RESUMEN
Epidermal hyperinnervation is a critical feature of pruritus during skin inflammation. However, the mechanisms underlying epidermal hyperinnervation are unclear. This study investigates the role of the transcription factor EGR1 in epidermal innervation by utilizing wild-type (Egr1+/+) and Egr1-null (Egr1â/â) mice topically applied Dermatophagoides farinae extract from dust mite. Our findings revealed that Egr1â/â mice exhibited reduced scratching behaviors and decreased density of epidermal innervation compared with Egr1+/+ mice. Furthermore, we identified artemin, a neurotrophic factor, as an EGR1 target responsible for Dermatophagoides farinae extract-induced hyperinnervation. It has been demonstrated that Dermatophagoides farinae extract stimulates toll-like receptors in keratinocytes. To elucidate the cellular mechanism, we stimulated keratinocytes with Pam3CSK4, a toll-like receptor 1/2 ligand. Pam3CSK4 triggered a toll-like receptor 1/2-mediated signaling cascade involving IRAK4, IκB kinase, MAPKs, ELK1, EGR1, and artemin, leading to increased neurite outgrowth and neuronal migration. In addition, increased expression of EGR1 and artemin was observed in the skin tissues of patients with atopic dermatitis. These findings highlight the significance of the EGR1-artemin axis in keratinocytes, promoting the process of epidermal innervation and suggesting it as a potential therapeutic target for alleviating itch and pain associated with house dust mite-induced skin inflammation.
Asunto(s)
Proteína 1 de la Respuesta de Crecimiento Precoz , Epidermis , Queratinocitos , Proteínas del Tejido Nervioso , Células Receptoras Sensoriales , Animales , Queratinocitos/metabolismo , Ratones , Proteínas del Tejido Nervioso/metabolismo , Epidermis/inervación , Epidermis/metabolismo , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismo , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Células Receptoras Sensoriales/metabolismo , Dermatophagoides farinae/inmunología , Prurito/inmunología , Prurito/etiología , Prurito/metabolismo , Modelos Animales de Enfermedad , Humanos , Antígenos Dermatofagoides/inmunología , Transducción de Señal , Ratones Noqueados , Masculino , Dermatitis Atópica/inmunología , Dermatitis Atópica/metabolismo , Dermatitis Atópica/patologíaRESUMEN
Bone marrow-derived mesenchymal stem cells (BM-MSCs) can differentiate into endothelial cells in an inflammatory microenvironment. However, the regulatory mechanisms underlying this process are not entirely understood. Here, we found that TIE2 in BM-MSCs was upregulated at the transcriptional level after stimulation with tumor necrosis factor-alpha (TNFα), a major pro-inflammatory cytokine. Additionally, the STAT-binding sequence within the proximal region of TIE2 was necessary for TNFα-induced TIE2 promoter activation. TIE2 and STAT3 knockdown reduced TNFα-induced endothelial tube formation in BMMSCs. Among the major TNFα-activated MAP kinases (ERK1/2, JNK1/2, and p38 MAPK) in BM-MSCs, only inhibition of the p38 kinase abrogated TNFα-induced TIE2 upregulation by inhibiting the JAK-STAT signaling pathway. These findings suggest that p38 MAP contributes to the endothelial differentiation of BM-MSCs by activating the JAK-STAT-TIE2 signaling axis in the inflammatory microenvironment.
RESUMEN
Vasculogenic mimicry (VM) is an intriguing phenomenon observed in tumor masses, in which cancer cells organize themselves into capillary-like channels that closely resemble the structure and function of blood vessels. Although VM is believed to contribute to alternative tumor vascularization, the detailed regulatory mechanisms controlling these cellular processes remain poorly understood. Our study aimed to investigate the role of Early Growth Response 1 (EGR1) in regulating VM in aggressive cancer cells, specifically MDA-MB-231 triple-negative breast cancer cells. Our study revealed that EGR1 promotes the formation of capillary-like tubes by MDA-MB-231 cells in a 3-dimensional Matrigel matrix. EGR1 was observed to upregulate Kruppel-like factor 4 (KLF4) expression, which regulates the formation of the capillary-like tube structure. Additionally, our findings highlight the involvement of the ERK1/2 and p38 mitogen-activated protein kinase pathways in mediating the expression of EGR1 and KLF4, underscoring their crucial role in VM in MDA-MB-231 cells. Understanding these regulatory mechanisms will provide valuable insights into potential therapeutic targets for preventing VM during the treatment of triple-negative breast cancer.
