Differential chemokine expression patterns in tonsillar disease.

Expression profiles of CXC- and CC-chemokines in various forms of tonsillar disease were studied to evaluate whether certain chemokines play a predominant role in a specific subset of tonsillar disease. Total RNA was isolated from 89 biopsies (21 hyperplastic palatine tonsils, 25 adenoids, 16 chronic inflammatory palatine tonsils and 27 chronic inflammatory palatine tonsils with histological prove of acute inflammation), reverse transcribed and subjected to PCR amplifying IL-8, Gro-alpha, eotaxin-1, eotaxin-2, MCP-3, MCP-4 and RANTES. 2% agarose gel electrophoresis revealed a predominance of IL-8 in the chronic inflammatory palatine tonsil group compared to tonsillar hyperplasia. Furthermore, eotaxin-2 was strongly overexpressed in adenoid samples compared to chronic inflammatory specimens. Our data suggest that the majority of diseases related to adenoid formation are mediated via an eotaxin-2 expression, whereas chronic inflammatory tonsillitis is associated with IL-8 upregulation. These data imply that adenoids are related to a Th-2, and chronic inflammatory tonsillitis to a Th-1 based immune response.

[An assessment tool for analysing the early vocal development of young children with cochlear implant]. / Ein Untersuchungsverfahren zur Analyse der vorsprachlichen Entwicklung bei Kleinkindern mit Cochlea-Implantat.

BACKGROUND: Early vocal development of German-speaking cochlear implant recipients has rarely been assessed so far. There-fore the purpose of this study was to describe the early vocal development following successful implantation. METHODS: A case study was designed to assess the temporal progression of early vocal development in a young cochlear implant recipient who was bilaterally implanted at the age of 8;3 months. Data were collected during one year by recording parent-child interactions on a monthly basis. The first recording was made before the onset of the signal-processors, the 12 following recordings were made during the first year of implant use. The child's vocalizations were classified according to the vocalization categories and developmental levels from the Stark Assessment of Early Vocal Development--Revised (SAEVD-R). This assessment tool was translated into German in this study and used with German-speaking children for the first time. It allows a coding of prelinguistic utterances via auditory perceptual analysis. RESULTS: The results show an overall decrease of early vocalizations and an increase of speech-like vowels and consonants. In the first six months no apparent progress took place; The child produced almost exclusively vocalizations from Levels 1-3. In the second half of the year an increase of canonical utterances (Level 4) and advanced forms (Level 5) was observed. However, vocalizations beyond the canonical babbling phase, especially vocants and closants as well as their combinations, continued to be dominant throughout the first year of implant use. The progress of development of the child investigated in this study is comparable to other children implanted at young age who had also been assessed with the SAEVD-R. In comparison to normal-hearing children, the implanted child's development seemed to progress slightly faster. Interrater- and intrarater-reliability using the SAEVD-R were measured for two independent observers and for a first and second coding procedure and revealed to be acceptable to good. CONCLUSION: The use of SAEVD-R for an implanted German-speaking child allowed the investigation of prelinguistic vocal development before the onset of words. The fact that early vocalizations remain the dominant form throughout the first year of hearing experience emphasizes the importance of documenting and analysing prelinguistic vocal development in order to monitor progression of speech acquisition.

[Modeling developmental aspects of sensorimotor control of speech production]. / Ein neuronales Modell zur sensomotorischen Entwicklung des Sprechens.

BACKGROUND: Detailed knowledge of the neurophysiology of speech acquisition is important for understanding the developmental aspects of speech perception and production and for understanding developmental disorders of speech perception and production. METHOD: A computer implemented neural model of sensorimotor control of speech production was developed. The model is capable of demonstrating the neural functions of different cortical areas during speech production in detail. RESULTS: (i) Two sensory and two motor maps or neural representations and the appertaining neural mappings or projections establish the sensorimotor feedback control system. These maps and mappings are already formed and trained during the prelinguistic phase of speech acquisition. (ii) The feedforward sensorimotor control system comprises the lexical map (representations of sounds, syllables, and words of the first language) and the mappings from lexical to sensory and to motor maps. The training of the appertaining mappings form the linguistic phase of speech acquisition. (iii) Three prelinguistic learning phases--i. e. silent mouthing, quasi stationary vocalic articulation, and realisation of articulatory protogestures--can be defined on the basis of our simulation studies using the computational neural model. These learning phases can be associated with temporal phases of prelinguistic speech acquisition obtained from natural data. CONCLUSIONS: The neural model illuminates the detailed function of specific cortical areas during speech production. In particular it can be shown that developmental disorders of speech production may result from a delayed or incorrect process within one of the prelinguistic learning phases defined by the neural model.

