Relevance of humanized three-dimensional tumor tissue models: a descriptive systematic literature review.

Despite numerous advances in tumor screening, diagnosis, and treatment, to date, tumors remain one of the leading causes of death, principally due to metastasis and the physiological damage produced by tumor growth. Among the main limits related to the study of tumor physiology there is the complex and heterogeneity nature of its environment and the absence of relevant, simple and inexpensive models able to mimic the biological processes occurring in patients allowing the correct clinical translation of results. To enhance the understanding of the mechanisms of tumors and to develop and evaluate new therapeutic approaches the set-up of advanced and alternative models is mandatory. One of the more translational approaches seems to be the use of humanized three-dimensional (3D) tissue culture. This model allows to accurately mimic tumor morphology and biology, maintaining the native microenvironment without any manipulation. However, little is still known on the real clinical relevance of these models for the study of tumor mechanisms and for the screening of new therapy. The aim of this descriptive systematic literature review was to evaluate and summarize the current knowledge on human 3D tumor tissue culture models. We reviewed the strategies employed by researchers to set-up these systems, also considering the different approaches and culture conditions used. All these aspects greatly contribute to the existing knowledge on tumors, providing a specific link to clinical scenarios and making the humanized 3D tumor tissue models a more attractive tool both for researchers and clinicians.

Helminth infections in Italian donkeys: Strongylus vulgaris more common than Dictyocaulus arnfieldi.

Donkeys have been used as working animals for transport and farm activities worldwide. Recently, in European countries, there has been an increasing interest in donkeys due to their use as pets, onotherapy or milk production. During 2014-2016, a countrywide survey was conducted to determine prevalence and risk factors of principal helminth infections in 1775 donkeys in 77 Italian farms. A questionnaire on management and parasite control practices was filled out for each farm. Faecal samples were examined using a modified McMaster technique, a centrifugation/flotation method and a sedimentation technique. Pooled coprocultures were performed for differentiation of strongylid eggs. Strongyles were the most common parasites detected (84.9%), followed by Dictyocaulus arnfieldi (6.9%), Oxyuris equi (5.8%), Parascaris spp. (3.6%), Anoplocephala spp. (1.0%), Strongyloides westeri (0.3%). Coprocultures revealed an omnipresence of cyathostomins (100%), followed by Strongylus vulgaris (31.0%), Poteriostomum spp. (25.0%), Triodontophorus spp. (9.0%), Strongylus edentatus (7.0%), Strongylus equinus (5.0%). Logistic regression analysis identified breed, co-pasture with horses, living area, herd size and number of treatments as significantly associated with strongyles. Sex, age, living area and herd size were significantly associated with Parascaris spp. Dictyocaulus arnfieldi was significantly associated with sex, grass, co-pasture with horses, living area and herd size. Strongylus vulgaris was significantly associated with living area and herd size. The mean number of anthelmintic treatments/year was 1.4; most of the donkeys (71.8%) were dewormed using an ivermectin drug. It is important to design parasite programs to specifically address both D. arnfieldi and S. vulgaris in donkeys, and this is especially important if donkeys co-graze with horses.

Gene therapy for chondral and osteochondral regeneration: is the future now?

Gene therapy might represent a promising strategy for chondral and osteochondral defects repair by balancing the management of temporary joint mechanical incompetence with altered metabolic and inflammatory homeostasis. This review analysed preclinical and clinical studies on gene therapy for the repair of articular cartilage defects performed over the last 10 years, focussing on expression vectors (non-viral and viral), type of genes delivered and gene therapy procedures (direct or indirect). Plasmids (non-viral expression vectors) and adenovirus (viral vectors) were the most employed vectors in preclinical studies. Genes delivered encoded mainly for growth factors, followed by transcription factors, anti-inflammatory cytokines and, less frequently, by cell signalling proteins, matrix proteins and receptors. Direct injection of the expression vector was used less than indirect injection of cells, with or without scaffolds, transduced with genes of interest and then implanted into the lesion site. Clinical trials (phases I, II or III) on safety, biological activity, efficacy, toxicity or bio-distribution employed adenovirus viral vectors to deliver growth factors or anti-inflammatory cytokines, for the treatment of osteoarthritis or degenerative arthritis, and tumour necrosis factor receptor or interferon for the treatment of inflammatory arthritis.

