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1.
Biotechnol Prog ; : e3419, 2024 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-38247123

RESUMO

Extracellular vesicles (EVs) derived from mesenchymal stromal cells (MSCs) have shown increasing therapeutic potential in the last years. However, large production of EV is required for therapeutic purposes. Thereby, scaling up MSC cultivation in bioreactors is essential to allow culture parameters monitoring. In this study, we reported the establishment of a scalable bioprocess to produce MSC-EV in suspension cultures using spinner flasks and human collagen-coated microcarriers (3D culture system). We compared the EV production in this 3D culture system with the standard static culture using T-flasks (2D culture system). The EV produced in both systems were characterized and quantify by western blotting and nanoparticle tracking analysis. The presence of the typical protein markers CD9, CD63, and CD81 was confirmed by western blotting analyses for EV produced in both culture systems. The cell fold-increase was 5.7-fold for the 3D culture system and 4.6-fold for the 2D culture system, signifying a fold-change of 1.2 (calculated as the ratio of fold-increase 3D to fold-increase 2D). Furthermore, it should be noted that the total cell production in the spinner flask cultures was 4.8 times higher than that in T-flask cultures. The total cell production in the spinner flask cultures was 5.2-fold higher than that in T-flask cultures. While the EV specific production (particles/cell) in T-flask cultures (4.40 ± 1.21 × 108 particles/mL, p < 0.05) was higher compared to spinner flask cultures (2.10 ± 0.04 × 108 particles/mL, p < 0.05), the spinner flask culture system offers scalability, making it capable of producing enough MSC-EV at a large scale for clinical applications. Therefore, we concluded that 3D culture system evaluated here serves as an efficient transitional platform that enables the scaling up of MSC-EV production for therapeutic purposes by utilizing stirred tank bioreactors and maintaining xeno-free conditions.

2.
Macromol Biosci ; 19(10): e1900117, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31402631

RESUMO

The development of polymers with low toxicity and efficient gene delivery remains a significant barrier of nonviral gene therapy. Modification and tuning of chemical structures of carriers is an attractive strategy for efficient nucleic acid delivery. Here, polyplexes consisting of plasmid DNA (pDNA) and dodecylated or non-dodecylated polysuccinimide (PSI)-based polycations are designed, and their transfection ability into HeLa cells is investigated by green fluorescent protein (GFP) expressing cells quantification. All cationic polymers show lower cytotoxicity than those of branched polyethyleneimine (bPEI). PSI and bPEI-based polyplexes have comparable physicochemical properties such as size and charge. Interestingly, a strong interaction between dodecylated polycations and pDNA caused by the hydrophobic moiety is observed in dodecylated PSI derivatives. Moreover, the decrease of GFP expression is associated with lower dissociation of pDNA from polyplexes according to the heparin displacement assay. Besides, a hydrophobization of PSI cationic derivatives with dodecyl side chains can modulate the integrity of polyplexes by hydrophobic interactions, increasing the binding between the polymer and the DNA. These results provide useful information for designing polyplexes with lower toxicity and greater stability and transfection performance.


Assuntos
Ácido Aspártico/análogos & derivados , DNA , Vetores Genéticos , Plasmídeos , Transfecção , Ácido Aspártico/química , Ácido Aspártico/farmacologia , DNA/química , DNA/farmacologia , Vetores Genéticos/química , Vetores Genéticos/farmacologia , Células HeLa , Humanos , Plasmídeos/química , Plasmídeos/farmacologia
3.
Photodiagnosis Photodyn Ther ; 24: 262-273, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30290231

RESUMO

The objective of this study was to develop and characterize lipid nanoparticles (LNs) containing chloroaluminum phthalocyanine (ClAlPc) to reduce the aggregation of the drug and improve its skin penetration and its antitumor effect. LNs were prepared and characterized by using stearic acid (SA) as solid lipid and oleic acid (OA) as liquid lipid in different proportions. in vitro and in vivo skin penetration was evaluated using modified Franz diffusion cells and fluorescence microscopy, respectively. in vitro biocompatibility and Photodynamic Therapy (PDT) were performed using L929-fibroblasts cell line and A549 cancer cell line and melanoma BF16-F10, respectively. OA promoted the increase in the encapsulation efficiency and drug loading, reaching values of 95.8% and 4%, respectively. The formulation with 40% OA (NLC 40) showed a significantly higher (p < 0.01) amount of drug retained in the skin compared to other formulations. All formulations developed were considered biocompatible. PDT evidenced the antitumor efficacy of NLC 40 with reduced cell viability for approximately 10% of cancer cells, demonstrating that the presence of OA in the NLC seems to potentialize this antitumor effect. PDT in BF16-F10 melanoma using NLC 40 resulted in a reduction in mean cell viability of approximately 99%. According to the results obtained, the systems developed may be promising for the incorporation of ClAlPc in the treatment of skin cancer by photodynamic therapy.


