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1.
APMIS ; 128(11): 583-592, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32865844

RESUMO

Multiple sclerosis (MS) is an immune-mediated inflammatory disease which affects the central nervous system (CNS). In the present study, the in vivo effects of ATRA, calcitriol, and their combinations on the expression of murine CD4+ T cell cytokines and their specific transcription factors in experimental autoimmune encephalomyelitis (EAE)-induced mice were explored. Thirty-two EAE induced inbred C57BL/6 female mice with an age ranged from 8 to 10 weeks were divided into four categories in a random manner. The first, second, and third groups received ATRA, calcitriol, ATRA+ calcitriol, respectively, and the fourth group received vehicle. The treatment started on the day prior to immunization and through the IP injections every other days for 21 days. The dosages of administration for calcitriol, ATRA, and calcitriol+ ATRA were 100 ng, 250 µg, and 50ng + 125 µg, respectively per mouse. An equal volume of excipient was administered for the vehicle group. T-bet, IFN-γ, GATA-3, and IL-4 genes expression were assessed in the splenocytes of EAE -induced mice. The expression of T-bet and IFN-γ genes in the splenocytes of ATRA, calcitriol and combination- treated mice were significantly reduced compared to vehicle group (p < 0.05). A significant decrease in T-bet expression was observed in the combination-treated group compared to the ATRA-treated group (p < 0.05). The expression of GATA3 and IL-4 genes was significantly increased in the ATRA-, calcitriol-, and combination-treated mice when compared with the control group (p < 0.05). Furthermore, the effect of calcitriol alone and in combination with ATRA was more considerable than that of ATRA alone. The nutraceutical approaches may be promising in the prevention and/or treatment of MS.


Assuntos
Calcitriol/farmacologia , Encefalomielite Autoimune Experimental/tratamento farmacológico , Regulação da Expressão Gênica/efeitos dos fármacos , Fatores Imunológicos/farmacologia , Fármacos Neuroprotetores/farmacologia , Tretinoína/farmacologia , Animais , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Esquema de Medicação , Quimioterapia Combinada , Encefalomielite Autoimune Experimental/induzido quimicamente , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Feminino , Fator de Transcrição GATA3/genética , Fator de Transcrição GATA3/imunologia , Interferon gama/genética , Interferon gama/imunologia , Interleucina-4/genética , Interleucina-4/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Glicoproteína Mielina-Oligodendrócito/administração & dosagem , Transdução de Sinais , Medula Espinal/efeitos dos fármacos , Medula Espinal/imunologia , Medula Espinal/patologia , Proteínas com Domínio T/genética , Proteínas com Domínio T/imunologia
2.
Yonsei Med J ; 61(9): 762-773, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32882760

RESUMO

PURPOSE: Pharmacological inhibition of mutant isocitrate dehydrogenase (IDH) reduces R-2-hydroxyglutarate (2-HG) levels and restores cellular differentiation in vivo and in vitro. The IDH2 inhibitor enasidenib (AG-221) has been approved by the FDA as a first-in-class inhibitor for the treatment of relapsed or refractory (R/R) IDH2-mutant acute myeloid leukemia (AML). In this study, the effects of a combination of all-trans retinoic acid (ATRA) and AG-221 on AML cell differentiation was explored, along with the mechanisms employed by IDH2-mutant cells in AML. MATERIALS AND METHODS: We treated the human AML cell line, IDH2-mutant-TF-1, and primary human AML cells carrying IDH2 mutation with 30 µM AG-221 and 100 nM ATRA, alone or in combination. RESULTS: Combined treatment with AG-221 and ATRA inhibited 2-HG production and resulted in synergistic effects on differentiation among IDH2-mutant AML cells and primary AML cells expressing IDH2 mutation. Combined treatment with AG-221 and ATRA altered autophagic activity. AG-221 and ATRA treatment-induced differentiation of IDH2-mutant AML cells was associated with autophagy induction, without suppressing autophagy flux at maturation and degradation stages. A RAF-1/MEK/ERK pathway was founded to be associated with AG-221 and ATRA-induced differentiation in IDH2-mutant AML cells. IDH-associated changes in histone methylation markers decreased after AG-221 and ATRA combination treatment. CONCLUSION: Our preliminary evidence indicates that the addition of ATRA to treatments with IDH2 inhibitor may lead to further improvements or increases in response rates in IDH2-mutant AML patients who do not appear to benefit from treatments with IDH2 inhibitor alone.


Assuntos
Aminopiridinas/farmacologia , Antineoplásicos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Isocitrato Desidrogenase/genética , Leucemia Mieloide Aguda/tratamento farmacológico , Tretinoína/farmacologia , Triazinas/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Isocitrato Desidrogenase/antagonistas & inibidores , Isocitrato Desidrogenase/metabolismo , Leucemia Mieloide Aguda/genética , Mutação
3.
Gene ; 755: 144889, 2020 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-32534056

RESUMO

Ferroptosis, a newly discovered form of non-apoptotic cell death, is induced by an excessive degree of iron-dependent lipid peroxide. ATPR, a novel all-trans retinoic acid (ATRA) derivative, has been extensively developed to show superior anticancer effect than ATRA in acute myeloid leukemia (AML). However, whether ferroptosis exists during ATPR treatment of AML remains unclear. Herein, we found that ferroptosis occurred in an AML xenograft mouse model of ATPR treatment. In vitro, ATPR was verified to induce ferroptosis in a dose-dependent manner by proferroptotic protein marker, lipid peroxidation, and lipid ROS, which could be significantly reversed by ferrostatin-1. Using lysosomal inhibitor chloroquine and iron chelator desferrioxamine, we further revealed that ATPR-induced ferroptosis was regulated by autophagy via iron homeostasis, especially Nrf2. Furthermore, targeting ferroptosis contributes to ATPR-induced AML differentiation. In conclusion, these results indicated that ferroptosis play an important role in ATPR-induced differentiation, and suggested that ATPR would provide a potential therapeutic value for AML treatment.