Seroepidemiological and biomolecular survey on Toxoplasma gondii infection on organic pig farms.

Pigs are an important reservoir of Toxoplasma gondii, and pork meat is considered one of the main sources of human infection. The present survey assesses the prevalence of T. gondii in organically reared pigs in Sardinia found on the island using various diagnostic methods. Firstly, a seroepidemiological survey was carried out on 63 farms, examining individual blood samples from 414 adult pigs. Secondly, a biomolecular investigation carried out on another 62 farms, examining the brain samples from 107 adult pigs. The seroprevalence detected for T. gondii at the farm level was 85.7% (54 out of the 63 pig farms examined). However, the prevalence rate in the investigated pigs was 51.7% (214 positives out of 414 examined). A total of 54.8% (34/62) of the monitored pig farms were found to be positive by PCR. The presence of T. gondii DNA in individual brain samples was detected in 47.7% (51/107) of the examined animals. The high prevalence of T. gondii found is alarming and suggests that pigs reared on organic farms should be considered as a significant source of infection for humans in the areas investigated.

Chondroprotective activity of N-acetyl phenylalanine glucosamine derivative on knee joint structure and inflammation in a murine model of osteoarthritis.

OBJECTIVE: Osteoarthritis (OA), the most common chronic degenerative joint disease, is characterized by joint structure changes and inflammation, both mediated by the IκB kinase (IKK) signalosome complex. The ability of N-acetyl phenylalanine derivative (NAPA) to increase cartilage matrix components and to reduce inflammatory cytokines, inhibiting IKKα kinase activity, has been observed in vitro. The present study aims to further clarify the effect of NAPA in counteracting OA progression, in an in vivo mouse model after destabilization of the medial meniscus (DMM). DESIGN: 26 mice were divided into three groups: (1) DMM surgery without treatment; (2) DMM surgery treated after 2 weeks with one intra-articular injection of NAPA (2.5 mM) and (3) no DMM surgery. At the end of experimental times, both knee joints of the animals were analyzed through histology, histomorphometry, immunohistochemistry and microhardness of subchondral bone (SB) tests. RESULTS: The injection of NAPA significantly improved cartilage thickness (CT) and reduced Chambers and Mankin modified scores and fibrillation index (FI), with weaker MMP13, ADAMTS5, MMP10 and IKKα staining. The microhardness measurements did not shown statistically significant differences between the different groups. CONCLUSIONS: NAPA markedly improved the physical structure of articular cartilage while reducing catabolic enzymes, extracellular matrix (ECM) remodeling and IKKα expression, showing to be able to exert a chondroprotective activity in vivo.

The recombinant protein rSP03B is a valid antigen for screening dog exposure to Phlebotomus perniciosus across foci of canine leishmaniasis.

The frequency of sandfly-host contacts can be measured by host antibody levels against sandfly salivary proteins. Recombinant salivary proteins are suggested to represent a valid replacement for salivary gland homogenate (SGH); however, it is necessary to prove that such antigens are recognized by antibodies against various populations of the same species. Phlebotomus perniciosus (Diptera: Psychodidae) is the main vector of Leishmania infantum (Trypanosomatida: Trypanosomatidae) in southwest Europe and is widespread from Portugal to Italy. In this study, sera were sampled from naturally exposed dogs from distant regions, including Campania (southern Italy), Umbria (central Italy) and the metropolitan Lisbon region (Portugal), where P. perniciosus is the unique or principal vector species. Sera were screened for anti-P. perniciosus antibodies using SGH and 43-kDa yellow-related recombinant protein (rSP03B). A robust correlation between antibodies recognizing SGH and rSP03B was detected in all regions, suggesting substantial antigenic cross-reactivity among different P. perniciosus populations. No significant differences in this relationship were detected between regions. Moreover, rSP03B and the native yellow-related protein were shown to share similar antigenic epitopes, as canine immunoglobulin G (IgG) binding to the native protein was inhibited by pre-incubation with the recombinant form. These findings suggest that rSP03B should be regarded as a universal marker of sandfly exposure throughout the geographical distribution of P. perniciosus.

