Analysis of ATG8 Family Members Using LC3-Interacting Regions (LIR)-Based Molecular Traps.

The ATG8 family of proteins regulates the autophagy process from the autophagosome maturation and cargo recruitment up to degradation. Autophagy dysfunction is involved in the development of multiple diseases. The LC3 interacting region (LIR)-based molecular traps have been designed to isolate endogenous ATG8 proteins and their interactors in order to facilitate the study of selective autophagy events. Here, we summarize protocols describing LC3 traps and sample preparation as well as adaptations for the analysis of ATG8 proteins in different biological models. This protocol was optimized to prepare affinity columns, reduce background, and improve the protein recovery to be analyzed by immunodetection with antibodies recognizing proteins of interest.

Exploring selective autophagy events in multiple biologic models using LC3-interacting regions (LIR)-based molecular traps.

Autophagy is an essential cellular pathway that ensures degradation of a wide range of substrates including damaged organelles or large protein aggregates. Understanding how this proteolytic pathway is regulated would increase our comprehension on its role in cellular physiology and contribute to identify biomarkers or potential drug targets to develop more specific treatments for disease in which autophagy is dysregulated. Here, we report the development of molecular traps based in the tandem disposition of LC3-interacting regions (LIR). The estimated affinity of LC3-traps for distinct recombinant LC3/GABARAP proteins is in the low nanomolar range and allows the capture of these proteins from distinct mammalian cell lines, S. cerevisiae and C. elegans. LC3-traps show preferences for GABARAP/LGG1 or LC3/LGG2 and pull-down substrates targeted to proteaphagy and mitophagy. Therefore, LC3-traps are versatile tools that can be adapted to multiple applications to monitor selective autophagy events in distinct physiologic and pathologic circumstances.

[Evaluation of Western Blot method for the detection of antibodies to Helicobacter pylori antigens in patients with gastric carcinoma and cases with epigastric complaints]. / Gastrik karsinomali hastalar ile epigastrik yakinmalari olan olgularda Helicobacter pylori antijenlerine karsi antikor varliginin saptanmasinda western blot yönteminin degerlendirilmesi.

Helicobacter pylori proteins CagA (cytotoxin-associated gene A) and VacA (vacuolating cytotoxin A) are among the virulence factors of this species. CagA gene carrying H. pylori strains are particularly associated with gastric adenocarsinoma. This study was conducted to evaluate Western Blot (WB) method to determine specific H. pylori antibodies in a group of patients with gastric cancer and in a control group with no malignancy. A total of 99 patients with gastric cancer (94 adenocarcinoma, 2 adenosquamous cell carcinoma, 3 non-Hodgkin lymphoma) and 150 control cases with epigastric complaints such as nausea, vomiting, diarrhea, gastroesophageal reflux and abdominal pain, were included to the study. H. pylori IgG-ELISA was positive in all study (mean age: 56.7 +/- 1.2 years, 62 male) and control (mean age: 24.2 +/- 1.3 years, 64 male) patients. Specific antibodies against CagA, VacA, OMP (outer membrane protein)-67, urease-A, urease-B, HSP (heat shock protein) and flagellin antigens determined by a commercial WB-based kit (RIDA Blot Helicobacter, R-Biopharm GmbH, Germany). Interestingly, no anti-VacA positivity was detected in none of the patient and control groups. The positivity rates for H. pylori CagA, OMP-67, urease A, urease-B, flagellin and HSP specific antibodies were as 78%, 54%, 37%, 60%, 53% and 82% in the gastric cancer group and 85%, 71%, 55%, 43%, 61% and 75% in the control group, respectively. There was no statistically significant difference (p > 0.05) between gastric carcinoma and control groups in terms of CagA, HSP and flagellin antibodies (p > 0.05). On the other hand, a statistically significant difference was found between the 2 groups in terms of urease-A, urease-B and OMP-67 (p < 0.01). These results suggested that this test should be assessed again by the manufacturer for its detection power directed towards specific H. pylori antibodies, especially for Vac-A. Further molecular and clinical studies are necessary to determine the factors that affect H. pylori virulence and disease prognosis.

The regulation of mitochondrial homeostasis by the ubiquitin proteasome system.

