[Type and frequency of cardiac symptoms in patients with pulmonary sarcoidosis]. / Rodzaj i czestosc wystepowania objawów kardiologicznych u chorych na sarkoidoze pluc.

AIM: The aim of the study was to assess the type and frequency of prevalence of cardiac symptoms in patients with pulmonary sarcoidosis. MATERIALS AND METHODS: The study group consisted of 54 patients (21 female, 33 male), with biopsy-proven pulmonary sarcoidosis. Mean age was 45.85 +/-11.77 years. According to time passed from diagnosis of sarcoidosis patients were divided into 2 subgroups. Additionally, age, sex, left ventricular ejection fraction, cigarette smoking and comorbidities were analyzed. Course and stage of sarcoidosis were also included in the analysis. RESULTS: The frequency of cardiovascular symptoms in the study group was 87.04%. 59.26% of patients reported chest pain, the same subjects reported dyspnoea. 48.15% of respondents reported heart palpitations, 33.33% pre-syncope states, 12.96% syncope, and 37.04% edema of lower limbs. There were no statistically significant differences in the incidence of analyzed symptoms, depending on the duration of lung sarcoidosis. CONCLUSIONS: Patients with pulmonary sarcoidosis who report cardiovascular symptoms require further diagnostics.

[Evaluation of the clinical suitability of automated left ventricle's fraction and volume measurements in 3-dimensional echocardiography compared to values obtained in magnetic resonance imaging (pilot study)]. / Ocena przydatnosci klinicznej automatycznego pomiaru frakcji i objetosci lewej komory w echokardiografii przezklatkowej 3D w porównaniu do wartosci uzyskanych w rezonansie magnetycznym (badanie pilotazowe).

Recently a lot of authors have been trying to determine the usefulness of 3-dimensional echocardiography (TTE 3D) in evaluation of ejection fraction and left ventricular volume, but few attempt to compare it to the current gold standard of cardiac magnetic resonance (CMR). 3D imaging technics allows to avoid errors caused by geometry of the heart chambers and foreshortened views. American Echocardiographic Guidelines recommend the use of 3-dimensional echocardiography in daily clinical practice. AIM: The aim of the study was to establish clinical usefulness of automated 3D TTE software to calculate left ventricle ejection fraction (LVEF), left ventricle end diastolic volume (LVEDV) and left ventricle end systolic volume (LVESV), and to compare those measurements calculated in CMR and in 3D TTE. MATERIALS AND METHODS: The aim of the study was to establish clinical usefulness of automated 3D TTE software to calculate left ventricle ejection fraction (LVEF), left ventricle end diastolic volume (LVEDV) and left ventricle end systolic volume (LVESV), and to compare those measurements calculated in CMR and in 3D TTE. RESULTS: The mean LVEF in 3D TTE was 65% +/- 12%; LVEDV 123 ml +/- 67 ml, LVESV 42 ml +/- 29 ml. The CMR LVEF in the study group was 61% +/- 9%, LVEDV 134 ml +/- 51ml, LVESV 54 ml +/- 33 ml. Wilcoxon rank test showed no difference between medians of the measurements, the correlation coefficient between LVEF in 3D TTE and CMR was R = 0.84 (p = 0.036). LVEF calculated in 3D TTE shows good correlation with LVEF computed in CMR. However good visualization of the endocardium, especially in the apex, is essential. The volume of left ventricle is underestimated in 3D TTE. In previous studies underestimation of LVEDV and LVESV was explained by exclusion of endocardial trabeculae from the left ventricle cavity in automated measurement. CONCLUSIONS: The automated 3D TTE software allows simple, fast and precise evaluation of parameters of the left ventricle - especially LVEF. Automated 3D TTE software gives hope for the inclusion of 3D TTE in routine clinical practice due to its repeatability and easy use of the Heart Model software.

[Exercise capacity in patients with pulmonary sarcoidosis]. / Wydolnosc fizyczna u chorych na sarkoidoze pluc.

