Post-partum testosterone administration partially reverses the effects of perinatal cadmium exposure on sexual behavior in rats

This study investigated the effects of perinatal cadmium exposure on sexual behavior, organ weight, and testosterone levels in adult rats. We examined whether immediate postpartum testosterone administration is able to reverse the toxic effects of the metal. Forty pregnant Wistar rats were divided into three groups: 1) control, 2) 10 mg kg-1 cadmium chloride per day, and 3) 20 mg kg-1 cadmium chloride per day. These dams were treated on gestational days 18 and 21 and from lactation 1 to 7. Immediately after birth, half of the offspring from the experimental and control groups received 50 μl (i.p.) of 0.2% testosterone. Male sexual behavior, histological analysis and weight of organs as well as serum testosterone levels were assessed. Results showed that both cadmium doses disrupted sexual behavior in male rats, and postnatal treatment with testosterone reversed the toxic effects of 10 mg kg-1 cadmium and attenuated the effects of 20 mg kg-1 cadmium. Body weight and absolute testis, epididymis, and seminal vesicle weight were decreased by the higher cadmium dose, and testosterone supplementation did not reverse these effects. Serum testosterone levels were unaffected by both cadmium doses. No histological changes were detected in all organs analyzed. Maternal cadmium exposure effects in sexual parameters of male rat offspring were explained by the altered masculinization of the hypothalamus. We suggest that cadmium damaged cerebral sexual differentiation by its actions as an endocrine disruptor and supported by the changes discretely observed from early life during sexual development to adult life, reflected by sexual behavior. Testosterone supplementation after birth reversed some crucial parameters directly related to sexual behavior.

Stability of maxillary anterior crowding treatment

OBJECTIVE: To evaluate the stability and the relapse of maxillary anterior crowding treatment on cases with premolar extraction and evaluate the tendency of the teeth to return to their pretreatment position. METHODS: The experimental sample consisted of 70 patients of both sex with an initial Class I and Class II maloclusion and treated with first premolar extractions. The initial mean age was 13,08 years. Dental casts' measurements were obtained at three stages (pretreatment, posttreatment and posttreatment of 9 years on average) and the variables assessed were Little Irregularity Index, maxillary arch length and intercanine. Pearson correlation coefficient was used to know if some studied variable would have influence on the crowding in the three stages (LII1, LII2, LII3) and in each linear displacement of the Little irregularity index (A, B, C, D, E) in the initial and post-retention phases. RESULTS: The maxillary crowding relapse ( LII3-2) is influenced by the initial ( LII1), and the teeth tend to return to their pretreatment position. CONCLUSION: The results underline the attention that the orthodontist should be given to the maxillary anterior relapse, primarily on those teeth that are crowded before the treatment.

Cephalometric effects of the use of 10-hour Force Theory for Class II treatment

OBJECTIVE: This study aimed to evaluate the cephalometric effects promoted by the orthodontic treatment of Class II malocclusion patients with the use of the 10-Hour Force Theory, that consists in the use of fixed appliances with 8 hours a day using a cervical headgear appliance and 16 hours a day using Class II elastics, 8 hours on the first mandibular molar and 8 hours in the second mandibular molar. METHODS: Sample comprised 31 patients with mean initial age of 14.90 years, final mean age of 17.25 years and mean treatment time of 2.35 years. The lateral cephalograms in pre-treatment and post-treatment stages were evaluated. Evaluation of cephalometric changes between initial and final treatment phases was performed by paired t test. RESULTS: The cases treated with the 10-Hour Force Theory presented a slight restriction of anterior displacement of the maxilla, increase in the effective length of the mandible, significant improvement of the maxillomandibular relationship, significant increase in anterior lower face height, distal tipping of the maxillary premolar crowns, extrusion and distal tipping of the roots of maxillary molars, significant proclination and protrusion of mandibular incisors, significant extrusion and mesialization of mandibular molars, besides a significant correction of the molar relationship, overjet and overbite. CONCLUSION: The use of the 10-Hour Force Theory in treatment of Class II malocclusion provided satisfactory results.