Toxoplasma gondii and Neospora caninum infection in epileptic dogs.

OBJECTIVES: Studies in humans have demonstrated the role of Toxoplasma gondii, a protozoan parasite, in epileptic seizures. This study aimed to investigate the serological correlation between T. gondii and N. caninum and epilepsy in dogs. MATERIALS AND METHODS: The medical record database of the Veterinary Teaching Hospital, University of Perugia, was searched for dogs serologically tested by IFAT for T. gondii and N. caninum and following specific inclusion criteria. Dogs were stratified by having a clinical diagnosis of epilepsy or suffering different conditions. RESULTS: One-hundred and twenty-eight dogs were included, 64 with epilepsy and 64 without clinical signs of epilepsy. Seventeen of the 64 epileptic dogs (26.6%; 95% CI: 15.7% to 37.4%) and twenty-one of the 64 non-epileptic dogs (32.8%; 95% CI: 21.3% to 44.3%) tested positive for T. gondii. Eight of the epileptic dogs (12.5%; 95% CI: 4.4% to 20.6%) and three of the non-epileptic dogs (4.7%; 95% CI: 0% to 9.9%) tested positive for N. caninum. There was no statistically significant difference in the rate of T. gondii or N. caninum seroreactivity between epileptic and non-epileptic dogs. CLINICAL SIGNIFICANCE: The results obtained do not seem to support the role of T. gondii and N. caninum as causative agents of dog epilepsy.

Biosynthetic scaffolds for partial meniscal loss: A systematic review from animal models to clinical practice.

Acute or degenerative meniscus tears are the most common knee lesions. Meniscectomy provides symptomatic relief and functional recovery only in the short- to mid-term follow-up but significantly increases the risk of osteoarthritis. For this reason, preserving the meniscus is key, although it remains a challenge. Allograft transplants present many disadvantages, so during the last 20 years preclinical and clinical research focused on developing and investigating meniscal scaffolds. The aim of this systematic review was to collect and evaluate all the available evidence on biosynthetic scaffolds for meniscus regeneration both in vivo and in clinical studies. Three databases were searched: 46 in vivo preclinical studies and 30 clinical ones were found. Sixteen natural, 15 synthetic, and 15 hybrid scaffolds were studied in vivo. Among them, only 2 were translated into clinic: the Collagen Meniscus Implant, used in 11 studies, and the polyurethane-based scaffold Actifit®, applied in 19 studies. Although positive outcomes were described in the short- to mid-term, the number of concurrent procedures and the lack of randomized trials are the major limitations of the available clinical literature. Few in vivo studies also combined the use of cells or growth factors, but these augmentation strategies have not been applied in the clinical practice yet. Current solutions offer a significant but incomplete clinical improvement, and the regeneration potential is still unsatisfactory. Building upon the overall positive results of these "old" technologies to address partial meniscal loss, further innovation is urgently needed in this field to provide patients better joint sparing treatment options.

Clinical, serological and molecular investigations of EHV-1 and EHV-4 in 15 unweaned thoroughbred foals.

Fifteen unweaned thoroughbred foals, born on a stud farm to vaccinated mares, were clinically monitored during their first six months of life and repeatedly tested for equine herpesvirus type 1 (EHV-1) and equine herpesvirus type 4 (EHV-4). Nasopharyngeal swabs and blood samples were collected and screened respectively by PCR and seroneutralisation to detect the presence of the virus, explore its role as a possible cause of respiratory disease, and to assess the efficiency of the pcr for the diagnosis of this disease. The foals were divided into three groups on the basis of their clinical signs and whether they had seroconverted to EHV-1 and/or EHV-4: first, foals with no clinical signs of disease that had not seroconverted; secondly, foals with clinical signs that had seroconverted, and thirdly, foals with clinical signs that had not seroconverted. The results indicated that the viruses circulated on the stud farm despite stringent vaccination regimens against them, and confirmed their association with respiratory disease. The absence of significantly different pcr results among the three groups of foals showed that the pcr was effective in confirming the circulation of the viruses on the premises without being particularly helpful as a diagnostic tool.

