Functional Diversity of Mitochondrial Peptidyl-tRNA Hydrolase ICT1 in Human Cells.

Mitochondria are energy producing organelles of the eukaryotic cell, involved in the synthesis of key metabolites, calcium homeostasis and apoptosis. Protein biosynthesis in these organelles is a relic of its endosymbiotic origin. While mitochondrial translational factors have homologues among prokaryotes, they possess a number of unique traits. Remarkably as many as four mammalian mitochondrial proteins possess a clear similarity with translation termination factors. The review focuses on the ICT1, which combines several functions. It is a non-canonical termination factor for protein biosynthesis, a rescue factor for stalled mitochondrial ribosomes, a structural protein and a regulator of proliferation, cell cycle, and apoptosis. Such a diversity of roles demonstrates the high functionality of mitochondrial translation associated proteins and their relationship with numerous processes occurring in a living cell.

Protein Biosynthesis in Mitochondria: Past Simple, Present Perfect, Future Indefinite.

Mitochondria are obligate organelles of most eukaryotic cells that perform many different functions important for cellular homeostasis. The main role of mitochondria is supplying cells with energy in a form of ATP, which is synthesized in a chain of oxidative phosphorylation reactions on the organelle inner membrane. It is commonly believed now that mitochondria have the endosymbiotic origin. In the course of evolution, they have lost most of their genetic material as a result of genome reduction and gene transfer to the nucleus. The majority of mitochondrial proteins are synthesized in the cytosol and then imported to the mitochondria. However, almost all known mitochondria still contain genomes that are maintained and expressed. The processes of protein biosynthesis in the mitochondria - mitochondrial translation - substantially differs from the analogous processes in bacteria and the cytosol of eukaryotic cells. Mitochondrial translation is characterized by a high degree of specialization and specific regulatory mechanisms. In this review, we analyze available information on the common principles of mitochondrial translation with emphasis on the molecular mechanisms of translation initiation in the mitochondria of yeast and mammalian cells.

Mitochondrial Translation Initiation Factor 3: Structure, Functions, Interactions, and Implication in Human Health and Disease.

Mitochondria are essential organelles of eukaryotic cell that provide its respiratory function by means of the electron transfer chain. Expression of mitochondrial genes is organized in a bacterial-like manner; however multiple evolutionary differences are observed between the two systems, including translation initiation machinery. This review is dedicated to the mitochondrial translation initiation factor 3 (IF3mt), which plays a key role in the protein synthesis in mitochondria. Involvement of IF3mt in human health and disease is discussed.

Factors beyond Enolase 2 and Mitochondrial Lysyl-tRNA Synthetase Precursor Are Required for tRNA Import into Yeast Mitochondria.

In yeast, the import of tRNALys with CUU anticodon (tRK1) relies on a complex mechanism where interaction with enolase 2 (Eno2p) dictates a deep conformational change of the tRNA. This event is believed to mask the tRNA from the cytosolic translational machinery to re-direct it towards the mitochondria. Once near the mitochondrial outer membrane, the precursor of the mitochondrial lysyl-tRNA synthetase (preMsk1p) takes over enolase to carry the tRNA within the mitochondrial matrix, where it is supposed to participate in translation following correct refolding. Biochemical data presented in this report focus on the role of enolase. They show that despite the inability of Eno2p alone to form a complex with tRK1, mitochondrial import can be recapitulated in vitro using fractions of yeast extracts sharing either recombinant or endogenous yeast Eno2p as one of the main components. Taken together, our data suggest the existence of a protein complex containing Eno2p that is involved in RNA mitochondrial import.

Procedure for Purification of Recombinant preMsk1p from E. coli Determines Its Properties as a Factor of tRNA Import into Yeast Mitochondria.

Mitochondrial genomes of many eukaryotic organisms do not code for the full tRNA set necessary for organellar translation. Missing tRNA species are imported from the cytosol. In particular, one out of two cytosolic lysine tRNAs of the yeast Saccharomyces cerevisiae is partially internalized by mitochondria. The key protein factor of this process is the precursor of mitochondrial lysyl-tRNA synthetase, preMsk1p. In this work, we show that recombinant preMsk1p purified from E. coli in native conditions, when used in an in vitro tRNA import system, demonstrates some properties different from those shown by the renatured protein purified from E. coli in the denatured state. We also discuss the possible mechanistic reasons for this phenomenon.

Morbidity and mortality in the antiphospholipid syndrome during a 10-year period: a multicentre prospective study of 1000 patients.

