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Nucleosomes are a crucial platform for the recruitment and assembly of protein complexes that process the DNA. Mechanistic and structuralin vitrostudies typically rely on recombinant nucleosomes that are reconstituted using artificial, strong-positioning DNA sequences. To facilitate such studies on native, genomic nucleosomes, there is a need for methods to produce any desired DNA sequence in an efficient manner. The current methods either do not offer much flexibility in choice of sequence or are less efficient in yield and labor. Here, we show that using ramified rolling circle amplification (RCA) milligram amounts of a genomic nucleosomal DNA fragment can be produced in a scalable, one-pot reaction overnight. The ramified RCA reaction is more efficient than the existing methods, is flexible in DNA sequence and shows a 10-fold increase in yield compared to PCR, rivalling the production using plasmids. We demonstrate the method by producing the genomic DNA from the human LIN28B locus and show that it forms functional nucleosomes capable of binding pioneer transcription factor Oct4.

The regulation of chromatin biology ultimately depends on the manipulation of its smallest subunit, the nucleosome. The proteins that bind and operate on the nucleosome do so, while their substrate is part of a polymer embedded in the dense nuclear environment. Their molecular interactions must in some way be tuned to deal with this complexity. Due to the rapid increase in the number of high-resolution structures of nucleosome-protein complexes and the increasing understanding of the cellular chromatin structure, it is starting to become clearer how chromatin factors operate in this complex environment. In this review, we analyze the current literature on the interplay between nucleosome-protein interactions and higher-order chromatin structure. We examine in what way nucleosomes-protein interactions can affect and can be affected by chromatin organization at the oligonucleosomal level. In addition, we review the characteristics of nucleosome-protein interactions that can cause phase separation of chromatin. Throughout, we hope to illustrate the exciting challenges in characterizing nucleosome-protein interactions beyond the nucleosome.

A pregnancy related cancer is defined as a neoplasm diagnosed in a woman during gestation or within 12 months following childbirth or abortion. Breast, thyroidal, melanoma, cervical and hematological malignancies are those most commonly diagnosed during pregnancy, with a reported incidence considered lower than those estimated of 1 case on 1000 pregnancies. The clinical complexity is generated by the co-presence of mother and fetus, and the consequent conflict represented by the need to identify a therapeutic strategy that is effective for the maternal prognosis and, at the same time, safeguards the fetal well-being. Moreover, the counseling, diagnosis, staging and management of these cases are further complicated because most of the scientific literature consists in case reports or small retrospective studies and lacks in guidelines and prospective clinical trials. On that basis, the Study Group on Cancer in Pregnancy (members of S.I.G.O.-Italian Society of Gynecology and Obstetrics) presents the TIGRE Trial, an Italian multicenter observational study aimed at establishing a shared database for the collection of epidemiological and clinical data of the patients affected by pregnancy related cancer in Italy, with the final objective to improve the outcome of a mother and a child involved in a such pathology.

China is rapidly becoming a major industrial competitor in high tech and growth sectors. Its economic success and related industrial policies have received a high degree of attention, especially in light of its capacity to challenge the leading position of advanced economies in several fields. China aims, through the 'Made in China 2025' strategy, to become a world leader in key industrial sectors. In these sectors, it strives to strengthen its domestic innovation capacity, to reduce its reliance on foreign technologies while moving up in global value chains. This report analyses China's approach to attain a dominant position in international markets through a combination of industrial, R&I, trade and foreign direct investment policies. It offers an assessment of China's current position compared to the EU and US innovation systems across a range of dimensions. It concludes that China has become a major industrial competitor in several rapidly expanding high tech sectors, which may well result in attaining China's goal of becoming an innovation leader in specific areas. As a response, the EU will need to boost its industrial and R&I performance and develop a trade policy that can ensure a level playing field for EU companies in China and for Chinese companies in the EU., JRC.B.7-Knowledge for Finance, Innovation and Growth