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Neoplasias de la Mama Triple Negativas , Humanos , Línea Celular , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Factor 4 Similar a Kruppel , Activación Transcripcional , Neoplasias de la Mama Triple Negativas/genética , Regulación hacia ArribaRESUMEN
INTRODUCTION: Tubulin polymerization inhibitors induce cancer cell death; therefore, they can be developed as chemotherapeutic agents. We hypothesized that hybrid compounds, including the trans-stilbene moiety contained in resveratrol and penta-1,4-dien-3-one contained in curcumin, could inhibit tubulin polymerization. METHODS: Twenty-six hybrid stilbene and pentadienone compounds were designed and synthesized. The cytotoxicity of the hybrid compounds against MDA-MB-231 human breast cancer cells was determined using a clonogenic long-term survival assay. The relationship between cytotoxicity and structural properties was evaluated. Biological activities, including inhibition of tubulin polymerization and cell cycle progression, were investigated to select compounds with excellent anticancer properties. The molecular binding mode between the selected compound and the α, ß-tubulin dimers was investigated. RESULTS: Twenty-six hybrid stilbene and pentadienone compounds were designed and synthesized. Among them, compound 13 exhibited the highest inhibitory effect on the clonogenicity of MDA-MB-231 cells. Compound 13 induced the destabilization of tubulins and inhibited cell cycle progression at the G2/M phase. Through in silico molecular docking analysis, compound 13 was predicted to bind to the colchicine binding site of α, ß-tubulin. CONCLUSION: The stilbene and pentadienone hybrid compound 13 has a binding mode similar to that of colchicine. Compound 13 inhibited the clonogenicity of MDA-MB-231 cells through a mechanism that destabilizes tubulin polymerization, leading to cell cycle arrest at the G2/M phase.
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Antineoplásicos , Estilbenos , Humanos , Relación Estructura-Actividad , Tubulina (Proteína)/metabolismo , Proliferación Celular , Polimerizacion , Simulación del Acoplamiento Molecular , Estilbenos/farmacología , Ensayos de Selección de Medicamentos Antitumorales , Moduladores de Tubulina/farmacología , Moduladores de Tubulina/química , Colchicina/farmacología , Antineoplásicos/farmacología , Antineoplásicos/químicaRESUMEN
Mesenchymal stem cells (MSCs) can differentiate into endothelial cells; however, the mechanisms underlying this process in the tumor microenvironment (TME) remain elusive. This study shows that tumor necrosis factor alpha (TNF-α), a key cytokine present in the TME, promotes the endothelial differentiation of MSCs by inducing vascular endothelial growth factor receptor 2 (VEGFR2) gene expression. EGR1 is a member of the zinc-finger transcription factor family induced by TNF-α. Our findings indicate that EGR1 directly binds to the VEGFR2 promoter and transactivates VEGFR2 expression. We also demonstrate that EGR1 forms a complex with c-JUN activated by c-JUN N-terminal kinase (JNK) to promote VEGFR2 transcription and endothelial differentiation in MSCs in response to TNF-α stimulation. The shRNA-mediated silencing of EGR1 or c-JUN abrogates TNF-α-induced VEGFR2 transcription and the endothelial differentiation of MSCs. To further evaluated the role of EGR1 in the endothelial differentiation of BM-MSCs, we used a syngenic tumor implantation model. 4T1 mouse mammary tumor cells were injected subcutaneously into BALB/c mice with primary mBM-MSCs isolated from wild-type (Egr1+/+) or Egr1-null (Egr1-/-) mice. CD31-positive cells were predominantly observed at the border of the tumor in the 4T1 plus wild-type MSC group, while staining less in the 4T1 alone or 4T1 plus Egr1-null MSC group. Collectively, these findings demonstrate that the JNK-EGR1 signaling axis plays a crucial role in the TNF-α-induced endothelial differentiation of MSCs in the TME, which could be a potential therapeutic target for solid tumors vasculatures.