OBJECTIVES: To assess the prevalence of the main causes of morbi-mortality in the antiphospholipid syndrome (APS) during a 10-year-follow-up period and to compare the frequency of early manifestations with those that appeared later. METHODS: In 1999, we started an observational study of 1000 APS patients from 13 European countries. All had medical histories documented when entered into the study and were followed prospectively during the ensuing 10 years. RESULTS: 53.1% of the patients had primary APS, 36.2% had APS associated with systemic lupus erythematosus and 10.7% APS associated with other diseases. Thrombotic events appeared in 166 (16.6%) patients during the first 5-year period and in 115 (14.4%) during the second 5-year period. The most common events were strokes, transient ischaemic attacks, deep vein thromboses and pulmonary embolism. 127 (15.5%) women became pregnant (188 pregnancies) and 72.9% of pregnancies succeeded in having one or more live births. The most common obstetric complication was early pregnancy loss (16.5% of the pregnancies). Intrauterine growth restriction (26.3% of the total live births) and prematurity (48.2%) were the most frequent fetal morbidities. 93 (9.3%) patients died and the most frequent causes of death were severe thrombosis (36.5%) and infections (26.9%). Nine (0.9%) cases of catastrophic APS occurred and 5 (55.6%) of them died. The survival probability at 10 years was 90.7%. CONCLUSIONS: Patients with APS still develop significant morbidity and mortality despite current treatment. It is imperative to increase the efforts in determining optimal prognostic markers and therapeutic measures to prevent these complications.

[Antibodies against phospholipids in patients with normal pregnancy].

We investigated the serum levels of IgG and IgM anticardiolipin (ACL), anti-beta-2-glycoprotein I (B2GPI), anti-phosphatidyl serine (PS), anti-prothrombin(PT), anti-annexin V (AnV) and anti-ethanolamine (Eth) antibodies using an ELISA method (Orgentec, Germany) in 16 females with normal pregnancy in the I, II and Ill trimester. We observed the following changes: 1. Elevation of the IgG u IgMACL, IgG PS, IgM Pr antibodies in the II and decreasing in the Ill trimester. 2. Decreasing of IgG u IgM B2GPI, IgG u IgM AnV, IgG Pr, IgG u IgM Eth antibodies in the II trimester, maintainante of the levels or more decreasing in the Ill trimester. 3. Increasing of lgM PS in the II and more increasing in the Ill trimester. All of these changes have no significant values (p > 0,05). In 10/16 we found extreme values of different antibodies, but all of them had normal delivery.

The annexin A5 protective shield model revisited: inherited carriage of the M2/ANXA5 haplotype in placenta as a predisposing factor for the development of obstetric antiphospholipid antibodies.

This concise review summarizes the role of reduced ANXA5 expression through carriage of the M2/ANXA5 haplotype as a predisposing factor for various thrombophilia related obstetric complications. A revised ANXA5 'protective shield' model is emphasized, where decreased coverage resulting of M2 carriage at placental villi could lead directly to the observed pathology and on the other hand through exposing of antiphospholipid antigenic determinants, to the development of antiphospholipid antibodies (aPL). The aPL then can further disrupt the ANXA5 protective shield. Available and prospective evidence for this revised model is discussed. Conclusions are made about the diagnostic implications of M2 carriage and possible therapeutic strategies with anticoagulants, proven successful in obstetric antiphospholipid syndrome (APS) treatment.

Morbidity and mortality in the antiphospholipid syndrome during a 5-year period: a multicentre prospective study of 1000 patients.

OBJECTIVES: To identify the main causes of morbidity and mortality in patients with antiphospholipid syndrome (APS) during a 5-year period and to determine clinical and immunological parameters with prognostic significance. METHODS: The clinical and immunological features of a cohort of 1000 patients with APS from 13 European countries who had been followed up from 1999 to 2004 were analysed. RESULTS: 200 (20%) patients developed APS-related manifestations during the 5-year study period. Recurrent thrombotic events appeared in 166 (16.6%) patients and the most common were strokes (2.4% of the total cohort), transient ischaemic attacks (2.3%), deep vein thromboses (2.1%) and pulmonary embolism (2.1%). When the thrombotic events occurred, 90 patients were receiving oral anticoagulants and 49 were using aspirin. 31/420 (7.4%) patients receiving oral anticoagulants presented with haemorrhage. 3/121 (2.5%) women with only obstetric APS manifestations at the start of the study developed a new thrombotic event. A total of 77 women (9.4% of the female patients) had one or more pregnancies and 63 (81.8% of pregnant patients) had one or more live births. The most common fetal complications were early pregnancy loss (17.1% of pregnancies) and premature birth (35% of live births). 53 (5.3% of the total cohort) patients died. The most common causes of death were bacterial infection (21% of deaths), myocardial infarction (19%) and stroke (13%). No clinical or immunological predictor of thrombotic events, pregnancy morbidity or mortality was detected. CONCLUSION: Patients with APS still develop significant morbidity and mortality despite current treatment (oral anticoagulants or antiaggregants, or both).