Assuntos
Ferroptose/efeitos dos fármacos , Leucemia Mieloide Aguda/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Retinoides/farmacologia , Animais , Antineoplásicos/farmacologia , Autofagia/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Homeostase , Humanos , Ferro/metabolismo , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos Nus , Transdução de Sinais/efeitos dos fármacos , Tretinoína/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Zhongguo Ying Yong Sheng Li Xue Za Zhi ; 36(1): 45-50, 2020 Jan 28.
Artigo em Chinês | MEDLINE | ID: mdl-32476372

RESUMO

OBJECTIVE: To investigate the probable roles of the novel C2H2 zinc finger transcription factor ZFP580 on all-transretinoic acid (ATRA)-regulated VSMCs migration and underlying mechanisms. METHODS: Rat aortic VSMCs were isolated, cultured and identified. VSMCs were treated with ATRA at the concentrations of 0, 5, 10 or 20 µmol/L for 24 hours. The migration ability of VSMCs was observed in each group and compared with control group which was treated by 0 µmol/L ATRA. The mRNA and protein expression levels of ZFP580 were detected by QPCR and Western blot. ZFP580 protein expression in VSMCs was detected under ATRA stimulation when ERK inhibitor PD98059 was used to inhibit the protein expression of ERK. Adenovirus transfection technology was used to obtain VSMCs with overexpression or low expression of ZFP580, and QPCR and Western blot were used to detect the mRNA and protein levels of MMP-2, MMP-9 and ZFP580. RESULTS: On the 10th day of VSMCs culture, immunofluorescence showed that SM22 alpha antibody, as a specific marker of smooth muscle cells, was positive. Compared to the control group, VSMCs migration was reduced by 32%, 43%, and 59% in the group of 5, 10, and 20 µmol/L ATRA pretreatment. Compared with the control group, VSMCs treated by 20 µmol/L ATRA reduced the cell migration by 49%, 36% and 22% at 24, 48 and 72 h. The mRNA and protein expression levels of ZFP580 were increased with the increase of ATRA stimulation solubility and the extension of stimulation time. ERK was increased significantly after 15 min of ATRA stimulation. Pretreatment with ERK inhibitor PD98059 (20 µmol/L) inhibited the expression of ERK protein and reduced the expression of ATRA-induced ZFP580 protein. Overexpression of ZFP580 inhibited the expressions of MMP-2 and MMP-9, whereas down-expression of ZFP580 promoted the expressions of MMP-2 and MMP-9. CONCLUSION: ATRA increased the expression of ZFP580 through the ERK signaling pathway, while ZFP580 was involved in ATRA's inhibition of VSMCs migration by affecting the expression of downstream MMP-2 and MMP-9.


Assuntos
Movimento Celular , Miócitos de Músculo Liso/citologia , Fatores de Transcrição/metabolismo , Tretinoína/farmacologia , Animais , Células Cultivadas , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/efeitos dos fármacos , Ratos , Transdução de Sinais
5.
Nat Commun ; 11(1): 2082, 2020 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-32350257

RESUMO

Developmental progression depends on temporally defined changes in gene expression mediated by transient exposure of lineage intermediates to signals in the progenitor niche. To determine whether cell-intrinsic epigenetic mechanisms contribute to signal-induced transcriptional responses, here we manipulate the signalling environment and activity of the histone demethylase LSD1 during differentiation of hESC-gut tube intermediates into pancreatic endocrine cells. We identify a transient requirement for LSD1 in endocrine cell differentiation spanning a short time-window early in pancreas development, a phenotype we reproduced in mice. Examination of enhancer and transcriptome landscapes revealed that LSD1 silences transiently active retinoic acid (RA)-induced enhancers and their target genes. Furthermore, prolonged RA exposure phenocopies LSD1 inhibition, suggesting that LSD1 regulates endocrine cell differentiation by limiting the duration of RA signalling. Our findings identify LSD1-mediated enhancer silencing as a cell-intrinsic epigenetic feedback mechanism by which the duration of the transcriptional response to a developmental signal is limited.


