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1.
Pharmacol Res ; 153: 104657, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31982488

RESUMO

Nonalcoholic fatty liver disease (NAFLD) is the most common liver disease worldwide and constitutes a major risk factor for progression to cirrhosis, liver failure and hepatocellular carcinoma (HCC). The occurrence of NAFLD is closely associated with abnormal lipid metabolism and implies a high risk of type 2 diabetes and cardiovascular disease. Therefore, specific and effective drugs for the prevention and treatment of NAFLD are necessary. Hypericin (HP) is one of the main active ingredients of Hypericum perforatum L., and we previously revealed its protective role in islet ß-cells and its effects against type 2 diabetes. In this study, we aimed to explore the preventive and therapeutic effects of HP against NAFLD and the underlying mechanisms in vitro and in vivo. Here, we demonstrated that HP improved cell viability by reducing apoptosis and attenuated lipid accumulation in hepatocytes both in vitro and in vivovia attenuating oxidative stress, inhibiting lipogenesis and enhancing lipid oxidization. Thus, HP exhibited significant preventive and therapeutic effects against HFHS-induced NAFLD and dyslipidemia in mice. Furthermore, we demonstrated that HP directly bound to PKACα and activated PKA/AMPK signaling to elicit its effects against NAFLD, suggesting that PKACα is one of the drug targets of HP. In addition, the enhancing effect of HP on lipolysis in adipocytes through the activation of PKACα was also elucidated. Together, the conclusions indicated that HP, of which one of the targets is PKACα, has the potential to be used as a preventive or therapeutic drug against NAFLD or abnormal lipid metabolism in the future.

2.
Biomed Pharmacother ; 121: 109615, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31707343

RESUMO

Psoriasis is considered an immune-mediated inflammatory skin disorder that affects the quality of life of nearly four percent of the world population. Considering the side effects of existing therapeutic drugs and the urgent need for new drug development, we screened more than 250 traditional Chinese medicine compounds to identify drugs that significantly reduced the viability of human HaCaT keratinocytes, a psoriasis-related model cell line. Convallatoxin (CNT) was found to be a highly effective inhibitor of HaCaT cell viability. Subsequent mechanistic studies revealed that CNT induced HaCaT cell death by necroptosis rather than by apoptosis. CNT destroyed the membrane integrity of HaCaT cells, as detected by nuclear propidium iodide (PI) staining and lactate dehydrogenase (LDH) release. Additionally, the intercellular levels of adenosine triphosphate (ATP) were lower in HaCaT cells treated with CNT than in control HaCaT cells, and typical necroptosis-associated characteristics were observed by electron microscopy in cells treated with CNT. Furthermore, compared with control HaCaT cells, CNT-treated HaCaT cells produced more reactive oxygen species (ROS), but this effect was inhibited by the antioxidants N-acetyl-cysteine (NAC), diphenyleneiodonium chloride (DPI), and apocynin and the necroptosis inhibitor Nec-1. In addition, antioxidant treatment attenuated necroptotic cell death, suggesting that CNT-induced HaCaT necroptosis is mediated by oxidative stress. More importantly, CNT ameliorated skin lesions and inflammation in imiquimod (IMQ)- and 12-O-tetradecanoyl-phorbol-13-acetate (TPA)-induced psoriasis-like mouse models. In conclusion, our results demonstrate that CNT is cytotoxic against HaCaT cells in vitro and exerts antipsoriatic activities in two mouse models of psoriasis in vivo, making CNT a potential promising candidate drug for future research.

3.
Eur J Pharmacol ; 863: 172680, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31563649

RESUMO

Psoriasis is a common dermatosis causing considerable inconvenience to 4% of the general population. Traditional psoriasis treatments often cause side effects, drug resistance and complications, necessitating development of safer and more effective treatments. In this study, we screened over 600 natural compounds to identify viability inhibitors of human HaCaT keratinocytes cultured in vitro. The results showed that nitidine chloride was a highly effective inhibitor. Further studies revealed that nitidine chloride inhibited HaCaT proliferation and induced S phase cell cycle arrest; these effects were associated with reduced DNA synthesis, decreased Ki67, cyclin A, and cyclin D1 levels, and increased p53 protein expression. Nitidine chloride also significantly downregulated bcl-2 and upregulated bax, cleaved caspase-9 and cleaved caspase-3. Mechanistic studies revealed that nitidine chloride-induced apoptosis involved the c-Jun N-terminal kinase (JNK) pathway. More importantly, in 12-O-tetradecanoyl-phorbol-13-acetate (TPA)- and imiquimod (IMQ)-induced epidermal hyperplasia and inflammation models, nitidine chloride inhibited topical edema in mouse ear and back skin, substantially reducing tissue thickness and weight. In some cases, nitidine chloride also ameliorated conditions caused by TPA and IMQ, such as angiogenesis and infiltration of large numbers of inflammatory cells around blood vessels. Additionally, nitidine chloride inhibited the expression of various proinflammatory cytokines in the two animal models. In conclusion, our results are the first to demonstrate that nitidine chloride inhibits the proliferation of HaCaT cells, induces apoptosis partly via the JNK signaling pathway in vitro and ameliorates skin lesions and inflammation in vivo, making it an appropriate candidate for psoriasis treatment.

