How should children with speech sound disorders be classified? A review and critical evaluation of current classification systems.

BACKGROUND: Children with speech sound disorders (SSD) form a heterogeneous group who differ in terms of the severity of their condition, underlying cause, speech errors, involvement of other aspects of the linguistic system and treatment response. To date there is no universal and agreed-upon classification system. Instead, a number of theoretically differing classification systems have been proposed based on either an aetiological (medical) approach, a descriptive-linguistic approach or a processing approach. AIMS: To describe and review the supporting evidence, and to provide a critical evaluation of the current childhood SSD classification systems. METHODS & PROCEDURES: Descriptions of the major specific approaches to classification are reviewed and research papers supporting the reliability and validity of the systems are evaluated. MAIN CONTRIBUTION: Three specific paediatric SSD classification systems; the aetiologic-based Speech Disorders Classification System, the descriptive-linguistic Differential Diagnosis system, and the processing-based Psycholinguistic Framework are identified as potentially useful in classifying children with SSD into homogeneous subgroups. The Differential Diagnosis system has a growing body of empirical support from clinical population studies, across language error pattern studies and treatment efficacy studies. The Speech Disorders Classification System is currently a research tool with eight proposed subgroups. The Psycholinguistic Framework is a potential bridge to linking cause and surface level speech errors. CONCLUSIONS & IMPLICATIONS: There is a need for a universally agreed-upon classification system that is useful to clinicians and researchers. The resulting classification system needs to be robust, reliable and valid. A universal classification system would allow for improved tailoring of treatments to subgroups of SSD which may, in turn, lead to improved treatment efficacy.

Biomarkers of endocrine disruption: cluster analysis of effects of plasticisers on Phase 1 and Phase 2 metabolism of steroids.

Although some endocrine disruptors (EDs) act at steroid receptors, it is now apparent that compounds may have ED potential if they alter steroid synthesis or metabolism, particularly if they affect Phase 1 or Phase 2 pathways. In the ENDOMET project (EU-funded 5th Framework programme), 23 different assays were used on a wide range of EDs. Cluster analysis of the matrix results enabled identification of four integrated test systems that can be used to pinpoint compounds that are able to alter steroid metabolism or function. Critical pathways were shown to include oestrogen synthesis and sulphonation, synthesis of sulphate/PAPS and thyroid hormone regulation so that the activity profiles of some Phase 1 and Phase 2 reactions can be used as biomarkers for detection of compounds with ED potential.

Detection of endocrine disruptors - from simple assays to whole genome scanning.

Endocrine disruptors frequently bear little structural relationship to the hormone whose actions they disrupt. Consequently, the threat of an uninvestigated chemical cannot easily be assessed. Here three different approaches to assessment are discussed. The first presumes an endocrine-disrupting property, following which a cell model capable of responding to such a hormone is used. Although simple and cheap, it provides limited data. A second approach involves multiple assays to detect multiple hormones. Increasing the amount of data increased the difficulty in assessing the significance of results. To meet this problem, cluster analysis based on a simple mathematical matrix was adopted. The matrix was used to determine (i) a limited number of assays to identify a maximum number of endocrine disruptors and (ii) the chemicals with the most wide-ranging effects. A third approach was a whole genome expression analysis based on expression of mRNAs in human TE671 medulloblastoma cells. Expression of individual mRNAs was assessed using the Affymetrix GeneChip(®) Human Genome U133 Plus 2.0 chip. The significance of differential expressed genes was assessed based on gene ontology and pathways analyses using DAVID and GenMaPP programs. The results illustrated the very wide-ranging effects of these chemicals across the genome.

Alexander Ure MD, FRCS (1808-1866), and the beginning of drug metabolism studies.

Although many fields of endeavour emerge owing to the coalescence of the work and observations of numerous individuals, there is usually one seminal event that unites and acts as a catalyst to stimulate and advance the process. Such was the case with Alexander Ure. Up to this point it had been speculated that chemicals taken into the body may undergo bio-transformation, akin to the digestion of nutrients, but no unequivocal and quantitative experiments had been performed before those of Ure. Following his observations the subject began to flourish; to him may be attributed the beginnings of xenobiochemistry and the field now known as drug metabolism.

