Biocompatible chitosan-DNAzyme nanoparticle exhibits enhanced biological activity

DNAzymes are oligonucleotides capable of specific catalysis of target mRNA. To date, a delivery vehicle for DNAzymes has not been developed. Chitosan is a biomaterial obtained abundantly in nature. A biocompatible c-jun (an oncogene) DNAzyme nanoparticle formulated from chitosan was found to be more active against osteosarcoma (bone cancer) cells, inducing apoptotic cell death in these cells. The formulation was stable in serum for a week and at room temperature for a month. Clinically, knockdown of c-jun gene expression with chitosan nanobiotechnology may improve treatment outcome for tumours growing in bon

Downregulation of c-jun results in apoptosis-mediated anti-osteosarcoma activity in an orthotopic model

c-jun has been found to be upregulated in a variety of cancers including osteosarcoma. DNAzymes are oligonucleotides capable of specific downregulation of target genes. c-jun knockdownmediated apoptosis in osteosarcoma cells involved caspases-1, -2 and -8, but not the Fas/FasL pathway. A c-jun DNAzyme, encapsulated within a novel cationic multilamellar vesicle liposome, inhibited the growth and metastasis of osteosarcoma in an orthotopic spontaneously metastasising model of the disease. The 60 nm DDAB:DOPE liposome was formulated using ethanol injection/ extrusion. Clinically, downregulation of c-jun may proffer an improved treatment outcome for these tumors originating in bone

A nanoparticulate system that enhances the efficacy of the tumoricide Dz13 when administered proximal to the lesion site

We demonstrate that Dz13, a DNA enzyme that cleaves c-Jun mRNA, and is capable of inhibiting cancer cell growth in vitro, can be encapsulated into chitosan nanoparticles. For optimisation of this chitosan-based formulation, pH 6, 0.02% chitosan concentration, and 55 °C were found to be best among the variables tested. Particles were 50–300 nm in diameter and encapsulated Dz13 was active when particles were exposed to cancer cells. Nanoparticles were stable during storage even for a month, but were not stable in mouse and human serum. In two different clinically-relevant disease models, and using a clinically-adoptable dosing regimen, these Dz13-nanoparticles were shown to be efficacious against a bone tumour (osteosarcoma), for which no real cure exists currently. However, no toxicity against other bone-dwelling cells was observed with the formulation, and no side-effects were noted in vivo in lymphatic and reticuloendothelial tissues proximal and distal to the administration site

The performance of doxorubicin encapsulated in chitosan–dextran sulphate microparticles in an osteosarcoma model

Osteosarcoma (OS) is the most common primary bone cancer affecting children and adolescents. It is potentially debilitating and fatal due to pulmonary metastasis. A common management strategy, chemotherapy, has a 10-year disease-free survival of approximately 60%. However, a targeted approach to OS tumor inhibition is still lacking, calling for improved management strategies. A frontline drug for OS, doxorubicin (DOX), causes multiple side-effects (example myelosuppression, heart failure, hepatic toxicity, alopecia) in patients, especially in high doses required to control tumor growth. A drug delivery system (DDS) was developed to deliver DOX specifically to tumor sites. Through DOX encapsulation into chitosan DDS via the complex coacervation method with dextran sulphate, novel DOX microparticles (DMPs), with a DOX loading content of more than 99%, were formed. Multiple optimisation steps produced DMPs which caused OS cell death through apoptosis, necrosis and autophagic cell death. Treatment of mice bearing orthotopic OS with DMP decreased tumor volume, decreased bone lysis, and reduced secondary metastasis to the lungs. DMP-treated mice also maintained their weight and did not appear to suffer from any visible side-effects such as heart failure or dry skin. Thus, DMP may prove to be a useful DDS platform clinically provided further studies are performed to rigorously validate this technology

Journal of economics & management strategy : JEMS

Leukemia inhibitory factor (LIF) and Ciliary Neurotrophic factor (CNTF) are members of the interleukin-6 family of cytokines, defined by use of the gp130 molecule as an obligate receptor. In the murine experimental autoimmune encephalomyelitis (EAE) model, antagonism of LIF and genetic deletion of CNTF worsen disease. The potential mechanism of action of these cytokines in EAE is complex, as gp130 is expressed by all neural cells, and could involve immuno-modulation, reduction of oligodendrocyte injury, neuronal protection, or a combination of these actions. In this study we aim to investigate whether the beneficial effects of CNTF/LIF signalling in EAE are associated with axonal protection; and whether this requires signalling through oligodendrocytes. We induced MOG(35–55) EAE in CNTF, LIF and double knockout mice. On a CNTF null background, LIF knockout was associated with increased EAE severity (EAE grade 2.1±0.14 vs 2.6±0.19; P<0.05). These mice also showed increased axonal damage relative to LIF heterozygous mice, as indicated by decreased optic nerve parallel diffusivity on MRI (1540±207 µm(2)−/s vs 1310±175 µm(2)−/s; P<0.05), and optic nerve (−12.5%) and spinal cord (−16%) axon densities; and increased serum neurofilament-H levels (2.5 fold increase). No differences in inflammatory cell numbers or peripheral auto-immune T-cell priming were evident. Oligodendrocyte-targeted gp130 knockout mice showed that disruption of CNTF/LIF signalling in these cells has no effect on acute EAE severity. These studies demonstrate that endogenous CNTF and LIF act centrally to protect axons from acute inflammatory destruction via an oligodendrocyte-independent mechanism