Unrepairable comminuted fractures of the radial head Mason type III or type IV have poor outcomes when treated by open reduction and internal fixation. Radial head resection has been proposed as good option for surgical treatment, while in the last decades, the development of technology and design in radial head prosthesis has increased efficacy in prosthetic replacement. The present review was conducted to determine the best surgical treatment for comminuted radial head when ORIF is not possible. Better outcomes are reported for radial head arthroplasty in terms of elbow stability, range of motion, pain, and fewer complications compared to radial head excision. Nevertheless, radial head resection still can be considered an option of treatment in isolated radial head fractures with no associated ligament injuries lesion of ligaments or in case of older patients with low demanding function.

Unrepairable comminuted fractures of the radial head Mason type III or type IV have poor outcomes when treated by open reduction and internal fixation. Radial head resection has been proposed as good option for surgical treatment, while in the last decades, the development of technology and design in radial head prosthesis has increased efficacy in prosthetic replacement. The present review was conducted to determine the best surgical treatment for comminuted radial head when ORIF is not possible. Better outcomes are reported for radial head arthroplasty in terms of elbow stability, range of motion, pain, and fewer complications compared to radial head excision. Nevertheless, radial head resection still can be considered an option of treatment in isolated radial head fractures with no associated ligament injuries lesion of ligaments or in case of older patients with low demanding function.

Nucleosomes are a crucial platform for the recruitment and assembly of protein complexes that process the DNA. Mechanistic and structural in vitro studies typically rely on recombinant nucleosomes that are reconstituted using artificial, strong-positioning DNA sequences. To facilitate such studies on native, genomic nucleosomes, there is a need for methods to produce any desired DNA sequence in an efficient manner. The current methods either do not offer much flexibility in choice of sequence or are less efficient in yield and labor. Here, we show that ramified rolling circle amplification (RCA) can be used to produce milligram amounts of a genomic nucleosomal DNA fragment in a scalable, one-pot reaction overnight. The protocol is efficient and flexible in choice of DNA sequence. It yields 10-fold more product than PCR, and rivals production using plasmids. We demonstrate the approach by producing the genomic DNA from the human LIN28B locus and show that it forms functional nucleosomes capable of binding pioneer transcription factor Oct4.

Chromatin state is highly dependent on the nucleosome binding proteins. Herein, we used a multipronged approach employing biophysical and in vivo experiments to characterize the effects of Nucleosome Binding Peptides (NBPeps) on nucleosome and cell activity. We performed a series of structure-based calculations on the nucleosome surface interaction with GMIP1 (a novel NBPep generated in silico), and HMGN2 (nucleosome binding motif of HMGN2), which contains sites that bind DNA and the acid patch, and also LANA and H4pep (nucleosome binding motif of H4 histone tail) that only bind to the acidic patch. Biochemical assays shows that H4pep, but not HMGN2, GMIP1 and LANA, is highly specific for targeting the nucleosome, with important effects on the final nucleosome structure and robust in vivo effects. These findings suggest that NBPeps might have important therapeutic implications and relevance as tools for chromatin investigation.

