RESUMO
Psoriasis is a chronic T-cell-mediated autoimmune skin disease. Tacrolimus (FK506) is commonly used treatment for psoriasis. However, since the molecular weight of FK506 is more than 500 Da, its skin penetration is limited, so that there is a need to improve the penetrability of FK506 to allow for more effective treatment. To this end, we employed iontophoresis (ItP), which is a physical, intradermal drug delivery technology that relies on the use of weak electric current. Previous findings suggest that activation of cell signaling by the weak electric current applied during ItP may affect the expression of inflammatory cytokines, leading to aggravation of psoriasis. In this study, we analyzed the effect of ItP on the expression of various inflammatory cytokines in the skin, and subsequently examined the therapeutic effect of ItP using negatively-charged liposomes encapsulating FK506 (FK-Lipo) in a rat psoriasis model induced by imiquimod. We found that ItP (0.34 mA/cm2, 1 h) did not affect mRNA levels of inflammatory cytokines or epidermis thickness, indicating that ItP is a safe technology for psoriasis treatment. ItP of FK-Lipo suppressed the expression of inflammatory cytokines induced by imiquimod treatment to a greater extent than skin treated with FK506 ointment for 1 h. Furthermore, epidermis thickening was significantly suppressed only by ItP of FK-Lipo. Taken together, results of this study demonstrate the successful development of an efficient treatment for psoriasis by combining FK-Lipo and ItP, without disease aggravation associated with the weak electric current.
Assuntos
Iontoforese , Psoríase , Animais , Ratos , Tacrolimo/uso terapêutico , Lipossomos , Imiquimode , Psoríase/tratamento farmacológico , CitocinasRESUMO
Antioxidants are promising therapeutics for treating oxidative stress-mediated liver diseases. Previously, we studied a potent natural antioxidant, ferulic acid, and developed a liposomal formulation of ferulic acid (ferulic-lipo) to improve its solubility. Ferulic-lipo significantly attenuated oxidative damage in the liver by inhibiting reactive oxygenase species (ROS). However, antioxidative liposomes must be less reactive with ROS prior to reaching the target sites to effectively neutralize existing ROS. But ferulic-lipo tends to be oxidized before reaching the liver. Besides, γ-oryzanol has been reported to decompose into ferulic acid in vivo; accordingly, we hypothesized that γ-oryzanol could be employed as a natural prodrug of ferulic acid to improve stability and antioxidative effectiveness. Therefore, in this study, we prepared a liposomal formulation of γ-oryzanol (γ-ory-lipo) and investigated its therapeutic effects in a CCl4-induced rat model of liver injury. We found that γ-ory-lipo has a higher chemical stability than does free γ-oryzanol. Although the antioxidative effect of γ-ory-lipo was lower than that of ferulic-lipo, pretreatment of the HepG2 cells with γ-ory-lipo improved the viability of CCl4-treated cells to a similar level as treatment with ferulic-lipo. γ-Oryzanol was shown to be converted into ferulic acid in vitro and in vivo. Furthermore, intravenous administration of γ-ory-lipo exhibited a similar effectiveness as ferulic-lipo against CCl4-induced hepatotoxicity, which should be the due to the conversion of γ-oryzanol into ferulic acid. These findings demonstrated that γ-ory-lipo could be a good natural prodrug of ferulic acid for eradicating its stability problem.
Assuntos
Hepatopatias , Nanopartículas , Fenilpropionatos , Pró-Fármacos , Ratos , Animais , Pró-Fármacos/farmacologia , Pró-Fármacos/uso terapêutico , Espécies Reativas de Oxigênio , Estresse Oxidativo , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Antioxidantes/metabolismo , Fenilpropionatos/farmacologia , Fenilpropionatos/uso terapêuticoRESUMO
Hyaluronic acid (HA) is a hydrophilic supra-macromolecule, with a molecular weight (MW) 1000000<. HA is recognized as a biomaterial for skin moisturization. HA solution is typically injected into the skin using a needle. However, needle injection is invasive and does not result in homogeneous distribution of HA over a large area of skin. Therefore, non-invasive and effective technologies for homogenous intradermal delivery of HA are needed. Recently, we demonstrated the use of iontophoresis (ItP) for non-invasive intradermal delivery of various macromolecules, such as small interfering RNA (siRNA) (MW: 12000) and antibodies (MW: 150000). Based on our previous studies, we hypothesized that HA can also be delivered non-invasively into the skin by ItP. In this study, we applied ItP to fluorescence-labeled HA (MW: 600000-1120000 and 1200000-1600000) on rat dorsal skin. Following treatment, fluorescence was observed to be widely distributed in the skin, demonstrating successful intradermal delivery of HA via ItP. In addition, the relative moisture content and elasticity of skin treated with ItP/HA was temporarily higher than that of control skin. This is the first report demonstrating successful non-invasive intradermal delivery of HA and improvement of skin conditions by high-molecular weight HA delivered by ItP. In conclusion, ItP would be a useful technology for non-invasive intradermal delivery of high-molecular weight HA for treatment of skin diseases and cosmetology applications.
