RESUMO
Fundamental to the design and development of nanoparticles for applications in nanomedicine is a detailed understanding of their biologic fate and potential toxic effects. Transient receptor potential (TRP) ion channels are a large superfamily of cation channels with varied physiologic functions. This superfamily is classified into six related subfamilies: TRP canonical, TRP vanilloid (TRPV), TRP melastatin (TRPM), TRP ankyrin (TRPA), TRP polycystin, and TRP mucolipin. TRPA1, TRPM2, and TRPM8 are nonselective Ca2+-permeable cation channels which regulate calcium pathways under oxidative stress, whereas TRPV4 can be activated by oxidative, osmotic, and thermal stress as well as different fatty acid metabolites. Using a series of well characterized silica nanoparticles with variations in size (approximately 50-350 nm in diameter) and porosity, as well as cationic and anionic poly(amido amine) (PAMAM) dendrimers of similar size, we examined the toxicity of these nanoparticles to human embryonic kidney-293 cells overexpressing different TRP channels. The data show that the toxicity of mesoporous silica nanoparticles was influenced by expression of the TRPA1 and TRPM2 channels, whereas the toxicity of smaller nonporous silica nanoparticles was only affected by TRPM8. Additionally, TRPA1 and TRPM2 played a role in the cytotoxicity of cationic dendrimers, but not anionic dendrimers. TRPV4 did not seem to play a significant role in silica nanoparticle or PAMAM toxicity.
Assuntos
Dendrímeros/toxicidade , Nanopartículas/toxicidade , Dióxido de Silício/toxicidade , Canais de Potencial de Receptor Transitório/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células HEK293 , Humanos , Porosidade , Canal de Cátion TRPA1/efeitos dos fármacos , Canais de Cátion TRPM/efeitos dos fármacosRESUMO
There is a limited amount of information available on gene expression regulation of macrophages in response to changing the time of exposure, concentration, and physicochemical properties of nanomaterials. In this study, RAW264.7 macrophages were treated with spherical nonporous and mesoporous silica nanoparticles of similar size at different incubation times and concentrations. RNA-sequencing was used to study transcriptional profiles. Bioinformatics analyses, functional annotation clustering, and network analyses were employed to understand signaling pathways of cellular response as a function of porosity, incubation time, and concentration. Porosity introduced drastic changes to the genomic response of macrophages at equitoxic concentrations and incubation times. Direct relations between increases in time and concentration with an increased number of differentially expressed genes were observed.
Assuntos
Macrófagos/efeitos dos fármacos , Nanopartículas/química , Dióxido de Silício/farmacologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica/efeitos dos fármacos , Camundongos , Nanopartículas/administração & dosagem , Porosidade , Células RAW 264.7 , RNA-Seq , Transdução de Sinais/efeitos dos fármacos , Dióxido de Silício/químicaRESUMO
Amorphous silica nanoparticles (SNPs) are widely used in biomedical applications and consumer products. Little is known, however, about their genotoxicity and potential to induce gene expression regulation. Despite recent efforts to study the underlying mechanisms of genotoxicity of SNPs, inconsistent results create a challenge. A variety of factors determine particle-cell interactions and underlying mechanisms. Further, high-throughput studies are required to carefully assess the impact of silica nanoparticle physicochemical properties on induction of genotoxic response in different cell lines and animal models. In this article, we review the strategies available for evaluation of genotoxicity of nanoparticles (NPs), survey current status of silica nanoparticle gene alteration and genotoxicity, discuss particle-mediated inflammation as a contributing factor to genotoxicity, identify existing gaps and suggest future directions for this research.
