Supermagnetic Human Serum Albumin (HSA) Nanoparticles and PLGA-Based Doxorubicin Nanoformulation: A Duet for Selective Nanotherapy.

Predicting the ability of nanoparticles (NP) to access the tumor is key to the success of chemotherapy using nanotherapeutics. In the present study, the ability of the dual NP-based theranostic system to accumulate in the tumor was evaluated in vivo using intravital microscopy (IVM) and MRI. The system consisted of model therapeutic doxorubicin-loaded poly(lactide-co-glycolide) NP (Dox-PLGA NP) and novel hybrid Ce3/4+-doped maghemite NP encapsulated within the HSA matrix (hMNP) as a supermagnetic MRI contrasting agent. Both NP types had similar sizes of ~100 nm and negative surface potentials. The level of the hMNP and PLGA NP co-distribution in the same regions of interest (ROI, ~2500 µm2) was assessed by IVM in mice bearing the 4T1-mScarlet murine mammary carcinoma at different intervals between the NP injections. In all cases, both NP types penetrated into the same tumoral/peritumoral regions by neutrophil-assisted extravasation through vascular micro- and macroleakages. The maximum tumor contrasting in MRI scans was obtained 5 h after hMNP injection/1 h after PLGA NP injection; the co-distribution level at this time reached 78%. Together with high contrasting properties of the hMNP, these data indicate that the hMNP and PLGA NPs are suitable theranostic companions. Thus, analysis of the co-distribution level appears to be a useful tool for evaluation of the dual nanoparticle theranostics, whereas assessment of the leakage areas helps to reveal the tumors potentially responsive to nanotherapeutics.

Novel Nanocarrier Platform for Effective Treatment of Visceral Leishmaniasis.

Leishmaniasis is among the five parasitic diseases that still require the development of new drugs. Ultrasmall cerium (Ce3/4+) cation-doped maghemite (γ-Fe2O3) nanoparticles (NPs) were tested as a potential drug to treat visceral leishmaniasis, a disease affecting millions of people worldwide. The NPs were engineered for binding a polycationic branched polyethylenimine (PEI) polymer, thereby rupturing the single lysosome of these parasites and enabling entry of the anti-Leishmania drug, pentamidine. Exploiting the known lanthanide cation/complex-based coordinative chemical reactivity enabled the binding of both active agents onto the surface of the NPs. To optimize the fabrication of the cytotoxic NPs, optimization via a DoE (Design of Experiments) process was used to identify the optimal NP with toxicity against the two stages of the parasite, promastigotes, which propagate in the insect, and amastigotes, which infect the mammalian host. The screen identified a single optimized NP (DoE Opt) that was further examined in a mouse model of visceral leishmaniasis. Intravenous injection of the NPs had no adverse effects on the cellular composition or biochemical parameters of the blood, demonstrating no signs of systemic toxicity. The optimized NP was able to eradicate visceral disease caused by Leishmania donovani infection. The study demonstrates the versatile ability of the cerium-doped NPs to bind at least two cytotoxic ligands. This approach could be used for optimizing the binding of different drugs for the treatment of other diseases, including cancer. Since resistance to treatment with nanocarriers was not reported to date, such an approach could potentially overcome drug resistance that emerges when using soluble small molecule drugs.

Ultrasensitive haptoglobin biomarker detection based on amplified chemiluminescence of magnetite nanoparticles.

BACKGROUND: Haptoglobin is an acute-phase protein used as predicting diagnostic biomarker both in humans (i.e., diabetes, ovarian cancer, some neurological and cardiovascular disorders) and in animals (e.g., bovine mastitis). The latter is a frequent disease of dairy industry with staggering economical losses upon decreased milk production and increased health care costs. Early stage diagnosis of the associated diseases or inflammation onset is almost impossible by conventional analytical manners. RESULTS: The present study demonstrates a simple, rapid, and cost-effective label-free chemiluminescence bioassay based on magnetite nanoparticles (MNPs) for sensitive detection of haptoglobin by employing the specific interaction of hemoglobin-modified MNPs. The resulting haptoglobin-hemoglobin complex inhibits the peroxidase-like activity of luminol/H2O2-hemoglobin-MNPs sensing scheme and reduces the chemiluminescence intensities correspondingly to the innate haptoglobin concentrations. Quantitative detection of bovine haptoglobin was obtained within the range of 1 pg mL-1 to 1 µg mL-1, while presenting 0.89 pg mL-1 limit of detection. Moreover, the influence of causative pathogenic bacteria (i.e., Streptococcus dysgalactiae and Escherichia coli) and somatic cell counts (depicting healthy, sub-clinical and clinical mastitis) on the emitted chemiluminescence radiation were established. The presented bioassay quantitative performances correspond with a standardized assay kit in differentiating dissimilar milk qualities. CONCLUSIONS: Overall, the main advantage of the presented sensing concept is the ability to detect haptoglobin, at clinically relevant concentrations within real milk samples for early bio-diagnostic detection of mastitis and hence adjusting the precise treatment, potentially initiating a positive influence on animals' individual health and hence on dairy farms economy.

