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1.
Mol Cell Neurosci ; : 103625, 2021 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-33933589

RESUMO

In a previous study, we demonstrated that neutrophil elastase is activated in the brain parenchyma after cerebral ischemia, which enzyme cleaves progranulin (PGRN), an anti-inflammatory factor. In that study, we also found that sivelestat, a selective neutrophil elastase inhibitor, attenuates ischemia-induced inflammatory responses. However, it was not clear whether this anti-inflammatory effect was due to the direct effect of sivelestat. In this study, we evaluated the effects of sivelestat or recombinant PGRN (rPGRN) on cell injuries in cultured neurons, astrocytes, and microglia under oxygen/glucose deprivation (OGD) conditions. We demonstrated that OGD-induced neuronal cell injury, astrocyte activation, and increased proinflammatory cytokines caused by microglial activation, were suppressed by rPGRN treatment, whereas sivelestat had no effect on any of these events. These results indicate that the anti-inflammatory responses after in vivo cerebral ischemia were not due to the direct action of sivelestat but due to the suppression of PGRN cleavage by inhibition of elastase activity. It was also suggested that the pleiotropic effect of rPGRN could be attributed to the differentiation of M1 microglia into anti-inflammatory type M2 microglia. Therefore, the inhibition of PGRN cleavage by sivelestat could contribute to the establishment of a new therapeutic approach for cerebral ischemia.

2.
Exp Cell Res ; 400(1): 112440, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33359470

RESUMO

Neurons require adhesive scaffolds for their growth and differentiation. Laminins are a major cell adhesive component of basement membranes and have various biological activities in the peripheral and central nervous systems. Here, we evaluated the biological activities of 5 peptides derived from laminin-111 as a scaffold for mouse neuroblastoma Neuro2a cells and rat neural stem/progenitor cells (NPCs). The 5 peptides showed Neuro2a cell attachment activity similar to that of poly-d-lysine. However, when NPCs were cultured on the peptides, 2 syndecan-binding peptides, AG73 (RKRLQVQLSIRT, mouse laminin α1 chain 2719-2730) and C16 (KAFDITYVRLKF, laminin γ1 chain 139-150), demonstrated significantly higher cell attachment and neurite extension activities than other peptides including integrin-binding ones. Long-term cell culture experiments showed that both AG73 and C16 supported the growth of neurons and astrocytes that had differentiated from NPCs. Furthermore, C16 markedly promoted the expression of neuronal markers such as synaptosomal-associated protein-25 and syntaxin 1A. These results indicate that AG73 and C16 are useful for NPC cultures and that C16 can be applied to specialized research on synapses in differentiated neurons. These peptides have the potential for use as valuable biomaterials for NPC research.

3.
Front Neurosci ; 14: 581915, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33177984

RESUMO

Cerebral ischemia induces neuronal cell death and causes various kinds of brain dysfunction. Therefore, prevention of neuronal cell death is most essential for protection of the brain. On the other hand, it has been reported that epigenetics including DNA methylation plays a pivotal role in pathogenesis of some diseases such as cancer. Accumulating evidences indicate that aberrant DNA methylation is related to cell death. However, DNA methylation after cerebral ischemia has not been fully understood yet. The aim of this present study was to investigate the relationships between DNA methylation and neuronal cell death after cerebral ischemia. We examined DNA methylation under the ischemic condition by using transient middle cerebral artery occlusion and reperfusion (MCAO/R) model rats and N-methyl-D-aspartate (NMDA)-treated cortical neurons in primary culture. In this study, we demonstrated that DNA methylation increased in these neurons 24 h after MCAO/R and that DNA methylation, possibly through activation of DNA methyltransferases (DNMT) 3a, increased in such neurons immediately after NMDA treatment. Furthermore, NMDA-treated neurons were protected by treatment with a DNMT inhibitor that were accompanied by inhibition of DNA methylation. Our results showed that DNA methylation would be an initiation factor of neuronal cell death and that inhibition of such methylation could become an effective therapeutic strategy for stroke.

