrene (BP) of chemical pollutants significantly impaired stem cell activity, the exact molecular mechanisms were not clearly elucidated. In the present study, we aimed to investigate the effects of BP on placenta-derived mesenchymal stem cells (PD-MSCs) in vitro. We found that the AhR in PD-MSCs was expressed under the treatment of BP, and its activation markedly disrupted osteogenic differentiation through the alteration of stemness activity of PD-MSCs. Moreover, BP treatment significantly reduced the proliferation activity of PD-MSCs and expression of pluripotent markers through the induction of AhR. Treatment with StemRegenin 1 (SR1), a purine derivative that antagonizes the AhR, effectively prevented BP-induced reduction of the proliferation and differentiation activity of PD-MSCs. In this study, we found that BP treatment in PD-MSCs markedly obstructs PD-MSC stemness through AhR signaling. Noteworthy, SR1-mediated MSC application will contribute to new perspectives on MSC-based therapies for air pollution-related bone diseases.
RESUMEN
Infection of hepatitis B virus (HBV) increase the incidence of chronic liver disease and hepatocellular carcinoma (HCC). The hepatitis B viral x (HBx) protein encoded by the HBV genome contributes to the pathogenesis of HCC and thus, negative regulation of HBx is beneficial for the alleviation of the disease pathogenesis. MARCH5 is a mitochondrial E3 ubiquitin ligase and here, we show that high MARCH5 expression levels are correlated with improved survival in HCC patients. MARCH5 interacts with HBx protein mainly accumulated in mitochondria and targets it for degradation. The N-terminal RING domain of MARCH5 was required for the interaction with HBx, and MARCH5H43W lacking E3 ligase activity failed to reduce HBx protein levels. High expression of HBx results in the formation of protein aggregates in semi-denaturing detergent agarose gels and MARCH5 mediates the elimination of protein aggregates through the proteasome pathway. HBx-induced ROS production, mitophagy, and cyclooxygenase-2 gene expression were suppressed in the presence of high MARCH5 expression. These results suggest MARCH5 as a target for alleviating HBV-mediated liver disease.
Asunto(s)
Carcinoma Hepatocelular/metabolismo , Virus de la Hepatitis B/química , Hepatitis B/metabolismo , Neoplasias Hepáticas/metabolismo , Proteínas de la Membrana/metabolismo , Agregado de Proteínas , Proteolisis , Transactivadores/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas Reguladoras y Accesorias Virales/metabolismo , Carcinoma Hepatocelular/etiología , Carcinoma Hepatocelular/mortalidad , Carcinoma Hepatocelular/patología , Células HEK293 , Células HeLa , Hepatitis B/complicaciones , Hepatitis B/virología , Humanos , Neoplasias Hepáticas/etiología , Neoplasias Hepáticas/mortalidad , Neoplasias Hepáticas/patología , Proteínas de la Membrana/genética , Mitocondrias/metabolismo , Agregación Patológica de Proteínas/metabolismo , Tasa de Supervivencia , Transfección , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
Mesenchymal stem cells (MSCs) are capable of differentiating into other cell types and exhibit immunomodulatory effects. MSCs are affected by several intrinsic and extrinsic signaling modulators, including growth factors, cytokines, extracellular matrix and hormones. Melatonin, produced by the pineal gland, is a hormone that regulates sleep cycles. Recent studies have shown that melatonin improves the therapeutic effects of stem cells. The present study aimed to investigate whether melatonin enhances the biological activities of human adiposederived MSCs. The results demonstrated that treatment with melatonin promoted cell proliferation by inducing SRYbox transcription factor 2 gene expression and preventing replicative senescence. In addition, melatonin exerted antiadipogenic effects on MSCs. PCR analysis revealed that the expression of the CCAAT enhancer binding protein a gene, a key transcription factor in adipogenesis, was decreased following melatonin treatment, resulting in reduced adipogenic differentiation in an in vitro assay. The present study also examined the effect of melatonin on the immunomodulatory response using a coculture system of human peripheral blood mononuclear cells and MSCs. Activated T cells were strongly inhibited following melatonin exposure compared with those in the control group. Finally, the favorable effects of melatonin on MSCs were confirmed using luzindole, a selective melatonin receptor antagonist. The proliferationpromoting, antiinflammatory effects of melatonin suggested that melatonintreated MSCs may be used for effective cell therapy.
Asunto(s)
Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Melatonina/farmacología , Células Madre Mesenquimatosas/efectos de los fármacos , Adipogénesis/efectos de los fármacos , Adipogénesis/genética , Adulto , Femenino , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Humanos , Leucocitos Mononucleares/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Persona de Mediana Edad , Factores de Transcripción SOXB1/genética , Transducción de Señal/efectos de los fármacos , Triptaminas/farmacologíaRESUMEN
Accumulating evidence has revealed that both high sensitivity C-reactive protein (hsCRP) and homocysteine (HCY) are associated with increased risk of metabolic syndrome (MetS) and cardiovascular disease. However, it is unclear whether the coexistence of these conditions accelerates the risk of metabolic syndrome (MetS). We hypothesized that the combination of high sensitivity C-reactive protein (hsCRP) and homocysteine (HCY) levels could exacerbate the development of MetS in a large prospective cohort study. We selected data from 3,170 individuals (1,614 men and 1,556 women) who participated in the Korean Genome and Epidemiology Study. Participants with high hsCRP and HCY levels were categorized into quartiles. MetS was defined based on the criteria of the modified National Cholesterol Education Program, Adult Treatment Panel III. The prevalence of MetS was higher in participants with concurrent high hsCRP and HCY compared to those with low hsCRP and HCY levels. The incidence of MetS at the 6-year follow-up was the highest in participants with concomitant high hsCRP and HCY levels, regardless of obesity. Even after adjusting for potential confounding factors including body mass index in a multivariate logistic regression model, subjects with elevated hsCRP and HCY levels had a 2.50-fold increased risk of developing MetS at the six-year follow-up compared to those who did not have high hsCRP and HCY level. MetS is more prevalent in the concurrent presence of elevated hsCRP and HCY levels. The combination of the two conditions may contribute to an increased risk of MetS, but these factors may not be synergistic.