Conditional responding is impaired in chronic alcoholics.

Bechara (2003) describes a model for disturbances in executive functions related to addiction. This model involves deficits in decision-making and in suppressing pre-potent representations or response patterns. We tested this model in 29 individuals with long-term heavy alcohol dependency and compared their performance with that of 20 control subjects. Only individuals without memory impairment, with normal intelligence and normal visual response times were included. We examined word fluency, object alternation, spatial stimulus-response incompatibility, extra-dimensional shift learning and decision-making using the Gambling task. We subtracted the performance in a control condition from that of the executive condition, in order to focus specifically on the executive component of each task. Only the object alternation and incompatibility tasks revealed significant differences between the group of alcoholics and the control group. Moreover, response times in the object alternation task correlated with duration of alcohol dependency. The results do not argue in favor of a specific deficit in decision-making or in shifting between relevant representations. We conclude that long-term alcohol abuse leads to an impairment in conditional responding, provided the response depends on former reactions or the inhibition of pre-potent response patterns.

[MRT sequences as a database for a visual articulatory model]. / MRT-Sequenzen als Datenbasis eines visuellen Artikulationsmodells.

Articulatory models can be used in phoniatrics for the visualisation of speech disorders, and can thus be used in teaching, the counselling of patients and their relatives, and in speech therapy. The articulatory model developed here was based on static MRI data of sustained sounds. MRI sequences are now being used to further refine the model with respect to speech movements. Medio-sagittal MRI sections were recorded for 12 consonants in the symmetrical context of the three point vowels [i:], [a:] and [u:] for this corpus. The recording-rate was eight images/s. The data show a strong influence of the vocalic context on the articulatory target-positions of all consonants. A method for the reduction of the MRI data for subsequent qualitative and quantitative analyses is presented.

[A visual model of articulation]. / Ein visuelles Modell der Artikulation.

Speech disorders become more and more important in clinical area of phoniatrics. It is striking during consultation and treatment of patients that there exist only humble concrete notions concerning speech organs and articulatory movements. Using the computational model introduced here speech organs and speech movements of unrestricted utterances can be displayed in the mediosagittal plane. Articulation is described concisely by using nine basic parameters. The time pattern of these parameter-values defines the articulatory motion plan of an utterance and gives a comprehensive overview of the spatio-temporal structure of the appertaining speech movements. The use of the model in speech therapy shall make patients sensitive for the existence of speech articulators--especially for those which are normally not directly visible (back tongue region, soft palate)--and help to understand and memorize the appertaining movement patterns.

[Bradycardia despite hyperthyroidism]. / Bradykardie trotz Hyperthyreose.

Hyperthyroidism is usually associated with tachycardia, hypothyroidism with bradycardia. After observing clinically inapparent hyperthyroidism in patients requiring pacemaker implantation, we studied the occurrence of hyperthyroidism in patients receiving a first permanent pacemaker. Of 237 patients (age 71.4 +/- 8.9 years; 54.9% females), 16 (6.75%) had subclinical (TSH < 0.1 mE/l and fT3 < or = 9.0 pmol/l) and 4 (1.69%) overt hyperthyroidism (TSH < 0.1 mE/l and fT3 > 9.0 pmol/l). Prevalence of hyperthyroidism was similar to that in the general population. Compared to euthyroid patients, in the patients with subclinical or overt hyperthyroidism there were significantly more females (n = 16) than males (n = 4; p = 0.018). Hyperthyroid patients were older (75.0 +/- 9.6 vs. 70.7 +/- 8.9 years; p = 0.015). At follow-up, all patients had a relevant proportion of pacemacer-induced beats. Clinical signs of hyperthyroidism or cardiac symptoms were not different between groups. In conclusion, bradycardia does not exclude the presence of hyperthyroidism. Temporary pacing is recommended in thyreotoxicosis with bradycardia. In contrast, primary implantation of a permanent pacemaker appears to be adequate in patients with bradycardia, cardiovascular disease and an additional diagnosis of hyperthyroidism.

Crystal structure of the human alpha-thrombin-haemadin complex: an exosite II-binding inhibitor.