Assuntos
Nanopartículas/química , Nanopartículas/toxicidade , Dióxido de Silício/química , Animais , Humanos , Inflamação/metabolismo , Inflamação/fisiopatologia , Relação Estrutura-AtividadeRESUMO
Careful evaluation of the toxicological response of engineered nanomaterials (ENMs) as a function of physicochemical properties can aid in the design of safe platforms for biomedical applications including drug delivery. Typically, in vitro ENM cytotoxicity assessments are performed under conventional static cell culture conditions. However, such conditions do not take into account the sedimentation rate of ENMs. Herein, we synthesized four types of similar size silica nanoparticles (SNPs) with modified surface roughness, charge, and density and characterized their cytotoxicity under static and dynamic conditions. Influence of particle density on sedimentation and diffusion velocities were studied by comparing solid dense silica nanoparticles of approximately 350 nm in diameter with hollow rattle shape particles of similar size. Surface roughness and charge had negligible impact on sedimentation and diffusion velocities. Lower cellular uptake and toxicity was observed by rattle particles and under dynamic conditions. Dosimetry of ENMs are primarily reported by particle concentration, assuming homogeneous distribution of nanoparticles in cell culture media. However, under static conditions, nanoparticles tend to sediment at a higher rate due to gravitational forces and hence increase effective doses of nanoparticles exposed to cells. By introducing shear flow to SNP suspensions, we reduced sedimentation and nonhomogeneous particle distribution. These results have implications for design of in vitro cytotoxicity assessment of ENMs and suggest that among other factors, sedimentation of nanoparticles in toxicity assessment should be carefully considered.
Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Nanopartículas/toxicidade , Dióxido de Silício/toxicidade , Animais , Técnicas de Cultura de Células , Camundongos , Nanopartículas/química , Células RAW 264.7 , Dióxido de Silício/química , Propriedades de Superfície , Testes de Toxicidade/métodosRESUMO
Little is known about the global gene expression profile of macrophages in response to changes in size and porosity of silica nanoparticles (SNPs). Spherical nonporous SNPs of two different diameters, and mesoporous spherical SNPs with comparable size were characterized. Reactive oxygen species, mitochondrial membrane potential, lysosome degradation capacity, and lysosome pH were measured to evaluate the influence of nonporous and mesoporous SNPs on mitochondrial and lysosomal function. RNA-sequencing was utilized to generate transcriptional profiles of RAW264.7 macrophages exposed to non-toxic SNP doses. DESeq2, limma, and BinReg2 software were used to analyze the data based on both unsupervised and supervised strategies to identify genes with greatest differences among NP treatments. Utilizing GATHER and DAVID software, possible induced pathways were studied. We found that mesoporous silica nanoparticles are capable of altering gene expression in macrophages at doses that do not elicit acute cytotoxicity, while gene transcription was minimally affected by nonporous SNPs.
Assuntos
Biomarcadores/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Macrófagos/metabolismo , Nanopartículas/administração & dosagem , Dióxido de Silício/química , Sobrevivência Celular , Células Cultivadas , Humanos , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Nanopartículas/química , Porosidade , Espécies Reativas de Oxigênio/metabolismoRESUMO
A solvent-free microsphere sintering technique was developed to fabricate scaffolds with pore size gradient for tissue engineering applications. Poly(D,L-Lactide) microspheres were fabricated through an emulsification method where TiO2 nanoparticles were employed both as particulate emulsifier in the preparation procedure and as surface modification agent to improve bioactivity of the scaffolds. A fine-tunable pore size gradient was achieved with a pore volume of 30±2.6%. SEM, EDX, XRD and FTIR analyses all confirmed the formation of bone-like apatite at the 14th day of immersion in Simulated Body Fluid (SBF) implying the ability of our scaffolds to bond to living bone tissue. In vitro examination of the scaffolds showed progressive activity of the osteoblasts on the scaffold with evidence of increase in its mineral content. The bioactive scaffold developed in this study has the potential to be used as a suitable biomaterial for bone tissue engineering and hard tissue regeneration.
