Biomedical subjects

Vincent Thomas

Publications and source records attributed to Vincent Thomas.

11 recordsLinked to original sources

PubMed2006

Estimation of arterial PCO2 from a lung model during ramp exercise in healthy young subjects.

The aim of this study is to propose a new approach to estimate non-invasively arterial carbon dioxide partial pressure (P(a)CO2) approach was based on the reconstruction of alveolar gas composition over each breath from a tidally ventilated lung model (P(M)(CO2)). Eight healthy young subjects were studied during a ramp exercise test on a cycle ergometer. Arterial samples were drawn at rest and every minute during the exercise test for determination of P(a)CO2 . P(a)CO2 was compared with indirect estimates of P(CO2) : P(M)(CO2), end-tidal P(CO2) (P(ET)(CO2)) and an empirical equation involving P(ET)(CO2) and tidal volume (P(J)(CO2)). The difference between estimated and measured P(a)CO2 on the whole ramp exercise was -0.3+/-1.9mmHg for P(M)(CO2), 1.0+/-2.2mmHg for P(ET)(CO2) and -1.7+/-1.7mmHg for P(J)(CO2) . P(ET)(CO2) and P(J)(CO2) were significantly different from actual P(a)CO2 (P<0.001). It is concluded that, on the basis of the bias, the breathing lung model gave better estimates of P(a)CO2 than the two other indirect methods during ramp exercise.

Adult↗

PubMed2006

Modifications to the Arabidopsis defense proteome occur prior to significant transcriptional change in response to inoculation with Pseudomonas syringae.

Alterations in the proteome of Arabidopsis (Arabidopsis thaliana) leaves during responses to challenge by Pseudomonas syringae pv tomato DC3000 were analyzed using two-dimensional gel electrophoresis. Protein changes characteristic of the establishment of disease, basal resistance, and resistance-gene-mediated resistance were examined by comparing responses to DC3000, a hrp mutant, and DC3000 expressing avrRpm1, respectively. The abundance of each protein identified was compared with that of selected transcripts obtained from comparable GeneChip experiments. We report changes in three subcellular fractions: total soluble protein, chloroplast enriched, and mitochondria enriched over four time points (1.5-6 h after inoculation). In total, 73 differential spots representing 52 unique proteins were successfully identified. Many of the changes in protein spot density occurred before significant transcriptional reprogramming was evident between treatments. The high proportion of proteins represented by more than one spot indicated that many of the changes to the proteome can be attributed to posttranscriptional modifications. Proteins found to show significant change after bacterial challenge are representative of two main functional groups: defense-related antioxidants and metabolic enzymes. Significant changes to photosystem II and to components of the mitochondrial permeability transition were also identified. Rapid communication between organelles and regulation of primary metabolism through redox-mediated signaling are supported by our data.

Arabidopsis↗

PubMed2006

'Candidatus Rhabdochlamydia crassificans', an intracellular bacterial pathogen of the cockroach Blatta orientalis (Insecta: Blattodea).

The genus Rickettsiella comprises various intracellular bacterial pathogens of arthropods, exhibiting a chlamydia-like developmental cycle. Species may be divided into two main groups, the R. popilliae-R. grylli group and the R. chironomi group. Previous phylogenetic studies based on the 16S ribosomal RNA encoding gene showed that two Rickettsiella species, one from each group, belong in reality to two distantly related lineages, the gamma-Proteobacteria (R. grylli) and the Chlamydiales ('Candidatus Rhabdochlamydia porcellionis', a pathogen of terrestrial isopods). In the present work, the 16S rDNA sequence of another Rickettsiella-like species, causing abdominal swelling to its cockroach host Blatta orientalis, was determined and phylogenetic analysis performed. Identical 16S rDNA sequences of 1495 nucleotides were obtained from fat body and ovary tissues of both healthy and diseased cockroach individuals. The sequence shared only 73% of similarity with R. grylli, but 82-87% with most Chlamydiales, and even 96.3% with 'Candidatus Rhabdochlamydia porcellionis'. Phylogenetic analyses confirmed the affiliation of the cockroach pathogen within the order Chlamydiales, and based on ultrastructural characteristics and genetic analyses, we propose its inclusion in the 'Candidatus Rhabdochlamydia' as a distinct taxon, 'Candidatus Rhabdochlamydia crassificans'. These results extend our knowledge of the phylogenetic diversity of the Chlamydiales.

