Paternal Obesity Induced by High-Fat Diet Impairs the Metabolic and Reproductive Health of Progeny in Rats.

Due to the increased incidence of obesity, it is of great importance to identify all the possible consequences in those who suffer from it and their descendants. This study aimed to investigate how paternal obesity, resulting from an 18-week high-fat diet (HFD), affects the metabolic and reproductive health of offspring. In the fathers (F0 generation), the HFD led to significant weight gain, primarily due to increased visceral fat. It also resulted in impaired glucose control and reduced insulin sensitivity. Furthermore, F0 males from the HFD group had reduced sperm concentration and lower sperm viability but were still able to sire litters. F1 offspring were monitored during 18 weeks; F1 offspring from obese fathers displayed increased body weight during the experimental window, especially in males, without significant metabolic disturbances. Additionally, F1 males showed reduced sperm viability, indicating potential reproductive implications. On the other hand, F1 females showed normal estrous cycle patterns but had a reduced number of primordial follicles, suggesting a decrease in their follicular reserve and reproductive potential. This study highlights that metabolic and reproductive issues may be passed down to future generations through the paternal line.

Bioremediation of copper using indigenous fungi Aspergillus species isolated from an abandoned copper mine soil.

Bioremediation of mining soils using metal tolerant fungi is widely considered as a promising cost-effective and ecofriendly approach. This study assessed the copper removal efficiency and bioaccumulation ability of the indigenous species Aspergillus hiratsukae LF1 and Aspergillus terreus LF2 isolated from the soils of an abandoned copper mine in Oman. Nutrient medium containing five different Cu (II) levels (0 - control, 100, 200, 300 and 500 mg/L) was employed for assessing both parameters. The removal efficiency from nutrient medium (100-500 mg Cu per L) ranged from 57% to 21% for A. hiratsukae LF1, and from 69% to 24% for A. terreus LF2. A. hiratsukae LF1 and A. terreus LF2 accumulated a maximum of 4.63 and 5.95 mg Cu/g,espectively, at 500 mg/L of Cu (II) concentration. The compositional analysis of extracellular polymeric substances excreted by both species revealed a hormetic response by A. hiratsukae LF1 at 100 mg/L; whereas increasing media Cu levels induced carbohydrates production in A. terreus LF2. These results hint at the involvement of carbohydrates in the Cu-tolerance mechanism of the latter. Copper accumulation in both species was further demonstrated through scanning electron microscopy and energy dispersive spectrometry. In line with the pertaining literature, our results are somewhat inconclusive concerning whether proteins or carbohydrates play a more pivotal role in copper complexation in both species; yet, FTIR analysis showed the participation of different functional groups in Cu sorption. Overall, although additional research is required to advance the knowledge about both Aspergillus species, our findings suggest that A. terreus LF2 presents greater promise for copper bioremediation due to enhanced tolerance and accumulation capacity.

Potential of Salvinia biloba Raddi for removing atrazine and carbendazim from aquatic environments.

In this exploratory study, naturally occurring Salvinia biloba Raddi specimens were assessed for atrazine and carbendazim polluted water remediation. Experiments were carried out over 21 days in glass vessels containing deionized water artificially contaminated with 0, 5, 10, and 20 mg L-1 of atrazine or carbendazim. Atrazine had a pronounced detrimental impact on S. biloba, as no biomass development was observed in all macrophytes exposed to this herbicide in the entire concentration range. However, carbendazim-treated plants were able to grow and survive in the polluted medium even when subjected to the highest concentration of this fungicide (i.e., 20 mg L-1). In addition, increased chlorosis and necrosis were also detected in plants subjected to carbendazim as a result of the high phytotoxicity caused by atrazine. A maximal removal efficiency of ~ 30% was observed for both pesticides at 5 mg L-1 and decreased with increasing concentrations of the pollutants. The spectrum of the FTIR-ATR analysis revealed the existence of various functional groups (e.g., amide, carboxyl, hydroxyl, phosphate, sulfate) on the plants, which could be related to pesticide biosorption. In addition, at the end of the 21-day assay, seven carbendazim-resistant bacteria could be isolated from the roots of fungicide-treated plants. Therefore, the use of autochthonous free-floating S. biloba macrophytes for phytoremediation of aquatic environments contaminated with carbendazim shows great promise. Still, additional research is required to further elucidate the plant-mediated carbendazim elimination process and the role of the herbicide-resistant bacteria, and seek alternative species capable of mitigating atrazine contamination.