4.
Chem Pharm Bull (Tokyo) ; 67(10): 1076-1081, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31406093

RESUMO

Histone deacetylases (HDACs) are enzymes that play a key role in structural modification and gene expression. The overexpression of HDAC is associated with cancer, and thus inhibiting the enzyme could be an efficient cancer therapy. To discover new HDAC inhibitors (HDACis), we proposed an improved protocol combining a hierarchical pharmacophore search, molecular docking, and molecular dynamic simulations. The test results showed that the improved screening protocol effectively reduced the false-positive rates of drug-like chemicals. Based on the protocol, we obtained 16 hit compounds as potential HDACis from the Life Chemicals database. Enzyme inhibition experiments showed that two of the hit chemical compounds had HDAC-inhibitory effects. In vitro assays showed that Z165155756 could selectively inhibit the proliferation of cancer cells and specifically promoted apoptosis and induced G1/S phase arrest in A2780 cells. It may have potential therapeutic effects in ovarian cancer and is worthy of further investigation.


Assuntos
Antineoplásicos/análise , Antineoplásicos/farmacologia , Descoberta de Drogas , Inibidores de Histona Desacetilases/análise , Inibidores de Histona Desacetilases/farmacologia , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores de Histona Desacetilases/química , Histona Desacetilases/metabolismo , Humanos , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Relação Estrutura-Atividade
5.
Int J Biol Sci ; 15(7): 1472-1487, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31337977

RESUMO

A decrease in islet ß-cell mass is closely associated with the development and progression of diabetes. Therefore, protection against ß-cell loss is an essential measure to prevent and treat diabetes. In this study, we investigated the protective effects of non-photoactivated hypericin, a natural compound, on ß-cells both in vitro and in vivo. In vitro, hypericin greatly improved INS-1 cell viability under high-glucose and high-fatty-acid conditions by inhibiting glucotoxicity- and lipotoxicity-induced apoptosis and nitric oxide (NO) production. Then, we further demonstrated that hypericin elicited its protective effects against glucotoxicity and lipotoxicity in INS-1 cells by attenuating the reduction in pancreatic duodenal homeobox-1 (PDX1) expression and Erk activity. In vivo, prophylactic or therapeutic use of hypericin inhibited islet ß-cell apoptosis and enhanced the anti-oxidative ability of pancreatic tissue in high-fat/high-sucrose (HFHS)-fed mice, thus alleviating ß-cell loss and maintaining or improving ß-cell mass and islet size. More importantly, hypericin treatment decreased fasting blood glucose, improved glucose intolerance and insulin intolerance, and alleviated hyperinsulinaemia in HFHS-fed mice. Therefore, hypericin showed preventive and therapeutic effects against HFHS-induced onset of type II diabetes in mice. Hypericin possesses great potential for development as an anti-diabetes drug in the future.

6.
FEBS J ; 286(18): 3718-3736, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31349381

RESUMO

Glucotoxicity or lipotoxicity leads to hyperglycemia and insulin secretion deficiency, which are important causes for the onset of type 2 diabetes mellitus (T2DM). Thus, the restoration of ß-cell function is a long-sought goal in diabetes research. Previous studies have implicated pancreatic and duodenal homeobox 1 gene (Pdx1) in ß-cell function and insulin secretion. In this study, we established a Pdx1 promoter-dependent luciferase system and identified the natural compound dracorhodin perchlorate (DP) as an effective promotor of Pdx1 expression. We further demonstrated that DP could significantly inhibit ß-cell apoptosis induced by 33 mm glucose or 200 µm palmitate by interfering with endoplasmic reticulum stress and mitochondrial pathways and enhance insulin secretion as well. These effects were associated with enhanced activities of Erk1/2, which in turn promoted Pdx1 expression and increased the ratio of Bcl2/Bax, since inhibition of the Erk1/2 pathway abolished the DP-induced expression of Pdx1 and suppression of apoptosis. In addition, our in vivo results in diabetic mice indicated that DP treatment lowered blood glucose, raised insulin levels, enhanced Pdx1 expression and increased islet size and number in the pancreas of diabetic mice. Our findings suggest that Pdx1 is a potential target molecule of DP in the treatment of T2DM via the inhibition of glucotoxicity- or lipotoxicity- induced ß-cell apoptosis and the attenuation of insulin secretion dysfunction.