Rosa De Vincenzo,1 Carmine Conte,1 Caterina Ricci,1 Giovanni Scambia,1 Giovanni Capelli2 1Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, Catholic University of the Sacred Heart, Rome, 2Department of Human Sciences, Society and Health, University of Cassino and Southern Lazio, Cassino, Italy Abstract: In this paper, we review the published evidence about the long-term efficacy of the available human papillomavirus (HPV) vaccines and their safety profile. Two prophylactic HPV vaccines – bivalent (bHPV) and quadrivalent (qHPV) – are now available, and vaccination programs are being widely implemented, primarily targeting adolescent girls. Efficacy has been widely demonstrated for both vaccines. Since the risk of HPV exposure potentially persists throughout a woman’s sexual life, vaccine duration of protection is critical to overall effectiveness. Interpreting the results of long-term efficacy studies for the two HPV vaccines can be puzzling, due to the heterogeneity of studies, different methods used in the assessment of immunogenicity, histopathological and virological end points, and statistical power issues. Moreover, an immunologic correlate of protection has not yet been established, and it is unknown whether higher antibody levels will really result in a longer duration of protection. Disease prevention remains the most important measure of long-term duration of vaccine efficacy. To date, the longest follow-up of an HPV vaccine has been 9.4 years for the bHPV vaccine. Long-term follow-up for qHPV vaccine goes up to 8 years. The vaccine continues to be immunogenic and well tolerated up to 9 years following vaccination. All randomized controlled clinical trials of the bHPV and the qHPV vaccines provide evidence of an excellent safety profile. The most common complaint reported is pain in the injection site, which is self-limiting and spontaneously resolved. The incidence of systemic adverse events (AEs), serious AEs, and discontinuations due to a serious AE reported in clinical studies are similar between the two vaccines and their control groups. In particular, no increased risk of autoimmune disease has been shown among HPV-vaccinated subjects in long-term observation studies. As these are crucial topics in HPV vaccination, it is important to establish systems for continued monitoring of vaccine immunogenicity, efficacy, and safety over time. Keywords: HPV vaccines, effectiveness, adverse events

BACKGROUND: Locally advanced rectal cancer (LARC) is a heterogeneous group of tumors where a risk-adapted therapeutic strategy is needed. Short-course radiotherapy (SCRT) is a more convenient option for LARC patients than preoperative long-course RT plus capecitabine. Histone-deacetylase inhibitors (HDACi) have shown activity in combination with RT and chemotherapy in the treatment of solid tumors. Valproic acid (VPA) is an anti-epileptic drug with HDACi and anticancer activity. In preclinical studies, our group showed that the addition of HDACi, including VPA, to capecitabine produces synergistic antitumour effects by up-regulating thymidine phosphorylase (TP), the key enzyme converting capecitabine to 5-FU, and by downregulating thymidylate synthase (TS), the 5-FU target. METHODS/DESIGN: Two parallel phase-1 studies will assess the safety of preoperative SCRT (5 fractions each of 5 Gy, on days 1 to 5) combined with (a) capecitabine alone (increasing dose levels: 500-825 mg/m2/bid), on days 1-21, or (b) capecitabine as above plus VPA (oral daily day -14 to 21, with an intra-patient titration for a target serum level of 50-100 microg/ml) followed by surgery 8 weeks after the end of SCRT, in low-moderate risk RC patients. Also, a randomized phase-2 study will be performed to explore whether the addition of VPA and/or capecitabine to preoperative SCRT might increase pathologic complete tumor regression (TRG1) rate. A sample size of 86 patients (21-22/arm) was calculated under the hypothesis that the addition of capecitabine or VPA to SCRT can improve the TRG1 rate from 5% to 20%, with one-sided alpha = 0.10 and 80% power.Several biomarkers will be evaluated comparing normal mucosa with tumor (TP, TS, VEGF, RAD51, XRCC1, Histones/proteins acetylation, HDAC isoforms) and on blood samples (polymorphisms of DPD, TS, XRCC1, GSTP1, RAD51 and XRCC3, circulating endothelial and progenitors cells; PBMCs-Histones/proteins acetylation). Tumor metabolism will be measured by 18FDG-PET at baseline and 15 days after the beginning of SCRT. DISCUSSION: This project aims to improve the efficacy of preoperative treatment of LARC and to decrease the inconvenience and the cost of standard long-course RT. Correlative studies could identify both prognostic and predictive biomarkers and could add new insight in the mechanism of interaction between VPA, capecitabine and RT.EudraCT Number: 2012-002831-28. TRIAL REGISTRATION: ClinicalTrials.gov number, NCT01898104.

