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The global dissemination of multi-drug resistant (MDR) pathogenic bacteria requires the rapid research and development of alternative therapies that can support or replace conventional antibiotics. Among MDR pathogens, carbapenem-resistant Klebsiella pneumoniae (CR-Kp) are of particular concern due to their extensive resistance profiles, global dissemination in hospital environments, and their major role in some life-threatening infections. Phages, or some of their components, are recognized as one of the potential alternatives that might be helpful to treat bacterial infections. In this study, we have isolated and characterized four lytic bacteriophages targeting K. pneumoniae strains of Sequence Type (ST) 307 or ST147, two predominant high-risk clones of CR-Kp. Phages, designated vB_KpS_GP-1, vB_KpP_GP-2, vB_KpP_GP-4, and vB_KpP_GP-5, were isolated from sewage wastewater samples. The vB_KpS_GP-1 phage was a siphovirus unable to establish lysogeny with its host, while the other three were podoviruses. While 85.7% of K. pneumoniae strains of ST307 were selectively lysed by the phages vB_KpS_GP-1 or vB_KpP_GP-5, the other two phages were able to lyse all the tested strains of ST147 (n = 12). Phages were stable over a broad pH and temperature range and were characterized by burst sizes of 10-100 plaque forming units and latency periods of 10-50 minutes. Genome sequencing confirmed the absence of antibiotic resistance genes, virulence factors or toxins and revealed that two phages were likely members of new genera. Given their strictly lytic nature and high selectivity towards two of the major high-risk clones of K. pneumoniae, cocktails of these phages could represent promising candidates for further evaluation in in vivo experimental models of K. pneumoniae infection. [ABSTRACT FROM AUTHOR]

Primary ciliary dyskinesia (PCD) is an inherited autosomal-recessive disorder characterized by abnormal ciliary motion, due to a defect in ciliary structure and/or function. This genetic condition leads to recurrent upper and lower respiratory infections, bronchiectasis, laterality defect, and subfertility. Male infertility is often associated with PCD, since the ultrastructure of the axoneme in the sperm tail is similar to that of the motile cilia of respiratory cells. We present the first reported case of a male patient from a non-consanguineous Italian family who exhibited a severe form of asthenozoospermia factor infertility but no situs inversus and absolutely no signs of the clinical respiratory phenotype, the proband being a professional basketball player. Whole-exome sequencing (WES) has identified a homozygote mutation (CCDC103 c.461 A>C, p.His154Pro) in the proband, while his brother was a heterozygous carrier for this mutation. Morphological and ultrastructural analyses of the axoneme in the sperm flagellum demonstrated the complete loss of both the inner and outer dynein arms (IDA and ODA, respectively). Moreover, immunofluorescence of DNAH1, which is used to check the assembly of IDA, and DNAH5, which labels ODA, demonstrated that these complexes are absent along the full length of the flagella in the spermatozoa from the proband, which was consistent with the IDA and ODA defects observed. Noteworthy, TEM analysis of the axoneme from respiratory cilia showed that dynein arms, although either IDAs and/or ODAs seldom missing on some doublets, are still partly present in each observed section. This case reports the total sperm immotility associated with the CCDC103 p.His154Pro mutation in a man with a normal respiratory phenotype and enriches the variant spectrum of ccdc103 variants and the associated clinical phenotypes in PCD, thus improving counseling of patients about their fertility and possible targeted treatments. [ABSTRACT FROM AUTHOR]

A coating technology based on low molecular weight hyaluronic acid (HA) and ferulic acid (FA) was applied to the coating of low generation poly(propylene imine) dendrimers through a biocompatible hexa(ethylene glycol) spacer. The ensuing HA‐FA‐HEG‐PPID dendrimeric materials showed interesting loading capability (between 7.65% and 9.08%) regarding anticancer agent doxorubicin, and their interactions with the drug appeared to hamper the drug release in the physiological environment. Thus, the stable nanostructured loaded delivery systems were able to internalize into cells expressing the HA receptor CD44 and to demonstrate high cytotoxicity comparable to that shown by equivalent amounts of free doxorubicin. Thus, HA‐FA‐HEG‐PPID dendrimeric materials were proposed as biocompatible drug carriers capable of transporting anticancer doxorubicin to tumor cells. [ABSTRACT FROM AUTHOR]