Assuntos
Indóis/farmacologia , Nanopartículas/química , Compostos Organometálicos/farmacologia , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/farmacologia , Absorção Cutânea/efeitos dos fármacos , Células A549 , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Fibroblastos , Humanos , Indóis/administração & dosagem , Camundongos , Ácido Oleico/química , Compostos Organometálicos/administração & dosagem , Tamanho da Partícula , Fármacos Fotossensibilizantes/administração & dosagem , Ácidos Esteáricos/química , Suínos
4.
Eur J Pharm Sci ; 111: 121-132, 2018 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-28951120

RESUMO

In vitro skin permeation/penetration studies may be affected by many sources of variation. Herein, we aimed to investigate the major critical procedures of in vitro skin delivery studies. These experiments were performed with model drugs according to official guidelines. The influence of skin source on penetration studies was studied as well as the use of a cryopreservation agent on skin freezing evaluated by transepidermal water loss, electrical resistance, permeation/penetration profiles and histological changes of the skin. The best condition for tape stripping procedure was validated through the evaluation of the distribution of corneocytes, mass of stratum corneum (SC) removed and amount of protein removed using finger pressure, a 2kg weight and a roller. The interchangeability of the tape stripping procedures followed by the epidermis and dermis homogenate and the micrometric horizontal cryostat skin sectioning methods were also investigated, besides the effect of different formulations. Noteworthy, different skin sources were able to ensure reliable interchangeability for in vitro permeation studies. Furthermore, an increased penetration was obtained for stored frozen skin compared to fresh skin, even with the addition of a cryoprotectant agent. The best method for tape stripping was the finger pressure followed by the addition of a propylene glycol solvent leading to better SC removal. Finally, no significant difference was found in skin penetration studies performed by different methods suggesting their possible interchangeability.


Assuntos
Estradiol/farmacocinética , Fluoresceínas/farmacocinética , Nicotina/farmacocinética , Absorção Cutânea , Pele/metabolismo , Administração Cutânea , Animais , Estradiol/administração & dosagem , Fluoresceínas/administração & dosagem , Técnicas In Vitro , Masculino , Camundongos Pelados , Modelos Animais , Nicotina/administração & dosagem , Serpentes , Suínos
5.
Mol Pharm ; 14(5): 1681-1690, 2017 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-28291360

RESUMO

An effective short interfering RNA (siRNA) delivery system protects the siRNA from degradation, facilitates its cellular uptake, and promotes its release into the cytoplasm. Local administration of siRNA presents advantages over systemic administration, such as the possibility to use lower doses and allow local and sustained release. In this context, in situ solidifying organogels based on monoglycerides (MO), polyethylenimine (PEI), propylene glycol (PG) and tris buffer are an attractive strategy for intratumoral delivery of siRNA. In this study, precursor fluid formulation (PFF) composed of MO/PEI/PG/tris buffer at 7.85:0.65:76.5:15 (w/w/w/w) was used to deliver siRNA to tumor cells. The internal structure of the gel obtained from PFF was characterized using small angle X-ray scattering (SAXS). In addition, its ability to complex siRNA, protect it from degradation, and functionally deliver it to tumor cells was investigated. Moreover, in vivo gel formation following intratumoral injection was evaluated. The gel formed in excess water from PFF was found to comprise a mixture of hexagonal and cubic phases. The system was able to complex high amounts of siRNA, protect it from degradation, promote siRNA internalization, and induce gene silencing in vitro in a variety of tumor cell lines. Moreover, a gel formed in situ following intratumoral injection in a murine xenograft model. In conclusion, PFF is a potential delivery system for local and sustained delivery of siRNA to tumor tissue after intratumoral administration.


Assuntos
Inativação Gênica/fisiologia , Cristais Líquidos/química , Monoglicerídeos/química , Polietilenoimina/química , Propilenoglicol/química , RNA Interferente Pequeno/genética
6.
Ther Deliv ; 6(9): 1053-61, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26458111

RESUMO

To achieve an efficient skin penetration of most compounds it is necessary to overcome the barrier function of the skin, provided mainly (but not only) by the stratum corneum. Among various strategies used or studied to date, chemical penetration enhancers are the most frequently employed with one of the longest histories of use. There is a multitude of agents described as penetration enhancers, and they present varying properties and structures. In this manuscript, we aim to provide a brief overview of traditional enhancers and some of their properties, focusing on the benefits of combination of chemical enhancers and on selected novel compounds that have shown promise to increase drug delivery into/across the skin.