Assuntos
Ácido Hialurônico , Dermatopatias , Animais , Ratos , Iontoforese , Pele , Administração Cutânea , Absorção Cutânea , Dermatopatias/metabolismoRESUMO
mRNA vaccines have attracted considerable attention as a result of the 2019 coronavirus pandemic; however, challenges remain regarding use of mRNA vaccines, including insufficient delivery owing to the high molecular weights and high negative charges associated with mRNA. These characteristics of mRNA vaccines impair intracellular uptake and subsequent protein translation. In the current study, we prepared a minimal mRNA vaccine encoding a tumor associated antigen human gp10025-33 peptide (KVPRNQDWL), as a potential treatment for melanoma. Minimal mRNA vaccines have recently shown promise at improving the translational process, and can be prepared via a simple production method. Moreover, we previously reported the successful use of iontophoresis (IP) technology in the delivery of hydrophilic macromolecules into skin layers, as well as intracellular delivery of small interfering RNA (siRNA). We hypothesized that combining IP technology with a newly synthesized minimal mRNA vaccine can improve both transdermal and intracellular delivery of mRNA. Following IP-induced delivery of a mRNA vaccine, an immune response is elicited resulting in activation of skin resident immune cells. As expected, combining both technologies led to potent stimulation of the immune system, which was observed via potent tumor inhibition in mice bearing melanoma. Additionally, there was an elevation in mRNA expression levels of various cytokines, mainly interferon (IFN)-γ, as well as infiltration of cytotoxic CD8+ T cells in the tumor tissue, which are responsible for tumor clearance. This is the first report demonstrating the application of IP for delivery of a minimal mRNA vaccine as a potential melanoma therapeutic.
Assuntos
Vacinas Anticâncer , Melanoma , Vacinas de mRNA , Animais , Humanos , Camundongos , Vacinas Anticâncer/genética , Linfócitos T CD8-Positivos , Iontoforese , Melanoma/terapia , Melanoma/genética , Vacinas de mRNA/genéticaRESUMO
Since small extracellular vesicle (sEVs) are involved in cell-to-cell communication via transfer of certain bioactive molecules and have the capability to overcome biological barriers against drug transport, their use as a drug delivery system (DDS) has been demonstrated in treatment of a diverse range of diseases. However, some issues in drug encapsulation have been pointed out, including low encapsulation efficiency and poor reproducibility. It was previously reported that liposomes containing phosphatidylserine (PS) can fuse together in the presence of calcium ion, which allows for drug encapsulation into the resultant liposomes (i.e., calcium fusion method). On the other hand, PS is reportedly present in lipid membrane of sEVs as a distinct lipid composition. We therefore hypothesized that PS-mediated membrane fusion of sEVs with PS-liposomes encapsulating therapeutic agents via the calcium fusion method can be applied to convenient drug encapsulation into sEVs. Membrane fusion of PS-liposomes and sEVs derived from murine melanoma B16F1 cells (B16-sEVs) was firstly confirmed. The obtained nanoparticles, termed chimeric nanoparticles (CM-NP), showed comparable cellular uptake to B16-sEVs into B16F1 cells. Moreover, CM-NP encapsulating an anticancer drug doxorubicin (DOX) (CM-NP-DOX) could be prepared by membrane fusion of PS-liposomes encapsulating DOX (PS-Lipo-DOX) and B16-sEVs. CM-NP-DOX exhibited a superior anticancer effect on B16F1 cells in vitro compared with PS-Lipo-DOX. These findings suggest that the calcium fusion method could be applied for membrane fusion of sEVs and PS-liposomes, and that this approach would likely be useful for efficient drug encapsulation into sEVs, as well as increasing liposome functionality.