Pulsed electromagnetic fields and platelet rich plasma alone and combined for the treatment of wear-mediated periprosthetic osteolysis: An in vivo study.

Wear-mediated osteolysis is a common complication occurring around implanted prosthesis, which ultimately leads to bone loss with mechanical instability and the need for surgical revision. At the moment, revision surgery is the only effective treatment. The aim of this study was to assess the efficacy of pulsed electromagnetic fields (PEMFs) and platelet rich plasma (PRP), alone and in association, in a clinically relevant in vivo model of periprosthetic osteolysis. Titanium alloy pins were implanted intramedullary in distal femurs of male inbred rats and, after osseointegration, polyethylene particles were injected intra-articularly to induce osteolysis. Animals were divided in four groups of treatment: PEMFs, PRP, PEMFs + PRP and no treatment. Microtomography was performed during the course of experiments to monitor bone stock and microarchitecture. Histology, histomorphometry, immunohistochemistry and biomechanics were evaluated after treatments. Biophysical and biological stimulations significantly enhanced bone to implant contact, bone volume and bone microhardness and reduced fibrous capsule formation and the number of osteoclasts around implants. Among treatments, PEMFs alone and in association with PRP exerted better results than PRP alone. Present data suggest that biophysical stimulation, with or without the enrichment with platelet derived growth factors, might be a safe, mini-invasive and conservative therapy for counteracting osteolysis and prompting bone formation around implants. STATEMENT OF SIGNIFICANCE: Pulsed electromagnetic fields (PEMFs) and platelet rich plasma (PRP) show anabolic and anti-inflammatory effects and they are already been used in clinical practice, but separately. To date, there are no preclinical in vivo studies evaluating their combined efficacy in periprosthetic osteolysis, in bone tissue microarchitecture and in biomechanics. The aim of the present study was to evaluate the effects of PEMFs and PRP in vivo, when administered individually and in combination in the treatment of periprosthetic wear mediated ostelysis, and in restoring the osteogenetic properties of perimplant bone tissue and its biomechanical competence. The combination of PEMFs and PRP could be employed for counteracting the ostelysis process in a conservative and non surgical manner.

Evaluation of alterations on mitral annulus velocities, strain, and strain rates due to abrupt changes in preload elicited by parabolic flight.

We tested the hypothesis that in normal subjects, cardiac tissue velocities, strain, and strain rates (SR), measured by Doppler tissue echocardiography (DTE), are preload dependent. To accomplish it, immediately preceding image acquisition, reversible, repeatable, acute nonpharmacological changes in preload were induced by parabolic flight. DTE has been proposed as a new approach to assess left ventricular regional myocardial function by computing tissue velocities, strain, and SR. However, preload dependence of these parameters in normal subjects still remains controversial. DTE images (Philips) were obtained in 10 normal subjects in standing upright position at normogravity (1 Gz), hypergravity (1.8 Gz), and microgravity (0 Gz) with and without -50 mmHg lower body negative pressure (LBNP). Myocardial velocity curves in the basal interventricular septum were reconstituted offline from DTE images, from which peak systolic (S'), early (E') and late (A') diastolic velocities, SR, and peak systolic strain (PSepsilon) were measured and averaged over four beats. At 1.8 Gz (reduced venous return), S', E', and A' decreased by 21%, 21%, and 26%, respectively, compared with 1-Gz values, while at 0 Gz (augmented venous return), E', A', and PSepsilon increased by 57%, 53%, and 49%, respectively. LBNP reduced E' and PSepsilon. In conclusion, our results were in agreement with those obtained in animal models, in which preload was changed in a controlled, acute, and reversible manner, and image acquisition was performed immediately following preload modifications. The hypothesis of preload dependence was confirmed for S', E', A', and PSepsilon, while SR appeared to be preload independent, probably reflecting intrinsic myocardial properties.