HBDI‐like chromophores represent a novel set of biomimetic switches mimicking the fluorophore of the green fluorescent protein that are currently studied with the hope to expand the molecular switch/motor toolbox. However, until now members capable of absorbing visible light in their neutral (i. e. non‐anionic) form have not been reported. In this contribution we report the preparation of an HBDI‐like chromophore based on a 3‐phenylbenzofulvene scaffold capable of absorbing blue light and photoisomerizing on the picosecond timescale. More specifically, we show that double‐bond photoisomerization occurs in both the E‐to‐Z and Z‐to‐E directions and that these can be controlled by irradiating with blue and UV light, respectively. Finally, as a preliminary applicative result, we report the incorporation of the chromophore in an amphiphilic molecule and demonstrate the formation of a visible‐light‐sensitive nanoaggregated state in water. [ABSTRACT FROM AUTHOR]

Despite the major target of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, being the respiratory system, clinical evidence suggests that the male reproductive system may represent another viral target organ. Revealing the effect of SARS-CoV-2 infection on testis and sperm is a priority for reproductive biology, as well as for reproductive medicine. Here, we confirmed that the SARS-CoV-2 receptor angiotensin-converting enzyme 2 (ACE2) is highly expressed on human testis and ejaculated sperm; moreover, we provide evidence for the expression of the co-receptors transmembrane protease/serine (TMPRSS2), Basigin (BSG), and Catepsin L (CTSL). Human sperm were readily infected, both in vivo and in vitro, by SARS-CoV-2, as demonstrated by confocal and electron microscopy. The demonstration that the seminiferous epithelium and sperm support SARS-CoV-2 viral replication suggests the possibility that the spermatogenetic process may be detrimentally affected by the virus, and at the same time, supports the need to implement safety measures and guidelines to ensure specific care in reproductive medicine. [ABSTRACT FROM AUTHOR]

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may affect female reproductive health. Here, we investigated the potential of SARS-CoV-2 to infect the follicular microenvironment, in particular granulosa (GCs) and cumulus cells (CCs), thus providing evidence for a productive infection. GCs and CCs were recovered from women (n = 25) who underwent in vitro fertilization at the Assisted Reproductive Unit, Siena University Hospital. Follicular ovarian cells were co-cultured with SARS-CoV-2 and then analyzed by qPCR, immunofluorescence (IF), western blot (WB) and transmission electron microscopy (TEM). In addition, cell culture supernatant was used to infect VERO6 cells. We demonstrated the expression of cell host factors ACE2, TRPMSS2, BSG and CTSL, which are pivotal for the virus life cycle. Cultured GCs and CCs incubated with SARS-CoV-2 revealed productive SARS-CoV-2 infection at 24 h, 48 h and 72 h post-adsorption. Indeed, SARS-CoV-2 RNA, spike and nucleocapsid proteins were detected in GCs and CCs, and their cell culture supernatant successfully infected the standard VERO E6 cells. Finally, TEM showed full-size virions attached to the membrane and located inside the cytoplasm. This in vitro study reveals the susceptibility of human ovarian cells to SARS-CoV-2 infection, suggesting a potential detrimental effect of COVID-19 infection on female human fertility. [ABSTRACT FROM AUTHOR]

Water-soluble MBHA derivatives were found to self-assemble in a water environment to generate aggregates showing core–shell architectures. The aggregates appeared to be capable of working as nanoreactors performing a multi-functionalization of poly-histidine fragments, which after an initial interaction with the solvated oligo(ethylene glycol) shell reach the reactive core. [ABSTRACT FROM AUTHOR]

Intraflagellar transport (IFT) is responsible for the bidirectional trafficking of molecular components required for the elongation and maintenance of eukaryotic cilia and flagella. Cargo is transported by IFT 'trains', linear rows of multiprotein particles moved by molecular motors along the axonemal doublets. We have previously described two structurally distinct categories of 'long' and 'short' trains. Here, we analyse the relative number of these trains throughout flagellar regeneration and show that long trains are most abundant at the beginning of flagellar growth whereas short trains gradually increase in number as flagella elongate. These observations are incompatible with the previous hypothesis that short trains are derived solely from the reorganization of long trains at the flagellar tip. We demonstrate with electron tomography the existence of two distinct ultrastructural organizations for the short trains, we name these 'narrow' and 'wide', and provide the first 3D model of the narrow short trains. These trains are characterized by tri-lobed units, which repeat longitudinally every 16 nm and contact protofilament 7 of the B-tubule. Functional implications of the new structural evidence are discussed. [ABSTRACT FROM AUTHOR]