Assuntos
Vesículas Extracelulares , Nanopartículas , Animais , Camundongos , Lipossomos , Cálcio , Fusão de Membrana , Reprodutibilidade dos Testes , Doxorrubicina/farmacologia , LipídeosRESUMO
Although the strategy in cancer vaccination is to provide a therapeutic effect against an established tumor, there is an urgent need to develop prophylactic vaccines for non-viral cancers. In this study, we prepared polyplex nanoparticles through electrostatic interactions between a positively-charged modified tumor associated antigen, namely human derived melanoma gp10025-33 peptide (KVPRNQDWL-RRRR), and a negatively charged cytosine-phosphate-guanosine motif (CpG-ODN) adjuvant. We previously demonstrated successful transdermal delivery of various hydrophilic macromolecules by iontophoresis (IP) using weak electricity. Herein, we investigated the effectiveness of IP in the transdermal delivery of a prophylactic polyplex vaccine. IP was successful in establishing a homogenous distribution of the vaccine throughout skin. Efficacy of the vaccine was demonstrated against melanoma growth. A significant tumor regression effect was observed, which was confirmed by elevated mRNA expression levels of various cytokines, mainly interferon (IFN)-γ, as well as infiltration of cytotoxic CD8+ T cells. Additionally, we evaluated the therapeutic effect of the vaccine and we found a significant reduction in tumor burden. Stimulation of systemic immunity was confirmed by upregulation of IFN-γ. This is the first report to demonstrate the use of IP in the transdermal delivery of a prophylactic melanoma vaccine.
Assuntos
Vacinas Anticâncer , Melanoma , Humanos , Iontoforese , Linfócitos T CD8-Positivos , Interferon gamaRESUMO
Heat is a detrimental environmental stressor that disrupts spermatogenesis and results in male infertility. Previous investigations have shown that heat stress reduces the motility, number, and fertilization ability of living spermatozoa. Sperm hyperactivation, capacitation, acrosomal reaction, and chemotaxis towards the ova are regulated by the cation channel of sperm (CatSper). This sperm-specific ion channel triggers the influx of calcium ions into sperm cells. The aim of this study in rats was to investigate whether heat treatment affected the expression levels of CatSper-1 and -2, together with the sperm parameters, testicular histology and weight. The rats were exposed to heat stress for 6 days and the cauda epididymis and testis were collected 1, 14, and 35 days after heat treatment to measure sperm parameters, gene and protein expression, testicular weight, and histology. Interestingly, we found that heat treatment caused a notable downregulation of CatSper-1 and -2 expression at all three time points. In addition, there were significant reductions in sperm motility and number and an increase in the percentage of abnormal sperm at 1 and 14 days, with cessation of sperm production at 35 days. Furthermore, expression of the steroidogenesis regulator, 3 beta-hydroxysteroid dehydrogenase (3ß-HSD) was upregulated in the 1-, 14- and 35-day samples. Heat treatment also upregulated the expression of the apoptosis regulator, BCL2-associated X protein (BAX), decreased testicular weight, and altered testicular histology. Therefore, our data showed for the first time that heat stress downregulated CatSper-1 and -2 in the rat testis, and that this may be a mechanism involved in heat stress-induced impairment of spermatogenesis.