Assuntos
Materiais Biocompatíveis/química , Nanopartículas/química , Osteoblastos/citologia , Poliésteres/química , Alicerces Teciduais/química , Titânio/química , Animais , Apatitas/análise , Apatitas/metabolismo , Linhagem Celular , Camundongos , Microesferas , Osteoblastos/metabolismo , Porosidade , Propriedades de Superfície , Engenharia Tecidual/métodosRESUMO
Considering the well-known phenomenon of enhancing bone healing by applying electromagnetic stimulation, manufacturing conductive bone scaffolds is on demand to facilitate the delivery of electromagnetic stimulation to the injured region, which in turn significantly expedites the healing procedure in tissue engineering methods. For this purpose, hybrid conductive scaffolds composed of poly(3,4-ethylenedioxythiophene), poly(4-styrene sulfonate) ( PEDOT: PSS), gelatin (Gel), and bioactive glass (BaG) were produced employing freeze drying technique. Concentration of PEDOT: PSS were optimized to design the most appropriate conductive scaffold in terms of biocompatibility and cell proliferation. More specifically, scaffolds with four different compositions of 0, 0.1, 0.3 and 0.6% (w/w) PEDOT: PSS in the mixture of 10% (w/v) Gel and 30% (w/v) BaG were synthesized. Immersing the scaffolds in simulated body fluid (SBF), we evaluated the bioactivity of samples, and the biomineralization were studied in details using scanning electron microscopy, energy dispersive spectroscopy, X-ray diffraction analysis and Fourier transform infrared spectroscopy. By performing cytocompatibility analyses for 21 days using adult human mesenchymal stem cells, we concluded that the scaffolds with 0.3% (w/w) PEDOT: PSS and conductivity of 170 µS/m has the optimized composition and further increasing the PEDOT: PSS content has inverse effect on cell proliferation. Based on our finding, addition of this optimized amount of PEDOT: PSS to our composition can increase the cell viability more than 4 times compared to a nonconductive composition.
Assuntos
Materiais Biocompatíveis , Compostos Bicíclicos Heterocíclicos com Pontes/química , Calcificação Fisiológica , Polímeros/química , Ácidos Sulfônicos/química , Alicerces Teciduais , Adesão Celular , Proliferação de Células , Células Cultivadas , Humanos , Microscopia Eletrônica de VarreduraRESUMO
Infiltration of immunosuppressive cells into the breast tumor microenvironment (TME) is associated with suppressed effector T cell (Teff) responses, accelerated tumor growth, and poor clinical outcomes. Previous studies from our group and others identified infiltration of immunosuppressive myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs) as critical contributors to immune dysfunction in the orthotopic claudin-low tumor model, limiting the efficacy of adoptive cellular therapy. However, approaches to target these cells in the TME are currently lacking. To overcome this barrier, polymeric micellular nanoparticles (PMNPs) were used for the co-delivery of small molecule drugs activating Toll-like receptors 7 and 8 (TLR7/8) and inhibiting PI3K delta (PI3Kδ). The immunomodulation of the TME by TLR7/8 agonist and PI3K inhibitor led to type 1 macrophage polarization, decreased MDSC accumulation and selectively decreased tissue-resident Tregs in the TME, while enhancing the T and B cell adaptive immune responses. PMNPs significantly enhanced the anti-tumor activity of local radiation therapy (RT) in mice bearing orthotopic claudin-low tumors compared to RT alone. Taken together, these data demonstrate that RT combined with a nanoformulated immunostimulant diminished the immunosuppressive TME resulting in tumor regression. These findings set the stage for clinical studies of this approach.
