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Diffusion cells are routinely used in pharmacology to measure the permeation of pharmaceutical compounds and contaminants across membranes (biological or synthetic). They can also be used to study drug release from excipients. The device is made of a donor (DC) and an acceptor (AC) compartment, separated by a membrane. Usually, permeation of molecules across membranes is measured by sampling from the AC at different time points. However, this process disturbs the equilibrium of the cell. Furthermore, analytical techniques used in association with diffusion cells sometimes lack either accuracy, sensitivity, or both. This work reports on the development of nuclear imaging – compatible diffusion cells. The cell is made of a polymer transparent to high-energy photons typically detected in positron emission tomography (PET). It was tested in a finite-dose set-up experiment with a pre-clinical PET system. Porous cellulose membranes (3.5, 25 and 300 kDa), a common excipient in pharmacology, as well as for dialysis membranes, were used as test membranes. The radioisotope 89Zr chelated with deferoxamine B (DFO; 0.65 kDa), was used as an imaging probe (7–10 MBq; 0.2–0.3 nMol 89Zr-DFO). In medicine, DFO is also commonly used for iron removal treatments and pharmacological formulations often require the association of this molecule with cellulose. Permeation profiles were obtained by measuring the radioactivity in the DC and AC for up to 2 weeks. The kinetic profiles were used to extract lag time, influx, and diffusion coefficients of DFO across porous cellulose membranes. A sensitivity threshold of 0.005 MBq, or 3.4 fmol of 89Zr-DFO, was revealed. The lag time to permeation (τ) measured in the AC compartment, was found to be 1.33, 0.5, and 0.19 h with 3.5, 25, and 300 kDa membranes, respectively. Diffusion coefficients of 3.65 × 10−6, 8.33 × 10−6, and 4.74 × 10−5 cm2 h−1 where revealed, with maximal pseudo steady-state influx values (Jpss) of 6.55 × 10−6, 1.76 × 10−5, and 1.29 × 10−5 nmol cm−2 h−1. This study confirms the potential of the technology for monitoring molecular diffusion and release processes at low concentrations, high sensitivities, in real time and in a visual manner.

Peut-être pour que dans vingt ans, dans trente ans, si les règles changent, l’île pourrait en avoir besoin. En même temps vous allez aider à amener du touriste icitte, si vous prenez quelques-uns marsouins, pour faire faire de l’argent. Alexis Tremblay dans Pour la suite du monde de Pierre Perreault (1963) Le tourisme est passé d’un appel à voir le monde à l’invitation de faire partie de la trame même du monde. Et puis, comme nous l’avons lu tout au long de ce numéro spécial, la COVID-19 a ro...

Gold nanoparticles (Au NPs) distributed in the vicinity of low-dose rate (LDR) brachytherapy seeds could multiply their efficacy thanks to the secondary emissions induced by the photoelectric effect. Injections of radioactive LDR gold nanoparticles (LDR Au NPs), instead of conventional millimeter-size radioactive seeds surrounded by Au NPs, could further enhance the dose by distributing the radioactivity more precisely and homogeneously in tumors. However, the potential of LDR Au NPs as an emerging strategy to treat cancer is strongly dependent on the macroscopic diffusion of the NPs in tumors, as well as on their microscopic internalization within the cells. Understanding the relationship between interstitial and intracellular distribution of NPs, and the outcomes of dose deposition in the cancer tissue is essential for considering future applications of radioactive Au NPs in oncology. Here, LDR Au NPs (103Pd:Pd@Au-PEG NPs) were injected in prostate cancer tumors. The particles were visualized at time-po...

Ultra-small superparamagnetic iron oxide nanoparticles (USPIOs) have been used as vascular contrast agents in magnetic resonance imaging (MRI), mainly for their capacity to generate negative contrast. To use USPIOs as positive contrast agents, it is necessary to achieve increased colloidal stability and signal-enhancement performance. Their molecular coatings must be carefully chosen so the vascular blood-pool contrast agents must lead to long blood turnover times. However, to avoid long-term toxicological effects, they must also be cleared rapidly through the urinary or the gastrointestinal pathways. In this context, highly stable USPIOs showing “positive” contrast in MRI and optimal clearance rates, call for the development of robust biocompatible molecular coatings. In the present study, USPIOs were stabilized with multidentate block copolymer (MDBC), using a one-pot polyol synthesis method in the presence of MDBC. Two types of MDBC having pendant COOH groups in the anchoring block were developed: a po...