7.
Sci Rep ; 9(1): 2350, 2019 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-30787420

RESUMO

Histone deacetylases (HDACs) can enzymatically transferred acetyl functional group from protein or lysine residues of histone, so they can regulate the expression of lots of genes. Now HDACs are used as drug targets and many HDAC inhibitors (HDACis) were approved for cancer therapy or in clinical trials. However, the physiological mechanisms and regulatory processes of HDACi anti-cancer effects are largely unexplored and uncompleted. Here we use the virtual screening workflow obtained 25 hit compounds and ZINC24469384 can significantly inhibit HDAC activity while arrest cell cycle at G1/S phase and significantly induced HepG2 cell apoptosis, time-course RNA-seq demonstrate that HepG2 cells transcriptionally respond to ZINC24469384. Pathway analysis of DEGs and DASGs reveal that NR1H4 may play an important role in ZINC24469384-induced anti-proliferation effect and is dramatically alleviated by down-regulating the SOCS2 expression and promoting STAT3 phosphorylation in knockdown NR1H4 HepG2 cells. Analysis based on TCGA database indicated that NR1H4 and SOCS2 were downregulated in liver cancer, this suggest NR1H4 and SOCS2 may play an important role in tumorigenesis. These results indicated that ZINC24469384 is a novel benzamine lead compound of HDACi and provides a novel mechanism for HDACi to inhibit cancer.

8.
Dig Dis Sci ; 64(6): 1548-1559, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30627917

RESUMO

BACKGROUND/AIM: The miR-181 family plays an important role in the regulation of various cellular functions. However, whether miR-181b-5p mediates hepatic insulin resistance remains unknown. In this study, we investigated the effect of miR-181b-5p on the regulation of hepatic glycogen synthesis. METHODS: The miR-181b-5p levels in the livers of diabetic mice were detected by real-time PCR. The glycogen levels and AKT/GSK pathway activation were examined in human hepatic L02 cells and HepG2 cells transfected with miR-181b-5p mimic or inhibitor. The potential target genes of miR-181b-5p were evaluated using a luciferase reporter assay and Western blot analysis. EGR1-specific siRNA and pCMV-EGR1 were used to further determine the role of miR-181b-5p in hepatic glycogen synthesis in vitro. Hepatic inhibition of miR-181b-5p in mice was performed using adeno-associated virus 8 (AAV8) vectors by tail intravenous injection. RESULTS: The miR-181b-5p levels were significantly decreased in the serum and livers of diabetic mice as well as the serum of type 2 diabetes patients. Importantly, inhibition of miR-181b-5p expression impaired the AKT/GSK pathway and reduced glycogenesis in hepatocytes. Moreover, upregulation of miR-181b-5p reversed high-glucose-induced suppression of glycogenesis. Further analysis revealed that early growth response 1 (EGR1) was a downstream target of miR-181b-5p. Silencing of EGR1 expression rescued miR-181b-5p inhibition-reduced AKT/GSK pathway activation and glycogenesis in hepatocytes. Hepatic inhibition of miR-181b-5p led to insulin resistance in C57BL/6 J mice. CONCLUSION: We demonstrated that miR-181b-5p contributes to glycogen synthesis by targeting EGR1, thereby regulating PTEN expression to mediate hepatic insulin resistance.


Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Glicogênio/biossíntese , Resistência à Insulina , Fígado/metabolismo , MicroRNAs/metabolismo , Adulto , Animais , Diabetes Mellitus Tipo 2/genética , Modelos Animais de Doenças , Proteína 1 de Resposta de Crescimento Precoce/genética , Feminino , Células Hep G2 , Humanos , Masculino , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Pessoa de Meia-Idade , PTEN Fosfo-Hidrolase/metabolismo , Transdução de Sinais
9.
Eur J Pharmacol ; 836: 75-82, 2018 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-30096296

RESUMO

Endoplasmic reticulum (ER) stress, together with unfolded protein response (UPR), can remove unfolded proteins and promote survival. However, severe and prolonged ER stress leads to cell death, tissue injury, and many serious diseases. Therefore, it is essential to identify drugs that can attenuate ER stress for ER-related disease treatment. A great deal of research shows that selenoprotein S (SelS) is a sensitive and ideal marker of ER stress. Here, we used a firefly luciferase reporter driven by SelS gene promoter to screen natural compounds that can attenuate ER stress. Then we identified compound 20(S)-25-methoxyl-dammarane-3ß,12ß,20-triol (25-OCH3-PPD) could inhibit the promoter activity of SelS, further results showed that 25-OCH3-PPD effectively inhibited tunicamycin (TM) induced up-regulation of SelS expression in both mRNA and protein levels. Moreover, 25-OCH3-PPD significantly inhibited glucose-regulated protein 78 (GRP78; the major ER stress marker) expression in TM-induced ER stress in HepG2 and HEK293T cells, suggesting that 25-OCH3-PPD could attenuate ER stress in these cells. Mechanism studies showed that 25-OCH3-PPD significantly activated ERK/MAPK signaling pathway, and the inhibition of ERK/MAPK by U0126 dramatically abolished the inhibitory effect of 25-OCH3-PPD on ER stress, suggesting that 25-OCH3-PPD attenuated ER stress at least partially through activation of ERK/MAPK signaling pathway. Taken together, our studies indicate that 25-OCH3-PPD is a novel small molecular compound reducing ER stress, and a potential drug for treating diseases associated with ER stress.


