Neuronal-specific septin-3 binds Atg8/LC3B, accumulates and localizes to autophagosomes during induced autophagy.

In synapses that show signs of local apoptosis and mitochondrial stress and undergo neuro-immunological synapse pruning, an increase in the levels of the presynaptic protein, neuronal-specific septin-3 can be observed. Septin-3 is a member of the septin GTPase family with the ability to form multimers and contribute to the cytoskeleton. However, the function of septin-3 remains elusive. Here, we provide evidence that septin-3 is capable of binding the most-studied autophagy protein Atg8 homolog microtubule-associated protein 1 light chain 3B (LC3B), besides another homolog, GABA receptor-associated protein-like 2 (GABARAPL2). Moreover, we demonstrate that colocalization of septin-3 and LC3B increases upon chemical autophagy induction in primary neuronal cells. Septin-3 is accumulated in primary neurons upon autophagy enhancement or blockade, similar to autophagy proteins. Using electron microscopy, we also show that septin-3 localizes to LC3B positive membranes and can be found at mitochondria. However, colocalization results of septin-3 and the early mitophagy marker PTEN-induced kinase 1 (PINK1) do not support that binding of septin-3 to mitochondria is mitophagy related. We conclude that septin-3 correlates with synaptic/neuronal autophagy, binds Atg8 and localizes to autophagic membranes that can be enhanced with chemical autophagy induction. Based on our results, elevated septin-3 levels might indicate enhanced or impeded autophagy in neurons.

The Effect of Sleep Deprivation and Subsequent Recovery Period on the Synaptic Proteome of Rat Cerebral Cortex.

Sleep deprivation (SD) is commonplace in the modern way of life and has a substantial social, medical, and human cost. Sleep deprivation induces cognitive impairment such as loss of executive attention, working memory decline, poor emotion regulation, increased reaction times, and higher cognitive functions are particularly vulnerable to sleep loss. Furthermore, SD is associated with obesity, diabetes, cardiovascular diseases, cancer, and a vast majority of psychiatric and neurodegenerative disorders are accompanied by sleep disturbances. Despite the widespread scientific interest in the effect of sleep loss on synaptic function, there is a lack of investigation focusing on synaptic transmission on the proteome level. In the present study, we report the effects of SD and recovery period (RP) on the cortical synaptic proteome in rats. Synaptosomes were isolated after 8 h of SD performed by gentle handling and after 16 h of RP. The purity of synaptosome fraction was validated with western blot and electron microscopy, and the protein abundance alterations were analyzed by mass spectrometry. We observed that SD and RP have a wide impact on neurotransmitter-related proteins at both the presynaptic and postsynaptic membranes. The abundance of synaptic proteins has changed to a greater extent in consequence of SD than during RP: we identified 78 proteins with altered abundance after SD and 39 proteins after the course of RP. Levels of most of the altered proteins were upregulated during SD, while RP showed the opposite tendency, and three proteins (Gabbr1, Anks1b, and Decr1) showed abundance changes with opposite direction after SD and RP. The functional cluster analysis revealed that a majority of the altered proteins is related to signal transduction and regulation, synaptic transmission and synaptic assembly, protein and ion transport, and lipid and fatty acid metabolism, while the interaction network analysis revealed several connections between the significantly altered proteins and the molecular processes of synaptic plasticity or sleep. Our proteomic data implies suppression of SNARE-mediated synaptic vesicle exocytosis and impaired endocytic processes after sleep deprivation. Both SD and RP altered GABA neurotransmission and affected protein synthesis, several regulatory processes and signaling pathways, energy homeostatic processes, and metabolic pathways.

Chronic stepwise cerebral hypoperfusion differentially induces synaptic proteome changes in the frontal cortex, occipital cortex, and hippocampus in rats.

During chronic cerebral hypoperfusion (CCH), the cerebral blood flow gradually decreases, leading to cognitive impairments and neurodegenerative disorders, such as vascular dementia. The reduced oxygenation, energy supply induced metabolic changes, and insufficient neuroplasticity could be reflected in the synaptic proteome. We performed stepwise bilateral common carotid occlusions on rats and studied the synaptic proteome changes of the hippocampus, occipital and frontal cortices. Samples were prepared and separated by 2-D DIGE and significantly altered protein spots were identified by HPLC-MS/MS. We revealed an outstanding amount of protein changes in the occipital cortex compared to the frontal cortex and the hippocampus with 94, 33, and 17 proteins, respectively. The high alterations in the occipital cortex are probably due to the hypoxia-induced retrograde degeneration of the primary visual cortex, which was demonstrated by electrophysiological experiments. Altered proteins have functions related to cytoskeletal organization and energy metabolism. As CCH could also be an important risk factor for Alzheimer's disease (AD), we investigated whether our altered proteins overlap with AD protein databases. We revealed a significant amount of altered proteins associated with AD in the two neocortical areas, suggesting a prominent overlap with the AD pathomechanism.