4.
Front Pharmacol ; 11: 1087, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32765280

RESUMO

Novel therapeutic strategies for breast cancer are urgently needed due to the sustained development of drug resistance and tumor recurrence. Trivalent arsenic derivative (arsenite, AsIII) has been reported to induce cytotoxicity in breast cancer cells. We recently demonstrated that AsIII plus tetrandrine (Tetra), a Chinese plant-derived alkaloid, exerted potent antitumor activity against human breast cancer cells, however, the underlying mechanisms for their action have not been well defined. In order to provide fundamental insights for understanding the action of AsIII plus Tetra, the effects of the combined regimen on two breast cancer cell lines T47D and MDA-MB-231 were evaluated. Compared to T47D cells, MDA-MB-231 cells were much more susceptible to the synergistic cytotoxic effects of AsIII and Tetra. Besides the induction of apoptotic/necrotic cell death, S-phase arrest and autophagic cell death were also observed in MDA-MB-231 cells. Exposure of MDA-MB-231 cells to AsIII and Tetra caused the activation of MAPKs. Cytotoxicity of the combined regimen in MDA-MB-231 cell was significantly abrogated by SP600125, a potent c-Jun N-terminal kinase (JNK) inhibitor. However, similar abrogation was not caused by p38 and ERK inhibitors. The addition of either autophagy inhibitors (3-methyladenine or wortmannin) or SP600125 corrected the combined regimen-triggered S-phase arrest, whereas had little effect on the apoptosis/necrosis induction in the cells. Surprisingly, SP600125NC, a negative control for SP600125, significantly strengthened S-phase arrest and the cytotoxicity induced by the combined regimen. The addition of SP600125 did not alter autophagy induction. In conclusion, the cytotoxicity of AsIII combined with Tetra was attributed to the induction of S-phase arrest, apoptotic/necrotic and autophagic cell death. The enhanced cytotoxicity of the two drugs by SP600125NC might be explained by its capability to strengthen S-phase arrest. Our results suggested that JNK and autophagy independently contributed to the cytotoxicity via modulating cell cycle progression. The study further provides fundamental insights for the development of AsIII in combination with Tetra for patients with different types of breast cancer.

5.
Oncol Rep ; 44(3): 1293, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32582993

RESUMO

Following the publication of the above paper, an interested reader drew to our attention the fact that the same ß­actin bands had been included in the western blots featured in Figs. 7 and 10. Upon consulting the authors in relation to this matter, they were able to offer a legitimate explanation for this apparent duplication of the bands; essentially, in the western blotting experiments, different antibodies were used in one membrane to present different target proteins by stripping and reprobing the membrane. That the authors had performed this additional step in their protocol was inadvertently omitted from the 'Western blot analyses' subsection of the Materials and methods section. The authors are grateful to the interested reader for drawing this matter to their attention, and apologize for any inconvenience caused to the readership of the Journal. [the original article was published in Oncology Reports 41: 27-42, 2019; DOI: 10.3892/or.2018.6780].

6.
Biomed Pharmacother ; 123: 109812, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31945696

RESUMO

Wu-tou decoction (WTD), a classic Traditional Chinese medicine formula, has been extensively used in the treatment of neuropathic pain (NP) such as chronic inflammatory pain, trigeminal neuralgia, and cancer-induced pain. Our previous studies have shown that the severity of mechanical allodynia and thermo hypersensitivity in NP rats are reduced by WTD, of which analgesic candidates are paeoniflorin (Pae) and liquiritin (Liq). The aim of this study was to clarify the molecular mechanisms of WTD, Pae and Liq against NP based on the primary rat glial cells in vitro. The gene expression levels of neurotrophic factors such as nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and Artemin and C-C chemokine receptor type 5 (CCR5) were augmented by inflammatory cytokines, while chemokines increased only CCR5 gene expression. The constitutive and cytokine-augmented neurotrophic factor gene expression was enhanced by WTD, Pae, and Liq through PI3K- and PKA-dependent pathways in rat glial cells, leading to the increase of NGF and BDNF production. Furthermore, the CCR5 gene expression under basal and chemokine-treated conditions was suppressed by these reagents, in which signal pathway(s) was independent on the activation of PI3K and PKA. Moreover, there was no cytotoxicity in the WTD, Pae, and Liq treatments in glial cells. Thus, these results provide a novel evidence that WTD may exert the anti-NP actions by predominantly increasing the production of neurotrophic factors through PI3K- and PKA-signaling pathways in rat glial cells. Furthermore, Pae and Liq may play as analgesic candidates in WTD-mediated NP management.


Assuntos
Medicamentos de Ervas Chinesas/uso terapêutico , Fatores de Crescimento Neural/metabolismo , Neuralgia/tratamento farmacológico , Neuroglia/metabolismo , Neuroglia/patologia , Receptores CCR5/metabolismo , Animais , Células Cultivadas , Citocinas/metabolismo , Medicamentos de Ervas Chinesas/farmacologia , Flavanonas/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Glucosídeos/farmacologia , Mediadores da Inflamação/metabolismo , Monoterpenos/farmacologia , Fatores de Crescimento Neural/genética , Neuralgia/genética , Neuralgia/patologia , Neuroglia/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Receptores CCR5/genética , Transdução de Sinais/efeitos dos fármacos
7.
J Ethnopharmacol ; 249: 112385, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31730888

