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Background and purpose: Accurate instance segmentation of teeth in panoramic dental X-rays is a challenging task due to variations in tooth morphology and overlapping regions. In this study, we propose a new algorithm, for instance, segmentation of the different teeth in panoramic dental X-rays. Methods: An instance segmentation model was trained using the architecture of a Mask Region-based Convolutional Neural Network (Mask-RCNN). The data for the training, validation, and testing were taken from the Tuft dental database (1000 panoramic dental radiographs). The number of the predicted label was 52 (20 deciduous and 32 permanent). The size of the training, validation, and test sets were 760, 190, and 70 images, respectively, and the split was performed randomly. The model was trained for 300 epochs, using a batch size of 10, a base learning rate of 0.001, and a warm-up multistep learning rate scheduler (gamma = 0.1). Data augmentation was performed by changing the brightness, contrast, crop, and image size. The percentage of correctly detected teeth and Dice in the test set were used as the quality metrics for the model. Results: In the test set, the percentage of correctly classified teeth was 98.4%, while the Dice score was 0.87. For both the left mandibular central and lateral incisor permanent teeth, the Dice index result was 0.91 and the accuracy was 100%. For the permanent teeth right mandibular first molar, mandibular second molar, and third molar, the Dice indexes were 0.92, 0.93, and 0.78, respectively, with an accuracy of 100% for all three different teeth. For deciduous teeth, the Dice indexes for the right mandibular lateral incisor, right mandibular canine, and right mandibular first molar were 0.89, 0.91, and 0.85, respectively, with an accuracy of 100%. Conclusions: A successful instance segmentation model for teeth identification in panoramic dental X-ray was developed and validated. This model may help speed up and automate tasks like teeth counting and identifying specific missing teeth, improving the current clinical practice.

Mammographic breast density (BD) is commonly visually assessed using the Breast Imaging Reporting and Data System (BI-RADS) four-category scale. To overcome inter- and intraobserver variability of visual assessment, the authors retrospectively developed and externally validated a software for BD classification based on convolutional neural networks from mammograms obtained between 2017 and 2020. The tool was trained using the majority BD category determined by seven board-certified radiologists who independently visually assessed 760 mediolateral oblique (MLO) images in 380 women (mean age, 57 years ± 6 [SD]) from center 1; this process mimicked training from a consensus of several human readers. External validation of the model was performed by the three radiologists whose BD assessment was closest to the majority (consensus) of the initial seven on a dataset of 384 MLO images in 197 women (mean age, 56 years ± 13) obtained from center 2. The model achieved an accuracy of 89.3% in distinguishing BI-RADS a or b (nondense breasts) versus c or d (dense breasts) categories, with an agreement of 90.4% (178 of 197 mammograms) and a reliability of 0.807 (Cohen κ) compared with the mode of the three readers. This study demonstrates accuracy and reliability of a fully automated software for BD classification. Keywords: Mammography, Breast, Convolutional Neural Network (CNN), Deep Learning Algorithms, Machine Learning Algorithms Supplemental material is available for this article. © RSNA, 2022

Cytochrome c oxidase (CcO), the CuA, heme a, heme a3, CuB enzyme of respiratory chain, converts the free energy released by aerobic cytochrome c oxidation into a membrane electrochemical proton gradient (ΔμH+). ΔμH+ derives from the membrane anisotropic arrangement of dioxygen reduction to two water molecules and transmembrane proton pumping from a negative (N) space to a positive (P) space separated by the membrane. Spectroscopic, potentiometric, and X-ray crystallographic analyses characterize allosteric cooperativity of dioxygen binding and reduction with protonmotive conformational states of CcO. These studies show that allosteric cooperativity stabilizes the favorable conformational state for conversion of redox energy into a transmembrane ΔμH+. (i) Dioxygen binding to Fea3 at BNC selects a CcO state in which the conformation of the subunit I-helix X, next in sequence to the hemes a and a3 histidine axial ligands, opens access of pumped protons from the N space to a Mg2+-storage site at the opposite (P) membrane side. (ii) Sequential electron transfer at CuA and heme a is associated with additional CcO conformational change in which protons are translocated from the Mg2+-site to the propionate(s) of heme a. From there, protons are expelled in the P space on heme a oxidation by electrostatic repulsion along a hydrogen bond network of amino acid residues and/or structured water molecules.

