Evaluation of the LIM homeobox genes LHX6 and LHX8 as candidates for Tourette syndrome.

The etiology and pathophysiology of Tourette Syndrome (TS) remain poorly understood. Multiple lines of evidence suggest that a complex genetic background and the cortico-striato-thalamo-cortical circuit are involved. The role of Lhx6 and Lhx8 in the development of the striatal interneurons, prompted us to investigate them as novel candidate genes for TS. We performed a comparative study of the expression of Lhx6 and Lhx8 and investigated genetic association with TS using two samples of trios (TSGeneSEE and German sample - 222 families). We show that Lhx6 and Lhx8 expression in the forebrain is evolutionarily conserved, underlining their possible importance in TS-related pathophysiological pathways. Our tagging-single nucleotide polymorphism (tSNP)-based association analysis was negative for association with LHX8. However, we found positive association with LHX6 in the TSGeneSEE sample (corrected P-value = 0.006 for three-site haplotype around SNP rs3808901) but no association in the sample of German families. Interestingly, the SNP allele that was identified to be significantly associated in the TSGeneSEE dataset, showed an opposite trend of transmission in the German dataset. Our analysis of the correlation of the LHX6 region with individual ancestry within Europe, revealed the fact that this particular SNP demonstrates a high degree of population differentiation and is correlated with the North to South axis of European genetic variation. Our results indicate that further study of the LHX6 gene in relation to the TS phenotype is warranted and suggest the intriguing hypothesis that different genetic factors may contribute to the etiology of TS in different populations, even within Europe.

Depletion of CD4+CD25+CD127lo regulatory T cells does not increase allergen-driven T cell activation.

BACKGROUND: It has been suggested that allergic diseases are caused by defective suppression of allergen-specific Th2 cells by CD4(+)CD25(+) regulatory T cells. However, such studies have been hampered by the difficulty in distinguishing regulatory T cells from CD25-expressing activated T cells. Recently, it was shown that conventional T cells expressed high levels of CD127, whereas regulatory T cells were CD127(lo), allowing discrimination between these distinct T cell subpopulations. OBJECTIVE: The aim of this study was to study whether the putative regulatory subset defined as CD4(+)CD25(+)CD127(lo) was involved in grass pollen-reactive T cell responses. METHODS: Peripheral blood mononuclear cells (PBMCs) were obtained from allergic donors and non-atopic controls out of season. Grass pollen-induced cytokine production and proliferation were compared in cultures of undepleted cells and cells depleted of CD4(+)CD25(+), CD4(+)CD25(+)CD127(hi) or CD4(+)CD25(+)CD127(lo) T cells. RESULTS: Undepleted cell cultures from allergic patients showed significantly increased proliferation and Th2 cytokine production compared with non-atopic controls. Depletion of all CD25(+) T cells did not increase cytokine production or proliferation, and more importantly, no increase in Th2 cytokine production or proliferation was observed in cell cultures depleted of CD4(+)CD25(+)CD127(lo) cells (putative regulatory T cells) compared with undepleted PBMCs in both the allergic and the non-atopic group. CONCLUSION: Our study showed that T cells from grass pollen-allergic patients and non-atopic controls responded very differently to grass pollen extract, but this difference could not be explained by differences in regulatory T cell function. Further studies are needed to understand the importance of regulatory T cells in allergy.

Muramyldipeptide modulates CXCL-8 release of BEAS-2B cells via NOD2.

Chronic inflammation and acute exacerbations are pathophysiological features of chronic obstructive pulmonary disease (COPD). An impaired immune response to bacterial pathogens can contribute to both of them. Nucleotide oligomerization domain 2 (NOD2) is an intracellular receptor of innate immunity for muramyldipeptide (MDP). Mutations of the NOD2 gene followed by decreased recognition of MDP are associated with chronic intestinal inflammation and pulmonary complications of patients with allogenic stem cell transplant and sepsis. Our study provides evidence that NOD2, toll-like receptor 4 (TLR4) and the adapter protein receptor-interacting protein 2 (RIP2) are induced by tumor-necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) in the bronchial epithelial cell line BEAS-2B. We also demonstrate that lipopolysaccharide (LPS) can further increase NOD2 transcription in a TNF-alpha and IFN-gamma-induced activation state. In addition, we show that, while MDP fails to enhance CXCL-8 release from otherwise unstimulated BEAS-2B cells, a 12 h prestimulation period with TNF-alpha and IFN-gamma primes the cells for an additional increase of CXCL-8 secretion via induction of NOD2 and RIP2. LPS itself significantly augments CXCL-8 production and co-administration of MDP further increases cytokine secretion. Finally, overexpression of an SNP13 mutant decreased MDP-induced chemokine production in BEAS-2B cells compared with NOD2 wild type overexpression. Taken together, our work indicates that MDP and NOD2 play an important role for CXCL-8 release of BEAS-2B cells following LPS-challenge via synergistic interactions between MDP and LPS.

