Vancomycin-Loaded, Nanohydroxyapatite-Based Scaffold for Osteomyelitis Treatment: In Vivo Rabbit Toxicological Tests and In Vivo Efficacy Tests in a Sheep Model.

The treatment for osteomyelitis consists of surgical debridement, filling of the dead space, soft tissue coverage, and intravenous administration of antimicrobial (AM) agents for long periods. Biomaterials for local delivery of AM agents, while providing controllable antibiotic release rates and simultaneously acting as a bone scaffold, may be a valuable alternative; thus, avoiding systemic AM side effects. V-HEPHAPC is a heparinized nanohydroxyapatite (nHA)/collagen biocomposite loaded with vancomycin that has been previously studied and tested in vitro. It enables a vancomycin-releasing profile with an intense initial burst, followed by a sustained release with concentrations above the Minimum Inhibitory Concentration (MIC) for MRSA. In vitro results have also shown that cellular viability is not compromised, suggesting that V-HEPHAPC granules may be a promising alternative device for the treatment of osteomyelitis. In the present study, V-HEPHAPC (HEPHAPC with vancomycin) granules were used as a vancomycin carrier to treat MRSA osteomyelitis. First, in vivo Good Laboratory Practice (GLP) toxicological tests were performed in a rabbit model, assuring that HEPHAPC and V-HEPHAPC have no relevant side effects. Second, V-HEPHAPC proved to be an efficient drug carrier and bone substitute to control MRSA infection and simultaneously reconstruct the bone cavity in a sheep model.

Microplastic appraisal of soil, water, ditch sediment and airborne dust: The case of agricultural systems.

Although microplastic pollution jeopardizes both terrestrial and aquatic ecosystems, the movement of plastic particles through terrestrial environments is still poorly understood. Agricultural soils exposed to different managements are important sites of storage and dispersal of microplastics. This study aimed to identify the abundance, distribution, and type of microplastics present in agricultural soils, water, airborne dust, and ditch sediments. Soil health was also assessed using soil macroinvertebrate abundance and diversity. Sixteen fields were evaluated, 6 of which had been exposed to more than 5 years of compost application, 5 were exposed to at least 5 years of plastic mulch use, and 5 were not exposed to any specific management (controls) within the last 5 years. We also evaluated the spread of microplastics from the farms into nearby water bodies and airborne dust. We found 11 types of microplastics in soil, among which Light Density Polyethylene (LDPE) and Light Density Polyethylene covered with pro-oxidant additives (PAC) were the most abundant. The highest concentrations of plastics were found in soils exposed to plastic mulch management (128.7 ± 320 MPs.g-1 soil and 224.84 ± 488 MPs.g-1 soil, respectively) and the particles measured from 50 to 150 µm. Nine types of microplastics were found in water, with the highest concentrations observed in systems exposed to compost. Farms applying compost had higher LDPE and PAC concentrations in ditch sediments as compared to control and mulch systems; a significant correlation between soil polypropylene (PP) microplastics with ditch sediment microplastics (r2 0.7 p < 0.05) was found. LDPE, PAC, PE (Polyethylene), and PP were the most abundant microplastics in airborne dust. Soil invertebrates were scarce in the systems using plastic mulch. A cocktail of microplastics was found in all assessed matrices.

Eyelid Reconstruction and Infraorbital Repair Using a Mustardé Flap Technique in a Dog.

This case report describes a 5-year-old dog with a defect in the right eyelid, absence of orbicularis musculature, and absence of cutaneous tissue in the infraorbital region, submitted to the Mustardé flap technique. A large rotational flap was performed, including a cartilage graft from the outer ear, to correct the defect in the lower eyelid and infraorbital region. Also, euryblepharon correction of the upper eyelid was performed with wedge excision. The techniques were performed in a single surgical step and with appropriated functional and aesthetic results. The described approach may be employed as a surgical option in large eyelid defects.

Evaluation of eye melanoma treatments in rabbits: A systematic review.

