By Altering Ocular Immune Privilege, Bone Marrow–derived Cells Pathogenically Contribute to DBA/2J Pigmentary Glaucoma

Pigment dispersion syndrome causes iris pigment release and often progresses to elevated intraocular pressure and pigmentary glaucoma (PG). Because melanin pigment can have adjuvant like properties and because the Gpnmb gene, which contributes to pigment dispersion in DBA/2J (D2) mice, is expressed in dendritic cells, we tested the hypothesis that ocular immune abnormalities participate in PG phenotypes. Strikingly, we show that D2 eyes exhibit defects of the normally immunosuppressive ocular microenvironment including inability of aqueous humor to inhibit T cell activation, failure to support anterior chamber (AC)-associated immune deviation, and loss of ocular immune privilege. Histologic analysis demonstrates infiltration of inflammatory leukocytes into the AC and their accumulation within the iris, whereas clinical indications of inflammation are typically very mild to undetectable. Importantly, some of these abnormalities precede clinical indications of pigment dispersal, suggesting an early role in disease etiology. Using bone marrow chimeras, we show that lymphohematopoietic cell lineages largely dictate the progression of pigment dispersion, the ability of the eye to support induction of AC-associated immune deviation, and the integrity of the blood/ocular barrier. These results suggest previously unsuspected roles for bone marrow–derived cells and ocular immune privilege in the pathogenesis of PG

A bovine lymphosarcoma cell line infected with theileria annulata exhibits an irreversible reconfiguration of host cell gene expression

Theileria annulata, an intracellular parasite of bovine lymphoid cells, induces substantial phenotypic alterations to its host cell including continuous proliferation, cytoskeletal changes and resistance to apoptosis. While parasite induced modulation of host cell signal transduction pathways and NFκB activation are established, there remains considerable speculation on the complexities of the parasite directed control mechanisms that govern these radical changes to the host cell. Our objectives in this study were to provide a comprehensive analysis of the global changes to host cell gene expression with emphasis on those that result from direct intervention by the parasite. By using comparative microarray analysis of an uninfected bovine cell line and its Theileria infected counterpart, in conjunction with use of the specific parasitacidal agent, buparvaquone, we have identified a large number of host cell gene expression changes that result from parasite infection. Our results indicate that the viable parasite can irreversibly modify the transformed phenotype of a bovine cell line. Fifty percent of genes with altered expression failed to show a reversible response to parasite death, a possible contributing factor to initiation of host cell apoptosis. The genes that did show an early predicted response to loss of parasite viability highlighted a sub-group of genes that are likely to be under direct control by parasite infection. Network and pathway analysis demonstrated that this sub-group is significantly enriched for genes involved in regulation of chromatin modification and gene expression. The results provide evidence that the Theileria parasite has the regulatory capacity to generate widespread change to host cell gene expression in a complex and largely irreversible manner

Probiotic consortiums: Structure and antagonistic activity against opportunistic bacteria and human normobiota (using the example of <i>Escherichia coli</i>) <i>in vitro</i>

Background. Using probiotic preparations based on consortia of microorganisms not only helps to restore the balance of the intestinal microbiota, but also increases the therapeutic effect of probiotics. Promising sources for obtaining probiotic consortia are milk products that have undergone natural fermentation with the help of spontaneously formed microbial consortia. The aim. To study the structure of five microbial consortia with probiotic properties from naturally fermented milk products and to assess in vitro their antagonistic activity against opportunistic bacteria and a representative of the human normobiota – Escherichia coli. Materials and methods. The structure of bacterial consortia was analyzed by sequencing methods. The antagonistic activity of the consortia was assessed by the disk diffusion method. Results. It has been established that the studied microbial consortiums are represented by Enterococcus spp. and Streptococcus spp. bacteria. In consortiums No. 1, No.  2, and No.  3, Enterococcus bacteria dominated, while in consortiums No.  4 and No. 5, Streptococcus dominated. Antagonistic activity was shown against four isolates of opportunistic bacteria: Klebsiella pneumoniae No.  493, Enterobacter hormaechei No. 372, Staphylococcus aureus No. 4 and Pseudomonas aeruginosa No. 25 IMB, as well as against one representative of the human normobiota – Escherichia coli No. 495. The highest growth delay zone is found in E. coli No. 495 isolate. Three test cultures (K. pneumoniae No. 509, E. coli ATCC25922 and P. aeruginosa No. 3 IMB) exhibited more dense growth around probiotic consortia. Conclusion. The results of the study showed that the effect of probiotic consortia differing in the composition of microorganisms can be neutral and bactericidal. The presence of antagonistic activity in the studied microbial consortia against multiresistant isolates of opportunistic bacteria is a prospect for creating probiotics with antibacterial properties