Evaluation of the stability of open bite treatment using a removable appliance with palatal crib combined with high-pull chincup

OBJECTIVE: The aim of this prospective study was to cephalometrically analyze the stability of dentoalveolar and skeletal changes produced by a removable appliance with palatal crib associated to high-pull chincup in individuals with anterior open bite treated for 12 months, and compare them to individuals with similar malocclusion and age, not submitted to orthodontic treatment, also followed for the same period. METHODS: Nineteen children with a mean age of 9.78 years old treated for 12 months with a removable appliance with palatal crib associated with chincup therapy were evaluated after 15 months (post-treatment period) and compared with a control group of 19 subjects with mean age of 9.10 years with the same malocclusion that was followed-up for the same period. Seventy-six lateral cephalograms were evaluated at T1 (after correction) and T2 (follow-up) and cephalometric variables were analyzed by statistical tests. RESULTS: The results did not show significant skeletal, soft tissue or maxillary dentoalveolar changes. Overall, treatment effects on the experimental group were maintained at T2 evaluation with an increase of 0.56 mm in overbite. Overjet and maxillary incisors/molars position (vertical and sagittal) remained essentially unchanged during the study period. Only mandibular incisors showed significant changes (labial inclination and protrusion) compared to control group. CONCLUSIONS: Thus, it can be concluded that the early open bite treatment with a removable appliance and palatal crib associated with high-pull chincup therapy provided stability of 95%.

Proctocolectomy and ileal J-pouch anal anastomosis on the surgical treatment of familial adenomatous polyposis and ulcerative colitis: analysis of 49 cases

OBJECTIVE: To evaluate the results of ileal J-pouch anal anastomosis in ulcerative colitis and familial adenomatous polyposis. METHOD: Retrospective analysis of medical records of 49 patients submitted to ileal J-pouch anal anastomosis. RESULTS: Ulcerative colitis was diagnosed in 65% and familial adenomatous polyposis in 34%. Mean age was 39.5 years. 43% were male. Among familial adenomatous polyposis, 61% were diagnosed with colorectal cancer. Thirty-one percent of patients with ulcerative colitis was submitted to a previous surgical approach and 21% of these had toxic megacolon. Average hospital stay was 10 days. Post-operative complications occurred in 50% of patients with ulcerative colitis and 29.4% with familial adenomatous polyposis. Intestinal diversion was performed in 100% of ulcerative colitis and 88% of familial adenomatous polyposis. Pouchitis occurred in eight cases (seven ulcerative colitis and one FAP), requiring excision of the pouch in three ulcerative colitis. Mortality rate was 7.6%: two cases of carcinoma on the pouch and two post-operative complications. Late post-operative complications occurred in 22.4%: six familial adenomatous polyposis and five ulcerative colitis). Two patients had erectile dysfunction, and one retrograde ejaculation. One patient with severe perineal dermatitis was submitted to excision of the pouch. Incontinence occurred in four patients and two reported soil. Mean bowel movement was five times a day. CONCLUSION: Ileal J-pouch anal anastomosis is a safe surgery with acceptable morbidity and good functional results, if well indicated and performed in referral centers.

LH surge in response to the treatment with GnRH analog or estradiol in ovariectomized buffaloes with or without progesterone pre-exposition

The aim of the present study was to evaluate the LH surge after EB (estradiol benzoate) or GnRH administration with or without P4 (progesterone) pre-exposure in ovariectomized (OVX) buffalo cows. Females were randomly assigned to receive an intravaginal P4 device (D0–D9). They were then given EB 24 h or GnRH 36 h post-P4 device removal (factorial 2×2, n=6 per group). Blood collection for LH measurement began 36 h after the P4 device removal and continued at 3 h intervals. The area under the LH curve (AUC; 30.2 ng2 and 13.41 ng2; P=0.007) and the area of the LH peak (AP; 19.0 ng2 and 8.9 ng2; P=0.009) were greater for EB than GnRH. We did not observe an effect of P4 pre-exposure on the AUC and AP. Furthermore, there was no interaction between P4 pre-exposure and EB or GnRH treatment on the AUC and AP. However, there was an interaction (P<0.01) between P4 pre-exposure and the type of inducer (EB or GnRH) to release a preovulatory-like LH surge at the beginning (BP), final (FP) and time (TP) of the LH peak. The P4 pre-exposure anticipated the BP (2.5 and 7.4 h), TP (6.0 and 12.0 h) and FP (11.5 and 17.1 h) when EB was used to induce a preovulatory-like LH surge (P<0.01). However, there was no effect of P4 pre-exposure on BP (0.4 and 0.4 h), TP (3.0 and 3.0 h) and FP (5.9 and 6.1 h) with GnRH treatment. There was also no effect of the pre-exposure to P4, type of inducer or interaction on the amplitude of the LH peak. We concluded that EB therefore led to greater LH release than GnRH, and pre-exposure to P4 before EB administration anticipated the preovulatory-like LH surge in buffalo cows.