Noninvasive Nanodiamond Skin Permeation Profiling Using a Phase Analysis Method: Ex Vivo Experiments.

Carbon-based nanoparticles (NPs) are widely used in nanotechnology. Among them, nanodiamonds (NDs) are suitable for biotechnology and are especially interesting for skin delivery and topical treatments. However, noninvasive detection of NDs within the different skin layers or analyzing their penetration ability is complicated due to the turbid nature of the tissue. The iterative multiplane optical properties extraction (IMOPE) technique detects differences in the optical properties of the measured item by a phase-image analysis method. The phase image is reconstructed by the multiplane Gerchberg-Saxton algorithm. This technique, traditionally, detects differences in the reduced scattering coefficients. Here, however, due to the actual size of the NDs, the IMOPE technique's detection relies on absorption analysis rather than relying on scattering events. In this paper, we use the IMOPE technique to detect the presence of the NDs within tissue-like phantoms. In addition, we perform ex vivo pigskin experiments to estimate the penetration of the NDs to the different skin layers and show that their presence reduces at deeper layers. The significance signal of the NDs within the epidermis, dermis, and fat layers gradually reduces, with t test significance values that are smaller than 10-4, 10-3, and 10-2, respectively. The IMOPE results are corroborated by TEM results and Franz-cell experiments. These results confirm that the IMOPE profiled the skin-permeation of the NDs noninvasively.

Nano-Leish-IL: A novel iron oxide-based nanocomposite drug platform for effective treatment of cutaneous leishmaniasis.

Kinetoplastids are infamous parasites that include trypanosomes and Leishmania species. Here, we developed an anti-Leishmania nano-drug using ultra-small functional maghemite (γ-Fe2O3) nanoparticles (NPs) that were surface-doped by [CeLn]3/4+ to enable effective binding of the polycationic polyethylenebyimine (PEI) polymer by coordinative chemistry. This resulting nano-drug is cytolytic in-vitro to both Trypanosoma brucei parasites, the causative agent of sleeping sickness, as well as to three Leishmania species. The nano-drug induces the rupture of the single lysosome present in these parasites attributed to the PEI, leading to cytolysis. To evaluate the efficacy of a "cream-based" version of the nano-drug, which was termed "Nano-Leish-IL" for topical treatment of cutaneous leishmaniasis (CL), we developed a rapid screening method utilizing T. brucei parasites involved in social motility and demonstrated that functional NPs arrested the migration of the parasites. This assay presents a surrogate system to rapidly examine the efficacy of "cream-based" drugs in topical preparations against leishmaniasis, and possibly other dermal infectious diseases. The resulting Nano-Leish-IL topical preparation eliminated L. major infection in mice. Thus, this study presents a novel efficient nano-drug targeting the single lysosome of kinetoplastid parasites.

Iterative optical technique for detecting anti-leishmania nanoparticles in mouse lesions.

Nanoparticles (NPs) based drugs for topical administration are gaining interest in the biomedical world. However, a study tool of their penetration depth to the different tissue layers without additional markers or contrast agents is required in order to relieve safety concerns. While common diagnostic tools, e.g. X-ray, computed tomography or magnetic resonance imaging, can provide in vivo detection of the metallic NPs, their resolution cannot determine the exact penetration depth to the thin skin layers. In this work, we propose the noninvasive nanophotonics iterative multi-plane optical property extraction (IMOPE) technique for the novel iron-based NPs detection in leishmaniasis lesions. The optical properties of the different tissue layers: epidermis, dermis, subcutaneous fat and muscle, were examined before and after topical drug administration. The potential topical drug was detected in the epidermis (∼13µm) and dermis (∼160µm) layers in mice lesions at different stages of the disease (two or four weeks post infection). The lesion size influence on the detection was also observed, where in larger lesions the IMOPE senses a greater presence of the topical drug.

Innovative functional polymerization of pyrrole-N-propionic acid onto WS2 nanotubes using cerium-doped maghemite nanoparticles for photothermal therapy.