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Células Madre Mesenquimatosas , Factor de Necrosis Tumoral alfa , Humanos , Ratones , Animales , Factor de Necrosis Tumoral alfa/farmacología , Factor de Necrosis Tumoral alfa/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Células Endoteliales/metabolismo , Diferenciación Celular , Células Madre Mesenquimatosas/metabolismo , Células Cultivadas , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismoRESUMEN
Prospero homeobox 1 (PROX1) is a member of the homeobox transcription factor family that plays a critical role in the development of multiple tissues and specification of cell fate. PROX1 expression is differentially regulated based on the cellular context and plays an antagonistic role as a tumour promoter or suppressor in different tumour types. In human breast cancer, PROX1 expression is suppress-ed; however, the molecular mechanism by which it is down-regulated remains poorly understood. Here, we show that ectopic expression of PROX1 reduces the motility and invasiveness of MDA-MB-231 human breast cancer cells, suggesting that PROX1 functions as a negative regulator of tumour invasion in MDA-MB-231 cells. Treatment with histone deacetylase (HDAC) inhibitors up-regulates PROX1 mRNA and protein expression levels. Knockdown of HDAC1 using short hairpin RNA also up-regulates PROX1 mRNA and protein expression levels. We found that HDAC1 interacted with c-JUN at the activator protein (AP)-1-binding site located at -734 to -710 in the PROX1 promoter region to suppress PROX1 expression. In addition, c-JUN N-terminal kinase-mediated c-JUN phosphorylation was found to be crucial for silencing PROX1 expression. In conclusion, PROX1 expression can be silenced by the epigenetic mechanism involved in the complex formation of HDAC1 and c-JUN at the AP-1 site in the PROX1 promoter region in MDA-MB-231 human breast cancer cells. Therefore, this study revealed the epigenetic regulatory mechanism involved in the suppression of PROX1 expression in breast cancer cells.
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Neoplasias de la Mama , Proteínas de Homeodominio , Femenino , Humanos , Neoplasias de la Mama/genética , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Genes Homeobox , Histona Desacetilasa 1/genética , Histona Desacetilasa 1/metabolismo , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Células MDA-MB-231 , ARN Mensajero/genética , Factores de Transcripción/genéticaRESUMEN
Atopic dermatitis (AD) is one of the most common inflammatory skin diseases accompanied by severe itching. ß-caryophyllene (BCP), which displays anti-inflammatory activity, is a natural agonist of cannabinoid receptor 2. However, the therapeutic effects of BCP on atopic dermatitis (AD) remain poorly understood. The current study aimed to evaluate the topical therapeutic efficacy of BCP in an AD-like mouse model. Thymic Stromal Lymphopoietin (TSLP) is a keratinocyte-derived cytokine that drives AD pathogenesis. This study also investigated the effect of BCP on the interleukin 4 (IL-4)-induced expression of TSLP in HaCaT keratinocytes. We found that the topical application of BCP alleviated AD-like skin inflammation and inhibited the infiltration of proinflammatory cells into skin lesions. Moreover, the topical application of BCP reduced EGR1 (Early Growth Response 1) and TSLP expression in AD-like skin lesions. We also found that BCP inhibited IL-4-induced TSLP expression by downregulating mitogen-activated protein kinase (MAPK)-mediated EGR1 expression in HaCaT keratinocytes. These findings demonstrate that BCP ameliorates DNCB-induced AD-like skin lesions through the downregulation of the MAPK/EGR1/TSLP signaling axis. BCP may be applicable for developing topical therapeutic agents for chronic skin inflammatory diseases, such as AD.
Asunto(s)
Dermatitis Atópica , Ratones , Animales , Dermatitis Atópica/inducido químicamente , Dermatitis Atópica/tratamiento farmacológico , Dermatitis Atópica/metabolismo , Dinitroclorobenceno , Interleucina-4/metabolismo , Linfopoyetina del Estroma Tímico , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Citocinas/metabolismo , Queratinocitos/metabolismo , Piel/metabolismo , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismoRESUMEN
Filaggrin (FLG), an essential structural protein for skin barrier function, is down-regulated under chronic inflammatory conditions, leading to disruption of the skin barrier. However, the detailed molecular mechanisms of how FLG changes in the context of chronic inflammation are poorly understood. Here, we identified the molecular mechanisms by which inflammatory cytokines inhibit FLG expression in the skin. We found that the AP1 response element within the -343/+25 of the FLG promoter was necessary for TNFα + IFNγ-induced down-regulation of FLG promoter activity. Using DNA affinity precipitation assay, we observed that AP1 subunit composition binding to the FLG promoter was altered from c-FOS:c-JUN (at the early time) to FRA1:c-JUN (at the late time) in response to TNFα + IFNγ stimulation. Knockdown of FRA1 or c-JUN abrogated TNFα + IFNγ-induced FLG suppression. Histone deacetylase (HDAC) 1 interacted with FRA1:c-JUN under TNFα + IFNγ stimulation. Knockdown of HDAC1 abrogated the inhibitory effect of TNFα + IFNγ on FLG expression. The altered expression of FLG, FRA1, c-JUN, and HDAC1 was confirmed in mouse models of 2,4-dinitrochlorobenzene-induced atopic dermatitis and imiquimod-induced psoriasis. Thus, the current study demonstrates that TNFα + IFNγ stimulation suppresses FLG expression by promoting the FRA1:c-JUN:HDAC1 complex. This study provides insight into future therapeutic strategies targeting the FRA1:c-JUN:HDAC1 complex to restore impaired FLG expression in chronic skin inflammation.