Raman scattering of Si matrix with randomly distributed nanoparticles of semiconducting silicides.

Samples, representing Si matrix with nanoformations of the semiconducting silicides beta-FeSi2 and Mg2Si are studied by Raman scattering. The unpolarized Raman spectra of the samples are interpreted in the framework of the appearance of interface-phonon polaritons. The theoretical dispersion relations of the interface-phonon polaritons in the system Si/silicide/Si are obtained from the Maxwell equations. The correspondence of the theoretical calculations and the experimental observations appeared to be sufficiently good. An evolution of the features in the Raman spectra on the experimental conditions is observed.

Resonant Raman and micro-Raman scattering from Si matrix with unburied beta-FeSi2 nanolayers.

Samples, representing Si matrix with nanolayers of the semiconducting beta-FeSi2 silicide are studied by Raman scattering. The unpolarized Raman spectra of the samples are measured in two different configurations. It is found that the characteristic beta-FeSi2 Raman modes are seen in the spectra, taken at incident angle of about 45 degrees , while only comparatively intensive broad feature is detected in a back-scattering geometry. The difference in the spectra is interpreted with the appearance of surface polariton modes of the optical phonons in the nanosized layers in near back-scattering geometry. The resonant Raman scattering is investigated at incident light angle of about 45 degrees and the energies of the interband transitions in the investigated energy range are determined. It is known that the resonant Raman scattering appears to be even more precise method for the determination of the interband transitions energies than the modulation spectroscopy. Thus we claim that the energies determined here are firstly determined with such a precision.

Inflammatory activity and anticardiolipin antibodies during tibolone treatment of healthy postmenopausal women.

The aim of the study was to investigate the inflammatory activity and anticardiolipin antibodies (Acl) during tibolone administration. Twenty seven clinically healthy postmenopausal women were included in the study and were divided into two groups: 16 women (mean age 56.4 +/- 4.6 years) who received tibolone at a dose of 2.5 mg/day for 6 months and an untreated control group (n = 11, mean age 54.8 +/- 4.0 years). Acl of IgG and IgM isotype, and C-reactive protein (CRP) were determined at baseline, 1, 3, and 6 months after treatment. Acl did not change during tibolone treatment, while CRP increased significantly at the 1st, 3rd, and 6th month compared with that at the baseline values. These preliminary data indicate that tibolone administration does not induce increase in Acl. This may counterbalance the adverse influence on CRP.

Prevalence of factor V Leiden and prothrombin 20210 A variant in Bulgarian patients with pulmonary thromboembolism and deep venous thrombosis.

Factor V Leiden mutation and prothrombin variant 20210 A are well-known risk factors for venous thrombosis (DVT). Recent papers have reported a lower prevalence of factor V Leiden in patients with pulmonary thromboembolism (PTE) than in patients with deep venous thrombosis. The aim of the present study was to compare the prevalence of factor V Leiden and the prothrombin 20210 G <-- A mutation in patients with DVT and in patients with PTE. We studied 128 consecutive patients (45 with DVT, 40 with PTE, and 43 with DVT and PTE) for factor V Leiden and prothrombin 20210 A. One hundred healthy persons matched by age and sex were used as controls. Factor V Leiden was present in five of the patients with PTE [12.5%; 95% confidence interval (CI), 1.5-23.5%; not significant], 15 of the patients with DVT (33.3%; 95% CI, 9.6-38.7%; P < 0.001), and 12 of the patients with DVT and PTE (27.9%; 95% CI, 4.8-33%; P = 0.001). Results for the prothrombin 20210 A mutation were as follows: four of 40 patients with PTE (10%; 95% CI, 0-13.3%; P = 0.46), nine of 45 (20%) of the patients with DVT (95% CI, 0.5-25.5%; P < 0.05) and eight of 43 with DVT and PTE were heterozygous (18.6%; 95% CI, 0-23.9%; P = 0.02). In conclusion, there is a significantly higher frequency of factor V Leiden among patients with DVT than in patients with PTE. However, there is no significant difference of factor V Leiden or 20210 A prothrombin mutation in patients with DVT than in patients with combined DVT/PTE, therefore patients with DVT, carriers of the mutations, do not appear to be at lower risk for pulmonary embolism.