Exertional dyspnea is a common manifestation of sarcoidosis. Cardiopulmonary exercise testing (CPET) is a useful tool to evaluate exercise tolerance of sarcoid patients. AIM: The aim of the study was to evaluate of exercise capacity in patients with pulmonary sarcoidosis with regard to duration of the disease. Analysis of differences in physical tolerance between patients with cardiac sarcoidosis and without cardiac sarcoidosis. MATERIALS AND METHODS: 39 patients diagnosed with pulmonary sarcoidosis were enrolled to our study. Cardiopulmonary exercise test was used to assess exercise capacity. According to time passed from diagnosis of sarcoidosis patients were grouped into 2 groups. Cardiac sarcoidosis was diagnosed in 9 subjects (23.07%). The control group consisted of 33 healthy volunteers. RESULTS: The results of the ergospirometry test in patients with lung sarcoidosis showed statistically significant differences in comparison to the control group. The age of patients with pulmonary sarcoidosis correlated negatively with maximal heart rate, oxygen consumption at peak exercise and at anaerobic threshold, breathing frequency at peak exercise, minute ventilation at peak exercise and metabolic equivalent. There was a negative, statistically significant correlation between the oxygen pulse at the peak of exercise and the treatment of steroids in the past. CONCLUSIONS: Exercise limitation in patients with pulmonary sarcoidosis is a consequence of both ventilatory and cardiocirculatory impairment. Patients with longer disease history of sarcoidosis achieved worse results at cardiopulmonary exercise test than patients with shorter disease history. Trend towards worse exercise tolerance in patients with cardiac sarcoidosis compared to patients without cardiac sarcoidosis was observed.

Intrinsically unstructured phosphoprotein TSP9 regulates light harvesting in Arabidopsis thaliana.

Thylakoid-soluble phosphoprotein of 9 kDa, TSP9, is an intrinsically unstructured plant-specific protein [Song, J., et al. (2006) Biochemistry 45, 15633-15643] with unknown function but established associations with light-harvesting proteins and peripheries of both photosystems [Hansson, M., et al. (2007) J. Biol. Chem. 282, 16214-16222]. To investigate the function of this protein, we used a combination of reverse genetics and biochemical and fluorescence measurement methods in Arabidopsis thaliana. Differential gene expression analysis of plants with a T-DNA insertion in the TSP9 gene using an array of 24000 Arabidopsis genes revealed disappearance of high light-dependent induction of a specific set of mostly signaling and unknown proteins. TSP9-deficient plants had reduced levels of in vivo phosphorylation of light-harvesting complex II polypeptides. Recombinant TSP9 was phosphorylated in light by thylakoid membranes isolated from the wild-type and mutant plants lacking STN8 protein kinase but not by the thylakoids deficient in STN7 kinase, essential for photosynthetic state transitions. TSP9-lacking mutant and RNAi plants with downregulation of TSP9 showed reduced ability to perform state transitions. The nonphotochemical quenching of chlorophyll fluorescence at high light intensities was also less efficient in the mutant compared to wild-type plants. Blue native electrophoresis of thylakoid membrane protein complexes revealed that TSP9 deficiency increased relative stability of photosystem II dimers and supercomplexes. It is concluded that TSP9 regulates plant light harvesting acting as a membrane-binding protein facilitating dissociation of light-harvesting proteins from photosystem II.

Sulfur starvation in rice: the effect on photosynthesis, carbohydrate metabolism, and oxidative stress protective pathways.

Sulfur-deficient plants generate a lower yield and have a reduced nutritional value. The process of sulfur acquisition and assimilation play an integral role in plant metabolism, and response to sulfur deficiency involves a large number of plant constituents. Rice (Oryza sativa) is the second most consumed cereal grain, and the effects of sulfur deprivation in rice were analyzed by measuring changes in photosynthesis, carbohydrate metabolism, and antioxidants. The photosynthetic apparatus was severely affected under sulfur deficiency. The Chl content was reduced by 49% because of a general reduction of PSII and PSI and the associated light-harvesting antenna. The PSII efficiency was 31% lower at growth light, and the ability of PSI to photoreduce NADP+ was decreased by 61%. The Rubisco content was also significantly reduced in the sulfur-deprived plants. The imbalances between PSII and PSI, and between photosynthesis and carbon fixation led to a general over-reduction of the photosynthetic electron carriers (higher 1-q(P)). Chromatographic analysis showed that the level of monosaccharides was lower and starch content higher in the sulfur-deprived plants. In contrast, no changes in metabolite levels were found in the tricarboxylic acid or Calvin cycle. The level of the thiol-containing antioxidant, GSH, was 70% lower and the redox state was significantly more oxidized. These changes in GSH status led to an upregulation of the cytosolic isoforms of GSH reductase and monodehydroascorbate reductase. In addition, alternative antioxidants like flavonoids and anthocyanins were increased in the sulfur-deprived plants.