Outcome measures and survivorship following total hip arthroplasty in adolescent population.

BACKGROUND: The most frequent indication for total hip arthroplasty (THA) in adolescent patients has been juvenile idiopathic arthritis (JIA). However, in recent years, other causes have become more prevalent. QUESTIONS/PURPOSES: (1) What is the survivorship following THA across the diagnostic spectrum in adolescent population? (2) How are quality of life results affected by systemic medical conditions? METHODS: We retrospectively reviewed all consecutive THA in patients under 21 years of age, performed at our Institution between 1993 and 2018. There were 34 prostheses implanted in 26 patients with a mean age of 18.4 years (range 11 to 21). The most frequent diagnosis was JIA (14 hips), followed by avascular necrosis (10 hips). Patient reported outcomes were assessed using Harris and Oxford Hip Scores (HHS and OHS), Visual Analogue Scale (VAS) and EuroQol-5D. Survivorship for revision and aseptic loosening was determined with Kaplan Meier analysis. RESULTS: At final follow-up, the overall survival rate was of 89.3% at 12-year follow-up. Three acetabular components underwent revision surgery for aseptic loosening. Clinical HHS significantly improved from 37.5 to 90.6 points (p < 0.001). Mean OHH was 37.4 points, with a final VAS of 1.64 points. Mean 5Q-5D was 0.704 with an interquartile range of 0.4 to 1.0. JIA patients displayed worse pre-operative HHS scores, and at final follow-up had worse HHS, OHS, VAS and EQ-5D scores compared to the rest of the patients. Complications included three intra-operative femoral fractures, one sciatic nerve palsy and one adductor contracture. CONCLUSION: THA in adolescent patients provides improved functional outcomes with acceptable revision rates at mid-term follow-up.

Deposition of trace metals associated with atmospheric particulate matter: Environmental fate and health risk assessment.

Anthropogenic and natural sources influence trace metals (TMs) bound to different sized particulate matter (PM) in dry and wet atmospheric deposition, which can create ecosystem and human health issues in the long run. Limited reviews are available summarizing worldwide concentrations in TMs in atmospheric PMs, their sources and pathways. Simultaneously, quantitative assessment of the potential human and ecosystem health risks imposed by the atmospheric particulate matter has not been adequately reviewed. Addressing this gap, here we review, the concentration of TMs in dry deposition mainly varies with the responsible sources, whereas, in wet deposition, it depends on the solubility of TMs. Other than deposition on impervious surfaces, the TMs incorporated PM can be deposited on biological agents. Health risk assessments show that ingestion and dermal contact pathways are more likely to cause health issues, however, the probability of occurring ingestion and dermal contact pathways is limited. Attention must be paid to the contribution from non-exhaust and exhaust vehicular emissions for TMs in atmospheric deposition, understanding their impact on stormwater management and urban agriculture. Behaviors of TMs in the atmosphere depends on many complex factors including origin, wind patterns, and weather conditions. Therefore, future research needs to be carried to model and predict the fate and transfer of TMs once they are generated through natural and anthropogenic sources. We believe that such research would allow establishing pollution control policies and measures in urban environments which will be critical to reduce the levels of TMs associated with atmospheric deposition in the environment.

Plant growth-promoting bacteria in phytoremediation of metal-polluted soils: Current knowledge and future directions.