Synaptic mitochondrial dysfunction and septin accumulation are linked to complement-mediated synapse loss in an Alzheimer's disease animal model.

Synaptic functional disturbances with concomitant synapse loss represent central pathological hallmarks of Alzheimer's disease. Excessive accumulation of cytotoxic amyloid oligomers is widely recognized as a key event that underlies neurodegeneration. Certain complement components are crucial instruments of widespread synapse loss because they can tag synapses with functional impairments leading to their engulfment by microglia. However, an exact understanding of the affected synaptic functions that predispose to complement-mediated synapse elimination is lacking. Therefore, we conducted systematic proteomic examinations on synaptosomes prepared from an amyloidogenic mouse model of Alzheimer's disease (APP/PS1). Synaptic fractions were separated according to the presence of the C1q-tag using fluorescence-activated synaptosome sorting and subjected to proteomic comparisons. The results raised the decline of mitochondrial functions in the C1q-tagged synapses of APP/PS1 mice based on enrichment analyses, which was verified using flow cytometry. Additionally, proteomics results revealed extensive alterations in the level of septin protein family members, which are known to dynamically form highly organized pre- and postsynaptic supramolecular structures, thereby affecting synaptic transmission. High-resolution microscopy investigations demonstrated that synapses with considerable amounts of septin-3 and septin-5 show increased accumulation of C1q in APP/PS1 mice compared to the wild-type ones. Moreover, a strong positive correlation was apparent between synaptic septin-3 levels and C1q deposition as revealed via flow cytometry and confocal microscopy examinations. In sum, our results imply that deterioration of synaptic mitochondrial functions and alterations in the organization of synaptic septins are associated with complement-dependent synapse loss in Alzheimer's disease.

Identification of Neuronal Pentraxins as Synaptic Binding Partners of C1q and the Involvement of NP1 in Synaptic Pruning in Adult Mice.

Elements of the immune system particularly that of innate immunity, play important roles beyond their traditional tasks in host defense, including manifold roles in the nervous system. Complement-mediated synaptic pruning is essential in the developing and healthy functioning brain and becomes aberrant in neurodegenerative disorders. C1q, component of the classical complement pathway, plays a central role in tagging synapses for elimination; however, the underlying molecular mechanisms and interaction partners are mostly unknown. Neuronal pentraxins (NPs) are involved in synapse formation and plasticity, moreover, NP1 contributes to cell death and neurodegeneration under adverse conditions. Here, we investigated the potential interaction between C1q and NPs, and its role in microglial phagocytosis of synapses in adult mice. We verified in vitro that NPs interact with C1q, as well as activate the complement system. Flow cytometry, immunostaining and co-immunoprecipitation showed that synapse-bound C1q colocalizes and interacts with NPs. High-resolution confocal microscopy revealed that microglia-surrounded C1q-tagged synapses are NP1 positive. We have also observed the synaptic occurrence of C4 suggesting that activation of the classical pathway cannot be ruled out in synaptic plasticity in healthy adult animals. In summary, our results indicate that NPs play a regulatory role in the synaptic function of C1q. Whether this role can be intensified upon pathological conditions, such as in Alzheimer's disease, is to be disclosed.

The short- and long-term proteomic effects of sleep deprivation on the cortical and thalamic synapses.

Acute total sleep deprivation (SD) impairs memory consolidation, attention, working memory and perception. Structural, electrophysiological and molecular experimental approaches provided evidences for the involvement of sleep in synaptic functions. Despite the wide scientific interest on the effects of sleep on the synapse, there is a lack of systematic investigation of sleep-related changes in the synaptic proteome. We isolated parietal cortical and thalamic synaptosomes of rats after 8h of total SD by gentle handling and 16h after the end of deprivation to investigate the short- and longer-term effects of SD on the synaptic proteome, respectively. The SD efficiency was verified by electrophysiology. Protein abundance alterations of the synaptosomes were analyzed by fluorescent two-dimensional differential gel electrophoresis and by tandem mass spectrometry. As several altered proteins were found to be involved in synaptic strength regulation, our data can support the synaptic homeostasis hypothesis function of sleep and highlight the long-term influence of SD after the recovery sleep period, mostly on cortical synapses. Furthermore, the large-scale and brain area-specific protein network change in the synapses may support both ideas of sleep-related synaptogenesis and molecular maintenance and reorganization in normal rat brain.