Reactive oxygen species and oxidative DNA damage mediate the cytotoxicity of tungsten-nickel-cobalt alloys in vitro.

Tungsten alloys (WA) have been introduced in an attempt to find safer alternatives to depleted uranium and lead munitions. However, it is known that at least one alloy, 91% tungsten-6% nickel-3% cobalt (WNC-91-6-3), causes rhabdomyosarcomas when fragments are implanted in rat muscle. This raises concerns that shrapnel, if not surgically removable, may result in similar tumours in humans. There is therefore a clear need to develop rapid and robust in vitro methods to characterise the toxicity of different WAs in order to identify those that are most likely to be harmful to human health and to guide development of new materials in the future. In the current study we have developed a rapid visual in vitro assay to detect toxicity mediated by individual WA particles in cultured L6-C11 rat muscle cells. Using a variety of techniques (histology, comet assay, caspase-3 activity, oxidation of 2'7'-dichlorofluorescin to measure the production of reactive oxygen species and whole-genome microarrays) we show that, in agreement with the in vivo rat carcinogenicity studies, WNC-91-6-3 was the most toxic of the alloys tested. On dissolution, it produces large amounts of reactive oxygen species, causes significant amounts of DNA damage, inhibits caspase-3, triggers a severe hypoxic response and kills the cells in the immediate vicinity of the alloy particles within 24h. By combining these in vitro data we offer a mechanistic explanation of the effect of this alloy in vivo and show that in vitro tests are a viable alternative for assessing new alloys in the future.

Evergreen coniferous forests of the pacific northwest.

The massive, evergreen coniferous forests in the Pacific Northwest are unique among temperate forest regions of the world. The region's forests escaped decimation during Pleistocene glaciation; they are now dominated by a few broadly distributed and well-adapted conifers that grow to large size and great age. Large trees with evergreen needle- or scale-like leaves have distinct advantages under the current climatic regime. Photosynthesis and nutrient uptake and storage are possible during the relatively warm, wet fall and winter months. High evaporative demand during the warm, dry summer reduces photosynthesis. Deciduous hardwoods are repeatedly at a disadvantage in competing with conifers in the regional climate. Their photosynthesis is predominantly limited to the growing season when evaporative demand is high and water is often limiting. Most nutrients needed are also less available at this time. The large size attained by conifers provides a buffer against environmental stress (especially for nutrients and moisture). The long duration between destructive fires and storms permits conifers to outgrow hardwoods with more limited stature and life spans.

Plant moisture stress: evaluation by pressure bomb.

The recently developed technique for determining the water stress of a plant by measuring the pressure necessary to force water back to the cut surface of a severed twig is adaptable to both field and laboratory experiments. We have designed and operated an efficient portable system weighing less than 18 kilograms. Sampling variation within and among Douglas fir trees varies from less than +/- I atmosphere under low stress conditions to +/- 10 atmospheres under high stress conditions. In the measurement of plants of comparable height and similar exposure, the variation is reduced to a minimum. Values in internal water stress of Douglas fir vary from 3 to more than 40 atmospheres. Both duration and magnitude of stress are important ecologically. Pressure-bomb measurements are used to demonstrate a relation between plant distribution and internal water stress.

Sulfotransferase inhibition: potential impact of diet and environmental chemicals on steroid metabolism and drug detoxification.

