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Biomedical subjects

M Chamorro

Publications and source records attributed to M Chamorro.

At least 19 recordsLinked to original sources

The sweating response of elite professional soccer players to training in the heat.

Sweat rate and sweat composition vary extensively between individuals, and quantification of these losses has a role to play in the individualisation of a hydration strategy to optimise training and competitive performance. Data were collected from 26 male professional football (soccer) players during one 90 min pre-season training session. This was the 2nd training session of the day, carried out between 19.30 and 21.00 h when the mean +/- SD environment was 32 +/- 3 degrees C, 20 +/- 5 %rh and WBGT 22 +/- 2 degrees C. Training consisted of interval running and 6-a-side games during which the average heart rate was 136 +/- 7 bpm with a maximum rate of 178 +/- 7 bpm (n = 19). Before and after training all players were weighed nude. During training all players had free access to sports drinks (Gatorade) and mineral water (Solan de Cabras). All drink bottles were weighed before and after training. Players were instructed to drink only from their own bottles and not to spit out any drink. No player urinated during the training session. Sweat was collected by patches from the chest, arm, back, and thigh of a subgroup of 7 players. These remained in place for the first 15 - 30 min of the training session, and sweat was analysed for sodium (Na (+)) and potassium (K (+)) concentration. Body mass loss was 1.23 +/- 0.50 kg (ranging from 0.50 to 2.55 kg), equivalent to dehydration of 1.59 +/- 0.61 % of pre-training body mass. The sweat volume lost was 2193 +/- 365 ml (1672 to 3138 ml), but only 972 +/- 335 ml (239 to 1724 ml) of fluid was consumed. 45 +/- 16 % of the sweat volume loss was replaced, but this ranged from 9 % to 73 %. The Na (+) concentration of the subgroup's sweat was 30.2 +/- 18.8 mmol/l (15.5 to 66.3 mmol/l) and Na (+) losses averaged 67 +/- 37 mmol (26 to 129 mmol). The K (+) concentration of the sweat was 3.58 +/- 0.56 mmol/l (2.96 to 4.50 mmol/l) and K (+) losses averaged 8 +/- 2 mmol (5 to 12 mmol). The drinking employed by these players meant that only 23 +/- 21 % of the sweat Na (+) losses were replaced: This ranged from replacing virtually none (when water was the only drink) to replacing 62 % when the sports drink was consumed. These elite soccer players did not drink sufficient volume to replace their sweat loss. This, however, is in accord with data in the literature from other levels of soccer players and athletes in other events. These measurements allow for an individualisation of the club's hydration strategy.

Adult↗

Measurement of serum levels of eosinophil cationic protein in the diagnosis of acute gastrointestinal anisakiasis.

Thirty-two patients with abdominal pain and/or intestinal pseudo-obstruction who had consumed raw or undercooked fish in the previous 72 h, were included in a study of anisakiasis, a parasitation of the human gastrointestinal tract by third stage Anisakis simplex larvae. Skin prick test (SPT) against A. simplex were positive in all the patients. High median eosinophil cationic protein (ECP) serum concentrations (> 15 mg/L) at day 0 with normal serum levels at day 30 and a rise of median total and specific IgE against A. simplex at day 30, were observed. We conclude that a raised serum level of ECP in the first 72 h from the onset of symptoms coinciding with a positive SPT against A. simplex and high total and specific immunoglobulin (IgE) in the first month after the parasitation, could be a useful tool in the diagnosis of gastrointestinal anisakiasis, even if the parasite cannot be isolated.

Acute Disease↗

Requirements for activation and RAFT localization of the T-lymphocyte kinase Rlk/Txk.

BACKGROUND: The Tec family kinases are implicated in signaling from lymphocyte antigen receptors and are activated following phosphorylation by Src kinases. For most Tec kinases, this activation requires an interaction between their pleckstrin homology (PH) domains and the products of phosphoinositide 3-Kinase, which localizes Tec kinases to membrane RAFTs. Rlk/Txk is a Tec related kinase expressed in T cells that lacks a pleckstrin homology domain, having instead a palmitoylated cysteine-string motif. To evaluate Rlk's function in T cell receptor signaling cascades, we examined the requirements for Rlk localization and activation by Src family kinases. RESULTS: We demonstrate that Rlk is also associated with RAFTs, despite its lack of a pleckstrin homology domain. Rlk RAFT association requires the cysteine-string motif and is independent of PI3 Kinase activity. We further demonstrate that Rlk can be phosphorylated and activated by Src kinases, leading to a decrease in its half-life. A specific tyrosine in the activation loop of Rlk, Y420, is required for phosphorylation and activation, as well as for decreased stability, but is not required for lipid RAFT association. Mutation of this tyrosine also prevents increased tyrosine phosphorylation of Rlk after stimulation of the T cell receptor, suggesting that Rlk is phosphorylated by Src family kinases in response to T cell receptor engagement. CONCLUSIONS: Like the other related Tec kinases, Rlk is associated with lipid RAFTs and can be phosphorylated and activated by Src family kinases, supporting a role for Rlk in signaling downstream of Src kinases in T cell activation.