Assuntos
Excipientes/química , Absorção Cutânea/efeitos dos fármacos , Administração Cutânea , Animais , Sistemas de Liberação de Medicamentos , Excipientes/efeitos adversos , Humanos
7.
Eur J Pharm Biopharm ; 88(1): 28-33, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24509413

RESUMO

The glycoalkaloids solasonine (SN) and solamargine (SM) have been studied for their antiparasitic, antifungal, and anticancer properties, especially in vitro and in vivo against non-melanoma skin cancer. Thus, the alkaloidic extract of Solanum lycocarpum, which contains approximately 45% each of SN and SM, was used to define the best experimental conditions for in vitro and in vivo assays. The in vitro assays were performed with the Franz cell diffusion porcine skin model to evaluate the effects of different pHs and the presence of monoolein, ethoxydiglycol or ethanol penetration enhancers on the skin penetration and retention of SN and SM after 3, 6, 9 and 12h of exposure. The in vivo assay was performed on hairless mice with the formulation selected in the in vitro assays. The results showed that pH 6.5 was optimal for SM penetration. The formulation containing 5% alkaloidic extract, 5% propylene glycol, 5% monoolein and a hydroxyethyl cellulose gel base (Natrosol) (pH 6.5) was optimal for the delivery of SN and SM into the skin, and this formulation is potentially useful for the topical therapy of several skin disorders.


Assuntos
Administração Tópica , Frutas/química , Extratos Vegetais/farmacologia , Alcaloides de Solanáceas/administração & dosagem , Solanum/química , Alcaloides/química , Animais , Antifúngicos/administração & dosagem , Antineoplásicos/farmacologia , Antiparasitários/administração & dosagem , Celulose/análogos & derivados , Celulose/química , Difusão , Etanol/química , Glicerídeos/química , Glicóis/química , Concentração de Íons de Hidrogênio , Técnicas In Vitro , Masculino , Camundongos , Pele/efeitos dos fármacos , Suínos
8.
J Liposome Res ; 23(1): 47-53, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23363304

RESUMO

Drug administration through the transdermal route has optimized for the comfort of patients and easy application. However, the main limitation of transdermal drug delivery is the impermeability of the human skin. Recent advances on improvement of drug transport through the skin include elastic liposomes as a penetration enhancer. Entrapment of ferrofluids in the core of liposomes produces magnetoliposomes, which can be driven by a high-gradient magnetic field. The association of both strategies could enhance the penetration of elastic liposomes. This work relies on the preparation and characterization of elastic-magnetic liposomes designed to permeate through the skin. The incorporation of colloidal magnetite and the elastic component, octaethylene glycol laurate (PEG-8-L), in the structure of liposomes were evaluated. The capability of the elastic magnetoliposomes for permeation through nanopores of two stacked polycarbonate membranes was compared to conventional and elastic liposomes. Magnetite incorporation was dependent on vesicle diameter and size distribution as well as PEG-8-L incorporation into liposomes, demonstrating the capability of the fluid bilayer to accommodate the surfactant without disruption. On the contrary, PEG-8-L incorporation into magnetoliposomes promoted a decrease of average diameter and a lower PEG-8-L incorporation percentage as a result of reduction on the fluidity of the bilayer imparted by iron incorporation into the lipid structure. Elastic liposomes demonstrated an enhancement of the deformation capability, as compared with conventional liposomes. Conventional and elastic magnetoliposomes presented a reduced capability for deformation and permeation.


Assuntos
Coloides/química , Lipossomos/administração & dosagem , Surfactantes Pulmonares/administração & dosagem , Administração Cutânea , Coloides/administração & dosagem , Óxido Ferroso-Férrico/administração & dosagem , Óxido Ferroso-Férrico/química , Humanos , Lauratos/administração & dosagem , Lauratos/química , Lipossomos/química , Campos Magnéticos , Farmacocinética , Polietilenoglicóis/administração & dosagem , Polietilenoglicóis/química , Surfactantes Pulmonares/química
9.
Drug Dev Ind Pharm ; 38(3): 259-63, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-21854223