Asunto(s)
Proteínas Filagrina , Histona Desacetilasa 1 , Queratinocitos , Proteínas Proto-Oncogénicas c-fos , Proteínas Proto-Oncogénicas c-jun , Animales , Enfermedad Crónica , Dermatitis/genética , Dermatitis/metabolismo , Regulación hacia Abajo , Proteínas Filagrina/genética , Proteínas Filagrina/metabolismo , Histona Desacetilasa 1/genética , Histona Desacetilasa 1/metabolismo , Interferón gamma/farmacología , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Ratones , Complejos Multiproteicos/genética , Complejos Multiproteicos/metabolismo , Proteínas Proto-Oncogénicas c-fos/genética , Proteínas Proto-Oncogénicas c-fos/metabolismo , Proteínas Proto-Oncogénicas c-jun/genética , Proteínas Proto-Oncogénicas c-jun/metabolismo , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
Atopic dermatitis (AD) is one of the most common chronic inflammatory skin diseases worldwide, characterized by intense pruritus and eczematous lesions. Aberrant expression of thymic stromal lymphopoietin (TSLP) in keratinocytes is associated with the pathogenesis of AD and is considered a therapeutic target for the treatment of this disease. Saikosaponin A (SSA) and saikosaponin C (SSC), identified from Radix Bupleuri, exert anti-inflammatory effects. However, the topical effects of SSA and SSC on chronic inflammatory skin diseases are unclear. In this study, we investigated the effects of SSA and SSC on TSLP suppression in an AD-like inflammatory environment. We observed that SSA and SSC suppressed tumor necrosis factor-α-induced TSLP expression by downregulating the expression of the transcription factor early growth response 1 (EGR1) via inhibition of the extracellular signal-regulated kinase 1/2, c-Jun N-terminal kinase 1/2, and p38 mitogen-activated protein kinase pathways. We also confirmed that topical application of SSA or SSC reduced AD-like skin lesions in BALB/c mice challenged with 2,4-dinitrochlorobenzene. Our findings suggest that suppression of EGR1-regulated TSLP expression in keratinocytes might be attributable to the anti-inflammatory effects of SSA and SSC in AD-like skin lesions.
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Dermatitis Atópica , Enfermedades de la Piel , Animales , Antiinflamatorios/farmacología , Citocinas/metabolismo , Dermatitis Atópica/inducido químicamente , Dermatitis Atópica/tratamiento farmacológico , Dermatitis Atópica/metabolismo , Proteína 1 de la Respuesta de Crecimiento Precoz/antagonistas & inhibidores , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismo , Células HaCaT , Humanos , Queratinocitos/metabolismo , Ratones , Ácido Oleanólico/análogos & derivados , Saponinas , Enfermedades de la Piel/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Linfopoyetina del Estroma TímicoRESUMEN
PER2 is a core circadian clock gene that regulates circadian rhythms. IL-4 plays a critical role in the pathogenesis of skin inflammation, including atopic dermatitis. IL-4 enhances PER2 expression, suggesting a relationship between inflammation and the circadian clock. However, little is known about the molecular and cellular mechanisms regulating PER2 expression by inflammatory cytokines. This study showed that transcription factor EGR1 interacted with the PER2 promoter and promoted IL-4âinduced transcriptional activation of the PER2, as revealed by promoterâreporter assay, electrophoretic mobility shift assay, DNA affinity precipitation assay, and chromatin immunoprecipitation analysis. We also found that IL-4 can use both MAPK and Jak signaling pathways to induce EGR1-mediated PER2 expression, and c-Jun N-terminal kinase 1/2 can augment IL-4âinduced activation of the Jakâsignal transducer and activator of transcription 3 pathway. Consistently, Per2 expression was reduced in dinitrochlorobenzene-induced atopic dermatitisâlike skin lesions in Egr1â/â mice compared with that in Egr1+/+ mice. In addition, using a real-time bioluminescence assay, we observed that EGR1 is required for rhythmic oscillation of PER2 expression under IL-4 exposure. These findings provide further insight into the role of EGR1 in regulating PER2 expression in impaired circadian rhythm in skin inflammation.
