Young Tag Ko

Gene delivery into ischemic myocardium by double-targeted lipoplexes with anti-myosin antibody and TAT peptide

The treatment of myocardial ischemia using gene therapy is a rather novel but promising approach. Gene delivery to target cells may be enhanced by using double-targeted delivery systems simultaneously capable of extracellular accumulation and intracellular penetration. With this in mind, we have used low cationic liposomes–plasmid DNA complexes (lipoplexes) modified with cell-penetrating transactivating transcriptional activator (TAT) peptide (TATp) and/or with monoclonal anti-myosin monoclonal antibody 2G4 (mAb 2G4) specific toward cardiac myosin, for targeted gene delivery to ischemic myocardium. In vitro transfection of both normoxic and hypoxic cardiomyocytes was enhanced by the presence of TATp as determined by fluorescence microscopy and ELISA. The in vitro transfection was further enhanced by the additional modification with mAb 2G4 antibody in the case of hypoxic, but not normoxic cardiomyocytes. However, we did not observe a synergism between TATp and mAb 2G4 ligands under our experimental condition. In in vivo experiments, we have clearly demonstrated an increased accumulation of mAb 2G4-modified TATp lipoplexes in the ischemic rat myocardium and significantly enhanced transfection of cardiomyocytes in the ischemic zone. Thus, the genetic transformation of normoxic and hypoxic cardiomyocytes can be enhanced by using lipoplexes modified with TATp and/or mAb 2G4. Such complexes also demonstrate an increased accumulation in the ischemic myocardium and effective transfection of hypoxic cardiomyocytes in vivo.

Cationic Liposomes Loaded with Proapoptotic Peptide d-(KLAKLAK)2 and Bcl-2 Antisense Oligodeoxynucleotide G3139 for Enhanced Anticancer Therapy

The treatment of cancer using macromolecular therapeutics such as oligonucleotides or peptides requires efficient delivery systems capable of intracellular penetration and may also benefit from use of a combination of therapeutics with different mechanisms of action. With this possibility in mind, we constructed cationic liposome loaded with the proapoptotic peptide, d-(KLAKLAK)(2) and the Bcl-2 antisense oligodeoxynucleotide, G3139, and determined whether the combination of the proapoptotic macromolecules in a single cationic liposome can enhance antitumor efficacy. Advantage was taken of alternating charge interaction to entrap macromolecules of opposite charge. The polycationic peptide d-(KLAKLAK)(2) was first condensed with the polyanionic oligodeoxynucleotide G3139 to obtain overall negatively charged peptide/oligodeoxynucleotide complexes. The complexes were then entrapped into DOTAP/DOPE cationic liposomes (CL). This sequential charge interaction ensured efficient entrapment of d-(KLAKLAK)(2) and G3139 with a high loading efficiency (50%) and capacity (7.5 wt %). In vitro treatment of mouse melanoma B16(F10) with CL loaded with d-(KLAKLAK)(2)/G3139 led to significantly enhanced antitumor efficacy, mediated by stimulated induction of apoptotic (caspase 3/7) activity, when compared to CL loaded with G3139 alone. Intratumoral injection of CL loaded with d-(KLAKLAK)(2)/G3139 in B16(F10) mice xenograft also led to suppressed tumor growth associated with enhanced apoptotic activity. Thus, the combination of proapoptotic peptide d-(KLAKLAK)(2) and antisense oligonucleotide G3139 in a cationic liposome led to enhanced apoptotic/antitumor efficacy and may provide a promising tool for cancer treatment.

Liposomal co-delivery of curcumin and albumin/paclitaxel nanoparticle for enhanced synergistic antitumor efficacy.

Paclitaxel (PTX) and curcumin (CUR) are potent chemotherapeutic agents used in the treatment of cancer. In the present study, hybrid polymer-lipid nanoparticles co-loaded with PTX and CUR were developed to investigate the therapeutic potential of a combination drug regimen. For this purpose, PTX-loaded albumin nanoparticles (APN) were prepared and encapsulated in PEGylated hybrid liposomes containing CUR (CL-APN) via a thin-film hydration technique. CL-APN was nanosized with a uniform spherical morphology. PTX and CUR release was sustained and occurred in a sequential manner, wherein CUR was expected to downregulate the nuclear factor NF-κB and Akt pathways and increase the therapeutic efficacy of PTX. The ratiometric combination of PTX and CUR was significantly more cytotoxic than the individual drugs. Importantly, dual-drug-loaded nanocarriers exhibited a superior cytotoxic effect than a cocktail combination at a lower dose. CL-APN induced significantly higher early and late apoptosis, induced a stronger G2/M arrest, and significantly increased the subG1 cell population. By combining CUR, an effective NF-κB inhibitor, with PTX, a powerful anticancer drug, in a polymer-lipid hybrid nanoparticle system, we could improve the therapeutic efficacy in cancer treatments. Our results showed that such co-loaded delivery systems could serve as a promising therapeutic approach to improve clinical outcomes against various malignancies.

