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A study is presented on the morphology and respiratory functions of mitochondria from Torpedo marmorata red blood cells. In vivo staining of red blood cells and transmission electron microscopy showed the existence of a considerab...
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A study is presented on the morphology and respiratory functions of mitochondria from Torpedo marmorata red blood cells. In vivo staining of red blood cells and transmission electron microscopy showed the existence of a considerable number of vital and orthodox mitochondria which decreased from young erythroblasts to mature erythrocytes from t~)—50 to 30—20 per cell. In erythrocytes mitochondria exhibited a canonical, functional respiratory chain. The content and activity of cytoebromes in erytbrocytes were, however, significantly lower as compared to mammalian tissues.
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Results of studies on the role of the 18 kDa (IP) polypeptide subunit of complex I, encoded by the nuclear NDUFS4 gene, in isolated bovine heart mitochondria and human and murine cell cultures are presented. The mammalian 18 kDa s...
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Results of studies on the role of the 18 kDa (IP) polypeptide subunit of complex I, encoded by the nuclear NDUFS4 gene, in isolated bovine heart mitochondria and human and murine cell cultures are presented. The mammalian 18 kDa subunit has in the carboxy-terminal sequence a conserved consensus site (RVS), which in isolated mitochondria is phosphorylated by cAMP-dependent protein kinase (PKA). The catalytic and regulatory subunits of PKA have been directly immunodetected in the inner membrane/matrix fraction of mammalian mitochondria. In the mitochondrial inner membrane a PP2Cγ-type phosphatase has also been immunodetected, which dephosphorylates the 18 kDa subunit, phosphorylated by PKA. This phosphatase is Mg~(2+)-dependent and inhibited by Ca~(2+). In human and murine fibroblast and myoblast cultures "in vivo", elevation of intracellular cAMP level promotes phosphorylation of the 18 kDa subunit and stimulates the activity of complex I and NAD-linked mitochondrial respiration. Four families have been found with different mutations in the cDNA of the NDUFS4 gene. These mutations, transmitted by autosomal recessive inheritance, were associated in homozygous children with fatal neurological syndrome. All these mutations destroyed the phosphorylation consensus site in the C terminus of the 18 kDa subunit, abolished cAMP activation of complex I and impaired its normal assembly.
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摘要 :
Results of studies on the role of the 18 kDa (IP) polypeptide subunit of complex I, encoded by the nuclear NDUFS4 gene, in isolated bovine heart mitochondria and human and murine cell cultures are presented.The mammalian 18 kDa su...
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Results of studies on the role of the 18 kDa (IP) polypeptide subunit of complex I, encoded by the nuclear NDUFS4 gene, in isolated bovine heart mitochondria and human and murine cell cultures are presented.The mammalian 18 kDa subunit has in the carboxy-terminal sequence a conserved consensus site (RVS), which in isolated mitochondria is phosphorylated by cAMP-dependent protein kinase (PKA). The catalytic and regulatory subunits of PKA have been directly immunodetected in the inner membrane/matrix fraction of mammalian mitochondria. In the mitochondrial inner membrane a PP2Cgamma-type phosphatase has also been immunodetected, which dephosphorylates the 18 kDa subunit, phosphorylated by PKA. This phosphatase is Mg(2+)-dependent and inhibited by Ca(2+). In human and murine fibroblast and myoblast cultures "in vivo", elevation of intracellular cAMP level promotes phosphorylation of the 18 kDa subunit and stimulates the activity of complex I and NAD-linked mitochondrial respiration.Four families have been found with different mutations in the cDNA of the NDUFS4 gene. These mutations, transmitted by autosomal recessive inheritance, were associated in homozygous children with fatal neurological syndrome. All these mutations destroyed the phosphorylation consensus site in the C terminus of the 18 kDa subunit, abolished cAMP activation of complex I and impaired its normal assembly.
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F3/Contactin is a neuronal surface glycoprotein, which plays a general role in neural development and, in particular, in neuronal and oligodendrocyte differentiation. In a previous study using the F3/EGFP transgenic mice, which ex...
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F3/Contactin is a neuronal surface glycoprotein, which plays a general role in neural development and, in particular, in neuronal and oligodendrocyte differentiation. In a previous study using the F3/EGFP transgenic mice, which express an EGFP reporter under control of the regulatory region from the mouse F3/Contactin gene, the activation of the F3/Contactin promoter was found to correlate with granule and Purkinje neuron differentiation in developing cerebellar cortex. Here we report that in developing cerebral cortex and basal ganglia the F3/Contactin gene is mostly activated during early commitment of neuronal precursors, thus indicating a region-specific profile of its developmental activation. We also report that, in the same structures of F3/EGFP mice, a downregulation of the endogenous F3/Contactin gene occurs, which correlates with upregulation of the dopaminergic phenotype and with locomotor pattern abnormalities. Therefore, F3/EGFP transgenic mice exhibit morphological and functional phenotypes recapitulating those arising from imbalance of the striatal dopaminergic pathway. As for the underlying mechanisms, we postulate that in F3/EGFP mice F3/Contactin downregulation results from the ability of transgene promoter sequences to interfere with the activation of the endogenous gene, thus realizing an F3/Contactin knockdown model, while dopaminergic upregulation is consistent with a general F3/Contactin inhibitory effect on the neuronal phenotype.