Soil metal contamination is a major concern due to the ever-rising number of areas afflicted worldwide and the detrimental effects of metals to the environment and human health. Due to their non-biodegradability and toxicity, it is paramount to prevent further metal contamination and remediate the thousands of contaminated sites across the planet. Yet, conventional reclamation based on physical and chemical methods is often expensive, impractical, and triggers secondary pollution issues. Hence, microbe-aided phytoremediation has been gaining significant traction due to its environment-friendly character, cost-effectiveness, and the breakthroughs achieved during the past few decades. Microorganisms are an essential part of natural ecosystems and play a crucial role in their restoration. Indeed, plant-microbe associations in metal-polluted soils are pivotal for plants to tolerate metal toxicity and thrive in these harsh environments. Therefore, improving the understanding of this intricate relationship is invaluable for boosting phytoremediation. In this review, we focus on the potential of plant growth promoting bacteria (PGPB) for enhancing phytoremediation of metal-polluted soils. We discuss the mechanisms employed by microbes to promote plant growth and assist the removal or immobilization of metals in soil, thereby enhancing phytoextraction and phytostabilization, respectively. Microbe-mediated metal removal and detoxification through processes entailing adsorption, chelation, transformation, and precipitation, to list but a few, are also critically examined. Moreover, this work covers the direct and indirect mechanisms used by PGPB to facilitate plant acquisition of nutrients like nitrogen and phosphorus, supply and regulate phytohormones, and exert control over antagonistic microorganisms. Lastly, we provide an outlook on the future directions of microbe-aided phytoremediation and phytomining. Clearly, to fully validate and comprehend the potential of PGPB-aided phytoremediation, a considerable shift from bench-scale to field research is necessary. What's more, it is envisaged that recent advancements in genetic engineering may soon help furthering the efficiency of microbe-assisted phytoremediation.

Evaluation of Cadmium Bioaccumulation-Related Physiological Effects in Salvinia biloba: An Insight towards Its Use as Pollutant Bioindicator in Water Reservoirs.

Free-living macrophytes play an important role in the health of aquatic ecosystems. Therefore, the use of aquatic plants as metal biomonitors may be a suitable tool for the management of freshwater reservoirs. Hence, in this study, we assessed the effects of cadmium (Cd) in Salvinia biloba specimens collected from the Middle Paraná River during a 10-day experiment employing artificially contaminated water (100 µM Cd). S. biloba demonstrated a great ability for Cd bioaccumulation in both the root-like modified fronds (named "roots") and the aerial leaf-like fronds (named "leaves") of the plants. Additionally, Cd toxicity was determined by the quantification of photosynthetic pigments (chlorophylls a and b, and carotenoids), flavonoids, and soluble carbohydrate contents in S. biloba over time (1, 3, 5, 7, and 10 days). In general, deterioration was more pronounced in leaves than in roots, suggesting a greater implication of the former in long-term Cd sequestration in S. biloba. Deleterious effects in the appraised parameters were well correlated with the total amount of Cd accumulated in the leaves, and with the qualitative changes observed in the plants' phenotype during the 10-day metal exposure assay. The flavonoids and carotenoids in leaves were highly affected by low Cd levels followed by root carbohydrates. In contrast, chlorophylls and root flavonoids were the least impacted physiological parameters. Therefore, our results demonstrate that S. biloba displays dissimilar organ-linked physiological responses to counteract Cd phytotoxicity and that these responses are also time-dependent. Though further research is needed, our work suggests that easy-handled physiological data obtained from autochthonous free-floating S. biloba specimens may be used as a valuable tool for metal-polluted water biomonitoring.

Metal accumulation in roadside soils of Rio de Janeiro, Brazil: impact of traffic volume, road age, and urbanization level.