Background: Immune protection against pathogenic organisms has been shown to incur costs. Previous studies investigating the cost of immunity have mostly focused on the metabolic requirements of immune maintenance and activation. In addition to these metabolic costs, the immune system can induce damage to the host if the immune response is mis-targeted or over-expressed. Given its non-specific nature, an over-expressed inflammatory response is often associated with substantial damage for the host. Here, we investigated the cost of an over-expressed inflammatory response in the reproductive function of male mice. Methodology/Principal Findings: We experimentally blocked the receptors of an anti-inflammatory cytokine (IL-10) in male mice exposed to a mild inflammatory challenge, with each treatment having an appropriate control group. The experiment was conducted on two age classes, young (3 month old) and old (15 month old) mice, to assess any age-related difference in the cost of a disrupted immune regulation. We found that the concomitant exposure to an inflammatory insult and the blockade of IL-10 induced a reduction in testis mass, compared to the three other groups. The frequency of abnormal sperm morphology was also higher in the group of mice exposed to the inflammatory challenge but did not depend on the blockade of the IL-10. Conclusions: Our results provide evidence that immune regulation confers protection against the risk of inflammation-induced infertility during infection. They also suggest that disruption of the effectors involved in the regulation of the inflammatory response can have serious fitness consequences even under mild inflammatory insult and benign environmental conditions. [ABSTRACT FROM AUTHOR]

Background. Alkaptonuria, a rare autosomal recessive metabolic disorder caused by deficiency in homogentisate 1,2-dioxygenase activity, leads to accumulation of oxidised homogentisic acid in cartilage and collagenous structures present in all organs and tissues, especially joints and heart, causing a pigmentation called ochronosis. A secondary amyloidosis is associated with AKU. Here we report a study of an aortic valve froman AKU patient. Results. Congo Red birefringence,Th-T fluorescence, and biochemical assays demonstrated the presence of SAA-amyloid deposits in AKU stenotic aortic valve. Light and electron microscopy assessed the colocalization of ochronotic pigment and SAA-amyloid, the presence of calcified areas in the valve. Immunofluorescence detected lipid peroxidation of the tissue and lymphocyte/macrophage infiltration causing inflammation. High SAA plasma levels and proinflammatory cytokines levels comparable to those fromrheumatoid arthritis patients were found in AKU patient. Conclusions. SAA-amyloidosis was present in the aortic valve from an AKU patient and colocalized with ochronotic pigment as well as with tissue calcification, lipid oxidation,macrophages infiltration, cell death, and tissue degeneration. A local HGD expression in human cardiac tissue has also been ascertained suggesting a consequent local production of ochronotic pigment in AKU heart. [ABSTRACT FROM AUTHOR]

Abstract: During our researches on systematics and ecology of terrestrial isopods, carried out in western Sicily, some specimens showing a blue–purple coloration were collected; they belonged to four species: Armadillidium decorum Brandt, 1833, Trichoniscus panormidensis Montesanto et al., 2011, Philoscia affinis Verhoeff, 1908, Porcellio siculoccidentalis Viglianisi et al., 1992. We hypothesized that such coloration could be due, as reported in literature, to characteristic paracrystalline arrays of virions inside the tissues of blue colored specimens. Ultrastructural observations by transmission electron microscopy, on tissues of A. decorum, showed the presence of electron-dense viral particles, with a diameter of nearly 0.12μm. Dual-axis tomography, performed on specimens of A. decorum, evidenced an icosahedral structure of viral particles matching with that of Isopod Iridescent Virus (IIV). Molecular analysis, on 254bp portion of the major capsid protein (MCP) gene, allowed to place the virus into IIV-31 group, already known for other oniscidean species. The symptoms of infected individuals and the course of the disease were followed in laboratory, indicating similarities with other studies on Isopod Iridoviruses. Moreover, some notes on reproduction of infected ovigerous females are reported. Our data support unequivocal and direct evidences for the first case of IIV infection in terrestrial isopods reported in Italy. [Copyright &y& Elsevier]

Two novel benzofulvene monomers bearing propargyl or allyl groups have been synthesized by means of readily accessible reactions, and were found to polymerize spontaneously by solvent removal, in the apparent absence of catalysts or initiators, to give the corresponding polybenzofulvene derivatives bearing clickable propargyl or allyl moieties. The clickable propargyl and allyl groups were exploited in appropriate click reactions to develop a powerful and versatile 'grafting onto' synthetic methodology for obtaining tailored polymer brushes. [ABSTRACT FROM AUTHOR]