RESUMO
Infiltration of immunosuppressive cells into the breast tumor microenvironment (TME) is associated with suppressed effector T cell (Teff) responses, accelerated tumor growth, and poor clinical outcomes. Previous studies from our group and others identified infiltration of immunosuppressive myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs) as critical contributors to immune dysfunction in the orthotopic triple-negative breast cancer (TNBC) tumor model limiting the efficacy of adoptive cellular therapy. However, approaches to target these cells specifically in the TME are currently lacking. To overcome this barrier, polymeric micelles nanoparticles (PMNPs) were used for co-delivery of small molecule drugs activating Toll-like receptors 7 and 8 (TLR7/8) and inhibiting PI3K delta. The immunomodulation of the TME by TLR7/8 agonist and PI3K inhibitor altered macrophage polarization, reduced MDSC accumulation and selectively decreased tissue-resident Tregs in the TME, while enhancing the T and B cell adaptive immune response. PMNPs significantly enhanced the anti-tumor activity of local radiation therapy (RT) in mice bearing orthotopic TNBC tumors compared to RT alone. Taken together, these data demonstrate that RT combined with a nanoformulated immunostimulant restructured the TME and has promising potential for future translation combined with RT for patients with TNBC.
RESUMO
Type II innate lymphoid cells (ILC2s) maintain homeostasis and barrier integrity in mucosal tissues. In both mice and humans, ILC2s poorly reconstitute after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Determining the mechanisms involved in their impaired reconstitution could improve transplant outcomes. By integrating single-cell chromatin and transcriptomic analyses of transplanted ILC2s, we identify a previously unreported population of converted ILC1-like cells in the mouse small intestine post-transplant. Exposure of ILC2s to proinflammatory cytokines resulted in a mixed ILC1-ILC2 phenotype but was able to convert only a small population of ILC2s to ILC1s, which were found post-transplant. Whereas ILC2s protected against acute graft-versus-host disease (aGVHD) mediated mortality, infusion of proinflammatory cytokine-exposed ILC2s accelerated aGvHD. Interestingly, murine ILC2 reconstitution post-HSCT is decreased in the presence of alloreactive T cells. Finally, peripheral blood cells from human patients with aGvHD have an altered ILC2-associated chromatin landscape compared to transplanted controls. These data demonstrate that following transplantation ILC2s convert to a pro-pathogenic population with an ILC1-like chromatin state and provide insights into the contribution of ILC plasticity to the impaired reconstitution of ILC2 cells, which is one of several potential mechanisms for the poor reconstitution of these important cells after allo-HSCT.
Assuntos
Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Imunidade Inata , Linfócitos , Camundongos Endogâmicos C57BL , Transplante Homólogo , Transplante de Células-Tronco Hematopoéticas/métodos , Animais , Humanos , Doença Enxerto-Hospedeiro/imunologia , Camundongos , Linfócitos/imunologia , Citocinas/metabolismo , Plasticidade Celular , Feminino , Intestino Delgado/imunologia , Masculino , Camundongos Endogâmicos BALB C , Cromatina/metabolismoRESUMO
The presence of immunosuppressive immune cells in tumors is a significant barrier to the generation of therapeutic immune responses. Similarly, in vivo triple-negative breast cancer (TNBC) models often contain prevalent, immunosuppressive tumor-associated macrophages in the tumor microenvironment (TME), resulting in breast cancer initiation, invasion, and metastasis. Here, we test systemic chemoimmunotherapy using small-molecule agents, paclitaxel (PTX), and colony-stimulating factor 1 receptor (CSF1R) inhibitor, PLX3397, to enhance the adaptive T cell immunity against TNBCs in immunocompetent mouse TNBC models. We use high-capacity poly(2-oxazoline) (POx)-based polymeric micelles to greatly improve the solubility of insoluble PTX and PLX3397 and widen the therapeutic index of such drugs. The results demonstrate that high-dose PTX in POx, even as a single agent, exerts strong effects on TME and induces long-term immune memory. In addition, we demonstrate that the PTX and PLX3397 combination provides consistent therapeutic improvement across several TNBC models, resulting from the repolarization of the immunosuppressive TME and enhanced T cell immune response that suppress both the primary tumor growth and metastasis. Overall, the work emphasizes the benefit of drug reformulation and outlines potential translational path for both PTX and PTX with PLX3397 combination therapy using POx polymeric micelles for the treatment of TNBC.