Exome analysis of HIV patients submitted to dendritic cells therapeutic vaccine reveals an association of CNOT1 gene with response to the treatment

INTRODUCTION: With the aim of searching genetic factors associated with the response to an immune treatment based on autologous monocyte-derived dendritic cells pulsed with autologous inactivated HIV, we performed exome analysis by screening more than 240,000 putative functional exonic variants in 18 HIV-positive Brazilian patients that underwent the immune treatment. METHODS: Exome analysis has been performed using the ILLUMINA Infinium HumanExome BeadChip. zCall algorithm allowed us to recall rare variants. Quality control and SNP-centred analysis were done with GenABEL R package. An in-house implementation of the Wang method permitted gene-centred analysis. RESULTS: CCR4-NOT transcription complex, subunit 1 (CNOT1) gene (16q21), showed the strongest association with the modification of the response to the therapeutic vaccine (p=0.00075). CNOT1 SNP rs7188697 A/G was significantly associated with DC treatment response. The presence of a G allele indicated poor response to the therapeutic vaccine (p=0.0031; OR=33.00; CI=1.74-624.66), and the SNP behaved in a dominant model (A/A vs. A/G+G/G p=0.0009; OR=107.66; 95% CI=3.85-3013.31), being the A/G genotype present only in weak/transient responders, conferring susceptibility to poor response to the immune treatment. DISCUSSION: CNOT1 is known to be involved in the control of mRNA deadenylation and mRNA decay. Moreover, CNOT1 has been recently described as being involved in the regulation of inflammatory processes mediated by tristetraprolin (TTP). The TTP-CCR4-NOT complex (CNOT1 in the CCR4-NOT complex is the binding site for TTP) has been reported as interfering with HIV replication, through post-transcriptional control. Therefore, we can hypothesize that genetic variation occurring in the CNOT1 gene could impair the TTP-CCR4-NOT complex, thus interfering with HIV replication and/or host immune response. CONCLUSIONS: Being aware that our findings are exclusive to the 18 patients studied with a need for replication, and that the genetic variant of CNOT1 gene, localized at intron 3, has no known functional effect, we propose a novel potential candidate locus for the modulation of the response to the immune treatment, and open a discussion on the necessity to consider the host genome as another potential variant to be evaluated when designing an immune therapy study

Evaluation of fluid transport processes in dental enamel. Methods to assess the relevance of enamel permeability in caries prevention and etching treatments.

This thesis evaluated in vivo and in vitro enamel permeability in different physiological and clinical conditions by means of SEM inspection of replicas of enamel surface obtained from polyvinyl siloxane impressions subsequently later cast in polyether impression ma-terial. This technique, not invasive and risk-free, allows the evaluation of fluid outflow from enamel surface and is able to detect the presence of small quantities of fluid, visu-alized as droplets. Fluid outflow on enamel surface represents enamel permeability. This property has a paramount importance in enamel physiolgy and pathology although its ef-fective role in adhesion, caries pathogenesis and prevention today is still not fully under-stood. The aim of the studies proposed was to evaluate enamel permeability changes in differ-ent conditions and to correlate the findings with the actual knowledge about enamel physiology, caries pathogenesis, fluoride and etchinhg treatments. To obtain confirmed data the replica technique has been supported by others specific techniques such as Ra-man and IR spectroscopy and EDX analysis. The first study carried out visualized fluid movement through dental enamel in vivo con-firmed that enamel is a permeable substrate and demonstrated that age and enamel per-meability are closely related. Examined samples from subjects of different ages showed a decreasing number and size of droplets with increasing age: freshly erupted permanent teeth showed many droplets covering the entire enamel surface. Droplets in permanent teeth were prominent along enamel perikymata. These results obtained through SEM inspection of replicas allowed innovative remarks in enamel physiology. An analogous testing has been developed for evaluation of enamel permeability in primary enamel. The results of this second study showed that primary enamel revealed a substantive permeability with droplets covering the entire enamel sur-face without any specific localization accordingly with histological features, without changes during aging signs of post-eruptive maturation. These results confirmed clinical data that showed a higher caries susceptibility for primary enamel and suggested a strong relationship between this one and enamel permeability. Topical fluoride application represents the gold standard for caries prevention although the mechanism of cariostatic effect of fluoride still needs to be clarified. The effects of topical fluoride application on enamel permeability were evaluated. Particularly two dif-ferent treatments (NaF and APF), with different pH, were examined. The major product of topical fluoride application was the deposition of CaF2-like globules. Replicas inspec-tion before and after both treatments at different times intervals and after specific addi-tional clinical interventions showed that such globule formed in vivo could be removed by professional toothbrushing, sonically and chemically by KOH. The results obtained in relation to enamel permeability showed that fluoride treatments temporarily reduced enamel water permeability when CaF2-like globules were removed. The in vivo perma-nence of decreased enamel permeability after CaF2 globules removal has been demon-strated for 1 h for NaF treated teeth and for at least 7 days for APF treated teeth. Important clinical consideration moved from these results. In fact the caries-preventing action of fluoride application may be due, in part, to its ability to decrease enamel water permeability and CaF2 like-globules seem to be indirectly involved in enamel protection over time maintaining low permeability. Others results obtained by metallographic microscope and SEM/EDX analyses of or-thodontic resins fluoride releasing and not demonstrated the relevance of topical fluo-ride application in decreasing the demineralization marks and modifying the chemical composition of the enamel in the treated area. These data obtained in both the experiments confirmed the efficacy of fluoride in caries prevention and contribute to clarify its mechanism of action. Adhesive dentistry is the gold standard for caries treatment and tooth rehabilitation and is founded on important chemical and physical principles involving both enamel and dentine substrates. Particularly acid etching of dental enamel enamel has usually employed in bonding pro-cedures increasing microscopic roughness. Different acids have been tested in the litera-ture suggesting several etching procedures. The acid-induced structural transformations in enamel after different etching treatments by means of Raman and IR spectroscopy analysis were evaluated and these findings were correlated with enamel permeability. Conventional etching with 37% phosphoric acid gel (H3PO4) for 30 s and etching with 15 % HCl for 120 s were investigated. Raman and IR spectroscopy showed that the treatment with both hydrochloric and phosphoric acids induced a decrease in the carbonate content of the enamel apatite. At the same time, both acids induced the formation of HPO42- ions. After H3PO4 treatment the bands due to the organic component of enamel decreased in intensity, while in-creased after HCl treatment. Replicas of H3PO4 treated enamel showed a strongly reduced permeability while replicas of HCl 15% treated samples showed a maintained permeability. A decrease of the enamel organic component, as resulted after H3PO4 treatment, involves a decrease in enamel permeability, while the increase of the organic matter (achieved by HCl treat-ment) still maintains enamel permeability. These results suggested a correlation between the amount of the organic matter, enamel permeability and caries. The results of the different studies carried out in this thesis contributed to clarify and improve the knowledge about enamel properties with important rebounds in theoretical and clinical aspects of Dentistry.