Assuntos
Estresse do Retículo Endoplasmático/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Triterpenos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Células HEK293 , Células Hep G2 , Humanos , Proteínas de Membrana/genética , Selenoproteínas/genética , Regulação para Cima/efeitos dos fármacos
10.
Biochem Biophys Res Commun ; 502(2): 283-288, 2018 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-29842883

RESUMO

TSP50, a testis-specific gene encoding a serine protease-like protein, was specifically expressed in the spermatocytes of testes but abnormally activated and expressed in many different kinds of cancers. Here, we aimed to analyze the expression of TSP50 in mouse embryo and its function in early embryonic development. Firstly, the distribution of TSP50 in oocytes and embryonic development was characterized by immunofluorescence, RT-PCR and western blotting, and the results showed that TSP50 was detected at all studied stages with a dynamic expression pattern. When overexpressed TSP50 in zygotes by microinjection, the zygotes development was highly accelerated. On the contrary, knocking down TSP50 expression by RNA interference greatly retarded the zygote development. Furthermore, TSP50 expression at embryonic day 6.5 (E6.5), day 8.5 (E8.5) and day 10.5 (E10.5) were increasingly enhanced, However, the expression of TSP50 decreased gradually in the development and differentiation of cardiac myocyte from E12.5 to postnatal (P0). Additionally, we found that TSP50 expression was decreased during cardiac myocyte differentiation of P19 cells. Overexpression of TSP50 could decrease the expression of GATA-4, and knockdown of TSP50 markedly increase the expression of GATA-4. Taken together, our data indicate that TSP50 may play an important role during the process of mouse embryonic development as well as myocardial cell differentiation.


Assuntos
Desenvolvimento Embrionário/genética , Desenvolvimento Embrionário/fisiologia , Miócitos Cardíacos/citologia , Miócitos Cardíacos/enzimologia , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Animais , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Transição Epitelial-Mesenquimal/genética , Transição Epitelial-Mesenquimal/fisiologia , Feminino , Coração Fetal/embriologia , Coração Fetal/enzimologia , Regulação da Expressão Gênica no Desenvolvimento , Regulação Enzimológica da Expressão Gênica , Técnicas de Silenciamento de Genes , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Gravidez
11.
Biomed Pharmacother ; 104: 729-737, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29807222

RESUMO

Cripto-1 is an oncogenic protein that belongs to the epidermal growth factor (EGF)-cripto-1/FRL1/cryptic (CFC) family. It has been shown to stimulate tumorigenesis and metastasis by promoting cancer cell proliferation, epithelial-to-mesenchymal transition (EMT), and tumor angiogenesis. However, the role of Cripto-1 in cell survival and apoptosis remains largely undefined. In the present study, we found that Cripto-1 is significantly upregulated in a number of human cancer cell lines. The membrane-associated but not the soluble form of Cripto-1 promotes resistance to drug-induced caspase-3 cleavage, an indicator of apoptosis. Consequently, Cripto-1 silencing sensitizes human cancer cells to chemotherapy drugs including cytarabine, cisplatin and taxol. Our mechanistic studies revealed that Cripto-1 promotes apoptosis resistance by inducing NF-κB-mediated Survivin expression through activation of TAK-1. We also found that Cripto-1 silencing does not affect growth of un-treated cancer cells, and Cripto-1 forms self-assembled punctiforms and changes its subcellular distribution upon cytarabine treatment. Thus, the anti-apoptotic activity of Cripto-1 could be an inducible function that can be activated by external stimuli such as drug stimulation. Our findings suggested that targeting the Cripto-1/TAK-1/NF-κB/Survivin pathway may be an effective approach to combat apoptosis resistance in cancer.