The “bovine” aortic arch is an anatomic variant consisting in a common origin of the innominate and left carotid artery (CILCA), associated with a greater risk of thoracic aortic diseases (aneurysms and dissections), stroke, and complications after endovascular procedures. CILCA can be detected by visual assessment of computed tomography (CT) chest scans, but it is rarely reported. We developed a deep learning (DL) segmentation-plus-classification system to automatically detect CILCA based on 302 CT studies acquired at 2 centers. One model (3D U-Net) was trained from scratch (supervised by manual segmentation), validated, and tested for the automatic segmentation of the aortic arch and supra-aortic vessels. Three DL architectures (ResNet50, DenseNet-201, and SqueezeNet), pre-trained over millions of common images, were trained, validated, and tested for the automatic classification of CILCA versus non-CILCA, supervised by radiologist’s classification. The 3D U-Net-plus-DenseNet-201 was found to be the best system (Dice index 0.912); its classification performance obtained from internal, independent testing on 126 patients gave a receiver operating characteristic area under the curve of 87.0%, sensitivity 66.7%, specificity 90.5%, positive predictive value 87.5%, negative predictive value 73.1%, positive likelihood ratio 7.0, and negative likelihood ratio 0.4. In conclusion, a combined DL system applied to chest CT scans was developed and proven to be an effective tool to detect individuals with “bovine” aortic arch with a low rate of false-positive findings.

Insight into mammalian respiratory complexes defines the role of allosteric protein interactions in their proton-motive activity. In cytochrome c oxidase (CxIV) conformational change of subunit I, caused by O2 binding to heme a32+-CuB+ and reduction, and stereochemical transitions coupled to oxidation/reduction of heme a and CuA, combined with electrostatic effects, determine the proton pumping activity. In ubiquinone-cytochrome c oxidoreductase (CxIII) conformational movement of Fe-S protein between cytochromes b and c1 is the key element of the proton-motive activity. In NADH-ubiquinone oxidoreductase (CxI) ubiquinone binding and reduction result in conformational changes of subunits in the quinone reaction structure which initiate proton pumping.

Digital breast tomosynthesis (DBT) studies were introduced as a successful help for the detection of calcification, which can be a primary sign of cancer. Expert radiologists are able to detect suspicious calcifications in DBT, but a high number of calcifications with non-malignant diagnosis at biopsy have been reported (false positives, FP). In this study, a radiomic approach was developed and applied on DBT images with the aim to reduce the number of benign calcifications addressed to biopsy and to give the radiologists a helpful decision support system during their diagnostic activity. This allows personalizing patient management on the basis of personalized risk. For this purpose, 49 patients showing microcalcifications on DBT images were retrospectively included, classified by BI-RADS (Breast Imaging-Reporting and Data System) and analyzed. After segmentation of microcalcifications from DBT images, radiomic features were extracted. Features were then selected with respect to their stability within different segmentations and their repeatability in test–retest studies. Stable radiomic features were used to train, validate and test (nested 10-fold cross-validation) a preliminary machine learning radiomic classifier that, combined with BI-RADS classification, allowed a reduction in FP of a factor of 2 and an improvement in positive predictive value of 50%.

Purpose To assess the presence and pattern of incidental interstitial lung alterations suspicious of COVID-19 on fluorine-18-fluorodeoxyglucose positron emission tomography (PET)/computed tomography (CT) ([18F]FDG PET/CT) in asymptomatic oncological patients during the period of active COVID-19 in a country with high prevalence of the virus. Methods This is a multi-center retrospective observational study involving 59 Italian centers. We retrospectively reviewed the prevalence of interstitial pneumonia detected during the COVID period (between March 16 and 27, 2020) and compared to a pre-COVID period (January–February 2020) and a control time (in 2019). The diagnosis of interstitial pneumonia was done considering lung alterations of CT of PET. Results Overall, [18F]FDG PET/CT was performed on 4008 patients in the COVID period, 19,267 in the pre-COVID period, and 5513 in the control period. The rate of interstitial pneumonia suspicious for COVID-19 was significantly higher during the COVID period (7.1%) compared with that found in the pre-COVID (5.35%) and control periods (5.15%) (p

