EBV reduces autophagy, intracellular ROS and mitochondria to impair monocyte survival and differentiation.

EBV has been reported to impair monocyte in vitro differentiation into dendritic cells (DCs) and reduce cell survival. In this study, we added another layer of knowledge to this topic and showed that these effects correlated with macroautophagy/autophagy, ROS and mitochondrial biogenesis reduction. Of note, autophagy and ROS, although strongly interconnected, have been separately reported to be induced by CSF2/GM-CSF (colony stimulating factor 2) and required for CSF2-IL4-driven monocyte in vitro differentiation into DCs. We show that EBV infects monocytes and initiates a feedback loop in which, by inhibiting autophagy, reduces ROS and through ROS reduction negatively influences autophagy. Mechanistically, autophagy reduction correlated with the downregulation of RAB7 and ATG5 expression and STAT3 activation, leading to the accumulation of SQSTM1/p62. The latter activated the SQSTM1-KEAP1- NFE2L2 axis and upregulated the anti-oxidant response, reducing ROS and further inhibiting autophagy. ROS decrease correlated also with the reduction of mitochondria, the main source of intracellular ROS, achieved by the downregulation of NRF1 and TFAM, mitochondrial biogenesis transcription factors. Interestingly, mitochondria supply membranes and ATP required for autophagy execution, thus their reduction may further reduce autophagy in EBV-infected monocytes. In conclusion, this study shows for the first time that the interconnected reduction of autophagy, intracellular ROS and mitochondria mediated by EBV switches monocyte differentiation into apoptosis, giving new insights into the mechanisms through which this virus reduces immune surveillance. Abbreviations: ACTB: actin beta; ATG5: autophagy related 5; BAF: bafilomycin A1; BECN1: beclin 1; CAT: catalase; CSF2: colony stimulating factor 2; CT: control; CYCS (cytochrome C: somatic); DCs: dendritic cells; EBV: Epstein-Barr virus; GSR: glutathione-disulfide reductase; KEAP1: kelch like ECH associated protein 1; IL4: interleukin 4; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MET: metformin; NAC: N-acetylcysteine; NFE2L2/NRF2 nuclear factor: erythroid 2 like 2; NRF1 (nuclear respiratory factor 1); clPARP1: cleaved poly(ADP-ribose) polymerase; Rapa: Rapamycin; ROS: reactive oxygen species; SQSTM1/p62: sequestosome 1; TFAM: (transcription factor A: mitochondrial); TUBA1A: tubulin alpha 1a.

Three-point bending test simulation on implant fpds with a bio-faithful model.

AIM OF THE STUDY: It is well known by previous important studies that mandible flexes during different jaw movements. According to this assumption it is very important to know how implant supported fixed partial dentures could restrict mandibular movements and, could lead to excess strain accumulation that could modify the resolution of implant treatment. The aim of our project is to create a bio-faithful model able to recreate mandibular movements, during three point bending test methods of (FIXED -PARTIAL -DENTURES) FPDs, to avoid a not flexible metal base, where models' properties doesn't allow to obtain a bio-faithful simulation during testing phases. MATERIALS AND METHODS: 2 implants (premium Sweden and Martina®) were embedded in mandible resin section to mimic osteointegrated implants in premolar and molar areas, in order to recreate a Kennedy Class II configuration. Our mandible test simulator was creating according to the measurement obtained according to the study of Schwartz-Dabney and Dechow (2002). Sample so created is tested with testing machine (Instron 5566®, UK) adopting the three point bending mechanical tests configuration. DISCUSSION AND CONCLUSION: We can admit that oral cavity is a bio-dynamic system, where different variables incurr, so it's very important that experimental conditions simulate clinical environment. Experimentation should be based on the correlation between the failure mechanisms exhibited for in vitro samples and those observed in fractured clinical prostheses made of the same composition and processing conditions. A bio-faithful model could reduce this wide range between in vitro and in vivo study experimentation.

KSHV reduces autophagy in THP-1 cells and in differentiating monocytes by decreasing CAST/calpastatin and ATG5 expression.

We have previously shown that Kaposi sarcoma-associated herpesvirus (KSHV) impairs monocyte differentiation into dendritic cells (DCs). Macroautophagy/autophagy has been reported to be essential in such a differentiating process. Here we extended these studies and found that the impairment of DC formation by KSHV occurs through autophagy inhibition. KSHV indeed reduces CAST (calpastatin) and consequently decreases ATG5 expression in both THP-1 monocytoid cells and primary monocytes. We unveiled a new mechanism put in place by KSHV to escape from immune control. The discovery of viral immune suppressive strategies that contribute to the onset and progression of viral-associated malignancies is of fundamental importance for finding new therapeutic approaches against them.

