Water with food intake does not influence caloric intake after gastric bypass (GBP): a cross-over trial.

BACKGROUND: Bariatric patients seeking information meet very different recommendations on postoperative diet and eating behaviour. A reason for variability may be lack of hard evidence. A national survey on current dietary advice was conducted to serve as background for the present study on how drinking during a meal influenced caloric consumption. METHODS: A standardised questionnaire was sent to all units in the Scandinavian Obesity surgery registry (SOReg) in order to obtain information regarding current diet advice after gastric bypass. Twenty-eight patients, 14 in each group, were studied either 2 months or 1 year after a standard gastric bypass (GBP). A standardised lunch was served on two separate days with or without water in randomised order. Meal and water weights were measured before and after. Hunger/satiety scores were obtained using visual analogue scales. RESULTS: Response rate for surgeons was low, for dieticians 75 %. No clear consensus for liquid intake during meals was found; few surgeons advised patients whether or not to drink with meals. All patients ate to full satiety. Two months post-GBP, 7/14 patients consumed more solid food when allowed drinking water; the increase in caloric consumption was not significant. One year post-GBP, 5/14 patients consumed more solid food when allowed drinking water, the difference not reaching statistical significance. CONCLUSION: Our study does not indicate that patients should refrain from drinking during meals the first year after a GBP, at least not from a caloric intake point of view.

Proximity of brain infarcts to regions of endogenous neurogenesis and involvement of striatum in ischaemic stroke.

BACKGROUND AND PURPOSE: Clinical stroke trials with stem cell-based approaches aiming for trophic actions, modulation of inflammation and neuroprotection are ongoing. However, experimental studies also suggest that neuronal replacement by grafted neural stem cells (NSCs) and possibly by endogenous NSCs from the subventricular zone (SVZ) may restore function in the stroke-damaged striatum. To evaluate the potential clinical impact of these findings, we analyzed the spatial relationship of infarcts to the SVZ and the proportion of individuals with striatal lesions in a consecutive series of ischaemic stroke patients. METHODS: Patients aged 20-75 years with first-ever ischaemic stroke underwent DW-MRI of the brain within 4 days after stroke onset. We analyzed location, size, number of acute focal ischaemic abnormalities and their spatial relationship to the SVZ. Stroke severity was assessed using NIH Stroke Scale (NIHSS). RESULTS: Of 108 included patients, the distance from the nearest margin of the infarct(s) to the SVZ was ≤2 mm in 51/102 patients with visible ischaemic lesions on DW-MRI. Twenty-four patients had involvement of striatum. Eight of these had predominantly striatal lesions, that is >50% of the total ischaemic lesion volume was located in caudate nucleus and/or putamen. These 8 patients had a median NIHSS of 3. CONCLUSIONS: Many stroke patients have infarcts located close to the SVZ, providing some supportive evidence that optimized endogenous neurogenesis may have therapeutic potential. However, predominantly striatal infarcts are rare and tend to give mild neurological deficits, indicating that striatum should not be the primary target for neuronal replacement efforts in humans.

First Report of "Candidatus Liberibacter solanacearum" Associated with Psyllid-Affected Carrots in Sweden.