Morphological, ultrastructural, and molecular-sequence data were used to assess the phylogenetic position of a tetraflagellate green alga isolated from soil samples of a saline dry basin near F'derick, Mauritania. This alga can grow as individual cells or form non-coenobial colonies of up to 12 individuals. It has a parietal chloroplast with an embedded pyrenoid covered by a starch sheath and traversed by single parallel thylakoids, and an eyespot located in a parietal position opposite to the flagellar insertion. Lipid vacuoles are present in the cytoplasm. Microspectroscopy indicated the presence of chlorophylls a and b, with lutein as the major carotenoid in the chloroplast, while the eyespot spectrum has a shape typical of green-algal eyespots. The cell has four flagella, two of them long and two considerably shorter. Sequence data from the 18S r RNA gene and ITS2 were obtained and compared with published sequences for green algae. Results from morphological and ultrastructural examinations and sequence analysis support the placement of this alga in the Chlorophyceae, as Tetraflagellochloris mauritanica L. Barsanti et A. Barsanti, gen. et sp. nov. [ABSTRACT FROM AUTHOR]

Autism spectrum disorders (ASDs) are a complex group of neurodevelopment disorders steadily rising in frequency and treatment refractory, where the search for biological markers is of paramount importance. Although red blood cells (RBCs) membrane lipidomics and rheological variables have been reported to be altered, with some suggestions indicating an increased lipid peroxidation in the erythrocyte membrane, to date no information exists on how the oxidative membrane damage may affect cytoskeletal membrane proteins and, ultimately, RBCs shape in autism. Here, we investigated RBC morphology by scanning electron microscopy in patients with classical autism, that is, the predominant ASDs phenotype (age range: 6-26 years), nonautistic neurodevelopmental disorders (i.e., "positive controls"), and healthy controls (i.e., "negative controls"). A high percentage of altered RBCs shapes, predominantly elliptocytes, was observed in autistic patients, but not in both control groups. The RBCs altered morphology in autistic subjects was related to increased erythrocyte membrane F2-isoprostanes and 4-hydroxynonenal protein adducts. In addition, an oxidative damage of the erythrocyte membrane β-actin protein was evidenced. Therefore, the combination of erythrocyte shape abnormalities, erythrocyte membrane oxidative damage, and β-actin alterations constitutes a previously unrecognized triad in classical autism and provides new biological markers in the diagnostic workup of ASDs. [ABSTRACT FROM AUTHOR]

Aims: Atrial Natriuretic Peptide (ANP)-containing amyloid is frequently found in the elderly heart. No data exist regarding ANP aggregation process and its link to pathologies. Our aims were: i) to experimentally prove the presumptive association of Congestive Heart Failure (CHF) and Isolated Atrial Amyloidosis (IAA); ii) to characterize ANP aggregation, thereby elucidating IAA implication in the CHF pathogenesis. Methods and Results: A significant prevalence (85%) of IAA was immunohistochemically proven ex vivo in biopsies from CHF patients. We investigated in vitro (using Congo Red, Thioflavin T, SDS-PAGE, transmission electron microscopy, infrared spectroscopy) ANP fibrillogenesis, starting from &agr;-ANP as well as the ability of dimeric b-ANP to promote amyloid formation. Different conditions were adopted, including those reproducing &bgr;-ANP prevalence in CHF. Our results defined the uncommon rapidity of &agr;-ANP self-assembly at acidic pH supporting the hypothesis that such aggregates constitute the onset of a fibrillization process subsequently proceeding at physiological pH. Interestingly, CHF-like conditions induced the production of the most stable and time-resistant ANP fibrils suggesting that CHF affected people may be prone to develop IAA. Conclusions: We established a link between IAA and CHF by ex vivo examination and assessed that &bgr;-ANP is, in vitro, the seed of ANP fibrils. Our results indicate that &bgr;-ANP plays a crucial role in ANP amyloid deposition under physiopathological CHF conditions. Overall, our findings indicate that early IAA-related ANP deposition may occur in CHF and suggest that these latter patients should be monitored for the development of cardiac amyloidosis. [ABSTRACT FROM AUTHOR]

Abstract: Spermatogenesis and the sperm structure of the terebrantian Aeolothrips intermedius Bagnall are described. Spermatogenesis consists of two mitotic divisions; the second is characterized by the loss of half of the spermatids, which have pyknotic nuclei. Early spermatids have two centrioles, but when spermiogenesis starts, a third centriole is produced. The three basal bodies give rise to three flagella; later these fuse into a single flagellum which contains three 9+0 axonemes. The basal bodies are surrounded by a large amount of centriole adjunct material. During spermiogenesis this material contributes to the shifting of the three axonemes towards the anterior sperm region parallel to the elongating nucleus, and it is transformed into a dense cylinder. In the mature spermatids the three axonemes amalgamate to create a bundle of 27 doublet microtubules. Near the end of spermiogenesis the dense cylinder of the centriole adjunct lies parallel to the nucleus and the axonemes. It ends where the mitochondrion appears at half-sperm length. We confirm that Terebrantia testes have a single sperm cyst; their sperm are characterized by a cylindrical nucleus, three axonemes fused into one, a small mitochondrion and a short cylindrical centriole adjunct which corresponds to the dense body described in a previous work. The acrosome is lacking. At the midpoint of the anterior half of the sperm the outline of the cross-section is bilobed, with the nucleus contained in a pocket evagination of the plasma membrane. These characters are discussed in light of a comparison between Tubulifera and Terebrantia. [ABSTRACT FROM AUTHOR]