T- and B-cell responses and previous exposure to hepatitis B virus in 'anti-HBc alone' patients.

A serologic response to hepatitis B virus (HBV) defined as 'anti-HBc alone' is commonly observed, but its significance remains unclear. This study aimed to define the relationship between 'anti-HBc alone' serostatus and HBV infection, including HBV-specific T- and B-cell memory responses. We enrolled 31 'anti-HBc alone' patients. Total HBV DNA and cccDNA were tested by nested polymerase chain reaction (PCR) analysis in liver samples from 22 'anti-HBc alone' patients vs controls (chronic or resolved HBV infection), followed by HBsAg/HBcAg immunohistochemical (IHC) staining. IFN-γ secretion by HBV-specific T cells was compared in individuals who were 'anti-HBc alone' (n = 27), resolved HBV (n = 21), chronic HBV (n = 24) and 12 healthy controls using enzyme-linked immunospot (ELISpot) assays. An HBsAg-IgG B-cell ELISpot assay was performed in 'anti-HBc alone' patients before and after one dose of recombinant HBsAg vaccine. The majority (23/31, 74.2%) of the 'anti-HBc alone' individuals were co-infected with HCV. Infrequent intrahepatic total HBV DNA (2/22, 9.1%) and cccDNA (1/22, 4.5%) were detected in biopsies; HBsAg and HBcAg IHC staining was negative. HBV-specific T-cell responses were similar between 'anti-HBc alone' individuals and HBV resolvers. Circulating HBV-memory B-cell responses were detected in all 'anti-HBc alone' individuals, consistent with an HBsAg-specific memory pool. After one HBV vaccine dose, increased anti-HBs antibody levels were observed, accompanied by an expansion of HBsAg-specific memory B cells (P = 0.0226). 'Anti-HBc alone' individuals showed HBV-specific T-cell and memory B-cell responses typical of previous viral exposure and protective memory, suggesting a resolved infection.

Comprehensive validation of published immunohistochemical prognostic biomarkers of prostate cancer -what has gone wrong? A blueprint for the way forward in biomarker studies.

BACKGROUND: Treatment planning of localised prostate cancer remains challenging. Besides conventional parameters, a wealth of prognostic biomarkers has been proposed so far. None of which, however, have successfully been implemented in a routine setting so far. The aim of our study was to systematically verify a set of published prognostic markers for prostate cancer. METHODS: Following an in-depth PubMed search, 28 markers were selected that have been proposed as multivariate prognostic markers for primary prostate cancer. Their prognostic validity was examined in a radical prostatectomy cohort of 238 patients with a median follow-up of 60 months and biochemical progression as endpoint of the analysis. Immunohistochemical evaluation was performed using previously published cut-off values, but allowing for optimisation if necessary. Univariate and multivariate Cox regression were used to determine the prognostic value of biomarkers included in this study. RESULTS: Despite the application of various cut-offs in the analysis, only four (14%) markers were verified as independently prognostic (AKT1, stromal AR, EZH2, and PSMA) for PSA relapse following radical prostatectomy. CONCLUSIONS: Apparently, many immunohistochemistry-based studies on prognostic markers seem to be over-optimistic. Codes of best practice, such as the REMARK guidelines, may facilitate the performance of conclusive and transparent future studies.

Sem analysis zirconia-ceramic adhesion interface.

OBJECTIVES: Modern dentistry increasingly tends to use materials aesthetically acceptable and biomimetic. Among these are zirconia and ceramics for several years, a combination that now has becoming synonym of aesthetic; however, what could be the real link between these two materials and especially its nature, remains a controversial topic debated in the literature. The aim of our study was to "underline" the type of bonding that could exist between these materials. MATERIALS AND METHODS: To investigate the nature of this bond we used a SEM microscopy (Zeiss SUPRA 25). Different bilaminar specimens: "white" zirconia Zircodent® and ceramic "Noritake®", after being tested with loading test in bending (three-point-bending) and FEM analysis, were analyzed by SEM. Fragments' analysis in closeness of the fracture's point has allowed us to be able to "see" if at large magnifications between these two materials, and without the use of linear, could exist a lasting bond and the possible type of failure that could incur. RESULTS: From our analysis of the specimens' fragments analyzed after test Equipment, it is difficult to highlight a clear margin and no-adhesion zones between the two materials, although the analysis involving fragments adjacent to the fracture that has taken place at the time of Mechanical test Equipment. CONCLUSIONS: According to our analysis and with all the clarification of the case, we can assume that you can obtain a long and lasting bond between the zirconia and ceramics. Agree to the data present in the literature, we can say that the type of bond varies according to the type of specimens and of course also the type of failure. In samples where the superstructure envelops the ceramic framework Zirconium we are in the presence of a cohesive failure, otherwise in a presence of adhesive failure.