Fresh osteochondral allotransplants: Outcomes, failures and future developments.

Osteochondral allografts are used to treat many different conditions as acute traumatic large-sized lesions, degenerative osteoarthritis, osteochondritis dissecans, avascular necrosis or in case of failure of previous procedures particularly in young patients for whom primary prosthesis is not desirable. Fresh allografts present the advantage of having mature viable hyaline cartilage, not causing donor morbidity, allowing the restoration of even large defects in a single surgical session. Conversely, they could account for risks of disease transmission, immunologic reactions, and for limited availability. The present review aimed to analyze published studies of the last decade in which patients received fresh osteochondral allografts by dividing them for knee or ankle regenerative purposes. We wish to report the observed failure rates and particularly to collect any other reported side effect or outcome for identifying major problems and limits linked to the procedure and for delineating possible future researches and approaches. The overall success rates resulted ranging from 5.3% to 48.3% in the ankle at a mean follow up of 3.3 years and from 0% to 85.7% in the knee at a mean follow up of 7.1 years. Among other outcomes, occurrence or progression of arthritis, osteolysis, graft instability, fractures, nonunions, edema and infections were recorded. Overall, the lack of well designed randomized and controlled clinical trials, of immunological determination of the anti-donor antibodies development and of local and systemic biomarkers to detect reaction to the graft seems to be the major drawback. Improvements in these limiting factors might be desirable in order to enhance the clinical scenario of a well-established and successful procedure to give, especially for young patients, a real regeneration of the joint.

Correlation between coronal and apical arch form and degree of divergence: a volumetric computed tomography analysis.

PURPOSE: To determine the existence of any correlation between the degree of divergence and apical and coronal arch form using cone-beam computed tomography (CBCT). MATERIALS AND METHODS: A total of 176 (88 coronal and 88 apical) CBCT scans pertaining to a sample of 44 subjects (16 males and 28 females), of which 26 were Class I, 14 Class II and 4 Class III, were analysed. A lateral projection of the skull was obtained from each VCT and cephalometric tracing was performed (according to Ricketts) so as to divide the sample into subgroups based on the degree of divergence (11 short-faced, 18 norm-faced and 15 long-faced subjects). Dahlberg's index values were calculated and Student's t-test for paired data was applied. RESULTS: On the whole, the hyperbolic cosine curve was found to be the most representative of the arch forms considered. CONCLUSIONS: A correlation between degree of divergence and the arch form of the apical and, especially, coronal portions of both the upper and lower jaws was revealed.

Tracking of left ventricular long axis from real-time three-dimensional echocardiography using optical flow techniques.

Two-dimensional echocardiography (2DE) is routinely used in clinical practice to measure left ventricular (LV) mass, dimensions, and function. The reliability of these measurements is highly dependent on the ability to obtain nonforeshortened long axis (LA) images of the left ventricle from transthoracic apical acoustic windows. Real time three-dimensional echocardiography (RT3DE) is a novel imaging technique that allows the acquisition of dynamic pyramidal data structures encompassing the entire ventricle and could potentially overcome the effects of LA foreshortening. Accordingly, the aim of this paper was to develop a nearly automated method based on optical flow techniques for the measurement of the left ventricular (LV) LA throughout the cardiac cycle from RT3DE data. The LV LA measurements obtained with the automated technique has been compared with LA measurements derived from manual selection of the LA from a volumetric display of RT3DE data. High correlation (r = .99, SEE = 1.8%, y = .94x + 5.3), no significant bias (-0.18 mm), and narrow limits of agreement (SD: 1.91 mm) were found. The comparison between the LA length derived from 2DE and RT3DE data showed significant underestimation of the 2DE based measurements. In conclusion, this study proves that RT3DE data overcome the effects of foreshortening and indicates that the method we propose allows fast and accurate quantification of LA length throughout the cardiac cycle.