The aim of this paper was to compare the open 1-T (O-1T) versus the closed 1.5-T (C-1.5T) cardiac magnetic resonance (MR).The MR examinations of two concurrent cohorts (each including 100 subjects) of patients with suspected or known cardiac disease were reviewed. Such examinations were obtained using O-1T or C-1.5T MRI. The bright-blood cine, T1-weighted (T1), T2-weighed short-tau inversion recovery (T2-STIR), late gadolinium enhancement (LGE) sequences were performed. Signal-to-noise ratio of blood (SNRb) or myocardium (SNRm), and contrast-to-noise ratio of myocardium (CNRm) were calculated. Subjective image quality (SIQ) of each sequence was graded as 0 = poor, 1 = intermediate, or 2 = optimal. Each examination was considered as diagnostic when the report answered the clinical question.C-1.5T was better than O-1T on cine for SNRb(median 172 versus 452), SNRm(71 versus 160) and CNRm (107 versus 265) and on T2-STIR for SNRb(10 versus 29), SNRm(74 versus 261) and CNRm(-67 versus -233)(P 0.001). On LGE, SNRm was higher with O-1T than for C-1.5T (312 versus 79, P 0.001) while CNR was lower (158 versus 389; P 0.001). No significant differences were found for SNRb on LGE and both SNRm and CNRm on T1 (P ≥ 0.215). SIQ of O-1T was not significantly different from that of C-1.5T for both R1 and R2 for cine, T1, and LGE (P ≥ 0.157); for T2-STIR, SIQ of O-1T was significantly lower (P = 0.003). R1-R2 concordance was almost perfect (κ = 0.816-0.894), and all examinations were diagnostic.Even though quantitative measurements mostly favored C-1.5T, the SIQ of O-1T was not significantly different for any sequence, with the only exception of T2-STIR.

Background Differentiate malignant from benign enhancing foci on breast magnetic resonance imaging (MRI) through radiomic signature. Methods Forty-five enhancing foci in 45 patients were included in this retrospective study, with needle biopsy or imaging follow-up serving as a reference standard. There were 12 malignant and 33 benign lesions. Eight benign lesions confirmed by over 5-year negative follow-up and 15 malignant histopathologically confirmed lesions were added to the dataset to provide reference cases to the machine learning analysis. All MRI examinations were performed with a 1.5-T scanner. One three-dimensional T1-weighted unenhanced sequence was acquired, followed by four dynamic sequences after intravenous injection of 0.1 mmol/kg of gadobenate dimeglumine. Enhancing foci were segmented by an expert breast radiologist, over 200 radiomic features were extracted, and an evolutionary machine learning method (“training with input selection and testing”) was applied. For each classifier, sensitivity, specificity and accuracy were calculated as point estimates and 95% confidence intervals (CIs). Results A k-nearest neighbour classifier based on 35 selected features was identified as the best performing machine learning approach. Considering both the 45 enhancing foci and the 23 additional cases, this classifier showed a sensitivity of 27/27 (100%, 95% CI 87–100%), a specificity of 37/41 (90%, 95% CI 77–97%), and an accuracy of 64/68 (94%, 95% CI 86–98%). Conclusion This preliminary study showed the feasibility of a radiomic approach for the characterisation of enhancing foci on breast MRI.

Graphical abstract, Recent epidemiological data report that worldwide more than 53 million people have been infected by SARS-CoV-2, resulting in 1.3 million deaths. The disease has been spreading very rapidly and few months after the identification of the first infected, shortage of hospital resources quickly became a problem. In this work we investigate whether artificial intelligence working with chest X-ray (CXR) scans and clinical data can be used as a possible tool for the early identification of patients at risk of severe outcome, like intensive care or death. Indeed, further to induce lower radiation dose than computed tomography (CT), CXR is a simpler and faster radiological technique, being also more widespread. In this respect, we present three approaches that use features extracted from CXR images, either handcrafted or automatically learnt by convolutional neuronal networks, which are then integrated with the clinical data. As a further contribution, this work introduces a repository that collects data from 820 patients enrolled in six Italian hospitals in spring 2020 during the first COVID-19 emergency. The dataset includes CXR images, several clinical attributes and clinical outcomes. Exhaustive evaluation shows promising performance both in 10-fold and leave-one-centre-out cross-validation, suggesting that clinical data and images have the potential to provide useful information for the management of patients and hospital resources.

We report on a family with occipital horn syndrome (OHS) diagnosed in the proband's late fifties. A novel ATP7A pathogenic variant (c.4222A > T, p.(Lys1408*)), representing the first nonsense variant and the second late truncation causing OHS rather than classic Menkes disease, was found to segregate in the family. The predicted maintenance of transmembrane domains is consistent with a residual protein activity, which may explain the mild clinical presentation.