Animals↗

PubMed2006

Development and evaluation of a modified colorimetric solid-phase microassay for measuring the activity of cellular and plasma (Factor XIII) transglutaminases.

Plasma TG (transglutaminase) [FXIII (Factor XIII)] stabilizes fibrin and plays an essential role in haemostasis. In the present paper, we report a simple colorimetric assay for measuring FXIII activity. The advantage of this approach, compared with all the other solid-phase assays described so far for measuring TG activity, is that the first substrate, namely the synthetic dipeptide, N-benzyloxycarbonyl-L-Gln-L-Gly, is coupled covalently by its C-terminus with amine-substituted polystyrene plates. Covalent coupling eliminates the problem of desorption of proteins such as casein and N,N'-dimethylcasein (first substrates) when 'blocking' buffers containing proteins, e.g. BSA, are employed, or when samples of plasma or cell homogenates are used. The principle of the assay itself is based on the incorporation of the well-known second substrate, 5-(biotinamido)pentylamine, into the gamma-carboxamide of glutamine in the immobilized dipeptide. The amount of biotinylated amine bound to the plate, as measured by the phosphatase activity of Extravidin phosphatase attached to the biotin moiety, is directly proportional to the TG activity. The method shows strong correlation (r = 0.96) with the radiometric assay for FXIII activity. For plasma samples, a linear response was obtained in the range 0-1.33 i.u. (international unit)/ml, versus the Behring standard. In a preliminary clinical investigation, the method was applied to normal and pathological plasma samples from patients with liver failure and disseminated intravascular coagulation. In the isolation of TG from guinea-pig liver, it was used to measure enzyme activity after each purification step. This method is rapid, sensitive and can easily be applied to routine clinical analyses and to specific research problems.

Animals↗

PubMed2006

Criblamydia sequanensis, a new intracellular Chlamydiales isolated from Seine river water using amoebal co-culture.

Accumulating evidence supports a role for Chlamydia-related organisms as emerging pathogens for human and animals. Assessment of their pathogenicity requires strain availability, at least for animal models and serological studies. As these obligate intracellular species are able to grow inside amoebae, we used co-culture with Acanthamoeba castellanii in an attempt to recover new Chlamydia-related species from river water. We isolated two strains from eight water samples. The first strain is a new Parachlamydia acanthamoebae strain that differs from previously described isolates by only two bases in the complete 16S rRNA gene sequence. The second isolate is the first representative of a new Chlamydiales family, as demonstrated by genetic and phylogenetic analyses of the 16S rRNA, 23S rRNA, ADP/ATP translocase and RnpB encoding genes. Using fluorescent in situ hybridization and electron microscopy, we demonstrated that it grows in high numbers in amoebae, where it exhibits a Chlamydia-like developmental cycle with reticulate bodies and star-like elementary bodies. Based on these results, we propose to name this new species 'Criblamydia sequanensis'. This work confirmed that amoebal co-culture is a relevant method to isolate new chlamydiae, and that it can be successfully applied to ecosystems colonized with a complex microbial community.

Amoeba↗

PubMed2006

Biodiversity of amoebae and amoeba-resisting bacteria in a hospital water network.

Free-living amoebae (FLA) are ubiquitous organisms that have been isolated from various domestic water systems, such as cooling towers and hospital water networks. In addition to their own pathogenicity, FLA can also act as Trojan horses and be naturally infected with amoeba-resisting bacteria (ARB) that may be involved in human infections, such as pneumonia. We investigated the biodiversity of bacteria and their amoebal hosts in a hospital water network. Using amoebal enrichment on nonnutrient agar, we isolated 15 protist strains from 200 (7.5%) samples. One thermotolerant Hartmannella vermiformis isolate harbored both Legionella pneumophila and Bradyrhizobium japonicum. By using amoebal coculture with axenic Acanthamoeba castellanii as the cellular background, we recovered at least one ARB from 45.5% of the samples. Four new ARB isolates were recovered by culture, and one of these isolates was widely present in the water network. Alphaproteobacteria (such as Rhodoplanes, Methylobacterium, Bradyrhizobium, Afipia, and Bosea) were recovered from 30.5% of the samples, mycobacteria (Mycobacterium gordonae, Mycobacterium kansasii, and Mycobacterium xenopi) were recovered from 20.5% of the samples, and Gammaproteobacteria (Legionella) were recovered from 5.5% of the samples. No Chlamydia or Chlamydia-like organisms were recovered by amoebal coculture or detected by PCR. The observed strong association between the presence of amoebae and the presence of Legionella (P < 0.001) and mycobacteria (P = 0.009) further suggests that FLA are a reservoir for these ARB and underlines the importance of considering amoebae when water control measures are designed.