Tungsten disulfide nanotubes (WS2-NTs) were found to be very active for photothermal therapy. However, their lack of stability in aqueous solutions inhibits their use in many applications, especially in biomedicine. Few attempts were made to chemically functionalize the surface of the NTs to improve their dispersability. Here, we present a new polymerization method using cerium-doped maghemite nanoparticles (CM-NPs) as magnetic nanosized linkers between the WS2-NT surface and pyrrole-N-propionic acid monomers, which allow in situ polymerization onto the composite surface. This unique composite is magnetic, and contains two active entities for photothermal therapy-WS2 and the polypyrrole. The photothermal activity of the composite was tested at a wavelength of 808 nm, and significant thermal activity was observed. Moreover, the polycarboxylated polymeric coating of the NTs enables effective linkage of additional molecules or drugs via covalent bonding. In addition, a new method was established for large-scale synthesis of CM-NPs and WS2-NT-CM composites.

Magnetic Targeting of mTHPC To Improve the Selectivity and Efficiency of Photodynamic Therapy.

Photodynamic therapy (PDT) is a promising recognized treatment for cancer. To date, PDT drugs are injected systemically, and the tumor area is irradiated to induce cell death. Current clinical protocols have several drawbacks, including limited accessibility to solid tumors and insufficient selectivity of drugs. Herein, we propose an alternative approach to improve PDT effectiveness by magnetic targeting of responsive carriers conjugated to the PDT drug. We coordinatively attached a meso-tetrahydroxyphenylchlorin (mTHPC) photosensitizer to Ce-doped-γ-Fe2O3 maghemite nanoparticles (MNPs). These MNPs are superparamagnetic and biocompatible, and the resulting mTHPC-MNPs nanocomposites are stable in aqueous suspensions. MDA-MB231 (human breast cancer) cells incubated with the mTHPC-MNPs showed high uptake and high death rates in cell population after PDT. The exposure to external magnetic forces during the incubation period directed the nanocomposites to selected sites enhancing drug accumulation that was double that of cells with no magnetic exposure. Next, breast cancer tumors were induced subcutaneously in mice and treated magnetically. In vivo results showed accelerated drug accumulation in tumors of mice injected with mTHPC-MNP nanocomposites, compared to the free drug. PDT irradiation led to a decrease in tumor size of both groups, whereas treatment with the focused magnetic nanocomposites led to significant tumor regression. Our results demonstrate a method to improve the current PDT treatments by applying magnetic forces to effectively direct the drug to cancerous tissue. This approach leads to a highly localized and effective PDT process, opening new directions for clinical PDT protocols.

Stimulating brain recovery after stroke using theranostic albumin nanocarriers loaded with nerve growth factor in combination therapy.

For many years, delivering drug molecules across the blood brain barrier has been a major challenge. The neuropeptide nerve growth factor is involved in the regulation of growth and differentiation of cholinergic neurons and holds great potential in the treatment of stroke. However, as with many other compounds, the biomolecule is not able to enter the central nervous system. In the present study, nerve growth factor and ultra-small particles of iron oxide were co-encapsulated into a chemically crosslinked albumin nanocarrier matrix which was modified on the surface with apolipoprotein E. These biodegradable nanoparticles with a size of 212 ±â€¯1 nm exhibited monodisperse size distribution and low toxicity. They delivered NGF through an artificial blood brain barrier and were able to induce neurite outgrowth in PC12 cells in vitro. In an animal model of stroke, the infarct size was significantly reduced compared to the vehicle control. The combination therapy of NGF and the small-molecular MEK inhibitor U0126 showed a slight but not significant difference compared to U0126 alone. However, further in vivo evidence suggests that successful delivery of the neuropeptide is possible as well as the synergism between those two treatments.

Biocatalytic carbon nanotube paper: a 'one-pot' route for fabrication of enzyme-immobilized membranes for organophosphate bioremediation.

Recent world events have demonstrated the critical need for facile and miniaturized bioremediation technologies for organophosphates (OPs). These compounds are among the most toxic substances synthesized to date and are used as pesticides and nerve agents. Biotechnological methods based on the use of organophosphate hydrolase (OPH) for detoxification of OPs have drawn significant attention. This work presents a new 'one-pot' methodology for a rapid and straightforward fabrication of enzymatically active carbon nanotube (CNT) paper for OP bioremediation. Carboxylated CNTs are ultrasonically dispersed in an aqueous surfactant solution followed by a microfiltration process to generate a paper-like membrane, which is assembled from entangled nanotubes. Herein, OPH conjugation to the CNTs is carried out by carbodiimide chemistry during the microfiltration process. Successful covalent immobilization of the enzyme onto the nanotube surface is confirmed by cryo-transmission electron microscopy and infrared spectroscopy. To study the potential of this platform for OP bioremediation, an aqueous solution of methyl paraoxon (used as a model OP) is filtered with the resulting OPH-CNT membranes. A significant decrease of methyl paraoxon concentration is achieved, which is ascribed to its in situ hydrolysis by the immobilized OPH during the filtration process. These thin membranes allow many subsequent filtration cycles to be performed, while maintaining their enzymatic activity, owing to the unique combination of the mechanically robust CNT scaffold and high OPH loading. This study presents a new generic approach for the design of bioactive paper-like scaffolds, which can be rationally tailored for a variety of applications.