From mitochondrial quality control pathways to the regulation of specific functions, the Ubiquitin Proteasome System (UPS) could be compared to a Swiss knife without which mitochondria could not maintain its integrity in the cell. Here, we review the mechanisms that the UPS employs to regulate mitochondrial function and efficiency. For this purpose, we depict how Ubiquitin and the Proteasome participate in diverse quality control pathways that safeguard entry into the mitochondrial compartment. A focus is then achieved on the UPS-mediated control of the yeast mitofusin Fzo1 which provides insights into the complex regulation of this particular protein in mitochondrial fusion. We ultimately dissect the mechanisms by which the UPS controls the degradation of mitochondria by autophagy in both mammalian and yeast systems. This organization should offer a useful overview of this abundant but fascinating literature on the crosstalks between mitochondria and the UPS.

Transcatheter versus Surgical Aortic Valve Replacement after Previous Cardiac Surgery: A Systematic Review and Meta-Analysis.

AIM: Aortic valve replacement (AVR) in patients with prior cardiac surgery might be challenging. Transcatheter aortic valve replacement (TAVR) offers a promising alternative in such patients. We therefore aimed at comparing the outcomes of patients with aortic valve diseases undergoing TAVR versus those undergoing surgical AVR (SAVR) after previous cardiac surgery. METHODS AND RESULTS: MEDLINE, EMBASE, and the Cochrane Central Register were searched. Seven relevant studies were identified, published between 01/2011 and 12/2015, enrolling a total of 1148 patients with prior cardiac surgery (97.6% prior CABG): 49.2% underwent TAVR, whereas 50.8% underwent SAVR. Incidence of stroke (3.8 versus 7.9%, p=0.04) and major bleeding (8.3 versus 15.3%, p=0.04) was significantly lower in the TAVR group. Incidence of mild/severe paravalvular leakage (14.4/10.9 versus 0%, p < 0.0001) and pacemaker implantation (11.3 versus 3.9%, p=0.01) was significantly higher in the TAVR group. There were no significant differences in the incidence of acute kidney injury (9.7 versus 8.7%, p=0.99), major adverse cardiovascular events (8.7 versus 12.3%, p=0.21), 30-day mortality (5.1 versus 5.5%, p=0.7), or 1-year mortality (11.6 versus 11.8%, p=0.97) between the TAVR and SAVR group. CONCLUSIONS: TAVR as a redo procedure offers a safe alternative for patients presenting with aortic valve diseases after previous cardiac surgery especially those with prior CABG.

Impact of previous cardiac surgery in patients undergoing transcatheter aortic valve implantation: a systematic review.

INTRODUCTION: Redo surgical aortic valve replacement after prior cardiac surgery is usually related to a higher risk of mortality and morbidity. Transcatheter aortic valve implantation (TAVI) became an alternative therapy for those patients in the past couple of years. EVIDENCE ACQUISITION: We aimed in this study to analyze the outcomes of patients undergoing TAVI after a prior cardiac surgery especially those who underwent coronary artery bypass grafting (CABG) and to see if TAVI offers any advantages for those patients than conventional surgical aortic valve replacement. EVIDENCE SYNTHESIS: We searched for relevant articles in Medline and abstracted clinical information based on pre-defined criteria and endpoints. Data of nine studies including the baseline characteristics, implantation data, postoperative outcomes and major adverse cardiac complications, which were published between 2011 and 2015 were collected and evaluated. From all reviewed studies, 769 patients had a prior cardiac surgery and underwent TAVI for symptomatic severe aortic stenosis. Of these, 738 patients (96%) had prior CABG. Patients' age ranged from 78±3 to 82±5.8 years. The STS and EuroSCORE ranged from 4.5±3% to 14.7±12.3% and 25.6±16.2% to 37±18%, respectively. In all reviewed studies the 30-day mortality was about 5.6% and was not significantly higher compared to patients with no history of prior cardiac surgery. The total incidence of stroke was about 3.6%, myocardial infarction was 1.7%, acute kidney injury was 13.8% and permanent pacemaker implantation was about 14.2%. CONCLUSIONS: However, patients presented with severe aortic valve disease after a previous cardiac surgery exhibited a higher preoperative STS and EuroSCORE than those without previous cardiac surgery. The 30-day mortality was not significantly higher in comparison to those patients without history of prior cardiac surgery. According to that, transcatheter aortic valve implantation should be considered as an attractive alternative for those patients.

