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1.
Int J Mol Sci ; 20(24)2019 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-31817146

RESUMO

Ginsenosides are known to have various highly pharmacological activities, such as anti-cancer and anti-inflammatory effects. However, the search for the most effective ginsenosides against the pathogenesis of atopic dermatitis (AD) and the study of the effects of ginsenosides on specific cytokines involved in AD remain unclear. In this study, ginsenoside Rh2 was shown to exert the most effective anti-inflammatory action on thymic stromal lymphopoietin (TSLP) and interleukin 8 in tumor necrosis factor-alpha and polyinosinic: polycytidylic acid induced normal human keratinocytes by inhibiting proinflammatory cytokines at both protein and transcriptional levels. Concomitantly, Rh2 also efficiently alleviated 2,4-dinitrochlorobenzene-induced AD-like skin symptoms when applied topically, including suppression of immune cell infiltration, cytokine expression, and serum immunoglobulin E levels in NC/Nga mice. In line with the in vitro results, Rh2 inhibited TSLP levels in AD mice via regulation of an underlying mechanism involving the nuclear factor κB pathways. In addition, in regard to immune cells, we showed that Rh2 suppressed not only the expression of TSLP but the differentiation of naïve CD4+ T-cells into T helper type 2 cells and their effector function in vitro. Collectively, our results indicated that Rh2 might be considered as a good therapeutic candidate for the alternative treatment of AD.

2.
Int J Mol Sci ; 20(24)2019 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-31817281

RESUMO

To improve the therapeutic potential of hepatocyte transplantation, the effects of the mitogen-activated protein kinase kinase 4 (MKK4) inhibitor, myricetin (3,3',4',5,5',7-hexahydroxylflavone) were examined using porcine and human hepatocytes in vitro and in vivo. Hepatocytes were cultured, showing the typical morphology of hepatic parenchymal cell under 1-10 µmol/L of myricetin, keeping hepatocyte specific gene expression, and ammonia removal activity. After injecting the hepatocytes into neonatal Severe combined immunodeficiency (SCID) mouse livers, cell colony formation was found at 10-15 weeks after transplantation. The human albumin levels in the sera of engrafted mice were significantly higher in the recipients of myricetin-treated cells than non-treated cells, corresponding to the size of the colonies. In terms of therapeutic efficacy, the injection of myricetin-treated hepatocytes significantly prolonged the survival of ornithine transcarbamylase-deficient SCID mice from 32 days (non-transplant control) to 54 days. Biochemically, the phosphorylation of MKK4 was inhibited in the myricetin-treated hepatocytes. These findings suggest that myricetin has a potentially therapeutic benefit that regulates hepatocyte function and survival, thereby treating liver failure.

3.
ACS Appl Mater Interfaces ; 11(47): 44582-44592, 2019 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-31682097

RESUMO

Janus nanoparticles with an anisotropic feature concentrated multiple properties on a single carrier, providing synergistic effects. In this study, dual-functionalized Janus nanoparticles (HA-JMSN/DOX-DMMA) were constructed with a tumor-targeting ligand (hyaluronic acid, HA) modified on the one side and a charge reversal group (2,3-dimethylmaleic anhydride, DMMA) on the other side. The drug release of HA-JMSN/DOX-DMMA was positively correlated with the acidity of the environment. The cytotoxicity and cell uptake of HA-JMSN/DOX-DMMA were superior to the isotropous nanoparticles. The endocytosis pathway of HA-JMSN/DOX-DMMA involved the clathrin-mediated endocytosis (HA) and the micropinocytosis (DMMA) at the same time, which indicated that they both participated in the interaction between nanoparticles and tumor cells. After being injected intravenously in mice, the distribution of HA-JMSN/DOX-DMMA in tumor was enhanced significantly. The antitumor therapy study in vivo showed that HA-JMSN/DOX-DMMA inhibited tumor growth and improved the survival rate of tumor-bearing mice effectively. In general, HA-JMSN/DOX-DMMA could take the synergistic effect of active targeting and charge reversal to deliver drug in tumor cells and kill them efficiently, which was a promising antitumor nanodrug.