The serine proteinase alpha-thrombin plays a pivotal role in the regulation of blood fluidity, and therefore constitutes a primary target in the treatment of various haemostatic disorders. Haemadin is a slow tight- binding thrombin inhibitor from the land-living leech Haemadipsa sylvestris. Here we present the 3.1 A crystal structure of the human alpha-thrombin- haemadin complex. The N-terminal segment of haemadin binds to the active site of thrombin, forming a parallel beta-strand with residues Ser214-Gly216 of the proteinase. This mode of binding is similar to that observed in another leech-derived inhibitor, hirudin. In contrast to hirudin, however, the markedly acidic C-terminal peptide of haemadin does not bind the fibrinogen-recognition exosite, but interacts with the heparin-binding exosite of thrombin. Thus, haemadin binds to thrombin according to a novel mechanism, despite an overall structural similarity with hirudin. Haemadin inhibits both free and thrombomodulin-bound alpha-thrombin, but not intermediate activation forms such as meizothrombin. This specific anticoagulant ability of haemadin makes it an ideal candidate for an antithrombotic agent, as well as a starting point for the design of novel antithrombotics.

An isopeptide bond splitting enzyme from Hirudo medicinalis similar to gamma-glutamyl transpeptidase.

A new enzyme from Hirudo medicinalis capable of splitting gamma-glutamyl-p-nitroanilide and Glu--Lys-(N6-gamma-glutamyllysine) (isopeptidic bond between the epsilon-amino group of lysine and the gammacarboxylic group of glutamic acid) isopeptide bonds was purified. The protein was partially sequenced at the amino acid level, and the complete nucleotide and amino acid sequences were determined after cDNA cloning. The new enzyme has more than 60% similarity at the amino acid level to vertebrate gammaglutamyl transpeptidase (gamma-GT). According to the cDNA, the new protein has a molecular mass of 65 521 Da and a length of 600 amino acids.

Monoclonal antibodies against human endothelin-converting enzyme-1.

Endothelin-converting enzyme-1 (ECE-1) is a membrane-bound metalloprotease which specifically converts the inactive precursor big-endothelin-1 (big ET-1) to the vasoactive endothelin-1 (ET-1). Six different mouse hybridoma cell lines have been generated secreting monoclonal antibodies specific to human ECE-1. These antibodies have been proven useful in a fast and efficient one-step purification of membrane-bound ECE-1 as well as of artificial soluble ECE-1 by immunoaffinity chromatography. The antibodies are suitable for a quantification of ECE-1 in solution by a sandwich-ELISA and for the immunohistochemical detection of ECE-1 in the cell membrane.

Evidence of alternative promoters directing isoform-specific expression of human endothelin-converting enzyme-1 mRNA in cultured endothelial cells.

The endothelins, a family of closely related vasoactive and mitogenic peptides, are thought to play an important role in cardiovascular pathophysiology. The conversion of the inactive precursor "big endothelin" to the biologically active peptide is catalyzed in vitro and in vivo by endothelin-converting enzymes (ECE). Recently the cDNA cloning of two homologous proteins, termed ECE-1 and ECE-2, has been reported. ECE-1 may play a key role in the activation and regulation of the cardiovascular endothelin proteolytic cascade. ECE-1 mRNA is expressed in two isoforms, termed alpha and beta, which are identical except for the 5'-terminal regions. To investigate the transcriptional regulation of isoform-specific ECE-1 mRNA expression we isolated phage clones from a human genomic library and identified the alpha- and beta-specific exons of ECE-1. The exon/intron organization of the 5'-terminal region of the human ECE-1 gene in conjunction with putative transcription initiation start sites suggests the existence of two alternative promoters, each directing the expression of either isoform. A reverse transcription/polymerase chain reaction assay indicated differential mRNA expression of ECE-1 isoforms. Using a luciferase reporter gene assay, we found that the genomic region upstream of exon 1 alpha confers strong promoter activity in the human endothelial cell line ECV 304, which was previously shown to express predominantly ECE-1 alpha mRNA. Transfection of serial deletion mutants in ECV304 cells indicated the existence of three positive and also one negative regulating element within 2 kb of the alpha-promoter region. Luciferase reporter gene studies also revealed that the genomic region upstream of exon 3, which encodes the putative ECE-1 beta specific N-terminus, was able to direct luciferase expression in primary cultured bovine aortic endothelial cells, indicating the existence of an alternative promoter. Transfection of nested deletions spanning 1.2 kb upstream of the putative translation initiation codon of ECE-1 beta suggested the existence of three positive regulating regions within the beta-specific promoter. Both ECE-1 promoters lack TATA or CAAT boxes, and the two show different patterns of consensus sequences for transcription factors, suggesting a differential transcriptional regulation of isoform-specific ECE-1 mRNA expression.