Assuntos
Apoptose/fisiologia , Resistencia a Medicamentos Antineoplásicos/fisiologia , Proteínas Ligadas por GPI/metabolismo , Proteínas Inibidoras de Apoptose/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , MAP Quinase Quinase Quinases/genética , NF-kappa B/metabolismo , Proteínas de Neoplasias/metabolismo , Transdução de Sinais/fisiologia , Apoptose/efeitos dos fármacos , Carcinogênese/efeitos dos fármacos , Carcinogênese/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cisplatino/farmacologia , Citarabina/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Fator de Crescimento Epidérmico/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Células HEK293 , Humanos , Neovascularização Patológica/metabolismo , Paclitaxel/farmacologia , Transdução de Sinais/efeitos dos fármacos , Survivina
12.
Sci Rep ; 8(1): 1479, 2018 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-29367618

RESUMO

Prohibitin 2 (PHB2), as a conserved multifunctional protein, is traditionally localized in the mitochondrial inner membrane and essential for maintenance of mitochondrial function. Here, we investigated the role of PHB2 in human rhabdomyosarcoma (RMS) RD cells and found substantial localization of PHB2 in the nucleolus. We demonstrated that PHB2 knockdown inhibited RD cell proliferation through inducing cell cycle arrest and suppressing DNA synthesis. Meanwhile, down-regulation of PHB2 also induced apoptosis and promoted differentiation in fractions of RD cells. In addition, PHB2 silencing led to altered nucleolar morphology, as observed by transmission electron microscopy, and impaired nucleolar function, as evidenced by down-regulation of 45S and 18S ribosomal RNA synthesis. Consistently, upon PHB2 knockdown, occupancy of c-Myc at the ribosomal DNA (rDNA) promoter was attenuated, while more myoblast determination protein 1 (MyoD) molecules bound to the rDNA promoter. In conclusion, our findings suggest that nucleolar PHB2 is involved in maintaining nucleolar morphology and function in RD cells by regulating a variety of transcription factors, which is likely to be one of the underlying mechanisms by which PHB2 promotes tumor proliferation and represses differentiation. Our study provides new insight into the pathogenesis of RMS and novel characterizations of the highly conserved PHB2 protein.


Assuntos
Nucléolo Celular/metabolismo , Proliferação de Células , RNA Ribossômico/genética , Proteínas Repressoras/metabolismo , Rabdomiossarcoma/patologia , Transcrição Genética , Apoptose , Diferenciação Celular , Humanos , Proteína MyoD/metabolismo , RNA Interferente Pequeno/genética , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/genética , Rabdomiossarcoma/genética , Rabdomiossarcoma/metabolismo , Células Tumorais Cultivadas
13.
Apoptosis ; 22(11): 1404-1418, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28864870

RESUMO

The pro-inflammatory cytokine interleukin 6 (IL-6), via activating its downstream JAK/STAT3 and Ras/ERK signaling pathways, is involved in cell growth, proliferation and anti-apoptotic activities in various malignancies. To screen inhibitors of IL-6 signaling, we constructed a STAT3 and ERK dual-pathway responsive luciferase reporter vector (Co.RE). Among several candidates, the natural compound 20(S)-25-methoxyl-dammarane-3ß, 12ß, 20-triol (25-OCH3-PPD, GS25) was identified to clearly inhibit the luciferase activity of Co.RE. GS25 was confirmed to indeed inhibit activation of both STAT3 and ERK pathways and expression of downstream target genes of IL-6, and to predominantly decrease the viability of HepG2 cells via induction of cell cycle arrest and apoptosis. Interestingly, GS25 showed preferential inhibition of HepG2 cell viability relative to normal liver L02 cells. Further investigation showed that GS25 could not induce apoptosis and block activation of STAT3 and ERK pathways in L02 cells as efficiently as in HepG2 cells, which may result in differential effects of GS25 on malignant and normal liver cells. In addition, GS25 was found to potently suppress the expression of endogenous STAT3 at a higher concentration and dramatically induce p38 phosphorylation in HepG2 cells, which could mediate its anti-cancer effects. Finally, we demonstrated that GS25 also inhibited tumor growth in HepG2 xenograft mice. Taken together, these findings indicate that GS25 elicits its anti-cancer effects on HepG2 cells through multiple mechanisms and has the potential to be used as an inhibitor of IL-6 signaling. Thus, GS25 may be developed as a treatment for hepatocarcinoma with low toxicity on normal liver tissues as well as other inflammation-associated diseases.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Regulação Neoplásica da Expressão Gênica , Ginsenosídeos/farmacologia , Hepatoblastoma/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Fator de Transcrição STAT3/genética , Animais , Antineoplásicos Fitogênicos/síntese química , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Proliferação de Células/efeitos dos fármacos , Feminino , Genes Reporter , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Ginsenosídeos/química , Células Hep G2 , Hepatoblastoma/genética , Hepatoblastoma/metabolismo , Hepatoblastoma/patologia , Humanos , Interleucina-6/genética , Interleucina-6/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Luciferases/genética , Luciferases/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Camundongos Nus , Fator de Transcrição STAT3/agonistas , Fator de Transcrição STAT3/metabolismo , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
14.
Int J Mol Sci ; 18(4)2017 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-28394269