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Dermatitis Atópica , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismo , Interleucina-4/metabolismo , Proteínas Circadianas Period , Animales , Ritmo Circadiano/fisiología , ADN/genética , Dermatitis Atópica/genética , Dinitroclorobenceno , Humanos , Inflamación , Queratinocitos/metabolismo , Ratones , Proteína Quinasa 8 Activada por Mitógenos/metabolismo , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Factor de Transcripción STAT3/metabolismoRESUMEN
SignificanceSimilar to mammalian TLR4/MD-2, the Toll9/MD-2-like protein complex in the silkworm, Bombyx mori, acts as an innate pattern-recognition receptor that recognizes lipopolysaccharide (LPS) and induces LPS-stimulated expression of antimicrobial peptides such as cecropins. Here, we report that papiliocin, a cecropin-like insect antimicrobial peptide from the swallowtail butterfly, competitively inhibits the LPS-TLR4/MD-2 interaction by directly binding to human TLR4/MD-2. Structural elements in papiliocin, which are important in inhibiting TLR4 signaling via direct binding, are highly conserved among insect cecropins, indicating that its TLR4-antagonistic activity may be related to insect Toll9-mediated immune response against microbial infection. This study highlights the potential of papiliocin as a potent TLR4 antagonist and safe peptide antibiotic for treating gram-negative sepsis.
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Antiinfecciosos Locales/farmacología , Péptidos Antimicrobianos/farmacología , Mariposas Diurnas/inmunología , Inmunidad Innata/efectos de los fármacos , Proteínas de Insectos/farmacología , Receptor Toll-Like 4/antagonistas & inhibidores , Animales , Antiinfecciosos Locales/química , Péptidos Antimicrobianos/química , Péptidos Antimicrobianos/metabolismo , Infecciones por Escherichia coli/tratamiento farmacológico , Femenino , Proteínas de Insectos/química , Proteínas de Insectos/metabolismo , Lipopolisacáridos/metabolismo , Ratones , Ratones Endogámicos ICR , Simulación del Acoplamiento Molecular , Unión Proteica , Conformación Proteica , Receptor Toll-Like 4/metabolismoRESUMEN
Novel (Z)-3-((4,6-diphenylpyrimidin-2-ylamino)methylene)-2,3-dihydrochromen-4-one derivatives were designed and synthesized to find chemotherapeutic agents. Derivative 9 was selected based on its clonogenicity against cancer cells and synthetic yield for further biological experiments. It showed decreases in aurora kinase A, B, and C phosphorylation from western blot analysis. Derivative 9 upregulated the expression of G1 cell cycle inhibitory proteins including p21 and p27, and G1 progressive cyclin D1, and downregulated G1-to-S progressive cyclins, resulting in cell cycle arrest at the G1/S boundary. It stimulated the cleavage of caspase-9, -3, -7, and poly (ADP-ribose) polymerase, resulting in triggering apoptosis through a caspase-dependent pathway. In addition, derivative 9 inhibited in vivo tumor growth in a syngeneic tumor implantation mouse model. The findings of this study suggest that derivative 9 can be considered as a lead compound for chemotherapeutic agents.
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Antineoplásicos , Caspasas , Animales , Antineoplásicos/farmacología , Apoptosis , Caspasas/metabolismo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/farmacología , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/farmacología , Ratones , Poli(ADP-Ribosa) Polimerasas/metabolismoRESUMEN
Thymic stromal lymphopoietin (TSLP) is an epithelial cell-derived cytokine that acts as a critical mediator in the pathogenesis of atopic dermatitis (AD). Various therapeutic agents that prevent TSLP function can efficiently relieve the clinical symptoms of AD. However, the downregulation of TSLP expression by therapeutic agents remains poorly understood. In this study, we investigated the mode of action of chrysin in TSLP suppression in an AD-like inflammatory environment. We observed that the transcription factor early growth response (EGR1) contributed to the tumor necrosis factor alpha (TNFα)-induced transcription of TSLP. Chrysin attenuated TNFα-induced TSLP expression by downregulating EGR1 expression in HaCaT keratinocytes. We also showed that the oral administration of chrysin improved AD-like skin lesions in the ear and neck of BALB/c mice challenged with 2,4-dinitrochlorobenzene. We also showed that chrysin suppressed the expression of EGR1 and TSLP by inhibiting the extracellular signal-regulated kinase (ERK) 1/2 and c-Jun N-terminal kinase (JNK) 1/2 mitogen-activated protein kinase pathways. Collectively, the findings of this study suggest that chrysin improves AD-like skin lesions, at least in part, through the downregulation of the ERK1/2 or JNK1/2-EGR1-TSLP signaling axis in keratinocytes.