Plasmacytoid dendritic cells induce a distinct cytokine pattern in virus-specific CD4+ memory T cells that is modulated by CpG oligodeoxynucleotides.

Inherent properties of dendritic cell (DC) subsets are important in the regulation of naïve T-cell differentiation (e.g. Th1 versus Th2 cells), whereas effector memory T cells are believed to produce a fixed cytokine repertoire independent of the type of antigen presenting cell. Here we show that two distinct human DC subsets, plasmacytoid DC (PDC) and myeloid CD11c+ DC, induced autologous mumps virus-specific memory CD4(+) T cells to produce markedly different cytokine patterns upon antigen stimulation. PDC stimulated the T cells to produce gamma-interferon (IFN-gamma) and interleukin-(IL)-10, whereas CD11c+ DC induced lower levels of IFN-gamma, virtually no IL-10, but significant levels of IL-5. Analysis of intracellular cytokine production showed simultaneous production of IL-10 and IFN-gamma in mumps-specific T cells activated by PDC, a cytokine pattern similar to that described for Th1-like regulatory cells. Introduction of CpG oligodeoxynucleotides in PDC/T-cell co-cultures had synergistic effect on virus-dependent IFN-gamma production, whereas the other cytokines remained unchanged. Together, our results show that the type of DC involved in reactivation of previously primed T cells may have significant impact on the resulting cytokine response and suggest that targeting of viral antigens and adjuvant to specific DC subsets should be considered in the design of therapeutic antiviral vaccines.

Omeprazole and talampanel as two examples of retrometabolic drug design.

The goal of retrometabolic drug design is: "to design safe, locally active compounds with an improved therapeutic index". Here we describe two cases from our own practice, talampanel and omeprazole.

Heparin-binding epidermal growth factor-like growth factor: a component in chromaffin granules which promotes the survival of nigrostriatal dopaminergic neurones in vitro and in vivo.

Chromaffin cells can restore function to the damaged nigrostriatal dopaminergic system in animal models of Parkinson's disease. It has been reported that a protein which is released from chromaffin granules can promote the survival of dopaminergic neurones in vitro and protect them against N-methylpyridinium ion toxicity. This neurotrophic effect has been found to be mediated by astroglial cells and blocked by inhibitors of the epidermal growth factor (EGF) receptor signal transduction pathway. Here we report the identification of bovine heparin-binding EGF-like growth factor (HB-EGF) in chromaffin granules and the cloning of the respective cDNA from bovine-derived adrenal gland. Protein extracts from bovine chromaffin granules were found to promote the survival of embryonic dopaminergic neurones in culture, to the same extent as recombinant human HB-EGF. Furthermore, the neurotrophic action of the chromaffin granule extract could be abolished by antiserum to recombinant human HB-EGF. We also show that intracerebral injection of recombinant human HB-EGF protected the nigrostriatal dopaminergic system in an in vivo adult rat model of Parkinson's disease. Intracerebral administration of this protein at the same time as a 6-hydroxydopamine lesion of the medial forebrain bundle was found to spare dopamine levels in the striatum and tyrosine hydroxylase-immunopositive neurones in the midbrain. This study has found that the main component in chromaffin granules responsible for their neurotrophic effect on dopaminergic neurones is HB-EGF. Furthermore, HB-EGF has significant protective effects on nigrostriatal dopaminergic neurones in vivo, making it a potential candidate for use in the treatment of Parkinson's disease.

Differences in LPS-induced activation of bronchial epithelial cells (BEAS-2B) and type II-like pneumocytes (A-549).

Lipopolysaccharide (LPS) as a major component of the outer membrane of gram-negative bacteria stimulates various cells to initiate a signalling cascade which ultimately leads to cell activation and expression of immunoregulatory or inflammatory cytokines. The human respiratory epithelium is an important environmental interface, but differences in LPS-induced cell activation between bronchial and alveolar epithelial cells have not yet been investigated in detail. First, the expression of Toll-like receptors (TLRs), as pattern-recognition receptors, was investigated for the bronchial epithelial cells and type II-like pneumocytes, demonstrating that they fulfil the prerequisites for LPS signalling. Thereafter, the effects of LPS, soluble CD14 (sCD14) and LPS-binding protein (LBP) on the release of interleukin-6 (IL-6) and IL-8 were studied. In the presence of LPS, sCD14 induced a significant and concentration-dependent cytokine release in type II-like pneumocytes, whereas the response of bronchial epithelial cells to sCD14 stimulation was low, implicating sCD14-independent activation mechanisms. Furthermore, LBP revealed inhibitory effects on the activation of alveolar epithelial cells, which may represent a novel local defence mechanism during gram-negative infection. We conclude that distinct pathways exist for LPS-induced activation of bronchial and alveolar epithelial cells.

