Some non-pathogenic trypanosomatids maintain a mutualistic relationship with a betaproteobacterium of the Alcaligenaceae family. Intensive nutritional exchanges have been reported between the two ...partners, indicating that these protozoa are excellent biological models to study metabolic co-evolution. We previously sequenced and herein investigate the entire genomes of five trypanosomatids which harbor a symbiotic bacterium (SHTs for Symbiont-Haboring Trypanosomatids) and the respective bacteria (TPEs for Trypanosomatid Proteobacterial Endosymbiont), as well as two trypanosomatids without symbionts (RTs for Regular Trypanosomatids), for the presence of genes of the classical pathways for vitamin biosynthesis. Our data show that genes for the biosynthetic pathways of thiamine, biotin, and nicotinic acid are absent from all trypanosomatid genomes. This is in agreement with the absolute growth requirement for these vitamins in all protozoa of the family. Also absent from the genomes of RTs are the genes for the synthesis of pantothenic acid, folic acid, riboflavin, and vitamin B6. This is also in agreement with the available data showing that RTs are auxotrophic for these essential vitamins. On the other hand, SHTs are autotrophic for such vitamins. Indeed, all the genes of the corresponding biosynthetic pathways were identified, most of them in the symbiont genomes, while a few genes, mostly of eukaryotic origin, were found in the host genomes. The only exceptions to the latter are: the gene coding for the enzyme ketopantoate reductase (EC:1.1.1.169) which is related instead to the Firmicutes bacteria; and two other genes, one involved in the salvage pathway of pantothenic acid and the other in the synthesis of ubiquinone, that are related to Gammaproteobacteria. Their presence in trypanosomatids may result from lateral gene transfer. Taken together, our results reinforce the idea that the low nutritional requirement of SHTs is associated with the presence of the symbiotic bacterium, which contains most genes for vitamin production.
Abstract The aim of this study was to investigate the suitability of the North Star Ambulatory Assessment as a possible outcome measure in multicentric clinical trials. More specifically we wished to ...investigate the level of training needed for achieving a good interobserver reliability in a multicentric setting. The scale was specifically designed for ambulant children with Duchenne Muscular Dystrophy and includes 17 items that are relevant for this cohort. Thirteen Italian centers participated in the study. In the first phase of the study we provided two training videos and an example of the scale performed on a child. After the first session of training, all the 13 examiners were asked to send a video with an assessment performed in their centre and to score all the videos collected. There were no difficulties in performing the items and in obtaining adequate videos with a hand held camera but the results showed a poor interobserver reliability (<.5). After a second training session with review and discussion of the videos previously scored, the same examiners were asked to score three new videos. The results of this session had an excellent interobserver reliability (.995). The level of agreement was maintained even when the same videos were rescored after a month, showing a significant intra-observer reliability (.95). Our results suggest that the NSAA is a test that can be easily performed, completed in 10 min and can be used in a multicentric setting, providing that adequate training is administered.
Trypanosomatids of the genera Angomonas and Strigomonas live in a mutualistic association characterized by extensive metabolic cooperation with obligate endosymbiotic Betaproteobacteria. However, the ...role played by the symbiont has been more guessed by indirect means than evidenced. Symbiont-harboring trypanosomatids, in contrast to their counterparts lacking symbionts, exhibit lower nutritional requirements and are autotrophic for essential amino acids. To evidence the symbiont's contributions to this autotrophy, entire genomes of symbionts and trypanosomatids with and without symbionts were sequenced here.
Analyses of the essential amino acid pathways revealed that most biosynthetic routes are in the symbiont genome. By contrast, the host trypanosomatid genome contains fewer genes, about half of which originated from different bacterial groups, perhaps only one of which (ornithine cyclodeaminase, EC:4.3.1.12) derived from the symbiont. Nutritional, enzymatic, and genomic data were jointly analyzed to construct an integrated view of essential amino acid metabolism in symbiont-harboring trypanosomatids. This comprehensive analysis showed perfect concordance among all these data, and revealed that the symbiont contains genes for enzymes that complete essential biosynthetic routes for the host amino acid production, thus explaining the low requirement for these elements in symbiont-harboring trypanosomatids. Phylogenetic analyses show that the cooperation between symbionts and their hosts is complemented by multiple horizontal gene transfers, from bacterial lineages to trypanosomatids, that occurred several times in the course of their evolution. Transfers occur preferentially in parts of the pathways that are missing from other eukaryotes.
