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Biomedical subjects

T A Williams

Publications and source records attributed to T A Williams.

At least 19 recordsLinked to original sources
PubMed2006

Data linkage enables evaluation of long-term survival after intensive care.

Outcomes of intensive care are important to the patient and for assessment of benefit. Short-term outcomes after critical illness are well described, but less is known about long-term outcomes. This study describes the use of data linkage, combining intensive care unit (ICU) clinical data with administrative morbidity and mortality data, to assess long-term outcomes after treatment in ICU. The hospital-based cohort study was conducted in a 22-bed general ICU in a metropolitan teaching hospital. All patient admissions admitted to ICU from 1 January 1987 to 31 December 2002 were included. The prospective ICU clinical database with patient demographics, ICU diagnoses, severity of illness, daily assessment of organ failures and common daily treatments used was linked using probabilistic methods to the state-wide hospital morbidity and mortality databases to describe long-term survival. There were 26,019 ICU admissions (22,980 patients) with 25,972 records (99.8%) linked to a hospitalization event that included the index ICU admission. Unadjusted survival was 84.7% at 1 year decreasing progressively to 50.7% at 15 years. Age, type of admission, severity of illness (measured by Acute Physiologic and Chronic Health Evaluation (APACHE) II and the presence of organ failure), ICU length of stay, comorbidity (Chronic Health Evaluation and Charlson comorbidity index) and ICU admission diagnosis, were all associated with survival at 1, 3, 5, 10, and 15 year follow-up (P<0.001 at all time points). Linkage of clinical and administrative data provides a feasible method for ascertaining long-term survival after critical illness. Age, admission severity of illness, diagnosis and comorbidity influenced long-term unadjusted survival.

APACHE↗
PubMed2004

Coexistence of different phenotypes in a family with glucocorticoid-remediable aldosteronism.

In glucocorticoid-remediable aldosteronism (GRA), there is a large interfamily variation of phenotype. We report three subjects with GRA in a single family (parents, two brothers and two sisters), of whom only one (proband) displayed classical features of the mineralocorticoid excess. The proband was a man found to be hypertensive and hypokalaemic at the age of 24 years. Plasma renin activity was suppressed and plasma aldosterone was repeatedly elevated. Blood pressure and aldosterone levels normalized within 5 days of dexamethasone therapy. The presence of a chimaeric CYP11B1/CYP11B2 gene was demonstrated by long-PCR and Southern blotting (crossover site at the end of intron 3) in the proband, in the younger sister (sibling 1) and in the father. In these patients, sequencing of the chimaeric portion of CYP11B1 did not reveal any mutation, while sequencing of the chimaeric portion of CYP11B2 showed a V386A polymorphism in exon 7, known to cause only a minimal impairment of enzymatic activity. Sibling 1 was normotensive, normokalaemic and had normal PRA and aldosterone. The father had normal blood pressure and potassium, low-normal PRA and normal aldosterone. All three subjects had elevated levels of urinary 18-hydroxycortisol and 18-oxocortisol. Baseline 11-deoxycorticosterone (DOC), corticosterone (B) and aldosterone were high in the proband and normal in the father and sibling 1; 11-deoxycortisol (S) and cortisol (F) were normal. ACTH induced a normal increase of B, DOC, S and F, and an excessive aldosterone increase in all three patients. Abnormalities in the chimaeric portions of CYB11B1 or CYP11B2 genes did not account for the phenotypic disparity of the different members in a single GRA family. Altered regulation of the chimaeric gene may be responsible for differences in its activity.

Adult↗
PubMed2004

Bradykinin B2 receptor gene C-58T polymorphism and insulin resistance. A study on obese patients.

The bradykinin B2 receptor (B2R) gene is a candidate in the pathogenesis of insulin resistance, which often clusters with other abnormalities in metabolic syndrome. We investigated the distribution of the C-58T B2R gene polymorphism within a population of overweight/obese patients (BMI > or = 25 kg/m2) potentially characterised by different levels of insulin resistance. Patients with type 2 diabetes, dyslipidemia and hypertension were excluded in order to distinguish the effect of obesity on insulin sensitivity from that of confounding factors. Ninety-two unrelated adults (41 men and 51 women, aged 33.7 +/- 11.6 years) were recruited by random sampling from a general population evaluated for cardiovascular risk stratification. Measurements included BMI, waist circumference, body composition, blood pressure, serum leptin, and lipid profile. Insulin sensitivity was calculated according to the homeostasis model assessment (HOMA) method. C-58T genotypes--CC (n = 20), CT (n = 47) and TT (n = 25)--were determined by restriction fragment-length polymorphism PCR. Patients subdivided on the basis of C-58T polymorphism, showed no difference in any of the parameters examined, including HOMA index values, after adjustment for age, sex, BMI and waist circumference. The results indicate that the C-58T B2R gene polymorphism is not associated with different levels of insulin resistance within a population of obese patients.