4.
Colloids Surf B Biointerfaces ; 184: 110546, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31606701

RESUMO

CD47, a transmembrane protein overexpressed in most tumors, limits macrophage phagocytosis by interacting with macrophage signal-regulated protein α (SIRPα). In this study, we have developed CD47-targeted bismuth selenide nanoparticles (Ab-PEG-Bi2Se3) that increase phagocytosis of cancer cells by macrophages to actualize improved photothermal therapy (PTT). The functionalized nanoparticles were constructed by conjugating anti-CD47 antibody (Ab) to PEGylated bismuth selenide nanoparticles (PEG-Bi2Se3). The anti-CD47 antibody modified on the nanoparticles enhanced the phagocytic activity of macrophages toward tumor cells by specifically blocking the crosstalk between CD47 and SIRPα. Meanwhile, Ab-PEG-Bi2Se3 showed excellent photothermal performance including strong near infrared (NIR) absorbance, high photothermal conversion efficiency and photostability, and exhibited outstanding in vitro PTT effect under NIR laser irradiation. In vivo therapeutic experiments revealed that this CD47-targeted PTT nanoagent, with the assistance of enhanced macrophage phagocytosis, achieved the goal of tumor eradication. Besides, toxicity studies confirmed that Ab-PEG-Bi2Se3 had good biocompatibility. In conclusion, Ab-PEG-Bi2Se3 may serve as an efficient PTT platform in combination with macrophage-mediated immunotherapy to improve antitumor efficacy.

5.
Mol Brain ; 12(1): 77, 2019 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-31488185

RESUMO

Ginseng has been shown to produce a cognitive improvement effect. The key molecular components in ginseng that produce pharmacological effects are ginsenosides. Previous studies reported a memory improvement effect of a few major ginsenosides. However, the identity of specific minor ginsenosides mediating such function remains unknown. Here, we report that a minor ginsenoside F1 improves memory function in APPswe/PSEN1dE9 (APP/PS1) double-transgenic Alzheimer's disease (AD) model mice. After 8-wk oral administration of F1 jelly, we observed that spatial working memory, but not context-dependent fear memory, was restored in AD mice. To search for a possible underlying molecular and cellular mechanism, we investigated the effect of F1 on Aß plaque. We observed F1 administration reduced the Aß plaque area and density in the cortex, but not in the hippocampus of AD mice. Next, we tested for the effect of F1 on the expression level of key molecules involved in learning and memory. Results from Western blot assay revealed that an abnormally reduced level of a phosphorylated form of CREB in the hippocampus of AD mice was restored to a normal level by F1 administration. Moreover, in the same animals, BDNF level was augmented in the cortex. Our results, therefore, suggest that minor ginsenoside F1 constitutes a promising target to develop therapeutic agents for AD.

6.
Appl Microbiol Biotechnol ; 103(17): 7003-7015, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31289903

RESUMO

Use of recombinant glycosidases is a promising approach for the production of minor ginsenosides, e.g., Compound K (CK) and F1, which have potential applications in the food industry. However, application of these recombinant enzymes for food-grade preparation of minor ginsenosides are limited by the lack of suitable expression hosts and low productivity. In this study, Corynebacterium glutamicum ATCC13032, a GRAS strain that has been used extensively for the industrial-grade production of additives for foodstuffs, was employed to express a novel ß-glucosidase (MT619) from Microbacterium testaceum ATCC 15829 with high ginsenoside-transforming activity. A cellulose-binding module was additionally fused to the N-terminus of MT619 for immobilization on cellulose, which is an abundant and safe material. Via one-step immobilization, the fusion protein in cell lysates was efficiently immobilized on regenerated amorphous cellulose at a high density (maximum 984 mg/g cellulose), increasing the enzyme concentration by 286-fold. The concentrated and immobilized enzyme showed strong conversion activities against protopanaxadiol- and protopanaxatriol-type ginsenosides for the production of CK and F1. Using gram-scale ginseng extracts as substrates, the immobilized enzyme produced 7.59 g/L CK and 9.42 g/L F1 in 24 h. To the best of our knowledge, these are the highest reported product concentrations of CK and F1, and this is the first time that a recombinant enzyme has been immobilized on cellulose for the preparation of minor ginsenosides. This safe, convenient, and efficient production method could also be effectively exploited in the preparation of food-processing recombinant enzymes in the pharmaceutical, functional food, and cosmetics industries.

7.
Molecules ; 24(8)2019 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-31018484

RESUMO

Cutaneous wound healing is a well-orchestrated event in which many types of cells and growth factors are involved in restoring the barrier function of skin. In order to identify whether ginsenosides, the main active components of Panax ginseng, promote wound healing, the proliferation and migration activities of 15 different ginsenosides were tested by MTT assay and scratched wound closure assay. Among ginsenosides, gypenoside LXXV (G75) showed the most potent wound healing effects. Thus, this study aimed to investigate the effects of G75 on wound healing in vivo and characterize associated molecular changes. G75 significantly increased proliferation and migration of keratinocytes and fibroblasts, and promoted wound closure in an excision wound mouse model compared with madecassoside (MA), which has been used to treat wounds. Additionally, RNA sequencing data revealed G75-mediated significant upregulation of connective tissue growth factor (CTGF), which is known to be produced via the glucocorticoid receptor (GR) pathway. Consistently, the increase in production of CTGF was confirmed by western blot and ELISA. In addition, GR-competitive binding assay and GR translocation assay results demonstrated that G75 can be bound to GR and translocated into the nucleus. These results demonstrated that G75 is a newly identified effective component in wound healing.