Gabapentin as add-on to morphine for severe neuropathic or mixed pain in children from age 3 months to 18 years - Evaluation of the safety, pharmacokinetics, and efficacy of a new gabapentin liquid formulation: Study protocol for a randomized controlled trial

Background: Gabapentin has shown efficacy in the treatment of chronic neuropathic or mixed pain in adults. Although pediatric pain specialists have extensive experience with gabapentin for the treatment of neuropathic pain, its use is off-label. Its efficacy and safety in this context have never been shown. The aim of this trial is to compare gabapentin with placebo as add-on to morphine for the treatment of severe chronic mixed or neuropathic pain in children. This trial is part of the European Union Seventh Framework Programme project Gabapentin in Paediatric Pain (GAPP) to develop a pediatric use marketing authorization for a new gabapentin suspension. Methods/design: The GAPP-2 study is a randomized, double-blind, placebo-controlled, multicenter superiority phase II study in children with severe chronic neuropathic or mixed pain. Its primary objective is to evaluate the efficacy of a gabapentin liquid formulation as adjunctive therapy to morphine. Sixty-six eligible children 3 months to 18 years of age with severe pain (pain scores ≥ 7), stratified in three age groups, will be randomized to receive gabapentin (to an accumulating dose of 45 to 63 mg/kg/day, dependent on age) or placebo, both in addition to morphine, for 12 weeks. Randomization will be preceded by a short washout period, and treatment will be initiated by a titration period of 3 weeks. After the treatment period, medication will be tapered during 4 weeks. The primary endpoint is the average pain scores in the two treatment groups (average of two measures each day for 3 days before the end-of-study visit [V10] assessed by age-appropriate pain scales (Face, Legs, Activity, Cry, Consolability scale; Faces Pain Scale-Revised; Numeric Rating Scale). Secondary outcomes include percentage responders to treatment (subjects with 30% reduction in pain scale), number of episodes of breakthrough pain, number of rescue interventions, number of pain-free days, participant dropouts, quality of life (Pediatric Quality of Life Inventory), and acceptability of treatment. Outcomes will be measured at the end-of-study visit after 12 weeks of treatment at the optimal gabapentin dose. Groups will be compared on an intention-to-treat basis. Discussion: We hope to provide evidence that the combination of morphine and gabapentin will provide better analgesia than morphine alone and will be safe. We also aim to obtain confirmation of the recommended pediatric dose. Trial registration: EudractCT, 2014-004897-40. Registered on 7 September 2017. ClinicalTrials.gov, NCT03275012. Registered on 7 September 2017

A Dominant-Negative Mutation of Mouse Lmx1b Causes Glaucoma and Is Semi-lethal via LBD1-Mediated Dimerisation

Mutations in the LIM-homeodomain transcription factor LMX1B cause nail-patella syndrome, an autosomal dominant pleiotrophic human disorder in which nail, patella and elbow dysplasia is associated with other skeletal abnormalities and variably nephropathy and glaucoma. It is thought to be a haploinsufficient disorder. Studies in the mouse have shown that during development Lmx1b controls limb dorsal-ventral patterning and is also required for kidney and eye development, midbrain-hindbrain boundary establishment and the specification of specific neuronal subtypes. Mice completely deficient for Lmx1b die at birth. In contrast to the situation in humans, heterozygous null mice do not have a mutant phenotype. Here we report a novel mouse mutant Icst, an N-ethyl-N-nitrosourea-induced missense substitution, V265D, in the homeodomain of LMX1B that abolishes DNA binding and thereby the ability to transactivate other genes. Although the homozygous phenotypic consequences of Icst and the null allele of Lmx1b are the same, heterozygous Icst elicits a phenotype whilst the null allele does not. Heterozygous Icst causes glaucomatous eye defects and is semi-lethal, probably due to kidney failure. We show that the null phenotype is rescued more effectively by an Lmx1b transgene than is Icst. Co-immunoprecipitation experiments show that both wild-type and Icst LMX1B are found in complexes with LIM domain binding protein 1 (LDB1), resulting in lower levels of functional LMX1B in Icst heterozygotes than null heterozygotes. We conclude that Icst is a dominant-negative allele of Lmx1b. These findings indicate a reassessment of whether nail-patella syndrome is always haploinsufficient. Furthermore, Icst is a rare example of a model of human glaucoma caused by mutation of the same gene in humans and mice