Traffic-related metal emissions have become a global concern due to their deposition in roadside soils and potential hazardous effects. This study evaluates metal levels in soils adjoining four highways of Rio de Janeiro (Linha Vermelha, Via Dutra, BR-465, and Avenida Brasil), chosen for their diverse traffic volumes, age, and urban/rural settings. In addition to soil physicochemical properties, 11 elements (Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Sr, V, and Zn) were assessed on samples collected at different distances from the road (1, 3, 5, 10, 15 m) and soil depths (0-15 and 15-30 cm). Moreover, the geoaccumulation index was also computed to infer the soil contamination extent. The results indicate that soil metal levels at each highway are highly dependent on factors like traffic volume, distance to road, other anthropogenic sources of pollution, and their rural or urban location. The highways with greater traffic volume, Linha Vermelha and Avenida Brasil (154,000 and 126,000 vehicles day-1, respectively), clearly presented the highest soil metal concentrations. Still, as stressed by the principal component analysis, traffic volume alone fails to explain the distribution of metals in soils neighboring these highways. Thus, factors like their urban setting and larger exposure to anthropogenic activities may play a pivotal role. On the other hand, soils from Via Dutra and BR-465, both on a rural backdrop, were mostly influenced by traffic as their metal levels decreased with increasing distances from the road. Comparison with reference and preventive values for Brazilian soils and the assessment of the geoaccumulation index have shown that concentrations of Pb and V have reached concerning thresholds at Linha Vermelha and Avenida Brasil.

The effects of rhenium accumulation on Indian mustard.

Rhenium (Re) is one of Earth's scarcest and more largely scattered elements, with an estimate concentration of 0.4-0.6 µg kg-1 in the upper crust. Still, considerable concentrations of bioavailable ReO4- ions are often found, particularly in copper-molybdenum mines, where their uptake by plants of these regions has been reported. Yet, the impact of Re on plants remains a question mark, as the only available knowledge derives from a limited investigation carried out over 60 years ago. The aim of this study was to evaluate the ecophysiological response of Brassica juncea, a species known to endure and accumulate various metals, to a broad range of Re concentrations. B. juncea plants were allowed to grow and on a substrate amended with KReO4 to attain soil Re levels ranging from 0 to 80 mg kg-1. Plants were collected 45 days after sowing for analysis. The results have shown that greater Re levels reduce growth, photosynthetic activity, soluble carbohydrate mobilization, and protein contents, and increase the plant's oxidative stress (anthocyanins, H2O2, lipid peroxidation) and corresponding response (ascorbic acid, superoxide dismutase activity). Nevertheless, B. juncea exhibited a remarkable ability to endure and uptake Re, featuring shoot Re concentrations that ranged from 1615 to 24,987 mg kg-1 among the 5 and 80 mg kg-1 treatments.

Genome Editing Weds CRISPR: What Is in It for Phytoremediation?

The arrival of sequence-specific endonucleases that allow genome editing has shaken the pillars of basic and applied plant biology. Clustered regularly interspaced palindromic repeats (CRISPR) is a revolutionary genome-engineering tool that enables the enhancement of targeted traits in plants. Numerous plants, including energy crops, known for their potential to tolerate, immobilize, and stabilize inorganic and organic pollutants, have already been edited using different CRISPR systems. Moreover, a large array of genes responsible for increased metal tolerance, metal uptake and hyperaccumulation have already been identified. Thus, the CRISPR-mediated genome reprogramming of plants, including its use in gene expression regulation through transcriptional repression or activation (CRISPRi and CRISPRa), could be of paramount importance for phytoremediation. The simplicity, inexpensiveness, and capabilities of this gene editing technique could soon be used to enhance plants and bacteria involved in phytotechnologies, such as phystabilization, phytoextraction, phytomining, phytovolatilization, and bio-energy generation. In this brief viewpoint piece, we posit some of the potential benefits of CRISPR for phytoremediation.

Decationized polyplexes for gene delivery.