The cytosolic sulfotransferase enzymes (SULT isoforms) utilise PAPS (3'-phosphoadenosine-5'-phosphosulfate) as co-factor to transfer sulfonate groups onto a wide range of substrates. SULT1A3 has catecholamines such as dopamine as substrates while SULT 1E1 sulfonates oestrogens. SULT 1A1 sulfonates phenols and also oestrogens at a higher K(m) than SULT 1E1. SULT 2A1 mainly sulfonates DHEA and some steroids, with hydroxy derivatives of polycyclic aromatic hydrocarbons. Studies on these isoforms with a range of environmental chemicals and dietary components have shown that SULT 1A1 is significantly inhibited by flavonoids; all flavones and flavonols with a 3',4'-dihydroxy motif had an IC(50) of < 100 nm against 3 microM 4-nitrophenol as the standard substrate. SULTs 1A3 and 2A1 were less strongly inhibited by flavonoids or isoflavonoids although tricin (3',5'-dimethoxy-4',5,7-trihydroxyflavone is a competitive inhibitor of SULT 1E1 with an inhibition constant of approximately 1 nM. Fruit and vegetable cytosols also inhibit SULT isoforms, as do long-chain alkylphenols and chlorinated phenols. Phthalates (used as plasticisers) inhibited SULTs 1E1 and 2A1. As these environmental contaminants and dietary components all act at the same site, their effects would be expected to be additive and could potentially therefore reduce sulfonation of drugs and lead to altered pharmacological responses.

Wide-ranging genomic effects of plasticisers and related compounds.

UNLABELLED: The effects of four compounds, bis(2-ethylhexyl)phthalate (BEHP); diisodecylphthalate (DIP); 4-n-octylphenol (OP); 4-chloro-3-methylphenol (CMP), on gene expression (steady-state mRNA levels) across the whole human genome were studied in human TE671 cells. Effects were studied using the Affymetrics GeneChip Human Genome U133 Plus 2.0, HG-U133 Plus 2.0 arrays, The array analyses the expression of 47,000 transcripts and variants, including approximately 38,500 well characterised. All four compounds exerted statistically significant actions, affecting between 4 and 6.5% of all genes. Each compound had its own expression signature. In most instances where there was an effect, steady-state mRNA levels were decreased, although not always. CMP treatment caused most increases in mRNA levels. A mixture of DIP and CMP caused fewer changes in mRNA levels than either of the individual compounds. CONCLUSIONS: These plasticisers affected the steady-state mRNA levels of many human genes. Exposure to these compounds over many years has the potential to influence human health.

Phytoestrogens and xenoestrogens: the contribution of diet and environment to endocrine disruption.

Some endocrine disrupting compounds such as phthalates and phenols act non-genomically by inhibiting the sulfotransferase (SULT 1E1 and SULT 1A1) isoforms which inactivate estrogens by sulfonation. A range of environmental phenolic contaminants and dietary flavonoids was tested for inhibition of the human SULT 1A1, 1E1 and 2A1 isoforms. In particular, the plasticisers 4-n-octyl- and 4-n-nonyl-phenol inhibit SULT 1E1 with IC(50) values of 0.16 microM vs. 10nM estradiol while the 2-substituted chlorophenols show similar values. Flavonoids are also SULT inhibitors; tricin is a competitive inhibitor of SULT 1E1 with a K(i) of 1.5+/-0.8 nM. In a small pilot study to determine whether ingestion of soy flavonoids would affect SULT1A1 activity in vivo as well as in vitro, sulfonation of daidzein was reduced in a group of women 'at risk' of breast cancer, as compared with controls, although the SULT 1A1*1/SULT 1A1*2 allele ratio was not different. Endocrine disrupting effects in man may be multifactorial when components from both the diet and the environment act at the same point in steroid metabolism.

Allometric relationships predicting foliar biomass and leaf area:sapwood area ratio from tree height in five Costa Rican rain forest species.

We developed allometric equations to predict whole-tree leaf area (A(l)), leaf biomass (M(l)) and leaf area to sapwood area ratio (A(l):A(s)) in five rain forest tree species of Costa Rica: Pentaclethra macroloba (Willd.) Kuntze (Fabaceae/Mim), Carapa guianensis Aubl. (Meliaceae), Vochysia ferru-gi-nea Mart. (Vochysiaceae), Virola koshnii Warb. (Myristicaceae) and Tetragastris panamensis (Engl.) Kuntze (Burseraceae). By destructive analyses (n = 11-14 trees per species), we observed strong nonlinear allometric relationships (r(2) > or = 0.9) for predicting A(l) or M(l) from stem diameters or A(s) measured at breast height. Linear relationships were less accurate. In general, A(l):A(s) at breast height increased linearly with tree height except for Penta-clethra, which showed a negative trend. All species, however, showed increased total A(l) with height. The observation that four of the five species increased in A(l):A(s) with height is consistent with hypotheses about trade--offs between morphological and anatomical adaptations that favor efficient water flow through variation in the amount of leaf area supported by sapwood and those imposed by the need to respond quickly to light gaps in the canopy.