Assuntos
Células Endócrinas/citologia , Células Endócrinas/metabolismo , Elementos Facilitadores Genéticos/genética , Inativação Gênica , Histona Desmetilases/metabolismo , Ilhotas Pancreáticas/citologia , Transdução de Sinais , Tretinoína/metabolismo , Adulto , Animais , Sequência de Bases , Diferenciação Celular/efeitos dos fármacos , Células Endócrinas/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Embrionárias Humanas/efeitos dos fármacos , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Ilhotas Pancreáticas/embriologia , Masculino , Camundongos , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição/metabolismo , Tretinoína/farmacologia , Adulto Jovem
6.
Exp Parasitol ; 215: 107917, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32446699

RESUMO

Cystic echinococcosis (CE) is a worldwide hazardous zoonotic parasitosis caused by Echinococcus granulosus. CE development involves complex immunological mechanisms, including participation of multiple immune cells and effector molecules. Myeloid-derived suppressor cells (MDSCs) are known to be involved in chronic and acute inflammatory conditions. In this study, we aimed to characterize the immune function of MDSCs in CE to improve the understanding, prevention and treatment of CE. Our results indicated that MDSCs overexpressing Ly6C and Ly6G inhibit the formation and activity of T helper 2 cells in a NO-dependent manner during E. granulosus infection.


Assuntos
Equinococose/imunologia , Echinococcus granulosus/imunologia , Células Supressoras Mieloides/imunologia , Células Th2/imunologia , Análise de Variância , Animais , Anticorpos Monoclonais , Arginase/análise , Medula Óssea/efeitos dos fármacos , Medula Óssea/imunologia , Citocinas/análise , Feminino , Citometria de Fluxo , Humanos , Ceratolíticos/farmacologia , Camundongos , Células Supressoras Mieloides/efeitos dos fármacos , Células Supressoras Mieloides/enzimologia , Óxido Nítrico/análise , Espécies Reativas de Oxigênio/análise , Organismos Livres de Patógenos Específicos , Baço/citologia , Baço/efeitos dos fármacos , Baço/imunologia , Tretinoína/farmacologia
7.
Biochim Biophys Acta Mol Cell Res ; 1867(7): 118708, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32240712

RESUMO

Spermatogonia migrate to the microenvironment during the establishment from gonocytes and leave it when they differentiate. However, the mechanisms underlying the regulation of spermatogonial differentiation-associated migration remain mostly unknown. In this study, we show that spermatogonial differentiation induced by retinoic acid (RA) was accompanied with increased migration ability and elevated expression of connective tissue growth factor (CTGF), a member of the CCN family. CTGF was mainly expressed in the testicular somatic cells and committed spermatogonial progenitors. Recombinant CTGF (rCTGF) promoted the spermatogonial migration and silencing of endogenous CTGF suppressed the migration of homogenous spermatogonial cell lines. Moreover, depletion of CTGF by neutralizing antibody inhibited the elevated migration ability induced by RA, suggesting both the paracrine and autocrine roles of CTGF in spermatogonial migration associated with differentiation. Finally, CTGF interacted with ß1-integrin and regulated its level in spermatogonial cell lines. Together, our study provides novel insights into the regulation of spermatogonial migration by CTGF, which may shed light on the diagnosis and treatment of male infertility.


Assuntos
Movimento Celular/genética , Fator de Crescimento do Tecido Conjuntivo/genética , Integrina beta1/genética , Espermatogônias/crescimento & desenvolvimento , Animais , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Humanos , Infertilidade Masculina/diagnóstico , Infertilidade Masculina/genética , Infertilidade Masculina/patologia , Masculino , Camundongos , Espermatogônias/metabolismo , Tretinoína/farmacologia
8.
Acta Cir Bras ; 35(1): e202000106, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32236320

RESUMO

PURPOSE: To explore the role of all-trans retinoic acid (ATRA) in renal ischemia/reperfusion injury of diabetic rats. METHODS: Sixty adult male rats were randomly divided into 6 groups, including sham group (S group), ischemia-reperfusion group (I/R group), ischemia-reperfusion+ATRA group (A group), diabetic group (D group), diabetic ischemia-reperfusion group (DI/R group), diabetic ischemia-reperfusion +ATRA group (DA group). The levels of creatinine (Cr), cystatin C (Cys-C) and ß2-microglobulin (ß2-MG) were measured. Morphology of renal tissue was observed under light microscope. RESULTS: DJ-1, Nrf2, HO-1 and caspase-3 were detected by western blot. DJ-1, Nrf2, HO-1 and caspase-3 in I/R group, D group and DI/R group was higher than that in S group. Compared with I/R group, Nrf2 and HO-1 in A group was decreased, but caspase-3 was increased. However, Nrf2 in DA group was higher than that in DI/R group, HO-1 and caspase-3 in DA group were lower than that in DI/R group. Compared with group S, Cr, Cys-C and ß2-MG in I/R group, A group, D group, and DI/R group were higher. Whereas the levels of Cr, Cys-C, ß2-MG and renal injury score in DA group were lower than those in DI/R group. CONCLUSION: ATRA has a protective effect on renal ischemia-reperfusion injury in diabetic rats, maybe relating to DJ/Nrf2 pathway.