Different approaches to identify markers for meat quality and production traits in pigs: analysis of the transcriptome in post mortem skeletal muscle and investigation of candidate genes

Due to the growing attention of consumers towards their food, improvement of quality of animal products has become one of the main focus of research. To this aim, the application of modern molecular genetics approaches has been proved extremely useful and effective. This innovative drive includes all livestock species productions, including pork. The Italian pig breeding industry is unique because needs heavy pigs slaughtered at about 160 kg for the production of high quality processed products. For this reason, it requires precise meat quality and carcass characteristics. Two aspects have been considered in this thesis: the application of the transcriptome analysis in post mortem pig muscles as a possible method to evaluate meat quality parameters related to the pre mortem status of the animals, including health, nutrition, welfare, and with potential applications for product traceability (chapters 3 and 4); the study of candidate genes for obesity related traits in order to identify markers associated with fatness in pigs that could be applied to improve carcass quality (chapters 5, 6, and 7). Chapter three addresses the first issue from a methodological point of view. When we considered this issue, it was not obvious that post mortem skeletal muscle could be useful for transcriptomic analysis. Therefore we demonstrated that the quality of RNA extracted from skeletal muscle of pigs sampled at different post mortem intervals (20 minutes, 2 hours, 6 hours, and 24 hours) is good for downstream applications. Degradation occurred starting from 48 h post mortem even if at this time it is still possible to use some RNA products. In the fourth chapter, in order to demonstrate the potential use of RNA obtained up to 24 hours post mortem, we present the results of RNA analysis with the Affymetrix microarray platform that made it possible to assess the level of expression of more of 24000 mRNAs. We did not identify any significant differences between the different post mortem times suggesting that this technique could be applied to retrieve information coming from the transcriptome of skeletal muscle samples not collected just after slaughtering. This study represents the first contribution of this kind applied to pork. In the fifth chapter, we investigated as candidate for fat deposition the TBC1D1 [TBC1 (tre-2/USP6, BUB2, cdc16) gene. This gene is involved in mechanisms regulating energy homeostasis in skeletal muscle and is associated with predisposition to obesity in humans. By resequencing a fragment of the TBC1D1 gene we identified three synonymous mutations localized in exon 2 (g.40A>G, g.151C>T, and g.172T>C) and 2 polymorphisms localized in intron 2 (g.219G>A and g.252G>A). One of these polymorphisms (g.219G>A) was genotyped by high resolution melting (HRM) analysis and PCR-RFLP. Moreover, this gene sequence was mapped by radiation hybrid analysis on porcine chromosome 8. The association study was conducted in 756 performance tested pigs of Italian Large White and Italian Duroc breeds. Significant results were obtained for lean meat content, back fat thickness, visible intermuscular fat and ham weight. In chapter six, a second candidate gene (tribbles homolog 3, TRIB3) is analyzed in a study of association with carcass and meat quality traits. The TRIB3 gene is involved in energy metabolism of skeletal muscle and plays a role as suppressor of adipocyte differentiation. We identified two polymorphisms in the first coding exon of the porcine TRIB3 gene, one is a synonymous SNP (c.132T> C), a second is a missense mutation (c.146C> T, p.P49L). The two polymorphisms appear to be in complete linkage disequilibrium between and within breeds. The in silico analysis of the p.P49L substitution suggests that it might have a functional effect. The association study in about 650 pigs indicates that this marker is associated with back fat thickness in Italian Large White and Italian Duroc breeds in two different experimental designs. This polymorphisms is also associated with lactate content of muscle semimembranosus in Italian Large White pigs. Expression analysis indicated that this gene is transcribed in skeletal muscle and adipose tissue as well as in other tissues. In the seventh chapter, we reported the genotyping results for of 677 SNPs in extreme divergent groups of pigs chosen according to the extreme estimated breeding values for back fat thickness. SNPs were identified by resequencing, literature mining and in silico database mining. analysis, data reported in the literature of 60 candidates genes for obesity. Genotyping was carried out using the GoldenGate (Illumina) platform. Of the analyzed SNPs more that 300 were polymorphic in the genotyped population and had minor allele frequency (MAF) >0.05. Of these SNPs, 65 were associated (P<0.10) with back fat thickness. One of the most significant gene marker was the same TBC1D1 SNPs reported in chapter 5, confirming the role of this gene in fat deposition in pig. These results could be important to better define the pig as a model for human obesity other than for marker assisted selection to improve carcass characteristics.