A phase I/II study of idarubicin, dexamethasone and interferon-alpha (I-Dexa) in patients with relapsed or refractory multiple myeloma.

To improve the outcome for patients with relapsed or refractory multiple myeloma (MM), we combined idarubicin (Ida), dexamethasone and interferon (IFN) in a new regimen, 'I-Dexa'. Here we report our results of a phase I/II study using the I-Dexa protocol in an outpatient setting. A total of 31 patients were enrolled. Most patients were heavily pretreated with a median of two different chemotherapy regimen (range, 1-4). All but four patients had advanced disease (stage III). The dose of Ida was escalated to define the maximal tolerated dose (MTD) in this regimen. Four patients were treated with 5 mg/m2 and 27 patients with 7.5 mg/m2 Ida (day 1, i.v.). Dexamethasone was given at a fixed dose of 20 mg per os daily on days 2-5 and 11-14. Treatment was repeated at day 21 and consisted of up to six cycles. Patients who achieved a response or stable disease received IFN maintenance. IFN was administered three times weekly at a dose of 3 x 10(6) U/day s.c. until relapse. At the time of evaluation, 125 courses of I-Dexa were analyzed. The MTD of Ida in this regimen was 7.5 mg/m2. Hematological toxicity was mild including 5% leukocytopenia, 3% thrombocytopenia and 1% anemia (WHO grade III) at dose level 2. The MTD was defined by nonhematological toxicities including two WHO grade IV infections and one renal failure. The overall response rate including stable disease was 62.5% (PR: nine patients, 37.5%). So far, nine patients have been treated with IFN maintenance therapy with a median duration of 7 months (range, 1-16). In conclusion, I-Dexa can be given safely in an outpatient setting and is effective for the treatment of relapsed and refractory MM.

Two heads are better than one: crystal structure of the insect derived double domain Kazal inhibitor rhodniin in complex with thrombin.

Rhodniin is a highly specific inhibitor of thrombin isolated from the assassin bug Rhodnius prolixus. The 2.6 Angstrum crystal structure of the non-covalent complex between recombinant rhodniin and bovine alpha-thrombin reveals that the two Kazal-type domains of rhodniin bind to different sites of thrombin. The amino-terminal domain binds in a substrate-like manner to the narrow active-site cleft of thrombin; the imidazole group of the P1 His residue extends into the S1 pocket to form favourable hydrogen/ionic bonds with Asp189 at its bottom, and additionally with Glu192 at its entrance. The carboxy-terminal domain, whose distorted reactive-site loop cannot adopt the canonical conformation, docks to the fibrinogen recognition exosite via extensive electrostatic interactions. The rather acidic polypeptide linking the two domains is displaced from the thrombin surface, with none of its residues involved in direct salt bridges with thrombin. The tight (Ki = 2 x 10(-13) M) binding of rhodniin to thrombin is the result of the sum of steric and charge complementarity of the amino-terminal domain towards the active-site cleft, and of the electrostatic interactions between the carboxy-terminal domain and the exosite.

Acute and chronic effects of winter swimming on LH, FSH, prolactin, growth hormone, TSH, cortisol, serum glucose and insulin.

The present investigation is based on a 2.5 months selbstversuch (self-experiment) of the authors, between October 21 1992, and January 6 1993. 11 healthy students, five females and six males, age 24 to 29 years, and their teachers underwent regular winter swimming at least once a week, for 2 to 10 minutes, at the natural water temperature (6.8 degrees C (October 1992) to 2.0 degrees C (January 1993)) in the southern Baltic Sea. Blood samples were drawn before and 30 and 60 minutes after the cold bath, both at the first and the last day of the swimming season. TSH increased from 0.96 mU/l to 1.42 mU/l (p < 0.01) in the untrained, and from 0.93 mU/l to 1.43 mU/l (p < 0.01) in the cold-trained persons, and decreased thereafter (p < 0.01). Similar changes occurred in cortisol serum concentrations, though psychological stress seemed to interfere with cold stress. Cortisol increased from 99 ng/ml to 133 ng/ml in the untrained, and from 101 ng/ml to 137 ng/ml (p < 0.05) in the cold-trained persons within 30 minutes after cold water immersion, and decreased thereafter (p < 0.01). There were mild decreases in prolactin serum levels after cold stress, whereas FSH, LH and growth hormone remained unaltered. There was a mild initial elevation of serum glucose after cold stress (plus 12 mg/dl, (p < 0.01)) which disappeared after training. There were long term training effects besides the effects on glucose: Basal prolactin levels increased by almost the factor two, and insulin serum levels dropped by almost 50%.(ABSTRACT TRUNCATED AT 250 WORDS)