Computed tomography findings in liver fibrosis and cirrhosis.

PRINCIPLES: Computed tomography (CT) is inferior to the fibroscan and laboratory testing in the noninvasive diagnosis of liver fibrosis. On the other hand, CT is a frequently used diagnostic tool in modern medicine. The auxiliary finding of clinically occult liver fibrosis in CT scans could result in an earlier diagnosis. The aim of this study was to analyse quantifiable direct signs of liver remodelling in CT scans to depict liver fibrosis in a precirrhotic stage. METHODS: Retrospective review of 148 abdominal CT scans (80 liver cirrhosis, 35 precirrhotic fibrosis and 33 control patients). Fibrosis and cirrhosis were histologically proven. The diameters of the three main hepatic veins were measured 1-2 cm before their aperture into the inferior caval vein. The width of the caudate and the right hepatic lobe were divided, and measured horizontally at the level of the first bifurcation of the right portal vein in axial planes (caudate-right-lobe ratio). A combination of both (sum of liver vein diameters divided by the caudate-right lobe ratio) was defined as the ld/crl ratio. These metrics were analysed for the detection of liver fibrosis and cirrhosis. RESULTS: An ld/crl-r <24 showed a sensitivity of 83% and a specificity of 76% for precirrhotic liver fibrosis. Liver cirrhosis could be detected with a sensitivity of 88% and a specificity of 82% if ld/crl-r <20. CONCLUSION: An ld/crl-r <24 justifies laboratory testing and a fibroscan. This could bring forward the diagnosis and patients would profit from early treatment in a potentially reversible stage of disease.

Mechanistic evaluation of the transfection barriers involved in lipid-mediated gene delivery: interplay between nanostructure and composition.

Here we present a quantitative mechanism-based investigation aimed at comparing the cell uptake, intracellular trafficking, endosomal escape and final fate of lipoplexes and lipid-protamine/deoxyribonucleic acid (DNA) (LPD) nanoparticles (NPs) in living Chinese hamster ovary (CHO) cells. As a model, two lipid formulations were used for comparison. The first formulation is made of the cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and the zwitterionic lipid dioleoylphosphocholine (DOPC), while the second mixture is made of the cationic 3ß-[N-(N,N-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and the zwitterionic helper lipid dioleoylphosphatidylethanolamine (DOPE). Our findings indicate that lipoplexes are efficiently taken up through fluid-phase macropinocytosis, while a less efficient uptake of LPD NPs occurs through a combination of both macropinocytosis and clathrin-dependent pathways. Inside the cell, both lipoplexes and LPD NPs are actively transported towards the cell nucleus, as quantitatively addressed by spatio-temporal image correlation spectroscopy (STICS). For each lipid formulation, LPD NPs escape from endosomes more efficiently than lipoplexes. When cells were treated with DOTAP-DOPC-containing systems the majority of the DNA was trapped in the lysosome compartment, suggesting that extensive lysosomal degradation was the rate-limiting factors in DOTAP-DOPC-mediated transfection. On the other side, escape from endosomes is large for DC-Chol-DOPE-containing systems most likely due to DOPE and cholesterol-like molecules, which are able to destabilize the endosomal membrane. The lipid-dependent and structure-dependent enhancement of transfection activity suggests that DNA is delivered to the nucleus synergistically: the process requires both the membrane-fusogenic activity of the nanocarrier envelope and the employment of lipid species with intrinsic endosomal rupture ability.

Palatal obturators in patients after maxillectomy.

