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Enterococcus faecalis is a microorganism that can be found in the oral cavity, especially in secondary endodontic infections, with a prevalence ranging from 24–70%. The increase in the ability to form biofilms in the presence of subinhibitory antibiotic concentrations is a phenomenon that is observed for a wide variety of bacterial pathogens and is associated with increased resistance. In this study, therefore, six E. faecalis isolates from an endodontic environment and two control strains were exposed to subinhibitory concentrations of Penicillin G, Amoxicillin, Doxycycline, Fosfomycin, Tetracycline and Vancomycin and examined for their biofilm formation abilities. The minimum inhibitory concentration (MIC) was determined for all E. faecalis isolates. A culture of the isolate was mixed with a serial dilution series of the respective antibiotic, incubated overnight and the biofilm formation was analyzed using a microtiter plate assay. All isolates were able to form biofilms in the absence of an antibiotic. A significant increase in biofilm formation of up to more than 50% was found in the isolates exposed to subinhibitory concentrations of various antibiotics. Most isolates showed a significant increase in Fosfomycin (7/8), Doxycycline (6/8) and Tetracycline (6/8). Three endodontic isolates showed a significant increase in five of the antibiotics examined at the same time. On exposure to Vancomycin, three endodontic isolates and the two control strains showed an increase. The increase in the ability to form biofilms extended over a concentration range from 1/2 to 1/64 of the MIC concentration. Antibiotics may reach certain niches in the oral cavity at subinhibitory concentrations only. This can increase the biofilm formation by enterococci, and in turn lead to decreased susceptibility of these taxa to antibiotics.

Mancozeb is an ethylene bis-dithiocarbamate widely used as fungicide, also due to a low reported toxicity in mammals. However, reproductive toxicity has been demonstrated in vivo and in vitro in mouse oocytes, by the alteration of spindle morphology [1] and impairment of fertilizability [2]. Mancozeb exerted on mouse GCs cultured in vitro a premalignant-like status, indicated by reduced p53 expression [3] and a mild oxidative stress [4]. However, presence and extent of ultrastructural alterations induced in vitro by Mancozeb on GCs were not yet studied. To this aim, Transmission Electron Microscopy (TEM) and Scanning Electron Microscopy (SEM) were applied on mouse GCs cultured with increasing concentration of Mancozeb. GCs were obtained by puncturing antral follicles of PMSG-treated prepubertal CD1 female mice and cultured in vitro in DMEM+5%FBS+pen/strep without (control) or with increasing concentration of Mancozeb (0.001-to-1 µg/ml) for 48hrs, at 37°C and 5%CO2. At the end of the culture period, cells were washed in PBS, fixed in 2.5% glutaraldehyde/PBS and stored at 4°C until processing. GCs were, then, subjected to standard preparative for TEM [5] and SEM [6] observation. Results showed a dose-dependent toxicity of Mancozeb on mouse GCs. Ultrastructural data showed intercellular communication retraction, irregular nuclear membrane and chromatin marginalization at lower concentrations; chromatin condensation, membrane blebbing and cytoplasmic vacuolization at higher concentrations. In conclusion, Mancozeb showed a dose-dependent harmful effect on granulosa cells in vitro, probably due to the toxic breakdown product ethylenethiourea. TEM and SEM were again confirmed to be a valuable tool to study ultrastructural alterations after toxicants exposure., Italian Journal of Anatomy and Embryology, Vol. 121, No. 1 (Supplement) 2016

Cryopreservation of human spermatozoa is widely used in many assisted reproduction units to preserve male fertility [1]. Vitrification is based on the ultrarapid freezing and is routinely assayed for cryopreservation in assisted reproductive technology. Mohamed [2] showed that cryopreservation significantly affects progressive motility, viability and mitochondrial membrane potential of spermatozoa. Pentoxifylline (PX) is a phosphodiesterase considered to be a sperm movement enhancer, hyperactivation agent, inhibitor of reactive oxygen species and acrosome reaction-improving agent. The aim of our study was to evaluate the effect of in-vitro application of PX on sperm parameters and ultrastructure after vitrification. A total of 30 asthenozoospermic semen samples were selected and divided into two groups after vitrification: control (without PX) and experimental (with PX). A significant decrease in sperm motility, morphology and viability was observed post vitrification, but sperm motility was increased significantly following application of PX. On the other hand, PX did not exert any significant effect on the ultrastructure of the acrosome, plasma membrane and tail of vitrified spermatozoa., Italian Journal of Anatomy and Embryology, Vol. 122, No. 1 (Supplement) 2017

Anatomia Umana - Atlante si avvale di un supporto cartaceo e di una piattaforma web (Virtual Campus), combinati in modo innovativo per l’apprendimento dell’anatomia. L’Atlante cartaceo segue un’organizzazione regionale e topografica. Ciascun capitolo è illustrato con l’ausilio di: disegni anatomici magistralmente realizzati e di sicuro effetto didattico; sequenze stratigrafiche per ricostruire l’organizzazione delle regioni anatomiche; realistiche immagini di renderizzazione 2D o 3D a partire da esami TC o RM; fotografie di esami laparoscopici o endoscopici; leggende chiare, tavole riassuntive e schemi che favoriscono l’apprendimento; richiami didascalici alle risorse presenti sulla piattaforma web. La piattaforma web - Virtual campus, innovativa nel mondo degli atlanti di anatomia, raccoglie un ricco catalogo di materiale didattico: corso di Anatomia Topografica completo, organizzato in percorsi guidati che conducono lo studente attraverso specifiche risorse della piattaforma web; ricostruzioni 3D interattive di organi o regioni anatomiche, a partire da esami TC o RM; video commentati di laparoscopie ed endoscopie; video commentati di dissezioni anatomiche su cadavere; animazioni stratigrafiche interattive