Propensity score analysis of outcomes following minimal access versus conventional aortic valve replacement.

OBJECTIVES: Minimal access aortic valve replacement has become routine in many institutions. Aim of this study was to compare the clinical outcomes between conventional and minimal access aortic valve replacement. METHODS: We retrospectively analysed the data of 2103 patients who underwent primary, isolated aortic valve replacement (AVR) in our institution between January 2001 and May 2012 with a minimal access AVR (MAAVR) via the upper partial ministernotomy approach (n = 936) or conventional AVR (CAVR) via the full sternotomy approach (n = 1167). After propensity score matching considering potential confounders [age, sex (female), weight, height, preoperative serum creatinine level, previous myocardial infarction, LV-EF and aortic valve pathology (isolated AS)], 585 matched patients were included in each group. RESULTS: Mean age (65 ± 10.5 vs 65.7 ± 11.5 years, P = 0.23), gender (females 37.2%, P = 0.9), aortic cross-clamp time (65.6 ± 18.4 vs 64.3 ± 19.8 min, P = 0.25) and postoperative blood loss [median (IQR) 400 (224-683) vs 400 (250-610) ml, P = 0.83) were similar in MAAVR and CAVR group. Thirty-day mortality was also not significantly different (1.5 vs 1.7%, P = 0.74, respectively). In contrast, CPB times were significantly longer in MAAVR (93.5 ± 25 vs 88 ± 28 min, P < 0.001). Intraoperative and postoperative autologous blood transfusions were significantly lower in MAAVR (927.2 ± 425.6 vs 1036.4 ± 599.6 ml, P < 0.001 and 170.2 ± 47.6 vs 243.5 ± 89.3 ml, P < 0.001, respectively). Intubation time was significantly shorter in MAAVR [median (IQR) 7 (5-11) vs 8 (6-14) h, P = 0.01). The incidence of renal insufficiency (creatinine ≥1.5 mg/dl) and respiratory insufficiency (need for non-invasive ventilation, reintubation or tracheotomy) was significantly lower in MAAVR (9 vs 16%, P < 0.001 and 8.5 vs 11.8%, P = 0.03, respectively). CONCLUSIONS: In comparison with CAVR, our study shows that MAAVR is a safe and effective procedure associated with low mortality rate and good long-term survival rates. In addition to that, MAAVR was associated with shorter ventilation times, lower rate of autologous blood transfusion, as well as a lower rate of postoperative respiratory and renal insufficiency. Because of the superior cosmetic results, we therefore advocate MAAVR as the procedure of choice for primary isolated AVR.

[Investigation of intestinal parasites in students of Mustafa Cengiz primary school in Van]. / Van mustafa cengiz ilkögretim okulu ögrencilerinde bagirsak parazitlerinin arastirilmasi

Intestinal parasites still maintain as a major public health problem in our country. In this study, we aimed to investigate the distribution of intestinal parasites in 1st and 2nd grade students of Mustafa Cengiz Primary School, aged between 7-9 and to contribute to the parasitological data of our province. For this purpose, stool examinations of a total of 195 students, including 82 boys and 113 girls, were performed. The results of the microscopic analysis of stool samples revealed one or more parasites in a total of 117 (60%) samples including 45 male students (54.8%) and 72 female students (63.7%). The diagnosed parasites and their ratios in children were; Giardia intestinalis 36.4%, Entamoeba coli 17.9%, Blastocystis hominis 14.4%, Hymenolepis nana 10.8%, Chilomastix mesnili 3.6%, Ascaris lumbricoides 2.6%, Entamoeba hartmanni 1.5%, Trichuris trichiura 1%, Iodamoeba butschlii 0.5%, Retortamonas intestinalis 0.5% ve Endolimax nana 0.5%, respectively. From 117 positive samples for parasites, only one parasite was found in 71 (60.7%), and more than one parasites were found in 46 (39.3%). As a result, parasitic infectious diseases still maintain its importance in our region. We conclude that incidence of parasitic infectious diseases will be reduced with education about personal hygiene and improvement of physical conditions.