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) regulates survival of midbrain dopaminergic neurons.

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a member of the EGF-family of ligands signaling via the EGF-receptor tyrosine kinase. In the present study we show that HB-EGF which is expressed in close proximity of developing mesencephalic dopaminergic neurons promotes the survival of TH-positive neurons in vitro. The survival promoting effect of HB-EGF is mediated via astroglial cells and utilizes the MAPK as well as the Akt-signaling pathway. Most notably endogenous HB-EGF significantly contributes to the survival of TH-+ neurons in control cultures, suggesting a relevant developmental role of HB-EGF for dopaminergic neurons. These findings indicate that HB-EGF may be an important molecule for developing dopaminergic neurons of the ventral midbrain.

A study of anthropometric measures before and after external septoplasty in children: a preliminary study.

OBJECTIVE: To test the hypothesis that surgery on the growing nasal septum does not adversely affect nasal and midfacial dimensions. DESIGN: Paired study. SETTING: Tertiary care center. PARTICIPANTS: Children treated consecutively during a 4-year period; all had significant nasal obstruction and cosmetic disfigurement secondary to skeletal septal deformities. INTERVENTION: Nasal septal surgery (using an external approach), in which the quadrilateral cartilage was removed, remodeled, and reinserted as a free graft. OUTCOME MEASURES: Anthropometric linear measurements and indexes of the face and nose preoperatively and postoperatively; nasal dorsum length, nasal height, nasal dorsum index, nasal tip protrusion, columellar length, facial height, face width, upper face height, facial index, nose-upper face height index, and columellar length-nasal tip protrusion index. Continuous measurements were transformed into ordered categories with reference to normative data. Data were analyzed using Wilcoxon signed rank sum test (alpha level of.05) and by applying the Bonferroni adjustment for multiple testing. RESULTS: Twenty-six children were studied (12 females and 14 males); age at surgery ranged from 4.5 to 15.5 years (mean age, 9.5 years); average age at postoperative measurement, 12.5 years; mean follow-up, 3.1 years. Only nasal dorsum length (P =.007) and nasal tip protrusion (P =.04) were decreased by a statistically significant level before the Bonferroni adjustment. The change was not considered clinically significant. Thus, relative to age-appropriate norms, the dimensions of the nose and midface and their proportionality did not change after surgery. CONCLUSIONS: Appropriate nasal septal surgery involving excision and subsequent reinsertion of a remodeled segment of the quadrilateral cartilage has no deleterious effects on development of the nose and midface. We question the absolute dogma that nasal surgery in children must always be avoided.

The effects of electrical and controlled atmosphere stunning methods on meat and liver quality of geese.

The objective of this research was to investigate the effects of electrical and gas stunning on the meat and liver quality characteristics of liver geese. Sixty birds were slaughtered at 12 wk of age, in groups of 20 at three different times. Thirty birds each were subjected to one of the two stunning methods. Deboned breast fillets and thigh muscles were evaluated for hemorrhaging, amount of blood in the veins, and color by using a visual grading system. Livers were removed from carcasses during evisceration and were subsequently graded. Besides external color and hemorrhaging, the degree of liver weight loss due to removal of blood vessels was also determined. The use of controlled atmosphere stunning significantly reduced the incidence of muscle hemorrhages when compared to electrical stunning. However, no significant difference was found for color or amount of blood remaining in the veins of muscles between geese receiving electrical or controlled atmosphere stunning. The hemorrhaging and color scores of livers from gas-stunned birds did not differ from those of electrically stunned birds. As for the liver weight loss caused by removal of veins and capillaries, stunning treatment had no significant effect on this parameter. These results suggest that controlled atmosphere stunning produced slightly better quality goose meat but did not improve liver quality when compared to the electrical stunning method used.

Surface anatomy of the face in Down's syndrome: anthropometric proportion indices in the craniofacial regions.