Post-transkriptionelle Regulation der Expression der humanen induzierbaren NO-Synthase

Die Expression der humanen induzierbaren NO-Synthase (iNOS) wird sowohl über transkriptionelle als auch über post-transkriptionelle Mechanismen reguliert. Dabei spielt die Modulation der iNOS-mRNA-Stabilität durch RNA-bindende Proteine eine bedeutende Rolle. In dieser Arbeit konnte eine Beteiligung des p38-MAPK-Signaltransduktionsweges sowie der RNA-bindenden Proteine TTP, KSRP, HuR und PTB an der Regulation der iNOS-Expression dargestellt werden. Hemmung der p38-MAPK führte zu einer Reduktion der iNOS-mRNA-Expression, hatte aber keinen Effekt auf die iNOS-Promotoraktivität. Das RNA-bindende Protein Tristetraprolin (TTP) erhöhte die Stabilität der iNOS-mRNA nach Zytokin-Stimulation, ohne jedoch mit ihr zu interagieren. Die Proteinexpression von TTP war unter dem Einfluss von Zytokinen erhöht; Inhibition der p38-MAPK verursachte eine Verminderung der Zytokin-stimulierten TTP-Expression. Das „KH-type splicing regulatory protein" (KSRP) übte einen destabilisierenden Effekt auf die iNOS-mRNA aus. Der Abbau der mRNA wird dabei wahrscheinlich durch eine Zytokin-unabhängige Interaktion von KSRP mit dem Exosom vermittelt. Ebenso konnte zwischen KSRP und TTP eine Wechselwirkung beobachtet werden, die nach Induktion der iNOS-Expression mit Zytokinen verstärkt und durch p38-MAPK-Inhibitoren hemmbar war. Des Weiteren konnte gezeigt werden, dass die Bindung von KSRP an die iNOS-mRNA-3’-UTR für die Vermittlung des destabilisierenden Effekts essentiell ist. Eine genaue Lokalisierung der KSRP-Bindungsstelle ergab, dass KSRP ebenso wie HuR mit dem AU-reichen Element am 3’-Ende der 3’-UTR interagiert. KSRP und HuR sind in der Lage, um diese Bindungsstelle zu konkurrieren. Nach Zytokin-Stimulation war dementsprechend die endogene Bindung von KSRP an die iNOS-mRNA vermindert, während die endogene Bindung von HuR an die iNOS-mRNA verstärkt war. Die Stabilisierung der iNOS-mRNA nach Zytokin-Stimulation ergibt sich demnach aus einer Verminderung der Bindung des KSRP-Exosom-Komplexes an die iNOS-mRNA als Folge der verstärkten Interaktion von TTP und KSRP. Dies ermöglicht parallel eine vermehrte Bindung von HuR an die iNOS-3’-UTR und führt damit zu einer Stabilisierung der iNOS-mRNA und so letztendlich auch zu einer Erhöhung der iNOS-Expression. Außerdem konnte eine Beteiligung des Polypyrimidin-Trakt-bindenden Proteins (PTB) an der Regulation der humanen iNOS-Expression gezeigt werden. PTB erhöhte die Expression der iNOS und interagierte Zytokin-unabhängig mit KSRP. Zusammenfassend lässt sich schließen, dass ein Zusammenspiel verschiedener Proteine in einem komplexen Netzwerk für die fein abgestimmte Regulation der humanen iNOS-Expression auf post-transkriptioneller Ebene verantwortlich.