Assuntos
Diabetes Mellitus Experimental/induzido quimicamente , Rim/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/uso terapêutico , Traumatismo por Reperfusão/prevenção & controle , Tretinoína/uso terapêutico , Animais , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Rim/patologia , Masculino , Fator 2 Relacionado a NF-E2/farmacologia , Ratos , Traumatismo por Reperfusão/patologia , Estreptozocina , Tretinoína/farmacologia
9.
PLoS One ; 15(3): e0229892, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32231396

RESUMO

The specification of cell identity depends on the exposure of cells to sequences of bioactive ligands. All-trans retinoic acid (ATRA) affects neuronal development in the early stage, and it is involved in neuronal lineage reprogramming. We previously established a fibroblast-like dedifferentiated fat cells (DFATs) derived from highly homogeneous mature adipocytes, which are more suitable for the study of cellular reprogramming. Canine cognitive dysfunction is similar to human cognitive dysfunction, suggesting that dogs could be a pathological and pharmacological model for human neuronal diseases. However, the effect of ATRA on neuronal reprogramming in dogs has remained unclear. Therefore, in this study, we investigated the effect of ATRA on the neuronal reprogramming of canine DFATs. ATRA induced the expression of neuronal marker mRNA/protein. The neuron-like cells showed Ca2+ influx with depolarization (50 mM KCl; 84.75 ± 4.05%) and Na+ channel activation (50 µM veratridine; 96.02 ± 2.02%). Optical imaging of presynaptic terminal activity and detection of neurotransmitter release showed that the neuron-like cells exhibited the GABAergic neuronal property. Genome-wide RNA-sequencing analysis shows that the transcriptome profile of canine DFATs is effectively reprogrammed towards that of cortical interneuron lineage. Collectively, ATRA can produce functional GABAergic cortical interneuron-like cells from canine DFATs, exhibiting neuronal function with > 80% efficiency. We further demonstrated the contribution of JNK3 to ATRA-induced neuronal reprogramming in canine DFATs. In conclusion, the neuron-like cells from canine DFATs could be a powerful tool for translational research in cell transplantation therapy, in vitro disease modeling, and drug screening for neuronal diseases.


Assuntos
Desdiferenciação Celular/efeitos dos fármacos , Neurogênese/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Tretinoína/farmacologia , Animais , Diferenciação Celular/efeitos dos fármacos , Reprogramação Celular/efeitos dos fármacos , Cães , Neurogênese/genética , RNA Mensageiro/genética , Sinapses/efeitos dos fármacos , Sinapses/genética
10.
J Leukoc Biol ; 107(4): 685-693, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32125014

RESUMO

As a subtype of acute myeloid leukemia (AML), acute promyelocytic leukemia (APL) is characterized by a chromosomal translocation, most of which result in the production of a PML-RAR alpha fusion protein. Although the overall survival rate of APL patients has improved dramatically due to all-trans retinoic acid (ATRA) treatment, ATRA-resistance remains a clinical challenge in the management of APL. Therefore, alternative agents should be considered for ATRA-resistant APL patients. Here, we report that antimalaria drug primaquine phosphate (PRQ) exhibits an anti-leukemia effect on both ATRA-sensitive cell line NB4 and ATRA-resistant APL cell lines, NB4-LR2, NB4-LR1, and NB4-MR2. Moreover, PRQ significantly inhibited primary colony formation of untreated or relapsed APL patients. Further study showed that PRQ could induce the apoptosis of APL cells by inhibiting NF-κB signaling pathway. The in vivo study showed that PRQ significantly inhibited NB4-LR2 xenograft tumors growth. These results suggest that PRQ is a potential therapeutic agent for ATRA-resistant APL patients.


Assuntos
Apoptose/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Leucemia Promielocítica Aguda/patologia , NF-kappa B/metabolismo , Primaquina/farmacologia , Transdução de Sinais , Tretinoína/farmacologia , Adulto , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Feminino , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Primaquina/química , Transdução de Sinais/efeitos dos fármacos , Ensaio Tumoral de Célula-Tronco , Ensaios Antitumorais Modelo de Xenoenxerto
11.
Curr Med Sci ; 40(1): 55-62, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32166665

RESUMO

The present study aimed to explore the molecular mechanisms underlying the increase of nicotinamide adenine dinucleotide phosphate:quinine oxidoreductase 1 (NQO1) and γ-glutamylcysteine synthetase (γ-GCS) in brain tissues after intracerebral hemorrhage (ICH). The microglial cells obtained from newborn rats were cultured and then randomly divided into the normal control group (NC group), model control group (MC group), rosiglitazone (RSG) intervention group (RSG group), retinoic-acid intervention group (RSG+RA group), and sulforaphane group (RSG+SF group). The expression levels of NQO1, γ-GCS, and nuclear factor E2-related factor 2 (Nrf2) were measured by real-time polymerase chain reaction (RT-PCR) and Western blotting, respectively. The results showed that the levels of NQO1, γ-GCS and Nrf2 were significantly increased in the MC group and the RSG group as compared with those in the NC group (P<0.01). They were found to be markedly decreased in the RSG+RA group and increased in the RSG+SF group when compared with those in the MC group or the RSG group (P<0.01). The RSG+SF group displayed the highest levels of NQO1, γ-GCS, and Nrf2 among the five groups. In conclusion, a medium dose of RSG increased the anti-oxidative ability of thrombin-activated microglia by increasing the expression of NQO1 and γ-GCS. The molecular mechanisms underlying the increase of NQO1 and γ-GCS in thrombin-activated microglia may be associated with the activation of Nrf2.