RESUMO
Apoferritin (APO-Fr) is one of the most investigated proteins proposed as an advanced structure for drug delivery systems. Herein, molecular dynamics simulation was employed to compare the opening of 3-fold and 4-fold pores in APO-Fr during the partial disassembly process at an acidic pH. We showed that more hydrophilic residues in the surface of 3-fold pores compared to 4-fold pores facilitate increased flexibility and a higher tendency to open. In particular, dissociation is induced by the presence of Doxorubicin (DOX) close to 3-fold pores. Our simulations showed loaded DOXs on the APO-Fr surface were mainly involved in the hydrogen bond interactions with the hydrophilic residues, suggesting the difficulty of hydrophobic drugs loading in APO-Fr with the partial disassembly process. However, π-π interactions as well as hydrogen bonds between protein and DOXs were mediated by the basic and acidic amino acids such as HIP128, GLU17, and LYS143 at the open pores, providing penetration of DOXs into the H-Apo-Fr. We conclude that increased drug encapsulations and loading capacity of hydrophobic drugs into the cavity of APO-Fr are feasible by further disassembly of openings to access the internal hydrophobic portions of the protein.
Assuntos
Apoferritinas , Doxorrubicina , Apoferritinas/química , Doxorrubicina/química , Sistemas de Liberação de Medicamentos , Concentração de Íons de Hidrogênio , Simulação de Dinâmica MolecularRESUMO
Liver metastasis is a leading cause of cancer morbidity and mortality. Thus, there has been strong interest in the development of therapeutics that can effectively prevent liver metastasis. One potential strategy is to utilize molecules that have broad effects on the liver microenvironment, such as miR-122, a liver-specific miRNA that is a key regulator of diverse hepatic functions. Here we report the development of a nanoformulation miR-122 as a therapeutic agent for preventing liver metastasis. We engineered a galactose-targeted lipid calcium phosphate (Gal-LCP) nanoformulation of miR-122. This nanotherapeutic elicited no significant toxicity and delivered miR-122 into hepatocytes with specificity and high efficiency. Across multiple colorectal cancer liver metastasis models, treatment with Gal-LCP miR-122 treatment effectively prevented colorectal cancer liver metastasis and prolonged survival. Mechanistic studies revealed that delivery of miR-122 was associated with downregulation of key genes involved in metastatic and cancer inflammation pathways, including several proinflammatory factors, matrix metalloproteinases, and other extracellular matrix degradation enzymes. Moreover, Gal-LCP miR-122 treatment was associated with an increased CD8+/CD4+ T-cell ratio and decreased immunosuppressive cell infiltration, which makes the liver more conducive to antitumor immune response. Collectively, this work presents a strategy to improve cancer prevention and treatment with nanomedicine-based delivery of miRNA. SIGNIFICANCE: Highly specific and efficient delivery of miRNA to hepatocytes using nanomedicine has therapeutic potential for the prevention and treatment of colorectal cancer liver metastasis.
Assuntos
Neoplasias Colorretais/complicações , Neoplasias Colorretais/genética , Neoplasias Hepáticas/secundário , MicroRNAs/metabolismo , Nanopartículas/metabolismo , Animais , Humanos , Camundongos , Metástase Neoplásica , Microambiente TumoralRESUMO
Poor water solubility, off-target toxicity, and small therapeutic window are among major obstacles for the development of drug products. Redox-responsive drug delivery nanoplatforms not only overcome the delivery and pharmacokinetic pitfalls observed in conventional drug delivery, but also leverage the site-specific delivery properties. Cleavable diselenide and disulfide bonds in the presence of elevated reactive oxygen species (ROS) and glutathione concentration are among widely used stimuli-responsive bonds to design nanocarriers. This review covers a wide range of redox-responsive chemical structures and their properties for designing nanoparticles aiming controlled loading, delivery, and release of hydrophobic anticancer drugs at tumor site.