Prosthodontic management of palatal defects is fundamental to improve patient's life undergoing to a maxillary surgical treatment. A lot of maxillary defects are a direct consequence of surgical treatment of malformations, neoplasms or trauma. The obturators are prosthesis used to close palatal defects after maxillectomy, to restore masticatory function and to improve speech. The primary goals of the obturator prosthesis are to preserve the remaining teeth and tissue and to provide comfort, function, and aesthetics to the patients. Different materials and retention methods are a characteristic of new types of obturators.

Mechanistic understanding of gene delivery mediated by highly efficient multicomponent envelope-type nanoparticle systems.

We packaged condensed DNA/protamine particles in multicomponent envelope-type nanoparticle systems (MENS) combining different molar fractions of the cationic lipids 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and 3ß-[N-(N,N-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and the zwitterionic lipids dioleoylphosphocholine (DOPC) and dioleoylphosphatidylethanolamine (DOPE). Dynamic light scattering (DLS) and microelectrophoresis allowed us to identify the cationic lipid/DNA charge ratio at which MENS are small sized and positively charged, while synchrotron small-angle X-ray scattering (SAXS) and atomic force microscopy (AFM) revealed that MENS are well-shaped DNA/protamine particles covered by a lipid monobilayer. Transfection efficiency (TE) experiments indicate that a nanoparticle formulation, termed MENS-3, was not cytotoxic and highly efficient to transfect Chinese hamster ovary (CHO) cells. To rationalize TE, we performed a quantitative investigation of cell uptake, intracellular trafficking, endosomal escape, and final fate by laser scanning confocal microscopy (LSCM). We found that fluid-phase macropinocytosis is the only endocytosis pathway used by MENS-3. Once taken up by the cell, complexes that are actively transported by microtubules frequently fuse with lysosomes, while purely diffusing systems do not. Indeed, spatiotemporal image correlation spectroscopy (STICS) clarified that MENS-3 mostly exploit diffusion to move in the cytosol of CHO cells, thus explaining the high TE levels observed. Also, MENS-3 exhibited a marked endosomal rupture ability resulting in extraordinary DNA release. The lipid-dependent and structure-dependent TE boost suggests that efficient transfection requires both the membrane-fusogenic activity of the nanocarrier envelope and the employment of lipid species with intrinsic endosomal rupture ability.

Clinical assessment of submerged vs non-submerged implants placed in pristine bone.

The medium-long term success of osseointegrated dental implants is evaluated on the basis of the degree of osseointegration over time, assessed by radiographic or instrumental analysis (ISQ). Over the years, the question has always been which surgical technique can provide a better performance in the medium-long term and, thanks to literature studies, it has been evidenced that there are no differences between "one stage" and "two stage" interventions. The purpose of this study is to evaluate clinical and radiographic parameters, referring to interventions for the insertion of dental implants characterized by a new kind of implant surface (Synthegra® GEASS, Udine). The prospective study, not randomized and controlled, referred to the insertion of 18 implants on 9 patients with mono or bilateral edentulism, with measurements at 1, 3, 6 and 12 months and an overall follow-up at 3 years, in order to evaluate the different degree of crestal bone resorption using the submerged and transmucosal surgical technique. The results of our study show that there are no differences in the resorption of the two surgical techniques, with an average bone resorption of 2,05±0,16 mm, comparable with values reported in literature.

Mechanical evaluation and fem analysis of stress in fixed partial dentures zirconium-ceramic.

OBJECTIVE: Over the last several years, the Finite Element Analysis (FEM) has been widely recognized as a reference method in different fields of study, to simulate the distribution of mechanical stress, in order to evaluate the relative distribution of loads of different nature. The aim of this study is to investigate through the FEM analysis the stress distribution in fixed prostheses that have a core in Zirconia and a ceramic veneer supported by implants. MATERIALS AND METHODS: In this work we investigated the mechanical flexural strength of a ceramic material (Noritake(®)) and a of zirconium framework (Zircodent(®)) and the effects of the manufacturing processes of the material commonly performed during the production of fixed prostheses with CAD/CAM technology. Specifically three point bending mechanical tests were performed (three-point-bending) (1-3), using a machine from Test Equipment Instron 5566(®), on two structures in zirconium framework-ceramic (structures supported by two implant abutments with pontic elements 1 and 2). A further in-depth analysis on the mechanical behavior in flexure of the specimens was conducted carrying out FEM studies in order to compare analog and digital data. RESULTS: The analysis of the data obtained showed that the stresses are distributed in a different way according to the intrinsic elasticity of the structure. The analysis of FPD with four elements, the stresses are mainly concentrated on the surface of the load, while, in the FPD of three elements, much more rigid, the stresses are concentrated near the inner margins of the abutments. The concentration of many stresses in this point could be correlated to chipping (4) that is found in the outer edges of the structure, as a direct result of the ceramic brittleness which opposes the resilience of the structure subjected to bending. CONCLUSIONS: The analysis of the UY linear displacement confirms previous data, showing, in a numerical way, that the presence of the ceramic is related to the lowering of the structure. So, the reference values are those of the linear lowering obtained in the Mechanical Test and in our FEM analysis. zirconium framework with four elements 4,227 10(-2)mm.zirconium framework with ceramic structure with four elements 2,266 10(-2) mm.That suggests that the presence of ceramics halves the flexion capabilities of the prosthetic materials.