Poly- BF3a, a new hydrophobic polymer obtained by spontaneous polymerization of 1-methylene-3-phenyl-1 H-indene, was found to give nanoparticles characterized by favorable shape and dimensions. Poly- BF3a nanoparticles were loaded with CR3124, a potent 5HT3 antagonist, as a drug model by desolvation methods either in the absence or in the presence of polyethylene glycol (PEG1000) as a wetting agent. The SEM studies showed that the introduction of CR3124 into the preparation led to a variable degree of aggregation–cementation, which afforded a sort of nanocomposite material. In the absence of PEG1000, the drug molecule was found to stay in the amorphous state (DSC studies) when its percentage is not higher than 10% by weight. In vitro release experiments showed that the formation and stability of the dispersion as well as the drug release were remarkably affected by the presence of PEG1000, demonstrating its beneficial effect to the nanoparticle morphology and disaggregation. [ABSTRACT FROM AUTHOR]

Abstract: We present a strategy for the alignment of dual-axis tomographic series, based on reference points and simultaneous alignment of both series. Each series is first aligned individually, an affine transformation is determined to bring the two series in a unique reference system, and all experimental coordinates are combined in a single system of equations. In case of severe shrinkage, a global and a local refinement of the orientation parameters are performed to correct all minors misalignments. The strategy is illustrated on tomographic experiments performed on sections from plastic-embedded biological samples. The efficiency in correcting the misalignment of gold particles and in improving the quality of the reconstruction is documented both visually and quantitatively. In our approach every region of the tomogram is associated with its own orientation parameters and can be eventually reconstructed with the preferred algorithm. This is convenient in the computation of 3D averages of equivalent structures. A simulation experiment is presented to show that the performances of this approach are superior to those of the method of rotation in direct space. [Copyright &y& Elsevier]

Abstract: Sperm ultrastructure of several species in each of the two suborders of Thysanoptera Tubulifera and Terebrantia shows a distinctive and unusual architecture. Members of the whole order share a bizarre axoneme consisting of 27 microtubular elements derived from the amalgamation of 3 (9+0) axonemes present in each spermatid at the beginning of spermiogenesis. The reciprocal shifting of these axonemes along the length of the sperm, together with their possible shortening and overlapping for short distances, could explain why in some species it is never possible to observe the complete set of 27 microtubular elements in any one cross section. Tubuliferan sperm have a small elliptical (in cross section) acrosome extending the length of the sperm. In Bolothrips insularis and Compsothrips albosignatus this structure is larger and is associated with an external, flattened vesicle throughout its length. Terebrantian sperm lack an acrosome, but display for half their length a dense body running parallel to the nucleus. The sperm, in members of this suborder, are also characterized by possession of a small mitochondrion and by the unusual bilobed outline of cross sections through the anterior sperm region, with the nucleus located in one of the two lobes. Structures serving to anchor sperm to the inner surface of the cyst cell have been observed at their anterior tips in the testes of tubuliferans. In B. insularis, an anterior appendage is formed in immature sperm and is maintained in the mature spermatozoon parallel to its long axis in the most anterior region. Such an anchoring structure has not been observed in sperm of the terebrantian species examined, probably because the testis of terebrantians contains only a single cyst of developing gametes. [Copyright &y& Elsevier]

Placental viviparity is a reproductive strategy usually attributed to mammals. However, it is also present in other vertebrate species, e.g. in Squamate reptiles. Although the immunological mechanisms that allow the survival of the semi-allogenic embryo in maternal tissues are still largely unknown, cytokines seem to play an important role in mammalian reproduction. Previous studies in our laboratory showed that interleukin-1 (IL-1), a cytokine associated with implantation in mice, is also expressed at the materno-fetal interface of placental viviparous Squamates. In this study, we used the model of Lacerta vivipara, which exhibits reproductive bimodality, that is, the coexistence of oviparous and viviparous populations. By means of immunohistochemistry and anti-human antibodies, we showed that uterine tissues of L. vivipara (seven oviparous and six viviparous animals) expressed the two IL-1 isoforms, IL-1α and IL-1β, and the type I IL-1 receptor (IL-1R tI) both at the pre-ovulatory stage and during gestation, with no significant difference between oviparous and viviparous females. In L. vivipara, as in most oviparous Squamates, an important phase of embryonic development takes place in the mother's oviduct, before egg-laying. Moreover, although thinner than in oviparous females, an eggshell membrane persists throughout gestation in viviparous females also, which develop a very simple type of placenta. The data suggest that immunological mechanisms that allow the survival of the semi-allogenic embryo in maternal tissues are independent of the timing or intimacy of contact between maternal and fetal tissues. [ABSTRACT FROM AUTHOR]