An extra-plastidial alpha-glucan, water dikinase from Arabidopsis phosphorylates amylopectin in vitro and is not necessary for transient starch degradation.

Starch phosphorylation catalysed by the alpha-glucan, water dikinases (GWD) has profound effects on starch degradation in plants. The Arabidopsis thaliana genome encodes three isoforms of GWD, two of which are localized in the chloroplast and are involved in the degradation of transient starch. The third isoform, termed AtGWD2 (At4g24450), was heterologously expressed and purified and shown to have a substrate preference similar to potato GWD. Analyses of AtGWD2 null mutants did not reveal any differences in growth or starch and sugar levels, when compared to the wild type. Subcellular localization studies in Arabidopsis leaves and in vitro chloroplast import assays indicated that AtGWD2 was not targeted to the chloroplasts. The AtGWD2 promoter showed a highly restricted pattern of activity, both spatially and temporally. High activity was observed in the companion cells of the phloem, with expression appearing just before the onset of senescence. Taken together, these data indicate that, although AtGWD2 is capable of phosphorylating alpha-glucans in vitro, it is not directly involved in transient starch degradation.

Knock-out of the chloroplast-encoded PSI-J subunit of photosystem I in Nicotiana tabacum.

The plastid-encoded psaJ gene encodes a hydrophobic low-molecular-mass subunit of photosystem I (PSI) containing one transmembrane helix. Homoplastomic transformants with an inactivated psaJ gene were devoid of PSI-J protein. The mutant plants were slightly smaller and paler than wild-type because of a 13% reduction in chlorophyll content per leaf area caused by an approximately 20% reduction in PSI. The amount of the peripheral antenna proteins, Lhca2 and Lhca3, was decreased to the same level as the core subunits, but Lhca1 and Lhca4 were present in relative excess. The functional size of the PSI antenna was not affected, suggesting that PSI-J is not involved in binding of light-harvesting complex I. The specific PSI activity, measured as NADP(+) photoreduction in vitro, revealed a 55% reduction in electron transport through PSI in the mutant. No significant difference in the second-order rate constant for electron transfer from reduced plastocyanin to oxidized P700 was observed in the absence of PSI-J. Instead, a large fraction of PSI was found to be inactive. Immunoblotting analysis revealed a secondary loss of the luminal PSI-N subunit in PSI particles devoid of PSI-J. Presumably PSI-J affects the conformation of PSI-F, which in turn affects the binding of PSI-N. This together renders a fraction of the PSI particles inactive. Thus, PSI-J is an important subunit that, together with PSI-F and PSI-N, is required for formation of the plastocyanin-binding domain of PSI. PSI-J is furthermore important for stability or assembly of the PSI complex.

The properties of the positively charged loop region in PSI-G are essential for its "spontaneous" insertion into thylakoids and rapid assembly into the photosystem I complex.

The PSI-G subunit of photosystem I (PSI) is an 11-kDa membrane protein that plays an important role in electron transport between plastocyanin and PSI and is involved in the stability of the PSI complex. Within the complex, the PSI-G subunit is bound to PSI-B and is in contact with Lhca1. PSI-G has two transmembrane spans connected by a positively charged stromal loop. The loop is inaccessible to proteases, indicating a tightly bound location within the PSI complex. Here, we have studied the insertion mechanism and assembly of PSI-G. We show that the protein inserts into thylakoids by a direct or "spontaneous" pathway that does not involve the activities of any known chloroplast protein-targeting machinery. Surprisingly, the positively charged stromal loop region plays a major role in this process. Mutagenesis or deletions within this region almost invariably lead to a marked lowering of insertion efficiency, strongly indicating a critical role for the loop in the organization of the transmembrane regions prior to or during membrane insertion. Finally, we have examined the assembly of newly inserted PSI-G into the PSI complex, since very little is known of the assembly pathway for this large multimeric complex. Interestingly, we find that inserted PSI-G can be found within the full PSI complex within the import assay time frame after insertion into thylakoids, strongly suggesting that PSI-G normally associates at the end of the assembly process. This is consistent with its location on the periphery of the complex.

