Amino-functionalized poly(l-lactide) lamellar single crystals as a valuable substrate for delivery of HPV16-E7 tumor antigen in vaccine development

Background: Poly(L-lactide) (PLLA) is a biodegradable polymer currently used in many biomedical applications, including the production of resorbable surgical devices, porous scaffolds for tissue engineering, nanoparticles and microparticles for the controlled release of drugs or antigens. The surfaces of lamellar PLLA single crystals (PLLAsc) were provided with amino groups by reaction with a multifunctional amine and used to adsorb an Escherichia coli-produced human papillomavirus (HPV)16-E7 protein to evaluate its possible use in antigen delivery for vaccine development. Methods: PLLA single crystals were made to react with tetraethylenepentamine to obtain amino-functionalized PLLA single crystals (APLLAsc). Pristine and amino-functionalized PLLAsc showed a two-dimensional microsized and one-dimensional nanosized lamellar morphology, with a lateral dimension of about 15–20 µm, a thickness of about 12 nm, and a surface specific area of about 130 m2/g. Both particles were characterized and loaded with HPV16-E7 before being administered to C57BL/6 mice for immunogenicity studies. The E7-specific humoral-mediated and cell-mediated immune response as well as tumor protective immunity were analyzed in mice challenged with TC-1 cancer cells. Results: Pristine and amino-functionalized PLLAsc adsorbed similar amounts of E7 protein, but in protein-release experiments E7-PLLAsc released a higher amount of protein than E7-APLLAsc. When the complexes were dried for observation by scanning electron microscopy, both samples showed a compact layer, but E7-APLLAsc showed greater roughness than E7-PLLAsc. Immunization experiments in mice showed that E7-APLLAsc induced a stronger E7-specific immune response when compared with E7-PLLAsc. Immunoglobulin G isotyping and interferon gamma analysis suggested a mixed Th1/Th2 immune response in both E7-PLLAsc-immunized and E7-APLLAsc-immunized mice. However, only the mice receiving E7-APLLAsc were fully protected from TC-1 tumor growth after three doses of vaccine. Conclusion: Our results show that APLLA single crystals improve the immunogenicity of HPV16-E7 and indicate that E7-APLLAsc could be used for development of an HPV16 therapeutic vaccine against HPV16-related tumors

Dynamics of the Sphere Model of Consciousness: Silence, Space, and Self

The Sphere Model of Consciousness (SMC) delineates a sphere-shaped matrix that aims to describe subjective experiences using geometric coordinates, in accordance with a neurophenomenological perspective. According to the SMC, an experience of overcoming the habitual self and the conditioning of memories could be placed at the center of the matrix, which can then be called the Place of Pre-Existence (PPE). The PPE is causally associated with self-determination. In this context, we suggest that silence could be considered an intentional inner environment enabling self-perception to focus on the “here and now,” which in turn improves perception of one’s own body in space. To investigate the hypotheses grounded in the model, the current preliminary study examined the Place of Pre-Existence Technique (PPEt), in which practitioners are guided to focus on a self-defined aim, reach a state of detachment from the habitual self, and envision the future. Four-hundred eighty-one volunteer PPEt practitioners completed self-report questionnaires before and after an intensive 3-day meditative training. We analyzed potential relationships between subjective experiences related to physical, emotional, mental, temporal, and spatial components, as well as self-determination and silence, before and after training. The results indicated a transition from a prevailing influence of mental dimension on the other aspects of experience, to a prevailing influence of the spatial dimension. Silence was reported more often following the training and was predominantly related to mental and emotional experiences. The results are discussed in the framework of the SMC, as compared to other models, and in relation to the shift from Narrative to Minimal Self and to increased balance among the considered dimensions

Identification of murine phosphodiesterase 5A isoforms and their functional characterization in HL-1 cardiac cell line

Phosphodiesterase 5A (PDE5A) specifically degrades the ubiquitous second messenger cGMP and experimental and clinical data highlight its important role in cardiac diseases. To address PDE5A role in cardiac physiology, three splice variants of the PDE5A were cloned for the first time from mouse cDNA library (mPde5a1, mPde5a2 and mPde5a3). The predicted amino acidic sequences of the three murine isoforms are different in the N-terminal regulatory domain. mPDE5A isoforms were transfected in HEK293T cells and they showed high affinity for cGMP and similar sensitivity to sildenafil inhibition. RT-PCR analysis showed that mPde5a1, mPde5a2 and mPde5a3 had differential tissue distribution. In the adult heart, mPde5a1 and mPde5a2 were expressed at different levels whereas mPde5a3 was undetectable. Overexpression of mPDE5As induced an increase of HL-1 number cells which progress into cell cycle. mPDE5A1 and mPDE5A3 overexpression increased the number of polyploid and binucleated cells, mPDE5A3 widened HL-1 areas and modulated hypertrophic markers more efficiently respect to the other mPDE5A isoforms. Moreover, mPDE5A isoforms had differential subcellular localization: mPDE5A1 was mainly localized in the cytoplasm, mPDE5A2 and mPDE5A3 were also nuclear localized. These results demonstrate for the first time the existence of three PDE5A isoforms in mouse and highlight their potential role in the induction of hypertrophy. This article is protected by copyright. All rights reserved