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Cerebral ischemia, also known as stroke, can stimulate the proliferation and migration of endogenous neural stem cells (NSCS) in subventricular zone of the lateral ventricle and subgranularzone of the dentate gyrus in the adult hippocampus as a defense response to damage. However, the proliferation of endogenous NSCS is insufficient for central nervous system repair. Neurogenesis and anti-neuroinflammation are two important aspects for neuroprotection. Rhizome Ligusticum chuanxiong (LC), the dried rhizomes of Ligusticum striatum DC., has been widely used to treat stroke for over hundreds of years in Traditional Chinese Medicine. PURPOSE: of the study: Previous reports on pharmacological mechanism of LC mainly focus on the cerebral blood flow and thrombolysis. We aim to explore whether LC provides neuroprotective effect by increasing neurogenesis and inhibiting the IL-1ß, TNF-α and expressions of glial fibrillary acidic protein. MATERIALS AND METHODS: LC extract was delivered to microsphere-embolized (ME) cerebral ischemia Wister rats to examine its neuroprotection. Body weight, neurological scores, hematoxylin-eosin staining (HE), TUNEL assay were conducted for neurological damage. Neurogenesis was evaluated by assessing the expression of Doublecortin (DCX) and neurogenic differentiation1 (NeuroD1) through immunofluorescence staining. Western blot performed to measure the protein levels of growth associated protein-43(GAP-43), glial fibrillary acidic protein (GFAP). IL-1ß and TNF-α was detected by Elisa. RESULTS: LC alleviated pathomorphological change and apoptosis of neurons in the hippocampus caused by ME surgery. Furthermore, LC significantly increased the DCX in the DG of adult rat hippocampus at 14 days after surgery. A significant upregulation of GAP-43 compared to the ME after LC was administered. Besides, LC decreased pro-inflammatory cytokine (IL-1ß, TNF-α) and protein level of GFAP. CONCLUSION: The finding suggested that LC had the ability to protect neurons by promoting the endogenous proliferation of neuroblast and production of neural differentiation factor in rats after ischemia injury. Meanwhile, LC can anti-neuroinflammation, which is important for the treatment of neuron injury. Accordingly, LC perhaps a promising medicine for neuron damage therapy after cerebral ischemia.


Assuntos
Isquemia Encefálica/prevenção & controle , Ligusticum/química , Fármacos Neuroprotetores/farmacologia , Acidente Vascular Cerebral/prevenção & controle , Animais , Apoptose/efeitos dos fármacos , Modelos Animais de Doenças , Hipocampo/efeitos dos fármacos , Hipocampo/patologia , Inflamação/prevenção & controle , Masculino , Microesferas , Células-Tronco Neurais/metabolismo , Neurogênese/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/patologia , Ratos , Ratos Wistar
8.
Chem Biol Interact ; 314: 108849, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31610157

RESUMO

To provide novel insight into approaches designed to combat glioblastoma, the molecular details of the cytotoxicity of gamabufotalin, were investigated in the human glioblastoma cell line U-87. A dose-dependent cytotoxicity was observed in the cells, whereas no detectable toxicity was confirmed in mouse primary astrocytes. LDH leakage was only observed in the cells treated with a relatively high concentration (>80 ng/ml). Downregulation of the expression levels of Aurora B, cdc25A, cdc25C, cdc2, Cyclin B1 and survivin, and upregulation of the expression level of p21 were observed in treated cells and occurred in parallel with G2/M phase arrest. Treatment with gamabufotalin also downregulated the expression level of uPA, CA9, and upregulated the expression level of TIMP3, all of which are closely associated with invasion/metastasis. Autophagy induction was observed in the treated cells and the addition of wortmannin, a potent autophagy inhibitor, significantly rescued U-87 cells. These results indicate that gamabufotalin exhibits cytotoxicity against cancerous glial cells with high potency and selectivity through multiple cytotoxic signaling pathways. The activation of p38 MAPK pathway along with the upregulation of VEGF/VEGFR2 was observed in the treated cells, both of which are likely to be compensatory changes in response to gamabufotalin treatment. Intriguingly, a specific inhibitor of p38 MAPK enhanced the cytotoxicity of the drug, suggesting an important prosurvival role for p38 MAPK. We thus suggest that developing a new combination regimen of gamabufotalin plus a p38 MAPK inhibitor and/or inhibitors for VEGF/VEGFR could improve the efficacy of the drug, and may provide more therapeutic benefits to patients with glioblastoma.


Assuntos
Apoptose/efeitos dos fármacos , Bufanolídeos/farmacologia , Animais , Astrócitos/citologia , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Autofagia/efeitos dos fármacos , Bufanolídeos/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Camundongos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Wortmanina/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
9.
Int J Mol Sci ; 20(20)2019 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-31640144

RESUMO

Progranulin (PGRN) plays a crucial role in diverse biological processes, including cell proliferation and embryonic development. PGRN can be cleaved by neutrophil elastase to release granulin (GRN). PGRN has been found to inhibit inflammation. Whereas, GRN plays a role as a pro-inflammatory factor. However, the pathophysiological roles of PGRN and GRN, at early stages after cerebral ischemia, have not yet been fully understood. The aim of this study was to obtain further insight into the pathologic roles of PGRN and GRN. We demonstrated that the amount of PGRN was significantly increased in microglial cells after cerebral ischemia in rats and that neutrophil elastase activity was also increased at an early stage after cerebral ischemia, resulting in the production of GRN. The inhibition of neutrophil elastase activity suppressed PGRN cleavage and GRN production, as well as the increase in pro-inflammatory cytokines, after cerebral ischemia. The administration of an elastase inhibitor decreased the number of injured cells and improved the neurological deficits test scores. Our findings suggest that an increase in the activity of elastase to cleave PGRN, and to produce GRN, was involved in an inflammatory response at the early stages after cerebral ischemia, and that inhibition of elastase activity could suppress the progression of cerebral ischemic injury.