Purpose CyberKnife can perform fiducial-free lung tracking, even when the target is visible from only one of the double imaging view, thanks to the Lung-Optimized-Treatment package. However, the uncertainty along the blind view (out-of-plane direction) must be taken into account in margin definition. The goal was to determine the accuracy of target localization in 1-view treatments (33% of all lung cases at Centro Diagnostico Italiano). Methods The actual tumor position, stored in logfiles from 2-view treatments, was projected using a geometric relation onto both image planes for each of 26 patients’ data. In the same plots exhale and inhale planned tumor positions, extracted from TPS and based on breath-hold TC, were novelty represented preserving metric with respect to imaging center. Actual vs. planned positions were compared through a home-made software in Matlab able to calculate also the extra-margin to estimated ITV which would have been necessary to cover the 95% of target positions for the 95% of patient population in case of 1-view treatment. The program was validated using data from 1-view and 2-view lung treatments with XLT Phantom (CIRS). Results The validation test confirmed the reliability of the technique. Expansions were equal to 5 mm and 7 mm for lesions in superior and inferior lobe respectively. The method proved to overestimate margins due to imaging sampling algorithm, data dependence on 2-view cases and the small amount of patients. However, results showed clearly that biphasic TC acquired in breath-hold way was rarely a good model for breath amplitude and breath center position in relation to spine alignment center. Conclusions This graphic comparison method is a useful tool to check localization accuracy of CyberKnife system for 1-view treatments and identify movement error components. It could be employed in clinical workflow to get patient-related information for customized margin definition.

The respiratory chain of mitochondria and bacteria is made up of a set of membrane-associated enzyme complexes which catalyse sequential, stepwise transfer of reducing equivalents from substrates to oxygen and convert redox energy into a transmembrane protonmotive force (PMF) by proton translocation from a negative (N) to a positive (P) aqueous phase separated by the coupling membrane. There are three basic mechanisms by which a membrane-associated redox enzyme can generate a PMF. These are membrane anisotropic arrangement of the primary redox catalysis with: (i) vectorial electron transfer by redox metal centres from the P to the N side of the membrane; (ii) hydrogen transfer by movement of quinones across the membrane, from a reduction site at the N side to an oxidation site at the P side; (iii) a different type of mechanism based on co-operative allosteric linkage between electron transfer at the metal redox centres and transmembrane electrogenic proton translocation by apoproteins. The results of advanced experimental and theoretical analyses and in particular X-ray crystallography show that these three mechanisms contribute differently to the protonmotive activity of cytochrome c oxidase, ubiquinone-cytochrome c oxidoreductase and NADH-ubiquinone oxidoreductase of the respiratory chain. This review considers the main features, recent experimental advances and still unresolved problems in the molecular/atomic mechanism of coupling between the transfer of reducing equivalents and proton translocation in these three protonmotive redox complexes.

Vimentin, a member of cytoskeleton intermediate filaments proteins, plays a critical role in cell structure and dynamics. The present proteomic study reveals reduced amount of six different lengths, N-terminal truncated proteolytic products of vimentin, in the primary skin fibroblasts from two unrelated PD patients, as compared to control fibroblasts. The decreased amount of N-terminal truncated forms of vimentin in parkin-mutant fibroblasts, could contribute to impairment of cellular function, potentially contributing to the pathogenesis of Parkinson disease.

Purpose When a lung lesion is detected by only one couple of X-ray tube and image detector integrated with CyberKnife®, the fiducial-less tracking is limited to 1-view (34% of lung treatments at Centro Diagnostico Italiano). The aim of the study was mainly to determine the margin needed to take into account the localization uncertainty along the blind view (out-of-plane direction). Methods 36 patients treated in 2-view tracking modality (127 fractions in total) were included in the study. The actual tumor positions were determined retrospectively through logfile analysis and were projected onto 2D image planes. In the same plots the planned target positions based on biphasic breath-hold CT scans were represented preserving the metric with respect to the imaging center. The internal margin necessary to cover in out-of-plane direction the 95% of the target position distribution in the 95% of cases was calculated by home-made software in Matlab®. A validation test was preliminarily performed using XLT Phantom (CIRS) both in 2-view and 1-view scenarios. Results The validation test proved the reliability of the method, in spite of some intrinsic limitations. Margins were estimated equal to 5 and 6 mm for targets in upper and lower lobe respectively. Biphasic breath-hold CT led to underestimate the target movement in the hypothetical out-of-plane direction. The inter-fractional variability of spine-target distance was an important source of uncertainty for 1-view treatments. Conclusion This graphic comparison method preserving metric could be employed in the clinical workflow of 1-view treatments to get patient-related information for customized margin definition.