Influence of genetic variants and post transcriptional factors on imatinib transporters as determinants of the pharmacological response

Introduction – Although imatinib (IM) is a recognized gold standard in chronic myeloid leukemia (CML) therapy, resistance has emerged in a significant proportion of patients. Aim – The aim of this study was: (1) to investigate the role of genetic variants in genes encoding for IM transporters, as candidate of IM responsiveness and (2) to test the influence of miRNAs on IM response, focusing on efflux transporters. Methods – As a first step, a panel of polymorphisms (SNPs) was genotyped in a subgroup population of 189 patients enrolled in the Tyrosine Kinase Inhibitor Optimization and Selectivity (TOPS) trial. The association with cytogenetic response and molecular response (MR) was assessed for each SNP. As a second step, an in vitro IM-resistant model (K-562 CML cell line) was established. miRNAs profiles were analyzed using Taqman arrays and in silico search was performed for miRNAs deregulated after IM treatment. mRNA and protein expression were quantified using TaqMan realtime PCR and Western blotting, respectively. Results – (1) Among Caucasian patients, ABCB1 rs60023214 significantly correlated with complete MR (P = 0.005). Concerning SNPs combination in IM uptake transporters, the associations with treatment outcomes were statistically significant for both major and complete MR (P = 0.005 and P = 0.01, respectively). (2) ABCB1 protein was not expressed under any conditions of treatment, differently from ABCG2. Two deregulated miRNAs, namely miR-212 and miR-328, were identified to be inversely correlated with ABCG2 (r2= 0.57; p=0.03 and r2=0.47; p=0.06, respectively). Experiments of loss and gain of function confirmed the functional influence of these miRNAs on ABCG2. Conclusion – The multiple candidate gene approach identified single and combination of SNPs that can be proposed as predictor of IM response. The in vitro study suggested that IM resistance could be mediated by miRNA-dependent mechanism. Further studies are needed to validate these preliminary findings.

The stratigraphic record of the quaternary sea level fluctuations and the impact of the post-glacial sea level rise (Termination I) in the Adriatic basin (Mediterranean sea)

The modern stratigraphy of clastic continental margins is the result of the interaction between several geological processes acting on different time scales, among which sea level oscillations, sediment supply fluctuations and local tectonics are the main mechanisms. During the past three years my PhD was focused on understanding the impact of each of these process in the deposition of the central and northern Adriatic sedimentary successions, with the aim of reconstructing and quantifying the Late Quaternary eustatic fluctuations. In the last few decades, several Authors tried to quantify past eustatic fluctuations through the analysis of direct sea level indicators, among which drowned barrier-island deposits or coral reefs, or indirect methods, such as Oxygen isotope ratios (δ18O) or modeling simulations. Sea level curves, obtained from direct sea level indicators, record a composite signal, formed by the contribution of the global eustatic change and regional factors, as tectonic processes or glacial-isostatic rebound effects: the eustatic signal has to be obtained by removing the contribution of these other mechanisms. To obtain the most realistic sea level reconstructions it is important to quantify the tectonic regime of the central Adriatic margin. This result has been achieved integrating a numerical approach with the analysis of high-resolution seismic profiles. In detail, the subsidence trend obtained from the geohistory analysis and the backstripping of the borehole PRAD1.2 (the borehole PRAD1.2 is a 71 m continuous borehole drilled in -185 m of water depth, south of the Mid Adriatic Deep - MAD - during the European Project PROMESS 1, Profile Across Mediterranean Sedimentary Systems, Part 1), has been confirmed by the analysis of lowstand paleoshorelines and by benthic foraminifera associations investigated through the borehole. This work showed an evolution from inner-shelf environment, during Marine Isotopic Stage (MIS) 10, to upper-slope conditions, during MIS 2. Once the tectonic regime of the central Adriatic margin has been constrained, it is possible to investigate the impact of sea level and sediment supply fluctuations on the deposition of the Late Pleistocene-Holocene transgressive deposits. The Adriatic transgressive record (TST - Transgressive Systems Tract) is formed by three correlative sedimentary bodies, deposited in less then 14 kyr since the Last Glacial Maximum (LGM); in particular: along the central Adriatic shelf and in the adjacent slope basin the TST is formed by marine units, while along the northern Adriatic shelf the TST is represented by costal deposits in a backstepping configuration. The central Adriatic margin, characterized by a thick transgressive sedimentary succession, is the ideal site to investigate the impact of late Pleistocene climatic and eustatic fluctuations, among which Meltwater Pulses 1A and 1B and the Younger Dryas cold event. The central Adriatic TST is formed by a tripartite deposit bounded by two regional unconformities. In particular, the middle TST unit includes two prograding wedges, deposited in the interval between the two Meltwater Pulse events, as highlighted by several 14C age estimates, and likely recorded the Younger Dryas cold interval. Modeling simulations, obtained with the two coupled models HydroTrend 3.0 and 2D-Sedflux 1.0C (developed by the Community Surface Dynamics Modeling System - CSDMS), integrated by the analysis of high resolution seismic profiles and core samples, indicate that: 1 - the prograding middle TST unit, deposited during the Younger Dryas, was formed as a consequence of an increase in sediment flux, likely connected to a decline in vegetation cover in the catchment area due to the establishment of sub glacial arid conditions; 2 - the two-stage prograding geometry was the consequence of a sea level still-stand (or possibly a fall) during the Younger Dryas event. The northern Adriatic margin, characterized by a broad and gentle shelf (350 km wide with a low angle plunge of 0.02° to the SE), is the ideal site to quantify the timing of each steps of the post LGM sea level rise. The modern shelf is characterized by sandy deposits of barrier-island systems in a backstepping configuration, showing younger ages at progressively shallower depths, which recorded the step-wise nature of the last sea level rise. The age-depth model, obtained by dated samples of basal peat layers, is in good agreement with previous published sea level curves, and highlights the post-glacial eustatic trend. The interval corresponding to the Younger Dyas cold reversal, instead, is more complex: two coeval coastal deposits characterize the northern Adriatic shelf at very different water depths. Several explanations and different models can be attempted to explain this conundrum, but the problem remains still unsolved.