Assuntos
Antineoplásicos , Nanopartículas , Preparações Farmacêuticas , Portadores de Fármacos , Sistemas de Liberação de Medicamentos , OxirreduçãoRESUMO
Regional delivery of chimeric antigen receptor (CAR) T cells in glioblastoma represents a rational therapeutic approach as an alternative to intravenous administration to avoid the blood-brain barrier impediment. Here, we developed a fibrin gel that accommodates CAR-T cell loading and promotes their gradual release. Using a model of subtotal glioblastoma resection, we demonstrated that the fibrin-based gel delivery of CAR-T cells within the surgical cavity enables superior antitumor activity compared to CAR-T cells directly inoculated into the tumor resection cavity.
RESUMO
Chronic toxicity evaluations of nanotechnology-based drugs are essential to support initiation of clinical trials. Ideally such evaluations should address the dosing strategy in human applications and provide sufficient information for long-term usage. Herein, we investigated one-year toxicity of non-surface modified silica nanoparticles (SNPs) with variations in size and porosity (Stöber SNPs 46 ± 4.9 and 432.0 ± 18.7 nm and mesoporous SNPs 466.0 ± 86.0 nm) upon single dose intravenous administration to female and male BALB/c mice (10 animal/sex/group) along with their human blood compatibility. Our evidence of clinical observation and blood parameters showed no significant changes in body weight, cell blood count, nor plasma biomarker indices. No significant changes were noted in post necropsy examination of internal organs and organ-to-body weight ratio. However, microscopic examination revealed significant amount of liver inflammation and aggregates of histocytes with neutrophils within the spleen suggesting an ongoing or resolving injury. The fast accumulation of these plain SNPs in the liver and spleen upon IV administration and the duration needed for their clearance caused these injuries. There were also subtle changes which were attributed to prior infarctions or resolved intravascular thrombosis and included calcifications in pulmonary vessels, focal cardiac fibrosis with calcifications, and focal renal injury. Most of the pathologic lesions were observed when large, non-porous SNPs were administered. Statistically significant chronic toxicity was not observed for the small non-porous particles and for the mesoporous particles. This one-year post-exposure evaluation indicate that female and male BALB/c mice need up to one year to recover from acute tissue toxic effects of silica nanoparticles upon single dose intravenous administration at their 10-day maximum tolerated dose. Further, ex vivo testing with human blood and plasma revealed no hemolysis or complement activation following incubation with these silica nanoparticles. These results can inform the potential utility of silica nanoparticles in biomedical applications such as controlled drug delivery where intravenous injection of the particles is intended.
Assuntos
Nanopartículas , Dióxido de Silício , Animais , Sistemas de Liberação de Medicamentos , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Nanopartículas/toxicidade , Porosidade , Dióxido de Silício/toxicidadeRESUMO
Despite increasing reports of using silica nanoparticles (SNPs) for controlled drug delivery applications, their long-term toxicity profile following intravenous administration remains unexplored. Herein, we investigated the acute (10-day) and subchronic (60-day and 180-day) toxicity of nonporous SNPs of approximately 50â¯nm (Stöber SNPs50) and approximately 500â¯nm in diameter (Stöber SNPs500), and mesoporous SNPs of approximately 500â¯nm in diameter (MSNPs500) upon single-dose intravenous injection into male and female immune-competent inbred BALB/c mice. The Maximum Tolerated Dose (MTD) of the particles was determined 10â¯days post-injection. The MTD of SNPs was administered and toxicity evaluated over 60 and 180â¯days. Results demonstrate that Stöber SNPs50 exhibit systemic toxicity with MTD of 103⯱â¯11â¯mg.kg-1 for female and 100⯱â¯6â¯mg.kg-1 for male mice, respectively. Toxicity was alleviated by increasing the size of the particles (Stöber SNPs500). MTD values of 303⯱â¯4â¯mg.kg-1 for female and 300⯱â¯13â¯mg.kg-1 for male were observed for Stöber SNPs500. Mesoporous