Apres l’obtention de son baccalaureat en gestion du tourisme et de l’hotellerie a l’Ecole des sciences de la gestion (ESG) de l’Universite du Quebec a Montreal (UQAM), Kevin Sauvageau a occupe des postes au sein de departements commerciaux d’etablissements hoteliers en plus de travailler en tant que consultant chez Horwath HTL, ou il a effectue des etudes de marche et de faisabilite de projets. Aujourd’hui, Kevin travaille chez Stay22 en tant que charge des partenariats et du developpement de...

Mesoporous silica nanoparticles (MSNs) are being developed as drug delivery vectors. Biomedical imaging (MRI and PET) enables their tracking in vivo, provided their surface is adequately grafted with imaging probes (metal chelates). However, MSNs are characterized by huge specific surfaces, and high-quality metal chelate anchoring procedures must be developed and validated, to demonstrate that their detection in vivo is associated to the presence of nanoparticles and not to detached metal chelates. MCM-48 nanospheres (M48SNs, 150 nm diam., 3-D pore geometry) were synthesized and functionalized with diethylenetriaminepentaacetic acid (DTPA). The strong grafting of DTPA was confirmed by 29Si MAS-NMR, XPS, FTIR and TGA. The particles were labeled with paramagnetic ions Gd3+ (for MRI) as well as radioactive ions 64Cu2+ (for PET; half-life: 12.7 h). Gd3+-DTPA-M48SNs formed a stable colloid in saline media for at least 6 months, without any sign of aggregation. The relaxometric properties were measured at various magnetic fields. The strength of DTPA binding at the surface of MSNs was also assessed in vivo, by injecting mice (i.v.) with Gd3+/64Cu2+-DTPA-M48SNs. Vascular retention and urinary clearance were monitored by MRI, whereas the PET modality provided dynamic and quantitative assessment of biodistribution and blood/organ clearance. No significant 64Cu activity was detectable in the bladder. The study confirmed the very limited detachment of DTPA from M48SNs cores once injected in vivo. The transit of MSNs through the liver and intestinal tract, does not lead to evidence of Gd3+/64Cu2+-DTPA in the urine. This physico-chemical and biodistribution study confirms the quality of DTPA attachment at the surface of the particles, necessary to allow further development of PET/MRI-assisted MSN-vectorized drug delivery procedures.

The radioisotope palladium (103Pd), encapsulated in millimetre-size seed implants, is widely used in prostate cancer brachytherapy. Gold nanoparticles (Au NPs) distributed in the vicinity of 103Pd radioactive implants, strongly enhance the therapeutic dose of radioactive implants (radiosensitisation effect). A new strategy under development to replace millimetre-size implants, consist in injecting radioactive NPs in the affected tissues. The development of 103Pd@Au NPs distributed in the diseased tissue, could increase the uniformity of treatment (compared with massive seeds), while enhancing the radiotherapeutic dose to the cancer cells (through Au-mediated radiosensitisation effect). To achieve this goal, it is necessary to develop a rapid, efficient, one-pot and easy-to-automatise procedure, allowing the synthesis of core-shell Pd@Au NPs. The novel synthesis route proposed here enables the production of Pd@Au NPs in not more than 4 h, in aqueous media, with minimal manipulations, and relying on biocompatible and non-toxic molecules. This rapid multi-step process consists of the preparation of ultra-small Pd NPs by chemical reduction of an aqueous solution of H2PdCl4 supplemented with ascorbic acid (AA) as reducing agent and 2,3-meso-dimercaptosuccinic acid (DMSA) as a capping agent. Pd conversion yields close to 87% were found, indicating the efficiency of the reaction process. Then Pd NPs were used as seeds for the growth of a gold shell (Pd@Au), followed by grafting with polyethylene glycol (PEG) to ensure colloidal stability. Pd@Au-PEG (TEM: 20.2 ± 12.1 nm) formed very stable colloids in saline solution as well as in cell culture medium. The physico-chemical properties of the particles were characterised by FTIR, XPS, and UV-vis. spectroscopies. The viability of PC3 human prostate cancer cells was not affected after a 24 h incubation cycle with Pd@Au-PEG NPs to concentrations up to 4.22 mM Au. Finally, suspensions of Pd@Au-PEG NPs measured in computed tomography (CT) are found to attenuate X-rays more efficiently than commercial Au NPs CT contrast media. A proof-of-concept was performed to demonstrate the possibility synthesise radioactive 103Pd:Pd@Au-PEG NPs. This study reveals the possibility to synthesise Pd@Au NPs rapidly (including radioactive 103Pd:Pd@Au-PEG NPs), and following a methodology that respects all the strict requirements underlying the production of NPs for radiotherapeutic use (rapidity, reaction yield, colloidal stability, NPs concentration, purification).