Assuntos
Canais de Cálcio , Sêmen , Masculino , Ratos , Animais , Canais de Cálcio/genética , Canais de Cálcio/metabolismo , Sêmen/metabolismo , Motilidade dos Espermatozoides , Espermatozoides/fisiologia , Espermatogênese , Testículo/metabolismo , CálcioRESUMO
Antioxidants are useful for the treatment of oxidative stress mediated liver damage. A naturally occurring antioxidant γ-oryzanol is rapidly hydrolyzed to its active hydrophobic metabolite, ferulic acid, inside the body. Limitations associated with the hydrophobicity of ferulic acid can be overcome by encapsulating in a liposomal formulation. As intravenously administered nanoparticles (including liposomes) can effectively reach the liver, such systems may be suitable drug delivery carriers to treat liver injury. In this study, we prepared a liposomal formulation of ferulic acid (ferulic-lipo) and examined its effects on liver damage induced by CCl4. Ferulic-lipo were ~100â nm in size and drug encapsulation efficiency was about 92%. Ferulic-lipo showed potent scavenging efficacy against hydroxyl radical compared to α-tocopherol liposomes. Ferulic-lipo significantly prevented CCl4-mediated cytotoxicity in human hepatocarcinoma cells. Furthermore, intravenous administration of ferulic-lipo significantly reduced alanine aminotransferase and aspartate amino transferase levels in a rat model of liver injury. CCl4-mediated reactive oxygen species generation in liver was also reduced by intravenous administration of ferulic-lipo. Hepatoprotective effects of ferulic-lipo were demonstrated by histological observation of CCl4-induced liver tissue damage. Therefore, ferulic-lipo exhibit potent antioxidative capacity and were suggested to be an effective formulation for prevention of oxidative damage of liver tissue.
RESUMO
Intraperitoneal administration of anticancer nanoparticles is a rational strategy for preventing peritoneal dissemination of colon cancer due to the prolonged retention of nanoparticles in the abdominal cavity. However, instability of nanoparticles in body fluids causes inefficient retention, reducing its anticancer effects. We have previously developed anticancer nanoparticles containing tocopheryl succinate, which showed high in vivo stability and multifunctional anticancer effects. In the present study, we have demonstrated that peritoneal dissemination derived from colon cancer was prevented by intraperitoneal administration of tocopheryl succinate nanoparticles. The biodistribution of tocopheryl succinate nanoparticles was evaluated using inductively coupled plasma mass spectroscopy and imaging analysis in mice administered quantum dot encapsulated tocopheryl succinate nanoparticles. Intraperitoneal administration of tocopheryl succinate nanoparticles showed longer retention in the abdominal cavity than by its intravenous (i.v.) administration. Moreover, due to effective biodistribution, tumor growth was prevented by intraperitoneal administration of tocopheryl succinate nanoparticles. Furthermore, the anticancer effect was attributed to the inhibition of cancer cell proliferation and improvement of the intraperitoneal microenvironment, such as decrease in the levels of vascular endothelial growth factor A, interleukin 10, and M2-like phenotype of tumor-associated macrophages. Collectively, intraperitoneal administration of tocopheryl succinate nanoparticles is expected to have multifaceted antitumor effects against colon cancer with peritoneal dissemination.
Assuntos
Neoplasias do Colo , Nanopartículas , Animais , Neoplasias do Colo/tratamento farmacológico , Humanos , Camundongos , Nanopartículas/química , Succinatos/farmacologia , Distribuição Tecidual , Microambiente Tumoral , Fator A de Crescimento do Endotélio Vascular , alfa-Tocoferol/química , alfa-Tocoferol/farmacologiaRESUMO
Delivery of cerebroprotective agents using liposomes has been demonstrated to be useful for treating cerebral ischemia/reperfusion (I/R) injury. We previously reported that intravenous administration of liposomes with diameters of 100 nm showed higher accumulation in the I/R region compared with larger liposomes (>200 nm) by passage through the disintegrated blood-brain barrier, suggesting a size-dependence for liposome-mediated drug delivery. Based on these findings, we hypothesized that regulation of liposomal particle size (<100 nm) may enhance the therapeutic efficacy of encapsulated drugs on cerebral I/R injury. Herein, we prepared lipid nanoparticles (LNP) with particle sizes <100 nm by the microfluidics method and compared their therapeutic potential with LNP exhibiting sizes >100 nm in cerebral I/R model rats. Intravenously administered smaller LNP (ca. 60 nm) exhibited wider accumulation and diffusivity in the brain parenchyma of the I/R region compared with larger LNP (>100 nm). Importantly, treatment with LNP encapsulating the cerebroprotective agent FK506 (FK-LNP) with particle sizes <100 nm showed greater cerebroprotective effects than FK-LNP with sizes >100 nm, and also significantly ameliorated brain injury. These results suggest that particle size regulation of LNP to sizes <100 nm can enhance the therapeutic effect of encapsulated drugs for treatment of cerebral I/R injury, and that FK-LNP could be a promising cerebroprotective agent.
