Detoxification of toxin A and toxin B by copper ion-catalyzed oxidation in production of a toxoid-based vaccine against Clostridioides difficile.

Clostridioides difficile infections (CDI) has emerged worldwide as a serious antimicrobial-resistant healthcare-associated disease resulting in diarrhea and pseudomembranous colitis. The two cytotoxic proteins, toxin A (TcdA) and toxin B (TcdB) are the major virulence factor responsible for the disease symptoms. We examined time-dependent oxidative detoxification of TcdA and TcdB using different molar ratios of protein:Cu2+:H2O2. The metal-catalyzed oxidation (MCO) reaction in molar ratios of 1:60:1000 for protein:Cu2+:H2O2 at pH 4.5 resulted in a significant 6 log10 fold reduction in cytotoxicity after 120-min incubation at 37 °C. Circular dichroism revealed that MCO-detoxified TcdA and TcdB had secondary and tertiary structural folds similar to the native proteins. The conservation of immunogenic epitopes of both proteins was tested using monoclonal antibodies in an ELISA, comparing our MCO-detoxification approach to a conventional formaldehyde-detoxification method. The oxidative detoxification of TcdA and TcdB led to an average 2-fold reduction in antibody binding relative to native proteins, whereas formaldehyde cross-linking resulted in 3-fold and 5-fold reductions, respectively. Finally, we show that mice immunized with a vaccine consisting of MCO-detoxified TcdA and TcdB were fully protected against disease symptoms and death following a C. difficile infection and elicited substantial serum IgG responses against both TcdA and TcdB. The results of this study present copper ion-catalyzed oxidative detoxification of toxic proteins as a method highly suitable for the rapid production of safe, immunogenic and irreversible toxoid antigens for future vaccine development and may have the potential for replacing cross-linking reagents like formaldehyde.

Mitral valve navigator. A new diagnostic tool for effective regurgitant orifice quantification in mitral regurgitation.

INTRODUCTION: Mitral regurgitation severity assessment is usually carried out using qualitative, semiquantitative, and quantitative parameters. The mitral valve navigation (MVN) tool allows to measure the mitral effective regurgitant orifice (MERO) from 3D echo datasets. Our aim was to validate the MVN as a new tool to quantify MERO. A secondary aim was to assess the intra- and interobserver variability. METHODS: This is a retrospective study in which consecutive subjects undergoing a transoesophageal echocardiogram for more than mild mitral regurgitation evaluation were included. MERO measurement obtained by means of 3D color Doppler was used as the gold standard method for comparison. In every patient, MERO was also obtained using the MVN tool. RESULTS: Fifty-nine consecutive patients were analyzed (47.5% female; mean age 50.8 years). Mitral regurgitation was moderate in 23 (39%) and severe in 36 (61%) patients. Forty patients (67.8%) had a primary and 19 (32.2%) a secondary mitral regurgitation. The intraclass correlation coefficient (ICC) between 3D color Doppler and MVN was excellent (ICC: 0.95; 95% CI: 0.82 to 0.98; P < 0.001) in the total group and for patients with primary and secondary mitral regurgitation. Intra- and interobserver agreements were also good. CONCLUSIONS: Mitral valve navigator shows an excellent accuracy for measuring MERO when the transoesophageal 3D color Doppler is used as the reference method, either primary or secondary mitral regurgitation. Intraobserver reproducibility and interobserver reproducibility are also excellent. These findings make this software a good alternative method to measure mitral regurgitation severity.

[Factors associated with success of a smoking cessation program]. / Factores asociados con el éxito de la terapia antitabaco en pacientes tratados con vareniclina: 10 años de experiencia en un programa multidisciplinario.