Ultrasmall paramagnetic Gd(2)O(3) nanoparticles have been developed as contrast agents for molecular and cellular preclinical MRI procedures. These small particles (mean diameter

Several studies have shown that 65 % of expanded poly (tetrafluoroethylene) (ePTFE) vascular prostheses had to be explanted within 10 years of implantation in humans. The reasons for these explantations relied on thrombosis formation and poor hemocompatibility of synthetic polymers. It has been shown that surface modification of ePTFE arterial prostheses could enable their endothelialization therefore improving their biocompatibility and hemocompatibility. Indeed, endothelial cells naturally cover the biological blood vessel wall and consequently, an endothelial layer constitutes the best achievable hemocompatible surface. In this context, our strategy consisted in micropatterning cell adhesion (RGD) and proliferation (WQPPRARI) peptides on the surface of plasma-functionalized PTFE, therefore enabling covalent conjugation of the peptides. Basically, the technology consisted in spraying a solution of the adhesion peptide, therefore leading to 10 µm-diameter RGD spots semi-randomly distributed over the sample and covering 20 % of the whole polymer surface. In a second step, proliferation peptide was applied to the remaining surface by soaking, therefore covering the unreacted surface. The 20 % coverage was obtained by using an x-y table, programmed to move from side to side of the surface on x value, with an increment on y value that has been calibrated.

Cardiovascular diseases are increasingly becoming the main cause of death all over the world, leading to an increase in the economical and social burden. Vascular tissue engineering (VTE) is paving its routes toward challenging applications, focused mainly on substitutions of small-diameter blood vessels (

In recent years, pulsed laser ablation in liquids (PLAL) has emerged as a new green chemistry method to produce different types of nanoparticles (NPs). It does not require the use of reducing or stabilizing agents, therefore enabling the synthesis of NPs with highly-pure surfaces. In this study, pure Au NPs were produced by PLAL in aqueous solutions, sterically stabilized using minimal PEG excess, and functionalized with manganese chelates to produce a dual CT/MRI contrast agent. The small hydrodynamic size (36.5 nm), low polydispersity (0.2) and colloidal stability of Au NPs@PEG-Mn2+ were demonstrated by DLS. The particles were further characterized by TEM, XPS, FTIR and 1H NMR to confirm the purity of the Au surfaces (i.e. free from the common residual chemicals found after NP synthesis) and the presence of the different surface molecules. The potential of these particles as contrast agents for CT/MRI was assessed in vivo (e.g. chicken embryo). Au NPs@PEG-Mn2+ also demonstrated strong blood retention for at least 90 minutes following intravenous injection in mouse models. The promising performance of PEGylated PLAL-synthesized Au NPs containing manganese chelates could open new possibilities for the production of purer dual imaging contrast agents based on Au colloids.

Progresses in cold atmospheric plasma technologies have made possible the synthesis of nanoparticles in aqueous solutions using plasma electrochemistry principles. In this contribution, a reactor based on microhollow cathodes and operating at atmospheric pressure was developed to synthesize iron-based nanoclusters (nanoparticles). Argon plasma discharges are generated at the tip of the microhollow cathodes, which are placed near the surface of an aqueous solution containing iron salts (FeCl2 and FeCl3) and surfactants (biocompatible dextran). Upon reaction at the plasma-liquid interface, reduction processes occur and lead to the nucleation of ultrasmall iron-based nanoclusters (IONCs). The purified IONCs were investigated by XPS and FTIR, which confirmed that the nucleated clusters contain a highly hydrated form of iron oxide, close to the stoichiometric constituents of α-FeOOH (goethite) or Fe5O3(OH)9 (ferrihydrite). Relaxivity values of r1 = 0.40 mM(-1) s(-1) and r2/r1 = 1.35 were measured (at 1.41 T); these are intermediate values between the relaxometric properties of superparamagnetic iron oxide nanoparticles used in medicine (USPIO) and those of ferritin, an endogenous contrast agent. Plasma-synthesized IONCs were injected into the mouse model and provided positive vascular signal enhancement in T1-w. MRI for a period of 10-20 min. Indications of rapid and strong elimination through the urinary and gastrointestinal tracts were also found. This study is the first to report on the development of a compact reactor suitable for the synthesis of MRI iron-based contrast media solutions, on site and upon demand.