Assuntos
Anti-Inflamatórios/farmacologia , Fator de Crescimento do Tecido Conjuntivo/genética , Fármacos Dermatológicos/farmacologia , Receptores de Glucocorticoides/genética , Ferida Cirúrgica/tratamento farmacológico , Cicatrização/efeitos dos fármacos , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/isolamento & purificação , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Fármacos Dermatológicos/química , Fármacos Dermatológicos/isolamento & purificação , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Regulação da Expressão Gênica , Ginsenosídeos/química , Ginsenosídeos/isolamento & purificação , Ginsenosídeos/farmacologia , Gynostemma/química , Humanos , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos ICR , Panax/química , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/farmacologia , Transporte Proteico , Receptores de Glucocorticoides/metabolismo , Transdução de Sinais , Pele/efeitos dos fármacos , Pele/lesões , Pele/metabolismo , Ferida Cirúrgica/genética , Ferida Cirúrgica/metabolismo , Ferida Cirúrgica/patologia , Cicatrização/fisiologia
8.
Int J Mol Sci ; 20(5)2019 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-30871042

RESUMO

The anthracycline antibiotic doxorubicin is commonly used antineoplastic drug in breast cancer treatment. Like most chemotherapy, doxorubicin does not selectively target tumorigenic cells with high proliferation rate and often causes serve side effects. In the present study, we demonstrated the cellular senescence and senescence associated secretory phenotype (SASP) of both breast tumor cell MDA-MB-231 and normal epithelial cell MCF-10A induced by clinical dose of doxorubicin (100 nM). Senescence was confirmed by flattened morphology, increased level of beta galactose, accumulating contents of lysosome and mitochondrial, and elevated expression of p16 and p21 proteins. Similarly, SASP was identified by highly secreted proteins IL-6, IL-8, GRO, GM-CSF, MCP-1, and MMP1 by antibody array assay. Reciprocal experiments, determined by cell proliferation and apoptosis assays and cell migration and cell invasion, indicated that SASP of MDA-MB-231 cell induces growth arrest of MCF-10A, whereas SASP of MCF-10A significantly stimulates the proliferation of MDA-MB-231. Interestingly, SASP from both cells powerfully promotes the cell migration and cell invasion of MDA-MB-231 cells. Treatment with the natural product ginsenoside Rh2 does not prevent cellular senescence or exert senolytic. However, SASP from senescent cells treated with Rh2 greatly attenuated the above-mentioned bystander effect. Altogether, Rh2 is a potential candidate to ameliorate this unwanted chemotherapy-induced senescence bystander effect.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Efeito Espectador/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Doxorrubicina/farmacologia , Células Epiteliais/efeitos dos fármacos , Ginsenosídeos/farmacologia , Apoptose/efeitos dos fármacos , Mama/efeitos dos fármacos , Mama/metabolismo , Neoplasias da Mama/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Humanos , Interleucina-6/metabolismo , Invasividade Neoplásica/patologia
9.
Int J Pharm ; 557: 66-73, 2019 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-30580088

RESUMO

Graphene oxide (GO) owns huge surface area and high drug loading capacity for aromatic molecules, such as doxorubicin (DOX). However, its biocompatibility is poor and it might agglomerate in physiological conditions. Chemical modification of GO with hydrophilicpolymer, especially PEGylation, was a common method to improve its biocompatibility. But the chemical modification of GO was complicated, and its drug loading capacity might be reduced because of the occupation of its functional groups. In this study, DOX-PEG polymers with different PEG molecular weights were synthesized to modify nano graphene oxide (NGO) to simultaneously realize the solubilization of NGO and the high loading capacity of DOX. The result showed that the drug release of NGO@DOX-PEG was pH sensitive. NIR irradiation could augment the drug release, cellular uptake, cytotoxicity and nuclear translocation of nanodrugs. Among the three kinds of nanodrugs, NGO@DOX-PEG5K was superior to others. It suggested that after conjugating with PEG, the bond between DOX-PEG and NGO was weakened, which resulted in a better drug release and treatment effect. In summary, the NIR and pH dual-responsive NGO@DOX-PEG nanodrugs were developed by noncovalent modification, and it demonstrated excellent biocompatibility and photochemical therapeutic effect, presenting a promising candidate for antitumor therapy, especially NGO@DOX-PEG5K.


