Magnetic Nanoemulsions for the Intra-Articular Delivery of Ascorbic Acid and Dexamethasone.

(1) Osteoarthritis (OA) is a progressive joint degenerative disease that currently has no cure. Limitations in the development of innovative disease modifying therapies are related to the complexity of the underlying pathogenic mechanisms. In addition, there is the unmet need for efficient drug delivery methods. Magnetic nanoparticles (MNPs) have been proposed as an efficient modality for the delivery of bioactive molecules within OA joints, limiting the side effects associated with systemic delivery. We previously demonstrated MNP's role in increasing cell proliferation and chondrogenesis. In the design of intra-articular therapies for OA, the combined NE-MNP delivery system could provide increased stability and biological effect. (2) Proprietary Fe3O4 MNPs formulated as oil-in-water (O/W) magneto nanoemulsions (MNEs) containing ascorbic acid and dexamethasone were tested for size, stability, magnetic properties, and in vitro biocompatibility with human primary adipose mesenchymal cells (ADSC), cell mobility, and chondrogenesis. In vivo biocompatibility was tested after systemic administration in mice. (3) We report high MNE colloidal stability, magnetic properties, and excellent in vitro and in vivo biocompatibility. By increasing ADSC migration potential and chondrogenesis, MNE carrying dexamethasone and ascorbic acid could reduce OA symptoms while protecting the cartilage layer.

Unusual Canine Distemper Virus Infection in Captive Raccoons (Procyon lotor).

Canine morbillivirus, also known as canine distemper virus (CDV), is the causative agent of canine distemper (CD), which is a serious contagious disease of canines, large felids, and, occasionally, raccoons. This study included seven raccoons from the Timisoara Zoological Garden, Romania. CDV was detected using RT-qPCR on blood samples, but several other exams were also performed-clinical, bacteriological, immunohistochemistry (IHC) and histopathology, toxicological screening, and necropsy-which confirmed CDV infection. Severe digestive disorders (diarrhea and frequent hematemesis) were observed. The necropsy findings included pseudo membranous gastroenteritis, congestion, and pulmonary edema in two raccoons. Immunohistochemistry showed immunolabeled CDV antigenantibodies on the viral nucleocapsid. Histopathology revealed lymphocyte depletion in mesenteric lymphnodes and intranuclear and intracytoplasmic inclusions in the enterocytes of the small intestine. Based on the RT-qPCR assay, laboratory tests, and the lesions observed, it was established that the raccoons were infected with CDV, which was the cause of death in two cases. The results from the necropsy, histology, and immunohistochemistry in the raccoons are comparable with reported CDV lesions in dogs. In conclusion, several exams may be performed to establish the etiology of possible interspecific viral infection, but only very specific exams can identify aCDV infection. Laboratory analyses must be completed by RT-qPCR assay or IHC to establish infection with uncommon viruses in raccoons with high accuracy.

3D high-precision melt electro written polycaprolactone modified with yeast derived peptides for wound healing.

In this study melt electro written (MEW) scaffolds of poly(ε-caprolactone) PCL are decorated with anti-inflammatory yeast-derived peptide for skin wound healing. Initially, 13 different yeast-derived peptides were screened and analyzed using both in vitro and in vivo assays. The MEW scaffolds are functionalized with the selected peptide VLSTSFPPW (VW-9) with the highest activity in reducing pro-inflammatory cytokines and stimulating fibroblast proliferation, migration, and collagen production. The peptide was conjugated to the MEW scaffolds using carbodiimide (CDI) and thiol chemistry, with and without plasma treatment, as well as by directly mixing the peptide with the polymer before printing. The MEW scaffolds modified using CDI and thiol chemistry with plasma treatment showed improved fibroblast and macrophage penetration and adhesion, as well as increased cell proliferation and superior anti-inflammatory properties, compared to the other groups. When applied to full-thickness excisional wounds in rats, the peptide-modified MEW scaffold significantly enhanced the healing process compared to controls (p < 0.05). This study provides proof of concept for using yeast-derived peptides to functionalize biomaterials for skin wound healing.

Expression of matrix metalloproteinases (MMPs)-2/-7/-9/-14 and tissue inhibitors of MMPs (TIMPs)-1/-2 in bovine cutaneous fibropapillomas associated with BPV-2 infection.