Anatomia Umana - Atlante si avvale di un supporto cartaceo e di una piattaforma web, combinati in modo innovativo per l’apprendimento dell’anatomia. L’Atlante cartaceo segue un’organizzazione regionale e topografica. Ciascun capitolo è illustrato con l’ausilio di: disegni anatomici magistralmente realizzati e di sicuro effetto didattico; sequenze stratigrafiche per ricostruire l’organizzazione delle regioni anatomiche; realistiche immagini di renderizzazione 2D o 3D a partire da esami TC o RM; fotografie di esami laparoscopici o endoscopici; leggende chiare, tavole riassuntive e schemi che favoriscono l’apprendimento; richiami didascalici alle risorse presenti sulla piattaforma web. La piattaforma web, innovativa nel mondo degli atlanti di anatomia, raccoglie un ricco catalogo di materiale didattico: corso di Anatomia Topografica completo, organizzato in percorsi guidati che conducono lo studente attraverso specifiche risorse della piattaforma web; ricostruzioni 3D interattive di organi o regioni anatomiche, a partire da esami TC o RM; video commentati di laparoscopie ed endoscopie; video commentati di dissezioni anatomiche su cadavere; animazioni stratigrafiche interattive.

The excessive exposure to pesticides in the Aral Sea area was correlated to the increased reproductive pathologies in those regions [1]. One of the principal chemical employed was the gamma-hexachlorocycloexane herbicide Lindane (L), a persistent organochlorine that may induces alterations in granulosa cell (GCs) survival [2, 3]. However, a comprehensive experimental study on the L-induced dose-effect morpho- logical alterations, has not yet addressed. Therefore, we studied by means of trans- mission and scanning electron microscopy, the morphological changes of mouse GCs, matured in vitro with increasing concentrations of L [4-6].GCs showed several dose-dependent changes, in respect to controls. In particular, we observed significant reduction of GC microvilli and decrease of cytoplasmic pro- cesses between adjacent GCs. In addition, peripheral aggregation of chromatin under the nuclear membrane, extensive plasma membrane blebbing, abundant GC remnants and cellular debris were also present. Mitochondria, endoplasmic reticula and Golgi apparatuses did not show significant changes. In conclusion, our results showed a dose-dependent toxicity of L on GCs, associated to morphological signs of apoptosis. Alterations of GCs may be associated to impaired oocyte competence and sterility [7]., Italian Journal of Anatomy and Embryology, Vol. 120, No. 1 (Supplement) 2015

During the past two decades important advances have been made in the field of assisted reproduction by using oocyte cryopreservation. However, mature (metaphase II) oocytes are very susceptible to cryodamage. In order to contribute to the identification of a cryopreservation protocol with minimal side effects on the oocyte structure and function, we evaluated and compared the subcellular features of human oocytes cryopreserved either with slow (controlled rate) freezing or vitrification (ultrarapid freezing). Supernumerary human metaphase II oocytes were donated by consenting patients enrolled in an IVF program. The age of these women ranged from 27 to 32 years old. The eggs were cryopreserved using slow freezing with 1.5M propanediol + 0.2M sucrose concentration or a closed vitrification system (Cryotip Irvine Scientific CA). Fresh oocytes were used as controls. Samples were fixed and prepared for light and transmission electron microscopy (LM and TEM) observations. By LM, all the oocytes were generally rounded, 90-100 microns in diameter, with regular ooplasm showing uniform distribution of organelles. By TEM, mitochondria-smooth endoplasmic reticulum (M-SER) aggregates were the most common structures found in all the oocytes fixed or cryopreserved within 3-4 hours after the retrieval. M-SER aggregates appeared instead partially replaced by mitochondria-vesicle complexes when oocytes were maintained in culture for a prolonged period of time. A slight to moderate vacuolization was found in the cytoplasm of the oocytes subjected to slow freezing. Slight microvacuolization was also found in vitrified oocytes, whereas vacuoles were almost completely absent in fresh controls. Amount and density of cortical granules (CGs) appeared abnormally reduced in all cryopreserved oocytes, irrespective of the protocol applied. In conclusion, it has been evidenced that prolonged stay in culture induces an intracellular membrane “recycling” in the oocytes, that causes the transformation of slender, anastomosed SER tubules into rounded vesicles surrounded by mitochondria, whose role is still uncertain. In addition, even though all cryopreservation protocols studied ensured a good overall preservation of the oocyte, vacuolization appears as a recurrent form of cell damage. This happens both during slow freezing and, at a lesser extent, during vitrification using a closed device. In addition, premature CG exocytosis was observed in both groups., Italian Journal of Anatomy and Embryology, Vol 117, No 2 (Supplement) 2012