Assuntos
Isquemia Encefálica/metabolismo , Granulinas/metabolismo , Elastase de Leucócito/metabolismo , Progranulinas/metabolismo , Animais , Células Cultivadas/citologia , Citocinas/metabolismo , Modelos Animais de Doenças , Glicina/análogos & derivados , Glicina/farmacologia , Elastase de Leucócito/antagonistas & inibidores , Masculino , Microglia/metabolismo , Ratos , Sulfonamidas/farmacologia , Regulação para Cima/efeitos dos fármacos
10.
BMC Complement Altern Med ; 19(1): 216, 2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31412862

RESUMO

BACKGROUND: Breast cancer is still the most common malignant tumor that threatens the female's life in the world, especially triple-negative breast cancer (TNBC), one of the most difficult subtypes. Lack of targeted therapies brings about urgent demand for novel treatments. In this study we aim to investigate the anti-tumor activity of Berberine (BBR), a Chinese plant-derived alkaloid, against the TNBC cell line MDA-MB-231 and elucidate its mechanism referring to anti-inflammation. METHODS: Cell inhibition rate was measured by Cell Proliferation Assay, the cytotoxic effects was detected by Lactate dehydrogenase (LDH) leakage assay, the colony formation and migration potential were evaluated by colony formation assay and wound healing assay, the release of inflammatory cytokines was detected by EMD multifactor detection, and alterations of proteins and genes related to the NLR family pyrin domain containing 3 (NLRP3) inflammasome pathway were analyzed using western blotting and real-time Polymerase Chain Reaction (PCR). RESULTS: BBR reduce the viability of MDA-MB-231 cells and increased the release of LDH from the cells in a dose-dependent manner, with and inhibition of colony formation potential and migration of the cells. BBR also caused a marked reduction in the secretion of proinflammatory cytokines, Interleukin-1α (IL-1α), Interleukin-1ß (IL-1ß), Interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). Besides, a down-regulated behavior was observed with the expression of P2X purinoceptor 7 (P2X7), NLRP3, pro-caspase-1, apoptosis-associated speck-like protein containing a caspase-activation and recruitment domain (ASC), caspase-1 p20, Interleukin-18 (IL-18), IL-1ß proteins and NLRP3, Caspase-1 and ASC mRNAs in the NLRP3 inflammasome cascade. CONCLUSIONS: Our results confirmed that BBR can effectively affect both tumor outgrowth and spontaneous metastasis in TNBC, and that we identified a new mechanism associated with inhibition the NLRP3 inflammasome pathway, suggesting its potential therapeutic relevance in clinical use.


Assuntos
Berberina/farmacologia , Medicamentos de Ervas Chinesas/farmacologia , Inflamassomos/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Neoplasias de Mama Triplo Negativas/imunologia , Apoptose/efeitos dos fármacos , Caspase 1/genética , Caspase 1/imunologia , Feminino , Humanos , Inflamassomos/genética , Inflamassomos/imunologia , Interleucina-18/genética , Interleucina-18/imunologia , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Interleucina-6/genética , Interleucina-6/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/fisiopatologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia
11.
Sci Rep ; 9(1): 11782, 2019 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-31409872

RESUMO

The N-methyl-D-aspartate (NMDA) receptor has been implicated in several neurodegenerative diseases, including stroke. Low-density lipoprotein receptor-related protein 1 (LRP1) plays pivotal roles in endocytosis and signaling in the cell. Immature LRP1 is processed by furin in the trans-Golgi network (TGN) and transported to the cell surface as its mature form. Activation of mature LRP1 exerts a protective effect against glutamate-induced degeneration of the rat retinal ganglion cells, as was shown in our previous study. However, the roles of LRP1 in the pathogenesis of excitotoxic neuronal injuries remain to be determined. The aim of this present study was to achieve further insight into the pathophysiologic roles of LRP1 after excitotoxic neuronal injuries. Our findings are the first to demonstrate that LRP1 was significantly cleaved by furin after cerebral ischemia in rats as well as after exposure of cultured cortical neurons to NMDA. It was noteworthy that the intracellular domain (ICD) of LRP1 was co-localized with TGN and furin. Furthermore, a furin inhibitor inhibited the cleavage of LRP1 and co-localization of LRP1-ICD with TGN or furin. Our findings suggest that furin-mediated cleavage of LRP1 and changes in the localization of LRP1-ICD were involved in the excitotoxic neuronal injury.