Adult↗
PubMed2003

Earlier flowering between 1962 and 2002 in agricultural varieties of white clover.

Date of first flowering was recorded for 40 years in the same agricultural varieties of white clover (Trifolium repens L.). Standard procedures, experimental conditions and germplasm allowed the unmediated effects of climatic trends to be observed. Flowering dates of the same varieties at the same site have become significantly earlier over this period, with an advance of first flowering of approximately 7.5 days per decade since 1978. Annual maximum, minimum and soil temperatures at the site increased during the 40-year period. First flowering dates (FFDs) were significantly negatively correlated with minimum and maximum temperatures during February and March and soil temperatures between January and April. Maximum, minimum and soil temperatures increased between 1962 and 2002 for these months. No significant correlation was seen between FFD and soil temperatures at other times of the year including the period of vernalisation during winter. The occurrence of ground frost in February was significantly correlated with later flowering. Total annual rainfall and the number of wet days in a year increased between 1962 and 2002. However, only rainfall in February had a significant impact on FFD.

Agriculture↗
PubMed2001

Developmental modularity and the evolutionary diversification of arthropod limbs.

Segmentation is one of the most salient characteristics of arthropods, and differentiation of segments along the body axis is the basis of arthropod diversification. This article evaluates whether the evolution of segmentation involves the differentiation of already independent units, i.e., do segments evolve as modules? Because arthropod segmental differentiation is commonly equated with differential character of appendages, we analyze appendages by comparing similarities and differences in their development. The comparison of arthropod limbs, even between species, is a comparison of serially repeated structures. Arthropod limbs are not only reiterated along the body axis, but limbs themselves can be viewed as being composed of reiterated parts. The interpretation of such reiterated structures from an evolutionary viewpoint is far from obvious. One common view is that serial repetition is evidence of a modular organization, i.e., repeated structures with a common fundamental identity that develop semi-autonomously and are free to diversify independently. In this article, we evaluate arthropod limbs from a developmental perspective and ask: are all arthropod limbs patterned using a similar set of mechanisms which would reflect that they all share a generic coordinate patterning system? Using Drosophila as a basis for comparison, we find that appendage primordia, positioned along the body using segmental patterning coordinates, do indeed have elements of common identity. However, we do not find evidence of a single coordinate system shared either between limbs or among limb branches. Data concerning the other diagnostic of developmental modularity--semi-autonomy of development--are not currently available for sufficient taxa. Nonetheless, some data comparing patterns of morphogenesis provide evidence that limbs cannot always be temporally or spatially decoupled from the development of their neighbors, suggesting that segment modularity is a derived character.

Adaptation, Physiological↗
PubMed2001

Osteopontin is an autocrine mediator of hepatocyte growth factor-induced invasive growth.

In epithelial cells, hepatocyte growth factor (HGF) activates a genetic program involving cell-cell dissociation ("scattering"), growth and invasiveness. The full program is not elicited by other growth factors like epidermal growth factor, and is aberrantly activated during cancer progression to the invasive-metastatic phenotype. To identify genes involved in the onset of invasive growth, we explored by cDNA microarrays the in vitro transcriptional response to HGF of mouse embryo liver cells. We identified osteopontin (OPN), a secreted matrix protein, as a major HGF transcriptional target. The wave of OPN induction is maximal at 6 h, in concomitance with the initiation of scattering, and is specific, because no other matrix protein among those explored by the microarray is affected. Interestingly, HGF, but not epidermal growth factor, promotes cell adhesion to OPN via the CD44 receptor. Scattering is significantly impaired by antibodies against OPN and CD44; conversely, constitutive OPN overexpression dramatically increases the motile and invasive responses to HGF, leading to disruption of the ordered morphogenetic program triggered by this ligand.

Animals↗
PubMed2000

Functional conservation of the active sites of human and Drosophila angiotensin I-converting enzyme.