Carrot (Daucus carota) plants with symptoms resembling those associated with the carrot psyllid Trioza apicalis and the bacterium "Candidatus Liberibacter solanacearum" (1-4) were observed in 70% of commercial fields in southern Sweden in August 2011, with approximately 1 to 45% symptomatic plants per field. T. apicalis, a pest of carrot in northern and central Europe, including Sweden, can cause as much as 100% crop loss and is associated with "Ca. L. solanacearum" (1-4). Symptoms on affected plants include leaf curling, yellow and purple discoloration of leaves, stunted growth of shoots and roots, and proliferation of secondary roots (3). Carrot plant and psyllid samples were collected from fields in the province of Halland. Total DNA was extracted from petiole and root tissues of 33 symptomatic and 16 asymptomatic plants (cvs. Nevis and Florida), with the cetyltrimethylammonium bromide (CTAB) buffer extraction method (2,3). DNA was also extracted from 155 psyllids (3). DNA samples were tested by PCR using primer pairs OA2/OI2c (5''-GCGCTTATTTTTAATAGGAGCGGCA-3'/5'-GCCTCGCGACTTCGCAACCCAT-3') and CL514F/R (5'-CTCTAAGATTTCGGTTGGTT-3'/5'-TATATCTATCGTTGCACCAG-3'), to amplify a portion of 16S rDNA and rplJ/rplL ribosomal protein genes, respectively, of "Ca. L. solanacearum" (2,3). A 1,168-bp 16S rDNA fragment was detected in the DNA from all 33 symptomatic and two asymptomatic plants, and a 668-bp rplJ/rplL fragment was amplified from the DNA of all 33 symptomatic and four asymptomatic plants, indicating the presence of liberibacter. DNA from 23 and 49 psyllid samples yielded similar amplicons with OA2/OI2c and CL514F/R primer pairs, respectively. Amplicons from the DNA of four carrot roots and three T. apicalis with each primer pair were cloned (pCR2.1-TOPO; Invitrogen, Carlsbad, CA) and three clones of each of the 14 amplicons were sequenced (MCLAB, San Francisco, CA). BLAST analysis of the 16S rDNA consensus sequences from carrot (GenBank Accession No. JN863095) and T. apicalis (GenBank Accession No. NJ863096) showed 100% identity to those of "Ca. L. solanacearum" previously amplified from carrot (GU373048 and GU373049) and T. apicalis (GU477254 and GU477255) from Finland (2,3). The rplJ/rplL consensus sequences from carrot (GenBank Accession No. JN863093) and T. apicalis (GenBank Accession No. JN863094) were 99% identical to the sequences of rplJ/rplL "Ca. L. solanacearum" ribosomal protein gene from carrots in Finland (GU373050 and GU373051). To our knowledge, this is the first report of "Ca. L. solanacearum" associated with carrot and T. apicalis in Sweden. The disease associated with this bacterium caused millions of dollars in losses to potato and several other solanaceous crops in North and Central America and New Zealand (1). This plant pathogen is also associated with significant economic damage to carrot crops observed in Finland (2,3). References: (1) J. E. Munyaneza. Southwest. Entomol. 35:471, 2010. (2) J. E. Munyaneza et al. Plant Dis. 94:639, 2010. (3) J. E. Munyaneza et al. J. Econ. Entomol. 103:1060, 2010. (4) A. Nissinen et al. Entomol. Exp. Appl. 125:277, 2007.

An in vitro comparison of possibly bioactive titanium implant surfaces.

The aim of the study was to compare Ca and P formation (CaP) and subsequent bone cell response of a blasted and four different possibly bioactive commercially pure (cp) titanium surfaces; 1. Fluoride etched (Fluoride), 2. Alkali-heat treated (AH), 3. Magnesium ion incorporated anodized (TiMgO), and 4. Nano HA coated and heat treated (nano HA) in vitro. Furthermore, to evaluate the significance of the SBF formed CaP coat on bone cell response. The surfaces were characterized by Optical Interferometry, Scanning Electron Microscopy (SEM) and X-ray Photoelectron Spectroscopy (XPS). CaP formation was evaluated after 12, 24 and 72 h in simulated body fluid (SBF). Primary human mandibular osteoblast-like cells were cultured on the various surfaces subjected to SBF for 72 h. Cellular attachment, differentiation (osteocalcin) and protein production (TGF-beta(1)) was evaluated after 3 h and 10 days respectively. Despite different morphological appearances, the roughness of the differently modified surfaces was similar. The possibly bioactive surfaces gave rise to an earlier CaP formation than the blasted surface, however, after 72 h the blasted surface demonstrated increased CaP formation compared to the possibly bioactive surfaces. Subsequent bone cell attachment was correlated to neither surface roughness nor the amount of formed CaP after SBF treatment. In contrast, osteocalcin and TGF-beta(1) production were largely correlated to the amount of CaP formed on the surfaces. However, bone response (cell attachment, osteocalcin and TGF-F production) on the blasted controls were similar or increased compared to the SBF treated fluoridated, AH and TiMgO surface.

Precipitation of calcium phosphate in the presence of albumin on titanium implants with four different possibly bioactive surface preparations. An in vitro study.