Comparison of working length control consistency between hand K-files and Mtwo NiTi rotary system.

BACKGROUND: The purpose of this study was to investigate the consistency of working length control between hand instrumentation in comparison to engine driven Mtwo nickel-titanium rotary files. MATERIAL AND METHODS: Forty extracted maxillary molars were selected and divided onto two parallel groups. The working lengths of the mesiobuccal root canals were estimated. The teeth were fixed in a phantom head. The root canal preparation was carried out group 1 (n=20) with hand K-files, (VDW, Munich, Germany) and group 2 (n=20) with Mtwo instruments (VDW, Munich, Germany). Vestibulo-oral and mesio-distal directional x-ray images were taken before the preparation with #10 K-file, inserted into the mesiobuccal root canal to the working length, and after preparation with #25, #30 and #40 files. Working lenght changes were detected with measurements between the radiological apex and the instrument tips. RESULTS: In the Mtwo group a difference in the working competency (p<0.05) could be noticed only in the vestibulo-oral direction from #10 to #40 file. The hand instrument group showed a significant difference in working length competency for each larger file size (p<0.05) (ANOVA). Regression analysis in the hand instrumentation group indicated a working length decrease with a mean of 0,2 mm after each consecutive file size (p<0.01). CONCLUSIONS: The outcome of our trial indicated a high consistency in working length control for root canal preparation under simulated clinical condition using Mtwo rotary files. Mtwo NiTi rotary file did therefore proved to be more accurate in comparison to the conventional hand instrumentation. KEY WORDS: Working length, Mtwo, nickel-titanium, hand preparation, engine driven preparation.

[Retrospective study of direct composite restorations according to the USPHS criteria]. / Direkt kompozit restaurációk retrospektív vizsgálata USPHS kritériumrendszer alapján.

The purpose of this retrospective study was to evaluate and describe the occurrence of different deficiencies of composite restorations in molar and premolar teeth. Further aim was to investigate possible correlations between occurring malformations and the localization or size of the restorations. 240 class II composite restorations (in 85 patients) were involved in the study. Control examinations were carried out five years after restorations, according to the United States Public Health Services' criteria. Namely, anatomic form, marginal integrity, marginal stain, color stability, surface smoothness, and the presence of secondary caries or fractures of the restorations. The associations between variables were calculated by bivariate analyses using either Pearson chi-square or Fisher tests. P < 0.05 was considered significant. In 0.8% of the fillings, secondary caries and in 0.4% of the cases, fracture was found as a failure. The frequency of adjacent deficiencies were found as follows: color instability, 12.5%; marginal stain, 20.8%; anatomic deformity, 15.0%; failure of marginal integrity, 8.8%; and surface roughness, 2.5%. Color instability was significantly more frequent in premolar teeth, than in molars (P = 0.031). Color instability (P = 0.015), marginal stain (P < 0.001) and anatomic form malformation (P = 0.002) occurred more frequently in MOD restorations than in MO/OD fillings. Our results suggest that class II restorations are correct both functionally and esthetically in 98.8% of the cases, even after a 5-year-period.

Chemomechanical preparation by hand instrumentation and by Mtwo engine-driven rotary files, an ex vivo study.

OBJECTIVE: To compare the disinfecting efficacy of the sodium hypochlorite irrigation by root canal preparation with stainless steel hand files, taper 0.02 and nickel-titanium Mtwo files with taper 0.04-0.06. STUDY DESIGN: 40 extracted human teeth were sterilized, and then inoculated with Enterococcus faecalis (ATCC 29212). After 6 day incubation time the root canals were prepared by hand with K-files (n=20) and by engine-driven Mtwo files (VDW, Munich, Germany) (n=20). Irrigation was carried out with 2.5% NaOCl in both cases. Samples were taken and determined in colony forming units (CFU) from the root canals before and after the preparation with instruments #25 and #35. RESULTS: Significant reduction in bacterial count was determined after filing at both groups. The number of bacteria kept on decreasing with the extension of apical preparation diameter. There was no significant difference between the preparation sizes in the bacterial counts after hand or engine-driven instrumentation at the same apical size. Statistical analysis was carried out with Mann-Whitney test, paired t-test and independent sample t-test. CONCLUSIONS: Significant reduction in CFU was achieved after the root canal preparation completed with 2.5% NaOCl irrigation, both with stainless steel hand or nickel-titanium rotary files. The root canal remained slightly infected after chemo mechanical preparation in both groups. Key words:Chemomechanical preparation, root canal disinfection, nickel-titanium, conicity, greater taper, apical size.