Melanomas are the most common cancer of the eye in canines, felines and humans. The treatment approaches vary, since no gold standard exists. Therefore, this systematic review aimed to compare the treatment modalities in ocular melanoma in rabbits. Medline/PubMed, Cochrane Library, Web of Science and Embase were searched for articles published until 21 April 2021 in English, Portuguese or Spanish, reporting animal studies evaluating photodynamic therapy (PDT), laser, radiotherapy or surgical excision. Twenty-seven articles were included for the qualitative synthesis, with publication dates from 1970 to 2018. Of the selected studies, 19 used PDT, six used radiotherapy and two used laser as treatment. No studies regarding surgical therapy that met the inclusion criteria were obtained. The tumour therapy results were evaluated in a heterogeneous manner for different periods and various methods, including microscopy, angiographic, histological examination, fundoscopy, ultrasound exam and electroretinogram. The treatment modalities analysed successfully treated the ocular melanoma, with tumour necrosis being commonly observed. Despite the therapeutic efficacy shown, side effects have been reported for all the therapies. The studies showed high heterogeneity, and therefore, in the future, new studies should be carried out to increase knowledge about ocular melanoma treatment. The analysed therapies can be used successfully in the treatment of ocular melanoma, with more conservative options such as PDT presenting great potential.

Current Therapeutics and Future Perspectives to Ocular Melanocytic Neoplasms in Dogs and Cats.

Neoplasms of melanocytic origin are diseases relevant to dogs and cats' ophthalmic oncology due to their incidence, potential visual loss, and consequent decrease in life quality and expectancy. Despite its non-specific clinical presentation, melanocytic neoplasms can be histologically distinguished in melanocytomas, which present benign characteristics, and malignant melanomas. The diagnosis often occurs in advanced cases, limiting the therapeutic options. Surgery, cryotherapy, radiotherapy, photodynamic therapy (PDT), and laser are currently available therapeutic strategies. As no clinical guidelines are available, the treatment choice is primarily based on the clinician's preference, proficiency, and the owner's financial constraints. While surgery is curative in benign lesions, ocular melanomas present a variable response to treatments, besides the potential of tumour recurrences or metastatic disease. This review presents the currently available therapies for ocular melanocytic neoplasms in dogs and cats, describing the therapeutic, indications, and limitations. Additionally, new therapeutics being developed are presented and discussed, as they can improve the current treatment options.

Prevalence of Radiographic Signs of Osteoarthritis in Lusitano Purebred Horses.

Radiographic prevalence and correlation of radiographic findings has not been performed in Lusitano Purebred horses. The aim of this study was to (1) evaluate the prevalence of primary osteoarthritis radiographic findings in Lusitano Purebred horses; (2) to assess correlations between radiographic findings in different joints of the same limb and different limbs; and (3) elucidate the effect of age in the radiographic findings. A radiographic protocol of the stifle, tarsi, fetlocks and distal limbs was done in 98 Lusitanos and the classification of the radiographs was performed using a 0-4 scale developed and applied blindly by three veterinarians. The distal interphalangeal, proximal interphalangeal, metacarpophalangeal, metatarsophalangeal, tarsometatarsal, distal intertarsal, proximal intertarsal/, tibiotarsal and femorotibial-patellar joints were evaluated. Most joints presented no abnormal findings or minor abnormal radiographic findings (82.86% grade ≤1). The most affected joint was tarsometatarsal and more severe lesions were found in tarsometatarsal and distal intertarsal. Femorotibial-patellar radiographic changes were rare (2.13%). A strong/moderate correlation was found between contralateral joints with exception hindlimb fetlocks. A moderate correlation was found between fore and hindlimbs for distal limb joints. When analyzing ipsilateral as well as diagonal distal limbs, a strong/moderate correlation was also found. The total score progressed in 0.2 score points per each year of age, revealing that age can be a statistically significant predictor for radiographic changes. Overall, Lusitano horses presented a low prevalence of severe radiographic sings of primary osteoarthritis. Findings in contralateral joints tend to be correlated.

