The mouse Mid1 gene: implications for the pathogenesis of Opitz syndrome and the evolution of the mammalian pseudoautosomal region

We have recently reported isolation of the gene responsible for X- linked Opitz G/BBB syndrome, a defect of midline development. MID1 is located on the distal short arm of the human X chromosome (Xp22. 3) and encodes a novel member of the B box family of zinc finger proteins. We have now cloned the murine homolog of MID1 and performed preliminary expression studies during development. Mid1 expression in undifferentiated cells in the central nervous, gastrointestinal and urogenital systems suggests that abnormal cell proliferation may underlie the defect in midline development characteristic of Opitz syndrome. We have also found that Mid1 is located within the mouse pseudoautosomal region (PAR) in Mus musculus , while it seems to be X- specific in Mus spretus. Therefore, Mid1 is likely to be a recent acquisition of the M. musculus PAR. Genetic and FISH analyses also demonstrated a high frequency of unequal crossovers in the murine PAR, creating spontaneous deletion/duplication events involving Mid1. These data provide evidence for the first time that genetic instability of the PAR may affect functionally important genes. In addition, we show that MID1 is the first example of a gene subject to X-inactivation in man while escaping it in mouse. These data contribute to a better understanding of the molecular content and evolution of the rodent PAR.      

Cellular basis of an auto-anti-allotypic mechanism for the maintenance of chronic allotype suppression in the rabbit.

Immunocytochemical identification of antibody-forming cells (AFCs) in situ was used to test the hypothesis that the maintenance of chronic allotype suppression in heterozygous rabbits is the result of an autoimmune B-cell-mediated response. Appreciable numbers of B cells with antibody activity directed against the suppressed allotypic determinant were found in spleen and bone marrow sections of all chronically suppressed rabbits examined. Appropriate double-staining was used to determine that such cells were of the non-suppressed allotype. These cells were indistinguishable from anti-allotypic AFCs found in larger numbers in spleens of normal heterozygous rabbits that had been immunized against a heterologous allotypic determinant. Auto-anti-allotypic AFCs were not found in suppressed rabbits less than 8 week old, nor were they found in normal (non-suppressed) heterozygous rabbits or chimeric rabbits formed by the injection of histocompatible but allotype-mismatched lymphoid cells at birth. The findings reported here support the hypothesis that the long-term maintenance of allotype suppression in the rabbit may result from the suppressive activities of autoimmune B cells. It is suggested that the suppression of an allotype during the first few weeks of life could result in a loss of tolerance to a self-determinant. The kinetics of auto-anti-AFC production support this idea in showing that such cells are generated following the decline of the antibody used to induce suppression. The triggering event may be the emergence of B cells expressing the previously suppressed gene product.     

The effect of anti-IgE treatment on in vitro leukotriene release in children with seasonal allergic rhinitis

BACKGROUND: Binding of allergens with IgE to the IgE receptors on mast cells and basophils results in the release of inflammatory mediators as sulfidoleukotrienes (SLTs), triggering allergic cascades that result in allergic symptoms, such as asthma and rhinitis. OBJECTIVE: We sought to investigate whether anti-IgE (Oma-lizumab), a humanized monoclonal anti-IgE antibody, in addition to specific immunotherapy (SIT) affects the leukotriene pathway. METHODS: Ninety-two children (age range, 6-17 years) with sensitization to birch and grass pollens and with seasonal allergic rhinitis were included in a phase III, placebo- controlled, multicenter clinical study. All subjects were randomized to one of 4 treatment groups. Two groups subcutaneously received birch SIT and 2 groups received grass SIT for at least 14 weeks before the start of the birch pollen season. After 12 weeks of SIT titration, placebo or anti-IgE was added for 24 weeks. The primary clinical efficacy variable was symptom load (ie, the sum of daily symptom severity score and rescue medication score during pollen season). Blood samples taken at baseline and at the end of study treatment after the grass pollen season were used for separation of leukocytes in this substudy. After in vitro stimulation of the blood cells with grass and birch pollen allergens, SLT release (LTC4, LTD4, and LTE4) was quantified by using the ELISA technique. RESULTS: Before the study treatment, SLT release to birch and grass pollen exposure did not differ significantly among the 4 groups. Under treatment with anti-IgE + SIT-grass (n = 23), a lower symptom load occurred during the pollen season compared to placebo + SIT-grass (n = 24, P =.012). The same applied to both groups receiving birch SIT (n = 23 and n = 22, respectively; P =.03). At the end of treatment, the combination of anti-IgE plus grass SIT, as well as anti-IgE plus birch SIT, resulted in significantly lower SLT release after stimulation with the corresponding allergen (416 ng/L [5th-95th percentile, 1-1168] and 207 ng/L [1-860 ng/L], respectively) compared with placebo plus SIT (2490 ng/L [384-6587 ng/L], P =.001; 2489 ng/L [1-5670 ng/L], P =.001). In addition, treatment with anti-IgE was also followed by significantly lower SLT releases to the allergens unrelated to SIT (grass SIT: 300 ng/L [1-2432 ng/L] in response to birch allergen; birch SIT: 1478 ng/L [1-4593 ng/L] in response to grass pollen) in comparison with placebo (grass SIT: 1850 ng/L [1-5499 ng/L], P =.001; birch SIT: 2792 ng/L [154-5839 ng/L], P =.04]. CONCLUSION: Anti-IgE therapy reduces leukotriene release of peripheral leukocytes stimulated with allergen in children with allergic rhinitis undergoing allergen immunotherapy independent of the type of SIT allergen used.     