Electrostatic supramolecular assembly of charged dendritic polymers and their biological application

The aim of this study was the development of functional multilayer films through electrostatic layer by layer (LbL) assembly of dendritic macromolecules, the investigation of the fundamental properties of these multilalyered films and the study of their biological applications. rnThe synthesis of the anionic hyperbranched polyglycerols (hbPG) and the preparation of multilayers made of hbPG/phosphorus dendrimer as well as the influences of deposition conditions on multilayers were reported. The thicknesses of multilayer films increase with a decrease of molecular weight of anionic hbPGs. The multilayer films fabricated by low molecular weight hbPGs grow less regularly due to the less charged carboxylic acid groups providing the relative weaker electrostatic forces for the deposition. The thicknesses of multilayer films are reduced with increasing pH values and decreasing the concentration of NaCl. The observed changes of multilayer thickness and surface morphology could be interpreted with the aid of theories regarding the charge density and conformation of the anionic hbPG chains in solution. rnBesides the study of fundamental properties of hbPG/phosphorus multilayer films, antifouling thin films derived from hbPG layers were developed. The antifouling properties of hbPG layers were found to correlate with factors of the molecular weight of anionic hbPG and the film thickness. It was demonstrated that anionic hbPG single layer with highest molecular weight can reduce non specific protein adsorption more efficiently than single layer with lower molecular weight and all the hbPG bilayers possessed excellent property of antifouling. rnPhosphorus dendrimer multilayers were successfully prepared as the platforms to detect DNA immobilization and hybridization. The effect of NaCl concentration on the multilayer film thickness was evaluated to obtain the optimized film thickness. Making use of the multilayer deposited at the optimized condition as a substrate, a high loading of DNA probes was achieved through covalent coupling of probe DNA with the as-formed multilayer films. The hybridization of target DNA with immobilized probe DNA was then carried out and studied by SPFS. The limit of detection upon hybridization was estimated on various dendrimer multilayer platforms. The minimum detection concentration for DNA hybridization is in the same order of magnitude compared with other neutral phosphorus dendrimer systems. Furthermore, the LbL deposition of phosphorus dendrimer multilayers provided a mild and simple way to prepare platforms as DNA microarrays. rnBased on the phosphorus dendrimer multilayer systems, dendritic star polymers were employed which have more reactive groups than that phosphorus dendrimers. The as-assembled dendritic star polymer multilayer films exhibited such distinct morphology characteristics that they underwent extensive structural reorganization upon post-treatment under different pH conditions. Kinetic binding of probe DNA molecules on the outermost negatively charged dendritic surface was studied by SPR as well. The binding capacities of probe DNA on the multilayer surfaces fabricated from the first-generation and the second-generation of dendritic star polymers were compared. The improved binding capacity was achieved from the second-generation of dendritic star polymer multilayer films due to their more reactive groups. DNA hybridization reaction on dendritic multilayer films was investigated by SPFS. The similar hybridization behaviors were found on both multilayer surfaces. Meanwhile, the hybridization kinetic affinities were compared with that of phosphorus dendrimer multilayer surfaces and showed improved detection sensitivity than phosphorus dendrimer multilayer films.rn