Gene therapy has received much attention in the field of drug delivery. Synthetic, nonviral gene delivery systems have gained increasing attention as vectors for gene therapy mainly due to a favorable immunogenicity profile and ease of manufacturing as compared to viral vectors. The great majority of these formulations are based on polycationic structures, due to their ability to interact with negatively charged nucleic acids to spontaneously form nanoparticles. In recent years, several polycationic systems have demonstrated high transfection in vitro. However, progress toward clinical applications has been slow, mainly because the cationic nature of these systems leads to intolerable toxicity levels, inappropriate biodistribution and unsatisfactory efficiency in vivo, particularly after systemic administration. Decationized polyplexes are a new class of gene delivery systems that have been developed as an alternative for conventional polycation-based systems. The major innovation introduced by decationized polyplexes is that these systems are based on neutral polymers, without any detrimental effect on the physicochemical stability or encapsulation ability, due to the transient presence of cationic charge and disulfide cross-links between the polymer chains by which the nucleic acids are physically entrapped in the particles. This editorial summarizes the most important features of decationized polyplexes and discusses potential implications for the development of new safe and efficient gene delivery systems.

Targeted decationized polyplexes for siRNA delivery.

The applicability of small interfering RNA (siRNA) in future therapies depends on the availability of safe and efficient carrier systems. Ideally, siRNA delivery requires a system that is stable in the circulation but upon specific uptake into target cells can rapidly release its cargo into the cytoplasm. Previously, we evaluated a novel generation of carrier systems ("decationized" polyplexes) for DNA delivery, and it was shown that folate targeted decationized polyplexes had an excellent safety profile and showed intracellular triggered release upon cell specific uptake. Targeted decationized polyplexes consist of a core of disulfide cross-linked poly(hydroxypropyl methacrylamide) (pHPMA) stably entrapping nucleic acids and a shell of poly(ethylene glycol) (PEG) decorated with folate molecules. In the present study, the applicability of folate targeted decationized polyplexes for siRNA delivery was investigated. This required optimization of the carrier system particularly regarding the cross-linking density of the core of the polyplexes. Stable and nanosized siRNA decationized polyplexes were successfully prepared by optimizing the cross-link density of their core. Upon incubation in human plasma, a significant portion of siRNA remained entrapped in the decationized polyplexes as determined by fluorescence correlation spectroscopy (FCS). When tested in a folate receptor overexpressing cell line stably expressing luciferase, Skov3-luc, sequence specific gene silencing was observed. As expected, neither interference on the intrinsic luciferase expression nor on the cell metabolic activity (determined by XTT) was induced by the free-polymer or the siRNA polyplexes. In conclusion, targeted decationized polyplexes are safe and stable carriers that interact with the targeted cells and rapidly disassemble upon cell entry making them promising siRNA delivery systems.

Germination and early growth of Brassica juncea in copper mine tailings amended with technosol and compost.

Mine tailings represent a serious threat to the environment and human health; thus their restoration has become a major concern. In this study, the interactions between Brassica juncea and different mine soil treatments were evaluated in order to understand their effect on germination and early growth. Three soil treatments containing 25% and 50% of technosol and 30% of compost were prepared. Germination and early growth were assessed in soil and pore water extracts from the treatments. Unlike the untreated mine soil, the three treatments allowed germination and growth, achieving levels comparable to those of seedlings from the same species developed in normal conditions. The seedlings grown in 50% of technosol and 30% of compost exhibited greater germination percentages, higher growth, and more efficient mechanisms against oxidative stress, ascribed to the organic matter and nutrients content of these treatments. Considering the unequivocal ability of B. juncea for phytoremediation, the results suggest that technosol and compost may be an auspicious solution to allow the germination and early growth of this species in mine tailings.

Decationized polyplexes as stable and safe carrier systems for improved biodistribution in systemic gene therapy.