Role of temperature-independent lipoplex-cell membrane interactions in the efficiency boost of multicomponent lipoplexes.

Multicomponent lipoplexes have recently emerged as especially promising transfection candidates, as they are from 10 to 100 times more efficient than binary complexes usually employed for gene delivery purposes. Previously, we investigated a number of chemical-physical properties of DNA-lipid complexes that were proposed to affect transfection efficiency (TE) of lipoplexes, such as nanoscale structure, size, surface potential, DNA-protection ability and DNA release from complexes upon interaction with cellular lipids. Although some minor differences between multicomponent and binary lipoplexes were found, they did not correlate clearly with efficiency. Instead, here we show that a marked difference between the cell internalization mechanism of binary and multicomponent lipoplexes does exist. Multicomponent lipoplexes significantly transfect cells at 4 °C, when endocytosis does not take place suggesting that they can enter cells via a temperature-independent mechanism. Confocal fluorescence microscopy experiments showed the existence of a correlation between endosomal escape and TE. Multicomponent lipoplexes exhibited a distinctive ability of endosomal escape and release DNA into the nucleus, whereas, poorly efficient binary lipoplexes exhibited minor, if any, endosomal rupture ability and remained confined in perinuclear late endosomes. Stopped-flow mixing measurements showed that the fusion rates of multicomponent cationic liposomes with anionic vesicles, used as model systems of cell membranes, were definitely shorter than those of binary liposomes. As either lipoplex uptake and endosomal escape involve fusion between lipoplex and cellular membranes, we suggest that a mechanism of lipoplex-cellular membrane interaction, driven by lipid mixing between cationic and anionic cellular lipids, does explain the TE boost of multicomponent lipoplexes.

Tumour-infiltrating lymphocytes and survival in patients with adenocarcinoma of the oesophagus.

INTRODUCTION: Tumor-infiltrating lymphocytes (TILs) and forkhead box transcription factor positive (FoxP3(+)) regulatory T-lymphocytes (TREGs) have been analyzed in a variety of tumors but not in oesophageal adenocarcinoma. PATIENTS AND METHODS: Tissue from 130 adenocarcinomas of the oesophagus was re-evaluated in the centre and periphery of tumour, respectively, using immunohistochemical staining with anti-CD3, anti-CD4, anti-CD8, anti-CD25 and anti-FoxP3 antibodies. Patients were stratified according neoadjuvant treatment. 106 patients proceeded directly to surgery and 24 underwent pre-operative radio-chemotherapy (RCT). RESULTS: In patients without RCT, TILs were found significantly more frequently in the periphery with the exception of CD25(+) cells. Patients with centrally low counts of FoxP3(+) TREGs had higher tumour stages than patients with high counts (p < 0.011). The number of FoxP3(+) TREGs was significantly associated with the number of CD8(+) cells (centre: p < 0.001, periphery: p = 0.002). The multivariate regression analysis identified UICC stage (IIB/III vs. I/IIA, hazard ratio 2.6, p = 0.011) and completeness of resection (no vs. yes, hazard ratio 2.3, p = 0.015) as independent predictors of survival. RCT significantly reduced the number of TREGs in the centre (p = 0.016) but not the number of the other TILs. CONCLUSION: UICC stage and completeness of resection but none of the TILs were prognostic markers for long-term survival. We found no morphologic evidence that TREGs suppress immunological response, represented by the infiltration of CD8(+) cells. Preoperative RCT affected the centre of tumours more than the periphery, which may indicate that it does not inhibit the host-to-tumour reaction. RCT affects TREGs more than the other TILs.