Assuntos
Isquemia Encefálica/genética , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , N-Metilaspartato/metabolismo , Acidente Vascular Cerebral/genética , Animais , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patologia , Modelos Animais de Doenças , Endocitose/efeitos dos fármacos , Furina/metabolismo , Humanos , N-Metilaspartato/farmacologia , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Neurônios/metabolismo , Neurônios/patologia , Ratos , Receptores de LDL/genética , Transdução de Sinais/efeitos dos fármacos , Acidente Vascular Cerebral/metabolismo , Acidente Vascular Cerebral/patologia , Rede trans-Golgi/efeitos dos fármacos , Rede trans-Golgi/genética
12.
J Ethnopharmacol ; 243: 112122, 2019 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-31356965

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Toad skin came from Bufo bufo gargarizans Cantor and Bufo melanostictus Schneider. As the traditional Chinese medicine, it had the effect of clearing away heat and detoxification. In traditional applications, toad skin was often used for the treatment of cancer and inflammation. Total indolealkylamines (IAAs) from this medicine were proved the main compounds exert anti-inflammatory activity in our previous research. AIM OF THE STUDY: In the present study, we aimed to investigate the potential mechanism of anti-inflammatory activity of IAAs on LPS induced zebrafish. MATERIALS AND METHODS: LPS induced zebrafish was applicated as an in vivo inflammation model to clarify the structure-activity relationship of 4 major IAAs (N-methyl serotonin, bufotenine, dehydrobufotenine and bufothionine) from toad skin. Quantitative RT-PCR was applied to detect key cytokines and members of the MyD88-dependent signaling pathway. In addition, the targeted lipidomics was conducted to find out the potential biomarkers in the inflammatory zebrafish. Network pharmacology was used to unveil the main enzymes closely related to the target lipids. RESULTS: Our results showed that the anti-inflammatory activity of free IAAs (N-methyl serotonin, bufotenine and dehydrobufotenine) was more potent than that of combined IAAs (bufothionine). RT-PCR demonstrated that 4 IAAs exerted antiendotoxin inflammatory effect via suppressing the TLR4/MyD88/NF-κB and TLR4/MyD88/MAPKs signaling pathway. A total of 33 possible inflammatory biomarkers, including 14 SM, 6 Cer, 11 PC and 2 GlcCer, triggered by LPS were screened out. The levels of most of candidates could be regulated toward a normal level by IAAs, especially in N-methyl serotonin and dehydrobufotenine groups. Enzymes especially LBP, PLA2, CERK, SMPD and SGMS were found closely associated with the regulation of most lipid markers. CONCLUSIONS: Overall, the mechanism underlying the anti-inflammatory activity of IAAs probably attributed to their capability to suppress NF-κB and MAPKs inflammatory pathway. Meanwhile, IAAs could also interfere the metabolism of SM, Cer and PC probably by regulating LBP, PLA2, CERK, SMPD and SGMS.


Assuntos
Anti-Inflamatórios/farmacologia , Indóis/farmacologia , Animais , Animais Geneticamente Modificados , Anti-Inflamatórios/química , Bufonidae , Citocinas/genética , Feminino , Indóis/química , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/genética , Larva , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipopolissacarídeos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Fator 88 de Diferenciação Mieloide/genética , NF-kappa B/metabolismo , Pele , Receptor 4 Toll-Like/genética , Peixe-Zebra
13.
Am J Chin Med ; 47(5): 1149-1170, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31311297

RESUMO

Three-dimensionally (3D) cultured tumor cells (spheroids) exhibit more resistance to therapeutic agents than the cells cultured in traditional two-dimensional (2D) system (monolayers). We previously demonstrated that arsenic disulfide (As2S2) exerted significant anticancer efficacies in both 2D- and 3D-cultured MCF-7 cells, whereas 3D spheroids were shown to be resistant to the As2S2 treatment. L-buthionine-(S, R)-sulfoximine (BSO), an inhibitor of glutathione (GSH) synthesis, has been regarded to be a potent candidate for combinatorial treatment due to its GSH modulation function. In the present study, we introduced BSO in combination with As2S2 at a low concentration to investigate the possible enhancing anticancer efficacy by the combinatorial treatment on 2D- and 3D-cultured MCF-7 cells. Our results presented for the first time that the combination of As2S2 and BSO exerted potent anticancer synergism in both MCF-7 monolayers and spheroids. The IC50 values of As2S2 in combinatorial treatment were significantly lower than those in treatment of As2S2 alone in both 2D- and 3D-cultured MCF-7 cells (P<0.01, respectively). In addition, augmented induction of apoptosis and enhanced cell cycle arrest along with the regulation of apoptosis- and cell cycle-related proteins, as well as synergistic inhibitions of PI3K/Akt signals, were also observed following co-treatment of As2S2 and BSO. Notably, the combinatorial treatment significantly decreased the cellular GSH levels in both 2D- and 3D-cultured MCF-7 cells in comparison with each agent alone (P<0.05 in each). Our results suggest that the combinatorial treatment with As2S2 and BSO could be a promising novel strategy to reverse arsenic resistance in human breast cancer.