Enhanced Oral Delivery of Curcumin from N-trimethyl Chitosan Surface-Modified Solid Lipid Nanoparticles: Pharmacokinetic and Brain Distribution Evaluations

ABSTRACTPurposeSolid lipid nanoparticles (SLNs) have been proposed as a colloidal carrier system that could enhance the oral bioavailability of curcumin. However, a burst release of the loaded drug, which occurs in acidic environments, has been a main obstacle to the oral delivery of curcumin by using SLNs as a carrier system. We hypothesized that a quarternized chitosan derivative could be used for acid-resistant coating to stabilize the SLNs and circumvent the burst release.MethodsN-trimethyl chitosan (TMC) was synthesized and determined by 1H-NMR and FT-IR. To investigate the details of chitosan and TMC surface modification on SLCNs composed of palmitic acid, cholesterol, TPGS and curcumin, a number of factors such as optimized SLNs composition, solid state characterization, stability, cell viability, in vitro release in GI conditions, curcumin oral bioavailability and brain distribution studies, were evaluated.ResultsThe TMC-SLCNs exhibited prolonged stability in room and refrigerated conditions, controlled drug release in simulated intestinal fluid, significantly higher oral bioavailability, and brain distribution of curcumin than free curcumin, chitosan and non-coated SLCNs.ConclusionsThese finding suggests that the TMC-SLCNs is a promising nanocarrier system for oral delivery and brain distribution of curcumin.

Matrix Metalloproteinase-8 Plays a Pivotal Role in Neuroinflammation by Modulating TNF-α Activation

Matrix metalloproteinases (MMPs) play important roles in normal brain development and synaptic plasticity, although aberrant expression of MMPs leads to brain damage, including blood–brain barrier disruption, inflammation, demyelination, and neuronal cell death. In this article, we report that MMP-8 is upregulated in LPS-stimulated BV2 microglial cells and primary cultured microglia, and treatment of MMP-8 inhibitor (M8I) or MMP-8 short hairpin RNA suppresses proinflammatory molecules, particularly TNF-α secretion. Subsequent experiments showed that MMP-8 exhibits TNF-α–converting enzyme (TACE) activity by cleaving the prodomain of TNF-α (A74/Q75, A76/V77 residues) and, furthermore, that M8I inhibits TACE activity more efficiently than TAPI-0, a general TACE inhibitor. Biochemical analysis of the underlying anti-inflammatory mechanisms of M8I revealed that it inhibits MAPK phosphorylation, NF-κB/AP-1 activity, and reactive oxygen species production. Further support for the proinflammatory role of microglial MMP-8 was obtained from an in vivo animal model of neuroinflammatory disorder. MMP-8 is upregulated in septic conditions, particularly in microglia. Administration of M8I or MMP-8 short hairpin RNA significantly inhibits microglial activation and expression/secretion of TNF-α in brain tissue, serum, and cerebrospinal fluid of LPS-induced septic mice. These results demonstrate that MMP-8 critically mediates microglial activation by modulating TNF-α activity, which may explain neuroinflammation in septic mouse brain.

Improved oral delivery of resveratrol from N-trimethyl chitosan-g-palmitic acid surface-modified solid lipid nanoparticles

Despite the therapeutic effects of resveratrol, its clinical application is restricted by its poor oral bioavailability, low water solubility, and instability. Solid lipid nanoparticles (SLNs)-based drug delivery systems have been shown to provide excellent support for the delivery of hydrophobic drugs. The poor stability and burst release behavior in stomach acidic pH conditions of SLNs result in increased aggregation of the particles in the gastrointestinal environment, limiting the success of these particles as an oral delivery system for hydrophobic drugs. N-trimethyl chitosan (TMC) graft palmitic acid (PA) (TMC-g-PA) mucoadhesive copolymer was hypothesized to be a promising candidate for the surface modification of PA-decorated resveratrol-loaded SLNs to stabilize SLNs and circumvent all the above mentioned obstacles. TMC and TMC-g-PA copolymers were therefore synthesized and characterized by (1)H-nuclear magnetic resonance ((1)H NMR) and Fourier-transformed infra-red (FT-IR) spectroscopy. Resveratrol-loaded SLNs (SLRNs) that comprised Precirol ATO 5, PA, Gelucire 50/13, Tween 80, and resveratrol as well as TMC-g-PA SLRNs were formulated and characterized in terms of physicochemical properties, stability, cytotoxicity, and in vitro and in vivo effects. The in vitro release studies of TMC-g-PA SLRNs demonstrated negligible release of resveratrol in simulated gastric and sustained release in simulated intestinal conditions and the relative bioavailability of resveratrol was furthermore found to be 3.8-fold higher from TMC-g-PA SLRNs than that from resveratrol suspension. Overall, the findings reported here indicate that TMC-g-PA SLRNs represent a potential oral drug delivery system for resveratrol.