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Melphalan has been a mainstay of multiple myeloma (MM) therapy for many years. However, following treatment with this alkylator, malignant plasma cells usually escape both apoptosis and cell cycle control, and acquire drug-resista...
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Melphalan has been a mainstay of multiple myeloma (MM) therapy for many years. However, following treatment with this alkylator, malignant plasma cells usually escape both apoptosis and cell cycle control, and acquire drug-resistance resulting in tumor progression. Bendamustine is being used in MM patients refractory to conventional DNA-damaging agents, although the mechanisms driving this lack of cross-resistance are still undefined. Here, we investigated the molecular pathway of bendamustine-induced cell death in melphalan-sensitive and melphalan-resistant MM cell lines. Bendamustine affected cell survival resulting in secondary necrosis, and prompted cell death primarily through caspase-2 activation. Also, bendamustine blocked the cell cycle in the G2/M phase and induced micronucleation, erratic chromosome spreading and mitotic spindle perturbations in melphalan-resistant MM cells. In these cells, both Aurora kinase A (AURKA) and Polo-like kinase-1 (PLK-1), key components of the spindle-assembly checkpoint, were down-regulated following incubation with bendamustine, whereas levels of Cyclin B1 increased as a consequence of the prolonged mitotic arrest induced by the drug. These findings indicate that, at least in vitro, bendamustine drives cell death by promoting mitotic catastrophe in melphalan-resistant MM cells. Hence, activation of this alternative pathway of cell death may be a novel approach to the treatment of apoptosis-resistant myelomas.
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PURPOSE: To evaluate the efficacy of bipolar diathermy in ensuring closure of leaking sclerotomies after complete 23-gauge transconjunctival sutureless vitrectomy. METHODS: In this prospective, interventional case series, in 136 e...
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PURPOSE: To evaluate the efficacy of bipolar diathermy in ensuring closure of leaking sclerotomies after complete 23-gauge transconjunctival sutureless vitrectomy. METHODS: In this prospective, interventional case series, in 136 eyes of 136 patients with at least one leaking sclerotomy at the end of a complete 23-gauge transconjunctival sutureless vitrectomy, external bipolar wet-field diathermy was applied to leaking sclerotomies, including the conjunctiva and sclera. Intraoperative wound closure, and postoperatively, at 6 hours, 1 day and 3 days, sclerotomies leakage, intraocular pressure, hypotony, and hypotony-related complications were evaluated. RESULTS: Intraoperative closure was achieved in 231 of 238 leaking sclerotomies (97%) that received diathermy. One of these with postoperative leakage needed suture. Compared with baseline (14.4 ± 2.8 mmHg), mean intraocular pressure was lower at 6 hours (13.2 ± 3.8 mmHg, Tukey-Kramer P < 0.001) and not different at 24 hours or 72 hours. Hypotony (intraocular pressure <5 mmHg) was observed in 6 eyes (4.5%) at 6 hours, in 2 (1.5%) at 24 hours, and in none at 3 days. Logistic regression analysis showed that, 6 hours postoperatively, hypotony was related to younger age (≤50 years) at surgery (P = 0.031). No hypotony-related complications were recorded. CONCLUSION: Bipolar wet-field diathermy of sutureless sclerotomies is an effective method for ensuring a leaking sclerotomies closure.
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Resveratrol, a polyphenol abundantly found in grapes and red wine, exhibits beneficial health effects due to its anti-inflammatory properties. In the present study, we evaluated the effect of resveratrol on inflammatory responses ...
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Resveratrol, a polyphenol abundantly found in grapes and red wine, exhibits beneficial health effects due to its anti-inflammatory properties. In the present study, we evaluated the effect of resveratrol on inflammatory responses induced by lipopolysaccharide (LPS) treatment of human intestinal Caco-2 and SW480 cell lines. In the LPS-treated intestinal cells, resveratrol dose-dependently inhibited the expression of inducible NO synthase (iNOS) mRNA as well as protein expression, resulting in a decreased production of NO. In addition, Toll-like receptor-4 expression was significantly diminished in LPS-stimulated cells after resveratrol pre-treatment. To investigate the mechanisms by which resveratrol reduces NO production and iNOS expression, we examined the activation of inhibitor of kappaB (I kappaB) in LPS-stimulated intestinal cells. Results demonstrated that resveratrol inhibited the phosphorylation, as well as the degradation, of the I kappaB complex. Overall, these results show that resveratrol is able to reduce LPS-induced inflammatory responses by intestinal cells, interfering with the activation of NF- kappaB-dependent molecular mechanisms. Copyright copyright The Authors 2012.