Parkinson's disease (PD) is the most common neurodegenerative movement disorder caused primarily by selective degeneration of the dopaminergic neurons in substantia nigra. In this work the proteomes extracted from primary fibroblasts of two unrelated, hereditary cases of PD patients, with different parkin mutations, were compared with the proteomes extracted from commercial adult normal human dermal fibroblasts (NHDF) and primary fibroblasts from the healthy mother of one of the two patients. The results show that the fibroblasts from the two different cases of parkin -mutant patients display analogous alterations in the expression level of proteins involved in different cellular functions, like cytoskeleton structure–dynamics, calcium homeostasis, oxidative stress response, protein and RNA processing.

Recently has been acknowledged the healthy use of Bacillus and related bacteria as probiotics. A mixture reported to contain four probiotic strains of Bacillus clausii is marketed as an OTC (Over The Counter) medicinal supplement for human use. Their poliantibiotic resistant property, useful for restoring the gut microbiota during antibiotic treatment, raises the question about the risk of resistance transfer. In order to better assess the risk-benefit ratio it is important to always monitoring the pattern and stability of resistance spectra in these bacteria. In this work, we have extensively redefined the antibiotic susceptibility profile of these four probiotic strains. Resistance phenotype has been determined by screening a large number of antibiotics, including natural products (such as penicillin, vancomycin and erythromycin), and completely synthetic molecules (such as fluoroquinolones). Extensive comparison with a wild type strain belonging to the normal intestinal microbiome was carried out. The molecular basis of some resistances was determined. Observed antibiotic resistances were correlated with previous and new data in safety evaluations of these strains for human use.

A study is presented on the expression of mitochondrial oxidative phosphorylation complexes in exponentially growing and serum-starved, quiescent human fibroblast cultures. The functional levels of respiratory complexes I and III and complex V (adenosine triphosphate (ATP) synthase) were found to be severely depressed in serum-starved fibroblasts. The depression of oxidative phosphorylation system (OXPHOS) complexes was associated with reduced levels of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) and the down-stream nuclear respiratory factor 1 (NRF1) and mitochondrial transcription factors (TFAM). In serum-starved fibroblasts decrease of the catalytic activity of AMP cyclic dependent protein kinase (PKA) and phosphorylation of cAMP response element-binding protein (CREB), the transcription coactivator of the PGC-1α gene, was found. Hydroxytyrosol prevented the decline in the expression of the PGC-1α transcription cascade of OXPHOS complexes in serum-starved fibroblast cultures. The positive effect of HT was associated with activation of PKA and CREB phosphorylation. These results show involvement of PKA, CREB and PGC-1α in the regulation of OXPHOS in cell transition from the replicating to the quiescent state.

Almost half of autosomal recessive early-onset parkinsonism has been associated with mutations in PARK2, coding for parkin, which plays an important role in mitochondria function and calcium homeostasis. Cyclic adenosine monophosphate (cAMP) is a major second messenger regulating mitochondrial metabolism, and it is strictly interlocked with calcium homeostasis. Parkin-mutant (Pt) fibroblasts, exhibiting defective mitochondrial respiratory/OxPhos activity, showed a significant higher value of basal intracellular level of cAMP, as compared with normal fibroblasts (CTRL). Specific pharmacological inhibition/activation of members of the adenylyl cyclase- and of the phosphodiesterase-families, respectively, as well as quantitative reverse transcription polymerase chain reaction (RT-qPCR) analysis, indicate that the higher level of cAMP observed in Pt fibroblasts can contribute to a higher level of activity/expression by soluble adenylyl cyclase (sAC) and to low activity/expression of the phosphodiesterase isoform 4 (PDE4). As Ca2+ regulates sAC, we performed quantitative calcium-fluorimetric analysis, showing a higher level of Ca2+ in the both cytosol and mitochondria of Pt fibroblasts as compared with CTRL. Most notably, inhibition of the mitochondrial Ca2+ uniporter decreased, specifically the cAMP level in PD fibroblasts. All together, these findings support the occurrence of an altered mitochondrial Ca2+-mediated cAMP homeostasis in fibroblasts with the parkin mutation.