Assuntos
Hemorragia Cerebral/genética , Glutamato-Cisteína Ligase/genética , Microglia/citologia , NAD(P)H Desidrogenase (Quinona)/genética , Fator 2 Relacionado a NF-E2/genética , PPAR gama/genética , Trombina/farmacologia , Animais , Animais Recém-Nascidos , Células Cultivadas , Hemorragia Cerebral/tratamento farmacológico , Hemorragia Cerebral/metabolismo , Modelos Animais de Doenças , Feminino , Glutamato-Cisteína Ligase/metabolismo , Isotiocianatos/administração & dosagem , Isotiocianatos/farmacologia , Masculino , Microglia/efeitos dos fármacos , Microglia/metabolismo , NAD(P)H Desidrogenase (Quinona)/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , PPAR gama/metabolismo , Cultura Primária de Células , Ratos , Ratos Sprague-Dawley , Rosiglitazona/administração & dosagem , Rosiglitazona/farmacologia , Tretinoína/administração & dosagem , Tretinoína/farmacologia
12.
Life Sci ; 249: 117479, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32119959

RESUMO

A concomitant change of nucleus shape and chromosome conformation often happens in all-trans retinoic acid (ATRA)-induced differentiation of acute myeloid leukemia cells. However, the relation between the 3D chromosome architecture and the genome-wide epigenetic pattern for transcriptional regulation is poorly understood. In this study, high-throughput chromosome conformation capture (Hi-C) and chromosome immunoprecipitation (ChIP-seq) were employed to investigate the landscape of chromosome distal interaction and H3K4/27me3 in HL-60 cells treated with ATRA. We observed a general loss of topological associated domains (TADs) at PTPN11 during the differentiation of HL-60 cells. Furthermore, the significantly reduced enrichment of CCCTC binding factor (CTCF) near the boundary where PTPN11 located, as well as the decreased H3K4me3 and increased H3K27me3 enrichment at PTPN11 upon ATRA treatment was observed. Taken together, our study indicated a regulatory mechanism behind the silenced PTPN11 in HL-60 cells differentiation.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Cromossomos Humanos , Leucemia Mieloide Aguda/patologia , Tretinoína/farmacologia , Imunoprecipitação da Cromatina , Células HL-60 , Histonas/metabolismo , Humanos , Leucemia Mieloide Aguda/genética
13.
Biochem J ; 477(4): 817-831, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-32016357

RESUMO

Inorganic phosphate (Pi) homeostasis is regulated by intestinal absorption via type II sodium-dependent co-transporter (Npt2b) and by renal reabsorption via Npt2a and Npt2c. Although we previously reported that vitamin A-deficient (VAD) rats had increased urine Pi excretion through the decreased renal expression of Npt2a and Npt2c, the effect of vitamin A on the intestinal Npt2b expression remains unclear. In this study, we investigated the effects of treatment with all-trans retinoic acid (ATRA), a metabolite of vitamin A, on the Pi absorption and the Npt2b expression in the intestine of VAD rats, as well as and the underlying molecular mechanisms. In VAD rats, the intestinal Pi uptake activity and the expression of Npt2b were increased, but were reduced by the administration of ATRA. The transcriptional activity of reporter plasmid containing the promoter region of the rat Npt2b gene was reduced by ATRA in NIH3T3 cells overexpressing retinoic acid receptor (RAR) and retinoid X receptor (RXR). On the other hand, CCAAT/enhancer-binding proteins (C/EBP) induced transcriptional activity of the Npt2b gene. Knockdown of the C/EBP gene and a mutation analysis of the C/EBP responsible element in the Npt2b gene promoter indicated that C/EBP plays a pivotal role in the regulation of Npt2b gene transcriptional activity by ATRA. EMSA revealed that the RAR/RXR complex inhibits binding of C/EBP to Npt2b gene promoter. Together, these results suggest that ATRA may reduce the intestinal Pi uptake by preventing C/EBP activation of the intestinal Npt2b gene.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Intestino Delgado/metabolismo , Rim/metabolismo , Regiões Promotoras Genéticas , Proteínas Cotransportadoras de Sódio-Fosfato Tipo IIb/genética , Transcrição Genética/efeitos dos fármacos , Tretinoína/farmacologia , Animais , Antineoplásicos/farmacologia , Proteínas Estimuladoras de Ligação a CCAAT/genética , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Hipofosfatemia Familiar/metabolismo , Hipofosfatemia Familiar/patologia , Hipofosfatemia Familiar/prevenção & controle , Intestino Delgado/efeitos dos fármacos , Rim/efeitos dos fármacos , Masculino , Camundongos , Células NIH 3T3 , Ratos , Ratos Wistar , Receptores do Ácido Retinoico/genética , Receptores do Ácido Retinoico/metabolismo , Receptores X Retinoide/genética , Receptores X Retinoide/metabolismo , Proteínas Cotransportadoras de Sódio-Fosfato Tipo IIb/metabolismo
14.
Development ; 147(4)2020 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-32001436

RESUMO

Proper organ development depends on coordinated communication between multiple cell types. Retinoic acid (RA) is an autocrine and paracrine signaling molecule essential for the development of most organs, including the lung. Despite extensive work detailing effects of RA deficiency in early lung morphogenesis, little is known about how RA regulates late gestational lung maturation. Here, we investigate the role of the RA catabolizing protein Cyp26b1 in the lung. Cyp26b1 is highly enriched in lung endothelial cells (ECs) throughout development. We find that loss of Cyp26b1 leads to reduction of alveolar type 1 cells, failure of alveolar inflation and early postnatal lethality in mouse. Furthermore, we observe expansion of distal epithelial progenitors, but no appreciable changes in proximal airways, ECs or stromal populations. Exogenous administration of RA during late gestation partially mimics these defects; however, transcriptional analyses comparing Cyp26b1-/- with RA-treated lungs reveal overlapping, but distinct, responses. These data suggest that defects observed in Cyp26b1-/- lungs are caused by both RA-dependent and RA-independent mechanisms. This work reports crucial cellular crosstalk during lung development involving Cyp26b1-expressing endothelium and identifies a novel RA modulator in lung development.