RESUMO

Conventional treatment of tuberculosis (TB) demands a long course therapy (6 months), known to originate multiple drug resistant strains (MDR-TB), which emphasizes the urgent need for new antituberculous drugs. The purpose of this study was to investigate a novel treatment for TB meant to improve patient compliance by reducing drug dosage frequency. Polymeric microparticles containing the synthetic analogue of neolignan, 1-phenyl-2-phenoxiethanone (LS-2), were obtained by a method of emulsification and solvent evaporation and chemically characterized. Only representative LS-2-loaded microparticles were considered for further studies involving experimental murine TB induced by Mycobacterium tuberculosis H37Rv ATCC 27294. The LS-2-loaded microparticles were spherical in shape, had a smooth wall and showed an encapsulation efficiency of 93% in addition to displaying sustained release. Chemotherapeutic potential of LS-2 entrapped in microparticles was comparable to control groups. These findings are encouraging and indicate that LS-2-loaded microparticles are a potential alternative to conventional chemotherapy of TB.


Assuntos
Antibacterianos/administração & dosagem , Portadores de Fármacos , Lignanas/administração & dosagem , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose Pulmonar/tratamento farmacológico , Animais , Materiais Biocompatíveis , Modelos Animais de Doenças , Farmacorresistência Bacteriana , Humanos , Ácido Láctico , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Microesferas , Modelos Animais , Tamanho da Partícula , Cooperação do Paciente , Ácido Poliglicólico , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Fatores de Tempo
10.
Microsc Res Tech ; 75(2): 145-52, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21761496

RESUMO

BACKGROUND: The aim of this work was to study the new bone tissue formation after bone morphogenetic protein type 2 (rhBMP-2) and P-1 application, using 5 and 10 µg of each, combined to a material carrier, in critical bone defects. METHODS: It was used 70 Wistar rats (male, ∼250 g) that were divided in 10 groups with seven animals on each. Groups are the following: critical bone defect only, pure monoolein gel, 5 µg of pure P-1, 5 µg of pure rhBMP-2, 5 µg of P-1/monoolein gel, 5 µg of rhBMP-2/monoolein gel, 10 µg of pure P-1, 10 µg of pure rhBMP-2, 10 µg of P-1/monoolein gel, 10 µg of rhBMP-2/monoolein gel. Animals were sacrificed after 4 weeks of the surgical procedure and the bone samples were submitted to histological, histomorphometrical, and immunohistochemical evaluations. RESULTS: Animals treated with pure P-1 protein, in both situations with 5 µg and 10 µg, had no significant difference (P > 0.05) for new bone formation; other groups treated with 10 µg were statistically significant (P < 0.05) among themselves and when compared with groups in which it was inserted the monoolein gel or critical bone defect only (P < 0.05). In the group involving the 10 µg rhBMP-2/monoolein gel association, it was observed an extensive bone formation, even when compared with the same treatment without the gel carrier. CONCLUSION: Using this experimental animal model, more new bone tissue was found when it was inserted the rhBMP-2, especially when this protein was combined to the vehicle, and this process seems to be dose dependent.


Assuntos
Proteína Morfogenética Óssea 2/farmacologia , Regeneração Óssea/efeitos dos fármacos , Látex/química , Proteínas de Plantas/farmacologia , Fator de Crescimento Transformador beta/farmacologia , Animais , Proteína Morfogenética Óssea 2/administração & dosagem , Relação Dose-Resposta a Droga , Portadores de Fármacos/administração & dosagem , Glicerídeos/administração & dosagem , Hevea/química , Humanos , Imuno-Histoquímica , Masculino , Modelos Animais , Proteínas de Plantas/administração & dosagem , Ratos , Ratos Wistar , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/farmacologia , Crânio/efeitos dos fármacos , Crânio/lesões , Fator de Crescimento Transformador beta/administração & dosagem
11.
J Microencapsul ; 26(3): 243-50, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-18923964

RESUMO

Chitosan treated alginate microparticles were prepared with the purpose of incorporating all-trans retinoic acid (ATRA) using an inexpensive, simple and fast method, enhancing dermal localization and sustaining the release of ATRA into the skin. Microparticles characterization, drug-polymer interaction, release profile and in vitro skin retention were investigated. Microparticles presented spherical shape and drug loading capacity of 47%. The drug content of these microparticles was affected by ATRA concentration and by the solvent used and it was more weakly affected by chitosan concentration. The release of ATRA was also affected by chitosan concentration. Microparticles prepared with 0.4% chitosan (w/w) resulted in drug release with a more sustained profile. The results of in vitro retention studies showed that chitosan treated alginate microparticles decreased the drug retention in the stratum corneum (SC), where occur the skin irritation, but maintained the ATRA concentration in the deeper skin layers, where occur the pathologies treated with ATRA. Then, the microparticles developed in this work can be a good candidate to improve the topical therapy with retinoid.