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Citocinas/metabolismo , Proteína 1 de la Respuesta de Crecimiento Precoz/antagonistas & inhibidores , Flavonoides/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Queratinocitos/efectos de los fármacos , Enfermedades de la Piel/tratamiento farmacológico , Factor de Necrosis Tumoral alfa/farmacología , Animales , Células Cultivadas , Citocinas/genética , Dinitroclorobenceno/toxicidad , Humanos , Queratinocitos/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Enfermedades de la Piel/inducido químicamente , Enfermedades de la Piel/metabolismo , Enfermedades de la Piel/patología , Linfopoyetina del Estroma TímicoAsunto(s)
Antiinflamatorios/farmacología , ADN/metabolismo , Dermatitis Atópica/tratamiento farmacológico , Proteína 1 de la Respuesta de Crecimiento Precoz/antagonistas & inhibidores , Animales , Antiinflamatorios/uso terapéutico , Dermatitis Atópica/inducido químicamente , Dermatitis Atópica/inmunología , Dermatitis Atópica/patología , Dinitroclorobenceno/administración & dosificación , Dinitroclorobenceno/toxicidad , Modelos Animales de Enfermedad , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismo , Células HaCaT , Células Endoteliales de la Vena Umbilical Humana , Humanos , Ratones , Ratones Noqueados , Simulación del Acoplamiento Molecular , Unión Proteica/efectos de los fármacos , Células THP-1 , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Matrix metalloproteinase 1 (MMP-1) initiates the breakdown of matrix networks by cleaving fibrillar collagen during the pathophysiological progression of skin aging. Ageratum houstonianum ethanol extract (AHE) has been used as a traditional herbal medicine to treat external wounds and skin diseases. However, the mechanism of action underlying A. houstonianum-mediated modulation of skin aging has not been investigated. In this study, we evaluated the effect of AHE on MMP-1 expression in HaCaT keratinocytes. Gene expression was analyzed by Reverse transcription-PCR (RT-PCR), Quantitative real-time PCR (Q-PCR), gene promoter-reporter assay, and immunoblotting. We found that AHE abrogated TNFα-induced MMP1 expression at the transcriptional level via the suppression of ERK1/2 mitogen-activated protein kinase (MAPK)-mediated Early Growth Response 1 (EGR1) expression. We also demonstrated that ß-caryophyllene, a cannabinoid receptor 2 (CB2) agonist, is a functional component of the AHE that inhibits TNFα-induced EGR-1 and MMP1 expression. AHE exerts inhibitory activity on TNFα-induced MMP1 expression at the transcription level through EGR-1 downregulation in keratinocytes. ß-Caryophyllene is a bioactive ingredient of AHE that is responsible for the inhibition of TNFα-induced EGR1 expression. ß-Caryophyllene can be used as a potential agent to prevent inflammation-induced skin aging.