Assuntos
Epitélio/embriologia , Pulmão/embriologia , Alvéolos Pulmonares/embriologia , Ácido Retinoico 4 Hidroxilase/genética , Ácido Retinoico 4 Hidroxilase/fisiologia , Animais , Sistemas CRISPR-Cas , Diferenciação Celular , Células Endoteliais/citologia , Células Epiteliais/citologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Rim/embriologia , Camundongos , Camundongos Endogâmicos C57BL , Organogênese/efeitos dos fármacos , Gravidez , Prenhez , Transdução de Sinais , Células-Tronco/citologia , Tretinoína/farmacologia
15.
J Exp Clin Cancer Res ; 39(1): 41, 2020 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-32087738

RESUMO

BACKGROUND: The oncogene MYCN is critical for tumorigenesis of several types of cancers including neuroblastoma. We previously reported that miR-506-3p repressed MYCN expression in neuroblastoma cells. However, the mechanism underlying such regulation was undetermined since there is no miR-506-3p target site in MYCN 3'UTR. METHODS: By a systematic investigation combining microarray, informatics and luciferase reporter assay, we identified that the transcriptional factor pleiomorphic adenoma gene-like 2 (PLAGL2) is a direct target of miR-506-3p that mediates its regulation on MYCN expression. Using CHIP-PCR and luciferase reporter assay, we validated the transcriptional regulation of MYCN by PLAGL2 and we further demonstrated the transcriptional regulation of PLAGL2 by MYCN. We examined the function of PLAGL2 in regulating neuroblastoma cell fate by cell viability assay, colony formation and Western blotting of differentiation markers. We examined the effect of retinoic acid, the differentiation agent used in neuroblastoma therapy, on miR-506-3p, PLAGL2 and MYCN expressions by quantitative PCR and Western blots. We investigated the clinical relevance of PLAGL2 expression by examining the correlation of tumor PLAGL2 mRNA levels with MYCN mRNA expression and patient survival using public neuroblastoma patient datasets. RESULTS: We found that miR-506-3p directly down-regulated PLAGL2 expression, and we validated a PLAGL2 binding site in the MYCN promoter region responsible for promoting MYCN transcription, thereby establishing a mechanism through which miR-506-3p regulates MYCN expression. Conversely, we discovered that MYCN regulated PLAGL2 transcription through five N-Myc-binding E-boxes in the PLAGL2 promoter region. We further confirmed the reciprocal regulation between endogenous PLAGL2 and MYCN in multiple neuroblastoma cell lines. Moreover, we found that PLAGL2 knockdown induced neuroblastoma cell differentiation and reduced cell proliferation, and combined knockdown of PLAGL2 and MYCN showed a synergistic effect. More strikingly, we found that high tumor PLAGL2 mRNA levels were significantly correlated with high MYCN mRNA levels and poor patient survival in neuroblastoma patients. Furthermore, we found that retinoic acid increased expression of miR-506-3p and repressed expression of MYCN and PLAGL2. CONCLUSIONS: Our findings altogether suggest that the interplay network formed by PLAGL2, MYCN and miR-506-3p is an important mechanism in regulating neuroblastoma cell fate, determining neuroblastoma prognosis, and mediating the therapeutic function of retinoic acid.


Assuntos
Proteínas de Ligação a DNA/genética , MicroRNAs/genética , Proteína Proto-Oncogênica N-Myc/genética , Neuroblastoma/genética , Proteínas de Ligação a RNA/genética , Fatores de Transcrição/genética , Regiões 3' não Traduzidas , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Proteínas de Ligação a DNA/metabolismo , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Proteína Proto-Oncogênica N-Myc/metabolismo , Neuroblastoma/metabolismo , Neuroblastoma/mortalidade , Regiões Promotoras Genéticas , Proteínas de Ligação a RNA/metabolismo , Análise de Sobrevida , Fatores de Transcrição/metabolismo , Tretinoína/farmacologia
16.
Chem Biol Interact ; 318: 108970, 2020 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-32007421