Cross-sectional survey on Tritrichomonas foetus infection in Italian cats.

The feline genotype of Tritrichomonas foetus is a widespread cause of large-bowel diarrhoea in cats. The aim of this study was to determine the prevalence of the T. foetus infection in cat populations across Italy. Fresh, individual faecal samples were collected from 267 cats, kept in different environments (i.e., private households, breeding structures, municipal catteries and colonies) in three different sites across Italy. The faecal samples were tested by PCR to detect T. foetus. Moreover, the same samples were subjected to a concentration-flotation technique and a commercial direct fluorescent-antibody (DFA) test to detect additional enteric parasites, including Giardia duodenalis. The overall prevalence of T. foetus infection was 5.2%. All the infected cats showed diarrhoea at the time of sampling: 9 out of 14 positive cats were co-infected with G. duodenalis, 1 with Toxocara cati and 3 with Dipylidium caninum. The risk factor analysis showed that not only the breed, but also co-infections with G. duodenalis and Dipylidium caninum were significantly associated with the presence of T. foetus. This study confirms the presence of T. foetus in cats living in Italy, suggesting that this protozoan parasite should always be included in the differential diagnosis of patients referred with large-bowel disease symptoms, especially if they were purebred animals, or affected by other enteric protozoa, such as G. duodenalis.

Needle-like ion-doped hydroxyapatite crystals influence osteogenic properties of PCL composite scaffolds.

Surface topography and chemistry both play a crucial role on influencing cell response in 3D porous scaffolds in terms of osteogenesis. Inorganic materials with peculiar morphology and chemical functionalities may be proficiently used to improve scaffold properties-in the bulk and along pore surface-promoting in vitro and in vivo osseous tissue in-growth. The present study is aimed at investigating how bone regenerative properties of composite scaffolds made of poly(Ɛ-caprolactone) (PCL) can be augmented by the peculiar properties of Mg(2+) ion doped hydroxyapatite (dHA) crystals, mainly emphasizing the role of crystal shape on cell activities mediated by microstructural properties. At the first stage, the study of mechanical response by crossing experimental compression tests and theoretical simulation via empirical models, allow recognizing a significant contribution of dHA shape factor on scaffold elastic moduli variation as a function of the relative volume fraction. Secondly, the peculiar needle-like shape of dHA crystals also influences microscopic (i.e. crystallinity, adhesion forces) and macroscopic (i.e. roughness) properties with relevant effects on biological response of the composite scaffold: differential scanning calorimetry (DSC) analyses clearly indicate a reduction of crystallization heat-from 66.75 to 43.05 J g(-1)-while atomic force microscopy (AFM) ones show a significant increase of roughness-from (78.15 ± 32.71) to (136.13 ± 63.21) nm-and of pull-off forces-from 33.7% to 48.7%. Accordingly, experimental studies with MG-63 osteoblast-like cells show a more efficient in vitro secretion of alkaline phosphatase (ALP) and collagen I and a more copious in vivo formation of new bone trabeculae, thus suggesting a relevant role of dHA to support the main mechanisms involved in bone regeneration.

Landscape scale estimation of soil carbon stock using 3D modelling.

Soil C is the largest pool of carbon in the terrestrial biosphere, and yet the processes of C accumulation, transformation and loss are poorly accounted for. This, in part, is due to the fact that soil C is not uniformly distributed through the soil depth profile and most current landscape level predictions of C do not adequately account the vertical distribution of soil C. In this study, we apply a method based on simple soil specific depth functions to map the soil C stock in three-dimensions at landscape scale. We used soil C and bulk density data from the Soil Survey for England and Wales to map an area in the West Midlands region of approximately 13,948 km(2). We applied a method which describes the variation through the soil profile and interpolates this across the landscape using well established soil drivers such as relief, land cover and geology. The results indicate that this mapping method can effectively reproduce the observed variation in the soil profiles samples. The mapping results were validated using cross validation and an independent validation. The cross-validation resulted in an R(2) of 36% for soil C and 44% for BULKD. These results are generally in line with previous validated studies. In addition, an independent validation was undertaken, comparing the predictions against the National Soil Inventory (NSI) dataset. The majority of the residuals of this validation are between ± 5% of soil C. This indicates high level of accuracy in replicating topsoil values. In addition, the results were compared to a previous study estimating the carbon stock of the UK. We discuss the implications of our results within the context of soil C loss factors such as erosion and the impact on regional C process models.