Alphaproteobacteria↗

PubMed2005

Comparison between maximal power in the power-endurance relationship and maximal instantaneous power.

The purpose of this study was to analyze the relevance of introducing the maximal power (P(m)) into a critical-power model. The aims were to compare the P(m) with the instantaneous maximal power (P(max)) and to determine how the P(m) affected other model parameters: the critical power ( P(c)) and a constant amount of work performed over P(c)(W'). Twelve subjects [22.9 (1.6) years, 179 (7) cm, 74.1 (8.9) kg, 49.4 (3.6) ml/min/kg] completed one 15 W/min ramp test to assess their ventilatory threshold (VT), five or six constant-power to exhaustion tests with one to measure the maximal accumulated oxygen deficit (MAOD), and six 5-s all-out friction-loaded tests to measure P(max) at 75 rpm, which was the pedaling frequency during tests. The power and time to exhaustion values were fitted to a 2-parameter hyperbolic model (NLin-2), a 3-parameter hyperbolic model (NLin-3) and a 3-parameter exponential model (EXP). The P(m) values from NLin-3 [760 (702) W] and EXP [431 (106) W] were not significantly correlated with the P(max) at 75 rpm [876 (82) W]. The P(c) value estimated from NLin-3 [186 (47) W] was not significantly correlated with the power at VT [225 (32) W], contrary to other models ( P <0.001). The W' from NLin-2 [25.7 (5.7) kJ] was greater than the MAOD [14.3 (2.7) kJ, P < 0.001] with a significant correlation between them (R = 0.76, P <0.01). For NLin-3, computation of W (P > P c), the amount of work done over P(C), yielded results similar to the W' value from NLin-2: 27.8 (7.4) kJ, which correlated significantly with the MAOD (R = 0.72, P <0.01). In conclusion, the P(m) was not related to the maximal instantaneous power and did not improve the correlations between other model parameters and physiological variables.

Adult↗

PubMed2005

Estimation of the parameters of the relationship between power and time to exhaustion from a single ramp test.

This study aimed to estimate the power/time relationship from a single ramp test (RT) assuming critical power (Pc) from ventilatory threshold (VT) and energy reserve (W') from total work during RT These estimates from single RT were compared to those from a series of 4 constant power exercises (CPT) and from a series of 4 RT. Only W' from CPT was higher than from the series of RT and from single RT using VT (p < 0.05).

Adult↗

PubMed2004

Definition of the fine specificity of the monoclonal antibody 81D4: its reactivity with lysine and polyamine isopeptide cross-links.

The 81D1C2 monoclonal antibody (Mab) directed against the Nepsilon-(gamma-L-glutamyl)-L-lysine isopeptide was found to cross-react on Enzyme Immuno Assay (EIA) with acylated lysines. Using a differential screening EIA procedure, a new Mab 81D4 was selected, which did not cross-react with acylated lysines but exhibited strong reactivity with Nepsilon-(gamma-L-glutamyl)-L-lysine formed by covalently coupling the gamma-carboxyl of NalphaCBZ OtBu glutamic acid to epsilon-NH2 derivatized microtiter plates. When Nepsilon-(gamma-L-glutamyl)-L-lysine isopeptides were generated on gamma-carboxyl derivatized plates, only lysine isopeptides with blocked alpha-amines were reactive, regardless of whether the bond formed by the amine blocking agent was a carbamate with carbobenzyloxychloride or an amide with acetic anhydride. The 81D4 Mab showed little or no affinity for free Nepsilon-(gamma-L-glutamyl)-L-lysine (IC50>5 mM), for N1 or N4 mono(gamma-Poly L-glutamyl)putrescine, and for N1 mono(gamma-Poly L-glutamyl)spermidine (IC50>5 mM). However, when these same isopeptides were synthesized as cross-links between two protein chains--Nepsilon-(gamma-L-glutamyl)-L-lysine between Poly L-glutamate and Poly L-lysine; N1N4 -bis(gamma-Poly L-glutamyl)putrescine, N1N8 -bis(gamma-Poly L-glutamyl)spermidine between Poly-L-glutamate chains--very good reactivity was observed (IC50 400 microM for lysine; 80 microM for putrescine and spermidine). In addition to the chemically synthesized isopeptide cross-links that were recognized by this Mab, the naturally occurring Nepsilon-(ã-L-glutamyl)-L-lysine isopeptide cross-links in D-dimer, which are formed by the action of plasma transglutaminase (Factor XIII) on fibrin, were also detected on immunoblots using 81D4 as the primary antibody.