Prognostic and predictive value of TOPK stratified by KRAS and BRAF gene alterations in sporadic, hereditary and metastatic colorectal cancer patients.

BACKGROUND: Our aim was to investigate the prognostic and predictive value of the oncogenic MAPKK-like protein T-cell-originated protein kinase (TOPK) stratified by KRAS and BRAF mutations in patients with sporadic, hereditary and metastatic colorectal cancer (CRC) treated with anti-EGFR therapy. METHODS: Immunohistochemistry (IHC) for TOPK was performed on four study groups. Group 1 included two subgroups of 543 and 501 sporadic CRC patients used to test the reliability of TOPK expression by IHC. In Group 2, representing an additional 222 sporadic CRCs, the prognostic effect of TOPK stratified by KRAS and BRAF was assessed. The prognostic effect of TOPK was further analysed in Group 3, representing 71 hereditary Lynch syndrome-associated CRC patients. In Group 4, the predictive and prognostic value of TOPK was analysed on 45 metastatic patients treated with cetuximab or panitumumab stratified by KRAS and BRAF gene status. RESULTS: In both sporadic CRC subgroups (Group 1), associations of diffuse TOPK expression with clinicopathological features were reproducible. Molecular analysis of sporadic CRCs in Group 2 showed that diffuse TOPK expression was associated with KRAS and BRAF mutations (p<0.001) and with poor outcome in patients with either mutation in univariate and multivariate analysis (P=0.017). In hereditary patients (Group 3), diffuse TOPK was linked to advanced pT stage. In metastatic patients treated with anti-EGFR therapy (Group 4), diffuse TOPK expression was linked to dismal outcome despite objective response to treatment (P=0.01). CONCLUSIONS: TOPK expression is an unfavourable prognostic indicator in sporadic patients with KRAS or BRAF mutations and also in patients with metastatic disease experiencing a response to anti-EGFR therapies. The inhibition of TOPK, which could benefit 30-40% of CRC patients, may represent a new avenue of investigation for targeted therapy.

Influence of neoadjuvant radio-chemotherapy on tumor-infiltrating lymphocytes in squamous esophageal cancer.

INTRODUCTION: Data on influence of radio-chemotherapy (RCT) on tumor-infiltrating lymphocytes (TILs) is scarce and no study addressed this issue in esophageal squamous cell cancer (SCC) so far. METHODS: Tumor specimens of 49 patients with SCC were re-evaluated with immunohistochemical staining with anti-CD3, anti-CD4, anti-CD8, anti-CD25 and anti-FOXP3 antibodies. Lymphocytes were counted in one high power field (0.189 mm(2)) at the periphery and in the centre of tumors. RESULTS: 21 patients received preoperative RCT, 28 proceeded directly to surgery. There was no significant difference in survival between the two groups (median survival 23.2 months vs. 22.1 months, log rank test p=n.s.). Cox regression analysis showed that no variable had a significant effect on survival. The infiltrating pattern of TILs revealed higher numbers peripherally independent of the administration of RCT. There was a significant decrease in all cell numbers except CD4(+) cells in the centre of the tumors after RCT (CD3(+)p=0.005; CD8(+)p=0.02; CD25(+)p=0.01; FoxP3(+)p=0.01). There were fewer TILs in the periphery after RCT; however, this difference only reached significance in FoxP3(+) cells (p=0.01). CONCLUSION: Neoadjuvant RCT reduced the number of TILs in esophageal SCC. This was primarily seen in the centre of tumors and suggests that the effect of RCT on immunological response is located in the centre of tumors.

Prognostic influence of immunohistochemically detected lymph node micrometastasis and histological subtype in pN0 oesophageal cancer.