Farmacocinetica della buprenorfina, e del suo metabolita norbuprenorfina, somministrata come infusione costante nel periodo post-operatorio in cagne sane sottoposte ad ovariectomia

Il dolore non è solo una conseguenza della malattia ma un fattore patogeno che è di per se stesso in grado di perpetuare il danno all’organismo. Il suo trattamento non è quindi solo un atto di umanità ma un contributo ad arrestare la malattia e restituire la salute al paziente. Tra i farmaci più popolari per la terapia del dolore negli animali da affezione si trova la buprenorfina. Questa molecola viene impiegata con successo da anni nel cane e nel gatto per motivi riconducibili, oltre che alla sua efficacia (la sua potenza è diverse volte quella della morfina), alla lunga durata d’azione e alla scarsità degli effetti collaterali. Nonostante l’ampia diffusione e longevità del suo utilizzo, però, sappiamo poco della farmacocinetica di questa molecola negli animali da affezione; i dosaggi clinicamente impiegati sono di fatto estrapolati dagli studi nell’uomo oppure basati su semplici osservazioni degli effetti; i pochi dati farmacocinetici ottenuti nel cane fanno riferimento a singoli boli di dosi che non sempre corrispondono a quelle clinicamente impiegate. Nonostante la buprenorfina trovi il suo principale impiego nelle somministrazioni protratte a lungo (durante il periodo post-operatorio o la degenza ospedaliera) non è mai stato indagato il profilo farmacocinetico della molecola somministrata a boli ripetuti o come infusione continua. Il nostro studio si pone come obiettivo di indagare la farmacocinetica della buprenorfina somministrata come bolo di carico seguito da un’infusione costante a dosaggi considerati clinici in cani sani nel periodo post operatorio. Lo scopo ultimo è quello di sviluppare un protocollo per la somministrazione di questa molecola in modo prolungato in pazienti degenti ed addolorati per poi, in futuro, confrontare la somministrazione come infusione continua con i tradizionali boli ripetuti. Per lo studio sono state utilizzate giovani cagne adulte di taglia media o grande sottoposte ad intervento di ovariectomia.

Il trattamento ortodontico nei bambini con particolari necessità sanitarie (SHCN): una valutazione della durata e del risultato clinico utilizzando l'indice PAR (Peer Assessment Rating), la componente DHC (Dental Health Component) e la componente AC (Aesthetic Component) dell'indice IOTN (Orthodontic Treatment Need Index)

Obbiettivo: Valutazione delle eventuali differenze nel trattamento ortodontico di un gruppo di bambini con particolari necessità sanitarie (SHCN) rispetto ad un gruppo di bambini non diagnosticati con SHCN. Materiali e Metodi: Il gruppo campione (SHCN) è costituito da 50 bambini con SHCN. Il gruppo di controllo (NO SHCN) è costituito da 50 bambini non diagnosticati con SHCN pienamente corrispondenti per età, genere e tipo di apparecchio ortodontico utilizzato con i pazienti del gruppo di studio. I dati riguardanti i gruppi SHCN e NO SHCN sono stati analizzati in modo retrospettivo, valutando: - il punteggio pre- e post-trattamento e la riduzione finale dei valori dell'indice PAR (Peer Assessment Rating), della componente DHC (Dental Health Component) e della componente AC (Aesthetic Component) dell'indice IOTN (Orthodontic Treatment Need Index), - il numero di appuntamenti, - il numero di sedute semplici e complesse, - la durata complessiva del trattamento, - l'età all’inizio ed alla fine della terapia. Risultati: Non sono state rilevate differenze statisticamente significative tra i due gruppi per quanto concerne il numero di appuntamenti, la durata complessiva del trattamento, l'età all’inizio ed alla fine della terapia ortodontica (valori del p-value:0.682, 0.458, 0.535, 0.675). Sono state rilevate differenze statisticamente significative tra i due gruppi per quanto riguarda i punteggi dell’indice PAR, delle componenti DHC e AC dello IOTN pre- e post-trattamento, il numero di sedute semplici e complesse (valori del p-value:0.030, 0.000, 0.020, 0.023, 0.000, 0.000, 0.043, 0.037). Per quanto concerne la riduzione finale del valore dell’indice PAR, della componente DHC e di quella AC dello IOTN non sono state riscontrate differenze statisticamente significative tra i due gruppi (valori del p-value:0.060, 0.765, 0.825). Conclusioni: Lo studio incoraggia gli ortodontisti a trattare i bambini con SHCN nell'obiettivo di migliorarne la qualità di vita, pur evidenziando la necessità di un maggior numero di sedute complesse.