NMR-based molecular ruler for determining the depth of intercalants within the lipid bilayer. Part IV: studies on ketophospholipids.

In our companion paper, we described the preparation and intercalation of two homologous series of dicarbonyl compounds, methyl n-oxooctadecanoates and the corresponding n-oxooctadecanoic acids (n=4-16), into DMPC liposomes. (13)C NMR chemical shift of the various carbonyls was analyzed using an E(T)(30) solvent polarity-chemical shift correlation table and the corresponding calculated penetration depth (in Å). An iterative best fit analysis of the data points revealed an exponential correlation between E(T)(30) micropolarity and the penetration depth (in Å) into the liposomal bilayer. However, this study is still incomplete, since the plot lacks data points in the important area of moderately polarity, i.e., in the E(T)(30) range of 51-45.5 kcal/mol. To correct this lacuna, a family of ketophospholipids was prepared in which the above n-oxooctadecanoic acids were attached to the sn-2 position of a phosphatidylcholine with a palmitic acid chain at sn-1. To assist in assignment and detection several derivatives were prepared (13)C-enriched in both carbonyls. The various homologs were intercalated into DMPC liposomes and give points specifically in the missing area of the previous polarity-penetration correlation graph. Interestingly, the calculated exponential relationship of the complete graph was essentially the same as that calculated in the companion paper based on the methyl n-oxooctadecanoates and the corresponding n-oxooctadecanoic acids alone. The polarity at the midplane of such DMPC systems is ca. 33 kcal/mol and is not expected to change very much if we extend the lipid chains. This paper concludes with a chemical ruler that maps the changing polarity experienced by an intercalant as it penetrates the liposomal bilayer.

NMR-based molecular ruler for determining the depth of intercalants within the lipid bilayer: Part III: studies on keto esters and acids.

The development of "molecular rulers" would allow one to quantitatively locate the penetration depth of intercalants within lipid bilayers. To this end, an attempt was made to correlate the (13)C NMR chemical shift of polarizable "reporter" carbons (e.g., carbonyls) of intercalants within DMPC liposomal bilayers - with the polarity it experiences, and with its Angstrom distance from the interface. This requires families of molecules with two "reporter carbons" separated by a known distance, residing at various depths/polarities within the bilayer. For this purpose, two homologous series of dicarbonyl compounds, methyl n-oxooctadecanoates and the corresponding n-oxooctadecanoic acids (n=4-16), were synthesized. To assist in assignment and detection several homologs in each system were prepared (13)C-enriched in both carbonyls. Within each family, the number of carbons and functional groups remains the same, with the only difference being the location of the second ketone carbonyl along the fatty acid chain. Surprisingly, the head groups within each family are not anchored near the lipid-water interface, nor are they even all located at the same depth. Nevertheless, using an iterative best fit analysis of the data points enables one to obtain an exponential curve. The latter gives substantial insight into the correlation between polarity (measured in terms of the Reichardt polarity parameter, ET(30)) and penetration depth into the liposomal bilayer. Still missing from this curve are data points in the moderate polarity range.

A dispersability study on poly(thiophen-3-yl-acetic acid) and PEDOT multi-walled carbon nanotube composites using an analytical centrifuge.

The present study investigated the influence of a covalently grown polythiophene (polyTh)-based adlayer on the N-methyl-2-pyrrolidone (NMP) dispersion of chemically modified multi-walled carbon nanotubes (MWCNTs). Poly(thiophen-3-yl-acetic acid) (PTAA), poly(ethylenedioxythiophene) (PEDOT), and PTAA/PEDOT co-polymer have been oxidatively grown from the surface of functional MWCNTs afforded corresponding polyTh(A)-, polyTh(B)-, and polyTh(A/B)-MWCNTs, respectively. To study and quantify the MWCNT particle sedimentation behavior of corresponding dispersion systems, an analytical centrifuge system was used. Comparison of dispersion quality of such polyTh-MWCNT composites with both commercial MWCNTs (COM-MWCNTs) and oxidized MWCNTs (polyCOOH-MWCNTs) was established. Analysis of sedimentation results combined with TGA, Elemental Analysis (EA), and Kaiser Test data showed that the COOH groups present on the surface of MWCNTs strongly promote the dispersability much beyond current sedimentation rate observed for COM-MWCNTs, - sedimentation rates were higher than 80% and below 2% for oxidized polyCOOH-MWCNTs. However, regarding MWCNTs coated with polyTh polymers in a core (MWCNT)-shell (polyTh) structure, outer functional groups are not the only influential factor for promoting high dispersion qualities. For these dual phase composite systems, the weight and thickness of polyTh-shell on the composite must be considered.