Assuntos
Antibióticos Antineoplásicos/administração & dosagem , Doxorrubicina/administração & dosagem , Portadores de Fármacos/administração & dosagem , Grafite/administração & dosagem , Nanopartículas/administração & dosagem , Óxidos/administração & dosagem , Polietilenoglicóis/administração & dosagem , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/efeitos da radiação , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Doxorrubicina/química , Doxorrubicina/efeitos da radiação , Portadores de Fármacos/química , Portadores de Fármacos/efeitos da radiação , Liberação Controlada de Fármacos , Grafite/química , Grafite/efeitos da radiação , Células HeLa , Humanos , Luz , Nanopartículas/química , Nanopartículas/efeitos da radiação , Óxidos/química , Óxidos/efeitos da radiação , Fotoquimioterapia , Polietilenoglicóis/química , Polietilenoglicóis/efeitos da radiação , Espécies Reativas de Oxigênio/metabolismo , Solubilidade
10.
J Mater Chem B ; 6(31): 5080-5090, 2018 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-30245822

RESUMO

Nano-graphene oxide (NGO) has been proposed as a novel drug carrier. However, the poor biocompatibility and physiological stability as well as lack of cancer targeting capability have limited its further applications in cancer therapy. To solve this problem, we developed a novel nanohybrid of NGO/DOX@SPC-FA by first allowing soy phosphatidylcholine membrane (SPC) to encapsulate DOX-loaded NGO (NGO/DOX) and then modifying the SPC membrane with PEGylated lipid-FA conjugate to achieve the display of cancer targeting FA on the nanohybrid surface. The SPC membrane (mimicking cell membrane) enabled the resultant nanohybrids (NGO/DOX@SPC-FA) to exhibit good stability and biocompatibility, high drug loading capability, efficient cellular uptake, and controlled drug release. Moreover, compared with NGO/DOX and SPC-modified NGO/DOX (NGO/DOX@SPC), the FA-modified NGO/DOX@SPC nanohybrids (NGO/DOX@SPC-FA) could deliver NGO/DOX to cancer cells with improved delivery and killing efficacy due to the presence of FA targeting motifs on the surface. The NGO/DOX@SPC-FA nanohybrids were found to be internalized specifically by FA-positive cancer cells (Hela cells) through both macropinocytosis-directed engulfment and clathrin-dependent endocytosis, and then become localized into the lysosomes. In vivo biodistribution study showed that NGO/DOX@SPC-FA had a high tumor targeting ability because of the active targeting mechanism with folate modification. In vivo antitumor therapy study demonstrated NGO/DOX@SPC-FA could significantly inhibit tumour growth and prolong the survival time of mice. Our results suggest that NGO/DOX@SPC-FA, as a novel drug delivery system with high drug loading and targeted delivery efficiency, holds promise for future cancer therapy.

11.
Chem Biol Interact ; 283: 75-83, 2018 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-29412148

RESUMO

Senescence is one of the hallmarks of aging and identified as a potential therapeutic target in the treatment of aging and aging-related diseases. Senescent cells accumulate with age in a variety of human tissues where they develop a complex senescence-associated secretory phenotype (SASP). SASP in brain could contribute to age-related inflammation and chronic neurodegenerative diseases. We confirmed that senescent astrocytes express a characteristic of SASP in vitro by human cytokine antibody array. Ginsenoside F1 suppresses the SASP from astrocytes induced by d-galactose via suppressing p38MAPK-dependent NF-κB activity. A specific inhibitor of p38MAPK, SB203580 significantly decreased the secretion of IL-6 and IL-8, the major components of SASPs. Additionally, treatment of senescent astrocytes with NF-κB inhibitor, BAY 11-7092, also suppressed the secretion of IL-6 and IL-8, suggesting NF-κB was required for SASP. Importantly, conditioned media from senescent astrocytes promoted the migration of glioblastoma cells, such as U373-MG, U251-MG and U87-MG assessed by scratch wound healing. This migration was significantly decreased by F1 treatment in senescent astrocytes. Interestingly, IL-8, the main mediator regulating glioblastoma cell invasion, was suppressed in both transcriptional and protein level. Herein, we propose ginsenoside F1 as a potential therapeutic strategy for reducing the deleterious contribution of senescent astrocytes in aged brain and related diseases.