Assuntos
Antineoplásicos/farmacologia , Arsenicais/farmacologia , Neoplasias da Mama/fisiopatologia , Butionina Sulfoximina/farmacologia , Sulfetos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Técnicas de Cultura de Células , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sinergismo Farmacológico , Feminino , Humanos , Células MCF-7 , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo
14.
Am J Transl Res ; 11(12): 7310-7323, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31934280

RESUMO

BACKGROUND: To provide novel insight into the development of new therapeutic strategies to combat breast cancer, differentiation-inducing activity of clinically achievable concentrations of arsenite (AsIII) and tetrandrine (Tetra) was investigated in breast cancer cell lines MDA-MB-231 and MCF-7. METHODS: Differentiation induction of cancer cells was analyzed by flow cytometer. Alterations of genes related to differentiation, and proliferation of human normal peripheral blood mononuclear cells (PBMCs) were analyzed using western blotting and cell viability assay, respectively. RESULTS: Exposure to Tetra alone or in combination with AsIII induced differentiation of both cells characterized by upregulation of ICAM-1, downregulation of Her2/neu. In comparison with MCF-7, the combination of lower concentrations of AsIII and Tetra induced differentiation of MDA-MB-231, indicating that MDA-MB-231 cells were highly susceptible to differentiation. The differentiation occurred in parallel with activation of Erk signaling pathway, and was abolished by PD98059, a potent Erk inhibitor. Consistent with in vitro experimental results, the upregulation of ICAM-1 and the activation of Erk signaling pathway were also observed in MDA-MB-231 breast tumors in xenograft mouse obtained from our previous study. No obvious proliferation inhibition of PBMCs was observed following the exposure to AsIII combined with Tetra at the concentrations capable of inducing differentiation of MDA-MB-231 cells. CONCLUSION: The Erk signaling pathway may be crucially involved in the differentiation induction of breast cancer cells in vitro and in vivo. Collectively, our results suggest that the combination can probably serve as promising candidates for the development of novel therapeutic approaches for different types of breast cancer.

15.
Mol Neurobiol ; 56(3): 1946-1956, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29981053

RESUMO

We previously demonstrated that proliferation of endogenous neural progenitor cells is enhanced by cerebral ischemia and that phosphatidylinositol 3-kinase (PI3-K)/Akt-dependent glycogen synthase kinase (GSK)-3ß signaling is involved in ischemia-induced neurogenesis. It is important to learn more about the regulation of proliferation and differentiation of neural progenitor cells under ischemic conditions, as such knowledge that may serve as the basis for the development of new therapeutic approaches for stroke. However, it remains to be addressed whether a change in that signaling pathway is induced in neural progenitor cells. We prepared neural progenitor cells by using the neurosphere method and conducted experiments to determine the relative contributions of the PI3-K/Akt-dependent GSK-3ß signaling pathway to the proliferation and differentiation of neural progenitor cells under the hypoxic condition in vitro. We showed that hypoxic exposure induced the proliferation of neural progenitor cells. This proliferation was accompanied by phosphorylation of Akt and GSK-3ß at its Ser9. Furthermore, treatment with a PI3-K inhibitor decreased the hypoxia-induced phosphorylation of GSK-3ß and proliferation of neural progenitor cells. Furthermore, hypoxic exposure enhanced the differentiation of neural progenitor cells, and this increased differentiation was not affected by treatment with the PI3-K inhibitor. Although the expression of NeuroD1 mRNA during cell differentiation was also enhanced by hypoxic exposure, this increased expression was not affected by treatment with the PI3-K inhibitor. Our findings suggest that the PI3K/Akt-dependent GSK-3ß signaling pathway was involved in the proliferation of neural progenitor cells under a pathologic condition, such as hypoxia and/or cerebral ischemia in vivo.


Assuntos
Hipóxia Celular/fisiologia , Proliferação de Células/fisiologia , Glicogênio Sintase Quinase 3 beta/metabolismo , Células-Tronco Neurais/metabolismo , Neurogênese/fisiologia , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais/fisiologia , Animais , Células-Tronco Neurais/citologia , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos
16.
Oncol Rep ; 41(1): 27-42, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30320388

RESUMO

In the present study, the antitumor effects of arsenic disulfide (As2S2) on the proliferative, survival and migratory ability of human breast cancer MCF­7 and MDA­MB­231 cells were investigated, and its potential underlying molecular mechanisms with an emphasis on cell cycle arrest, apoptosis induction, autophagy induction and reactive oxygen species (ROS) generation were determined. The results indicated that As2S2 significantly inhibited the viability, survival and migration of breast cancer cells in a dose­dependent manner. In addition, it was identified that As2S2 induced cell cycle arrest primarily at G2/M phase in the two breast cancer cell lines by regulating the expression of associated proteins, including cyclin B1 and cell division cycle protein 2. In addition to cell cycle arrest, As2S2 also triggered the induction of apoptosis in cells by activating the expression of pro­apoptotic proteins, including caspase­7 and ­8, as well as increasing the B­cell lymphoma 2 (Bcl­2)­associated X protein/Bcl­2 ratio, while decreasing the protein expression of anti­apoptotic B­cell lymphoma extra­large. In addition, As2S2 stimulated the accumulation of microtubule­associated protein 1A/1B­light chain 3 (LC3)­II and increased the LC3­II/LC3­I ratio, indicating the occurrence of autophagy. As2S2 treatment also inhibited the protein expression of matrix metalloproteinase­9 (MMP­9), but increased the intracellular accumulation of ROS in the two breast cancer cell lines, which may assist in alleviating metastasis and attenuating the progression of breast cancer. Taken together, the results of the present study suggest that As2S2 inhibits the progression of human breast cancer cells through the regulation of cell cycle arrest, intrinsic and extrinsic apoptosis, autophagy, MMP­9 signaling and ROS generation.