A study is presented on the regulation of alternative splicing (AS) of the Ndufb11 gene of complex I of the mitochondrial respiratory chain and the impact on this process of rotenone treatment in neuroblastoma cells. In physiological conditions the Ndufb11 gene produces at high level a short transcript isoform encoding for a 153 aa protein. This subunit is essential for the assembly of a functional and stable mammalian complex I. The gene produces also, at low level, a longer transcript isoform encoding for a 163 aa protein whose role is unknown. Evidence is presented here showing that the level of the two isoforms is regulated by three DGGGD ESS elements located in exon 2 which can bind the hnRNPH1 protein. In neuronal cells rotenone treatment affects the Ndufb11 alternative splicing pathway, with the increase of the 163/153 mRNAs ratio. This effect appears to be due to the down-regulation of the hnRNPH1 protein. Since rotenone induces apoptosis in neuronal cells, the post-transcriptional regulation of the Ndufb11 gene can be involved in the programmed cell death process.

The respiratory chain of mitochondria and bacteria is made up of a set of membrane-associated enzyme complexes which catalyse sequential, stepwise transfer of reducing equivalents from substrates to oxygen and convert redox energy into a transmembrane protonmotive force (PMF) by proton translocation from a negative (N) to a positive (P) aqueous phase separated by the coupling membrane. There are three basic mechanisms by which a membrane-associated redox enzyme can generate a PMF. These are membrane anisotropic arrangement of the primary redox catalysis with: (i) vectorial electron transfer by redox metal centres from the P to the N side of the membrane; (ii) hydrogen transfer by movement of quinones across the membrane, from a reduction site at the N side to an oxidation site at the P side; (iii) a different type of mechanism based on co-operative allosteric linkage between electron transfer at the metal redox centres and transmembrane electrogenic proton translocation by apoproteins. The results of advanced experimental and theoretical analyses and in particular X-ray crystallography show that these three mechanisms contribute differently to the protonmotive activity of cytochrome c oxidase, ubiquinone-cytochrome c oxidoreductase and NADH-ubiquinone oxidoreductase of the respiratory chain. This review considers the main features, recent experimental advances and still unresolved problems in the molecular/atomic mechanism of coupling between the transfer of reducing equivalents and proton translocation in these three protonmotive redox complexes.

ATP synthase, canonically mitochondrially located, is reported to be ectopically expressed on the plasma membrane outer face of several cell types. We analysed, for the first time, the expression and catalytic activities of the ecto- and mitochondrial ATP synthase during liver regeneration. Liver regeneration was induced in rats by two-thirds partial hepatectomy. The protein level and the ATP synthase and/or hydrolase activities of the hepatocyte ecto- and mitochondrial ATP synthase were analysed on freshly isolated hepatocytes and mitochondria from control, sham-operated and partial hepatectomized rats. During the priming phase of liver regeneration, 3 h after partial hepatectomy, liver mitochondria showed a marked lowering of the ATP synthase protein level that was reflected in the impairment of both ATP synthesis and hydrolysis. The ecto-ATP synthase level, in 3 h partial hepatectomized hepatocytes, was decreased similarly to the level of the mitochondrial ATP synthase, associated with a lowering of the ecto-ATP hydrolase activity coupled to proton influx. Noteworthily, the ecto-ATP synthase activity coupled to proton efflux was completely inhibited in 3 h partial hepatectomized hepatocytes, even in the presence of a marked intracellular acidification that would sustain it as in control and sham-operated hepatocytes. At the end of the liver regeneration, 7 days after partial hepatectomy, the level and the catalytic activities of the ecto- and mitochondrial ATP synthase reached the control and sham-operated values. The specific modulation of hepatocyte ecto-ATP synthase catalytic activities during liver regeneration priming phase may modulate the extracellular ADP/ATP levels and/or proton influx/efflux trafficking, making hepatocyte ecto-ATP synthase a candidate for a novel player in the liver regeneration process.

GRIM-19 is an oncosuppressor which antagonizes the oncogene Stat 3. GRIM19 is partitioned between the nucleus and mitochondria where it appears as a subunit of respiratory complex I. Our group has studied the impact of the chemiotherapic agent all-trans- retinoic acid (ATRA) on the subcellular distribution of GRIM-19. The results show that ATRA treatment of keratinocytes results in increased level of GRIM-19 and other subunits of complex I, in particular of their carbonylated forms, associated with inhibition of its enzymatic activity. In keratinocytes ATRA-promoted phosphatase activity appears to control the proteostasis and activity of complex I. Furthermore, the oncogene STAT-3, besides being localized in the nucleus, has also been found in mitochondria where it interacts with GRIM 19. Thus, perturbation of the interplay of GRIM 19 with factors, associated with complex I, can be involved in the ATRA suppression of keratinocyte growth.