The authors describe a case-oriented seminar on managed care designed to teach psychiatric residents and medical students the principles of cost containment and delivery of quality care. The participants engage in role-play (reviewer, provider, appeal arbitrator) to elucidate the process. The response of the participants indicates that this seminar is a useful teaching model. Salient points regarding the content and process of the seminar are discussed in some detail.

In this paper, we review the published evidence about the long-term efficacy of the available human papillomavirus (HPV) vaccines and their safety profile. Two prophylactic HPV vaccines – bivalent (bHPV) and quadrivalent (qHPV) – are now available, and vaccination programs are being widely implemented, primarily targeting adolescent girls. Efficacy has been widely demonstrated for both vaccines. Since the risk of HPV exposure potentially persists throughout a woman’s sexual life, vaccine duration of protection is critical to overall effectiveness. Interpreting the results of long-term efficacy studies for the two HPV vaccines can be puzzling, due to the heterogeneity of studies, different methods used in the assessment of immunogenicity, histopathological and virological end points, and statistical power issues. Moreover, an immunologic correlate of protection has not yet been established, and it is unknown whether higher antibody levels will really result in a longer duration of protection. Disease prevention remains the most important measure of long-term duration of vaccine efficacy. To date, the longest follow-up of an HPV vaccine has been 9.4 years for the bHPV vaccine. Long-term follow-up for qHPV vaccine goes up to 8 years. The vaccine continues to be immunogenic and well tolerated up to 9 years following vaccination. All randomized controlled clinical trials of the bHPV and the qHPV vaccines provide evidence of an excellent safety profile. The most common complaint reported is pain in the injection site, which is self-limiting and spontaneously resolved. The incidence of systemic adverse events (AEs), serious AEs, and discontinuations due to a serious AE reported in clinical studies are similar between the two vaccines and their control groups. In particular, no increased risk of autoimmune disease has been shown among HPV-vaccinated subjects in long-term observation studies. As these are crucial topics in HPV vaccination, it is important to establish systems for continued monitoring of vaccine immunogenicity, efficacy, and safety over time.

Gamma-hydroxybutyric acid is an allegedly benign illicit substance that is gaining increasing recognition and attention among substance abusers and athletes. Alongside foreign-made brands, the compound is also easily available, at low cost because of the facility with which it can be produced in one's kitchen. Named by some “Nature's Quaalude” or sold as a health product, it is often used with a false sense of security as it may cause serious and disabling complications, as illustrated by this clinical vignette.

Objective : The authors describe demographic data, the distribution of diagnoses, and comorbid psychoactive substance use in a large sample of patients involuntarily admitted to a psychiatric hospital from multiple crisis centers and explore the relative roles these variables may play in service utilization and admission rates. Method : Data on demographic characteristics and comorbid psychoactive substance use in 2,200 consecutive involuntary hospital admissions of 1,755 psychiatric patients were gathered. Pertinent demographic and comorbidity data at first admission for the 1,755 patients, 314 of whom were admitted more than once, were analyzed ; then the data for the 1,441 single-admission patients and the data at first admission for the 314 patients who had multiple admissions were compared. Finally, the diagnostic distribution and comorbid psychoactive substance use in all 2,200 admissions were investigated, with attention to a subgroup of 88 high-risk patients (those with three or more admissions) who represented a total of 307 admissions. Results : Specific demographic characteristics were represented in the patient group at a high level of statistical significance. The diagnosis of schizophrenia was significantly overrepresented. Schizophrenia and psychosis not elsewhere classified clustered in the subgroup with a high risk of readmission. Conclusions: The results suggest a specific profile for the patient with heightened risk of hospital admission : a young, unmarried, African American male who has schizophrenia without comorbid substance abuse. An effect size data analysis identified marital status and a diagnosis of schizophrenia as the variables associated with the greatest likelihood of admission. Unexpectedly, the impact of comorbid psychoactive substance use was relatively modest and showed a uniform distribution among diagnostic groups.