BACKGROUND: Smoking cessation therapies include counseling, psychological management and pharmacological therapy. Varenicline is the most effective and safe medication available. AIM: To study risk factors for the failure of pharmacological smoking cessation therapy with varenicline. PATIENTS AND METHODS: Retrospective analysis of 281 patients aged 45 ± 11 years (65% males) with a mean consumption of 31 ± 22 packs/year. They completed a smoking cessation program comprising psychological support and use of varenicline in a private clinic. Patients were followed with telephonic interviews during one year. A complete abstinence during one year was considered as a success of the program. RESULTS: The success rate of the program was 53.4%. The factors associated with failure were a high tobacco dependence rate determined with the Fageström test (Odds ratio (OR) 2.47, 95% confidence intervals (CI) 1.16-5.26, p = 0.02). An instruction level of more than 12 years was associated with a lower failure rate (OR 0.38 95% CI 0.18-0.82). CONCLUSIONS: A high tobacco dependence rate and a lower education were associated with a higher failure rate of this smoking cessation program.

Factores asociados con el éxito de la terapia antitabaco en pacientes tratados con vareniclina: 10 años de experiencia en un programa multidisciplinario / Factors associated with success of a smoking cessation program

Background: Smoking cessation therapies include counseling, psychological management and pharmacological therapy. Varenicline is the most effective and safe medication available. Aim: To study risk factors for the failure of pharmacological smoking cessation therapy with varenicline. Patients and Methods: Retrospective analysis of 281 patients aged 45 ± 11 years (65% males) with a mean consumption of 31 ± 22 packs/year. They completed a smoking cessation program comprising psychological support and use of varenicline in a private clinic. Patients were followed with telephonic interviews during one year. A complete abstinence during one year was considered as a success of the program. Results: The success rate of the program was 53.4%. The factors associated with failure were a high tobacco dependence rate determined with the Fageström test (Odds ratio (OR) 2.47, 95% confidence intervals (CI) 1.16-5.26, p = 0.02). An instruction level of more than 12 years was associated with a lower failure rate (OR 0.38 95% CI 0.18-0.82). Conclusions: A high tobacco dependence rate and a lower education were associated with a higher failure rate of this smoking cessation program.

[Results of a multidisciplinary program to quit smoking]. / Resultados de un programa multidisciplinario para el control del hábito tabáquico.

BACKGROUND: Therapies to quit smoking are based on counseling, psychological therapy (PT), nicotine replacement therapy, bupropion or varenidine. AIM: To report the results of a multidisciplinary program to quit smoking. MATERIAL AND METHODS: Patients aged l8 years or more, motivated to quit smoking were admitted in a program based in counseling and PT, with or without pharmacological therapy. They were assessed by telephone during one year of follow up. Patients with unstable psychiatric diseases were excluded. Results were considered as "successful" when patients maintained abstinence during the year of follow up. A logistic regression analysis was done to identify factors associated with treatment success. RESULTS: Between 2005 and 2011, 198 patients aged 45 ± 11 years (56% males), who smoked 31.5 ± 20.6 packages/year, were treated. Of these, 155 (78%) were treated with varenidine, 26 (13%) with bupropion and 17 (9%>) did not receive pharmacological therapy. One hundred sixty eight patients completed the year of follow up. In 82 (49%>), treatment was successful and was negatively associated with a history of depression (odds ratio = 4 (95% confidence intervals 1.23-38.33). The main side effeets associated to varenidine and bupropion were nausea in 37 and 23%o, sleep disorders in 20 and 19%o and headache in 12 and 0%>, respectively. CONCLUSIONS: A multidisciplinary program to quit smoking achieved a 49%> of abstinence during a year of follow up.