INTRODUCTION: Differences in frequency and clinical impact of lymph node micrometastasis between histological subtypes of oesophageal cancer have not been determined. METHODS: 1204 lymph nodes from 32 squamous cell carcinomas and 54 adenocarcinomas with complete resection and pN0 status were re-evaluated using a serial sectioning protocol including immunohistochemistry. Intra-nodal tumour cells were classified as micrometastases (0.2-2 mm) or isolated tumour cells (<0.2 mm). RESULTS: There was no significant difference in the frequency of micrometastases between adenocarcinoma and squamous cell carcinoma (11.3% vs. 3.1%, p=n.s.). In the squamous cell carcinoma group, Kaplan-Meier curves showed a significantly prolonged 5-year survival (p=0.02) and disease free interval (p<0.01) for immunohistochemically node negative versus node positive patients. In patients with adenocarcinoma, no such difference (p=n.s. and p=n.s., respectively) was seen. In patients who did not undergo pre-treatment, those with adenocarcinoma had a significant 5-year survival (65% vs. 53%; p=0.03) and disease free interval (83% vs. 58%; p<0.05) advantage over those with squamous cell carcinoma. After pre-treatment, no difference between the histological subtypes was detected. Regression analysis did not reveal any factors that significantly affected overall survival in node negative patients. However, four factors did significantly influence disease free interval: pre-treatment (HR 3.3 [95% CI 1.2-9.1], p=0.02); micrometastasis (HR 5.3 [95% CI 1.4-19.7], p=0.01); UICC stage II vs. 0/I (HR 2.2 [95% CI 1.1-4.4], p=0.03); and adenocarcinoma (HR 0.3 [95% CI 0.1-0.9], p=0.03). CONCLUSION: The difference in frequency and clinical impact of immunohistochemically detected micrometastasis may indicate that adenocarcinoma and squamous cell carcinoma should not be treated as one entity.

Angulated abutments and perimplants stress: F.E.M. analysis.

The long-term success of an implant supported prosthesis depends on many variables. In addition to the osseointegration rules that are commonly acknowledged, a clinician should consider also the biomechanical concepts related to the manufacturing process of a prosthetic structure assembly and their effects on the bone/implant interface. These concepts are particularly important when the anatomical structures leads to angulate the prosthetic component with respect to the fixture. In this study the stress distribution on the bone and on the prosthetic components was evaluated by means of a Finite Element Analysis (FEA). The effect of abutment axis angle was investigated considering three different measures: 15°, 25° and 35°. The simulation of both 450N maximal biting and 100N functional forces, highlighted the stress trend, which suggests as a maximum limit to choose an abutment with an angulation of 25°.

Mesenchymal stem cells share molecular signature with mesenchymal tumor cells and favor early tumor growth in syngeneic mice.

Tumor microenvironment in carcinomas recruits mesenchymal cells with an abnormal proangiogenic and invasive phenotype. It is not clear whether mesenchymal tumor cells (MTCs) derive from the activation of mature fibroblasts or from their stem cell precursors. However, stromal cell activation in tumors resembles in several aspects the mesenchymal rearrangement which normally occurs during reparative processes such as wound healing. Mesenchymal stem cells (MSCs) play a crucial role in developmental and reparative processes and have extraordinary proangiogenic potential, on the basis of which they are thought to show great promise for the treatment of ischemic disorders. Here, we show that MTCs have proangiogenic potential and that they share the transcriptional expression of the best-known proangiogenic factors with MSCs. We also found that MTCs and MSCs have the same molecular signature for stemness-related genes, and that when co-implanted with cancer cells in syngeneic animals MSCs determine early tumor appearance, probably by favoring the angiogenic switch. Our data (1) reveal crucial aspects of the proangiogenic phenotype of MTCs, (2) strongly suggest their stem origin and (3) signal the risk of therapeutic use of MSCs in tumor-promoting conditions.

IGFII and MIB1 immunohistochemistry is helpful for the differentiation of benign from malignant adrenocortical tumours.

AIMS: The differentiation of adrenocortical carcinomas from adenomas may be difficult based on morphology alone. Differential expression of insulin-like growth factor (IGF) II and cyclin-dependent kinase (CDK) 4 has recently been described in these tumours. The aim of this study was to investigate the diagnostic usefulness of these markers immunohistochemically. METHODS AND RESULTS: We examined 22 benign and 17 malignant adrenocortical tumours and compared IGFII and CDK4 expression with known immunohistochemical as well as morphological criteria of malignancy. Thirteen of 17 carcinomas showed immunohistochemical reactivity for IGFII, whereas all adenomas but one were negative. Intense CDK4 expression was detected in 11 of 17 carcinomas but was present in only three of 22 adenomas. The MIB1 index was >5% in 14 of 16 carcinomas and was <5% in all adenomas but one. The combination of IGFII immunohistochemistry with MIB1 index led to high sensitivity and specificity in detecting adrenocortical carcinomas. CONCLUSIONS: IGFII and MIB1 are helpful immunohistochemical markers to predict malignancy in adrenocortical neoplasms. These markers can be used in addition to clinical, gross and morphological features to establish a diagnosis in difficult cases.