The authors describe a sample of 815 acutely ill schizophrenia patients hospitalized under a petition for involuntary inpatient treatment and illustrate how this group of patients can be considered as more representative of involuntary commitment status than more traditional "involuntary" patients drawn from State hospitals' wards. Available demographics of the general population from which the sample was drawn are also presented for comparison. The authors then report the gender distribution of several variables--age, marital status, psychoactive substance use, comorbidity, length of stay, and frequency of readmissions--among the study sample and discuss the relevance of these findings.

We examined the levels of activity of methyl-p-hydroxyphenyl-lactate esterase (MeHPLA-ase) and cytosolic Type-II-estrogen-binding sites (Type-II EBS) in 61 and 71 cases, respectively, of primary ovarian cancer. MeHPLA-ase activity and Type-II EBS were seen to by asymmetrically distributed, in that levels were skewed towards the lower values. A statistically significant direct correlation was found between MeHPLA-ase activity and Type-II EBS. MeHPLA-ase activity and Type-II EBS were inversely correlated with ER and PR levels and showed a trend towards inverse correlation with the percentage of cells in S-phase of the cell cycle. MeHPLA-ase activity and Type-II EBS did not correlate with clinico-pathological parameters. The median MeHPLA-ase activity tended to be higher in responders than in unresponsive patients, but statistical significance was not reached. Higher Type-II-EBS levels were found in cases showing complete and partial response to chemotherapy than in cases which did not respond. A statistically significant relationship was found between high MeHPLA-ase activity and longer overall survival. © 1995 Wiley-Liss, Inc.

The bisdioxopiperazine ICRF 187 is a potent intracellular chelating agent which effectively diminishes Adriamycin cardiotoxicity without compromising its antitumor activity. Our study aimed at verifying whether ICRF 187 can modulate the cytotoxic action of cisplatin (CDDP) on ovarian cancer cells. We used the A2780 ovarian cancer cell line and a subline resistant to CDDP (A2780-CDDP) obtained in our laboratory by continuous exposure of the parenatal cells to progressively increasing CDDP doses. In both cell lines ICRF 187 (0.1-0.5 microgram/ml) used in combination with CDDP (0.01-1 microgram/ml) produced a dose-dependent reduction of CDDP IC50 (the concentration inhibiting 50% of cell growth). Moreover, when ICRF 187 was used in combination with CDDP, analysis of the data by the isobole method showed that the combination of the two drugs produced a synergistic antiproliferative activity in both cell lines, with a CDDP potentiation up to fivefold. Our in vitro data show that ICRF 187 can synergize with CDDP. Prospective clinical trials are now needed to verify whether the addition of ICRF 187 to CDDP-containing regimens will result in an improved clinical response in ovarian cancer.

Frequency and results of urine toxicology screens in a state-guided system of emergency rooms in a large city were compared with frequency and results of the same tests in the same population at a university-run intensive treatment psychiatric unit. Differences both in prevalence and in results were noted, with the emergency rooms testing significantly less patients than the intensive treatment unit and documenting a significantly lower rate of active substance abuse comorbidity to psychiatric disorders. Possible explanations for these differences are discussed, including clinical, attitudinal, and cost containment factors. These findings have a definite impact on treatment decision making, diagnostic precision, and validity of epidemiologic reports.

It has been demonstrated that the flavonoid quercetin (3,3',4',5,7-pentahydroxyflavone; Q) inhibits the growth of several cancer cell lines. There is evidence suggesting that the antiproliferative activity of this substance is mediated by the so-called type II estrogen-binding site (type II EBS). We looked for the presence of type II EBS and the effect of Q on the proliferation of an Adriamycin-resistant estrogen-receptor-negative human breast-cancer cell line (MCF-7 ADRr). By whole-cell assay using estradiol labelled with 6,7-tritium [( 3H]-E2) as a tracer, we demonstrated that MCF-7 ADRr cells contain type II EBSs. Competition analysis revealed that diethylstilbestrol (DES) and Q competed with similar potency for [3H]-Es binding to type II EBSs. The antiestrogen tamoxifen (TAM) competed for type II EBSs, albeit to a lesser extent than either DES or Q. Growth experiments demonstrated that Q and DES exerted a dose-dependent inhibition of cell proliferation in the range of concentrations between 10 nM and 10 microM, whereas TAM was less effective. Q could also inhibit colony formation in a clonogenic assay. Our results indicate that multidrug-resistant estrogen-receptor-negative MCF-7 cells express type II EBSs and are sensitive to the inhibitory effect of Q. This substance could be the parent compound of a novel class of anticancer agents.