Plastic contamination of the food chain: A threat to human health?

Macro-plastic pollution is found in terrestrial and marine environments and is degraded to micro-particles (MP) and nano-particles (NP) of plastic. These can enter the human food chain either by inhalation or by ingestion, particularly of shellfish and crustaceans. Absorption across the gastrointestinal tract is relatively low, especially for MPs, which appear to have little toxicity. However, NPs are more readily absorbed and may accumulate in the brain, liver and other tissues in aquatic species and other animals. Studies using nanoparticles of other materials suggest that toxicity could potentially affect the central nervous system and the reproductive system, although this would be unlikely unless exposure levels were very high and absorption was increased by physiological factors.

Base pairing between the 3' exon and an internal guide sequence increases 3' splice site specificity in the Tetrahymena self-splicing rRNA intron.

It has been proposed that recognition of the 3' splice site in many group I introns involves base pairing between the start of the 3' exon and a region of the intron known as the internal guide sequence (R. W. Davies, R. B. Waring, J. Ray, T. A. Brown, and C. Scazzocchio, Nature [London] 300:719-724, 1982). We have examined this hypothesis, using the self-splicing rRNA intron from Tetrahymena thermophila. Mutations in the 3' exon that weaken this proposed pairing increased use of a downstream cryptic 3' splice site. Compensatory mutations in the guide sequence that restore this pairing resulted in even stronger selection of the normal 3' splice site. These changes in 3' splice site usage were more pronounced in the background of a mutation (414A) which resulted in an adenine instead of a guanine being the last base of the intron. These results show that the proposed pairing (P10) plays an important role in ensuring that cryptic 3' splice sites are selected against. Surprisingly, the 414A mutation alone did not result in activation of the cryptic 3' splice site.

Drug metabolism in the elderly: A multifactorial problem?

Whether or not an individual's drug metabolising capacity declines with advancing age is a vexing question. There is no clear evidence that drug metabolism itself ('the biologically-assisted chemical alteration of the administered parent molecule') is less efficient in healthy old age than at younger ages, whereas a decreased capacity may be associated with ill-health and frailty. However, elderly individuals do show a reduced enzyme induction capability and are less able to tolerate overdoses. It appears that the majority of deleterious clinical outcomes related to drug therapy in an elderly (usually ill or frail) population may be ascribed to various anatomical and physiological age-related changes. These may affect both pharmacodynamics and pharmacokinetics, but not necessarily drug metabolism. Information gleaned from animal studies undertaken mainly in rodents does not seem to be of relevance to humans and studies in healthy aged human populations may not highlight possible problems. However, certain circumstances may influence metabolic competence, and phenotyping rather than genotyping is of more value in identifying those susceptible to adverse drug reactions. This short review discusses the potential contributions of four factors (inflammation, circadian rhythm, gut microbes, epigenetic aspects) which may lead to alterations in drug metabolism with increasing age.

In utero exposure to carcinogens: Epigenetics, developmental disruption and consequences in later life.

The uterine environment is often viewed as a relatively safe haven, being guarded by the placenta which acts as a filter, permitting required materials to enter and unwanted products to be removed. However, this defensive barrier is sometimes breached by potential chemical hazards to which the mother may be subjected. Many of these toxins have immediate and recognisable deleterious effects on the embryo, foetus or neonate, but a few are insidious and leave a legacy of health issues that may emerge in later life. Several substances, falling into the categories of metals and metalloids, endocrine disruptors, solvents and other industrial chemicals, have been implicated in the development of long-term health problems in the offspring following maternal and subsequent in utero exposure. The mechanisms involved are complex but often involve epigenetic changes which disrupt normal cell processes leading to the development of cancers and also dysregulation of biochemical pathways.