The aim of the present study was to compare the nucleating behaviour on four types of bioactive surfaces by using the simulated body fluid (SBF) model with the presence albumin. Titanium discs were blasted (B) and then prepared by alkali and heat treatment (AH), anodic oxidation (AO), fluoridation (F), or hydroxyapatite coating (HA). The discs were immersed in SBF with 4.5 mg/ml albumin for 3 days, 1, 2, 3 and 4 weeks and analysed with scanning electron microscopy/energy dispersive X-ray analysis (SEM/EDX) and X-ray photoelectron spectroscopy (XPS). Topographic surface characterisation was performed with a contact stylus profilometer. The results demonstrated that the bioactive surfaces initiated an enhanced calcium phosphate (CaP) formation and a more rapid increase of protein content was present on the bioactive surfaces compared to the blasted control surface. The observation was present on all bioactive surfaces. The fact that there was a difference between the bioactive surfaces and the blasted control surface with respect to precipitation of CaP and protein content on the surfaces support the fact that there may be biochemical advantages in vivo by using a bioactive surface.

Characterisation of structures in salivary secretion film formation. An experimental study with atomic force microscopy.

The purpose of the present study was to characterise the structure dynamics of pure salivary secretions retained on controlled surfaces with different surface energies in the early stage of salivary film formation. Germanium prisms prepared to have either low surface energy or medium surface energy were incubated in fresh secretions of either human parotid saliva (HPS) or human submandibular/sublingual saliva (HSMSLS) for 15, 90, and 180 min. After controlled rinsing with distilled water, the surfaces were air dried and thereafter imaged with atomic force microscopy (AFM). The amount of adsorbed material and the size of the structures detected increased with increased saliva exposure time. The film thicknesses varied from 10 to 150 nm, and both HPS and HSMSLS films contained structures with diameters varying from 40 nm to 2 microm. Some of these were clustered into special formations. The HPS films exhibited a more granular morphology than the HSMSLS films. Furthermore, branched lines were detected on the low surface energy germanium prisms incubated in saliva. The results indicate that exposure time, surface energy, and type of salivary secretion all are factors affecting the adsorption characteristics of salivary films.

A new method to quantify postexercise ST-deviation--the ST-deficit. A study in men at high and low-risk for coronary heart disease.

BACKGROUND: Quantitative heart rate adjusted exercise ST criteria like microV/beats per minute (bpm) improve the diagnostic accuracy of the exercise ECG. However, there are few quantitative HR adjusted postexercise variables available. The aim of the present exercise study was to evaluate a new such variable from computerized averaging of the postexercise ECG. METHODS: The presence of possible myocardial ischaemia in a population based sample of 74 elderly male hypertensives at high-risk of coronary heart disease, and in 42 age-matched clinically healthy males (reference group) at low-risk was assessed by exercise ECG. All men had a normal resting ECG without signs of ischaemia. VARIABLES STUDIED: standard ST-criteria, ST/HR slope < or =-2.4 microV. bpm-1, shape of the rate-recovery loop, the latter also with a new quantitative variable, the ST-deficit. RESULTS: In spite of a normal resting ECG many subjects showed an abnormal ST/HR slope during exercise, 43% in the hypertension group and 26% in the reference group. An abnormal rate-recovery loop (ST-deficit) also contributed substantially to identify patients with possible myocardial ischaemia, 30 vs. 10%, respectively (P<0.02); cumulatively for the two HR adjusted criteria 53% vs. 29%, respectively (P<0.02). Mean ST-deficit was significantly lower in the high-risk group. CONCLUSIONS: Effort-related myocardial ischaemia is frequently silent in elderly high-risk hypertensives and necessitates testing, preferably with computerized exercise ECG and heart rate adjusted ST criteria. A new quantitative variable to assess the postexercise rate-recovery loop in the time domain, the ST-deficit is described. This variable seems to effectively discriminate between subjects with low and high-risk for coronary heart disease and thus provides new information. Further studies are warranted to validate this variable against myocardial perfusion scintigraphy and coronary angiography.

Stroke induces widespread changes of gene expression for glial cell line-derived neurotrophic factor family receptors in the adult rat brain.