Cubic SrTiO3 particles are obtained by precipitation from an aqueous gel suspension. The gel suspension is prepared by hydrolyzing a TiOCl2 solution with NaOH and adding SrCl2. The addition of citric acid leads to spherical particles. The particle size can be tailored in the range 80-1400 nm by varying the temperature and the concentration. A careful control of the synthesis conditions is essential for producing particles with a narrow size distribution. The results of HRTEM and ED investigations provide strong evidence that the oriented aggregation of small (4-5 nm) nanocrystals is the dominant growth mechanism for the formation of the observed SrTiO3 particles. The primary nanocrystals self-assemble in a highly oriented fashion, producing defective single-crystal particles. The above results show that the directional aggregation process can be controlled by changing the temperature and concentration of the suspension as well as by adding organic molecules, obtaining SrTiO3 particles with a controlled size and shape.

The expression of transforming growth factor alpha (TGFalpha), amphiregulin (AR) and cripto-1 (CR-1) was assessed by immunohistochemistry in 83 specimens (59 primary ovarian tumors and 24 extra-ovarian carcinomas) that were obtained from 68 ovarian carcinoma patients. Within the 59 primary tumors, 54 (92%) expressed immunoreactive TGFalpha, 45 (76%) expressed AR, and 28 (47%) expressed CR-1. The expression of AR and CR-1 mRNAs in the ovarian carcinomas was also demonstrated by RT-PCR analysis. Seventeen extra-ovarian specimens (71%) were found to express CR-1, whereas AR and TGFalpha were expressed respectively in 21 (87%) and 22 (92%) extra-ovarian tissues. In 15 cases for whom both ovarian and extra-ovarian tissues were available, a statistically significant higher expression of CR-1 was found in extra-ovarian specimens. A statistically significant correlation was found between AR expression in the ovarian carcinomas and both low grade and low proliferative activity. Finally, expression of TGFalpha was predictive of longer progression-free survival. These data strongly suggest that the EGF-related peptides might be involved in the pathogenesis and outcome of human ovarian cancer.

Several programs that have had varying success in providing rapid intervention in the treatment of acutely mentally ill patients have been dcscribed in the last decade (1-3). Such programs often constitute a muchneeded bridge to ongoing psychiatric treatment and services in the public sector for chronic mentally ill patients. This paper describes a short-term intensive psychiatric treatment unit at a general hospital in Philadelphia and provides data on the number of admissions and their disposition in the first 35 months ofthc program’s operation. The unit was a joint effort of the city’s Office ofMental Health and the department of psychiatry at Jefferson Medical College.

A new series of N-deacetyl-N-(N-trifluoroacetylaminoacyl)thiocolchicine derivatives 9-15 have been synthesized starting from the corresponding N-deacetylthiocolchicine (3) and the N-trifluoroacetylamino acids 5-8 which were used as a racemic mixture. The trifluoroacetyl protecting group has been removed easily, giving the corresponding N-deacetyl-N-aminoacylthiocolchicines 16-22. Optical pure compounds 9-22 were isolated from the diastereoisomeric mixture or were prepared starting from compound 3 and an optical pure amino acid derivative; the configuration of each compound was assigned unequivocally. The diastereoisomeric couples of amino acids synthesized were tested, and their antiproliferative activity on MDR-positive and MDR-negative human cancer cell lines was evaluated.