RESUMO

Cardiovascular disorders constitute the principal cause of deaths worldwide and will continue as the major disease-burden by the year 2060. A significant proportion of heart failures occur because of use and misuse of drugs and most of the investigational agents fail to achieve any clinical relevance. Here, we investigated rosuvastatin and retinoic acid for their "pharmacological pleiotropy" against high dose ß-adrenergic agonist (isoproterenol)-induced acute myocardial insult. Rats were pretreated with rosuvastatin and/or retinoic acid for seven days and the myocardial injury was induced by administering isoproterenol on the seventh and eighth day. After induction, rats were anaesthetized for electrocardiography, then sacrificed and different samples were collected/stored for various downstream assays. Myocardial injury with isoproterenol resulted in increased cardiac mass, decreased R-wave amplitude, increased QRS and QT durations; elevated levels of cardiac markers like cTnI, CK-MB, ALT and AST; increased lipid peroxidation, protein carbonylation and tissue nitric oxide levels; decreased endogenous antioxidants like SOD, CAT, GR, GST, GPx and total antioxidant activity; increased inflammatory markers like TNF-α and IL-6; decreased the mRNA expression of Nrf2 and Bcl-2; increased the mRNA expression of Bax, eNOS and iNOS genes. Pretreatment with rosuvastatin and/or retinoic acid mitigated many of the above biochemical and pathological alterations. Our results demonstrate that rosuvastatin and retinoic acid exert cardioprotective effects and may act as potential agents in the prevention of ß-adrenergic agonist-induced acute myocardial injury in rats. Cardioprotective potential of rosuvastatin and retinoic acid could be attributed to their influence on the redox pathways, immunomodulation, membrane stability, Nrf2 preservation, iNOS and Bax expression levels. Thus, they may act directly or indirectly at various steps, the breakpoints, in the pathophysiological cascade responsible for cardiac injury. Our study gives insights about the pharmacological pleiotropism of rosuvastatin and retinoic acid.


Assuntos
Isoproterenol/toxicidade , Infarto do Miocárdio/induzido quimicamente , Infarto do Miocárdio/prevenção & controle , Rosuvastatina Cálcica/farmacologia , Transdução de Sinais/efeitos dos fármacos , Tretinoína/farmacologia , Agonistas Adrenérgicos beta/toxicidade , Animais , Anticolesterolemiantes/farmacologia , Antineoplásicos/uso terapêutico , Peso Corporal/efeitos dos fármacos , Coração/anatomia & histologia , Coração/efeitos dos fármacos , Masculino , Tamanho do Órgão , Distribuição Aleatória , Ratos , Ratos Wistar
17.
Macromol Rapid Commun ; 41(5): e1900585, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32009277

RESUMO

3D liquid crystal elastomer (3D-LCE) foams are used to support long-term neuronal cultures for over 60 days. Sequential imaging shows that cell density remains relatively constant throughout the culture period while the number of cells per observational area increases. In a subset of samples, retinoic acid is used to stimulate extensive neuritic outgrowth and maturation of proliferated neurons within the LCEs, inducing a threefold increase in length with cells displaying morphologies indicative of mature neurons. Designed LCEs' micro-channels have a similar diameter to endogenous parenchymal arterioles, ensuring that neurons throughout the construct have constant access to growth media during extended experiments. Here it is shown that 3D-LCEs provide a unique environment and simple method to longitudinally study spatial neuronal function, not possible in conventional culture environments, with simplistic integration into existing methodological pipelines.


Assuntos
Materiais Biocompatíveis/química , Elastômeros/química , Cristais Líquidos/química , Neurônios/citologia , Tecidos Suporte/química , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Cristais Líquidos/ultraestrutura , Porosidade , Tretinoína/farmacologia
18.
PLoS One ; 15(2): e0228436, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32027669

RESUMO

BACKGROUND: Cytochrome P450 1b1 (Cyp1b1) deletion and dietary retinol deficiency during pregnancy (GVAD) affect perinatal liver functions regulated by Srebp. Cyp1b1 is not expressed in perinatal liver but appears in the E9.5 embryo, close to sites of retinoic acid (RA) signaling. HYPOTHESIS: Parallel effects of Cyp1b1 and retinol on postnatal Srebp derive from effects in the developing liver or systemic signaling. APPROACH: Cluster postnatal increases in hepatic genes in relation to effects of GVAD or Cyp1b1 deletion. Sort expression changes in relation to genes regulated by Srebp1 and Srebp2.Test these treatments on embryos at E9.5, examining changes at the site of liver initiation. Use in situ hybridization to resolve effects on mRNA distributions of Aldh1a2 and Cyp26a1 (RA homeostasis); Hoxb1 and Pax6 (RA targets). Assess mice lacking Lrat and Rbp4 (DKO mice) that severely limits retinol supply to embryos. RESULTS: At birth, GVAD and Cyp1b1 deletion stimulate gene markers of hepatic stellate cell (HSC) activation but also suppress Hamp. These treatments then selectively prevent the postnatal onset of genes that synthesize cholesterol (Hmgcr, Sqle) and fatty acids (Fasn, Scd1), but also direct cholesterol transport (Ldlr, Pcsk9, Stard4) and retinoid synthesis (Aldh1a1, Rdh11). Extensive support by Cyp1b1 is implicated, but with distinct GVAD interventions for Srebp1 and Srebp2. At E9.5, Cyp1b1 is expressed in the septum transversum mesenchyme (STM) with ß-carotene oxygenase (Bco1) that generates retinaldehyde. STM provides progenitors for the HSC and supports liver expansion. GVAD and Cyp1b1-/- do not affect RA-dependent Hoxb1 and Pax6. In DKO embryos, RA-dependent Cyp26a1 is lost but Hoxb1 is sustained with Cyp1b1 at multiple sites. CONCLUSION: Cyp1b1-/- suppresses genes supported by Srebp. GVAD effects distinguish Srebp1 and Srebp2 mediation. Srebp regulation overlaps appreciably in cholesterol and retinoid homeostasis. Bco1/Cyp1b1 partnership in the STM may contribute to this later liver regulation.