Gene expression for glial cell line-derived neurotrophic factor (GDNF) family ligands and receptors was analyzed with in situ hybridization after two focal ischemic insults of different severities. Focal ischemia was induced in rats by either 30 min or 2 h of middle cerebral artery occlusion (MCAO), causing damage to the striatum only, or involving also the parietal cortex, respectively. We found modest, transient elevation of GDNF mRNA in the dentate granule cell layer. In addition, the number of GDNF mRNA-expressing cells increased in the cortex and striatum after 2 h or 30 min of MCAO, respectively. No changes of neurturin or persephin mRNA expression were detected. Both c-Ret and GFRalpha1 mRNA levels were markedly increased in the ipsilateral cortex outside the ischemic lesion at 6-24 h after the 2-h insult, whereas GFRalpha2 expression was decreased in cortical areas both within and outside the lesion. Similar increases of c-Ret and GFRalpha1 mRNA levels were detected in the striatum, and to a lesser extent, in the cortex following 30 min of MCAO. The 2-h insult also gave rise to transient increases of c-Ret and GFRalpha1 mRNA in hippocampal subregions. Thirty minutes and 2 h of MCAO lead to elevated c-Ret, and GFRalpha1 or GFRalpha2 mRNA expression, respectively, in the ipsilateral ventroposterolateral thalamic nucleus. Both insults induced increased levels of GFRalpha1 mRNA in the subventricular zone of the lateral ventricle. Our data indicate major changes of GDNF family signaling in the forebrain, regulated mainly through altered receptor levels, in the post-ischemic phase. These changes could enhance neuroprotective and neuroregenerative responses both to endogenous and exogenous GDNF ligands.

N-methyl-D-aspartate receptor-mediated increase of neurogenesis in adult rat dentate gyrus following stroke.

Neurogenesis in the adult rat dentate gyrus was studied following focal ischemic insults produced by middle cerebral artery occlusion (MCAO). Animals were subjected to either 30 min of MCAO, which causes damage confined to the striatum, or 2 h of MCAO, which leads to both striatal and cortical infarction. When compared to sham-operated rats, MCAO-rats showed a marked increase of the number of cells double-labelled for 5-bromo-2'-deoxyuridine-5'-monophosphate (BrdU; injected during 4-6 days postischemia) and neuronal-specific antigen (NeuN; a marker of postmitotic neurons) in the ipsilateral dentate granule cell layer and subgranular zone at 5 weeks following the 2 h insult. Only a modest and variable increase of BrdU-labelled cells was found after 30 min of MCAO. The enhanced neurogenesis was not dependent on cell death in the hippocampus, and its magnitude was not correlated to the degree of cortical damage. Systemic administration of the N-methyl-D-aspartate (NMDA) receptor blocker dizocilpine maleate (MK-801) completely suppressed the elevated neurogenesis following 2 h of MCAO. Our findings indicate that stroke leads to increased neurogenesis in the adult rat dentate gyrus through glutamatergic mechanisms acting on NMDA receptors. This modulatory effect may be mediated through changes in the levels of several growth factors, which occur after stroke, and could influence various regulatory steps of neurogenesis.

Reactions in the oral mucous membrane after exposure to Carisolv--combined results from a clinical screening test in humans and an experimental study in rats.

OBJECTIVE: To evaluate reactions in the oral mucosa after direct contact with Carisolv. SETTING: The Faculty of Odontology in Göteborg, Sweden. SUBJECTS: 34 healthy persons for a clinical screening test and 35 Sprague Dawley rats for a histological study. DESIGN: Mixed Carisolv or 0.5 % NaOCl were soaked in paper and applied to either side of the medial frenula of the lower lip of 34 persons. The solutions were left on the oral mucosa for three minutes. Inspection was made and photographs were taken immediately after exposure and also after 1 hour, 24 hours, and 72 hours. Mixed Carisolv was applied in a similar manner as described above to 35 adult Sprague Dawley rats. The animals were killed and biopsies were taken immediately after Carisolv exposure and also after 1 hour, 24 hours, and 48 hours. The biopsies were sectioned and prepared for histomorphometrical evaluation in light microscopy where cells were counted on regions from the epithelium layer deeper into the mucous membrane. RESULTS: Some adverse reactions were detected on the oral mucosa of humans up to 24 hours after Carisolv exposure for 3 minutes. The detected inflammatory reactions were slight and no patient felt any discomfort. The results of the histological study on rat did not show any statistically significant increase of the number of cells at any time after Carisolv exposure. CONCLUSIONS: If the oral mucosa gets in direct contact with Carisolv for 3 minutes no or only a weak inflammatory response may be expected.

Characterization of the cloned atlantic cod neuropeptide Y-Yb receptor: peptide-binding requirements distinct from known mammalian Y receptors.