A validated LC-MS/MS method for quantitative analysis of curcumin in mouse plasma and brain tissue and its application in pharmacokinetic and brain distribution studies.

Curcumin is a well-known multitherapeutic agent widely employed in neurodegenerative disorders and cancer. A selective, fast, and sensitive method employing liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) was developed and validated for the simultaneous determination of curcumin in mouse plasma and brain tissue, by using salbutamol as an internal standard. Triple quadrupole mass detection with multiple reaction monitoring (MRM) mode was used to monitor the ion transitions, m/z of 369>285 for curcumin, and m/z of 240>148 for salbutamol. The method was validated for recovery, accuracy, precision, linearity, and applicability. The lower limits of quantification (LLOQ) in both matrices were 2.5ng/mL. The inter-day and intra-day precisions and accuracy values were within the Food and Drug Administration (FDA) acceptance criteria, for both matrixes. The method was successfully applied in pharmacokinetics and brain distribution studies of curcumin after intravenous administration of free curcumin and curcumin-loaded solid lipid nanoparticles to mice. Furthermore, the application of this method along with serial blood sampling in mice has led to significant reduction in animal use and dosage and drastic improvement in speed, throughput, and quality of pharmacokinetic parameters.

“Physical Activity as a Luxury”: African American Women’s Attitudes Toward Physical Activity

The purpose of this study was to explore African American midlife women’s attitudes toward physical activity. Using a feminist perspective, a 6-month online forum was conducted with 21 African American midlife women recruited on the Internet. The data were analyzed using thematic analysis. Four themes emerged: (a) culturally acceptable body, (b) missed opportunity to learn, (c) physical activity as a luxury, and (d) want to do by myself. The women had positive body images regardless of their actual weight. The women considered physical activity “a luxury” in their busy lives and thought that they had already missed opportunities to learn. The women wanted to participate in physical activities alone because of their bad childhood experiences and hesitance to go out in public with sweaty, messy hair. The findings suggested that unique programs that promote physical activity should be developed that consider the women’s ethnic-specific attitudes.

Active-targeted pH-responsive albumin–photosensitizer conjugate nanoparticles as theranostic agents

The objective of this study was to develop an active-targeted, pH-responsive albumin-photosensitizer conjugate as a theranostic agent. Herein, a porphyrin derivative photosensitizer, pheophorbide-a (PheoA), was conjugated to bovine serum albumin (BSA) via a cis-aconityl linkage, and the conjugate was then linked with polyethylene glycosylated folate to improve targeting ability. Further, BSA-c-PheoA and folate (FA)-BSA-c-PheoA at a ratio of 2 : 1 were self-assembled to form nanoparticles with a mean hydrodynamic diameter of 121.47 ± 11.60 nm. The release study exhibited that the photosensitizer was released 4.5-fold faster at pH 5.0 than at pH 7.4 when incubated for 24 h. Cellular uptake results showed that the FA-BSA-c-PheoA nanoparticles were readily taken up by B16F10 and MCF7 cancer cells. In vitro phototoxicity results showed that FA-BSA-c-PheoA NPs have higher efficacy on cancer cells compared to simple BSA-c-PheoA NPs. In vivo bioimaging results exhibited that FA-BSA-c-PheoA NPs greatly accumulated into the tumor area as compared to free PheoA. These results show that our prepared FA-BSA-c-PheoA NPs have the potential to be applied as theranostic agents in photodynamic therapy and photodiagnosis of cancer.

A national internet survey on midlife women's attitudes toward physical activity

Background:Despite an increasing number of studies of midlife women’s physical activity, little is known about how attitudes toward physical activity of midlife women from diverse ethnic groups influence the women’s physical activity. Objectives:To explore ethnic differences in midlife women’s attitudes toward physical activity and determine the relationships between the attitudes and their actual participation in physical activity while considering other influencing factors. Methods:The Midlife Women’s Attitudes Toward Physical Activity model was used to guide the study. This was a cross-sectional Internet survey study of 542 midlife women. The instruments included questions on background characteristics and health and menopausal status; the Physical Activity Assessment Inventory; a modified Barriers to Health Activities Scale; the Questions on Attitudes Toward Physical Activity, Subjective Norm, Perceived Behavioral Control, and Behavioral Intention; and the Kaiser Physical Activity Survey. The data were analyzed using ANOVA, correlation, hierarchical multiple regression, and path analyses. Results:There were significant ethnic differences in the attitude scores (F = 2.58, p < .05), but no ethnic differences in the physical activity scores. Interestingly, there were significant ethnic differences in the occupational physical activity scores (F = 5.68, p < .01). Attitude scores accounted for 5% of total variances of the physical activity scores (Fch = 43.52, p < .01). The direct paths from the attitude scores (p < .01), the self-efficacy scores (p < .01), and the barrier scores (p < .05) to the physical activity scores were statistically significant. Discussion:Ethnic differences in the women’s attitudes toward physical activity need to be considered in promoting physical activity of midlife women.