The objective of the study was to identify the proportions closest to normal and those indicating mild-to-moderate and severe degrees of disproportion. Eight proportion indices were analyzed in five craniofacial regions of 125 Down's syndrome patients, based on a total of 985 data points. More than two thirds of the patients fell within the normal range, although more than one quarter were abnormal (disproportionate). All statistical summaries were based on z-scores (adjusting for age and sex differences), converted into descriptive anthropometric categories to yield a simplified frequency distribution for each proportion index. Normal proportions were harmonious in 55.9% of patients. Disproportions were mild to moderate in 66.4%, severe in 33.6%. The highest frequency of harmony was found in the head (70.2%), the lowest in the orbits (40.8%). The highest percentage of mild-moderate disproportion was found in the face (79.3%). The highest percentage of severe disproportions was recorded in the intercanthal index of the orbits (44.7%) and the smallest frequency in the face (20.7%). In the five craniofacial regions among the normal proportions, harmonies were more frequent than disharmonies. Among the disproportions, the percentage of mild-moderate ones was greater than those of severe degree.

Occurrence of anaerobic bacterial, clostridial, and Clostridium perfringens spores in raw goose livers from a poultry processing plant in Hungary.

Anaerobic bacterial, clostridial, and Clostridium perfringens spores were enumerated in raw goose liver samples taken after evisceration of the birds (EB) in the slaughterhouse and after removal of blood vessels from the liver (RBVL) in the cannery. The samples taken after RBVL had significantly higher (P < 0.05) spore counts than did those taken after EB, indicating contamination of livers during processing. The number of C. perfringens spores was one log cycle higher in the samples taken after RBVL than in those taken after EB (P < 0.05). The confirmation of C. perfringens according to the profiles of Rapid ID 32 A tests was carried out by means of the ATB Plus computer program. With an identification percentage of 99.9 and a T-value of 0.65, the suspect colonies proved to be C. perfringens. Therefore, the importance of an appropriate cleaning and sanitation program and of personnel hygiene should be emphasized in the industry.

Surface anatomy of the face in Down's syndrome: linear and angular measurements in the craniofacial regions.

Measurements (23 projective linear, 2 angular) taken in the 6 craniofacial regions of 127 patients with Down's syndrome showed that 63.1% (1,836 of 2,908) were within normal limits and 36.9% (1,072) were outside them. Abnormal measurements were subnormal in 90.8% (973) and supernormal in 9.2% (99). All statistical summaries were based on z scores (adjusting for age and sex differences) classified into a small number of ranges to yield a simplified frequency distribution for each measurement. The purpose of the study was to identify the measurements closest to normal and those indicating the most severe degrees of sub- or supernormality. Approximately a quarter of normal measurements were classified as optimal, and half the subnormal or supernormal measurements were classified as severe. Intercanthal width had the highest frequency of optimal measurements (93.7%, 119 of 127), head circumference the smallest (28.6%, 36 of 126). Knowledge of the frequency of extreme abnormalities in the craniofacial regions will help during visual examination of patients with Down's syndrome. This study found the highest percentage of severely subnormal measurements in the orbital region (57.8%, 74 of 128) and the smallest in the labio-oral region (32.7%, 16 of 49). The measurement with the highest proportion of severely subnormal to all subnormal values was the palpebral fissure length (68%, 51 of 75), and the nose width had the smallest proportion (14.3%, 1 of 7).

Short-term changes in renal function after extracorporeal shock wave lithotripsy in children.

PURPOSE: To our knowledge despite numerous studies the biological effect of extracorporeal shock wave lithotripsy (ESWL, Dornier Medical Systems, Inc., Marietta, Georgia) on the function of the immature kidney has not yet been evaluated. We determined the short-term effect of ESWL on renal function in children. MATERIALS AND METHODS: In a 5-year period 65 children were treated for 77 kidney stones by ESWL and followed regularly at our department. In 16 children the short-term effect of shock wave energy was studied by measuring blood parameters (sodium, potassium, urea, creatinine and C-reactive protein), urinary electrolytes (sodium, potassium and creatinine), urinary enzyme activity (aspartate transaminase, alanine transferase, alkaline phosphatase and lactate dehydrogenase) and the excretion of beta 2-microglobulin. Samples were obtained immediately before and 2 hours after ESWL, and on days 1, 2, 8, 15, 30 and 90 after treatment. RESULTS: Morphological changes in the kidneys were not detected by ultrasound during followup. No significant changes were noted in overall renal function, serum parameters or urine electrolytes. A significant elevation in the excretion of aspartate transaminase, alkaline phosphatase, lactate dehydrogenase and beta 2-microglobulin was observed, indicating proximal tubular dysfunction and cell destruction. Enzyme levels returned to baseline within 15 days. CONCLUSIONS: Our results demonstrate that shock wave energy induces transient functional damage of tubular function in children. Minimizing the kV. and number of shocks may decrease the deleterious effect. When considering functional regeneration time, the minimal interval between 2 shock wave treatments should be at least 15 days. The long-term effect needs further investigation.