RESUMO

Ginkgolide A (GA) is a natural compound isolated from Ginkgo biloba and has been used to treat cardiovascular diseases and diabetic vascular complications. However, only a few studies have been conducted on the anti-inflammatory effects of GA. In particular, no related reports have been published in a common inflammation model of lipopolysaccharide (LPS)-stimulated macrophages, and the anti-inflammatory mechanisms of GA have not been fully elucidated. In the present study, we extensively investigated the anti-inflammatory potential of GA in vitro and in vivo. We showed that GA could suppress the expression of pro-inflammatory mediators (cyclooxygenase-2 (COX-2) and nitric oxide (NO) and pro-inflammatory cytokines (tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-1ß) in LPS-treated mouse peritoneal macrophages, mouse macrophage RAW264.7 cells, and differentiated human monocytes (dTHP-1) in vitro. These effects were partially carried out via downregulating Nuclear factor kappa-B (NF-κB), Mitogen-activated protein kinases (MAPKs) (p38 mitogen-activated protein kinase and extracellular signal-regulated kinase (ERK), but not c-Jun N-terminal kinase (JNK), and activating the AMP-activated protein kinase (AMPK) signaling pathway also seems to be important. Consistently, GA was also shown to inhibit the LPS-stimulated release of TNF-α and IL-6 in mice. Taken together, these findings suggest that GA can serve as an effective inflammatory inhibitor in vitro and in vivo.


Assuntos
Ginkgolídeos/farmacologia , Inflamação/prevenção & controle , Lactonas/farmacologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Animais , Western Blotting , Linhagem Celular , Linhagem Celular Tumoral , Ciclo-Oxigenase 2/metabolismo , Citocinas/sangue , Citocinas/genética , Citocinas/metabolismo , Feminino , Expressão Gênica/efeitos dos fármacos , Humanos , Inflamação/sangue , Inflamação/induzido quimicamente , Mediadores da Inflamação/sangue , Mediadores da Inflamação/metabolismo , Lipopolissacarídeos , Macrófagos/metabolismo , Macrófagos Peritoneais/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Óxido Nítrico/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/efeitos dos fármacos
15.
Mol Immunol ; 82: 94-103, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-28064070

RESUMO

Maspin (mammary serine protease inhibitor) is a non-inhibitory member of the serine protease inhibitor superfamily and a tumor suppressor in several cancers due to its ability to inhibit cell invasion, angiogenesis, and promote apoptosis. However, its immunomodulatory function remains largely unexplored. Thus, we explored the potential link between Maspin and macrophage function, first evaluating the regulatory effects of conditioned medium (CM) of a Maspin-overexpressing CHO cell strain on mouse peritoneal macrophage phagocytosis and cytokine secretion. Next, we used a transwell co-culture system and recombinant Maspin (rMaspin) to confirm the effects of Maspin on macrophages, and attempted to clarify the underlying mechanisms. We found that irrespective of CM, rMaspin or co-culture of Maspin-overexpressing cells with macrophages impaired macrophages phagocytosing Saccharomyces cerevisiae. Furthermore, q-RT-PCR or ELISA confirmed increased IL-1ß, TNF-α, IFN-γ, IL-6, IL-12, IL-10, and M1 marker iNOS production in macrophages after Maspin stimulation, but TGF-ß and M2 marker Arg-1 production were suppressed. Western blot showed activated NF-κB signaling in Maspin-stimulated macrophages; upregulated cytokines were lowered, and impaired phagocytosis recovered after blocking NF-κB signaling with PDTC. Thus, Maspin mildly inhibited phagocytic activity, but markedly enhanced inflammatory cytokine production and likely skewed macrophages towards M1 polarization, partially due to activation of NF-κB signaling. These results reveal a novel biological function of Maspin in modulating macrophage activity and may open a new avenue for Maspin-based tumor therapy.


Assuntos
Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Fagocitose , Serpinas/imunologia , Transdução de Sinais/imunologia , Animais , Células CHO , Técnicas de Cocultura , Cricetulus , Citocinas/biossíntese , Ensaio de Imunoadsorção Enzimática , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/imunologia , NF-kappa B/metabolismo , Fagocitose/imunologia , Reação em Cadeia da Polimerase , Serpinas/metabolismo , Transfecção
16.
Tumour Biol ; 37(9): 11805-11813, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27039397