Adipose tissue (AT) substitutes are being developed to answer the strong demand in reconstructive surgery. To facilitate the validation of their functional performance in vivo, and to avoid resorting to excessive number of animals, it is crucial at this stage to develop biomedical imaging methodologies, enabling the follow-up of reconstructed AT substitutes. Until now, biomedical imaging of AT substitutes has scarcely been reported in the literature. Therefore, the optimal parameters enabling good resolution, appropriate contrast, and graft delineation, as well as blood perfusion validation, must be studied and reported. In this study, human adipose substitutes produced from adipose-derived stem/stromal cells using the self-assembly approach of tissue engineering were implanted into athymic mice. The fate of the reconstructed AT substitutes implanted in vivo was successfully followed by magnetic resonance imaging (MRI), which is the imaging modality of choice for visualizing soft ATs. T1-weighted images allowed clear delineation of the grafts, followed by volume integration. The magnetic resonance (MR) signal of reconstructed AT was studied in vitro by proton nuclear magnetic resonance ((1)H-NMR). This confirmed the presence of a strong triglyceride peak of short longitudinal proton relaxation time (T1) values (200 ± 53 ms) in reconstructed AT substitutes (total T1=813 ± 76 ms), which establishes a clear signal difference between adjacent muscle, connective tissue, and native fat (total T1 ~300 ms). Graft volume retention was followed up to 6 weeks after implantation, revealing a gradual resorption rate averaging at 44% of initial substitute's volume. In addition, vascular perfusion measured by dynamic contrast-enhanced-MRI confirmed the graft's vascularization postimplantation (14 and 21 days after grafting). Histological analysis of the grafted tissues revealed the persistence of numerous adipocytes without evidence of cysts or tissue necrosis. This study describes the in vivo grafting of human adipose substitutes devoid of exogenous matrix components, and for the first time, the optimal parameters necessary to achieve efficient MRI visualization of grafted tissue-engineered adipose substitutes.

Tripositive gadolinium-ion doped NIR-to-NIR upconverting paramagnetic nanoparticles are efficiently detected are NIR imaging techniques but can also provide efficient "positive" contrast in MRI. On page 1478 John A. Capobianco, Marc-Andre Fortin, and co-workers show that citrate-coated nanoparticles present the lowest relaxometric ratios reported for NaGdF4 nanoparticle suspensions. IV-injected nanoparticles evidence long blood retention times in mice while biodistribution studies show elimination through the reticuloendothelial and urinary systems.

The aryl hydrocarbon receptor (AHR) is a ligand-dependent transcriptional regulator of several genes including the cytochrome P4501 (CYP1) family as well as genes encoding factors involved in cell growth and differentiation. In mice, several polymorphic forms of the AHR are known, some of which have

Prenatal exposure to tobacco smoke represents an important confounding factor in epidemiological studies addressing developmental effects and requires careful controlling by the use of biomarkers. We compared the following biomarkers of exposure to tobacco smoke during pregnancy and related biological effects in 23 smokers and 17 non-smokers: placental concentrations of heavy metals (cadmium, chrome, lead and zinc), cotinine concentration in meconium, placental CYP1A1 activity (EROD) and bulky DNA adducts. Cadmium was detected in all samples and found in higher concentration in placentas of smokers compared with non-smokers (geometric mean ± GSD: 56.1 ± 1.8 vs 27.4 ± 1.6 μg kg -1 dry weight; p < 0.001). Cotinine was not detected in meconium samples from the non-smoker group, while samples from the smoker group contained a mean concentration of 114.1 ± 2.9 μg kg -1 . Correlation analysis of biomarkers among smokers revealed that daily cigarette consumption was strongly correlated to placental cadmium (Pearson's r = 0.83, p < 0.001) and to cotinine (r = 0.73, p < 0.001). EROD activity was also higher in smokers than in non-smokers (9.4 ± 3.4 vs 2.5 ± 1.8 pmol resorufin min -1 mg -1 protein; p < 0.001) and values were correlated to cotinine concentration in meconium (r = 0.80, p < 0.001) and placental cadmium level (r = 0.66, p < 0.001). The amount of bulky DNA adducts in placenta was highly variable and poorly associated with smoking status. Because of their high sensitivity and specificity to detect women who smoke during pregnancy, cotinine concentrations in meconium and placental EROD activity should be incorporated in epidemiological studies that investigate adverse developmental effects induced by in utero exposure to environmental contaminants.

Organochlorine compounds bioaccumulate in fishing and hunting products included in the daily diet of many coastal populations. Prenatal and perinatal exposure to large doses of PCBs and PCDFs was shown to be deleterious on fetal and neonatal development, but information is scarce regarding possible effects of chronic low-dose exposure. This study investigates biomarkers of early effects in newborns from women exposed to organochlorines through the consumption of species from marine food chains, in two remote coastal regions of the province of Quebec (Canada). A CYP1A1-dependent enzyme activity (EROD) and DNA adducts were measured in placenta samples obtained from 30 women living on the Lower-North-Shore of the St. Lawrence River and 22 Inuit women from Nunavik (Arctic Quebec). These biomarkers were also assessed in 30 women from a Quebec urban center (Sept-Iles) as a reference group. Prenatal organochlorine exposure was determined by measuring these compounds in umbilical cord plasma. The amount of bulky polycyclic aromatic hydrocarbon (PAH)-related DNA adducts was significantly greater in the Lower-North-Shore group than in the reference group. Placental EROD activity and the amount of less bulky (OC-related) DNA adducts were significantly higher in the Nunavik group than in the reference group. For both biomarkers, smoking was found to be an important confounding factor. Organochlorine exposure was significantly associated with EROD activity and DNA adduct levels when stratifying for smoking. This study confirms that CYP1A1 enzyme induction and DNA adducts in placental tissue constitute useful biomarkers of early effects induced by environmental exposure to organochlorines.