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Objectives: The purpose of this study was to describe the susceptibility of environmental strains of Legionella spp. to 10 antimicrobials commonly used for legionellosis therapy. A study of environmental strains could be useful to...
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Objectives: The purpose of this study was to describe the susceptibility of environmental strains of Legionella spp. to 10 antimicrobials commonly used for legionellosis therapy. A study of environmental strains could be useful to timely predict the onset of antibiotic resistance in the environment before it is evidenced in clinical specimens. Methods: The minimum inhibitory concentrations (MICs) of 100 environmental Legionella pneumophila (Lpn) strains belonging to serogroups (sgs) 1, 6, 8, and 10 were tested using the E-test methodology on buffered charcoal yeast extract agar supplemented with α-ketoglutarate. The most frequent sgs were selected from those obtained during microbiological surveillance conducted in 2014 in a hospital in Southern Italy. The MICs were read after 2 days of incubation at 35 ℃ in a humidified atmosphere without CO_2. Results: All isolates were inhibited by low concentrations of fluoroquinolones and macrolides. Rifampicin was the most active drug against the isolates in vitro. All Lpn isolates were inhibited by the following drugs (in decreasing order of their MICs): doxycycline > tigecycline > cefotaxime. The MICs of azi-thromycin, ciprofloxacin, levofloxacin, moxifloxacin, and tigecycline were significantly lower for Lpn non-sg 1 than Lpn sg 1 isolates. Conclusions: Susceptibility testing of Legionella strains to appropriate antibiotics should be performed often to evaluate the possible emergence of resistance, to improve the outcomes of patients, and to reduce the direct costs associated with hospitalization.
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Objectives: Diffusion imaging represents a new imaging tool for the diagnosis of breast cancer. This study aims to investigate the role of diffusion-weighted MRI with background body signal suppression (DWIBS) for evaluating breas...
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Objectives: Diffusion imaging represents a new imaging tool for the diagnosis of breast cancer. This study aims to investigate the role of diffusion-weighted MRI with background body signal suppression (DWIBS) for evaluating breast lesions. Methods: 90 patients were prospectively evaluated by MRI with STIR, TSE-T_2, contrast enhanced THRIVET1 and DWIBS sequences. DWIBS were analyzed searching for the presence of breast lesions and calculating the ADC value. ADC values of ≤1.44 × 10~(-3) mm~2/s were considered suspicious for malignancy. This analysis was then compared with the histological findings. Sensitivity, specificity, diagnostic accuracy (DA), positive predictive value (PPV) and negative (NPV) were calculated. Results: In 53/90 (59%) patients, DWIBS indicated the presence of breast lesions, 16 (30%) with ADC values of > 1.44 and 37 (70%) with ADC ≤ 1.44. The comparison with histology showed 25 malignant and 28 benign lesions. DWIBS sequences obtained sensitivity, specificity, DA, PPV and NPV values of 100, 82, 87, 68 and 100%, respectively. Conclusion: DWIBS can be proposed in theMRI breast protocol representing an accurate diagnostic complement.
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Sequence analysis of mitochondrial and nuclear candidate genes of complex I in children with deficiency of this complex and exhibiting Leigh-like syndrome has revealed, in one of them, a novel mutation in the NDUFS4 gene encoding ...
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Sequence analysis of mitochondrial and nuclear candidate genes of complex I in children with deficiency of this complex and exhibiting Leigh-like syndrome has revealed, in one of them, a novel mutation in the NDUFS4 gene encoding the 18 kDa subunit. Phosphorylation of this subunit by cAMP-dependent protein kinase has previously been found to activate the complex. The present mutation consists of a homozygous G-->A transition at nucleotide position +44 of the coding sequence of the gene, resulting in the change of a tryptophan codon to a stop codon. Such mutation causes premature termination of the protein after only 14 amino acids of the putative mitochondrial targeting peptide. Fibroblast cultures from the patient exhibited severe reduction of the rotenone-sensitive NADH-->UQ oxidoreductase activity of complex I, which was insensitive to cAMP stimulation. Two-dimensional electrophoresis showed the absence of detectable normally assembled complex I in the inner mitochondrial membrane. These findings show that the expression of the NDUFS4 gene is essential for the assembly of a functional complex I.
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