The search of suitable combinations of stem cells, biomaterials and scaffolds manufacturing methods have become a major focus of research for bone engineering. The aim of this study was to test the potential of dental pulp stem cells to attach, proliferate, mineralize and differentiate on 3D printed polycaprolactone (PCL) scaffolds. A 100% pure M_w: 84,500 ± 1000 PCL was selected. 5 × 10 × 5 mm³ parallelepiped scaffolds were designed as a wood-pilled structure composed of 20 layers of 250 μm in height, in a non-alternate order ([0,0,0,90,90,90°]). 3D printing was made at 170 °C. Swine dental pulp stem cells (DPSCs) were extracted from lower lateral incisors of swine and cultivated until the cells reached 80% confluence. The third passage was used for seeding on the scaffolds. Phenotype of cells was determined by flow Cytometry. Live and dead, Alamar blue™, von Kossa and alizarin red staining assays were performed. Scaffolds with 290 + 30 μm strand diameter, 938 ± 80 μm pores in the axial direction and 689 ± 13 μm pores in the lateral direction were manufactured. Together, cell viability tests, von Kossa and Alizarin red staining indicate the ability of the printed scaffolds to support DPSCs attachment, proliferation and enable differentiation followed by mineralization. The selected material-processing technique-cell line (PCL-3D printing-DPSCs) triplet can be though to be used for further modelling and preclinical experiments in bone engineering studies.

Skin infections have been associated with Acanthamoeba, nevertheless the events during skin invasion and UV-B light effects on it are unknown. The early morphological events of Acanthamoeba castellanii skin invasion are shown in SKH-1 mice that were chronically UV-B light irradiated. Mice that developed skin lesions (group 1) were topical and intradermally inoculated with A. castellanii trophozoites and sacrificed 48 h or 18 days later. Mice that showed no skin lesions (group 2) were intradermally inoculated and sacrificed 24, 48 or 72 h later. Mice ventral areas were considered controls with and without trophozoites intradermally inoculated. Skin samples were processed by histological and immunohistochemistry techniques. In group 1, trophozoites were immunolocalized in dermal areas, hair cysts, sebaceous glands, and blood vessels, and collagen degradation was observed. One of these mice shown trophozoites in the spleen, liver, and brain. In group 2, few trophozoites nearby collagenolytic activity zones were observed. In control samples, nor histological damage and no trophozoites were observed. Adherence and collagenolytic activity by A. castellanii were corroborated in vitro. We can infer that UV-B light irradiated skin could favor A. castellanii invasiveness causing damage in sites as far away as the brain, confirming the invasive capacity and pathogenic potential of these amphizoic amoebae.

Amoebae of the genus Acanthamoeba are etiological agents of amoebic keratitis, for which up to now there is no treatment of choice and one of its main risk factors is the use of contact lenses, including cosmetic contact lenses. Recently there has been an increase in amoebic keratitis cases due to the use of cosmetic contact lenses. Therefore, having a solution for the care of lenses with an efficient disinfectant effect that prevents the adhesion of trophozoites to lenses becomes essential. This study was carried out to determine the effect of 8 multipurpose contact lenses care solutions on Acanthamoeba castellanii trophozoites viability, and the efficiency of two of them to prevent the trophozoites adherence onto two cosmetic contact lenses (Acuvue 2, approved by the US Food and Drug Administration, and Magic Eye CCL, not approved). After 3 h of interaction, only AO Sept Plus, OPTI FREE Replenish, Renu Plus, Bio True and Multiplus significantly reduced the number of viable trophozoites with respect to the control; at 6 h Renu Plus, and at 12 h Conta Soft Plus and Multiplus, maintained the inhibitory effect. Only Opti Free Pure Moist did not significantly reduce the number of viable trophozoites. Multiplus and Opti Free Pure Moist (selected for their greater and lesser antiamibic effect) significantly reduced trophozoite adherence to both lenses; however, Opti Free Pure Moist was more efficient, despite the fact that A. castellanii adhered similarly to both lenses. Our results show that in all the multipurpose solutions evaluated, hundreds of viable A. castellanii trophozoites remain after several hours of incubation. Therefore, storage of the lenses in their case with MPS maintains the potential risk of amoebic keratitis in, cosmetic contact lenses wearers. Moreover, the use of CCL, not approved by the FDA, can increase the risk factor for AK since its poor manufacture can favor the permanence of amoebae, in addition to being a risk for corneal integrity.

Several species of Acanthamoeba genus are potential pathogens and etiological agents of several diseases. The pathogenic mechanisms carried out by these amoebae in different target tissues have been documented, evidencing the relevant role of contact-dependent mechanisms. With the purpose of describing the pathogenic processes carried out by these protozoans more precisely, we considered it important to determine the emission of extracellular vesicles (EVs) as part of the contact-independent pathogenicity mechanisms of A. culbertsoni, a highly pathogenic strain. Through transmission electronic microscopy (TEM) and nanoparticle tracking analysis (NTA), EVs were characterized. EVs showed lipid membrane and a size between 60 and 855 nm. The secretion of large vesicles was corroborated by confocal and TEM microscopy. The SDS-PAGE of EVs showed proteins of 45 to 200 kDa. Antigenic recognition was determined by Western Blot, and the internalization of EVs by trophozoites was observed through Dil-labeled EVs. In addition, some EVs biological characteristics were determined, such as proteolytic, hemolytic and COX activity. Furthermore, we highlighted the presence of leishmanolysin in trophozites and EVs. These results suggest that EVs are part of a contact-independent mechanism, which, together with contact-dependent ones, allow for a better understanding of the pathogenicity carried out by Acanthamoeba culbertsoni.

Diabetic nephropathy (DN) is a serious complication of diabetes mellitus; oxidative stress plays a key role in the pathogenesis of DN. The objective of this study was to evaluate the antioxidant effect of vitamin E on diabetic nephropathy. A control group and three groups of rats with streptozotocin-induced diabetes mellitus (untreated diabetic rats and diabetic rats treated with vitamin E 250 and 500 mg/kg) were studied. After 4 weeks of treatment, the kidneys were removed under anesthesia with sodium pentobarbital. The kidneys were weighed, the AT₁ and AT₂ receptor expression was measured by Western blot, and the activities of glutathione peroxidase, catalase, and superoxide dismutase were determined in the renal cortex. Rats with diabetes mellitus had hyperglycemia, increased food and water consumption, and higher urinary volume than control rats. In diabetic rats (DM), kidney hypertrophy was observed and measured by kidney weight, protein/DNA ratio in the renal cortex, and proximal tubular cell area; proteinuria and reduced creatinine clearance were observed. AT₁ and AT₂ receptor expression in the kidney cortex of DM rats increased significantly compared to normoglycemic rats; antioxidant enzyme activities were decreased; treatment with vitamin E reversed kidney hypertrophy and reduced proteinuria; reduction in expression of AT₁ and AT₂ receptors was associated with increased antioxidant activity. Thus, treatment with vitamin E slows the progress of DN.