In the longtime challenge of identifying specific, easily detectable and reliable biomarkers of IPF, BALF proteomics is providing interesting new insights into its pathogenesis. To the best of our knowledge, the present study is the first shotgun proteomic investigation of EVs isolated from BALF of IPF patients. Our main aim was to characterize the proteome of the vesicular component of BALF and to explore its individual impact on the pathogenesis of IPF. To this purpose, ultracentrifugation was chosen as the EVs isolation technique, and their purification was assessed by TEM, 2DE and LC-MS/MS. Our 2DE data and scatter plots showed considerable differences between the proteome of EVs and that of whole BALF and of its fluid component. Analysis of protein content and protein functions evidenced that EV proteins are predominantly involved in cytoskeleton remodeling, adenosine signaling, adrenergic signaling, C-peptide signaling and lipid metabolism. Our findings may suggest a wider system involvement in the disease pathogenesis and support the importance of pre-fractioning of complex samples, such as BALF, in order to let low-abundant proteins-mediated pathways emerge. [ABSTRACT FROM AUTHOR]

Marine diatoms have been identified among the most abundant taxa of microorganisms associated with plastic waste collected at sea. However, the impact of nano-sized plastic fragments (nanoplastics) at single cell and population level is almost unknown. We exposed the marine diatom Skeletonema marinoi to model polystyrene nanoparticles with carboxylic acid groups (PS–COOH NPs, 90 nm) for 15 days (1, 10, 50 μg/mL). Growth, reactive oxygen species (ROS) production, and nano-bio-interactions were investigated. No effect on diatom growth was observed, however Dynamic light scattering (DLS) demonstrated the formation of large PS aggregates which were localized at the diatoms' fultoportula process (FPP), as shown by TEM images. Increase production of ROS and reduction in chain length were also observed upon PS NPs exposure (p < 0.005). The observed PS-diatom interaction could have serious consequences on diatoms ecological role on the biogeochemical cycle of carbon, by impairing the formation of fast-sinking aggregates responsible for atmospheric carbon fixation and sequestration in the ocean sea floor. S. marinoi exposure to PS NPs caused an increase of intracellular and extracellular oxidative stress, the reduction of diatom's chain length and the adhesion of PS NPs onto the algal surface. Image 1 • PN NPs do not affect S. marinoi growth rate. • Fifteen days of exposure to PS NPs affect diatoms at cell and colony level. • PS NPs exposure resulted in a reduction of diatom's colony chain length. • PS NPs aggregates localized at diatom's fultoportula process. • PS NPs causes an increase in intracellular and extracellular ROS. [ABSTRACT FROM AUTHOR]

Embryo implantation has been defined as the "black box" of human reproduction. Most of the knowledge on mechanisms underlining this process derives from animal models, but they cannot always be translated to humans. Therefore, the development of an in vitro/ex vivo model recapitulating as closely and precisely as possible the fundamental functional features of the human endometrial tissue is very much desirable. Here, we have validated endometrial organoids as a suitable 3D-model to studying epithelial endometrial interface for embryo implantation. Transmission and scanning electron microscopy analyses showed that organoids preserve the glandular organization and cell ultrastructural characteristics. They also retain the responsiveness to hormonal treatment specific to the corresponding phase of the menstrual cycle, mimicking the in vivo glandular-like aspect and functions. Noteworthy, organoids mirroring the early secretive phase show the development of pinopodes, large cytoplasmic apical protrusions of the epithelial cells, traditionally considered as reliable key features of the implantation window. Moreover, organoids express glycodelin A (GdA), a cycle-dependent marker of the endometrial receptivity, with its quantitative and qualitative features accounting well for the profile detected in the endometrium in vivo. Accordingly, organoids deriving from the eutopic endometrium of women with endometriosis show a GdA glycosylation pattern significantly different from healthy organoids, confirming our prior data on endometrial tissues. The present results strongly support the idea that organoids may closely recapitulate the molecular and functional characteristics of their cells/tissue of origin. [ABSTRACT FROM AUTHOR]