Cell Line↗

Blockade of Wnt-5A/frizzled 5 signaling inhibits rheumatoid synoviocyte activation.

OBJECTIVE: It is not understood why cultured fibroblast-like synoviocytes (FLS) from patients with rheumatoid arthritis (RA) often display a persistently activated phenotype, despite removal from an inflammatory environment. Previously, we found that these FLS expressed high levels of both Wnt-5A and Frizzled 5 (Fz5), a receptor-ligand pair implicated in both limb bud and bone marrow stem cell development. The objective of the present experiments was to determine whether Wnt-5A/FzS signaling contributes to FLS activation. METHODS: Wnt-5A expression in FLS was inhibited by transfection with both antisense and dominant negative (dn) vectors. Fz5 signaling was blocked with an antibody to the extracellular domain of the receptor. The effects of these treatments on the expression of the proinflammatory cytokines interleukin-6 (IL-6) and IL-15 and on the expression of receptor activator of nuclear factor kappaB ligand (RANKL) were assessed by reverse transcriptase-polymerase chain reaction and immunoblotting. RESULTS: Both antisense Wnt-5A and dnWnt-5A vectors, but not empty vector, diminished IL-6 and IL-15 expression in RA FLS. Anti-Fz5 antibody exerted similar effects and also reduced RANKL expression. CONCLUSION: Wnt-5A/Fz5 signaling may contribute to the activated state of FLS in RA. Receptor antagonists of Fz5 should be considered for the treatment of refractory synovitis.

Antibodies, Blocking↗

Anaphylaxis due to hypersensitivity to Anisakis simplex.

Anaphylactic reactions are among the most frequent causes for requests for emergency medical care. It is possible that no clear precipitating factor can be found delaying the onset of adequate treatment and increasing the risk of new episodes of anaphylaxis in the future. Hypersensitivity to Anisakis simplex, an intestinal parasite found in fish, is an unusual cause of anaphylaxis but it should always be borne in mind in countries where a great deal of fish is consumed, especially if it is eaten raw or undercooked. 3 cases of anaphylaxis due to hypersensitivity to A. simplex are described.

Allergens↗

Transfer RNA modification status influences retroviral ribosomal frameshifting.

The possibility of whether tRNAs with and without a highly modified base in their anticodon loop may influence the level of retroviral ribosomal frameshifting was examined. Rabbit reticulocyte lysates were programmed with mRNA encoding UUU or AAC at the frameshift site and the corresponding Phe tRNA with or without the highly modified wyebutoxine (Y) base on the 3' side of its anticodon or Asn tRNA with or without the highly modified queuine (Q) base in the wobble position of its anticodon added. Phe and Asn tRNAs without the Y or Q base, respectively, stimulated the level of frameshifting, suggesting that the frameshift event is influenced by tRNA modification status. In addition, when AAU occurred immediately upstream of UUU as the penultimate frameshift site codon, addition of tRNAAsn without the Q base reduced the stimulatory effect of tRNAPhe without the Y base, whereas addition of tRNAAsn with the Q base did not alter the stimulatory effect. The addition of tRNAAsn without the Q base and tRNAPhe with the Y base inhibited frameshifting. The latter studies suggest an interplay between the tRNAs decoded at the penulimate frameshift and frameshift site codons that is also influenced by tRNA modification status. These data may be intrepreted as indicating that a hypomodified isoacceptor modulates frameshifting in an upward manner when utilized at the frameshift site codon, but modulates frameshifting in a downward manner when utilized at the penultimate frameshift site codon.

Animals↗

Occupational asthma caused by champignon flies.

BACKGROUND: Occupational bronchial asthma in mushroom (champignon) workers is unusual, although reports on it appeared in 1938 and 1951; we have not found any others since those dates. Here we report the case of a 52-year-old man who works as a champignon cultivator. He suffered rhinoconjunctivitis and asthma attacks whenever he entered the champignon culture caves. We studied flies as a possible antigen source. We collected these insects from the growing sites in order to identify them, and then prepare an extract; the samples turned out to be of two families of insects of the order Diptera, 98% from the Phoridae family (Brachycera suborder) and 2% from the Sciaridae (Nematocera suborder). METHODS: Skin prick tests, conjunctival provocation tests, serum specific IgE, specific IgE-binding fractions in immunoblotting, and monitoring of PEFR (at work and off work) were performed. RESULTS: IgE-mediated hypersensitivity to these flies was demonstrated by skin prick test, conjunctival provocation test, serum specific IgE, and IgE-binding fractions in immunoblotting. Monitoring of PEFR both at work and off work showed a clear relationship between symptoms, or fall in PEFR, and the workplace. CONCLUSIONS: We report the case of a patient suffering from asthma and rhinoconjunctivitis caused by hypersensitivity to fly proteins.