Five members of the neuropeptide Y (NPY) receptor family have been cloned in mammals. The recently cloned NPY receptor in the Atlantic cod seems to be distinct from the mammalian subtypes as it has only 50% identity to Y1, Y4, and y6 and only 30% to Y2 and Y5. In most of the other families of G-protein-coupled receptors, species homologues have 65-90% identity between fishes and mammals. The functional expression and detailed pharmacological characterization of this cod NPY receptor, designated Yb, is reported. Membranes of cells transiently transfected with cod Yb showed saturable [(125)I]PYY binding with a K(d) of 45 pM. The pharmacological profile is similar to those of both the zebrafish Yb and Yc receptors and distinct from those of the mammalian NPY receptors. In competition experiments the cod Yb receptor had the following rank order of potencies: porcine PYY = porcine NPY = p[Leu(31), Pro(34)]NPY > zebrafish PYY > zebrafish NPY >> NPY2-36 = NPY3-36 > NPY18-36 > bovine PP = [D-Trp(32)]NPY > BIBP3226. This is in sharp contrast to the high selectivity of BIBP3226 for the Y1 receptor from all mammalian species. Together with the low amino acid identity of cod Yb with the mammalian Y1, Y4, and y6 receptors, this is further support for the notion that fish Yb constitutes a distinct NPY receptor subtype.

Clinical implication of down-scatter in attenuation-corrected myocardial SPECT.

BACKGROUND: Interpretation of myocardial perfusion single photon emission computed tomography (SPECT) studies is hampered by attenuation artifacts. Attenuation correction methods with simultaneous emission and transmission are now commercially available. However, it has been observed in clinical practice that attenuation correction without down-scatter correction in a 1-day rest/stress myocardial perfusion protocol may lead to serious interpretation errors. Therefore the aim of this study was to study errors resulting from down-scatter under realistic conditions, thus providing a background for the assessment of further corrections. METHODS AND RESULTS: Forty-six patients underwent myocardial perfusion scintigraphy in a 1-day technetium 99m-tetrofosmin rest-stress SPECT protocol, with a moving 153Gd line-source device for attenuation correction without down-scatter correction. Short-axis slices were quantified as inferior/anterior, septal/lateral, and apical/remainder count ratios. The changes at rest (350 MBq) and exercise (900 MBq) induced by attenuation correction were studied. Attenuation correction gave differences in apparent perfusion between rest and exercise not seen before correction. The gender differences in inferior-anterior ratio were greatly reduced after correction at rest but remained at exercise. A torso phantom study indicated that these results were due to under-correction at exercise because of down-scatter. CONCLUSIONS: Down-scatter results in an underestimation of attenuation in simultaneous emission and transmission, if not accurately accounted for. Particularly, a high-dose study compared with a low-dose study, as in the 1-day protocol, might cause serious interpretation errors.

SHP-2 binds to Tyr763 and Tyr1009 in the PDGF beta-receptor and mediates PDGF-induced activation of the Ras/MAP kinase pathway and chemotaxis.

Activation of the beta-receptor for platelet-derived growth factor (PDGF) by its ligand leads to autophosphorylation on a number of tyrosine residues. Here we show that Tyr763 in the kinase insert region is a novel autophosphorylation site, which after phosphorylation binds the protein tyrosine phosphatase SHP-2. SHP-2 has also previously been shown to bind to phosphorylated Tyr1009 in the PDGF beta-receptor. Porcine aortic endothelial (PAE) cells transfected with a PDGF beta-receptor in which Tyr763 and Tyr1009 were mutated to phenylalanine residues failed to associate with SHP-2 after ligand stimulation. Moreover, PDGF-BB-induced Ras GTP-loading and Erk2 activation were severely compromised in the receptor mutant. Whereas the mitogenic response to PDGF-BB remained at the same level as in cells expressing wild-type PDGF beta-receptor, chemotaxis induced by PDGF-BB was significantly decreased in the case of the Y763F/Y1009F mutant cells, suggesting an important role for SHP-2 in chemotactic signaling.

Cloning of a neuropeptide Y/peptide YY receptor from the Atlantic cod: the Yb receptor.