Assuntos
Alginatos/química , Quitosana/química , Ceratolíticos/administração & dosagem , Pele/metabolismo , Tretinoína/administração & dosagem , Administração Tópica , Animais , Ácido Glucurônico/química , Ácidos Hexurônicos/química , Ceratolíticos/farmacocinética , Microesferas , Suínos , Tretinoína/farmacocinética
12.
Micron ; 39(7): 952-9, 2008 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-18083037

RESUMO

The purpose of this work was to evaluate the new bone tissue, comparing two different carriers for rhBMP-2, monoolein and chitosan gels, using the decortication and nondecorticatication surgical technique in rat mandibles, evaluated by histomorphometrical method. It was used 56 male Wistar rats (300 g), divided into 8 groups according to the rhBMP-2 carrier used, monoolein or chitosan gels; surgical technique, bone decortication or nondecortication; and period of time, 3 or 6 weeks until the sacrifice by perfusion. Results obtained in this study showed that the rhBMP-2/monoolein and rhBMP-2/chitosan used in this experimental model was able to induce osteogenesis, contributing to the bone healing process. The bone repair process was time dependent, so that at 6 weeks there was an improved amount of new bone in relation to 3 weeks, considered each analyzed group, and the decortication was able to expose the bone marrow and speed up the bone healing process, which was showed by histomorphometrical methods. Both of carriers were capable to adapt to the bone surgical area, according to the clinical observations, and had favorable properties in relation to protein releasing, revealed by the amount of new bone tissue found in the histological analysis.


Assuntos
Proteínas Morfogenéticas Ósseas/farmacologia , Quitosana/metabolismo , Portadores de Fármacos/metabolismo , Géis/química , Glicerídeos/metabolismo , Osteogênese/efeitos dos fármacos , Proteínas Recombinantes/farmacologia , Fator de Crescimento Transformador beta/farmacologia , Animais , Proteína Morfogenética Óssea 2 , Quitosana/farmacologia , Modelos Animais de Doenças , Portadores de Fármacos/farmacologia , Glicerídeos/farmacologia , Humanos , Masculino , Mandíbula/citologia , Mandíbula/cirurgia , Distribuição Aleatória , Ratos , Ratos Wistar
13.
Int J Pharm ; 310(1-2): 203-12, 2006 Mar 09.
Artigo em Inglês | MEDLINE | ID: mdl-16439076

RESUMO

Cryo-transmission electron microscopy (cryo-TEM) was used to image the microstructure in dilute sonicated dispersions of monoolein and oleic acid. The aim of the study was to explore how different experimental parameters, such as sample composition, total lipid concentration, pH, and ageing affect the phase behaviour and aggregate structure. Our investigations show that a rich variety of lamellar and non-lamellar structures, including liposomes and particles of cubic and inverted hexagonal phase, may form depending on the experimental conditions. The results are analyzed and discussed in relation to existing phase diagrams and earlier investigations concerning phase- and structural behaviour in monoolein/oleic acid/water systems.


Assuntos
Coloides/química , Glicerídeos/química , Ácido Oleico/química , Microscopia Crioeletrônica , Concentração de Íons de Hidrogênio , Lipossomos , Cristais Líquidos , Microscopia Eletrônica de Transmissão/métodos , Sonicação , Fatores de Tempo , Água/química
14.
RBCF, Rev. bras. ciênc. farm. (Impr.) ; 37(2): 189-196, maio-ago. 2001. graf
Artigo em Português | LILACS | ID: lil-314044

RESUMO

Géis de fase cúbica de monoleína e água têm sido propostos como sistemas de liberação de fármacos com diferentes características, incluindo moléculas protéicas e peptídicas. O presente trabalho estudou a incorporação de lactoferrina, usada como molécula protéica modelo em várias concentrações em géis de fase cúbica de monoleína e água. A influência da lactoferrina nos géis de monoleína/água foi avaliada por microscopia de luz polarizada e estudo de suas propriedades reológicas, asssim como estudos de liberação in vitro foram realizados. A fase cúbica foi observada na presença da lactoferrina sendo que esta proteína não modificou as transições de fase dos géis, quando observados ao microscópio de luz polarizada...


Assuntos
Géis/farmacocinética , Técnicas In Vitro , Lactoferrina , Lipídeos/biossíntese , Preparações de Ação Retardada/farmacocinética , Preparações Farmacêuticas/análise , Microscopia de Polarização/métodos , Química Farmacêutica/métodos , Reologia
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