Fully automated assessment of left ventricular volumes and mass from cardiac magnetic resonance images.

Quantification of left ventricular (LV) size and function from cardiac magnetic resonance (CMR) images requires manual tracing of LV borders on multiple 2D slices, which is subjective, tedious and time-consuming experience. This paper presents a fully automated method for endocardial and epicardial boundaries detection for the assessment of LV volumes, ejection fraction (EF) and mass from CMR images. The segmentation procedure is based on a combined level set approach initialized by an automatically detected point inside the LV cavity. To validate the proposed technique, myocardial boundaries were manually traced on end-diastolic (ED) and end-systolic (ES) frames by an experienced cardiologist. Bland-Altman analysis and linear regression were used to validate LV volumes, EF and mass and similarity metrics were applied to assess the agreement between manually and automatically detected contours. We found minimal biases and narrow limits of agreement for LV volumes, EF and mass; Dice coefficient, Jaccard index and Hausdorff distance evaluated for 2D ED and ES endocardial and epicardial boundaries showed adequate overlapping. The proposed technique allows fast and accurate assessment of LV volumes, EF and mass as a basis for accurate quantification of LV size and function, and myocardial scar from CMR images.

Expression of an evolved engineered variant of a bacterial glycine oxidase leads to glyphosate resistance in alfalfa.

The main strategy for resistance to the herbicide glyphosate in plants is the overexpression of an herbicide insensitive, bacterial 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). A glyphosate resistance strategy based on the ability to degrade the herbicide can be useful to reduce glyphosate phytotoxicity to the crops. Here we present the characterization of glyphosate resistance in transgenic alfalfa (Medicago sativa L.) expressing a plant-optimized variant of glycine oxidase (GO) from Bacillus subtilis, evolved in vitro by a protein engineering approach to efficiently degrade glyphosate. Two constructs were used, one with (GO(TP+)) and one without (GO(TP-)) the pea rbcS plastid transit peptide. Molecular and biochemical analyses confirmed the stable integration of the transgene and the correct localization of the plastid-imported GO protein. Transgenic alfalfa plants were tested for glyphosate resistance both in vitro and in vivo. Two GO(TP+) lines showed moderate resistance to the herbicide in both conditions. Optimization of expression of this GO variant may allow to attain sufficient field resistance to glyphosate herbicides, thus providing a resistance strategy based on herbicide degradation.

In vivo effect of two different pulsed electromagnetic field frequencies on osteoarthritis.

Osteoarthritis (OA) is a joint pathology characterized by fibrillation, reduced cartilage thickness and subchondral bone sclerosis. There is evidence that pulsed electromagnetic fields (PEMFs) counteract OA progression, but the effect of two different PEMF frequencies has not yet been shown. The aim of this study was to test the effectiveness of PEMFs at two different frequencies (37 and 75 Hz) in a late OA stage in 21-month-old Guinea pigs. After 3 months of 6 h/day PEMF stimulation, histological and histomorphometric analyses of the knees were performed. At both frequencies, PEMFs significantly reduced histological cartilage score, fibrillation index (FI), subchondral bone thickness (SBT) and trabecular number (Tb.N) and increased trabecular thickness (Tb.Th) and separation (Tb.Sp) in comparison to the not treated SHAM group. However, PEMFs at 75 Hz produced significantly more beneficial effects on the histological score and FI than 37 Hz PEMFs. At 75 Hz, PEMFs counteracted cartilage thinning as demonstrated by a significantly higher cartilage thickness values than either those of the SHAM or 37 Hz PEMF-treated groups. Although in severe OA both PEMF frequencies were able to limit its progression, 75 Hz PEMF stimulation achieved the better results.