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Ageratum/química , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Metaloproteinasa 1 de la Matriz/genética , Extractos Vegetales/farmacología , Envejecimiento de la Piel/efectos de los fármacos , Proteína 1 de la Respuesta de Crecimiento Precoz/antagonistas & inhibidores , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Inflamación/tratamiento farmacológico , Inflamación/genética , Inflamación/patología , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Queratinocitos/patología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Extractos Vegetales/química , Sesquiterpenos Policíclicos/farmacología , Envejecimiento de la Piel/patología , Transcripción Genética/efectos de los fármacos , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
Kallikrein-related peptidase 7 (KLK7) is a chymotrypsin-like serine peptidase that plays a crucial role in regulating skin desquamation. KLK7 expression is highly upregulated in atopic dermatitis (AD) skin lesions in both humans and mice. Th2-lymphocyte-derived cytokines, including interleukin (IL)-4 and IL-13, have been shown to promote KLK7 expression in keratinocytes in patients with AD. However, the molecular mechanism underlying KLK7 expression remains poorly understood. Here, we demonstrated that the EGR-1-binding sequence (EBS) in the promoter region of KLK7 played a crucial role in IL-13-induced KLK7 transcription. Disruption of the EBS induced by a point mutation inhibited IL-13-induced KLK7 promoter activity. EGR-1 was shown to directly bind to the EBS, and EGR1 knockdown with shRNA abrogated IL-13-induced KLK7 expression. Using Egr1 knockout mice, we showed that Egr-1 was necessary for KLK7 expression in AD-like lesions induced by the repeated topical application of 2,4-dinitrobenzene on the dorsal skin of mice. We also demonstrated that the ERK1/2 mitogen-activated protein kinase (MAPK) pathway was responsible for EGR-1-dependent KLK7 transcription in response to IL-13 stimulation. Our findings delineate a signaling pathway that contributes to the regulation of KLK7 expression through the IL13-ERK MAPK-EGR1 signaling axis.
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Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismo , Interleucina-13/metabolismo , Calicreínas/genética , Animales , Dermatitis Atópica/genética , Dermatitis Atópica/metabolismo , Dermatitis Atópica/patología , Modelos Animales de Enfermedad , Proteína 1 de la Respuesta de Crecimiento Precoz/antagonistas & inhibidores , Proteína 1 de la Respuesta de Crecimiento Precoz/deficiencia , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Técnicas de Silenciamiento del Gen , Células HaCaT , Humanos , Calicreínas/metabolismo , Queratinocitos/metabolismo , Queratinocitos/patología , Sistema de Señalización de MAP Quinasas , Ratones , Ratones Noqueados , Mutagénesis Sitio-Dirigida , Regiones Promotoras Genéticas , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Interferente Pequeño/genética , Transactivadores/antagonistas & inhibidores , Transactivadores/genética , Transactivadores/metabolismoRESUMEN
Estrogen overproduction is closely associated with the development of estrogen receptor-positive breast cancer. Aromatase, encoded by the cytochrome P450 19 (CYP19) gene, regulates estrogen biosynthesis. This study aimed to identify active flavones that inhibit CYP19 expression and to explore the underlying mechanisms. CYP19 expression was evaluated using reverse transcription PCR, quantitative real-time PCR, and immunoblot analysis. The role of transcription factor early growth response gene 1 (EGR-1) in CYP19 expression was assessed using the short-hairpin RNA (shRNA)-mediated knockdown of EGR-1 expression in estrogen receptor-positive MCF-7 breast cancer cells. We screened 39 flavonoids containing 26 flavones and 13 flavanones using the EGR1 promoter reporter activity assay and observed that chrysoeriol exerted the highest inhibitory activity on tumor necrosis factor alpha (TNFα)-induced EGR-1 expression. We further characterized and demonstrated that chrysoeriol inhibits TNFα-induced CYP19 expression through inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2)-mediated EGR-1 expression. Chrysoeriol may be beneficial as a dietary supplement for the prevention of estrogen receptor-positive breast cancer, or as a chemotherapeutic adjuvant in the treatment of this condition.
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Aromatasa/genética , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Flavonas/farmacología , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Factor de Necrosis Tumoral alfa/metabolismo , Aromatasa/metabolismo , Productos Biológicos/farmacología , Células Cultivadas , Ensayos de Selección de Medicamentos Antitumorales , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismo , Femenino , Flavonas/química , Silenciador del Gen , Humanos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Células MCF-7 , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
Chrysin (5,7-dihydroxyflavone) is a natural polyphenolic compound that induces an anti-inflammatory response. In this study, we investigated the molecular mechanism underlying the chrysin-induced suppression of C-C motif chemokine ligand 5 (CCL5) gene expression in atopic dermatitis (AD)-like inflammatory microenvironment. We showed that chrysin inhibited CCL5 expression at the transcriptional level through the suppression of nuclear factor kappa B (NF-κB) in the inflammatory environment. Chrysin could bind to the ATP-binding pocket of the inhibitor of κB (IκB) kinase (IKK) and, subsequently, prevent IκB degradation and NF-κB activation. The clinical efficacy of chrysin in targeting IKK was evaluated in 2,4-dinitrochlorobenzene-induced skin lesions in BALB/c mice. Our results suggested that chrysin prevented CCL5 expression by targeting IKK to reduce the infiltration of mast cells to the inflammatory sites and at least partially attenuate the inflammatory responses. These findings suggested that chrysin might be useful as a platform for the design and synthesis of small-molecule IKK-targeting drugs for the treatment of chronic inflammatory diseases, such as AD.