Introduction: Bovine papillomaviruses -1/-2 (BPVs) are small non-enveloped double-stranded DNA viruses able to infect the skin of bovids and equids, causing development of neoplastic lesions such as bovine cutaneous fibropapillomas and equine sarcoid. Matrix metalloproteinases (MMPs) are a group of zinc-dependent endopeptidases that degrade basal membrane and extracellular matrix, whose function is essential in physiological processes such as tissue remodeling and wound healing. MMPs activity is finely regulated by a balancing with expression of tissue inhibitors of MMPs (TIMPs), a process that is impaired during tumour development. BPV infection is associated with upregulation of MMPs and /or their unbalancing with TIMPs, contributing to local invasion and impairment of extracellular matrix remodeling in equine sarcoid; however, studies regarding this topic in bovine fibropapillomas are lacking. Methods: The aim of this study was to perform an immunohistochemical and biochemical analysis on a panel of MMPs and TIMPs in BPV-2 positive bovine cutaneous fibropapillomas vs. normal skin samples. Results: Immunohistochemistry revealed a cytoplasmic expression of MMP-2 (15/19), a cytoplasmic and perinuclear immunoreactivity of MMP-7 (19/19) and MMP-9 (19/19), along with a cytoplasmic and nuclear pattern of MMP-14 (16/19), accompanied by a cytoplasmic expression of TIMP-1 (14/19) and TIMP-2 (18/19) in tumour samples; western blotting revealed an overexpression of MMP-2 (8/9), MMP-7 (9/9) and MMP-9 (9/9), and a decreased level of MMP-14 (9/9), TIMP-1 (9/9) and TIMP-2 (9/9) in tumour versus normal skin samples. Moreover, gelatine zymography confirmed the expression of pro-active MMP-2 (9/9) and MMP-9 (9/9) and, most importantly, indicated the presence and increased activity of their active forms (82 and 62 kDa, respectively) in tumour samples. Discussion: This is the first study describing MMPs and TIMPs in bovine cutaneous fibropapillomas and our results suggest that their unbalanced expression in presence of BPV-2 may play a significant role in tumour development. A further analysis of supplementary MMPs and TIMPs could bring new important insights into the papillomavirus induced tumours.

Improving flexor tendon gliding by using the combination of carboxymethylcellulose-polyethylene oxide on murine model.

The current approach to flexor tendon injuries is complex and is no longer limited to suturing techniques. Strategies for improving hand function currently include rehabilitation protocols, appropriate suturing materials and techniques, changing the gliding surface by using lubricants and providing growth factors. One product, originally used in spinal surgery, has been shown to be effective in preventing postoperative adhesions. It is a combination of carboxymethylcellulose and polyethylene oxide-Dynavisc® (FzioMed, Inc.). The aim of the present study was to test the effect of Dynavisc® on acute injuries of the intrasynovial flexor tendons in the prevention of postoperative adhesions and the improvement of functional results. The study was performed on 20 Wistar rats distributed in two groups. The control group, represented by 10 rats, in which after the reconstruction of the flexor tendon, the peritendinous area was injected with saline solution and the study group, in which the peritendinous area was injected with a single administration of the lubricating gel, Dynavisc® (carboxymethylcellulose and polyethylene oxide). At 4 and 12 weeks, the rats were sacrificed and tissue biopsy consisted of tendon fragments and adjacent tissue. The evaluation of the results was performed by measuring the adhesion score and observing histological parameters. The presence of important adhesions was found in the control group compared with the group treated with Dynavisc®, where a supple and smooth tendon, with significantly fewer adhesions were found. The differences between the two groups were significant, thus indicating the efficiency of the lubricant in preventing adhesions. This study supported the important role of Dynavisc® in the regeneration of the tendon and the peritendinous structures, by limiting aberrant fibrous proliferation in the regeneration process and helping to build a peritendinous space.

Expression of collagenases (matrix metalloproteinase-1, -8, -13) and tissue inhibitor of metalloproteinase-3 (TIMP-3) in naturally occurring bovine cutaneous fibropapillomas.

Bovine cutaneous fibropapillomas are among the most common skin tumors in cattle; their etiology is associated with infection by bovine papillomavirus (BPV) types-1/-2 which are considered oncogenic. Degradation of the extracellular matrix (ECM), especially collagenolysis, is a key event during a series of relevant physiological processes, including tissue remodeling and repair. Various types of proteins are implicated in the regulation of ECM degradation: among these, matrix metalloproteinases (MMPs), a group of zinc-dependent endoenzymes, and tissue inhibitors of matrix metalloproteinases (TIMPs) are known to play a major role. Previous studies reported that aberrant expression of collagenolytic MMPs (MMP-1/-8/-13) and unbalancing between MMPs and TIMPs represent a critical step in tumor growth and invasion; however, studies regarding this topic in bovine cutaneous fibropapillomas are lacking. The aim of this study was to investigate the expression of the collagenases MMP-1/-8/-13 and TIMP-3 in naturally occurring fibropapillomas harboring BPV-2 DNA and normal skin samples. Here, by immunohistochemistry and western blotting analysis, we demonstrated overexpression of MMP-8/-13 along with a down-regulation of MMP-1, associated with a decrease in TIMP-3 levels in tumor compared with normal skin samples. This is the first study describing MMP-1/-8/-13 and TIMP-3 expression in bovine cutaneous fibropapillomas and our results suggest that an impaired expression of collagenases along with an imbalance between MMPs/TIMPs may contribute to an increased collagenolytic activity, which in turn could be important in ECM changes and tumors development.

Novel Mg-0.5Ca-xMn Biodegradable Alloys Intended for Orthopedic Application: An In Vitro and In Vivo Study.