Assuntos
Senescência Celular/efeitos dos fármacos , Ginsenosídeos/farmacologia , Astrócitos/citologia , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Humanos , Imidazóis/farmacologia , Interleucina-6/análise , Interleucina-6/metabolismo , Interleucina-8/análise , Interleucina-8/metabolismo , NF-kappa B/antagonistas & inibidores , NF-kappa B/metabolismo , Fosforilação/efeitos dos fármacos , Piridinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
12.
J Hematol Oncol ; 11(1): 30, 2018 02 27.
Artigo em Inglês | MEDLINE | ID: mdl-29482582

RESUMO

BACKGROUND: We recently identified the human leukocyte immunoglobulin-like receptor B2 (LILRB2) and its mouse ortholog-paired Ig-like receptor (PirB) as receptors for several angiopoietin-like proteins (Angptls). We also demonstrated that PirB is important for the development of acute myeloid leukemia (AML), but exactly how an inhibitory receptor such as PirB can support cancer development is intriguing. RESULTS: Here, we showed that the activation of Ca (2+)/calmodulin-dependent protein kinases (CAMKs) is coupled with PirB signaling in AML cells. High expression of CAMKs is associated with a poor overall survival probability in patients with AML. Knockdown of CAMKI or CAMKIV decreased human acute leukemia development in vitro and in vivo. Mouse AML cells that are defective in PirB signaling had decreased activation of CAMKs, and the forced expression of CAMK partially rescued the PirB-defective phenotype in the MLL-AF9 AML mouse model. The inhibition of CAMK kinase activity or deletion of CAMKIV significantly slowed AML development and decreased the AML stem cell activity. We also found that CAMKIV acts through the phosphorylation of one of its well-known target (CREB) in AML cells. CONCLUSION: CAMKs are essential for the growth of human and mouse AML. The inhibition of CAMK signaling may become an effective strategy for treating leukemia.


Assuntos
Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Leucemia Mieloide Aguda/metabolismo , Animais , Carcinogênese/metabolismo , Carcinogênese/patologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Progressão da Doença , Humanos , Leucemia Mieloide Aguda/patologia , Camundongos Endogâmicos C57BL , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Receptores Imunológicos/metabolismo , Transdução de Sinais
13.
Drug Deliv ; 25(1): 112-121, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29260912

RESUMO

How to overcome drug resistance and prevent tumor metastasis is key to the success of malignant tumor therapy. In this paper, ADH-1 peptide-modified liposomes (A-LP) have been successfully constructed for restoring chemosensitivity and suppressing cancer cell migration. With a particle size of about 90 nm, this functionalized nanocarrier was loaded with fluorescent probe or paclitaxel (PTX). Cellular uptake studies showed that A-LP facilitated the delivery of anticancer drug to tumor cells undergoing EMT. Interestingly, this nanocarrier enhanced chemosensitivity by assessing the cell activity using CCK-8 assay. Further, the results of Wound scratch assay and Transwell migration assay showed the inhibition effect of this nanocarrier on tumor cell migration. Moreover, this nanocarrier exhibited significant tumor-targeting ability and anti-tumor efficacy in vivo. Collectively, A-LP might be a novel targeted drug delivery system to enhance the efficacy of chemotherapy and prevent tumor metastasis.


Assuntos
Antineoplásicos/administração & dosagem , Antineoplásicos/química , Movimento Celular/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Lipossomos/química , Oligopeptídeos/administração & dosagem , Oligopeptídeos/química , Peptídeos Cíclicos/administração & dosagem , Peptídeos Cíclicos/química , Animais , Linhagem Celular Tumoral , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos/métodos , Feminino , Humanos , Lipossomos/administração & dosagem , Células MCF-7 , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Nanopartículas/química , Paclitaxel/química , Tamanho da Partícula
14.
Int J Pharm ; 534(1-2): 71-80, 2017 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-28958879

RESUMO

The invasion and metastasis of tumour cells are closely correlated with poor prognosis of cancer patients. In this study, a CD44 and N-cadherin dual targeting drug delivery system based on mesoporous silica nanoparticles (MSNs) has been successfully constructed for inhibiting tumour cell invasion and metastasis. Amino modified MSN (MSN/NH2) was first synthesized and then functionalized with hyaluronic acid (HA) and ADH-1, constructing the carrier ADH-1-HA-MSN. Doxorubicin hydrochloride (DOX) was selected as a model anticancer drug. The prepared vector had a spherical shape with a narrow distribution of particle size. Flow cytometry and confocal microscopy studies showed that the modification with HA significantly enhanced CD44-mediated cellular uptake of this nanocarrier. ADH-1-HA-MSN/DOX exhibited higher cytotoxicity compared to non-ADH-1 modified counterparts. Of note, a transwell chamber assay demonstrated that the migration and invasion of tumour cells were markedly inhibited by ADH-1-HA-MSN/DOX. Furthermore, Western blotting analysis revealed that ADH-1-HA-MSN/DOX inhibited tumour cell invasion and metastasis by down-regulating N-cadherin expression. Taken together, these results indicated that ADH-1-HA-MSN might be a promising targeted drug delivery system for inhibiting cancer invasion and metastasis.