SNPs500 showed considerable systemic sex-related toxicity, with MTDs ranging from 40⯱â¯2â¯mg.kg-1 to 95⯱â¯2â¯mg.kg-1 for male and female mice, respectively. Studies of SNPs showed blood toxicity as a function of physiochemical properties such as significant differences in the mean corpuscular hemoglobin (MCHC) and platelet number at day 10 and white blood cell count at day 60. Histological examination also showed size-, porosity- and time-dependent tissue toxicity. Stöber SNPs500 caused major toxic effects such as lung thrombosis, cardiac wall fibrosis and calcifications, brain infarctions with necrotizing inflammatory response, infiltrate, retinal injuries with calcification and focal gliosis, renal parenchymal damage and liver lobular inflammation dependent on the dose and time of exposure. However, tissue toxicity and accumulation of SNPs in liver observed at day 10 was greater than at day 60 and much greater than at day 180. In contrast, a dramatic increase in cytokine levels was observed at day 60. Despite the relatively high doses, SNPs did not cause subchronic toxicity at day 180 after single-dose intravenous injection. However, they showed distinct differences in the 60â¯day in vivo subchronic toxicity and inflammation profile as a function of surface area and size.
Assuntos
Sistemas de Liberação de Medicamentos , Nanopartículas/toxicidade , Dióxido de Silício/química , Animais , Citocinas/metabolismo , Feminino , Injeções Intravenosas , Masculino , Dose Máxima Tolerável , Camundongos , Camundongos Endogâmicos BALB C , Nanopartículas/administração & dosagem , Tamanho da Partícula , Porosidade , Fatores Sexuais , Fatores de Tempo , Distribuição Tecidual , Testes de Toxicidade SubcrônicaRESUMO
Tunable glutathione (GSH)-sensitive hollow mesoporous silica nanoparticles (HMSiO2 NPs) were developed using a structural difference-based selective etching strategy. These organosilica hollow nanoparticles contained disulfide linkages (S-S) in the outer shell which were degraded by GSH. The particles were compared with their nonGSH-sensitive tetraethyl orthosilicate (TEOS) HMSiO2 counterparts in terms of their synthesis method, characterization, doxorubicin (DOX) release profile, and in vitro cytotoxicity in MCF-7 breast cancer cells. Transmission electron microscopy (TEM) of the particles indicated that the fabricated HMSiO2 NPs had an average diameter of 130⯱â¯5â¯nm. Thermogravimetric analysis (TGA) revealed that GSH-sensitive particles had approximately 5.3% more weight loss than TEOS HMSiO2 NPs. Zeta potential of these redox-responsive particles was -23⯱â¯1â¯mV at pHâ¯6 in deionized (DI) water. Nitrogen adsorption-desorption isotherm revealed that the surface area of the hollow mesoporous nanoreservoirs was roughly 446⯱â¯6â¯m2â¯g-1 and the average diameter of the pores was 2.3⯱â¯0.5â¯nm. TEM images suggest that the nanoparticles started to lose mass integrity from Day 1. The particles showed a high loading capacity for DOX (8.9⯱â¯0.5%) as a model drug, due to the large voids existing in the hollow structures. Approximately 58% of the incorporated DOX released within 14â¯days in phosphate buffered saline (PBS) at pHâ¯6 and in the presence of 10â¯mM of GSH, mimicking intracellular tumor microenvironment while release from TEOS HMSiO2 NPs was only c.a. 18%. The uptake of these hollow nanospheres by MCF-7 cells and RAW 264.7 macrophages was evaluated using TEM and confocal microscopy. The nanospheres were shown to accumulate in the endolysosomal compartments after incubation for 24â¯h with the maximum uptake of c.a. 2.1⯱â¯0.3% and 5.2⯱â¯0.4%, respectively. Cytotoxicity of the nanospheres was investigated using CCK-8 assay. Results indicate that intact hollow particles (both GSH-sensitive and TEOS HMSiO2 NPs) were nontoxic to MCF-7 cells after incubation for 24â¯h within the concentration range of 0-1000⯵gâ¯ml-1. DOX-loaded GSH-sensitive nanospheres containing 6⯵gâ¯ml-1 of DOX killed c.a. 51% of MCF-7 cells after 24â¯h while TEOS HMSiO2 NPs killed c.a. 20% with the difference being statistically significant. Finally, cytotoxicity data in RAW 264.7 macrophages and NIH 3â¯T3 fibroblasts shows that intact GSH-sensitive HMSiO2 NPs did not show any toxic effects on these cells with the concentrations equal or <125⯵gâ¯ml-1.