Mg-based biodegradable materials, used for medical applications, have been extensively studied in the past decades. The in vitro cytocompatibility study showed that the proliferation and viability (as assessed by quantitative MTT-assay-3-(4,5-dimethyltiazol-2-yl)-2,5-diphenyl tetrazolium bromide) were not negatively affected with time by the addition of Mn as an alloying element. In this sense, it should be put forward that the studied alloys don't have a cytotoxic effect according to the standard ISO 10993-5, i.e., the level of the cells' viability (cultured with the studied experimental alloys) attained both after 1 day and 5 days was over 82% (i.e., 82, 43-89, 65%). Furthermore, the fibroblastic cells showed variable morphology (evidenced by fluorescence microscopy) related to the alloy sample's proximity (i.e., related to the variation on the Ca, Mg, and Mn ionic concentration as a result of alloy degradation). It should be mentioned that the cells presented a polygonal morphology with large cytoplasmic processes in the vicinity of the alloy's samples, and a bipolar morphology in the remote region of the wells. Moreover, the in vitro results seem to indicate that only 0.5% Mn is sufficient to improve the chemical stability, and thus the cytocompatibility; from this point of view, it could provide some flexibility in choosing the right alloy for a specific medical application, depending on the specific parameters of each alloy, such as its mechanical properties and corrosion resistance. In order to assess the in vivo compatibility of each concentration of alloy, the pieces were implanted in four rats, in two distinct body regions, i.e., the lumbar and thigh. The body's reaction was followed over time, 60 days, both by general clinical examinations considering macroscopic changes, and by laboratory examinations, which revealed macroscopic and microscopic changes using X-rays, CT(Computed Tomography), histology exams and SEM (Scanning Electron Microscopy). In both anatomical regions, for each of the tested alloys, deformations were observed, i.e., a local reaction of different intensities, starting the day after surgery. The release of hydrogen gas that forms during Mg alloy degradation occurred immediately after implantation in all five of the groups examined, which did not affect the normal functionality of the tissues surrounding the implants. Imaging examinations (radiological and CT) revealed the presence of the alloy and the volume of hydrogen gas in the lumbar and femoral region in varying amounts. The biodegradable alloys in the Mg-Ca-Mn system have great potential to be used in orthopedic applications.

Uncommon co-infection due to Moniezia expansa and Moniezia benedeni in young goats from Romania: morphological and histopathological analysis.

Digestive parasitoses negatively affect the goat's health, the gain weight of the kids, the efficiency of food conversion, fertility, and productivity, causing important economic losses. This investigation was carried out on a group of goats, Carpathian breed, in the hill area of Tg. Frumos-Iași, to specify the etiology of the acute digestive syndrome, triggered towards the end of the pasturing season, in the young goats. In this context, four sick animals, aged 6-8 months, were slaughtered. Morphological, pathological, and parasitological examination of dissected animals was conducted. Multiple tapeworms belonging to Moniezia expansa and Moniezia benedeni species were found. From the duodenum, jejunum, ileum, fragments were taken for the histopathological processing. The identification of the tapeworm species was carried out through morphological differentiation. The prevalence of tapeworm infection in the examined animals was 100% of which M. expansa was 56.10% and M. benedeni was 43.90%. The average infection intensity was 10.25 tapeworms/animal. The average intensity of infection with M. expansa was 7.5 tapeworms/ animal, and with M. benedeni it was 4.5 tapeworms/animal. Severe pathological lesions in the small intestine such as occlusion, intestinal obstruction, catarrhal inflammation, total atrophy of intestinal villi, ulcerations were produced by the parasites, which led to the clinical expression of the disease and to direct economic losses. The infection with the Moniezia benedeni species is noticed for the first time in Romania, in young goats.

In Vitro and In Vivo Antioxidant Activity of the New Magnetic-Cerium Oxide Nanoconjugates.

BACKGROUND: Cerium oxide nanoparticles present the mimetic activity of superoxide dismutase, being able to inactivate the excess of reactive oxygen species (ROS) correlated with a large number of pathologies, such as stents restenosis and the occurrence of genetic mutations that can cause cancer. This study presents the synthesis and biological characterisation of nanoconjugates based on nanoparticles of iron oxide interconnected with cerium oxide conjugates. METHODS: The synthesis of magnetite-nanoceria nanoconjugates has been done in several stages, where the key to the process is the coating of nanoparticles with polyethyleneimine and its chemical activation-reticulation with glutaraldehyde. The nanoconjugates are characterised by several techniques, and the antioxidant activity was evaluated in vitro and in vivo. RESULTS: Iron oxide nanoparticles interconnected with cerium oxide nanoparticles were obtained, having an average diameter of 8 nm. Nanoconjugates prove to possess superparamagnetic properties and the saturation magnetisation varies with the addition of diamagnetic components in the system, remaining within the limits of biomedical applications. In vitro free-radical scavenging properties of nanoceria are improved after the coating of nanoparticles with polyethylenimine and conjugation with magnetite nanoparticles. In vivo studies reveal increased antioxidant activity in all organs and fluids collected from mice, which demonstrates the ability of the nanoconjugates to reduce oxidative stress. CONCLUSION: Nanoconjugates possess magnetic properties, being able to scavenge free radicals, reducing the oxidative stress. The combination of the two properties mentioned above makes them excellent candidates for theranostic applications.