Campylobacter jejuni fatal sepsis in a patient with non-Hodgkin’s lymphoma: Case report and literature review of a difficult diagnosis

Campylobacter jejuni (C. jejuni) bacteremia is difficult to diagnose in individuals with hematological disorders undergoing chemotherapy. The cause can be attributed to the rarity of this infection, to the variable clinical presentation, and to the partial overlapping symptoms underlying the disease. Here, we report a case of a fatal sepsis caused by C. jejuni in a 76-year-old Caucasian man with non-Hodgkin's lymphoma. After chemotherapeutic treatment, the patient experienced fever associated with severe neutropenia and thrombocytopenia without hemodynamic instability, abdominal pain, and diarrhea. The slow growth of C. jejuni in the blood culture systems and the difficulty in identifying it with conventional biochemical phenotyping methods contributed to the delay of administering a targeted antimicrobial treatment, leading to a fatal outcome. Early recognition and timely intervention are critical for the successful management of C. jejuni infection. Symptoms may be difficult to recognize in immunocompromised patients undergoing chemotherapy. Thus, it is important to increase physician awareness regarding the clinical manifestations of C. jejuni to improve therapeutic efficacy. Moreover, the use of more aggressive empirical antimicrobial treatments with aminoglycosides and/or carbapenems should be considered in immunosuppressed patients, in comparison to those currently indicated in the guidelines for cancer-related infections supporting the use of cephalosporins as monotherapy

Antibiotics and pathogens, when the natural bacterial community resists!

The spread and persistence of antibiotic resistances (AR) in waters is a major threat for the environmental management, with direct risks for human health. Although antibiotic resistant bacteria (ARB) are found everywhere, their ecological success is related to anthropic impact, and to disturbed natural communities. In the same waters where AR can be a problem, other disturbances promoted by human activities can interfere with natural communities, e.g. the input of allochthonous (and potentially pathogenic) bacteria. The concomitant impact of those disturbances can have dramatic effects on the persistence and resistance of natural communities. With an experimental system on continuous cultures, we tested the impact on the microbial community from anthropized Lake Maggiore of enhanced tetracycline concentrations and of a subsequent invasion by a mixed population of E. coli. We measured the response in terms of bacterial and eukaryotic abundances, phenotypical distribution, bacterial community composition, spread and persistence of tetracycline resistance genes, and the fate of E. coli strains, by applying a number of techniques including flow-cytometry, microscopy, qPCR, and Illumina sequencing. We observed the preservation of AR genes, even when antibiotics were not in the system, suggesting a relatively small cost of maintenance of these genes, or their involvement in other genetic pathways. However, we did not see a general increase of e.g. tetA resistance genes, following the addition of antibiotics. In fact, it rather seemed that resistance to tetracycline was rather acquired by morphological changes and species interactions; we observed many more aggregated cells when the antibiotic was added. On the other hand, the invasion with E.coli seemed to generally promote the overall productivity of the systems without and with little antibiotic. Our results demonstrate that despite the heavy disturbance the natural microbial communities can develop a number of mechanisms of resistance in order to reduce the impact (e.g. aggregation), complicating the system modelling, thus reducing our forecasting possibilities

Insulin-like Growth Factor II mRNA-Binding Protein 1 Regulates Pancreatic Cancer Cell Growth through the Surveillance of CDC25A mRNA

: A number of data indicate that the sources of different kinds of PDAC may be discovered at the transcription/transduction stage. RNA metabolism is manipulated at various steps by different RNA-binding proteins (RBPs), and the deregulation or irregular activity of RBPs is known to contribute to tumor promotion and progression. The insulin-like growth factor 2 mRNA-binding protein family (IMPs), and IMP1 in particular, has been linked with a poor prognosis in PDAC patients; however, little is known about its contribution in PDAC carcinogenesis. In this study, we investigated the function of IMP1 in PDAC. To evaluate IMP1 expression and correlation with PDAC prognosis, we utilized several public databases. Using a specific siRNA IMP1, we analyzed cell death and cell cycle progression in PDAC cell lines and 3D spheroids. the role of IMP1 was also evaluated in vivo in a panc-1-derived tumor xenograft murine model. Public data suggest that PDAC patients with higher expression of IMP1 showed poor overall and progression-free survival. IMP1 silencing leads to reduced cell growth in PDAC cells and three-dimensional spheroids. Abrogation of IMP1 in PDAC cells showed lower levels of CDC25A, increased phosphorylation of the cyclin-dependent kinase (CDK)2, and accumulation of PDAC cells in the G1 phase. immunoprecipitation experiments revealed that IMP1 binds CDC25A mRNA, thus controlling cell-cycle progression. Ultimately, we proved that suppression of IMP1 blocked in vivo growth of Panc-1 transferred into immunodeficient mice. Our results indicate that IMP1 drives the PDCA cell cycle and represents a novel strategy for overcoming PDCA cell proliferation