We have herein uncovered the genetic and evolutionary bases of essential amino acid biosynthesis in several trypanosomatids with and without endosymbionts, explaining and complementing decades of experimental results. We uncovered the remarkable plasticity in essential amino acid biosynthesis pathway evolution in these protozoans, demonstrating heavy influence of horizontal gene transfer events, from Bacteria to trypanosomatid nuclei, in the evolution of these pathways.
Glycosomes are peroxisome-related organelles that have been identified in kinetoplastids and diplonemids. The hallmark of glycosomes is their harboring of the majority of the glycolytic enzymes. Our ...biochemical studies and proteome analysis of
glycosomes have located, in addition to enzymes of the glycolytic pathway, enzymes of several other metabolic processes in the organelles. These analyses revealed many aspects in common with glycosomes from other trypanosomatids as well as features that seem specific for
. Their enzyme content indicates that
glycosomes are multifunctional organelles, involved in both several catabolic processes such as glycolysis and anabolic ones. Specifically discussed in this minireview are the cross-talk between glycosomal metabolism and metabolic processes occurring in other cell compartments, and the importance of metabolite translocation systems in the glycosomal membrane to enable the coordination between the spatially separated processes. Possible mechanisms for metabolite translocation across the membrane are suggested by proteins identified in the organelle's membrane-homologs of the ABC and MCF transporter families-and the presence of channels as inferred previously from the detection of channel-forming proteins in glycosomal membrane preparations from the related parasite
. Together, these data provide insight in the way in which different parts of
metabolism, although uniquely distributed over different compartments, are integrated and regulated. Moreover, this information reveals opportunities for the development of drugs against Chagas disease caused by these parasites and for which currently no adequate treatment is available.
The research on a class of asymptotic exit-time problems with a vanishing Lagrangian, begun in M. Motta and C. Sartori, Nonlinear Differ. Equ. Appl. Springer (2014). for the compact control case, is ...extended here to the case of unbounded controls and data, including both coercive and non-coercive problems. We give sufficient conditions to have a well-posed notion of generalized control problem and obtain regularity, characterization and approximation results for the value function of the problem.
Purpose
This study was designed to describe the clinical, laboratorial, and urodynamic findings of prostatic artery embolization (PAE) in patients with urinary retention due to benign prostatic ...hyperplasia (BPH).
Methods
A prospective study of 11 patients with urinary retention due to BPH was conducted. Patients underwent physical examination, prostate specific antigen (PSA) measurement, transrectal ultrasound, and magnetic resonance imaging. International prostate symptom score (IPSS), quality of life (QoL), and urodynamic testing were used to assess the outcome before and after 1 year.
Results
Clinical success was 91 % (10/11 patients) with a mean follow-up of 22.3 months (range, 12–41 months). At the first year follow-up, the mean IPSS score was 2.8 points (
p
= 0.04), mean QoL was 0.4 points (
p
= 0.001), mean PSA decreased from 10.1 to 4.3 ng/mL (
p
= 0.003), maximum urinary flow (Qmax) improved from 4.2 to 10.8 mL/sec (
p
= 0.009), and detrusor pressure (Pdet) decreased from 85.7 to 51.5 cm H
2
O (
p
= 0.007). Before PAE, Bladder Outlet Obstruction Index (BOOI) showed values >40 in 100 % of patients. After PAE, 30 % of patients were >40 (obstructed), 40 % were between 20 and 40 (undetermined), and 30 % were <20 (unobstructed). Patients with a BOOI <20 had higher PSA values at 1-day after PAE.