Assuntos
Isquemia Encefálica , Nanopartículas , Fármacos Neuroprotetores , Traumatismo por Reperfusão , Animais , Isquemia Encefálica/tratamento farmacológico , Lipossomos/uso terapêutico , Fármacos Neuroprotetores/farmacologia , Tamanho da Partícula , Ratos , Ratos Wistar , Traumatismo por Reperfusão/tratamento farmacológico , Traumatismo por Reperfusão/prevenção & controle , Tacrolimo/farmacologia , Tacrolimo/uso terapêuticoRESUMO
Delivery of medicines using nanoparticles via the enhanced permeability and retention (EPR) effect is a common strategy for anticancer chemotherapy. However, the extensive heterogeneity of tumors affects the applicability of the EPR effect, which needs to overcome for effective anticancer therapy. Previously, we succeeded in the noninvasive transdermal delivery of nanoparticles by weak electric current (WEC) and confirmed that WEC regulates the intercellular junctions in the skin by activating cell signaling pathways (J. Biol. Chem., 289, 2014, Hama et al.). In this study, we applied WEC to tumors and investigated the EPR effect with polyethylene glycol (PEG)-modified doxorubicin (DOX) encapsulated nanoparticles (DOX-NP) administered via intravenous injection into melanoma-bearing mice. The application of WEC resulted in a 2.3-fold higher intratumor accumulation of nanoparticles. WEC decreased the amount of connexin 43 in tumors while increasing its phosphorylation; therefore, the enhancing of intratumor delivery of DOX-NP is likely due to the opening of gap junctions. Furthermore, WEC combined with DOX-NP induced a significant suppression of tumor growth, which was stronger than with DOX-NP alone. In addition, WEC alone showed tumor growth inhibition, although it was not significant compared with non-treated group. These results are the first to demonstrate that effective anticancer therapy by combination of nanoparticles encapsulating chemotherapeutic agents and WEC.
Assuntos
Doxorrubicina/administração & dosagem , Doxorrubicina/uso terapêutico , Técnicas Eletroquímicas , Melanoma/tratamento farmacológico , Nanopartículas/química , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/uso terapêutico , Sistemas de Liberação de Medicamentos/métodos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Nanopartículas/administração & dosagem , Neoplasias Experimentais , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Targeted drug delivery using nanoparticles has been applied for the treatment of diverse diseases, including cancer and inflammatory diseases. Nanoparticle-mediated delivery of therapeutic agents via the enhanced permeability and retention effect generally augments their therapeutic efficiency; however, limitations with passive entry of nanoparticles into diseased sites, due to the presence of biological barriers represented by the endothelial layer, remain to be addressed. To this end, development of nanoparticles with intrinsic characteristics similar to circulatory cells (e.g., leukocytes, platelets) for use as biomimetic drug delivery systems (DDS) has been focused as a means to overcome the issues of conventional DDS. In particular, synthetic biomimetic nanoparticles coated with cellular membranes were recently prepared and shown to actively overcome the inflamed vessels and tumor microenvironment as a result of the functionality of membrane proteins, which allowed secure drug delivery into diseased sites. We recently developed liposomes modified with leukocyte membrane proteins via intermembrane protein transfer, a simple method to reconstitute cellular membrane proteins onto lipid bilayers. The resultant liposomes demonstrated the ability to cross the inflamed endothelial layer and permeate into tumor tissue by mimicking the properties of leukocytes. Thus, biomimetic DDS offer promise as new therapeutic approaches for various diseases by overcoming biological barriers that typically inhibit drug delivery. Herein, we review recent approaches to develop biomimetic DDS using the cell membrane coating method, and highlight our recent findings on leukocyte-mimetic liposomes prepared via intermembrane protein transfer.