Commercial formulations of the pesticides: Guthion (azinphos methyl), Sencor (metribuzin), Lorox (linuron), Reglone (diquat), Daconil (chlorothalonil) and Admire (imidacloprid) were studied for their genotoxicity by 32P-postlabeling. Metabolites of the pesticides were obtained enzymatically using arochlor induced rat liver S9 fraction, in an NADPH generating system. The resulting metabolites were reacted with calf thymus DNA and the DNA was analyzed for presence of adducts by either the nuclease P1 or butanol enrichment. Nuclease P1 enrichment resulted in adducts for all the pesticides. Compared to the level of adducts in control DNA, the levels in pesticide-treated DNA were higher for all the pesticides, except Daconil. The increase in adduct numbers for pesticide-treated DNAs ranged from 4.9-12.4 times the control-DNA indicating pesticide genotoxicity in this in vitro system. Enrichment using butanol extraction gave three adducts unique to Sencor-DNA. These adducts were different from those obtained with nuclease P1 enrichment of the same. B(α)P was the positive control for the in vitro metabolism, and two adduct enrichment procedures: nuclease P1 digestion and butanol extraction.

A microfluorometric method for the detection of low levels of cytochrome P450 was developed to increase the sensitivity of the assay, since a low level of CYP450 associated enzymatic activities was expected in human placenta tissues and small samples of placenta (∼10 g) could be easily collected, stored and processed. The dual fluorescence assay of Kennedy et al. [1], which was developed to simultaneously quantitate microsomal proteins and ethoxyresorufin-O-deethylase (EROD) activity was adapted for 96 wells microtiter plates. Placental microsomes samples were analyzed. For samples obtained from non-smoking mothers from the general southern Quebec population, results ranged from less than 1–3.3 pmol/mg protein•min. Samples collected from smoking mothers showed activity levels ranging from 30–69 pmol/mg protein•min. These results showed the suitability of the microassay for measuring low level of CYP450 activity in tissues such as placenta. (Mol Cell Biochem 175: 131–136, 1997)

Mesoporous silica nanoparticles (MSNs) are used in drug delivery and cell tracking applications. As Mn(2+) is already implemented as a "positive" cell contrast agent in preclinical imaging procedures (in the form of MnCl2 for neurological studies), the introduction of Mn in the porous network of MSNs would allow labelling cells and tracking them using MRI. These particles are in general internalized in endosomes, an acidic environment with high saline concentration. In addition, the available MSN porosity could also serve as a carrier to deliver medical/therapeutic substances through the labelled cells. In the present study, manganese oxide was introduced in the porous network of MCM-48 silica nanoparticles (Mn-M48SNs). The particles exhibit a narrow size distribution (~140 nm diam.) and high porosity (~60% vol.), which was validated after insertion of Mn. The resulting Mn-M48SNs were characterized by TEM, N2 physisorption, and XRD. Evidence was found with H2-TPR, and XPS characterization, that Mn(II) is the main oxidation state of the paramagnetic species after suspension in water, most probably in the form of Mn-OOH. The colloidal stability as a function of time was confirmed by DLS in water, acetate buffer and cell culture medium. In NMR data, no significant evidence of Mn(2+) leaching was found in Mn-M48SNs in acidic water (pH 6), up to 96 hours after suspension. High longitudinal relaxivity values of r1 = 8.4 mM(-1) s(-1) were measured at 60 MHz and 37 °C, with the lowest relaxometric ratios (r2/r1 = 2) reported to date for a Mn-MSN system. Leukaemia cells (P388) were labelled with Mn-M48SNs and nanoparticle cell internalization was confirmed by TEM. Finally, MRI contrast enhancement provided by cell labelling with escalated incubation concentrations of Mn-M48SNs was quantified at 1 T. This study confirmed the possibility of efficiently confining Mn into M48SNs using incipient wetness, while maintaining an open porosity and relatively high pore volume. Because these Mn-labelled M48SNs express strong "positive" contrast media properties at low concentrations, they are potentially applicable for cell tracking and drug delivery methodologies.