A new polymer brush was synthesized by spontaneous polymerization of benzofulvene macromonomer 6-MOEG-9-T-BF3k bearing a nona(ethylene glycol) side chain linked to the 3-phenylindene scaffold by means of a triazole heterocycle. The polymer structure was studied by SEC-MALS, NMR spectroscopy, and MALDI-TOF MS techniques, and the results supported the role of oligomeric initiatory species in the spontaneous polymerization of polybenzofulvene derivatives. The aggregation features of high molecular weight poly-6-MOEG-9-T-BF3k-FE were investigated by pyrene fluorescence analysis, dynamic light scattering studies, and transmission electron microscopy, which suggested a tendency towards the formation of spherical objects showing dimensions in the range of 20–200 nm. Moreover, poly-6-MOEG-9-T-BF3k-FE showed an interesting cytocompatibility in the whole concentration range tested that, besides its aggregation features, makes this polybenzofulvene brush a good polymer candidate for nanoencapsulation and delivery of drug molecules. Finally, the photo-physical features of poly-6-MOEG-9-T-BF3k-FE could allow the biodistribution of the resulting drug delivery systems to be monitored by fluorescence microscopy techniques. [ABSTRACT FROM AUTHOR]

Despite the significant advances in the last decades, low implantation rate per transferred embryo still remains a major concern in assisted reproductive techniques, highlighting a need to better characterize endometrial receptivity also by mean of specific biomarkers. Based on physiology and on the intimate contact with endometrium as the tissue of interest, in this study we developed and validated an optimized protocol that uses extracellular vesicles (EVs) recovered from uterine flushings and from a cervical brush, the latter never used until now as an EVs source, as surrogates for endometrial biopsies. This method combines the safety of sampling with the ability to study the expression profile across the uterine cycle. We have compared the yield and composition of EVs recovered from different biofluids samples and fractions thereof, opting for chemical precipitation as the EV isolation procedure, assuring the highest yield without introducing any bias in specific EV recovery. Moreover, collected EVs, in particular exosome-like vesicles, express putative endometrial markers, such as glycodelin A and receptors for estrogen and progesterone, thus confirming their endometrial origin. We also identified uterine flushing EVs, in particular those recovered from its mucous fraction, as the richest source of endometrial transcripts, likely correlated to cellular (epithelial) origin of these vesicles. Finally, our pilot quantitative assessment of three endometrial gene profiles, in samples collected at different time points along the luteal phase, revealed the fluctuations apparently recapitulating gene expression variability prior reported during the menstrual cycle. Unlike tissue biopsy that is subjected to inter- and intra-sample differences, our data suggest that EVs from liquid biopsies (from uterine flushings and a cervical brush) obtained through less-invasive procedures, can be substrate to detect and track the tissue representative expression profiles, better depicting the total endometrium complexity. [ABSTRACT FROM AUTHOR]

Abstract: Background: Hypoxemia and increased oxidative stress (OS) have been reported in Rett Syndrome (RTT), a genetical neurodevelopmental disorder. Although OS and hypoxemia can lead to red blood cells (RBCs) shape abnormalities, no information on RBCs morphology in RTT exists. Here, RBCs shape was evaluated in RTT patients and healthy subjects as a function of OS markers, blood oxygenation, pulmonary gas exchange, and cardio-respiratory parameters. Methods: RBCs morphology was evaluated by Scanning Electron Microscopy. Intraerythrocyte and plasma non protein-bound iron (NPBI), esterified F2-Isoprostanes (F2-IsoPs), 4-HNE protein adducts (4-HNE PAs) were measured. Pulmonary oxygen gradients and PaO2 were evaluated by gas analyzers and cardiopulmonary variables by pulse oximetry. In RTT patients these parameters were assessed before and after ω-3 polyunsaturated fatty acids (ω-3 PUFAs) administration. Results: Altered RBCs shapes (leptocytes) and increased NPBI were present in RTT, together with increased erythrocyte membrane esterified F2-IsoPs and 4-HNE PAs. Abnormal erythrocyte shapes were related to OS markers levels, pulmonary gas exchange, PaO2 and cardio-respiratory variables. After ω-3 PUFAs, a decrease of leptocytes was accompanied by a progressive increase in reversible forms of RBCs. This partial RBCs morphology rescue was related to decreased OS damage markers, improved pulmonary oxygen exchange, and cardiopulmonary physiology. Conclusions: These findings indicate that in RTT 1) RBCs shape is altered; 2) the OS-hypoxia diad is critical in generating altered RBCs shape and membrane damage; 3) ω-3 PUFAs are able to partially rescue RBCs morphology and the OS-derived damage. General significance: RBCs morphology is an important biosensor for OS imbalance and chronic hypoxemia. [Copyright &y& Elsevier]