Enteral feeding reduces metabolic activity of the intestinal microbiome in Crohn's disease: an observational study.

BACKGROUND/OBJECTIVES: Enteral feeding will induce remission in as many as 80-90% of compliant patients with active Crohn's disease (CD), but its method of action remains uncertain. This study was designed to examine its effects on the colonic microbiome. METHODS/SUBJECTS: Healthy volunteers and patients with CD followed a regimen confined to enteral feeds alone for 1 or 2 weeks, respectively. Chemicals excreted on breath or in faeces were characterised at the start and at the end of the feeding period by gas chromatography/mass spectrometry. RESULTS: One week of feeding in healthy volunteers caused significant changes in stool colour and deterioration in breath odour, together with increased excretion of phenol and indoles on the breath. Feeding for 2 weeks in patients with CD produced significant improvements in symptoms and a decrease in the concentration of C-reactive protein. The faecal concentrations of microbial products, including short-chain fatty acids (SCFAs), and potentially toxic substances, including 1-propanol, 1-butanol and the methyl and ethyl esters of SCFAs, showed significant falls. CONCLUSIONS: A significant change occurs in the production of microbial metabolites after enteral feeding in both healthy volunteers and patients with CD. Many of those detected in CD are toxic and may feasibly lead to the immunological attack on the gut microbiota, which is characteristic of inflammatory bowel disease. The reduction in the production of such metabolites after enteral feeding may be the reason for its effectiveness in CD.

Primary structure of V-ATPase subunit B from Manduca sexta midgut.

The amino acid sequence of a vacuolar-type ATPase (V-ATPase) subunit B has been deduced from a cDNA clone isolated from a Manduca sexta larval midgut library. The library was screened by hybridization with a labeled cDNA encoding subunit B of Arabidopsis thaliana tonoplast V-ATPase. The M. sexta V-ATPase subunit B consists of 494 amino acids with a calculated M(r) of 54,902. The amino acid sequence deduced for V-ATPase subunit B of M. sexta is between 98% and 76% identical with that of seven other V-ATPase subunits B and greater than 52% identical with three archaebacterial ATPase subunits B.

A direct correlation between nicotinamide N-methyltransferase activity and protein levels in human liver cytosol.

Phenotypic differences in nicotinamide N-methyltransferase (NNMT, E. C. 2.1.1.1) activity may be due to a genetic polymorphism. We report the characterisation of the hepatic NNMT activity in cytosol from normal human livers, enzyme protein levels determined by Western blotting and ELISA and mRNA levels determined by SDS-PAGE/Northern blotting. Subjects with high NNMT activity had high levels of NNMT protein and NNMT mRNA levels in hepatic cytosol and the converse was true for individuals with low NNMT activity. No differences in sequences were seen when cDNAs of individuals with high and low NNMT activity were compared. Thus phenotypic differences in the general population are due to differences in steady-state mRNA levels and not because of a polymorphism in the coding region of the NNMT gene.

The maturase encoded by a group I intron from Aspergillus nidulans stabilizes RNA tertiary structure and promotes rapid splicing.

The AnCOB group I intron from Aspergillus nidulans self-splices, providing the Mg2+ concentration is >/= 15 mM. The splicing reaction is greatly stimulated by a maturase protein encoded within the intron itself. An initial structural and biochemical analysis of the splicing reaction has now been performed. The maturase bound rapidly to the precursor RNA (kon approximately 3 x 10(9) M(-1) min(-1)) and remained tightly bound (koff /= 150 mM) was tenfold slower, in part because of the existence of an equilibrium between folded and partially folded RNA. In contrast, the maturase very effectively stabilized tertiary structure in 5 mM Mg2+, a noticeable example being an interaction between the P8 helix and a GNRA sequence that constitutes the L2 terminal loop of the P2 helix. Formation of the 5' splice-site recognition helix was assisted by either the maturase or high concentrations of Mg2+. The maturase was required during splicing so it is not a true chaperone. However, RNase protection assays and kinetic studies suggest that the maturase recognizes and facilitates folding of an intron with limited tertiary structure and even incomplete secondary structure.