Assuntos
Antibióticos Antineoplásicos/administração & dosagem , Preparações de Ação Retardada/metabolismo , Doxorrubicina/administração & dosagem , Glutationa/metabolismo , Nanopartículas/metabolismo , Dióxido de Silício/metabolismo , Animais , Antibióticos Antineoplásicos/farmacocinética , Antibióticos Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Preparações de Ação Retardada/química , Preparações de Ação Retardada/toxicidade , Doxorrubicina/farmacocinética , Doxorrubicina/farmacologia , Feminino , Humanos , Células MCF-7 , Camundongos , Células NIH 3T3 , Nanopartículas/química , Nanopartículas/toxicidade , Porosidade , Células RAW 264.7 , Dióxido de Silício/química , Dióxido de Silício/toxicidadeRESUMO
Silica nanoparticles are extensively used in biomedical applications and consumer products. Little is known about the interaction of these NPs with the endothelium and effect on platelet adhesion under flow conditions in circulation. In this study, we investigated the effect of silica nanoparticles on the endothelium and its inflammation, and subsequent adhesion of flowing platelets in vitro. Platelet counts adhered onto the surface of endothelial cells in the presence of nanoparticles increased at both low and high concentrations of nanoparticles. Preincubation of endothelial cells with nanoparticles also increased platelet adhesion. Interestingly, platelet adhesion onto TNF-α-treated endothelial cells decreased in the presence of nanoparticles at different concentrations as compared with the absence of nanoparticles. We monitored the expression of different endothelial proteins, known to initiate platelet adhesion, in the presence and absence of silica nanoparticles. We found that silica nanoparticles caused changes in the endothelium such as overexpression of PECAM that promoted platelet adhesion to the endothelial cell.
RESUMO
OBJECTIVE: A systematic characterization of hybrid scaffolds, fabricated based on combinatorial additive manufacturing technique and freeze-drying method, is presented as a new platform for osteoblastic differentiation of dental pulp cells (DPCs). METHODS: The scaffolds were consisted of a collagenous matrix embedded in a 3D-printed beta-tricalcium phosphate (ß-TCP) as the mineral phase. The developed construct design was intended to achieve mechanical robustness owing to 3D-printed ß-TCP scaffold, and biologically active 3D cell culture matrix pertaining to the Collagen extracellular matrix. The ß-TCP precursor formulations were investigated for their flow-ability at various temperatures, which optimized for fabrication of 3D printed scaffolds with interconnected porosity. The hybrid constructs were characterized by 3D laser scanning microscopy, X-ray diffraction, Fourier transform infrared spectroscopy, and compressive strength testing. RESULTS: The in vitro characterization of scaffolds revealed that the hybrid ß-TCP/Collagen constructs offer superior DPCs proliferation and alkaline phosphatase (ALP) activity compared to the 3D-printed ß-TCP scaffold over three weeks. Moreover, it was found that the incorporation of TCP into the Collagen matrix improves the ALP activity. SIGNIFICANCE: The presented results converge to suggest the developed 3D-printed ß-TCP/Collagen hybrid constructs as a new platform for osteoblastic differentiation of DPCs for craniomaxillofacial bone regeneration.