Many polycation-based gene delivery vectors show high transfection in vitro, but their cationic nature generally leads to significant toxicity and poor in vivo performance which significantly hampers their clinical applicability. Unlike conventional polycation-based systems, decationized polyplexes are based on hydrophilic and neutral polymers. They are obtained by a 3-step process: charge-driven condensation followed by disulfide crosslinking stabilization and finally polyplex decationization. They consist of a disulfide-crosslinked poly(hydroxypropyl methacrylamide) (pHPMA) core stably entrapping plasmid DNA (pDNA), surrounded by a shell of poly(ethylene glycol) (PEG). In the present paper the applicability of decationized polyplexes for systemic administration was evaluated. Cy5-labeled decationized polyplexes were evaluated for stability in plasma by fluorescence single particle tracking (fSPT), which technique showed stable size distribution for 48 h unlike its cationic counterpart. Upon the incubation of the polymers used for the formation of polyplexes with HUVEC cells, MTT assay showed excellent cytocompatibility of the neutral polymers. The safety was further demonstrated by a remarkable low teratogenicity and mortality activity of the polymers in a zebrafish assay, in great contrast with their cationic counterpart. Near infrared (NIR) dye-labeled polyplexes were evaluated for biodistribution and tumor accumulation by noninvasive optical imaging when administered systemically in tumor bearing mice. Decationized polyplexes exhibited an increased circulation time and higher tumor accumulation, when compared to their cationic precursors. Histology of tumors sections showed that decationized polyplexes induced reporter transgene expression in vivo. In conclusion, decationized polyplexes are a platform for safer polymeric vectors with improved biodistribution properties when systemically administered.

Effect of salinity on zinc uptake by Brassica juncea.

Salinity is a major worldwide problem that affects agricultural soils and limits the reclamation of contaminated sites. Despite the large number of research papers published about salt tolerance in Brassica juncea L., there are very few accounts concerning the influence of salinity on the uptake of trace metals. In this study, B. juncea plants divided through soil sets comprising 0, 900 and 1800 mg Zn kg(-1), were treated with solutions containing 0, 60 and 120 mmol L(-1) of NaCl, with the purpose of observing the effect of salt on Zn uptake, and some physiological responses throughout the 90 days experiment. Increasing concentrations of NaCl and Zn produced a decline in the ecophysiological and biochemical properties of the plants, with observable synergistic effects on parameters like shoot dry weight, leaf area, or photochemical efficiency. Nevertheless, plants treated with 60 mmol L(-1) of NaCl accumulated striking harvestable amounts of Zn per plant that largely exceed those reported for Thlaspi caerulescens. It was concluded that salinity could play an important role on the uptake of Zn by B. juncea. The potential mechanisms behind these results are discussed, as well as the implications for phytoremediation of Zn on saline and non-saline soils.

Targeted decationized polyplexes for cell specific gene delivery.

Decationized polyplexes have previously shown unique features, especially regarding their excellent cytocompatibility and very low degree of nonspecific cellular uptake. In the present study, targeted disulfide cross-linked decationized polyplexes were composed of a core of disulfide cross-linked poly(hydroxypropyl methacrylamide) (pHPMA) stably entrapping plasmid DNA (pDNA) and a shell of poly(ethylene glycol) (PEG) decorated with folate molecules. Folate was used as targeting ligand because of its high binding affinity to its receptor, which is overexpressed in many tumors. Studies using folate receptor overexpressing cell lines (HeLa and OVCAR-3) showed significantly higher cell uptake for the folate-targeted decationized polyplexes, when compared to their nontargeted counterparts. On the contrary, for a nonexpressing folate receptor cell line (A549) similar uptake was observed for both targeted and nontargeted decationized polyplexes. Transfection studies using OVCAR-3 cells showed higher transfection efficiency for folate-targeted polyplexes, because of improved cellular uptake. Simultaneously, introduction of targeting moiety on polyplexes did not affect their good cytocompatibilty. The results reported in this paper demonstrate that coupling of folate to decationized polyplexes generates a potential system for targeted gene delivery.