Electrocatalytic Reduction of Peroxodisulfate in 0.5 M NaOH at Copper Electrodes. A Combined Quartz Microbalance and Rotating Ring/Disc Electrode Investigation

From an electrochemical investigation by means of an electrochemical quartz microbalance, a rotating disc electrode, and a ring/disc electrode, two mechanisms for the reduction of S2O82- became apparent. Besides the well-known outer-sphere cathodic reduction, a catalytic mechanism of S2O82- reduction operates in a potential range between the surface oxide region (≈-0.5 V/SCE) and -1.0 V/SCE. It involves the chemical oxidation of the copper surface to a soluble Cu(I) species. The catalytic mechanism is concluded to result from the specific interaction between S2O82- and the Cu surface modified by the presence of subsurface oxygen

Electrocatalytic Reduction Of Peroxodisulfate In 0.5 M Naoh At Copper Electrodes. A Combined Quartz Microbalance And Rotating Ring/disc Electrode Investigation

From an electrochemical investigation by means of an electrochemical quartz microbalance, a rotating disc electrode, and a ring/disc electrode, two mechanisms for the reduction of S2O82- became apparent. Besides the well-known outer-sphere cathodic reduction, a catalytic mechanism of S2O82- reduction operates in a potential range between the surface oxide region (≈-0.5 V/SCE) and -1.0 V/SCE. It involves the chemical oxidation of the copper surface to a soluble Cu(I) species. The catalytic mechanism is concluded to result from the specific interaction between S2O82- and the Cu surface modified by the presence of subsurface oxygen.1011424112414Frumkin, A.N., (1933) Z. Phys. Chem. (Munich), 164, p. 121Wolf, W., Ye, J., Purgand, M., Eiswirth, M., Doblhofer, K., (1992) Ber. Bunsen-Ges. Phys. Chem., 96, p. 1797Flätgen, G., Krischer, K., Ertl, G., (1995) Z. Naturforsch., 50 A, p. 1097Flätgen, G., Krischer, K., (1995) J. Chem. Phys., 103, p. 5428Flätgen, G., Krischer, K., Pettinger, B., Doblhofer, K., Junkes, H., Ertl, G., (1995) Science, 269, p. 668Flätgen, G., Krischer, K., (1995) Phys. Rev. E, 51, p. 3997Flätgen, G., Krischer, K., Ertl, G., J. Electroanal. Chem., , in pressKoper, M.T.M., (1996) Ber. Bunsen-Ges. Phys. Chem., 100, p. 497Damaskin, B.B., Safonov, V.A., Fedorovich, N.V., (1993) J. Electroanal. Chem., 349, p. 1Latimer, W.M., (1952) The Oxidation States of the Elements and Their Potentials in Aqueous Solutions, , Prentice-Hall: New YorkLevie, M.G., Migliorini, E., Ercolini, G., (1908) Gazz. Chim. Ital., 38, p. 583Bond, G.C., Hill, G.M., Tennison, R., (1959) J. Chem. Soc., p. 33Müller, L., (1967) J. Electroanal. Chem., 13, p. 275Müller, L., Wetzel, R., Otto, H., (1970) J. Electroanal. Chem., 24, p. 175Mark, H.B., Anson, F.C., (1963) J. Electroanal. Chem., 6, p. 251Burke, L.D., O'Sullivan, J.F., O'Dwyer, K.J., Scannell, R.A., Ahern, M.J.G., McCarthy, M.M., (1990) J. Electrochem. Soc., 137, p. 2476Desilvestro, J., Weaver, M.J., (1987) J. Electroanal. Chem., 234, p. 237Ye, J., Wolf, W., Doblhofer, K., Eiswirth, M., (1994), Unpublished work. Wolf, W. Ph.D. Thesis, Frei Universität Berlin, BerlinHärtinger, S., Pettinger, B., Doblhofer, K., (1995) J. Electroanal. Chem., 397, p. 335Soares, D.M., (1993) Meas. Sci. Technol., 4, p. 549Fruböse, C., Doblhofer, K., Soares, D.M., (1993) Ber. Bunsen-Ges. Phys. Chem., 97, p. 475Sauerbrey, G., (1959) Z. Phys., 155, p. 206Albery, W.J., Bruckenstein, S., (1966) Trans. Faraday Soc., 62, p. 1920Ikemiya, N., Kubo, T., Hara, S., (1995) Surf. Sci., 323, p. 81Strehblow, H.H., Titze, B., (1980) Electrochim. Acta, 25, p. 839Shirkhanzadeh, M., Thompson, G.E., Ashworth, V., (1990) Corr. Sci., 31, p. 293Miller, B., (1969) J. Electrochem. Soc., 116, p. 1675Tindall, G.W., Bruckenstein, S., (1969) J. Electroanal. Chem., 22, p. 367Schubert, H., Tegetmeyer, U., Schlögl, R., (1994) Cat. Lett., 28, p. 383Schedl-Niedrig, T., Bao, X., Muhler, M., Neisius, T., Schlögl, R., (1995) Annual Fachbeirat of Fritz-Haber-Institut of the Max-Planck-Gesellschaft, , Abstract AC12, BerlinWerner, H., Demuth, D., Schubert, H., Weinberg, G., Braun, T., Herein, D., Schlögl, R., (1995) Annual Fachbeirat of Fritz-Haber-Institut of the Max-Planck-Gesellschaft, , Abstract AC13, BerlinPolak, M., (1994) Surf. Sci., 321, p. 249Carley, A.F., Davies, P.R., Roberts, M.W., Vicent, D., (1994) Top. Catal., 1, p. 35Davies, P.R., Roberts, M.W., Shukla, N., Vicent, D., (1995) Surf. Sci., 325, p. 5