Resultados de un programa multidisciplinario para el control del hábito tabáquico / Results of a multidisciplinary program to quit smoking

Background: Therapies to quit smoking are based on counseling, psychological therapy (PT), nicotine replacement therapy, bupropion or varenidine. Aim: To report the results of a multidisciplinary program to quit smoking Material and Methods: Patients agedl8years or more, motivated to quit smoking were admitted in a program based in counseling and PT, with or without pharmacological therapy. They were assessed by telephone during one year offollow up. Patients with unstable psychiatric diseases were excluded. Results were considered as "successful" when patients maintained abstinence during the year offollow up. A logistic regression analysis was done to identify factors associated with treatment success. Results: Between 2005 and 2011, 198 patients aged 45 ± 11 years (56% males), who smoked 31.5 ± 20.6 packages/year, were treated. Ofthese, 155 (78%) were treated with varenidine, 26 (13%) with bupropion and 17 (9%>) did not receive pharmacological therapy. One hundred sixty eightpatients completed the year offollow up. In 82 (49%>), treatment was successful and was negatively associated with a history of depression (odds ratio = 4 (95% confidence intervals 1.23-38.33). The main side effeets associated to varenidine and bupropion were nausea in 37 and 23%o, sleep disorders in 20 and 19%o and headache in 12 and 0%>, respectively Conclusions: A multidisciplinary program to quit smoking achieved a 49%> of abstinence during a year offollow up.

Effect of subcutaneous lidocaine infiltration on blood loss secondary to corporal lipoaspiration: a prospective, randomized, double-masked clinical trial.

BACKGROUND: Lipoaspiration is one of the most frequently performed aesthetic surgical procedures worldwide. The use of tumescent solution containing lidocaine to infiltrate subcutaneous fat before surgery has been accepted as the standard of care for these procedures. Its objective is to diminish postoperative analgesic necessities and secondary blood loss, but its role in hematocrit reduction is not clear at this time. This study aimed to measure the effect of subcutaneous lidocaine infiltration on blood loss secondary to corporal liposuction. METHODS: A prospective, randomized, double-masked clinical trial was performed. Between November 2005 and July 2007, 70 consecutive female patients submitted to corporal liposuction as a single surgery were included in the study. All the patients were randomly assigned to two groups. The study group received tumescent solution containing lidocaine and epinephrine, whereas the control group received tumescent solution containing only epinephrine. Hematocrit was measured preoperatively, then 2 to 4, 12, and 24 h postoperatively. RESULTS: The comparison between the two groups did not show significant statistical differences with respect to age, body mass index, infiltrated volume, aspirated fat volume, surgical time, hospital stay, preoperative hematocrit, or comparison of the areas lipoaspirated. The mean hematocrit reduction in the study group was 10.8% +/- 2.9% compared with 8.8% +/- 2.6% in the control group (P = 0.004). CONCLUSION: The use of lidocaine in the tumescent solution for subcutaneous fat infiltration significantly increases postoperative anemia compared with the results obtained when epinephrine infiltration alone is used.

Targeted disruption of Dkc1, the gene mutated in X-linked dyskeratosis congenita, causes embryonic lethality in mice.

Dyskeratosis congenita (DC) is an inherited bone marrow failure syndrome associated with increased cancer susceptibility. The X-linked form is due to mutations in the DKC1 gene encoding dyskerin, a nucleolar protein predicted to be involved in rRNA processing and associated with the telomerase complex. Available evidence suggests the pathology of DC is due to telomerase defects. We have used the inducible Cre/loxP system to produce deletions in the murine Dkc1 gene in early embryogenesis. A large deletion lacking exons 12-15 and a small deletion lacking only the last exon, were produced. We found both deletions showed a parent-of-origin effect with 100% embryonic lethality when the mutation occurred on the maternal Dkc1. Embryonic analysis at day E7.5 and E9.5 showed no male embryos carrying either deletion whereas females with maternally derived deletions died around day E9.5, with degeneration of the extra embryonic tissue, in which the paternal X-chromosome is inactivated. Female mice carrying the deletion in the paternally derived Dkc1 show extreme skewing of X-inactivation with the wild type X-chromosome active in all cells. Since mice with no telomerase are viable in the first generations the lethality we observe is unlikely to be due to the effects of mutated dyskerin on telomerase activity.