Neuropeptide Y (NPY) belongs to a family of structurally related neuroendocrine peptides that bind to G protein-coupled receptors. Five different receptor subtypes have recently been cloned in mammals and we have found another three receptor genes in the zebrafish, called zYa, zYb, and zYc, that appear to be distinct subtypes as deduced from their widely different sequences. To elucidate the evolutionary relationships between the mammalian and zebrafish receptors, we have used the zebrafish probes to isolate genomic clones from another teleost fish, the Atlantic cod, Gadus morhua. We present here the sequence of the cod Yb gene, whose deduced protein sequence is equally identical to the zebrafish Yb (69%) and Yc proteins (66%). The two zebrafish receptors are 76% identical to each other, suggesting that they arose by gene duplication in the zebrafish lineage after divergence from the cod lineage. The five cloned mammalian NPY-family receptors and the three cloned zebrafish NPY receptors indicate that this is the largest receptor family among all peptide receptors that belong to the superfamily of G protein-coupled receptors.

Platelet-derived growth factor (PDGF)-induced actin rearrangement is deregulated in cells expressing a mutant Y778F PDGF beta-receptor.

Platelet-derived growth factor-stimulated actin rearrangement and edge ruffle formation have previously been shown to be dependent on activation of phosphatidylinositol 3'-kinase, the activity of which also is important for directed migration of cells. This lipid kinase binds to phosphorylated tyrosine residues Y740 and Y751 in the kinase insert of the human platelet-derived growth factor ss-receptor. We examined the role of two other tyrosine residues in the kinase insert of this receptor, Y775 and Y778, for ligand-induced actin rearrangement. Both were shown to be phosphorylation sites; Y775 was only marginally phosphorylated in cells expressing the wild-type ss-receptor, whereas Y778 was phosphorylated at higher stoichiometry. Mutant receptors Y775F, Y778F and Y775/778F were active kinases and mediated proliferative responses when expressed in porcine aortic endothelial cells. Fluorescence staining of actin in platelet-derived growth factor-stimulated PAE cells revealed that Y778 is involved in regulation of the actin cytoskeleton since the cells contained, apart from edge ruffles and circular ruffles, a novel type of giant ruffle on the dorsal side of the cell, which consisted of irregular multilayered actin structures. Mutation at Y778 had no effect on activation of phosphatidylinositol 3'-kinase, nor on the GTPase activating protein of Ras and phospholipase C(gamma), and the extent of directed migration towards platelet-derived growth factor of these cells was not changed. We conclude that actin rearrangement is regulated in part by Y778 in the platelet-derived growth factor ss-receptor, potentially through binding of a novel signaling molecule to this site.

A comparative analysis of the phosphoinositide binding specificity of pleckstrin homology domains.

Pleckstrin homology (PH) and phosphotyrosine binding (PTB) domains are structurally related regulatory modules that are present in a variety of proteins involved in signal transduction, such as kinases, phospholipases, GTP exchange proteins, and adapter proteins. Initially these domains were shown to mediate protein-protein interactions, but more recently they were also found to bind phosphoinositides. Most studies to date have focused on binding of PH domains to phosphatidylinositol (PtdIns)-4-P and PtdIns-4,5-P2 and have not considered the lipid products of phosphoinositide 3-kinase: PtdIns-3-P, PtdIns-3,4-P2, and PtdIns-3,4,5-P3. Here we have compared the phosphoinositide specificity of six different PH domains and the Shc PTB domain using all five phosphoinositides. We show that the Bruton's tyrosine kinase PH domain binds to PtdIns-3,4, 5-P3 with higher affinity than to PtdIns-4,5-P2, PtdIns-3,4-P2 or inositol 1,3,4,5-tetrakisphosphate (Ins-1,3,4,5-P4). This selectivity is decreased by the xid mutation (R28C). Selective binding of PtdIns-3,4,5-P3 over PtdIns-4,5-P2 or PtdIns-3,4-P2 was also observed for the amino-terminal PH domain of T lymphoma invasion and metastasis protein (Tiam-1), the PH domains of Son-of-sevenless (Sos) and, to a lesser extent, the PH domain of the beta-adrenergic receptor kinase. The oxysterol binding protein and beta-spectrin PH domains bound PtdIns-3,4,5-P3 and PtdIns-4,5-P2 with similar affinities. PtdIns-3,4,5-P3 and PtdIns-4,5-P2 also bound to the PTB domain of Shc with similar affinities and lipid binding was competed with phosphotyrosine (Tyr(P)-containing peptides. These results indicate that distinct PH domains select for different phosphoinositides.