Near-infrared (NIR)-to-NIR upconverting NaY(Gd)F4 :Tm(3+) ,Yb(3+) paramagnetic nanoparticles (NPs) are efficiently detected by NIR imaging techniques. As they contain Gd(3+) ions, they also provide efficient "positive" contrast in magnetic resonance imaging (MRI). Water-dispersible small (≈25 nm, "S-") and ultrasmall (5 nm diam., "US-") NaY(Gd)F4 :Tm(3+) ,Yb(3+) NPs are synthesized by thermal decomposition and capped with citrate. The surface of citrate-coated US-NPs shows sodium depletion and high Gd elemental ratios, as confirmed by a comparative X-ray photoelectron spectroscopy (XPS)/neutron absorption analysis study. US-NaGd0.745 F4 :Tm0.005 ,Yb0.25 NPs have hydrodynamic diameters close to that measured by TEM, with the lowest relaxometric ratios (r2 /r1 = 1.18) reported for NaGdF4 nanoparticle suspensions (r1 = 3.37 mM(-1) s(-1) at 1.4 T and 37 °C). Strong relaxivity peaks in the range of 20 (0.47 T) - 300 MHz (7.05 T) are revealed in nuclear magnetic resonance dispersion profiles, with high r2 /r1 ratios at increasing field strengths for S-NPs. This indicates the superiority of US-NPs over S-NPs for achieving high positive contrast at clinical MRI field strengths. I.-v. injected citrate-coated US-NPs evidence long blood retention times (90 min) in mice. Biodistribution studies (48 h, 8 d) show elimination through the reticuloendothelial and urinary systems, similarly to other citrate-capped US-NP systems. In summary, upconverting NaY(Gd)F4 :Tm(3+) ,Yb(3+) nanoparticles have promising luminescent, relaxometric and blood-retention properties for dual MRI/optical imaging.

Prostate cancer (PCa) is one of the leading causes of death among men. Low-dose brachytherapy is an increasingly used treatment for PCa, which requires the implantation of tens of radioactive seeds. This treatment causes discomfort; these implants cannot be removed, and they generate image artifacts. In this study, the authors report on intratumoral injections of radioactive gold nanoparticles (Au NPs) as an alternative to seeds. The particles (Pd-103:Pd@Au-PEG and Pd-103:Pd@Au-198:Au-PEG; 10-14 nm Pd@Au core, 36-48 nm hydrodynamic diameter) are synthesized by a one-pot process and characterized by electron microscopy. Administrated as low volume (2-4 mu L) single doses (1.6-1.7 mCi), the particles are strongly retained in PCa xenograft tumors, impacting on their growth rate. After 4 weeks, a tumor volume inhibition of 56% and of 75%, compared to the controls, is observed for Pd-103:Pd@Au-PEG NPs and Pd-103:Pd@Au-198: Au-PEG NPs, respectively. Skin necrosis is observed with Au-198; therefore, Au NPs labeled with Pd-103 only are a more advisable choice. Overall, this is the first study confirming the impact of Pd-103@Au NPs on tumor growth. This new brachytherapy procedure could allow tunable doses of radioactivity, administered with smaller needles than with the current technologies, and leading to fewer image artifacts.

Manganese oxide (MnO) nanoparticles have been suggested as a promising "positive" MRI contrast agent for cellular and molecular studies. Mn-based contrast agents could enable T(1)-weighted quantitative cell tracking procedures in vivo based on signal enhancement. In this study, ultrasmall MnO particles were synthesized and coated with thiolated molecules (DMSA) and polyethylene glycol (PEG) to allow enhanced cell labeling properties and colloidal stability. This coating allowed the fabrication of individual ultrasmall nanoparticles of MnO (USPMnO) as well as of nanoaggregates of the same material (SPMnO). Particle size was measured by TEM and DLS. Physico-chemical properties were characterized by XPS and FTIR. The relaxometric properties of these aqueous suspensions were measured at various magnetic fields. The suspensions provided strong positive contrast enhancement in T(1)-weighted imaging due to high longitudinal relaxivities (r(1)) and low r(2)/r(1) ratios (USPMnO: r(1) = 3.4 ± 0.1 mM(-1)s(-1), r(2)/r(1) = 3.2; SPMnO: r(1) = 17.0 ± 0.5 mM(-1)s(-1), r(2)/r(1) = 4.0, at 1.41T). HT-1080 cancer cells incubated with the contrast agents were clearly visualized in MRI for Mn contents1.1 pg Mn/cell. The viability of cells was not affected, contrarily to cells labeled with an equivalent concentration of Mn(2+) ions. A higher signal per cell was found for SPMnO-labeled compared with USPMnO-labeled cells, due to the higher relaxometric properties of the agglomerates. As a result, the "positive" signal enhancement effect is not significantly affected upon agglomeration of MnO particles in endosomes. This is a major requirement in the development of reliable cell tracking procedures using T(1)-weighted imaging sequences. This study confirms the potential of SPMnO and USPMnO to establish more quantitative cell tracking procedures with MRI.