Assuntos
Biomimética , Lipossomos , Sistemas de Liberação de Medicamentos , Proteínas de Membrana , Sistemas de Liberação de Fármacos por NanopartículasRESUMO
Anti-rheumatoid arthritis (RA) effects of α-tocopherol (α-T) have been shown in human patients in a double-blind trial. However, the effects of α-T and its derivatives on fibroblast-like synoviocytes (FLS) during the pathogenesis of RA remain unclear. In the present study, we compared the expression levels of genes related to RA progression in FLS treated with α-T, succinic ester of α-T (TS), and phosphate ester of α-T (TP), as determined via RT-PCR. The mRNA levels of interleukin (IL)-6, tumor necrosis factor-α (TNF-α), matrix metalloproteinase (MMP)-3, and MMP-13 were reduced by treatment with TP without cytotoxicity, while α-T and TS did not show such effects. Furthermore, intraperitoneal injection of TP ameliorated the edema of the foot and joint and improved the arthritis score in laminarin-induced RA model mice. Therefore, TP exerted anti-RA effects through by inhibiting RA-related gene expression.
Assuntos
Antirreumáticos/farmacologia , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , alfa-Tocoferol/análogos & derivados , Animais , Artrite Reumatoide/induzido quimicamente , Citocinas/biossíntese , Glucanos/toxicidade , Humanos , Metaloproteinase 13 da Matriz/biossíntese , Metaloproteinase 3 da Matriz/biossíntese , Camundongos , alfa-Tocoferol/farmacologiaRESUMO
Obesity is a pathological state related to various lifestyle-related diseases, such as diabetes and dyslipidemia, that may be prevented through the development of anti-obesity treatments. Lipid accumulation in cells could be affected by vitamin E ester α-tocopheryl succinate (TS), which has various biological activities, such as anti-cancer effect, via activation of cell signaling pathways, although the antioxidative activity of TS is lost due to esterification of the phenolic OH group. In this study, we found for the first time that TS significantly suppressed lipid accumulation in mouse 3T3-L1 adipocytes. TS treatment reduced the amount of triglycerides in the culture medium, and inhibited activity of glycerol-3-phosphate dehydrogenase, a marker of lipid synthesis. Furthermore, TS accelerated lipolysis. Treatment of adipocytes with TS for 24 h induced no significant cytotoxicity. In TS-treated cells, phosphorylation of Akt, which is involved in fatty acid synthesis via sterol regulatory element-binding proteins (SREBP), was prevented, while levels of phosphorylated protein kinase A (PKA) did not change. Taken together, these results suggest that vitamin E ester TS can suppress lipid accumulation in adipocytes by regulating lipid metabolic cell signaling.
Assuntos
Adipócitos/efeitos dos fármacos , Antioxidantes/farmacologia , Lipogênese/efeitos dos fármacos , Lipólise/efeitos dos fármacos , alfa-Tocoferol/farmacologia , Células 3T3 , Células 3T3-L1 , Adipócitos/metabolismo , Animais , Relação Dose-Resposta a Droga , Lipogênese/fisiologia , Lipólise/fisiologia , CamundongosRESUMO
α-Tocopheryl succinate (TS) is a tocopherol derivative and has multifaceted anti-cancer effects; TS not only causes cancer cell-specific apoptosis but also inhibits tumor angiogenesis. Although TS has the potential to be used as a well-tolerated anti-angiogenic drug, it is still unclear which step of the angiogenic process is inhibited by TS. Here, we show that TS inhibits the expression of angiopoietin (Ang)-2, which induces destabilization of vascular structure in the initial steps of the angiogenic process. In mouse melanoma cells, TS treatment decreased mRNA and extracellular protein levels of Ang-2; however, the mRNA level of Ang-1, which stabilizes the vascular structure, remained unchanged. Furthermore, aorta ring and Matrigel plug angiogenesis assays indicated that the conditioned medium from TS-treated cells (CM-TS) inhibited neovascularization and blood leakage from the existing blood vessels, respectively. Following immunohistochemical staining of the vessels treated with CM-TS, imaging studies showed that the vascular endothelial cells were highly packed with pericytes. In conclusion, we found that TS inhibits Ang-2 expression and, consequently, stabilizes the vascular structure during the initial step of tumor angiogenesis.