Agaricales↗

rlk/TXK encodes two forms of a novel cysteine string tyrosine kinase activated by Src family kinases.

Rlk/Txk is a member of the BTK/Tec family of tyrosine kinases and is primarily expressed in T lymphocytes. Unlike other members of this kinase family, Rlk lacks a pleckstrin homology (PH) domain near the amino terminus and instead contains a distinctive cysteine string motif. We demonstrate here that Rlk protein consists of two isoforms that arise by alternative initiation of translation from the same cDNA. The shorter, internally initiated protein species lacks the cysteine string motif and is located in the nucleus when expressed in the absence of the larger form. In contrast, the larger form is cytoplasmic. We show that the larger form is palmitoylated and that mutation of its cysteine string motif both abolishes palmitoylation and allows the protein to migrate to the nucleus. The cysteine string, therefore, is a critical determinant of both fatty acid modification and protein localization for the larger isoform of Rlk, suggesting that Rlk regulation is distinct from the other Btk family kinases. We further show that Rlk is phosphorylated and changes localization in response to T-cell-receptor (TCR) activation and, like the other Btk family kinases, can be phosphorylated and activated by Src family kinases. However, unlike the other Btk family members, Rlk is activated independently of the activity of phosphatidylinositol 3-kinase, consistent with its lack of a PH domain. Thus, Rlk has two distinct isoforms, each of which may have unique properties in signaling downstream from the TCR.

Animals↗

Migration of mitochondria to viral assembly sites in African swine fever virus-infected cells.

An examination by electron microscopy of the viral assembly sites in Vero cells infected with African swine fever virus showed the presence of large clusters of mitochondria located in their proximity. These clusters surround viral factories that contain assembling particles but not factories where only precursor membranes are seen. Immunofluorescence microscopy revealed that these accumulations of mitochondria are originated by a massive migration of the organelle to the virus assembly sites. Virus infection also promoted the induction of the mitochondrial stress-responsive proteins p74 and cpn 60 together with a dramatic shift in the ultrastructural morphology of the mitochondria toward that characteristic of actively respiring organelles. The clustering of mitochondria around the viral factory was blocked in the presence of the microtubule-disassembling drug nocodazole, indicating that these filaments are implicated in the transport of the mitochondria to the virus assembly sites. The results presented are consistent with a role for the mitochondria in supplying the energy that the virus morphogenetic processes may require and make of the African swine fever virus-infected cell a paradigm to investigate the mechanisms involved in the sorting of mitochondria within the cell.

African Swine Fever Virus↗

A characteristic bent conformation of RNA pseudoknots promotes -1 frameshifting during translation of retroviral RNA.

The structures of four different RNA pseudoknots that provide one of the signals required for ribosomal frameshifting in mouse mammary tumor virus have been determined by NMR. The RNA pseudoknots have similar sequences and assume similar secondary structures, but show significantly different frameshifting efficiencies. The three-dimensional structures of one frameshifting and one non-frameshifting RNA pseudoknot had been determined previously by our group. Here we determine the structures of two new RNA pseudoknots, and relate the structures of all four pseudoknots to their frameshifting abilities. The two efficient frameshifting pseudoknots adopt characteristic bent conformations with stem 1 bending towards the major groove of stem 2. In contrast, the two poor frameshifting pseudoknots have structures very different from each other and from the efficient frameshifters. One has linear, coaxially stacked stems, the other has stems twisted and bent, but in the opposite direction to the efficient frameshifters. Changes in loop size that favor bending (shorter loops) increase frameshifting efficiency; longer loops that allow linear arrangement of the stems decrease frameshifting. Frameshifting pseudoknots in feline immunodeficiency virus and simian retrovirus have different loop sequences, but the sequences at their stem junctions imply the same bent conformation as in the mouse mammary tumor viral RNA. The requirement for a precise pseudoknot conformation for efficient frameshifting strongly implies that a specific interaction occurs between the viral RNA pseudoknot and the host protein-synthesizing machinery.

Base Sequence↗

Structural and functional studies of retroviral RNA pseudoknots involved in ribosomal frameshifting: nucleotides at the junction of the two stems are important for efficient ribosomal frameshifting.