Plasmacytoid dendritic cells (natural interferon- alpha/beta-producing cells) accumulate in cutaneous lupus erythematosus lesions.

Plasmacytoid dendritic cell (P-DC) precursors in peripheral blood produce large amounts of interferon (IFN)-alpha/beta when triggered by viruses. However, when incubated with interleukin-3 and CD40 ligand, the same precursors differentiate into mature DCs that stimulate naïve CD4(+) T cells to produce Th2 cytokines. We recently reported that P-DCs accumulate in nasal mucosa of experimentally induced allergic rhinitis, supporting a role for this DC subset in Th2-dominated inflammation. Here we examined whether P-DCs accumulate in cutaneous lesions of lupus erythematosus (LE), a disorder associated with increased IFN-alpha/beta production. Our results showed that P-DCs were present in 14 out of 15 tissue specimens of cutaneous LE lesions, but not in normal skin. Importantly, the density of P-DCs in affected skin correlated well (r(s) = 0.79,P < 0.0005) with the high number of cells expressing the IFN-alpha/beta-inducible protein MxA, suggesting that P-DCs produce IFN-alpha/beta locally. Accumulation of P-DCs coincided also with the expression of L-selectin ligand peripheral lymph node addressin on dermal vascular endothelium, adding further support to the notion that these adhesion molecules are important in P-DC extravasation to peripheral tissue sites. Together, our findings suggested that P-DCs are an important source of IFN-alpha/beta in cutaneous LE lesions and may therefore be of pathogenic importance.

Repair of bilateral complete cleft lip: intraoperative nasolabial anthropometry.

Synchronous repair of bilateral complete cleft lip and nasal deformity requires conception of three-dimensional form and fourth-dimensional changes with growth, as distorted by the malformation. The aim is to obviate typical postoperative nasolabial stigmata. The strategy is to construct fast-growing features on a smaller scale and slow-growing features on a normal or slightly larger scale. In this study, intraoperative alterations in nasolabial dimensions were documented by anthropometry in 46 consecutive infants with bilateral complete cleft lip. These values were averaged and compared with measures from normal Caucasian infants at ages 0 to 5 months and 6 to 12 months. Nasal height (n-sn) and nasal width (al-al), both fast-growing features, were set smaller (88 percent and 96 percent, respectively) than those of age-matched normal infants. In contrast, the slow-growing features, nasal protrusion (sn-prn) and columellar length, were constructed longer than normal (130 percent and 167 percent, respectively). Because all labial features grow rapidly, they were made diminutive in this study, with the exception of central vermilion-mucosal height (median tubercle), which was purposively made full. These maneuvers resulted in a normal, average overall upper-lip height (sn-sto). Two technical refinements also are described: (1) construction of deepithelialized bands flanking the philtral flap to improve surface contour; and (2) positioning and fixation of the dislocated alar cartilages, performed entirely through superiomedial nostril rim incisions.

Experimentally induced recruitment of plasmacytoid (CD123high) dendritic cells in human nasal allergy.

Recent evidence suggests that the previously enigmatic cell type designated plasmacytoid monocytes can function as dendritic cells and contribute substantially to both innate and adaptive immunity. This cell type has previously been described only in bone marrow, blood, and organized lymphoid tissue, but not at effector sites with direct Ag exposure such as the mucosae. Plasmacytoid dendritic cells (P-DCs) matured in vitro can induce T cells to produce allergy-promoting Th2 cytokines; therefore, their possible occurrence in nasal mucosa during experimentally elicited allergic rhinitis was examined. Patients with silent nasal allergy were challenged topically with relevant allergen daily for 7 days. Biopsy specimens as well as blood samples were obtained before and during such provocation, and P-DCs were identified by their high expression of CD123 (IL-3R alpha-chain), together with CD45RA. Our results showed that P-DCs were present in low and variable numbers in normal nasal mucosa but increased dramatically during the allergic reaction. This accumulation concurred with the expression of the L-selectin ligand peripheral lymph node addressin on the mucosal vascular endothelium. The latter observation was particularly interesting in view of the high levels of L-selectin on circulating P-DC precursors and of previous reports suggesting that these cells can enter organized lymphoid tissue via high endothelial venules (which express peripheral lymph node addressin constitutively). Together, our findings suggested that P-DCs are involved in the triggering of airway allergy and that they are directed to allergic lesions by adhesion molecules that normally mediate leukocyte extravasation in organized lymphoid tissue.