Structural characterization of a complex of photosystem I and light-harvesting complex II of Arabidopsis thaliana.

Chloroplasts are central to the provision of energy for green plants. Their photosynthetic membrane consists of two major complexes converting sunlight: photosystem I (PSI) and photosystem II (PSII). The energy flow toward both photosystems is regulated by light-harvesting complex II (LHCII), which after phosphorylation can move from PSII to PSI in the so-called state 1 to state 2 transition and can move back to PSII after dephosphorylation. To investigate the changes of PSI and PSII during state transitions, we studied the structures and frequencies of all major membrane complexes from Arabidopsis thaliana chloroplasts at conditions favoring either state 1 or state 2. We solubilized thylakoid membranes with digitonin and analyzed the complete set of complexes immediately after solubilization by electron microscopy and image analysis. Classification indicated the presence of a PSI-LHCII supercomplex consisting of one PSI-LHCI complex and one LHCII trimer, which was more abundant in state 2 conditions. The presence of LHCII was confirmed by excitation spectra of the PSI emission of membranes in state 1 or state 2. The PSI-LHCII complex could be averaged with a resolution of 16 A, showing that LHCII has a specific binding site at the PSI-A, -H, -L, and -K subunits.

Insertion of the plant photosystem I subunit G into the thylakoid membrane.

Subunit G of photosystem I is a nuclear-encoded protein, predicted to form two transmembrane alpha-helices separated by a loop region. We use in vitro import assays to show that the positively charged loop domain faces the stroma, whilst the N- and C-termini most likely face the lumen. PSI-G constructs in which a His- or Strep-tag is placed at the C-terminus or in the loop region insert with the same topology as wild-type photosystem I subunit G (PSI-G). However, the presence of the tags in the loop make the membrane-inserted protein significantly more sensitive to trypsin, apparently by disrupting the interaction between the loop and the PSI core. Knock-out plants lacking PSI-G were transformed with constructs encoding the C-terminal and loop-tagged PSI-G proteins. Experiments on thylakoids from the transgenic lines show that the C-terminally tagged versions of PSI-G adopt the same topology as wild-type PSI-G, whereas the loop-tagged versions affect the sensitivity of the loop region to trypsin, thus confirming the in vitro observations. Furthermore, purification of PSI complexes from transgenic plants revealed that all the tagged versions of PSI-G are incorporated and retained in the PSI complex, although the C-terminally tagged variants of PSI-G were preferentially retained. This suggests that the loop region of PSI-G is important for proper integration into the PSI core. Our experiments demonstrate that it is possible to produce His- and Strep-tagged PSI in plants, and provide further evidence that the topology of membrane proteins is dictated by the distribution of positive charges, which resist translocation across membranes.

Photosystem I lacking the PSI-G subunit has a higher affinity for plastocyanin and is sensitive to photodamage.

PSI-G is an 11 kDa subunit of PSI in photosynthetic eukaryotes. Arabidopsis thaliana plants devoid of PSI-G have a decreased PSI content and an increased activity of NADP(+) photoreduction in vitro but otherwise no obvious phenotype. To investigate the biochemical basis for the increased activity, the kinetic parameters of the reaction between PSI and plastocyanin were determined. PSI-G clearly plays a role in the affinity for plastocyanin since the dissociation constant (K(D)) is only 12 muM in the absence of PSI-G compared to 32 muM for the wild type. On the physiological level, plants devoid of PSI-G have a more reduced Q(A). This indicates that the decreased PSI content is due to unstable PSI rather than an adaptation to the increased activity. In agreement with this indication of decreased stability, plants devoid of PSI-G were found to be more photo-inhibited both at low temperature and after high light treatment. The decreased PSI stability was confirmed in vitro by measuring PSI activity after illumination of a thylakoid suspension which clearly showed a faster decrease in PSI activity in the thylakoids lacking PSI-G. Light response of the P700 redox state in vivo showed that in the absence of PSI-G, P700 is more reduced at low light intensities. We conclude that PSI-G is involved in the binding dynamics of plastocyanin to PSI and that PSI-G is important for the stability of the PSI complex.