Low intrinsic running capacity is associated with reduced skeletal muscle substrate oxidation and lower mitochondrial content in white skeletal muscle

Chronic metabolic diseases develop from the complex interaction of environmental and genetic factors, although the extent to which each contributes to these disorders is unknown. Here, we test the hypothesis that artificial selection for low intrinsic aerobic running capacity is associated with reduced skeletal muscle metabolism and impaired metabolic health. Rat models for low- (LCR) and high- (HCR) intrinsic running capacity were derived from genetically heterogeneous N:NIH stock for 20 generations. Artificial selection produced a 530% difference in running capacity between LCR/HCR, which was associated with significant functional differences in glucose and lipid handling by skeletal muscle, as assessed by hindlimb perfusion. LCR had reduced rates of skeletal muscle glucose uptake (∼30%; P = 0.04), glucose oxidation (∼50%; P = 0.04), and lipid oxidation (∼40%; P = 0.02). Artificial selection for low aerobic capacity was also linked with reduced molecular signaling, decreased muscle glycogen, and triglyceride storage, and a lower mitochondrial content in skeletal muscle, with the most profound changes to these parameters evident in white rather than red muscle. We show that a low intrinsic aerobic running capacity confers reduced insulin sensitivity in skeletal muscle and is associated with impaired markers of metabolic health compared with high intrinsic running capacity. Furthermore, selection for high running capacity, in the absence of exercise training, endows increased skeletal muscle insulin sensitivity and oxidative capacity in specifically white muscle rather than red muscle. These data provide evidence that differences in white muscle may have a role in the divergent aerobic capacity observed in this generation of LCR/HCR

Co-nanoencapsulated doxorubicin and Dz13 control osteosarcoma progression in a murine model

Objectives: Chitosan is a green (natural, abundant, biodegradable, biocompatible) biopolymer that can be formulated to encapsulate a variety of therapeutic compounds. This study aimed to investigate chitosan nanoparticles (NPs) as a means of improving delivery of the clinically used anti-cancer agent doxorubicin (Dox) and the preclinical lead compound Dz13 oligonucleotide together. Methods: A novel chitosan NP system encapsulating Dox and Dz13 was designed, biophysically characterised and tested in a clinically relevant model of the metastasising bone tumour, osteosarcoma (OS). Key findings: By careful alteration of the concentration of the individual components, a final formulation of Dz13-Dox NPs (DDNPs) was achieved, with high (>91%) loading of both compounds, which consisted of individual 50-nm particles forming aggregates as large as 500 nm, with a large positive ζ-potential. The DDNPs could be stored at various temperatures for a week without loss in activity but were prone to degradation in serum. DDNPs successfully inhibited OS tumour growth more effectively than treatment with NPs of Dz13 and Dox-chitosan, as well as Dox administered intraperitoneally. Apart from inhibiting tumour growth, DDNPs protected the affected bone from substantial destruction by aggressive tumour growth and reduced the incidence of metastasis to the lungs without causing adverse effects in mice. Conclusion: This NP is a promising formulation that could be useful for clinical management of OS

EAE disease severity grades.

<p>EAE disease severity scores for CNTF WT/LIF WT (n = 20) vs CNTF WT/LIF HET (n = 9) mice (A); CNTF WT/LIF HET (n = 30) vs CNTF WT/LIF KO (n = 21) mice (B); CNTF KO/LIF WT (n = 19) vs CNTF KO/LIF HET mice (n = 17) (C); and CNTF KO/LIF HET (n = 30) vs CNTF KOLIF KO (18) mice (D), over time. In CNTF gene deficient mice, the presence of the LIF gene was associated with reduced EAE disease severity scores relative to LIF gene deficient mice, from EAE day 15–18 (D)(*P&lt;0.05). All data are presented as mean ± SEM.</p

Analyses of CNS cell responses: Representative images of CD3 labelled T-lymphocyte cells (A, B), IBA1 labelled microglia/macrophages (C, D), NIMP-R14 labelled neutrophils (E, F) and GFAP labelled astrocytes (G, H), in lumbar spinal cord sections from CNTF

<p>Cell counts were conducted in the dorsal column (Dorsal) and at the lateral edges (Lateral) for CD3 labelled T-lymphocytes (I, J), IBA1 labelled microglia/macrophages (K, L), NIMP-R14 labelled neutrophils (M, N), and GFAP labelled astrocytes (O, P). No differences in cell numbers were found between the groups, Data are presented as mean ± SEM. Scale bar represents 50 µm.</p

Optic nerve histological parameters for CNTF KO/LIF KO vs. CNTF KO/LIF HET healthy and EAE induced mice.

<p>Comparison of key morphometric measures in the optic nerve between CNTF KO/LIF KO (LIF KO) and CNTF KO/LIF HET (LIF HET) mice in the presence and absence of EAE. Data are presented as mean ± SEM values for each parameter for each group. Region of interest (ROI) represents an area of 3480 µm².</p>*<p>P&lt;0.05 by 2-sample t-test, assuming equal variances.</p