[Direct comparison of working length determination by ProPex electronic apex locator and radiographic method--an in vitro study]. / ProPex apexlokátorral és radiológiai módszerrel végzett munkahossz-meghatározás összehasonlítása in vitro direkt eljárással.

The purpose of this study was to compare the accuracy of electronic and radiographical working length determination in extracted molars. Palatal root canals of twenty maxillary molars were used. Teeth were randomly divided into two groups. In case of the first group the working length was measured by apex locator (Propex, Dentsply-Maillefer), and in the second group it was measured with radiographic method (RVG, Trophy). The files, used for measuring the working length, were cemented into the teeth. The apical half of the root was ground, and the tips of the files visualised as well as the apical region of the root canal with the apical constriction. Standard digital photographs were taken from the prepared samples, and the distance between the tip of the file and the physiological foramen was measured. Two sample t test was used for statistical analysis. The physiological foramen was approached on average to 0.11 mm by the electronic method and to 0.58 mm by the radiological method. There was no significant difference in the measuring accuracy between the two methods (p=0.10). The electronic and radiological working length determination may prove to show an equivalent consistency.

Comparison of radiographic and electronical working length determination on palatal and mesio-buccal root canals of extracted upper molars.

OBJECTIVE: The objective of this work was to compare the accuracy of working length determination of ProPex (Dentsply-Maillefer) electronic apex locator (EAL) to the radiographic method on extracted molars. STUDY DESIGN: Root canal length was determined using the electronic apex locator and conventional radiographic method. Digital radiographs were taken and the distance between the tip of the file and the radiographic apex was measured. Mann-Whitney and chi(2) tests were used for the statistical analysis. RESULTS: The average distance between the tip of the file and the radiographic apex in the mesio-buccal canals was 0.46 mm with the radiologic method, and 0.23 mm with EAL, and 0.43 mm and 0.31 mm in the palatal canals, respectively. The difference at the mesio-buccal canals was significant (Mann-Whitney test P = .048). The number of overextensions was higher in the EAL group. CONCLUSION: The desired position of the file achieved by the EAL proved to be more accurate in the mesio-buccal canals than with the radiologic method alone.

[Clinical evaluation of a quality criteria system for ceramic inlay restorations]. / Minoségi követelményrendszer alkalmazása kerámia-restaurációk ellenorzo vizsgálatában.

The aim of this study was to test the utilization of a generally accepted quality criteria system (United States Public Health Services Modified Quality Criteria) in the authors' practice, in the cases of all ceramic inlays and onlays. 41 ceramic inlays/onlays--29 pressed and 12 laminated--were made for 28 patients. Restorations were controlled after 2 years of cementation. According to the USPHS criteria system the following characteristics were controlled: anatomic contour, marginal integrity, marginal discoloration, color match, secondary caries, and surface roughness. In addition, postoperative sensitivity, patient's satisfaction and tooth vitality were examined as well. The USPHS quality criteria system together with the complementary data proved to be an objective examination method that was easily applicable to our clinical practice. On the basis of these studies the utilization of this system together with patient's satisfaction, tooth vitality and sensitivity records appeared a good basis for a regular quality control system of ceramic inlays and onlays.

[In vitro comparison of root canal preparation with step-back technique and GT rotary file--a nickel-titanium engine driven rotary instrument system]. / Step-back technikával és GT rotary file nikkel-titanium gépi forgómuszerrel végzett gyökércsatorna-elokészítés in vitro vizsgálata.

The aim of the authors' study was to compare the remaining root canal wall thickness and the preparation time of root canals, prepared either with step-back technique, or with GT Rotary File, an engine driven nickel-titanium rotary instrument system. Twenty extracted molars were decoronated. Teeth were divided in two groups. In Group 1 root canals were prepared with step-back technique. In Group 2 GT Rotary File System was utilized. Preoperative vestibulo-oral X-ray pictures were taken from all teeth with radiovisiograph (RVG). The final preparations at the mesiobuccal canals (MB) were performed with size #30 and palatinal/distal canals with size #40 instruments. Postoperative RVG pictures were taken ensuring the preoperative positioning. The working time was measured in seconds during each preparation. The authors also assessed the remaining root canal wall thickness at 3, 6 and 9 mm from the radiological apex, comparing the width of the canal walls of the vestibulo-oral projections on pre- and postoperative RVG pictures both mesially and buccally. The ratios of the residual and preoperative root canal wall thickness were calculated and compared. The largest difference was found at the MB canals of the coronal and middle third level of the root, measured on the distal canal wall. The ratio of the remaining dentin wall thickness at the coronal and the middle level in the case of step-back preparation was 0.605 and 0.754, and 0.824 and 0.895 in the cases of GT files respectively. The preparation time needed for GT Rotary File System was altogether 68.7% (MB) and 52.5% (D/P canals) of corresponding step-back preparation times. The use of GT Rotary File with comparison of standard step-back method resulted in a shortened preparation time and excessive damage of the coronal part of the root canal could be avoided.