[Frequency of antiphospholipid antibodies and Leiden mutation of hemostasis Factor V in unexplained recurrent fetal and embryo loss]. / Chestota na antifosfolipidni antitela i Leidenova mutatsiia Faktor V na kr'vos'sirvaneto pri povtariashta se neobiasnima fetalna i embrionalna zaguba.

Object of the study are women with a history of unexplained recurrent embryo, fetal and early neonatal death, severe preeclampsia, fetal growth retardation, abruptio placentae, puerperal thromboses. Quite often placental insufficiency is linked to abnormal vascular system and hemostatic disturbancies. In about 65% of the women with a complicated and in 18% of the women with a normal pregnancy are observed different genetic anomalies that lead to a hypercoagulative state. A major place is taken by the Leiden mutation of hemostasis factor V, by protein C and protein S deficiency, etc. Another disease that leads to arterial and venous thromboses and is most often linked to recurrent miscarriage is the antiphospholipid syndrome. Many authors confirm the findings of large placental infarctions and thromboses in women who are positive for antophospholipid antibodies.

[Alpha-2 macroglobulin in serum and synovial fluid of patients with rheumatoid arthritis and osteoarthrosis]. / Alpha2-makroglobulin v syvorotke i sinovial'noi zhidkosti bol'nykh revmatoidnym artritom i osteoartrozom.

AIM: To study concentrations of alpha-2-macroglobulin (A-2-MG) in blood serum and synovial fluid (SF) in patients with rheumatoid arthritis and formulation of basic clinical, laboratory and x-ray criteria of RA activity. MATERIALS AND METHODS: A-2-MG levels were measured in the serum and SF from 151 RA patients and in the serum of 20 patients with osteoarthrosis (OA) and 62 healthy donors. The serum concentration for RA patients was 166 +/- 65.3 mg%, for OA patients--175.26 +/- 36.99 mg% and for healthy donors--177.772 +/- 50 mg%. Mean concentration of SF A-2-MG in RA patients was 98.77 +/- 82.43 mg%. CONCLUSION: Changes in the concentration of A-2-MG are unrelated to inflammation activity in the joints of RA and OA patients. Serum and synovial concentration of this protein corresponds to changes in the concentration of IgM and rheumatoid factors in RA patients.

Anticardiolipin antibodies in spontaneously hypertensive rats (SHR), stroke-prone SHR and normal Wistar rats.

1. Anticardiolipin antibodies (ACA) can be detected in the serum of patients with autoimmune disturbances, ischaemic heart disease, myocardial infarction, neurological disorders and other medical conditions. Elevated values of these autoantibodies can be associated with recurrent fetal loss, arterial and venous thrombosis and thrombocytopenia. 2. In the present study, we investigated the presence of ACA in three rat strains, namely normal Wistar rats (WR), spontaneously hypertensive rats Okamoto-Aoki (SHR) and stroke-prone SHR (SHRSP). All animals were examined at four ages: 1, 4, 10 and 12 months of age. Anticardiolipin antibodies were determined by ELISA. 3. Anticardiolipin antibody levels in normal WR, which were used as controls, were lowest at 1 month and increased significantly from the 4th month on. At the prehypertensive age (1 month), ACA levels in SHR and SHRSP were significantly higher compared with control WR, decreased with age and were significantly lower at 4, 10 and 12 months compared with age-matched WR. 4. These differences may be a result of immunological disorders in SHR.

[Spontaneous abortions in Sneddon's syndrome]. / Spontanni aborti pri sindrom na Sneddon.

Following the publications on the connection between Sneddon's syndrome and the antiphospholipid++ syndrome, the attention of researchers concentrated on the reproductive problems of women comprising the greater part of patients suffering from Sneddon's syndrome. One of the main indications of these almost coinciding syndromes are miscarriages for no obvious reason. It is supposed that the general mechanism is thrombosis. In a group of 38 patients with Sneddon's syndrome 32 (84%) are women. Twenty-eight of them have altogether given bright 43 times. Seven of them have suffered miscarriages for no obvious reason and three have habitual miscarriages. Three women have sterility problems.

[Application of low-molecular-weight heparin during pregnancy and delivery in case of reproductive failure due to antiphospholipid syndrome]. / Prilozhenie na niskomolekuliaren kheparin po vreme na bremennost i razhdane pri sluchai na reproduktivni neblagopoluchiia, sv'rzani s p'rvichen antifosfolipiden sindrom.

The authors report the results of application of low molecular weight heparin during pregnancy, delivery and after birth to counter placental insufficiency in case of primary antiphospholipid syndrome. The patient had been treated with Fraxiparin for 126 days. No systemic or local side effects were spotted. There were no genital bleeding and tromboembolic accidents during pregnancy. A Caesaerean section was carried out under spinal anaesthesia. Fraxiparin treatment was not halted during the section, which was accompanied by a normal blood loss. The pregnancy yielded an eutrophic newborn and no complications were demonstrated during the neonatal period.