Human somatic angiotensin I-converting enzyme (sACE) has two active sites present in two homologous protein domains, resulting from a tandem gene duplication. It has been proposed that the N- and C-terminal active sites can have specific in vivo roles. In Drosophila melanogaster, Ance and Acercode for two ACE-like single-domain proteins, also predicted to have distinct physiological roles. We have investigated the relationship of Ance and Acer to the N- and C-domains of human sACE by genomic sequence analysis and by using domain-selective inhibitors, including RXP 407, a selective inhibitor of the human N-domain. These phosphinic peptides were potent inhibitors of Acer, but not of Ance. We conclude that the active sites of the N-domain and of Acer share structural features that permit the binding of the unusual RXP407 inhibitor and the hydrolysis of a broader range of peptide structures. In comparison, Ance, like the human C-domain of ACE, displays greater inhibitor selectivity. From the analysis of the published sequence of the Adh region of Drosophila chromosome 2, which carries Ance, Acer, and four additional ACE-like genes, we also suggest that this functional conservation is reflected in an ancestral gene structure identifiable in both protostome and deuterostome lineages and that the duplication seen in vertebrate genomes predates the divergence of these lineages. The conservation of ACE enzymes with distinct active sites in the evolution of both vertebrate and invertebrate species provides further evidence that these two kinds of active sites have different physiological functions.

Acetylation↗
PubMed2000

Sustained recruitment of phospholipase C-gamma to Gab1 is required for HGF-induced branching tubulogenesis.

A distinctive property of Hepatocyte Growth Factor (HGF) is its ability to induce differentiation of tubular structures from epithelial and endothelial cells (branching tubulogenesis). The HGF receptor directly activates PI3 kinase, Ras and STAT signalling pathways and phosphorylates the adaptator GRB2 Associated Binder-1 (Gab1). Gab1 is also phosphorylated in response to Epidermal Growth Factor (EGF) but is unable to induce tubule formation. Comparison of 32P-peptide maps of Gab1 from EGF- versus HGF-treated cells, demonstrates that the same sites are phosphorylated in vivo. However, while both EGF and HGF induce rapid tyrosine phosphorylation of Gab1 with a peak at 15 min, the phosphorylation persists for over 1 h, only in response to HGF. Nine tyrosines are phosphorylated by both receptors. Three of them (Y307, Y373, Y407) bind phospholipase C-gamma (PLC-gamma). Interestingly, the overexpression of a Gab1 mutant unable to bind PLC-gamma (Gab1 Y307/373/407F) did not alter HGF-stimulated cell scattering, only partially reduced the growth stimulation but completely abolished HGF-mediated tubulogenesis. It is concluded that sustained recruitment of PLCgamma to Gab1 plays an important role in branching tubulogenesis.

Animals↗
PubMed2000

Different point mutations in the met oncogene elicit distinct biological properties.

The MET proto-oncogene, encoding the tyrosine kinase receptor for HGF, controls genetic programs leading to cell growth, invasiveness, and protection from apoptosis. Recently, MET mutations have been identified in hereditary and sporadic forms of papillary renal carcinoma (PRC). Introduction of different naturally occurring mutations into the MET cDNA results in the acquisition of distinct biochemical and biological properties of transfected cells. Some mutations result in a high increase in tyrosine kinase activity and confer transforming ability in focus forming assays. These mutants hyperactivate the Ras signaling pathway. Other mutations are devoid of transforming potential but are effective in inducing protection from apoptosis and sustaining anchorage-independent growth. These Met(PRC) receptors interact more efficiently with the intracellular transducer Pi3Kinase. The reported results show that MET(PRC) mutations can be responsible for malignant transformation through different mechanisms, either by increasing the growth ability of cells or by protecting cells from apoptosis and allowing accumulation of other genetic lesions.-Giordano, S., Maffe, A., Williams, T. A., Artigiani, S., Gual, P., Bardelli, A., Basilico, C., Michieli, P., Comoglio, P. M. Different point mutations in the met oncogene elicit distinct biological properties.

Apoptosis↗
PubMed1999

A peptide representing the carboxyl-terminal tail of the met receptor inhibits kinase activity and invasive growth.

Interaction of the hepatocyte growth factor (HGF) with its receptor, the Met tyrosine kinase, results in invasive growth, a genetic program essential to embryonic development and implicated in tumor metastasis. Met-mediated invasive growth requires autophosphorylation of the receptor on tyrosines located in the kinase activation loop (Tyr(1234)-Tyr(1235)) and in the carboxyl-terminal tail (Tyr(1349)-Tyr(1356)). We report that peptides derived from the Met receptor tail, but not from the activation loop, bind the receptor and inhibit the kinase activity in vitro. Cell delivery of the tail receptor peptide impairs HGF-dependent Met phosphorylation and downstream signaling. In normal and transformed epithelial cells, the tail receptor peptide inhibits HGF-mediated invasive growth, as measured by cell migration, invasiveness, and branched morphogenesis. The Met tail peptide inhibits the closely related Ron receptor but does not significantly affect the epidermal growth factor, platelet-derived growth factor, or vascular endothelial growth factor receptor activities. These experiments show that carboxyl-terminal sequences impair the catalytic properties of the Met receptor, thus suggesting that in the resting state the nonphosphorylated tail acts as an intramolecular modulator. Furthermore, they provide a strategy to selectively target the MET proto-oncogene by using small, cell-permeable, peptide derivatives.