Poly[N-(2-aminoethyl)ethyleneimine] as a new non-viral gene delivery carrier: the effect of two protonatable nitrogens in the monomer unit on gene delivery efficiency.

PURPOSE: The aim of this study was to investigate the in vitro gene delivery efficiency of poly[N-(2-aminoethyl)ethylene-imine](PAEEI), a polymer with a linear Polyethyleneimine (LPEI) backbone and with aminoethyl side groups that has two protonatable nitrogen atoms per monomer unit instead of one as in LPEI (an established gene delivery polymer). Method. PAEEI (Mn=4.5 kDa, Mw= 10 kDa) was synthesized by ring-opening polymerization of N-(2-(1'-aziridino)ethyl)formamide followed by hydrolysis of the amide groups. The buffering capacity of the resulting polymer was determined by acid-base titration and consequently the percentage of the protonated nitrogen atoms was calculated. Polyplexes were prepared separately in buffers with different ionic strength including Hepes buffered saline (150 mM NaCl) and Hepes buffered glucose (5% glucose) and their zeta-potential, hydrodynamic diameter and colloidal stability were measured. Transfection activity (and toxicity in Hela cells) of the polyplexes were done in HeLa, CHO and HEK293T cells. Cell incubations with polyplexes were done both in the presence and absence (HeLa cells) of serum. Results. PAEEEI showed two times more buffering capacity than LPEI. PAEEI-based Polyplexes had about the same size and zeta-potential as those of LPEI, with a higher colloidal stability in saline buffer in continuous particle size measurement. Their transfection activity was slightly higher than 22-kDa LPEI polyplexes whereas their toxicity profiles were similar in cell lines studied. The PAEEI polyplexes showed gene expression activity both in the presence and absence of serum. Conclusion. Paying attention to the fact that LPEI molecules with smaller sizes than 22 kDa show less transfection efficiency than LPEI 22, the effect of smaller size of PAEEI (10 kDa) on the gene delivery efficiency was compensated by its higher buffering capacity due to carrying more protonatable nitrogen per monomeric unit comparing with LPEI (22 kDa). Having slightly higher transfection efficiency and better colloidal stability than PEI-based systems, PAEEI is an attractive candidate for future in vivo gene delivery studies.

Π-π stacking increases the stability and loading capacity of thermosensitive polymeric micelles for chemotherapeutic drugs.

Thermosensitive amphiphilic block copolymers self-assemble into micelles above their lower critical solution temperature in water, however, the micelles generally display mediocre physical stability. To stabilize such micelles and increase their loading capacity for chemotherapeutic drugs, block copolymers with novel aromatic monomers were synthesized by free radical polymerization of N-(2-benzoyloxypropyl methacrylamide (HPMAm-Bz) or the corresponding naphthoyl analogue (HPMAm-Nt), with N-(2-hydroxypropyl) methacrylamide monolactate, using a polyethylene glycol based macroinitiator. The critical micelle temperatures and critical micelle concentrations decreased with increasing the HPMAm-Bz/Nt content. The micelles of 30-50 nm were prepared by heating the polymer aqueous solutions from 0 to 50 °C and were colloidally stable for at least 48 h at pH 7.4 and 37 °C. Paclitaxel and docetaxel encapsulation was performed by mixing drug solutions in ethanol with polymer aqueous solutions and heating from 0 to 50 °C. The micelles had a drug loading capacity up to 34 wt % for docetaxel, which is among the highest loadings reported for polymeric micelles, with loaded micelle sizes ranging from 60 to 80 nm. The micelles without aromatic groups almost completely released loaded paclitaxel in 10 days, whereas the HPMAm-Bz/Nt containing micelles released 50% of the paclitaxel at the same time, which showed a better retention for the drug of the latter micelles. (1)H solid-state NMR spectroscopy data are compatible with π-π stacking between aromatic groups. The empty micelles demonstrated good cytocompatibility, and paclitaxel-loaded micelles showed high cytotoxicity to tumor cells. In conclusion, the π-π stacking effect introduced by aromatic groups increases the stability and loading capacity of polymeric micelles.