Col4a1 mutation causes endoplasmic reticulum stress and genetically modifiable ocular dysgenesis.

Ocular anterior segment dysgenesis (ASD) is a complex and poorly understood group of conditions. A large proportion of individuals with ASD develop glaucoma, a leading cause of blindness resulting from retinal ganglion cell death. Optic nerve hypoplasia is thought to have distinct causes and is a leading cause of blindness in children. Here, we show that a mutation in the type IV collagen alpha 1 (Col4a1) gene can cause both ASD and optic nerve hypoplasia. COL4A1 is a major component of almost all basement membranes. The mutation results in non-secretion of the mutant COL4A1 proteins, which instead accumulate within cells. Basement membrane abnormalities may, therefore, contribute to the phenotype. The mutation also induces endoplasmic reticulum stress and so intracellular stress may contribute to pathogenesis. The overall consequence of the Col4a1 mutation depends on genetic context. In one genetic context, the mutation causes severe ASD with intraocular pressure abnormalities and optic nerve hypoplasia. In a different genetic context, both the ASD and optic nerve hypoplasia are rescued, and we have identified a single dominant locus that confers the phenotypic modification

Intraocular pressure in genetically distinct mice: an update and strain survey.

BACKGROUND: Little is known about genetic factors affecting intraocular pressure (IOP) in mice and other mammals. The purpose of this study was to determine the IOPs of genetically distinct mouse strains, assess the effects of factors such as age, sex and time of day on IOP in specific strain backgrounds, and to assess the effects of specific candidate gene mutations on IOP. RESULTS: Based on over 30 studied mouse strains, average IOP ranges from approximately 10 to 20 mmHg. Gender does not typically affect IOP and aging results in an IOP decrease in some strains. Most tested strains exhibit a diurnal rhythm with IOP being the highest during the dark period of the day. Homozygosity for a null allele of the carbonic anhydrase II gene (Car2n) does not alter IOP while homozygosity for a mutation in the leptin receptor gene (Leprdb) that causes obesity and diabetes results in increased IOP. Albino C57BL/6J mice homozygous for a tyrosinase mutation (Tyrc-2J) have higher IOPs than their pigmented counterparts. CONCLUSIONS: Genetically distinct mouse strains housed in the same environment have a broad range of IOPs. These IOP differences are likely due to interstrain genetic differences that create a powerful resource for studying the regulation of IOP. Age, time of day, obesity and diabetes have effects on mouse IOP similar to those in humans and other species. Mutations in two of the assessed candidate genes (Lepr and Tyr) result in increased IOP. These studies demonstrate that mice are a practical and powerful experimental system to study the genetics of IOP regulation and disease processes that raise IOP to harmful levels