Assuntos
Antineoplásicos/administração & dosagem , Doxorrubicina/administração & dosagem , Nanopartículas/administração & dosagem , Invasividade Neoplásica/prevenção & controle , Metástase Neoplásica/tratamento farmacológico , Neoplasias/tratamento farmacológico , Dióxido de Silício/administração & dosagem , Células A549 , Caderinas/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos/métodos , Humanos , Receptores de Hialuronatos/metabolismo , Ácido Hialurônico/metabolismo , Concentração de Íons de Hidrogênio , Neoplasias/metabolismo , Tamanho da Partícula , Porosidade
15.
J Microbiol Biotechnol ; 27(9): 1559-1565, 2017 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-28633513

RESUMO

Naturally occurring ginsenoside F1 (20-O-ß-D-glucopyranosyl-20(S)-protopanaxatriol) is rare. Here, we produced gram-scale quantities of ginsenoside F1 from a crude protopanaxatriol saponin mixture comprised mainly of Re and Rg1 through enzyme-mediated biotransformation using recombinant ß-glucosidase (BgpA) cloned from a soil bacterium, Terrabacter ginsenosidimutans Gsoil 3082T. In a systematic step-by-step process, the concentrations of substrate, enzyme, and NaCl were determined for maximal production of F1. At an optimized NaCl concentration of 200 mM, the protopanaxatriol saponin mixture (25 mg/ml) was incubated with recombinant BgpA (20 mg/ml) for 3 days in a 2.4 L reaction. Following octadecylsilyl silica gel column chromatography, 9.6 g of F1 was obtained from 60 g of substrate mixture at 95% purity, as assessed by chromatography. These results represent the first report of gramscale F1 production via recombinant enzyme-mediated biotransformation.


Assuntos
Proteínas de Bactérias/metabolismo , Ginsenosídeos/metabolismo , Proteínas Recombinantes/metabolismo , beta-Glucosidase/metabolismo , Actinobacteria/enzimologia , Actinobacteria/genética , Proteínas de Bactérias/genética , Escherichia coli/genética , Ginsenosídeos/análise , Proteínas Recombinantes/genética , Sapogeninas/metabolismo , beta-Glucosidase/genética
16.
Molecules ; 22(5)2017 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-28534845

RESUMO

Minor ginsenosides, such as compound K, Rg3(S), which can be produced by deglycosylation of ginsenosides Rb1, showed strong anti-cancer effects. However, the anticancer effects of gypenoside LXXV, which is one of the deglycosylated shapes of ginsenoside Rb1, is still unknown due to the rarity of its content in plants. Here, we cloned and characterized a novel ginsenoside-transforming ß-glucosidase (BglG167b) derived from Microbacterium sp. Gsoil 167 which can efficiently hydrolyze gypenoside XVII into gypenoside LXXV, and applied it to the production of gypenoside LXXV at the gram-scale with high specificity. In addition, the anti-cancer activity of gypenoside LXXV was investigated against three cancer cell lines (HeLa, B16, and MDA-MB231) in vitro. Gypenoside LXXV significantly reduced cell viability, displaying an enhanced anti-cancer effect compared to gypenoside XVII and Rb1. Taken together, this enzymatic method would be useful in the preparation of gypenoside LXXV for the functional food and pharmaceutical industries.


Assuntos
Actinobacteria/química , Antineoplásicos Fitogênicos/metabolismo , Proteínas de Bactérias/metabolismo , Ginsenosídeos/metabolismo , beta-Glucosidase/metabolismo , Actinobacteria/enzimologia , Animais , Antineoplásicos Fitogênicos/isolamento & purificação , Antineoplásicos Fitogênicos/farmacologia , Proteínas de Bactérias/genética , Biotransformação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Clonagem Molecular , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Ginsenosídeos/isolamento & purificação , Ginsenosídeos/farmacologia , Gynostemma , Células HeLa , Humanos , Melanoma Experimental/tratamento farmacológico , Camundongos , Panax/química , Extratos Vegetais/biossíntese , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/farmacologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , beta-Glucosidase/genética
17.
PLoS One ; 12(4): e0176098, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28423055