[The effectiveness of a benzydamine rinse in the management of stomato-glossopyrosis]. / Benzidamin-tartalmú szájöblítovel szerzett klinikai tapasztalatok a stomato- glossopyrosis kezelésében.

Burning mouth syndrome (BMS) is a frequent source of oral discomfort and sensation mainly at middle-aged or elderly women without identifiable local pathology. Forty-two patients with stomato-glossopyrosis were registered at the Department of Periodontology, Dental School, University Pécs, between August 2002 and January 2003. Oral clinical examination failed to identify physical explanation for the complaint of painful or burning mouth. After complete examination (blood test included), scaling, root planning and oral hygiene instructions were given. Six patients reported decreasing mucosal pain or burning. Two patients had iron-deficiency and Sjögren's syndrome was suspected and verified at another two patients. Thirty-two patients completed the benzydamine-study. They were divided into two groups. The study group (16 patients) rinsed with benzydamine containing solution, the control group (16 patients) received placebo solution for rinsing the same way for ten days. Differences were found between the two groups favouring the benzydamine containing solution. Pain reduction, mouth dryness and diminution of burning sensation were detected by means of visual analog scale (VAS). Benzydamine rinses could be effective in the management of BMS, but clinicians need to monitor patients for possible side effects in the case of long-term use.

[Comparative analysis of the sensitivity and accuracy of clinical, radiographic and histometric measurements in assessing periodontal attachment levels]. / Klinikai radiológiai és histometriás paraméterek érzékenységének összehasonlító vizsgálata parodontális tapadásnyereség értékelésekor.

The ultimate goal of the regenerative periodontal treatment is the de novo formation of new connective tissue attachment with the regrowth of the alveolar bone. Clinically it is very difficult to evaluate the nature of clinical healing and prove the occurrence of real periodontal regeneration. The aim of the present study was to compare the clinical and radiographic parameters with the histometric findings following two different periodontal regenerative procedures in humans.

Comparison of clinical, radiographic, and histometric measurements following treatment with guided tissue regeneration or enamel matrix proteins in human periodontal defects.

BACKGROUND: The purpose of this study was to compare the clinical and radiographic parameters with the histometric findings following 2 different regenerative procedures in humans. METHODS: Fourteen advanced intrabony defects at teeth scheduled for extraction were randomly treated as follows: 8 with guided tissue regeneration (GTR) using bioabsorbable barriers and 6 with an enamel matrix protein derivative (EMD). Standardized radiographs, probing depths (PD), and attachment levels (CAL) at baseline and 6 months after therapy were evaluated and compared to the histometric measurements made following the removal of teeth and surrounding tissues 6 months after the surgery. RESULTS: Significant PD reductions (GTR: -5.62 mm; EMD: -5.00 mm) and CAL gains (GTR: 3.87 mm; EMD: 2.67 mm) were observed in both groups. Six months after surgery, minor resorptions of the alveolar crest (AC) (GTR: 0.40 mm; EMD: 0.33 mm) and bony gain at the bottom of the defects (GTR: 0.47 mm; EMD: 1.05 mm) were observed radiographically. No statistically significant differences in the change of clinical and radiographic parameters between the GTR and EMD groups were found. Histometrically, significant amounts of new connective tissue attachment (i.e., cementum with inserting collagen fibers) were observed in both groups (GTR: 2.29 mm; EMD: 1.81 mm). Bone regeneration was found to be significant only in the GTR group (GTR: 1.93 mm; EMD: 0.78 mm). However, the study lacked statistical power for determining equivalence between the groups. CONCLUSIONS: Within the limitations of the present study, it may be concluded that at 6 months after GTR or enamel matrix protein derivative therapy, formation of new cementum and bone may be histometrically demonstrated. Except for the formation of new bone, no statistically significant differences between both therapies could be seen for clinical, radiographic, and histometric results 6 months after surgery.