Intimal hyperplasia and thrombosis are responsible for the poor patency rates of small-diameter vascular grafts. These complications could be avoided by a rapid and strong adhesion of endothelial cells to the prosthetic surfaces, which typically consist of expanded polytetrafluoroethylene (PTFE) for small-diameter vessels. We have previously described two peptide micropatterning strategies that increase the endothelialization rates of PTFE. The micropatterns were generated either by inkjet printing 300 μm squares or by spraying 10.1 ± 0.1 μm diameter droplets of the CGRGDS cell adhesion peptide, while the remaining surface was functionalized using the CWQPPRARI cell migration peptide. We now directly compare these two micropatterning strategies and examine the effect of hydrodynamic stress on human saphenous vein endothelial cells grown on the patterned surfaces. No significant differences in cell adhesion were observed between the two micropatterning methods. When compared to unpatterned surfaces treated with a uniform mixture of the two peptides, the cell expansion was significantly higher on sprayed or printed surfaces after 9 days of static cell culture. In addition, after 6 h of exposure to hydrodynamic stress, the cell retention and cell cytoskeleton reorganization on the patterned surfaces was improved when compared to untreated or random treated surfaces. These results indicate that micropatterned surfaces lead to improved rates of PTFE endothelialization with higher resistance to hydrodynamic stress. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 101A: 694– 703, 2013.

The turnover of the homopolymer of ADP-ribose, which is known to be involved in many DNA-related functions, is controlled by 2 principal enzymes. Poly(ADP-ribose) polymerase (EC 2.4.2.30) synthesizes the polymer from NAD, and poly(ADP-ribose) glycohydrolase (PARG) is the major enzyme responsible for its catabolism (Thomassin et al. (1992) Biochim. Biophys. Acta 1137, 171–181). In vivo, poly(ADP-ribose) polymers constitute a heterogeneous population of branched polymers attaining sizes of 200–400 residues. They are rapidly degraded by PARG, displaying variable kinetic parameters as a function of polymer size. Several studies have suggested that PARG acts exoglycosidically on its substrate but others observed that it could act endo/exo-glycosidically. We analysed the mode of action of PARG under conditions most suitable for expression of all the activities of PARG, using HPLC purified long free polymer and very pure PARG. We conclusively show that on large free polymers, PARG exhibits endoglycosidic activity along with exoglycosidic activity. This endoglycosidic activity could have a significant role during cellular response to DNA damage.

The majority of contrast agents used in magnetic resonance imaging (MRI) is based on the rare-earth element gadolinium. Gadolinium-based nanoparticles could find promising applications in pre-clinical diagnostic procedures of certain types of cancer, such as glioblastoma multiforme. This is one of the most malignant, lethal and poorly accessible forms of cancer. Recent advances in colloidal nanocrystal synthesis have led to the development of ultra-small crystals of gadolinium oxide (US-Gd(2)O(3), 2-3 nm diameter). As of today, this is the smallest and the densest of all Gd-containing nanoparticles. Cancer cells labeled with a sufficient quantity of this compound appear bright in T(1)-weighted MRI images. Here we demonstrate that US-Gd(2)O(3) can be used to label GL-261 glioblastoma multiforme cells, followed by localization and visualization in vivo using MRI. Very high amounts of Gd are efficiently internalized and retained in cells, as confirmed with TEM and ICP-MS. Labeled cells were visualized in vivo at 1.5 T using the chicken embryo model. This is one more step toward the development of "positively contrasted" cell tracking procedures with MRI.

We have characterized the effect of poly(ADP-ribose) polymerase automodification on the enzyme's activities, which include poly(ADP-ribose) synthesis and NADase activity. The apparent Km of the enzyme for NAD+ during polymer synthesis is higher than the one measured for alternate NADase activity. Furthermore, we have found that there are 28 automodification sites, in contrast to the 15 sites (postulated to be on the 15 glutamic acids) reported to be present in the automodification domain. For the first time, we show that some of these acceptor sites are outside the reported automodification domain (15 kDa); we demonstrate automodification in the NAD+ binding domain (55.2 kDa) and the DNA binding domain (42.5 kDa). We have analyzed the relationship between the number of sites modified on poly(ADP-ribose) polymerase and its effect on the polymerization activity and its alternate NADase activity. Automodification greatly altered both enzyme activities, decreasing both polymer synthesis and alternate NADase activity.