Adaptor Proteins, Signal Transducing↗
PubMed1999

MET(PRC) mutations in the Ron receptor result in upregulation of tyrosine kinase activity and acquisition of oncogenic potential.

Ron and Met are structurally related receptor tyrosine kinases that elicit a complex biological response leading to invasive growth. Naturally occurring point mutations activate the Met kinase in papillary renal carcinomas (MET(PRC) mutations). By site-directed mutagenesis, we generated homologous amino acid substitutions in the Ron kinase domain and analyzed the biochemical and biological properties of the mutant receptors. Among the mutations studied, D(1232)H and M(1254)T displayed transforming activity in NIH3T3 cells, inducing focus formation and anchorage-independent growth. The D(1232)H and M(1254)T substitutions resulted in increased Ron autophosphorylation both in vivo and in vitro and constitutive binding to intracellular signal transducers. Both mutations yielded a dramatic increase in catalytic efficiency, indicating a direct correlation between kinase activity and oncogenic potential. Molecular modeling of the Ron D(1232)H mutation suggests that this single amino acid substitution favors the transition of the kinase from the inactive to the active state. These data demonstrate that point mutations can confer transforming activity to the Ron receptor and show that RON is a potential oncogene.

3T3 Cells↗
PubMed1999

Hydrolysis by somatic angiotensin-I converting enzyme of basic dipeptides from a cholecystokinin/gastrin and a LH-RH peptide extended at the C-terminus with gly-Arg/Lys-arg, but not from diarginyl insulin.

Endoproteolytic cleavage of protein prohormones often generates intermediates extended at the C-terminus by Arg-Arg or Lys-Arg, the removal of which by a carboxypeptidase (CPE) is normally an important step in the maturation of many peptide hormones. Recent studies in mice that lack CP activity indicate the existence of alternative tissue or plasma enzymes capable of removing C-terminal basic residues from prohormone intermediates. Using inhibitors of angiotensin I-converting enzyme (ACE) and CP, we show that both these enzymes in mouse serum can remove the basic amino acids from the C-terminus of CCK5-GRR and LH-RH-GKR, but only CP is responsible for converting diarginyl insulin to insulin. ACE activity removes C-terminal dipeptides to generate the Gly-extended peptides, whereas CP hydrolysis gives rise to CCK5-GR and LH-RH-GK, both of which are susceptible to the dipeptidyl carboxypeptidase activity of ACE. Somatic ACE has two similar protein domains (the N-domain and the C-domain), each with an active site that can display different substrate specificities. CCK5-GRR is a high-affinity substrate for both the N-domain and C-domain active sites of human sACE (Km of 9.4 microm and 9.0 microm, respectively) with the N-domain showing greater efficiency (kcat : Km ratio of 2.6 in favour of the N-domain). We conclude that somatic forms of ACE should be considered as alternatives to CPs for the removal of basic residues from some Arg/Lys-extended peptides.

Animals↗
PubMed1999

Rapid gas chromatographic analysis of drugs of forensic interest.

High-speed gas chromatographic (GC) screening for drugs of forensic relevance is performed using a commercial Flash GC instrument in which the chromatographic column is resistively heated at rates of up to 30 degrees C/s. Temperature programming conditions are varied in an experiment designed to evaluate trade-offs between resolution and analysis time for a mixture of 19 drugs of abuse. All 19 components can be separated with excellent resolution in 90 s. Specific analytes can be analyzed even faster; for example, amphetamine analysis is completed in less than 20 s. Case studies of confiscated street drugs containing amphetamine, cocaine, and heroin are analyzed to evaluate the retention time repeatability. Ten replicate injections over a 2-day period for 3 different drug samples achieved retention time relative standard deviations in the range of 0.48 to 0.81%.

Amphetamine↗
PubMed1998

The Drosophila melanogaster-related angiotensin-I-converting enzymes Acer and Ance--distinct enzymic characteristics and alternative expression during pupal development.