Molecular characterization of human and bovine endothelin converting enzyme (ECE-1).

A membrane-bound protease activity that specifically converts Big endothelin-1 has been purified from bovine endothelial cells (FBHE). The enzyme was cleaved with trypsin and the peptide sequencing analysis confirmed it to be a zinc chelating metalloprotease containing the typical HEXXH (HELTH) motif. RT-PCR and cDNA screens were employed to isolate the complete cDNAs of the bovine and human enzymes. This human metalloprotease was expressed heterologously in cell culture and oocytes. The catalytic activity of the recombinant enzyme is the same as that determined for the natural enzyme. The data suggest that the characterized enzyme represents the functional human endothelin converting enzyme ECE-1.

Murine protein which binds preferentially to oligo-C-rich single-stranded nucleic acids.

Two single-stranded nucleic acid binding proteins mCBP and mCTBP were identified by means of their binding to a potential recombination hotspot in LTRs of mouse retro-transposons. Both are nuclear proteins of 35 and 55 kDa respectively. mCBP binds preferentially to oligo dC, mCTBP to oligo dCdT. mCBP was purified and its cDNA was isolated and sequenced.

A Kazal-type inhibitor with thrombin specificity from Rhodnius prolixus.

A thrombin-specific inhibitor with an apparent molecular mass of 11 kDa has been purified from the insect Rhodnius prolixus. Amino-terminal protein sequence analysis allowed the molecular cloning of the corresponding cDNA. The open reading frame codes for a protein of about 103 amino acid residues and displays an internal sequence homology of residues 6-48 with residues 57-101 indicating a two-domain structure. Based on the amino acid sequence the two domains exhibit high homology to protease inhibitors belonging to the Kazal-type family. Model building suggests that the first domain binds to the active site with residue His10 pointing into the specificity pocket. From gel filtration and tight-binding inhibition experiments the inhibitor appears to form 1:1 complexes with thrombin. Periplasma-directed heterologous expression of the rhodniin cDNA in Escherichia coli yields the intact thrombin inhibitor. Natural and recombinant rhodniin both display inhibition constants of about 2 x 10(-13) M.

Isolation, sequence analysis, and cloning of haemadin. An anticoagulant peptide from the Indian leech.

A slow, tight-binding inhibitor of thrombin with an apparent molecular mass of about 5 kDa has been isolated from Haemadipsa sylvestris, an Indian leech of the family of Haemadipsidae. The inhibitory activity, called haemadin, is thrombin specific since it does not inhibit other proteases like trypsin, chymotrypsin, factor Xa, or plasmin. NH2-terminal amino acid sequence analysis (residues 1-45) does not reveal any homology to known serine protease inhibitors, including the thrombin-specific inhibitor hirudin. The haemadin cDNA cloned by polymerase chain reaction techniques codes for a polypeptide of 57 amino acid residues preceded by 20 residues of a signal peptide sequence. A synthetic gene coding for the mature haemadin was expressed in Escherichia coli. Recombinant haemadin displays a similar inhibition constant and specific activity as its natural counterpart. Although there is no obvious sequence identity between haemadin and hirudin, both proteins seem to share common mechanisms for thrombin inhibition.

A gestural production model and its application to reduction in German.

A quantitative speech production model has been computer-implemented based on specifications of gestures as its input. Articulatory gestures serve the central role of phonological/phonetic units in speech organization. The present model is not based on task dynamics, but it still assumes a critically damped linear second-order system. The capability of this model is partially demonstrated by its application to reduction phenomena in conversational speech in German. Explanations in terms of timing and magnitude alterations of gestures in different articulators use only a few general principles for a variety of apparent segmental alterations. Some salient examples have been synthesized based on the current model, which also contributed to the evaluation of the parameter values involved in sample gestures.