Poly(ADP-ribose) polymerase (PARP) and DNA-dependent protein kinase (DNA-PK) are important nuclear enzymes that cooperate to minimize genomic damage caused by DNA strand interruptions. DNA strand interruptions trigger the ADP-ribosylation activity and phosphorylation activity of PARP and DNA-PK respectively. In order to understand the relationship of PARP and DNA-PK with respect to DNA binding required for their activation, we analyzed the kinetics of the reactions and determined the apparent dissociation constants ( K d app ) of the enzymes for DNA strand interruptions. PARP has a high binding affinity for blunt ends of DNA ( K d app =116 pM) and 3′ single-base overhangs ( K d app =332 pM) in comparison to long overhangs ( K d app =2.6–5.0 nM). Nicks are good activators of PARP although the affinity of PARP for nicks ( K d app =467 pM) is 4-fold less than that for blunt ends. The K d app of DNA-PK for 3′ single-base overhangs, blunt ends and long overhangs is 704 pM, 1.3 nM and 1.4–2.2 nM respectively. These results demonstrate that (1) PARP, when compared to DNA-PK, has a greater preference for blunt ends and 3′ single-base overhangs but a weaker preference for long overhangs, and (2) nicks are effective in attracting and activating PARP. The possible implications of the preferences of PARP and DNA-PK for DNA strand interruptions in vivo are discussed.

Poly(ADP-ribose) metabolism plays an important role in numerous DNA-related functions. This homopolymer is synthesized by poly(ADP-ribose) polymerase and is degraded mainly by the poly(ADP-ribose) glycohydrolase. The activities of these two enzymes in the nucleus are closely coordinated. To better understand the interactions between these enzymes, we designed an in vitro system in which both enzymes are present at the same time. In this work, we report a model describing the synthesis and degradation of the poly(ADP-ribose) in turnover conditions. Because the half-life of the polymer in the cell is close to 1 min, we studied the very early kinetic interactions of these two enzymes.

Poly(ADP-ribose) catabolism is a complex situation involving many proteins and DNA. We have developed an in vitro turnover system where poly(ADP-ribose) metabolism is monitored in presence of different relative amounts of two principal enzymes poly(ADP-ribose) transferase and poly(ADP-ribose) glycohydrolase along with other proteins and DNA. Our current results reviewed here show that the quality of polymer, i.e. chain length and complexity, as well as preference for the nuclear substrate varies depending upon the availability of poly(ADP-ribose) glycohydrolase. These results are interpreted in the light of the recent data implicating poly(ADP-ribose) metabolism in DNA-repair.

In the past five years, poly(ADP-ribosyl)ation has developed greatly with the help of molecular biology and the improvement of biochemical techniques. In this article, we describe the physico-chemical properties of the enzymes responsible for the synthesis and degradation of poly(ADP-ribose), respectively poly(ADP-ribose) polymerase and poly(ADP-ribose) glycohydrolase. We then discuss the possible roles of this polymer in DNA repair and replication as well as in cellular differentiation and transformation. Finally, we put forward various hypotheses in order to better define the function of this polymer found only in eucaryotes. (Mol Cell Biochem122: 171–193, 1993)

Poly(ADP-ribose) polymerase is a nuclear enzyme which catalyses the hydrolysis of NAD+ to ADP-ribose and nicotinamide (1-3). Most of this ADP-ribose is covalently attached to nuclear proteins as poly(ADP-ribose) (4-5). The polymerase can carry out branching of ADP-ribose residues on average at every 20 residues (6). In this paper, we are analyzing the kinetic properties of poly(ADP-ribose) polymerase as a function of automodification and also the distribution of these on the various domains of the polymerase.

The catabolism of NAD+ in cells is carried out by poly(ADP-ribose) polymerase (PARP) which converts NAD+ into poly(ADP-ribose) in response to DNA damage. All the evidence that has accumulated up to now indicates that the degradation of poly(ADP-ribose) in vivo is carried out by poly(ADP-ribose) glycohydrolase (the glycohydrolase) and not by a phosphodiesterase. Moreover, we have recently found that inhibition of the glycohydrolase during heat shock in C3H 10T1/2 fibroblasts causes a 5-fold accumulation of polymer levels without any effect on the levels of NAD+ (1). The half-life of poly(ADP-ribose) in vivo after a carcinogenic treatment is very short, being close to 1 minute (2). Nevertheless, the decrease in NAD+ levels, which reflects the activity of the PARP, is not proportional to the accumulation of polymer suggesting that high turnover rates occur in intact cells. These high rates of synthesis and degradation of the polymer support the hypothesis of a close physical and temporal relationship between the PARP and the glycohydrolase. This hypothesis is further supported by the physical attachment of the PARP to nuclear structures (chromatin and matrix) and the presence of the glycohydrolase in the nucleus.