Androgen modulation of pro-inflammatory and antiinflammatory cytokines during preadipocyte differentiation

Background: Macrophages and adipocytes contribute to release of cytokines resulting in the chronic inflammatory profile of the metabolic syndrome. The local increase of proinflammatory cytokines impairs adipogenesis, resulting in formation of dysfunctional adipocytes that are unable to store and handle lipids. The altered lipid fluxes in/from adipocytes affect whole-body metabolism. We investigated the role of androgens on adipocyte-derived proinflammatory and anti-inflammatory cytokines during preadipocyte differentiation. Materials and methods: Various differentiation methods were used to obtain full conversion of 3T3-L1 into mature adipocytes. The degree of adipocyte conversion in the presence/absence of dihydrotestosterone (DHT) was analyzed by measuring intracellular triglycerides (Oil Red O staining). The effects of DHT administration on interleukin 1Β (IL-1Β), IL-2, IL-6, IL-10, IL-12, interferon γ (IFNΓ) and tumor necrosis factor α (TNFα) secretion was measured at days 0, 4, 6 and 8 of differentiation using the SearchLight multiplex protein array. Results: DHT regulates a number of cytokines in committed and mature 3T3-L1 adipocytes. IL-1Β and TNFα were readily suppressed at the very early stages of differentiation. IFNΓ release was inhibited at day 4, but the effect was no longer detectable on day 8. IL-6 and IL-12 were significantly reduced at day 8 of differentiation. Conversely, the differentiation-dependent increase of IL-2 and IL-10 was further stimulated by DHT since day 0. Conclusions: We provide evidence that androgens promote an anti-inflammatory profile that parallels the acquisition of a functional adipocyte phenotype. The crosstalk between androgens, adipocyte-derived mediators of inflammation and intracellular lipid fluxes could have profound implications on metabolism of men with obesity and metabolic syndrome. © 2010, by Walter de Gruyter Berlin New York. All rights reserved

Interplay between Tectonics and Mount Etna’s Volcanism: Insights into the Geometry of the Plumbing System

Mt. Etna lies in front of the southeast-verging Apennine-Maghrebian fold-and-thrust belt, where the NNW-trending Malta Escarpment separates the Sicilian continental crust from the Ionian Mesozoic oceanic basin, presently subducting beneath the Calabrian arc (Selvaggi and Chiarabba, 1995). Seismic tomographic studies indicate the presence of a mantle plume beneath the volcano with a Moho transition at depth less than 20 km (Nicolich et al.,2000; Barberi et al., 2006). Geophysical and geological evidences suggest that the Mt. Etna magma ascent mechanism is related to the major NNW-trending lithospheric fault (Doglioni et al., 2001). However, the reason for the Mt. Etna mantle plume draining and channeling the magma from the upper mantle source to the surface is not yet clear. All models proposed in literature (Rittmann, 1973; Tanguy et al., 1997; Monaco et al.; 1997; Gvirtzman and Nur, 1999; Doglioni et al., 2001) do not explain why such a mantle plume has originated in this anomalous external position with respect to the arc magmatism and back-arc spreading zones associated with the Apennines subduction. Some ideas on the subduction rollback must be better developed through the comparison with new regional tomographic studies that are being released. Moreover, tomographic studies reveal a complex and large plumbing system below the volcano from -2 to -7 km a.s.l., wide up to 60 km2 that reduces itself in size down to -18 km of depth close to the apex of the mantle plume. Chiocci et al. (2011) found a large bulge on the underwater continental margin facing Mt. Etna, and suggested that the huge crystallized magma body intruded in the middle and upper continental crust was able to trigger an instability process involving the Sicilian continental margin during the last 0.1 Ma. This phenomenon induces the sliding of the volcano eastern flank observed since the 90s (Borgia et al, 1992; Lo Giudice and Rasà, 1992) because the effects of the bulge collapse are propagating upslope, and the continuous decompression at the volcano summit favors the ascent of basic magma without lengthy storage in the upper crust, as one might expect in a compressive tectonic regime. Taken together, these new evidences (tomographic, tectonic, volcanic) are concerned with the exceptional nature of Mt. Etna and raise the need to explain the origin of the mantle plume that supplies its volcanism. The lower crust and the uppermost mantle need to be better resolved in future experiments and studies. The use of regional and teleseismic events for tomography and receiver function analyses is required to explore a volume that has only marginally been investigated to date. The relation between the magma source in the mantle and the upper parts of the system, as well as the hypothesis above reported on the relation between tectonics and volcanism and the role of lithospheric faults, could be resolved only by applying seismological techniques able to better constrain broader and deeper models. Finally, although the recent tomographic inversions have progressively improved our knowledge of Etna’s shallow structure, highlighting a complex pattern of magma chambers and conduits with variable dimensions, the geometry of the conduits and the dimensions and shapes of small magmatic bodies still require greater investigation. Their precise definition is crucial to delineate a working model of this volcano in order to understand its behaviour and evolution. For this purpose, at least within the volcanic edifice, the precise locations of the seismo-volcanic signals can be considered a useful tool to constrain both the area and the depth range of magma degassing and the geometry of the shallow conduits. In this work, we furnish evidences that the tremor and LP locations allowed to track magma migration during the initial phase of the 2008-2009 eruption and in particular the initial northward dike intrusion, also confirmed by other geophysical, structural and volcanological observations (Aloisi et al., 2009; Bonaccorso et al., 2011), and the following fissure opening east of the summit area at the base of SEC. All these evidences, obtained by the marked improvement in the monitoring system together with the development of new processing techniques, allowed us to constrain both the area and the depth range of magma degassing, highlighting the geometry of the magmatic system feeding the 2008-2009 eruption