Conclusions
Clinical and urodynamic parameters improved significantly after PAE in patients with acute urinary retention due to BPH. Total PSA at day 1 after PAE was higher in patients with unobstructed values in pressure flow studies.
Genome-editing technology has emerged as a powerful method that enables the generation of genetically modified cells and organisms necessary to elucidate gene function and mechanisms of human ...diseases. The clustered regularly interspaced short palindromic repeats- (CRISPR-) associated 9 (Cas9) system has rapidly become one of the most popular approaches for genome editing in basic biomedical research over recent years because of its simplicity and adaptability. CRISPR/Cas9 genome editing has been used to correct DNA mutations ranging from a single base pair to large deletions in both in vitro and in vivo model systems. CRISPR/Cas9 has been used to increase the understanding of many aspects of cardiovascular disorders, including lipid metabolism, electrophysiology and genetic inheritance. The CRISPR/Cas9 technology has been proven to be effective in creating gene knockout (KO) or knockin in human cells and is particularly useful for editing induced pluripotent stem cells (iPSCs). Despite these progresses, some biological, technical, and ethical issues are limiting the therapeutic potential of genome editing in cardiovascular diseases. This review will focus on various applications of CRISPR/Cas9 genome editing in the cardiovascular field, for both disease research and the prospect of in vivo genome-editing therapies in the future.
In trypanosomatids, the kinetoplast is the portion of the single mitochondrion that is connected to the basal body and contains the kDNA, a network composed by circular and interlocked DNA. The kDNA ...packing is conducted by Kinetoplast Associated Proteins (KAPs), which are similar to eukaryotic histone H1. In symbiont-harboring trypanosomatids (SHTs) such as Angomonas deanei and Strigomonas culicis, a ß-proteobacterium co-evolves with the host in a mutualistic relationship. The prokaryote confers nutritional benefits to the host and affects its cell structure. Atomic force microscopy showed that the topology of isolated kDNA networks is quite similar in the two SHT species. Ultrastructural analysis using high-resolution microscopy techniques revealed that the DNA fibrils are more compact in the kinetoplast region that faces the basal body and that the presence of the symbiotic bacterium does not interfere with kDNA topology. However, RT-PCR data revealed differences in the expression of KAPs in wild-type protozoa as compared to aposymbiotic cells. Immunolocalization showed that different KAPs present distinct distributions that are coincident in symbiont-bearing and in symbiont-free cells. Although KAP4 and KAP7 are shared by all trypanosomatid species, the expanded repertoire of KAPs in SHTs can be used as phylogenetic markers to distinguish different genera.
Gaucher disease (GD) is characterized by accumulation of glucosylceramide (GC) in the cells of monocyte/macrophage system. The degradation of GC is controlled by glucosylceramidase (GCase) and ...saposin (Sap) C, a member of a family of four small glycoproteins (Saps A, B, C and D), all derived by proteolytic processing of a common precursor, prosaposin (PSAP). Saps contain six cysteine residues, forming three disulfide bridges, that affect their structure and function. Sap C is an essential activator of GCase and its deficit impairs the GCase activity causing GD. In the present study the biological properties of cells from four recently described GD patients carrying mutations in the Sap C domain of the PSAP gene have been characterized. Two patients had mutations involving a cysteine residue, whereas the other two had a L349P mutation. It was found that: (i) in the four Sap C-deficient cells PSAP was normally processed and sorted, the lack of Sap C being mainly due to the Sap C instability in late endosomal/lysosomal environment; (ii) the decrease/absence of Sap C affected the GCase intracellular localization; (iii) the lowest level of Sap C and enhanced autophagy were observed in the cells, which carried a Sap C mutation involving a cysteine residue; (iv) the four Sap C-deficient fibroblasts stored GC, ceramide and cholesterol, the last two lipids being clearly localized in lysosomes; (v) a correlation was observed between the type of Sap C mutation and the Gaucher phenotype: apparently, mutations involving cysteine residues lead to a neurological variant of GD.
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