RESUMO

While the incidence of cancer continues to increase, the current therapeutic options remain imperfect. Therefore, there is an urgent need to discover new targeted anti-cancer therapies. Testes-specific protease 50 (TSP50) is abnormally expressed in most cancer tissues and downregulation of TSP50 expression can reduce cell proliferation and induce cell apoptosis, which makes it a potential target for cancer therapy. In this study, we constructed a firefly luciferase reporter pGL3-TSP50-3'-UTR as a drug screening model to screen potential candidate compounds that target TSP50 mRNA. We identified the compound 7P3A, which consists of 70 % 25-methoxyl-dammarane-3ß, 12ß, 20-triol and 30 % artemisinin, as being capable of inhibiting the TSP50-3'-UTR reporter activity, as well as the expression of TSP50. Further investigation revealed that 7P3A could inhibit MDA-MB-231 cell proliferation and induce cell cycle arrest, and over-expression of TSP50 partially reversed the effect of 7P3A. In vivo investigation showed that 7P3A could inhibit tumor growth in a xenograft model of breast cancer. These results suggest that 7P3A exhibits anti-cancer effects, in part, through downregulation of TSP50 expression.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias da Mama/tratamento farmacológico , Proliferação de Células/efeitos dos fármacos , Serina Endopeptidases/metabolismo , Regiões 3' não Traduzidas/genética , Animais , Antimaláricos/administração & dosagem , Artemisininas/administração & dosagem , Western Blotting , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação para Baixo/efeitos dos fármacos , Sinergismo Farmacológico , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Células MCF-7 , Camundongos Endogâmicos BALB C , Camundongos Nus , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sapogeninas/administração & dosagem , Serina Endopeptidases/genética , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/genética , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Mol Med Rep ; 13(6): 5118-24, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27109260

RESUMO

The primary effect of the endoplasmic reticulum (ER) stress response or unfolded protein response (UPR) is to reduce the load of unfolded protein and promote survival. However, prolonged and severe ER stress leads to tissue injury and serious diseases. Thus, it is important to identify drugs that can attenuate ER stress for the treatment of diseases. Natural products continue to provide lead compounds for drug discovery and front­line pharmacotherapy for people worldwide. Previous studies have indicated that selenoprotein S (SelS) is a sensitive and ideal maker of ER stress. In the present study, a firefly luciferase reporter driven by the SelS gene promoter was used to screen for natural compounds capable of attenuating ER stress. From this, paclitaxel (PTX) was identified to efficiently inhibit the promoter activity of the SelS gene, and further results revealed that PTX significantly inhibited the tunicamycin­induced upregulation of SelS at the mRNA and protein levels in HepG2 and HEK293T cells. In addition, PTX was able to efficiently inhibit the expression of the ER stress marker, glucose­regulated protein 78, in ER stress, indicating that PTX may reverse ER stress. Taken together, these results suggest that PTX is able to inhibit SelS expression during ER stress and attenuate ER stress.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Membrana/genética , Paclitaxel/farmacologia , Selenoproteínas/genética , Antineoplásicos Fitogênicos/química , Produtos Biológicos/química , Produtos Biológicos/farmacologia , Linhagem Celular , Descoberta de Drogas/métodos , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Humanos , Paclitaxel/química
18.
Int J Biochem Cell Biol ; 73: 63-71, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26855338

RESUMO

Previous studies have shown that testes-specific protease 50 (TSP50), a pro-oncogene overexpressed in many types of tumors, could promote cell proliferation, invasion, tumorigenesis, and tumor metastasis, suggesting that it is a potential cancer therapeutic target in drug discovery. Here, a luciferase assay system driven by the TSP50 gene promoter was used to screen the inhibitor of expression of TSP50. The study found that cardamonin, a flavone compound, could efficiently inhibit the expression of TSP50 in both mRNA and protein levels. Further results revealed that cardamonin also efficiently inhibited the viability of TSP50 high-expressing cancer cells by inducing G2/M-phase arrest and mitochondrial-dependent apoptosis. Surprisingly, knocking down the expression of TSP50 gene had the same effects as treatment with cardamonin. Moreover, it has been found that cardamonin had an inhibitory potency on TSP50 high-expressing tumor growth in vivo. In contrast, overexpression of TSP50 greatly decreased the cell sensitivity to the inhibitory effect of cardamonin and reversed the decreased tumor-inhibitory effect of cardamonin. Additionally, both TSP50 interference and treatment with cardamonin could suppress p65 nuclear translocation, and overexpression of TSP50 reversed the suppressive effect of cardamonin on p65 nuclear translocation. Taken together, these results suggest that cardamonin inhibited cell viability and tumorigenesis at least partially via blocking the activation of TSP50-mediated nuclear factor-kappaB signaling pathway, and cardamonin may be a promising anticancer drug candidate in the development of a novel agent for TSP50 high-expressing cancer cells.