Assuntos
Inibidores da Angiogênese/farmacologia , Angiopoietina-2/antagonistas & inibidores , Neovascularização Patológica/tratamento farmacológico , alfa-Tocoferol/farmacologia , Inibidores da Angiogênese/química , Angiopoietina-1/metabolismo , Angiopoietina-2/metabolismo , Animais , Aorta/efeitos dos fármacos , Linhagem Celular Tumoral , Meios de Cultivo Condicionados/farmacologia , Humanos , Masculino , Melanoma/irrigação sanguínea , Melanoma/tratamento farmacológico , Melanoma/patologia , Camundongos , Camundongos Endogâmicos BALB C , Neovascularização Patológica/patologia , Relação Estrutura-Atividade , alfa-Tocoferol/químicaRESUMO
Technologies that overcome the barrier presented by vascular endothelial cells are needed to facilitate targeted delivery of drugs into tissue parenchyma by intravenous administration. We previously reported that weak electric current treatment (ET: 0.3-0.5 mA/cm2) applied onto skin tissue in a transdermal drug delivery technique termed iontophoresis induces cleavage of intercellular junctions that results in permeation of macromolecules such as small interfering RNA and cytosine-phosphate-guanine (CpG) oligonucleotide through the intercellular space. Based on these findings, we hypothesized that application of ET to blood vessels could promote cleavage of intercellular junctions that artificially induces increase in vascular permeability to enhance extravasation of drugs from the vessels into target tissue parenchyma. Here we investigated the effect of ET (0.34 mA/cm2) on vascular permeability using embryonated chicken eggs, which have blood vessels in the chorioallantoic membrane (CAM), as an animal model. ET onto the CAM of the eggs significantly increased extravasation of intravenously injected calcein (M.W. 622.6), a low molecular weight compound model, and the macromolecule fluorescein isothiocyanate (FITC)-dextran (M.W. 10000). ET-mediated promotion of penetration of FITC-dextran through vascular endothelial cells was also observed in transwell permeability assay using monolayer of human umbilical vein endothelial cells without induction of obvious cellular damage. Confocal microscopy detected remarkable fluorescence derived from injected FITC-dextran in blood vessel walls. These results in embryonated chicken eggs suggest that ET onto blood vessels could artificially enhance vascular permeability to facilitate extravasation of macromolecules from blood vessels.
Assuntos
Permeabilidade Capilar , Dextranos/administração & dosagem , Endotélio Vascular/metabolismo , Fluoresceína-5-Isotiocianato/análogos & derivados , Animais , Galinhas , Membrana Corioalantoide/irrigação sanguínea , Dextranos/farmacocinética , Estimulação Elétrica , Endotélio Vascular/citologia , Fluoresceína-5-Isotiocianato/administração & dosagem , Fluoresceína-5-Isotiocianato/farmacocinética , Células Endoteliais da Veia Umbilical Humana , Humanos , Injeções Intravenosas , Microscopia ConfocalRESUMO
Our previous study reported that co-encapsulation of potent antioxidants astaxanthin (Asx) and capsaisin (Cap) into liposomes brought about synergistically higher antioxidative activity than the calculated additive activity of each single antioxidant encapsulating liposome. Based on the previous computational chemistry analysis, the synergistic effect was revealed to be resulted from intermolecular interactions between Asx, especially 3R,3'R-form of Asx stereoisomer (Asx-R), and Cap, by which changes of electronic states of the polyene moiety of Asx-R were induced. Although liposomes co-encapsulating Asx-R and Cap (Asx-R/Cap-Lipo) at an optimal ratio clearly showed synergistic antioxidative activity in vitro, it is unclear whether the effective antioxidative activity derived from intermolecular interaction between Asx-R and Cap is also exerted in vivo. Therefore, in this study, we investigated therapeutic potential of Asx-R/Cap-Lipo as an antioxidant formulation in vivo. For this purpose, we employed carbon tetrachloride (CCl4)-induced acute liver injury rat model, since CCl4 is known to cause oxidative damage in liver. CCl4 administration significantly increased the levels of aspartate transaminase (AST) and alanine aminotransferase (ALT). Intravenous combined administration of liposomes encapsulating Asx-R (Asx-R-Lipo) and liposomes encapsulating Cap (Cap-Lipo) significantly decreased CCl4-induced increase of AST and ALT levels. Importantly, the treatment with Asx-R/Cap-Lipo tended to show higher protective effect on acute liver injury than combined treatment with Asx-R-Lipo plus Cap-Lipo. These results suggest that co-encapsulated Asx-R and Cap in liposomal membranes could exert more effective antioxidative activities in vivo, and that Asx-R/Cap-Lipo would be a hopeful antioxidant formulation for treating reactive oxygen species-related diseases.