Long term performance evaluation of small-diameter vascular grafts based on polyvinyl alcohol hydrogel and dextran and MSCs-based therapies using the ovine pre-clinical animal model.

The functional and structural performance of a 5cm synthetic small diameter vascular graft (SDVG) produced by the copolymerization of polyvinyl alcohol hydrogel with low molecular weight dextran (PVA/Dx graft) associated to mesenchymal stem cells (MSCs)-based therapies and anticoagulant treatment with heparin, clopidogrel and warfarin was tested using the ovine model during the healing period of 24 weeks. The results were compared to the ones obtained with standard expanded polyetetrafluoroethylene grafts (ePTFE graft). Blood flow, vessel and graft diameter measurements, graft appearance and patency rate (PR), thrombus, stenosis and collateral vessel formation were evaluated by B-mode ultrasound, audio and color flow Doppler. Graft and regenerated vessels morphologic evaluation was performed by scanning electronic microscopy (SEM), histopathological and immunohistochemical analysis. All PVA/Dx grafts could maintain a similar or higher PR and systolic/diastolic laminar blood flow velocities were similar to ePTFE grafts. CD14 (macrophages) and α-actin (smooth muscle) staining presented similar results in PVA/Dx/MSCs and ePTFE graft groups. Fibrosis layer was lower and endothelial cells were only detected at graft-artery transitions where it was added the MSCs. In conclusion, PVA/Dx graft can be an excellent scaffold candidate for vascular reconstruction, including clinic mechanically challenging applications, such as SDVGs, especially when associated to MSCs-based therapies to promote higher endothelialization and lower fibrosis of the vascular prosthesis, but also higher PR values.

Long term performance evaluation of small-diameter vascular grafts based on polyvinyl alcohol hydrogel and dextran and MSCs-based therapies using the ovine pre-clinical animal model.

The functional and structural performance of a 5cm synthetic small diameter vascular graft (SDVG) produced by the copolymerization of polyvinyl alcohol hydrogel with low molecular weight dextran (PVA/Dx graft) associated to mesenchymal stem cells (MSCs)-based therapies and anticoagulant treatment with heparin, clopidogrel and warfarin was tested using the ovine model during the healing period of 24 weeks. The results were compared to the ones obtained with standard expanded polyetetrafluoroethylene grafts (ePTFE graft). Blood flow, vessel and graft diameter measurements, graft appearance and patency rate (PR), thrombus, stenosis and collateral vessel formation were evaluated by B-mode ultrasound, audio and color flow Doppler. Graft and regenerated vessels morphologic evaluation was performed by scanning electronic microscopy (SEM), histopathological and immunohistochemical analysis. All PVA/Dx grafts could maintain a similar or higher PR and systolic/diastolic laminar blood flow velocities were similar to ePTFE grafts. CD14 (macrophages) and α-actin (smooth muscle) staining presented similar results in PVA/Dx/MSCs and ePTFE graft groups. Fibrosis layer was lower and endothelial cells were only detected at graft-artery transitions where it was added the MSCs. In conclusion, PVA/Dx graft can be an excellent scaffold candidate for vascular reconstruction, including clinic mechanically challenging applications, such as SDVGs, especially when associated to MSCs-based therapies to promote higher endothelialization and lower fibrosis of the vascular prosthesis, but also higher PR values.

In vitro and in vivo evaluation of blood coagulation activation of polyvinyl alcohol hydrogel plus dextran-based vascular grafts.