AIM Extracorporeal shock wave lithotripsy (ESWL) and semirigid ureteroscopy lithotripsy (URSL) have become standards of treatment for ureteral calculi. The aim of this retrospective study was to compare ESWL vs. URSL in terms of safety and efficacy for treatment of large distal ureteral stones ≥ 1cm. PATIENTS AND METHODS This investigation assessed 637 patients with distal ureteral stones (10 to 15mm in size). 313 in the ESWL group were treated on an outpatient basis using the LithoDiamond machine without anaesthesia. URSL was performed in 324 patients with a 6-8 Fr semirigid ureterorenoscope and YAG laser under spinal anaesthesia. A successful outcome was defined as the patient being stone free 1 month after treatment. For all patients the parameters, including stone-free rate, operation time, complications, were inserted retrospectively in this study after review of medical records and operating room logs. RESULTS The stone-free rate after URSL was 77.5% and 45.4% after ESWL treatment (p

In this paper allosteric interactions in protonmotive heme aa 3 terminal oxidases of the respiratory chain are dealt with. The different lines of evidence supporting the key role of H + /e − coupling (redox Bohr effect) at the low spin heme a in the proton pump of the bovine oxidase are summarized. Results are presented showing that the I-R54M mutation in P. denitrificans aa 3 oxidase, which decreases by more than 200 mV the E m of heme a , inhibits proton pumping. Mutational aminoacid replacement in proton channels, at the negative (N) side of membrane-inserted prokaryotic aa 3 oxidases, as well as Zn 2 + binding at this site in the bovine oxidase, uncouples proton pumping. This effect appears to result from alteration of the structural/functional device, closer to the positive, opposite (P) surface, which separates pumped protons from those consumed in the reduction of O 2 to 2 H 2 O. This article is part of a Special Issue entitled: Respiratory Oxidases.

A comparative proteomic approach, using two dimensional gel electrophoresis and mass spectrometry, has been developed to compare and elucidate the differences among the cellular proteomes of four closely related isogenic O/C, SIN, N/R and T, B. clausii strains during both exponential and stationary phases of growth. Image analysis of the electropherograms reveals a high degree of concordance among the four proteomes, some proteins result, however, differently expressed. The proteins spots exhibiting high different expression level were identified, by mass-spectrometry analysis, as alcohol dehydrogenase (ADHA, EC1.2.1.3; ABC0046 isoform) aldehyde dehydrogenase (DHAS, EC 1.2.1.3; ABC0047 isoform) and flagellin-protein of B. clausii KSM-k16. The different expression levels of the two dehydrogenases were confirmed by quantitative RT-PCR and dehydrogenases enzymatic activity. The different patterns of protein expression can be considered as cell proteome signatures of the different strains.

Structural and functional observations are reviewed which provide evidence for a central role of redox Bohr effect linked to the low-spin heme a in the proton pump of bovine heart cytochrome c oxidase. Data on the membrane sidedness of Bohr protons linked to anaerobic oxido-reduction of the individual metal centers in the liposome reconstituted oxidase are analysed. Redox Bohr protons coupled to anaerobic oxido-reduction of heme a (and Cu A ) and Cu B exhibit membrane vectoriality, i.e. protons are taken up from the inner space upon reduction of these centers and released in the outer space upon their oxidation. Redox Bohr protons coupled to anaerobic oxido-reduction of heme a 3 do not, on the contrary, exhibit vectorial nature: protons are exchanged only with the outer space. A model of the proton pump of the oxidase, in which redox Bohr protons linked to the low-spin heme a play a central role, is described. This article is part of a Special Issue entitled: Allosteric cooperativity in respiratory proteins.

In mammals, complex I (NADH-ubiquinone oxidoreductase) of the mitochondrial respiratory chain has 31 supernumerary subunits in addition to the 14 conserved from prokaryotes to humans. Multiplicity of structural protein components, as well as of biogenesis factors, makes complex I a sensible pace-maker of mitochondrial respiration. The work reviewed here shows that the cAMP/PKA pathway regulates the biogenesis, assembly and catalytic activity of complex I and mitochondrial oxygen superoxide production. The structural, functional and regulatory complexity of complex I, renders it particularly vulnerable to genetic and sporadic pathological factors. Complex I dysfunction has, indeed, been found, to be associated with several human diseases. Knowledge of the pathogenetic mechanisms of these diseases can help to develop new therapeutic strategies. (C) 2011 Federation of European Biochemical Societies. Published by Elsevier B. V. All rights reserved.