Effects of endocrine disruptors on fetal testis development, male puberty, and transition age

Purpose Endocrine disruptors (EDs) are exogenous substances able to impair endocrine system; consequently, they may cause numerous adverse effects. Over the last years, particular focus has been given to their harmful effects on reproductive system, but very little is known, especially in males. The aim of this review is to discuss the detrimental effects of EDs exposure on fetal testis development, male puberty, and transition age. Methods A search for the existing literature focusing on the impact of EDs on fetal testis development, male puberty, andrological parameters (anogenital distance, penile length, and testicular volume), and testicular cancer with particular regard to pubertal age provided the most current information available for this review. Human evidence-based reports were given priority over animal and in vitro experimental results. Given the paucity of available articles on this subject, all resources were given careful consideration. Results Information about the consequences associated with EDs exposure in the current literature is limited and often conflicting, due to the scarcity of human studies and their heterogeneity. Conclusions We conclude that current evidence does not clarify the impact of EDs on human male reproductive health, although severe harmful effects had been reported in animals. Despite controversial results, overall conclusion points toward a positive association between exposure to EDs and reproductive system damage. Further long-term studies performed on wide number of subjects are necessary in order to identify damaging compounds and remove them from the environment

Procalcitonin as a marker of Candida species detection by blood culture and polymerase chain reaction in septic patients

Background: The aim of our study is to test procalcitonin (PCT) as surrogate marker of identification of Candida spp. by blood culture (BC) and real-time-polymerase chain reaction (PCR), whether alone or in association with bacteria, in septic patients. Methods: We performed a single-centre retrospective study. We reviewed the clinical charts of patients with a diagnosis of severe sepsis or septic shock treated at our general intensive care unit from March 2009 to March 2013. We analysed all diagnostic episodes consisting of BC, real-time PCR assay and dosage of PCT. We registered age, sex, white blood count, sequential organ failure assessment score and type of admission between medical or surgical. When inclusion criteria were met more than once, we registered the new diagnostic episode as subsequent diagnostic episode. The diagnostic performance of PCT to predict Candida spp. identification alone or in mixed infections by either BC or PCR was tested using the receiver-operative characteristic curve. Logistic regression was constructed using presence of Candida spp. as the dependent variable. Results: A total of 260 diagnostic episodes met the inclusion criteria. According to BC results classification, a significantly lower value of PCT was observed in Candida spp. BSI (0.99 ng/ml, 0.86 - 1.34) than in BSI caused by bacteria (16.7 ng/ml, 7.65 - 50.2) or in mixed infections (4.76 ng/ml, 2.98 - 6.08). Similar findings were observed considering PCR results. A cut-off of ≤ 6.08 ng/ml for PCT yielded a sensitivity of 86.8%, a specificity of 87.4%, a positive predictive value of 63.9%, a negative predictive value (NPV) of 96.3% and an area under the curve of 0.93 for Candida spp. identification by BC. A similar high NPV for a cut-off ≤ 6.78 ng/ml was observed considering the classification of diagnostic episodes according to PCR results, with an AUC of 0.85. A subsequent diagnostic episode was independently associated with Candida spp. detection either by BC or PCR. Conclusion: PCT could represent a useful diagnostic tool to exclude the detection of Candida spp. by BC and PCR in septic patients