Polyvinyl alcohol hydrogel (PVA) is a water-soluble synthetic polymer that is commonly used in biomedical applications including vascular grafting. It was argued that the copolymerization of PVA with dextran (Dx) can result in improvement of blood-biomaterial interactions. The focus of this experimental study was to assess that interaction through an in vivo and in vitro evaluation of the coagulation system activation. The thrombogenicity of the copolymer was determined by quantification of platelet adhesion through the lactate dehydrogenase assay, determination of whole blood clotting time, and by quantification of platelet activation by flow cytometry. The thrombin-antithrombin complex blood levels were also determined. The obtained results for the in vitro assays suggested a non-thrombogenic profile for PVA/Dx. Additionally in vivo coagulation and hematological parameters were determined in an animal model after PVA/Dx vascular graft implantation. For coagulation homeostasis assessment, the intrinsic and extrinsic pathway's activation was determined by measuring prothrombin time (PT) and activated partial thromboplastin time (aPTT). Other markers of coagulation and inflammation activation including d-dimers, interleukin-6, and C-reactive protein were also assessed. The PVA/Dx copolymer tended to inhibit platelet adhesion/activation process and the contact activation process for coagulation. These results were also confirmed with the in vivo experiments where the measurements for APTT, interleukin-6, and C-reactive protein parameters were normal considering the species normal range of values. The response to those events is an indicator of the in vitro and in vivo hemocompatibility of PVA/Dx and it allows us to select this biomaterial for further preclinical trials in vascular reconstruction.

Biocompatibility and hemocompatibility of polyvinyl alcohol hydrogel used for vascular grafting--In vitro and in vivo studies.

Polyvinyl alcohol hydrogel (PVA) is a synthetic polymer with an increasing application in the biomedical field that can potentially be used for vascular grafting. However, the tissue and blood-material interactions of such gels and membranes are unknown in detail. The objectives of this study were to: (a) assess the biocompatibility and (b) hemocompatibility of PVA-based membranes in order to get some insight into its potential use as a vascular graft. PVA was evaluated isolated or in copolymerization with dextran (DX), a biopolymer with known effects in blood coagulation homeostasis. The effects of the mesenchymal stem cells (MSCs) isolated from the umbilical cord Wharton's jelly in the improvement of PVA biocompatibility and in the vascular regeneration were also assessed. The biocompatibility of PVA was evaluated by the implantation of membranes in subcutaneous tissue using an animal model (sheep). Histological samples were assessed and the biological response parameters such as polymorphonuclear neutrophilic leucocytes and macrophage scoring evaluated in the implant/tissue interface by International Standards Office (ISO) Standard 10993-6 (annex E). According to the scoring system based on those parameters, a total value was obtained for each animal and for each experimental group. The in vitro hemocompatibility studies included the classic hemolysis assay and both human and sheep bloods were used. Relatively to biocompatibility results, PVA was slightly irritant to the surrounding tissues; PVA-DX or PVA plus MSCs groups presented the lowest score according to ISO Standard 10993-6. Also, PVA was considered a nonhemolytic biomaterial, presenting the lowest values for hemolysis when associated to DX.

Detection of Rickettsia conorii strains in Portuguese dogs (Canis familiaris).

This study reports a serological and molecular survey on the spotted fever group Rickettsia found in a group of 51 dogs with suspected tick-borne illness from the south of Portugal. Additionally, a prevalence of IgG antibodies to R. conorii was also estimated in a group of 400 healthy dogs from the same region. In the group of healthy dogs, the immunofluorescence test revealed that 154 (38.5%) of the 400 dogs had IgG antibodies reactive with R. conorii. The highest proportion of dogs (45%) with R. conorii antibodies was found in blood samples collected from October to December. Among the group suspected with tick-borne illness, 35 (62%) dogs showed to be seroreactive (IgG≥128) for antibodies against R. conorii, and the analysis of PCR-positive amplicons revealed that 5 dogs were infected with R. conorii Malish and 2 dogs were infected with R. conorii Israeli tick typhus strain. There was a higher prevalence of antibodies to R. conorii in the group of sick dogs, and the detection of R. conorii DNA in blood samples from this group points to their potential role as a reservoir and sentinel host helping to evaluate and characterize the distribution of circulating rickettsial strains.

Detection of Ehrlichia canis by polymerase chain reaction in dogs from Portugal.

Antibodies against Ehrlichia canis, the cause of canine monocytic ehrlichiosis, have been reported previously in clinically ill and stray dogs from Portugal. In this study, the 16S rRNA gene of E. canis was detected by the polymerase chain reaction (PCR) in 12/55 (22%) of dogs with suspected tick-borne disease in the Algarve region in Portugal.