Asunto(s)
Quimiocina CCL5/genética , Dermatitis Atópica/genética , Flavonoides/farmacología , Quinasa I-kappa B/genética , Inflamación/tratamiento farmacológico , Animales , Microambiente Celular/efectos de los fármacos , Microambiente Celular/genética , Quimiocina CCL5/antagonistas & inhibidores , Dermatitis Atópica/patología , Flavonoides/química , Humanos , Inflamación/genética , Inflamación/patología , Mastocitos/efectos de los fármacos , Mastocitos/metabolismo , Ratones , FN-kappa B/genética , Factor de Necrosis Tumoral alfaRESUMEN
BACKGROUND: Reactive oxygen species (ROS) generation specifically in cancer cells may be a promising strategy for their selective killing. The synthetic chalcone derivative (E)-3-(3,5-dimethoxyphenyl)-1-(2-methoxyphenyl)prop-2-en-1-one (DPP23) exerts antitumor activity through ROS-mediated apoptosis in cancer cells but not in healthy cells. However, the mechanism underlying ROS generation by DPP23 remains unknown. OBJECTIVE: The current study aims to identify possible DPP23 target genes responsible for ROS generation through the mining of microarray data stored in NCBI's Gene Expression Omnibus (GEO). METHODS: A comprehensive expression profile of genes modulated by DPP23 was examined by gene ontology analysis. DPP23-modulated genes in Mia-PaCa2 pancreatic cells were validated by reverse transcription-PCR. RESULTS: Multiple genes were up and downregulated by DPP23 treatment in MiaPaCa2 pancreatic cancer cells. Genes with absolute fold-change (FC) of > 2 were selected as the cut-off criteria and grouped into 10 clusters to analyze expression patterns systematically. We observed that genes with increased expression at 6 h were significantly affected by ROS increase, unfolded protein response, and cell death. Expression of 13 genes involved in glutathione metabolism, including CHAC1, GCLC, G6PD, GSTO2, GSTA5, GSTM2, GSR, GPX3/6/8, GGT1, PGD, ATF4, and NAT8B, are modulated by DPP23. Of these, CHAC1 was most highly upregulated upon DPP23 treatment. CONCLUSION: DPP23 alters global gene expression associated with multiple cellular responses, including oxidative stress and apoptosis. We found that DPP23 may induce GSH depletion through modulation of gene expression, which is especially involved in glutathione metabolism. Of these, CHAC1 emerged as the most prominent candidate for DPP23 as it was the most responsive to DPP23 treatment.
Asunto(s)
Chalconas/farmacología , Neoplasias Pancreáticas/tratamiento farmacológico , Transcriptoma/genética , gamma-Glutamilciclotransferasa/genética , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Perfilación de la Expresión Génica , Humanos , Proteínas de Neoplasias/clasificación , Proteínas de Neoplasias/genética , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/genética , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Especies Reactivas de Oxígeno/metabolismo , Transcriptoma/efectos de los fármacosRESUMEN
WNT11 is a member of the non-canonical Wnt family and plays a crucial role in tumor progression. However, the regulatory mechanisms underlying WNT11 expression are unclear. Tumor necrosis factor-alpha (TNFα) is a major inflammatory cytokine produced in the tumor microenvironment and contributes to processes associated with tumor progression, such as tumor invasion and metastasis. By using site-directed mutagenesis and introducing a serial deletion in the 5'-regulatory region of WNT11, we observed that TNFα activates the early growth response 1 (EGR1)-binding sequence (EBS) in the proximal region of WNT11 and that the transcription factor EGR1 is necessary for the TNFα-induced transcription of WNT11. EGR1 bound directly to the EBSs within the proximal 5'-regulatory region of WNT11 and ectopic expression of EGR1 stimulated WNT11 promoter activity, whereas the knockdown of EGR1 expression by RNA interference reduced TNFα-induced WNT11 expression in T47D breast cancer cells. We also observed that mitogen-activated protein kinases (MAPK), extracellular signalregulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 kinase mediated TNFα-induced transcription of WNT11 via EGR1. Our results suggest that EGR1 directly targets WNT11 in response to TNFα stimulation in breast cancer cells. [BMB Reports 2020; 53(12): 628-633].