Abstract: Carboplatin elicits minor side effects with respect to its first generation analogue cisplatin. Nevertheless, a dose-dependent nephrotoxicity of the drug has been reported to occur both in patients and in rats and a possible pathogenic role have been attributed to oxidative stress. We have studied the effect of carboplatin administration on the thiol/disulfide balance, on other biomarkers of oxidative stress and on antioxidant enzymes in the isolated perfused rat kidney. A 5–500μM carboplatin dose range did not alter renal function but significantly decreased levels of cysteine, glutathione and exposed protein sulfhydryl groups. Only a minimal increment in disulfides was observed, whereas malonyldialdehyde and protein carbonyls did not increase significantly. Among the antioxidant enzymes studied, only thioltransferase was inhibited by the treatment. Our data suggest that a minimal oxidative stress is present under our experimental conditions, thus indicating that platinum-based drugs do not produce significant amount of reactive oxygen species. [Copyright &y& Elsevier]

SET-M33 is a multimeric antimicrobial peptide active against Gram-negative bacteria in vitro and in vivo . Insights into its killing mechanism could elucidate correlations with selectivity. SET-M33 showed concentration-dependent bactericidal activity against colistin-susceptible and resistant isolates of P. aeruginosa and K. pneumoniae. Scanning and transmission microscopy studies showed that SET-M33 generated cell blisters, blebs, membrane stacks and deep craters in K. pneumoniae and P. aeruginosa cells. NMR analysis and CD spectra in the presence of sodium dodecyl sulfate micelles showed a transition from an unstructured state to a stable α-helix, driving the peptide to arrange itself on the surface of micelles. SET-M33 kills Gram-negative bacteria after an initial interaction with bacterial LPS. The molecule becomes then embedded in the outer membrane surface, thereby impairing cell function. This activity of SET-M33, in contrast to other similar antimicrobial peptides such as colistin, does not generate resistant mutants after 24 h of exposure, non-specific interactions or toxicity against eukaryotic cell membranes, suggesting that SET-M33 is a promising new option for the treatment of Gram-negative antibiotic-resistant infections. [ABSTRACT FROM AUTHOR]

Abstract: Alkaptonuria (AKU) is an ultra-rare disease developed from the lack of homogentisic acid oxidase activity, causing homogentisic acid (HGA) accumulation that produces a HGA-melanin ochronotic pigment, of unknown composition. There is no therapy for AKU. Our aim was to verify if AKU implied a secondary amyloidosis. Congo Red, Thioflavin-T staining and TEM were performed to assess amyloid presence in AKU specimens (cartilage, synovia, periumbelical fat, salivary gland) and in HGA-treated human chondrocytes and cartilage. SAA and SAP deposition was examined using immunofluorescence and their levels were evaluated in the patients'' plasma by ELISA. 2D electrophoresis was undertaken in AKU cells to evaluate the levels of proteins involved in amyloidogenesis. AKU osteoarticular tissues contained SAA-amyloid in 7/7 patients. Ochronotic pigment and amyloid co-localized in AKU osteoarticular tissues. SAA and SAP composition of the deposits assessed secondary type of amyloidosis. High levels of SAA and SAP were found in AKU patients'' plasma. Systemic amyloidosis was assessed by Congo Red staining of patients'' abdominal fat and salivary gland. AKU is the second pathology after Parkinson''s disease where amyloid is associated with a form of melanin. Aberrant expression of proteins involved in amyloidogenesis has been found in AKU cells. Our findings on alkaptonuria as a novel type II AA amyloidosis open new important perspectives for its therapy, since methotrexate treatment proved to significantly reduce in vitro HGA-induced A-amyloid aggregates. [Copyright &y& Elsevier]

The physicochemical and biopharmaceutical properties, such as pKa, crystal habit, water solubility, logD, molecular structure and dynamics, and membrane permeability of CR3124 (endo-2-(8-methyl-8-azabicyclo[3.2.1]oct-3-yl)-2,3-dihydro-1H-benz[e]isoindol-1-one, a novel potent 5-HT3 receptor antagonist) have been studied in order to obtain preformulation information. The study showed that CR3124 is a very rigid molecule in which conformational freedom due to the presence of a rotatable bond is restricted by the interaction between an activated hydrogen and the amide oxygen and the conformation of the tropane piperidine ring is regulated by the environment in such a manner as to optimize the intermolecular interactions with the solvent. This chameleon behavior appears to be capable of explaining the biopharmaceutical properties showed by CR3124, such as low wettability, relatively good solubility, and very high membrane permeability. Copyright © 2006 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 95:2706–2721, 2006 [ABSTRACT FROM AUTHOR]