RESUMO

The ginsenoside Rh2, a pharmaceutically active component of ginseng, is known to have anticancer and antitumor effects. However, white ginseng and red ginseng have extremely low concentrations of Rh2 or Rh2-Mix [20(S)-Rh2, 20(R)-Rh2, Rk2, and Rh3]. To enhance the production of food-grade ginsenoside Rh2, an edible enzymatic bioconversion technique was developed adopting GRAS host strains. A ß-glucosidase (BglPm), which has ginsenoside conversion ability, was expressed in three GRAS host strains (Corynebacterium glutamicum, Saccharomyces cerevisiae and Lactococus lactis) by using a different vector system. Enzyme activity in these three GRAS hosts were 75.4%, 11.5%, and 9.3%, respectively, compared to that in the E. coli pGEX 4T-1 expression system. The highly expressed BglPm_C in C. glutamicum can effectively transform the ginsenoside Rg3-Mix [20(S)-Rg3, 20(R)-Rg3, Rk1, Rg5] to Rh2-Mix [20(S)-Rh2, 20(R)-Rh2, Rk2, Rh3] using a scaled-up biotransformation reaction, which was performed in a 10-L jar fermenter at pH 6.5/7.0 and 37°C for 24 h. To our knowledge, this is the first report in which 50 g of PPD-Mix (Rb1, Rb2, Rb3, Rc, and Rd) as a starting substrate was converted to ginsenoside Rg3-Mix by acid heat treatment and then 24.5-g Rh2-Mix was obtained by enzymatic transformation of Rg3-Mix through by BglPm_C. Utilization of this enzymatic method adopting a GRAS host could be usefully exploited in the preparation of ginsenoside Rh2-Mix in cosmetics, functional food, and pharmaceutical industries, thereby replacing the E. coli expression system.


Assuntos
Proteínas de Bactérias/genética , Proteínas Fúngicas/genética , Ginsenosídeos/metabolismo , Microbiologia Industrial/métodos , beta-Glucosidase/genética , Proteínas de Bactérias/metabolismo , Biotransformação , Clonagem Molecular , Corynebacterium glutamicum/enzimologia , Corynebacterium glutamicum/genética , Escherichia coli/enzimologia , Escherichia coli/genética , Proteínas Fúngicas/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Ginsenosídeos/química , Concentração de Íons de Hidrogênio , Cinética , Lactococcus lactis/enzimologia , Lactococcus lactis/genética , Peso Molecular , Panax/química , Engenharia de Proteínas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/genética , Temperatura Ambiente , beta-Glucosidase/metabolismo
18.
Biomaterials ; 121: 55-63, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28081459

RESUMO

A Sleeping Beauty (SB) transposon system is made of a transposon plasmid (containing gene encoding a desired functional or therapeutic protein) and a transposase plasmid (encoding an enzyme capable of cutting and pasting the gene into the host cell genome). It is a kind of natural, nonviral gene delivery vehicle, which can achieve efficient genomic insertion, providing long-term transgenic expression. However, before the SB transposon system could play a role in promoting gene expression, it has to be delivered efficiently first across cell membrane and then into cell nuclei. Towards this end, we used a nanoparticle-like lipid-based protocell, a closed bilayer of the neutral lipids with the DNA encapsulated inside, to deliver the SB transposon system to cancer cells. The SB transposon system was amplified in situ inside the protocells by a polymerase chain reaction (PCR) process, realizing more efficient loading and delivery of the target gene. To reach a high transfection efficiency, we introduced two targeting moieties, folic acid (FA) as a cancer cell-targeting motif and Dexamethasone (DEX) as a nuclear localization signaling molecule, into the protocells. As a result, the FA enabled the modified targeting protocells to deliver the DNA into the cancer cells with an increased efficiency and the DEX promoted the DNA to translocate to cell nuclei, eventually leading to the increased chromosome insertion efficiency of the SB transposon. In vivo study strongly suggested that the transfection efficiency of FA-modified protocells in the tumor tissue was much higher than that in other tissues, which was consistent with the in vitro results. Our studies implied that with the targeting ligand modification, the protocells could be utilized as an efficient targeting gene carrier. Since the protocells were made of neutral lipids without cationic charges, the cytotoxicity of protocells was significantly lower than that of traditional cationic gene carriers such as cationic liposomes and polyethylenimine, enabling the protocells to be employed in a wider dosage range in gene therapy. Our work shows that the protocells are a promising gene carrier for future clinical applications.