Protein C activity in patients with antiphospholipid syndrome.

The interaction between antiphospholipid antibodies and the protein C system may explain at least a part of the mechanisms underlying thrombosis in the antiphospholipid syndrome (APS). We evaluated the protein C activity, factor V Leiden mutation and the presence of several types of antiphospholipid antibodies in 60 patients with antiphospholipid syndrome. Nineteen patients (31.6%) and 5 controls (8.3%) had decreased protein C activity (95% CI, 10%-37%). 14 patients with high levels of antiphospholipid antibodies and normal factor V molecule (37.8%) and none of the patients with antiphospholipid antibodies in normal ranges and normal factor V molecule had decreased protein C activity (95% CI, 24%-52%; (x 510.4; p 5 0.001). Leiden mutation was found in 5 controls (8.3%) and in 4 patients (8.3%; 95% CI, 8-24%). We found a strong association between decreased protein C activity and presence of anticardiolipin antibodies and antibodies against b2-glycoprotein I. The decreased protein C activity depended on the antibody titre. Although abnormalities of the natural anticoagulant proteins like protein C and protein S seem to be involved in the pathogenesis of the APS, activation of protein C also occurs during the process of clotting. Studies such as we describe may help us identify subsets of patients whose clotting is dependent on protein C-antiphospholipid interactions that may lead to future novel therapies.

Prevalence of IgM-, IgA- and IgG-rheumatoid factors in seronegative polyarticular disease compared to pauciarticular disease in juvenile chronic arthritis as measured by ELISA.

OBJECTIVE: To determine the prevalence of the rheumatoid factor isotypes measured by enzyme-linked immunosorbent assay (ELISA) in polyarticular and pauciarticular juvenile chronic arthritis (JCA), and evaluate the diagnostic test qualities. PATIENTS AND METHODS: 53 patients with JCA (20 with seronegative polyarticular disease at onset, 21 with pauciarticular onset and course of disease and 12 with extended pauciarticular disease), as well as 125 control children (58 healthy controls and 67 patients with other diseases) were tested. ELISA for the detection of IgM-, IgA- and IgG-isotypes of RFs was used. The diagnostic characteristics of the tests were evaluated by means of clinical epidemiology methods. RESULTS: The prevalence of the ELISA for IgG-, IgA-, and IgM-RF for JCA patients vs all controls at optimal cut-off titres was 13%, 29%, and 32%, respectively. The test for IgG-RF was established to be of no significance. IgA-RF had higher prevalence in the polyarticular and extended pauciarticular form, 40% and 33%, respectively. IgM-RF showed a prevalence of 55% for the polyarticular and 42% for the extended pauciarticular form. No significant prevalence has been found in the pauciarticular form. CONCLUSION: Our results indicate that ELISA for IgG-RF is of no diagnostic value for JCA. The ELISAs for IgM- and IgA-RFs demonstrated a diagnostic significance for the polyarticular and extended pauciarticular form. Juvenile chronic arthritis (JCA) is a heterogeneous disease which encompasses different forms defined by the type of onset. There is evidence, supported by immunogenetic studies that the various subgroups may represent distinct disease entities [1, 2]. Numerous immunological abnormalities have been detected in JCA, but the most characteristic serological findings are ANA and IgM-rheumatoid factor, thought to be useful in the classification of patients and their management. Antinuclear antibodies are universal in JCA, most commonly found in children with early onset pauciarthritis and late onset seropositive polyarthritis [1, 2, 3]. In contrast, the IgM-rheumatoid factor, measured by conventional agglutination techniques, is a hallmark only of polyarthritis with late onset resembling adult rheumatoid arthritis. This group of patients with "seropositive" disease represents less than 20% of all JCA children. Of those patients with "seronegative" disease 20-30% have a systemic onset and the remainder have either a pauciarticular or polyarticular form [2, 3]. Following the introduction of more sensitive techniques, it has already been established that rheumatoid arthritis (RA) patients' sera contain not only the "classical" 19S IgM-RF, but also other isotypes of the rheumatoid factor (RF). A number of studies have emphasized the presence of IgG-, IgA-, IgM- and even IgE- RF in patients with "seronegative" RA [4, 5, 6]. The aim of this study is to determine the prevalence of IgG, IgM and IgA RFs and to attempt at evaluating the diagnostic and prognostic qualities of the ELISA-tests for rheumatoid factor isotypes in polyarticular and pauciarticular forms at onset of JCA.