Assuntos
Colesterol/biossíntese , Citocromo P-450 CYP1B1/fisiologia , Desenvolvimento Fetal , Fígado/metabolismo , Proteínas de Ligação a Elemento Regulador de Esterol/fisiologia , Tretinoína/metabolismo , Animais , Animais Recém-Nascidos , Citocromo P-450 CYP1B1/genética , Embrião de Mamíferos , Feminino , Desenvolvimento Fetal/efeitos dos fármacos , Desenvolvimento Fetal/genética , Fígado/efeitos dos fármacos , Fígado/embriologia , Fígado/crescimento & desenvolvimento , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Gravidez , Proteínas Plasmáticas de Ligação ao Retinol/genética , Proteínas Plasmáticas de Ligação ao Retinol/metabolismo , Tretinoína/farmacologia
19.
Sci Adv ; 6(2): eaaw2746, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31934619

RESUMO

Creating complex multicellular kidney organoids from pluripotent stem cells shows great promise. Further improvements in differentiation outcomes, patterning, and maturation of specific cell types are, however, intrinsically limited by standard tissue culture approaches. We describe a novel full factorial microbioreactor array-based methodology to achieve rapid interrogation and optimization of this complex multicellular differentiation process in a facile manner. We successfully recapitulate early kidney tissue patterning events, exploring more than 1000 unique conditions in an unbiased and quantitative manner, and define new media combinations that achieve near-pure renal cell type specification. Single-cell resolution identification of distinct renal cell types within multilayered kidney organoids, coupled with multivariate analysis, defined the definitive roles of Wnt, fibroblast growth factor, and bone morphogenetic protein signaling in their specification, exposed retinoic acid as a minimal effector of nephron patterning, and highlighted critical contributions of induced paracrine factors on cell specification and patterning.


Assuntos
Rim/crescimento & desenvolvimento , Organoides/crescimento & desenvolvimento , Comunicação Parácrina , Perfusão , Células-Tronco Pluripotentes/citologia , Reatores Biológicos , Humanos , Rim/efeitos dos fármacos , Análise Multivariada , Néfrons/citologia , Néfrons/efeitos dos fármacos , Organoides/efeitos dos fármacos , Comunicação Parácrina/efeitos dos fármacos , Células-Tronco Pluripotentes/efeitos dos fármacos , Células-Tronco Pluripotentes/metabolismo , Tretinoína/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos
20.
Cell Death Dis ; 11(1): 66, 2020 01 27.
Artigo em Inglês | MEDLINE | ID: mdl-31988297

RESUMO

Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related deaths worldwide due to its high rate of recurrence, in part because of cancer stem cell (CSC)-dependent "field cancerization". Recently, we identified that the oncogene v-myc avian myelocytomatosis viral oncogene neuroblastoma derived homolog (MYCN) marked CSC-like subpopulations in heterogeneous HCC and served as a therapeutic target and prognostic marker for HCC. In this study, we explored the molecular basis of upregulated MYCN gene expression in HCC cells. Liquid chromatograph time-of-flight mass spectrometry-based metabolome analysis demonstrated that the content of unsaturated fatty acids was increased in MYCN high expression (MYCNhigh) CSC-like HCC cells. Inhibition of lipid desaturation using either the chemical inhibitor or siRNA/shRNA against stearoyl-CoA desaturase-1 (SCD1) suppressed cell proliferation as well as MYCN gene expression in MYCNhigh HCC cells, grown as both monolayer and spheres. Further mechanistic study using RNA-seq based transcriptome analysis revealed that endoplasmic reticulum (ER) stress related signaling networks such as endocannabinoid cancer inhibition pathway were under the control of SCD1 in MYCNhigh HCC cells. Furthermore, the expression of ER stress-inducible transcription suppressor cyclic AMP-dependent transcription factor (ATF3) was downregulated in MYCNhigh CSC-like HCC cells and CSC-rich spheroids, which was upregulated by inhibition of lipid desaturation or treatment with acyclic retinoid (ACR). Lipid profiling using NMR spectroscopy revealed that the ACR dramatically reduced the content of unsaturated fatty acids in HCC cells. The chemical inducer of ER stress inhibited MYCN gene expression, while the chemical inhibitor of ER stress or knockdown of ATF3 gene expression partially rescued the suppression of MYCN gene expression by ACR in MYCNhigh HCC cells. These data suggested that lipid desaturation-mediated ER stress signaling regulates MYCN gene expression in HCC cells and serves as a promising therapeutic target for the treatment and prevention of HCC.


Assuntos
Carcinoma Hepatocelular/metabolismo , Estresse do Retículo Endoplasmático/genética , Ácidos Graxos Insaturados/metabolismo , Neoplasias Hepáticas/metabolismo , Proteína Proto-Oncogênica N-Myc/metabolismo , Fator 3 Ativador da Transcrição/genética , Fator 3 Ativador da Transcrição/metabolismo , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Cromatografia Líquida , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Ácidos Graxos Insaturados/química , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Neoplasias Hepáticas/genética , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Metaboloma , Proteína Proto-Oncogênica N-Myc/genética , Células-Tronco Neoplásicas/metabolismo , Organoides/efeitos dos fármacos , Organoides/metabolismo , RNA Interferente Pequeno , RNA-Seq , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Estearoil-CoA Dessaturase/antagonistas & inibidores , Tretinoína/análogos & derivados , Tretinoína/metabolismo , Tretinoína/farmacologia
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