Assuntos
Chalconas/farmacologia , NF-kappa B/metabolismo , Testículo/enzimologia , Apoptose/efeitos dos fármacos , Carcinogênese/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Transformação Celular Neoplásica/efeitos dos fármacos , Células HEK293 , Humanos , Marcação In Situ das Extremidades Cortadas , Masculino , Regiões Promotoras Genéticas/genética , Serina Endopeptidases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Testículo/efeitos dos fármacos
19.
Biochem Pharmacol ; 105: 66-79, 2016 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-26850986

RESUMO

Psoriasis is a multifactorial skin disease that inconveniences many patients. Considering the side effects and drug resistance of the current therapy, it is urgent to discover more effective and safer anti-psoriatic drugs. In the present study, we screened over 250 traditional Chinese medicine compounds for their ability to inhibit the cell viability of cultured human HaCaT keratinocytes, a psoriasis-relevant in vitro model, and found that periplogenin was highly effective. Mechanistic studies revealed that apoptosis and autophagy were not induced by periplogenin in HaCaT cells. However, periplogenin caused PI to permeate into cells, increased lactate LDH release and rapidly increased the number of necrotic cells. Additionally, the typical characteristics of necrosis were observed in the periplogenin-treated HaCaT cells. Notably, the necroptosis inhibitor Nec-1 and NSA were able to rescue the cells from necrotic cell death, supporting that necroptosis was involved in periplogenin-induced cell death. Furthermore, the ROS levels were elevated in the periplogenin-treated cells, NAC (an antioxidant) and Nec-1 could inhibit the ROS levels, and NAC could attenuate necroptotic cell death, indicating that the periplogenin-induced necroptotic cell death was mediated by oxidative stress. More importantly, in the murine models of TPA-induced epidermal hyperplasia and IMQ-induced skin inflammation, topical administration of periplogenin ameliorated skin lesions and inflammation. In sum, our results indicate, for the first time, that periplogenin is a naturally occurring compound with potent anti-psoriatic effects in vitro and in vivo, making it a promising candidate for future drug research.


Assuntos
Aminoquinolinas/toxicidade , Digitoxigenina/análogos & derivados , Modelos Animais de Doenças , Estresse Oxidativo/fisiologia , Psoríase/metabolismo , Acetato de Tetradecanoilforbol/toxicidade , Animais , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Digitoxigenina/farmacologia , Digitoxigenina/uso terapêutico , Feminino , Humanos , Imiquimode , Camundongos , Camundongos Endogâmicos BALB C , Necrose/induzido quimicamente , Necrose/tratamento farmacológico , Necrose/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Psoríase/induzido quimicamente , Psoríase/tratamento farmacológico
20.
Apoptosis ; 21(3): 340-50, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26621097

RESUMO

Juglans mandshurica Maxim (Juglandaceae) is a famous folk medicine for cancer treatment and some natural compounds isolated from it have been studied extensively. Previously we isolated a type of ω-9 polyunsaturated fatty acid (JA) from the bark of J. mandshurica, however little is known about its activity and the underlying mechanisms. In this study, we studied anti-tumor activity of JA on several human cancer cell lines. Results showed that JA is cytotoxic to HepG2, MDA-MB-231, SGC-7901, A549 and Huh7 cells at a concentration exerting minimal toxic effects on L02 cells. The selective toxicity of JA was better than other classical anti-cancer drugs. Further investigation indicated that JA could induce cell apoptosis, characterized by chromatin condensation, DNA fragmentation and activation of the apoptosis-associated proteins such as Caspase-3 and PARP-1. Moreover, we investigated the cellular apoptosis pathway involved in the apoptosis process in HepG2 cells. We found that proteins involved in mitochondrion (cleaved-Caspase-9, Apaf-1, HtrA2/Omi, Bax, and Mitochondrial Bax) and endocytoplasmic reticulum (XBP-1s, GRP78, cleaved-Caspase-7 and cleaved-Caspase-12) apoptotic pathways were up-regulated when cells were treated by JA. In addition, a morphological change in the mitochondrion was detected. Furthermore, we found that JA could inhibit DNA synthesis and induce G2/M cell cycle arrest. The expression of G2-to-M transition related proteins, such as CyclinB1 and phosphorylated-CDK1, were reduced. In contrast, the G2-to-M inhibitor p21 was increased in JA-treated cells. Overall, our results suggest that JA can induce mitochondrion- and endocytoplasmic reticulum-mediated apoptosis, and G2/M phase arrest in HepG2 cells, making it a promising therapeutic agent against hepatoma.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/patologia , Ácidos Graxos Insaturados/farmacologia , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Juglans/química , Neoplasias Hepáticas/patologia , Medicina Tradicional Chinesa , Antineoplásicos/isolamento & purificação , Proteína Quinase CDC2 , Carcinoma Hepatocelular/metabolismo , Caspase 3/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Ciclina B1/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Quinases Ciclina-Dependentes/metabolismo , Retículo Endoplasmático/metabolismo , Ácidos Graxos Insaturados/isolamento & purificação , Células Hep G2 , Humanos , Neoplasias Hepáticas/metabolismo , Mitocôndrias/metabolismo , Poli(ADP-Ribose) Polimerase-1 , Poli(ADP-Ribose) Polimerases/metabolismo , Transdução de Sinais
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