The results of the present work show that the exposure of pregnant rats to low doses of all-trans-retinoic acid (ATRA) (2.5 mg/kg body weight) results in postnatal dysfunction of complex I of the respiratory chain in the cerebellum of the offspring. ATRA had no effect on the postnatal expression of complex I and did not exert any direct inhibitory effect on the enzymatic activity of the complex. The ATRA embryonic exposure resulted, however, in a marked increase in the level of carbonylated proteins in the mitochondrial fraction of the cerebellum, in particular of complex I subunits. The postnatal increase of the carbonylated proteins correlated directly with the inhibition of the activity of complex I. ATRA had, on the other hand, no effect on oxygen free-radical scavengers. It is proposed that embryonic exposure to ATRA results in impairment of protein surveillance in the cerebellum, which persists after birth and results in accumulation of oxidatively damaged complex I.

Work is presented on the role of cAMP-dependent protein phosphorylation in post-translational processing and biosynthesis of complex I subunits in mammalian cell cultures. PKA-mediated phosphorylation of the NDUFS4 subunit of complex I promotes in cell cultures in vivo import/maturation in mitochondria of the precursor of this protein. The import promotion appears to be associated with the observed cAMP-dependent stimulation of the catalytic activity of complex I. These effects of PKA are counteracted by activation of protein phosphatase(s). PKA and the transcription factor CREB play a critical role in the biosynthesis of complex I subunits. CREB phosphorylation, by PKA and/or CaMKs, activates at nuclear and mitochondrial level a transcriptional regulatory cascade which promotes the concerted expression of nuclear and mitochondrial encoded subunits of complex I and other respiratory chain proteins.

A summary is presented of the cellular function and topology of the protein products of genes whose mutations are associated with familial forms of parkinsonism, with particular emphasis on mitochondrial involvement. Observations are reviewed which show mitochondrial respiratory depression in the fibroblasts of a patient affected by familial parkinsonism associated with homozygous PINK1 mutation. The respiratory depression, which was due to loss of mitochondrial cytochrome c, was associated with decreased capacity of respiratory chain oxidative phosphorylation and enhanced cellular level of ROS. Sequence analysis of the overall mtDNA revealed coexistence with the PINK1 mutation of homoplasmic point mutations in the ND5 and ND6 genes of complex I. The presence of these mutations appears to have an impact on the development of the parkinsonism, which can also occur in the heterozygous PINK1 mutation state.

This paper covers genetic and biochemical aspects of mitochondrial bioenergetics dysfunction in hereditary neurological disorders associated with complex I defects. Three types of hereditary complex I dysfunction are dealt with: (i) homozygous mutations in the nuclear genes NDUFS1 and NDUFS4 of complex I, associated with mitochondrial encephalopathy; (ii) a recessive hereditary epileptic neurological disorder associated with enhanced proteolytic degradation of complex I; (iii) homoplasmic mutations in the ND5 and ND6 mitochondrial genes of the complex, cohexistent with mutation in the nuclear PINK1 gene in familial Parkinsonism. The genetic and biochemical data examined highlight different mechanisms by which mitochondrial bioenergetics is altered in these hereditary defects of complex I. This knowledge, besides clarifying molecular aspects of the pathogenesis of hereditary diseases, can also provide hints for understanding the involvement of complex I in sporadic neurological disorders and aging, as well as for developing therapeutical strategies.

Extracellular ATP formation from ADP and inorganic phosphate, attributed to the activity of a cell surface ATP synthase, has so far only been reported in cultures of some proliferating and tumoral cell lines. We now provide evidence showing the presence of a functionally active ecto-F(o)F(1)-ATP synthase on the plasma membrane of normal tissue cells, i.e. isolated rat hepatocytes. Both confocal microscopy and flow cytometry analysis show the presence of subunits of F(1) (alpha/beta and gamma) and F(o) (F(o)I-PVP(b) and OSCP) moieties of ATP synthase at the surface of rat hepatocytes. This finding is confirmed by immunoblotting analysis of the hepatocyte plasma membrane fraction. The presence of the inhibitor protein IF(1) is also detected on the hepatocyte surface. Activity assays show that the ectopic-ATP synthase can work both in the direction of ATP synthesis and hydrolysis. A proton translocation assay shows that both these mechanisms are accompanied by a transient flux of H(+) and are inhibited by F(1) and F(o)-targeting inhibitors. We hypothesise that ecto-F(o)F(1)-ATP synthase may control the extracellular ADP/ATP ratio, thus contributing to intracellular pH homeostasis.