Drosophila melanogaster express two distinct angiotensin-I-converting enzymes (ACEs) called Ance and Acer, which display a high level of primary structure similarity. We have expressed Acer in the yeast Pichia pastoris and purified the recombinant enzyme with a view to developing biochemical tools to distinguish between Acer and Ance. Purified Acer and Ance expressed in yeast were used to raise anti-Acer Ig and anti-Ance Ig that specifically cross-reacted with the respective enzyme on immunoblotting, but did not act as specific inhibitors. Acer cleaves the C-terminal dipeptides from benzoylglycyl-histidyl-leucine and [Leu5]enkephalin, and Acer and Ance are both able to act as endopeptidases, releasing the C-terminal dipeptideamide from [Leu5]enkephalinamide. However, Acer hydrolyses this substrate at a slightly faster rate than [Leu5]enkephalin, whereas Ance hydrolyses the peptide with a free C-terminus with a kcat 15-fold higher than [Leu5]enkephalinamide. In addition, Acer did not cleave angiotensin I. In contrast, Ance hydrolysed 25% of this substrate at an 8-fold lower enzyme concentration. Furthermore, Acer did not hydrolyse the synthetic substrates Phe-Ser-Pro-Arg-Leu-Gly-Arg-Arg and Phe-Ser-Pro-Arg-Leu-Gly-Lys-Arg, two partially processed putative locustamyotropin precursors, under conditions where Ance produced 82% substrate hydrolysis. Acer was inhibited by captopril, trandolaprilat and enalaprilat, with apparent Ki values in the nanomolar range, whereas lisinopril and fosinoprilat were less potent. We show that the two Drosophila ACEs are alternatively expressed in stages P1 (white puparium)-P15 (eclosion) of pupal development; Ance is expressed predominantly during stages P4-P7, whereas the ACE activity expressed during stages P9-P12 is mainly due to Acer suggesting different roles for the two enzymes during pupal development.

Amino Acid Sequence↗
PubMed1998

A novel peptide-processing activity of insect peptidyl-dipeptidase A (angiotensin I-converting enzyme): the hydrolysis of lysyl-arginine and arginyl-arginine from the C-terminus of an insect prohormone peptide.

Insect peptidyl-dipeptidase A [angiotensin I-converting enzyme (ACE)] is a soluble single-domain peptidyl-dipeptidase that has many properties in common with the C-domain of mammalian somatic ACE and with the single-domain mammalian germinal ACE. Mammalian somatic ACE is important in blood homoeostasis, but the role of ACE in insects is not known. Immunocytochemistry has been used to localize ACE in the neuroendocrine system of the locust, Locusta migratoria. Staining was observed in five groups of neurosecretory cells in the brain and suboesophageal ganglion, in the nervi corpori cardiaci, the storage part of the corpora cardiaca and in the nervi corpori allati. In three groups of neurosecretory cells, ACE co-localized with locustamyotropins, suggesting a possible role for the enzyme in the metabolism of these neuropeptides. We demonstrate in vitro a novel activity of ACE that removes pairs of basic amino acid residues from a locustamyotropin peptide extended at the C-terminus with either Gly-Lys-Arg or Gly-Arg-Arg, corresponding to a consensus recognition sequence for endoproteolysis of prohormone proteins by prohormone convertases. The low Km and high kcat values (Km 7.3 and 5.0 microM, kcat 226 and 207 s-1 for the hydrolysis of Phe-Ser-Pro-Arg-Leu-Gly-Lys-Arg and Phe-Ser-Pro-Arg-Leu-Gly-Arg-Arg, respectively) obtained for the hydrolysis of these two peptides by insect ACE means that these peptides, along with mammalian bradykinin, are the most favoured in vitro ACE substrates so far identified. The discovery of this in vitro prohormone-processing activity of insect ACE provides a possible explanation for the intracellular co-localization of the enzyme with locustamyotropin peptides, and provides evidence for a new role for ACE in the biosynthesis of peptide hormones and transmitters.

Animals↗
PubMed1998

Distalless expression in crustaceans and the patterning of branched limbs.

In Drosophila, Distalless (Dll) is critical in establishing the proximal/distal axis of the leg. Lack of proper Dll expression causes distal limb structures to be truncated or lost. Dll expression was examined through the course of development in the limbs of two crustaceans, Triops and Nebalia. Because the limbs of these two species are branched, they provide a comparison to the uniramous (unbranched) leg of Drosophila. In Triops and Nebalia, development of limb branches is not tightly coupled with Dll expression: in some cases, branches can arise prior to Dll expression and in others, certain branches never express Dll. These data suggest that, while Dll may indeed initiate overall limb outgrowth, limb branches are unlikely to be patterned by a simple iteration of the mechanism patterning the unbranched leg of Drosophila.

Animals↗