Animals↗

PubMed2004

Specific changes in the Arabidopsis proteome in response to bacterial challenge: differentiating basal and R-gene mediated resistance.

Alterations in the proteome of Arabidopsis thaliana leaves during early responses to challenge by Pseudomonas syringae pv. tomato DC3000 (DC3000) were analysed using two-dimensional (2D) gel electrophoresis. Protein changes characteristic of the establishment of basal resistance and R-gene mediated resistance were examined by comparing responses to DC3000, a hrp mutant and DC3000 expressing avrRpm1 respectively. The abundance of selected transcripts was also analysed in GeneChip experiments. Here we present data from the soluble fraction of leaf protein, highlighting changes in two antioxidant enzyme groups; the glutathione S-transferases (GSTs F2, F6, F7 and F8) and peroxiredoxins (PrxA, B and IIE). Members of both enzyme groups showed signs of specific post-translational modifications, represented by multiple spots on gels. We suggest that oxidation of specific residues is responsible for some of the spot shifts. All forms of the GST proteins identified here increased following inoculation with bacteria. GSTF8 showed particularly dynamic responses to pathogen challenge, the corresponding transcript was significantly up-regulated by 2 h after inoculation, and the protein showed post-translational modifications specific to an incompatible interaction. Differential changes were observed with the peroxiredoxin proteins; PrxIIE and to a lesser extent PrxB, no change was observed with PrxA, but a truncated form PrxA-L was greatly reduced in abundance following bacterial challenges. Our data suggest that bacterial challenge generally induces Prxs and the antioxidants GSTs, however individual members of these families may be specifically modified dependent upon the virulence of the DC3000 strain and outcome of the interaction. Finally, proteomic and transcriptomic data derived from the same inoculation system are compared and the advantages offered by 2D gel analysis discussed in light of our results.

Arabidopsis↗

PubMed2004

4-methylthio 2-oxobutanoate transaminase: a specific target for antiproliferative agents.

We have previously shown that the addition of 4-methylthio-2-oxobutanoate (MTOB) to cultures of methionine dependent neoplastic cells which lack endogenous MTOB restores their capacity to grow in the absence of exogenous methionine. Transition state inhibitors of the MTOB transaminase,responsible for the transamination of MTOB to methionine, had also been designed and selected for their capacity to inhibit the proliferation of methionine dependent neoplastic cells but not that of normal cells in culture. We now show that the transition state analogue : L-methionine ethyl esterpyridoxal(MEEP) with a structure corresponding to the oxo acid receptor covalently linked to pyridoxamine and the amine donor analogue: D-aspartate beta hydroxamate (D-AH) are efficient inhibitors of MTOB transaminase. [3H] MEEP uptake into transformed HeLa cells is similar to that in normal MRC5 cells, yet growth inhibition is seem in the transformed but not in the normal cells.MEEP irreversibly inhibits the activity of this enzyme when added to HeLa cells in culture but not that of the purified rat liver enzyme, probably due to pyridoxal phosphate already bound in the active site. On the contrary, D-AH is a noncompetitive reversible inhibitor of the purified rat liver enzyme in vitro and also inhibits intracellular HeLa MTOB transaminase. Furthermore, in HeLa cells both inhibitors induce DNA strand breaks typical of apoptotic cell death. These results provide evidence that MTOB transaminase is a potential target for antiproliferative agents which could selectively affect methionine-dependent neoplastic cells. The transition state intermediale : MEEP as an amine acceptor analogue was found to be 20 fold more effective than D-AH as the amine donor analogue in inducing apoptosis.

Animals↗