Assuntos
Antioxidantes/farmacologia , Capsaicina/administração & dosagem , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Doença Aguda , Animais , Capsaicina/química , Capsaicina/farmacologia , Tetracloreto de Carbono , Lipossomos , Masculino , Ratos , Ratos Wistar , Xantofilas/administração & dosagem , Xantofilas/química , Xantofilas/farmacologiaRESUMO
Oxidative stress induced by decreases in tear volume and excessive tear evaporation is a key factor in dry eye disease (DED). Previously, we reported that desiccation stress induces reactive oxygen species generation and up-regulated expression of age-related markers such as p53, p21 and p16. We also showed that the antioxidant astaxanthin prepared as a liposomal formulation could suppress these phenomena in the in vitro DED model. In this study, we evaluated the protective effect of liposomes encapsulating astaxanthin against superficial punctate keratopathy (SPK) in the in vivo rat DED model. This model of DED was characterized by decreased tear volume and increased fluorescein score as an indicator of SPK as well as upregulated expression of age-related markers. Repeat-dose of liposomal astaxanthin prevented increases in the fluorescein score and up-regulation of age-related markers. Liposomes bearing a slight positive surface charge had superior effects and higher affinity compared to neutral liposomes. Furthermore, fluorescence intensities in rat corneal epithelium after administration of high-affinity liposomes labeled with fluorescent dye were higher than those for neutral liposomes. In conclusion, we developed the high-affinity liposomal formulation that can prevent DED and promote antioxidative effects of astaxanthin.
RESUMO
Decreases in tear volume, unstable tear films and excessive tear evaporation are known to cause desiccation and hyperosmolar stress. These, in turn, induce oxidative stress that is thought to cause dry eye, which is also considered to be age-related disease. We hypothesized that oxidative stress induces up-regulation of age-related markers, and that the antioxidant astaxanthin prepared as a liposomal formulation may be a candidate for the treatment of dry eye. Herein, we examined age-related markers in an in vitro dry eye model, and evaluated the efficacy of high-affinity liposomes containing astaxanthin. The in vitro dry eye model showed desiccation time-dependent increases in reactive oxygen species. We confirmed the up-regulation of p53, p21 and p16 as a function of desiccation time. Pretreatment with both neutral and slightly-positively-charged astaxanthin liposomal formulations showed significant suppression of up-regulation of all markers, with the positively-charged liposomes exhibiting the greatest efficacy. Furthermore, positively-charged liposomes labeled with fluorescent dyes demonstrated much higher affinity to normal human corneal epithelial cells (HCECs) than neutral liposomes. Taken together, we confirmed the up-regulation of age-related markers, especially p16, in an in vitro dry eye model, and demonstrated the potential of high-affinity liposomal astaxanthin for the treatment of dry eye.
RESUMO
Lysophosphatidic acid (LPA) is a bioactive phospholipid that induces diverse biological responses. Recently, we found that LPA ameliorates NSAIDs-induced gastric ulcer in mice. Here, we quantified LPA in 21 medicinal herbs used for treatment of gastrointestinal (GI) disorders. We found that half of them contained LPA at relatively high levels (40-240⯵g/g) compared to soybean seed powder (4.6⯵g/g), which we previously identified as an LPA-rich food. The LPA in peony (Paeonia lactiflora) root powder is highly concentrated in the lipid fraction that ameliorates indomethacin-induced gastric ulcer in mice. Synthetic 18:1 LPA, peony root LPA and peony root lipid enhanced prostaglandin E2 production in a gastric cancer cell line, MKN74 cells that express LPA2 abundantly. These materials also prevented indomethacin-induced cell death and stimulated the proliferation of MKN74 cells. We found that LPA was present in stomach fluids at 2.4⯵M, which is an effective LPA concentration for inducing a cellular response in vitro. These results indicated that LPA is one of the active components of medicinal herbs for the treatment of GI disorder and that orally administered LPA-rich herbs may augment the protective actions of endogenous LPA on gastric mucosa.