Ribosomal frameshifting, a translational mechanism used during retroviral replication, involves a directed change in reading frame at a specific site at a defined frequency. Such programmed frameshifting at the mouse mammary tumor virus (MMTV) gag-pro shift site requires two mRNA signals: a heptanucleotide shifty sequence and a pseudoknot structure positioned downstream. Using in vitro translation assays and enzymatic and chemical probes for RNA structure, we have defined features of the pseudoknot that promote efficient frameshifting. Heterologous RNA structures, e.g. a hairpin, a tRNA or a synthetic pseudoknot, substituted downstream of the shifty site fail to promote frameshifting, suggesting that specific features of the MMTV pseudoknot are important for function. Site-directed mutations of the MMTV pseudoknot indicate that the pseudoknot junction, including an unpaired adenine nucleotide between the two stems, provides a specific structural determinant for efficient frameshifting. Pseudoknots derived from other retroviruses (i.e. the feline immunodeficiency virus and the simian retrovirus type 1) also promote frameshifting at the MMTV gag-pro shift site, dependent on the same structure at the junction of the two stems.

Animals↗

Mammalian adaptation to extrauterine environment: mitochondrial functional impairment caused by prematurity.

In this paper we report that, compared with term rat neonates, both mitochondrial content and function are diminished in liver of preterm neonates (delivered 24 h before full term) compromising cellular energy provision in the postnatal period. In addition, there is a parallel reduction in the content of mRNAs encoding mitochondrial proteins in preterm rats. Also, efficient oxidative phosphorylation is not attained in these pups until 3 h after birth. Although isolated liver mitochondria from preterm neonates show a two-fold increase in F1-ATPase beta-subunit and cytochrome c oxidase activity 1 h after birth, the abnormal coupling efficiency between respiration and oxidative phosphorylation (ADP/O ratio) is due to maintenance of high H(+)-leakage values in the inner mitochondrial membrane. Postnatal reduction of the H+ leak occurs concomitantly with an increase in intra-mitochondrial adenine nucleotide concentration. Accumulation of adenine nucleotides in preterm and term liver mitochondria parallels the postnatal increase in total liver adenine nucleotides. Delayed postnatal induction of adenine biosynthesis most likely accounts for the lower adenine nucleotide pool in the liver of preterm neonates. The delayed postnatal accumulation of adenine nucleotides in mitochondria is thus responsible for the impairment in oxidative phosphorylation displayed by organelles of the preterm liver.

Adaptation, Physiological↗

Identification of the major components of the high molecular weight crystallins from old human lenses.

High molecular weight aggregates from the water soluble portion of four lenses from older donors were examined by mass spectrometric techniques that permitted unambiguous identification of the principal components as alpha A-, alpha B- and gamma s-crystallins. Post-translational modifications that were located and identified included deamidations in alpha A and gamma s and an intramolecular disulfide bond in alpha A-crystallin. The C-terminus of alpha A was not found, supporting previous suggestions that this portion of the protein may be important in preventing aggregation.

Aged↗

The effect of omental wrapping on nerve graft regeneration.

We have performed 1 cm long nerve grafts in the sciatic nerves of 60 Wistar rats. In 30 cases (n = 30) the grafts were wrapped in omentum. We have studied the nerve regeneration accomplished after 30 and 90 postoperative days from the histological and neurophysiological points of views. Survival of the omentum has been excellent. The histological assessment showed a higher blood vessel population as well as a higher count of axons in the cases where the nerve graft was wrapped in omentum. Areas with neural fibrosis, which are a sign of poor vascularisation, were smaller in the cases with omentum. The amplitude of the contraction in the gastrocnemius muscle was higher in the cases where omentum was used. The endothelial cells of the omentum synthesis fibroblast growth factors (acid and basic) which, due to their angiogenic and neuronotrophic properties, may be the cause of the beneficial effects of omentum on peripheral nerve regeneration.

Animals↗

An RNA pseudoknot and an optimal heptameric shift site are required for highly efficient ribosomal frameshifting on a retroviral messenger RNA.

Synthesis of the pol gene products of most retroviruses requires ribosomes to shift frame once or twice in the -1 direction while translating gag-pol mRNA. The viral signals for frameshifting include a heptanucleotide sequence on which the shift occurs and higher-order RNA structure just downstream of the shift site. We have made site-directed mutations in two stems (S1 and S2) of a putative RNA pseudoknot that begins 7 nucleotides 3' of the previously identified shift site (A AAA AAC) in the gag-pro region of mouse mammary tumor virus (MMTV) RNA. The mutants confirm the predicted structure, show that loss of either S1 or S2 impairs frameshifting, and exclude alternative RNA structures as significant for frameshifting. The importance of the MMTV pseudoknot has been further demonstrated by showing that shift sites from two other retroviruses function more efficiently in the position of the MMTV site than in their native contexts. However, the MMTV pseudoknot cannot promote detectable frameshifting in the absence of a recognizable upstream shift site. In addition, the species of tRNA that reads the second codon in the shift site appears to be a critical determinant, since changing the 7th nucleotide in the MMTV gag-pro shift site from C to A, U, or G severely impairs frameshifting.

Base Sequence↗