Assuntos
Antineoplásicos/farmacologia , Proteínas Reguladoras de Apoptose/metabolismo , Arsenicais/farmacologia , Neoplasias da Mama/metabolismo , Sulfetos/farmacologia , Apoptose , Autofagia , Neoplasias da Mama/tratamento farmacológico , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Células MCF-7 , Espécies Reativas de Oxigênio/metabolismo
17.
Molecules ; 24(1)2018 Dec 27.
Artigo em Inglês | MEDLINE | ID: mdl-30591646

RESUMO

Toad skin and toad venom, as two kinds of Chinese medicine, are prepared from Bufo bufo gargarizans Cantor and Bufo melanostictus Schneider. However, they display distinct properties in traditional application, and the hydrophilic ingredients are the possible distinguishing chemicals between them. In this work, 36 and 22 hydrophilic components were characterized from toad skin and venom, respectively, by UHPLC-HR-MS/MS, including amino acids, nucleosides, polypeptides, and indolealkylamines (IAAs). Among them, 15 compounds were unambiguously confirmed by comparison with standards. The CID-MS/MS fragmentation behaviors of seven indolealkylamine references were investigated to ascertain three types of structures. Subsequently, 11 high abundance contents of hydrophilic ingredients were determined from 11 batches of toad skin and 4 batches of toad venom by UPLC-QqQ-MS/MS. The quantitative results showed that the content of main IAAs in toad venom was much higher than in skin. In addition, the N-methyl serotonin (free IAA), bufothionine (combined IAA), and total IAAs sample were selected for anti-inflammatory evaluation in lipopolysaccharide (LPS) stimulated zebrafish embryo models. The obvious anti-inflammatory activities of IAAs were observed, especially for the free IAAs. This study illustrated IAAs were the main distinct hydrophilic components that probably lead to the difference between toad skin and toad venom in traditional applications.


Assuntos
Aminas/farmacologia , Venenos de Anfíbios/química , Anti-Inflamatórios/farmacologia , Bufonidae/metabolismo , Interações Hidrofóbicas e Hidrofílicas , Pele/química , Espectrometria de Massas em Tandem/métodos , Aminas/química , Animais , Cromatografia Líquida de Alta Pressão , Alcaloides Indólicos/farmacologia , Lipopolissacarídeos , Análise de Componente Principal , Compostos de Quinolínio/farmacologia , Padrões de Referência , Serotonina/análogos & derivados , Serotonina/farmacologia , Peixe-Zebra
18.
Int J Oncol ; 53(6): 2488-2502, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30272276

RESUMO

Glioblastoma is the most common and lethal intracranial tumor type, characterized by high angiogenic and infiltrative capacities. To provide a novel insight into therapeutic strategies against glioblastoma, the cytotoxicity of arenobufagin and hellebrigenin was investigated in the human glioblastoma cell line, U-87. Similar dose-dependent cytotoxicity was observed in the cells, whereas no detectable toxicity was confirmed in mouse primary astrocytes. Treatment with each drug downregulated the expression levels of Cdc25C, Cyclin B1 and survivin, which occurred in parallel with G2/M phase arrest. Necrotic-like cell death was only observed in the cells treated with a relatively high concentration (>100 ng/ml). These results indicate that the two drugs exhibited distinct cytotoxicity against cancerous glial cells with high potency and selectivity, suggesting that growth inhibition associated with G2/M phase arrest and/or necrosis were attributed to their toxicities. Activation of the p38 mitogen activated protein kinase (MAPK) signaling pathway was also observed in treated cells. Notably, a specific inhibitor of p38 MAPK, SB203580, itself caused a significant decrease in cell viability, and further enhanced the cytotoxicity of the two drugs, suggesting an important pro-survival role for p38 MAPK. Given that p38 MAPK serves an essential role in promoting glioblastoma cell survival, developing a novel combination regimen of arenobufagin/hellebrigenin plus a p38 MAPK inhibitor may improve the efficacy of the two drugs, and may provide more therapeutic benefits to patients with glioblastoma. The qualitative assessment demonstrated the existence of arenobufagin in the cerebrospinal fluid of arenobufagin-treated rats, supporting its clinical application.