Decationized crosslinked polyplexes for redox-triggered gene delivery.

The clinical applicability of polymers as gene delivery systems depends not only on their efficiency, but also on their safety. The cytotoxicity of these systems remains a major issue, mainly due to their cationic nature. Therefore, it is highly preferable to have a system based on biocompatible neutral polymers, lacking polycations, without compromising the DNA condensing and protecting capacities. Here, we introduce a concept to obtain a neutral polymeric gene delivery system, through a 3-step process (charge-driven condensation; stabilization through disulfide crosslinking; polyplex decationization) to generate polyplexes with a core of disulfide crosslinked poly(hydroxypropyl methacrylamide) (pHPMA) in which plasmid DNA (pDNA) is entrapped and a shell of poly(ethylene glycol) (PEG). The resulting polyplexes combine beneficial features of high and stable DNA loading capacity, stealth behavior and reduced toxicity. The nanoparticles are designed to release the pDNA after cellular uptake through cleavage of disulfide crosslinks within the intracellular reducing environment. This was shown by forced introduction of the polyplexes into the cytosol of HeLa cells by electroporation, which resulted in a high level of expression of the reporter gene. Additionally, the decationized polyplexes showed no interference on the cellular cell viability or metabolic activity (even at high dose) and no complex-induced membrane destabilization. Furthermore, decationized polyplexes showed a low degree of non-specific uptake, which is a highly favorable property for targeted therapy. Summarizing, the stabilized, decationized polyplexes presented here contribute to solve the high toxicity, low stability and lack of cellular/tissue specificity of cationic polymer based gene delivery systems.

Complete molecular genome analyses of equine rotavirus A strains from different continents reveal several novel genotypes and a largely conserved genotype constellation.

In this study, the complete genome sequences of seven equine group A rotavirus (RVA) strains (RVA/Horse-tc/GBR/L338/1991/G13P[18], RVA/Horse-wt/IRL/03V04954/2003/G3P[12] and RVA/Horse-wt/IRL/04V2024/2004/G14P[12] from Europe; RVA/Horse-wt/ARG/E30/1993/G3P[12], RVA/Horse-wt/ARG/E403/2006/G14P[12] and RVA/Horse-wt/ARG/E4040/2008/G14P[12] from Argentina; and RVA/Horse-wt/ZAF/EqRV-SA1/2006/G14P[12] from South Africa) were determined. Multiple novel genotypes were identified and genotype numbers were assigned by the Rotavirus Classification Working Group: R9 (VP1), C9 (VP2), N9 (NSP2), T12 (NSP3), E14 (NSP4), and H7 and H11 (NSP5). The genotype constellation of L338 was unique: G13-P[18]-I6-R9-C9-M6-A6-N9-T12-E14-H11. The six remaining equine RVA strains showed a largely conserved genotype constellation: G3/G14-P[12]-I2/I6-R2-C2-M3-A10-N2-T3-E2/E12-H7, which is highly divergent from other known non-equine RVA genotype constellations. Phylogenetic analyses revealed that the sequences of these equine RVA strains are related distantly to non-equine RVA strains, and that at least three lineages exist within equine RVA strains. A small number of reassortment events were observed. Interestingly, the three RVA strains from Argentina possessed the E12 genotype, whereas the three RVA strains from Ireland and South Africa possessed the E2 genotype. The unusual E12 genotype has until now only been described in Argentina among RVA strains collected from guanaco, cattle and horses, suggesting geographical isolation of this NSP4 genotype. This conserved genetic configuration of equine RVA strains could be useful for future vaccine development or improvement of currently used equine RVA vaccines.