Characterization of three separated exons in the HLA class II DR region of the human major histocompatibility complex.

The human major histocompatibility complex, HLA, is a highly polymorphic gene region which includes the DRA and DRB genes. The number of DRB genes differs between haplotypes. The DR4 haplotype seems to be one of the most complex with five DRB loci, DRB1, DRB4, DRB7, DRB8, and DRB9, in addition to the single DRA locus. We determined the nucleotide sequences of three separated DRB exons located between the DRB4 locus and the DRA locus in the DR4 haplotype, two DRB signal-peptide exons (S1 and S3) and one DRB first-domain exon (locus designation DRB9). Sequence comparisons suggest the following order of events for the origin of these exons: DRB9 seems to be the oldest exon and has previously been detected in multiple HLA haplotypes. DRB9 is more divergent than the three other known DRB pseudogenes, all of which have been found in apes. This suggests that DRB9 arose prior to the hominoid divergence. An L1 repeat has been inserted 3' to DRB9. Subsequently, a LTR of the ERV9 retrovirus-like family was inserted into the L1 repeat. Such LTRs have recently been observed in some of the other DRB genes. The pseudogenes DRB7 and DRB8 (containing only exons 3-6) arose after DRB9. Finally, the separated signal peptide exons S1 and S3 were formed. The molecular characterization of these separated DRB exons and insertion elements further clarifies the complex evolutionary history of the HLA-DR region. These selectively neutral exons may serve as useful markers for tracing the phylogeny of HLA haplotypes.

Effects of maternal pethidine on infants' developing breast feeding behaviour.

The aim of this quasi-experimental study was to examine the effects of maternal pethidine during labour on the developing breast feeding behaviour in infants in the first 2 h after birth compared with infants not exposed to pethidine. Forty-four healthy infants were observed immediately after birth. They were placed skin-to-skin on their mothers' chests. The development of mouth and sucking movements as well as rooting behaviour and state of sleep/wakefulness were noted. The observer was blind as to the pain relief the mother had received during labour. Of the 44 mothers 18 had received pethidine. The main findings were that infants exposed to pethidine had delayed and depressed sucking and rooting behaviour. In addition, a smaller proportion of infants exposed to pethidine started to suckle the breast. Rooting movements which are expected to be vigorous at 30 min after birth were affected both by administration of pethidine and a longer second stage of labour. It is suggested that the differences found in sucking behaviour may be a central effect of pethidine. Depression of rooting movements in the pethidine group may be caused by exhaustion due to a longer second stage of labour and administration of pethidine. It is recommended that pethidine-exposed mother-infant couples stay together after birth long enough to enable the infant to make the choice to attach or not to attach to the nipple without the forceful helping hand of the health staff.

Tyr-716 in the platelet-derived growth factor beta-receptor kinase insert is involved in GRB2 binding and Ras activation.

Ligand stimulation of the platelet-derived growth factor (PDGF) beta-receptor leads to activation of its intrinsic tyrosine kinase and autophosphorylation of the intracellular part of the receptor. The autophosphorylated tyrosine residues mediate interactions with downstream signal transduction molecules and thereby initiate different signalling pathways. A pathway leading to activation of the GTP-binding protein Ras involves the adaptor molecule GRB2. Here we show that Tyr-716, a novel autophosphorylation site in the PDGF beta-receptor kinase insert, mediates direct binding of GRB2 in vitro and in vivo. In a panel of mutant PDGF beta-receptors, in which Tyr-716 and the previously known autophosphorylation sites were individually mutated, only PDGFR beta Y716F failed to bind GRB2. Furthermore, a synthetic phosphorylated peptide containing Tyr-716 bound GRB2, and this peptide specifically interrupted the interaction between GRB2 and the wild-type receptor. In addition, the Y716(P) peptide significantly decreased the amount of GTP bound to Ras in response to PDGF in permeabilized fibroblasts as well as in porcine aortic endothelial cells expressing transfected PDGF beta-receptors. The mutant PDGFR beta Y716F still mediated activation of mitogen-activated protein kinases and an increased DNA synthesis in response to PDGF, indicating that multiple signal transduction pathways transduce mitogenic signals from the activated PDGF beta-receptor.