Assuntos
Antineoplásicos/farmacologia , Neoplasias Encefálicas/metabolismo , Bufanolídeos/farmacologia , Ciclo Celular/efeitos dos fármacos , Glioblastoma/metabolismo , Animais , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ciclina B1/genética , Ciclina B1/metabolismo , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Humanos , Imidazóis/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Piridinas/farmacologia , Ratos , Survivina/genética , Survivina/metabolismo , Fosfatases cdc25/genética , Fosfatases cdc25/metabolismo
19.
Cancer Cell Int ; 18: 113, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30123091

RESUMO

Background: Triple-negative breast cancer (TNBC) is one of the most difficult subtypes of breast cancer to treat due to its aggressive, metastatic behavior, and a lack of a targeted therapy. Trivalent arsenic derivatives (arsenite, AsIII) with remarkable clinical efficacy in acute promyelocytic leukemia has been demonstrated to exhibit inhibitory effect against breast cancer cells. To provide novel insight into the development of new therapeutic strategies, antitumor activity of AsIII and tetrandrine (Tetra), a Chinese plant-derived alkaloid, against the TNBC cell line MDA-MB-231 in vitro and in vivo was investigated. Methods: Cytotoxicity was evaluated using cell viability, lactate dehydrogenase leakage and cell cycle assay. Alterations of genes related to cell proliferation and death were analyzed using western blotting. In vivo antitumor activity of AsIII alone or in combination with Tetra was studied using MDA-MB-231 xenografts in nude mice. Results: Synergistic cytotoxic effects of two drugs were observed in the cells. In vivo study also showed that co-administration of AsIII and Tetra significantly reduced tumor volume and weight, directly supporting its in vitro antitumor activity. No deaths and reduction of body-weight were observed after a long-term co-administration, indicating its good tolerability. S-phase arrest associated with the upregulation of FOXO3a, p27 along with decreased Cyclin D1 expression was observed in the cells treated with the combined regimen. A substantial upregulated p21 expression and downregulated phospho-FOXO3a and Cyclin D1 expression was observed in the tumor tissues of mice co-administered with AsIII and Tetra. Autophagy induction was observed in the combination treatment in vitro and in vivo. The addition of wortmannin, a potent autophagy inhibitor, significantly rescued MDA-MB-231 cells from their cytotoxicity of AsIII and Tetra. Conclusions: S-phase arrest, autophagic and necrotic cell death contribute to the cytocidal effects of the combined regimen of AsIII and Tetra. Considering our previous study showing synergistic cytotoxic effects of the combined regimen in estrogen receptor-positive breast cancer cell line MCF-7, these results suggest that development of the combination regimen of AsIII plus Tetra may offer many benefits to patients with different types of breast cancer.

20.
Chem Biol Interact ; 294: 9-17, 2018 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-30125548

RESUMO

Among five major anthocyanin compounds, delphinidin exhibited the most potent and selective cytocidal effect against HL-60, a trivalent arsenic (As(III))-resistant cell line. Co-treatment with delphinidin and As(III) resulted in the reduction of IC50 value for As(III) from 11.2 to 1.5 µM, which was considered as clinically achieved concentrations of As(III). The combination treatment strongly preferred to selectively enhance the cytotoxicity of As(III) against HL-60 cells rather than human peripheral blood mononuclear cells. The induction of apoptosis as evidenced by the increase of sub-G1 cells, DNA fragmentation, annexin V-positive cells and the activation of caspase-8, -9 and -3 was observed in HL-60 cells co-treated with As(III) and delphinidin. Similar to the activation pattern of caspases, a substantial decrease in the expression level of Bid along with the loss of mitochondrial membrane potential was also observed. These results suggested that the combination treatment triggered a convergence of the intrinsic and extrinsic pathways of apoptosis via the activation of caspase-8 and cleaved Bid. Delphinidin itself significantly decreased the intracellular GSH ([i]GSH) and nuclear factor-κB (NF-κB) binding activity, and further returned As(III)-triggered increment of [i]GSH and enhancement of NF-κB binding activity to control level. Additionally, buthionine sulfoximine, a GSH depletor; JSH-23, a NF-κB inhibitor, also mimicked the capacity of delphinidin to significantly induce the reduction of [i]GSH along with the potentiation of As(III) cytotoxicity in HL-60 cells. These observations suggested that delphinidin-induced sensitization of HL-60 cells to As(III) was caused by the reduction of [i]GSH, which was probably associated with the inhibitory effect of delphinidin on NF-κB binding activity. These findings further suggest that delphinidin-induced sensitization of HL-60 cells to As(III) may lead to dose reduction of As(III) in clinical application, and ultimately contribute to minimizing its side effects.


Assuntos
Antocianinas/química , Apoptose/efeitos dos fármacos , Arsenitos/farmacologia , Antocianinas/farmacologia , Arsenitos/química , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/metabolismo , Caspases/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Fragmentação do DNA/efeitos dos fármacos , Sinergismo Farmacológico , Glutationa/metabolismo , Células HL-60 , Humanos , Leucemia Promielocítica Aguda/metabolismo , Leucemia Promielocítica Aguda/patologia , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , NF-kappa B/química , NF-kappa B/metabolismo
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