Assuntos
Células Artificiais/química , Terapia Genética/métodos , Nanocápsulas/química , Neoplasias Experimentais/genética , Neoplasias Experimentais/terapia , Plasmídeos/administração & dosagem , Transposases/genética , Animais , Linhagem Celular Tumoral , Elementos de DNA Transponíveis/genética , Feminino , Marcação de Genes/métodos , Células HeLa , Humanos , Lipídeos/química , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Nanocápsulas/administração & dosagem , Plasmídeos/genética , Reação em Cadeia da Polimerase/métodos , Transfecção/métodos , Transposases/administração & dosagem , Resultado do Tratamento
19.
Biochem Biophys Res Commun ; 478(4): 1713-9, 2016 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-27596969

RESUMO

Topical use of ginsenosides, the major bioactive substances in Panax ginseng, has been used for the treatment of irritated skin complaints. However, the protective mechanisms of ginsenosides remain unclear. In the present study, we investigated the anti-inflammatory role of ginsenoside F2 (GF2) on the skin inflammation. To induce irritant dermatitis, 12-O-tetradecanoylphorbol-13-acetate (TPA) was applied on the surface of the mouse ears with or without treatments of GF2 and dexamethasone for 24 h. Protective effects of GF2 on edema and inflammation were assessed by measuring ear thickness, weights of skin punch, and inflammatory responses. In gross findings, treatments with GF2 significantly decreased skin thickness and weight compared to those of TPA-treated groups, which was comparable with the protective effects of dexamethasone. In addition, expression of inflammatory mediators was remarkably reduced in GF2-treated ears compared to that of vehicle-treated ears of mice. Interestingly, immunohistochemistry and flow cytometry analyses revealed that TPA treatment significantly increased infiltration of interleukin-17 (IL-17) producing dermal γδ T cells, while frequencies of γδ T cells was decreased by GF2 treatment, subsequently ameliorating inflammation in skin. Concomitantly, TPA-mediated skin inflammation was significantly ameliorated in IL-17A knock out mice. Furthermore, GF2 treatment inhibited infiltration and generation of reactive oxygen species (ROS) of neutrophils in damaged ears compared with vehicle-treated mice. These results clearly suggest that GF2 treatment ameliorates TPA-induced dermal inflammation by inhibiting production of IL-17 and ROS in γδ T cells and neutrophils, respectively. Therefore, as a natural compound, application of GF2 may be a novel therapeutic approach for treating skin inflammation.


Assuntos
Dermatite/prevenção & controle , Orelha Externa/efeitos dos fármacos , Edema/prevenção & controle , Ginsenosídeos/farmacologia , Administração Cutânea , Animais , Dermatite/etiologia , Dermatite/metabolismo , Orelha Externa/metabolismo , Orelha Externa/patologia , Edema/induzido quimicamente , Edema/metabolismo , Citometria de Fluxo , Expressão Gênica/efeitos dos fármacos , Ginsenosídeos/administração & dosagem , Interleucina-17/genética , Interleucina-17/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Substâncias Protetoras/administração & dosagem , Substâncias Protetoras/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Receptores de Antígenos de Linfócitos T gama-delta/genética , Receptores de Antígenos de Linfócitos T gama-delta/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Acetato de Tetradecanoilforbol/toxicidade
20.
Nat Cell Biol ; 17(5): 665-77, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25915125

RESUMO

Conventional strategies are not particularly successful in the treatment of leukaemia, and identification of signalling pathways crucial to the activity of leukaemia stem cells will provide targets for the development of new therapies. Here we report that certain receptors containing the immunoreceptor tyrosine-based inhibition motif (ITIM) are crucial for the development of acute myeloid leukaemia (AML). Inhibition of expression of the ITIM-containing receptor LAIR1 does not affect normal haematopoiesis but abolishes leukaemia development. LAIR1 induces activation of SHP-1, which acts as a phosphatase-independent signalling adaptor to recruit CAMK1 for activation of downstream CREB in AML cells. The LAIR1-SHP-1-CAMK1-CREB pathway sustains the survival and self-renewal of AML stem cells. Intervention in the signalling initiated by ITIM-containing receptors such as LAIR1 may result in successful treatment of AML.


Assuntos
Leucemia Mieloide Aguda/metabolismo , Células-Tronco Neoplásicas/metabolismo , Receptores Imunológicos/metabolismo , Adulto , Idoso , Motivos de Aminoácidos , Animais , Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina/genética , Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Feminino , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Células-Tronco Neoplásicas/patologia , Proteína Tirosina Fosfatase não Receptora Tipo 6/genética , Proteína Tirosina Fosfatase não Receptora Tipo 6/metabolismo , Interferência de RNA , Receptores Imunológicos/deficiência , Receptores Imunológicos/genética , Transdução de Sinais , Fatores de Tempo , Transfecção , Células Tumorais Cultivadas , Adulto Jovem
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