Primary sity immune response in popliteal lymph nodes and spleen of mice after subcutaneous immunization with thymus-dependent or thymus-independent (type 1 and 2) antigens

Mice were immunized subcutaneously with thymus-independent (TI)-type 1 antigen trinitrophenylated lipopolysacccgaride (TNP-LPS), TI-type 2 antigen TNP-Ficoll or thymus-dependnt (TD) antien TNP-keyhole limpet haenicyanin (TNP-KLH) in order to study the primary in situ immune response in popliteal lymph nodes (PLN) and spleen. The spleen responded more rapidly in developin specific antibody-forming cells (AFC) than the lymph nodes did, in spite of the fact that antigens reach the spleen only after pasing several lymplh several lymph node stations. This difference between lymph nodes and spleen in development AFC was partivularly significant with respect to the responses to the responses to TI (both type 1 and type 2) antigens. No differences in the distribyution of specific AFC in PLN and spleen were oseved after immunization with TI and TD antigens. Results are discussed with respect to the relative contributions of lymph nodes and spleen to immune responses to antigens injected subcutaneously.     

Development of a Cell Patterning Technique Using Poly(Ethylene Glycol) Disilane

A simple technique for controlling cell adhesion on glass substrates by surface modification using a commercially available poly(ethylene glycol) (PEG) disilane, which can bind directly to glass in a single step, in combination with photolithographic micropatterning, was developed, characterized, and evaluated for patterning of HepG2 hepatoma cells and 3T3 fibroblasts. The optimal concentration of PEG-disilane for surface modification was 5 mM, and patterning of strongly adherent cells such as HepG2 required the chelation of divalent metal cations in order to inhibit nonspecific binding and cell aggregation. Whereas the average thickness of the PEG-disilane layer was 18±3.5 nm, the perimeters of patterned areas of exposed glass exhibited ridges of average height 857±50 nm, which may have aided in constraining cell spreading and migration. Although unpatterned PEG-treated substrates were hydrophilic (contact angle 46±1°), micropatterned surfaces behaved as if they were somewhat hydrophobic (contact angle 90°), necessitating special protocols for preventing deleterious dewetting of cells. For optimized protocols, the probability of adhesion of HepG2 cells to a patterned area of exposed glass was almost 15 times higher than the probability of adhesion to a PEG-treated background region of equal area. Our technique is useful for short- to intermediate-term patterning of cell or tissue morphology, e.g., for investigation of the effects of cell–cell interactions or cell geometry.     

Severe calcification in left main stem coronary artery stenosis visible on routine chest radiograph

Coronary calcification is a strong predictor of significant coronary stenosis in symptomatic patients. While discrete calcification within coronary arteries is only detected by sensitive methods such as computed tomography, severe calcification can already be seen on the plain chest radiograph. In this article, we describe a patient with a high grade left main stem coronary artery stenosis who presented with a severe focal calcification on the plain chest radiograph in projection of the offspring of the left coronary artery.     

Production of prostaglandin E2 in monocytes stimulated in vitro by Chlamydia trachomatis, Chlamydophila pneumoniae, and Mycoplasma fermentans

Chlamydia trachomatis (CT) as well as Chlamydophila pneumoniae (CP) cause chronic inflammatory diseases in humans. Persistently infected monocytes are involved in the pathogenesis by inducing mediators of inflammation. An in vitro system of chlamydial persistence in human peripheral blood monocytes (HPBM) was used to investigate prostaglandin E(2) (PGE(2)) production and the expression of the key enzyme for prostaglandin production, cyclooxygenase-2 (COX-2). PGE(2) production was determined by PGE(2)-ELISA of HPBM-culture supernatants. Cox-2 mRNA expression was measured by real-time RT-PCR of total RNA isolated from HPBM. Both, CT and CP, stimulated PGE(2) production of HPBM in vitro. Equivalent numbers of CT per host cell induced a higher PGE(2)-response compared to CP. The amount of synthesized PGE(2) depended on the chlamydial multiplicity of infection (MOI). Even at an MOI of 10 the amount of CT- and CP-induced prostaglandin, respectively, was lower than the amount of prostaglandin induced by E. coli lipopolysaccharide (LPS) at a concentration of 10microg/ml. In contrast to stimulation with LPS, Chlamydia-induced PGE(2) production as well as cox-2 mRNA decreased after day 1 post infection (p.i.). These data indicate that Chlamydia stimulate PGE(2) production in human monocytes. Since Chlamydia are often contaminated by mycoplasma, the influence of mycoplasma on the prostaglandin production was investigated additionally. Mycoplasma fermentans (MF) also stimulated PGE(2) production. The co-infection of mycoplasma and Chlamydia resulted in an additive effect in the production of PGE(2). Thus it is important to use host cells and Chlamydia free of mycoplasma contamination for the analysis of Chlamydia-induced prostaglandin production.     

Boric acid single dose pharmacokinetics after intravenous administration to man

Eighth young adult male volunteers with a basic (alimentary) plasma boric acid concentration of <0.10–0.46 mg/l were given a single dose of boric acid (562–611 mg) by 20 min IV infusion. The plasma concentration curves, followed for 3 days, best fitted a three-compartment open model, although two subjects had to be left out due to inconstant basal plasma concentration values or failure to fit to the three-compartment model. The 120 h urinary excretion was 98.7±9.1% of dose, Cl_tot 54.6±8.0 ml/min/1.73 m², t_1/2 21.0±4.9 h and distribution volumes V₁, V₂, and V₃: 0.251±0.099, 0.456±0.067 and 0.340±0.128 l/kg.     

Deciphering the molecular effects of non-ablative Er:YAG laser treatment in an in vitro model of the non-keratinized mucous membrane

Lasers in Medical Science -