A paternal methyl donor-rich diet altered cognitive and neural functions in offspring mice.

Dietary intake of methyl donors, such as folic acid and methionine, shows considerable intra-individual variation in human populations. While it is recognized that maternal departures from the optimum of dietary methyl donor intake can increase the risk for mental health issues and neurological disorders in offspring, it has not been explored whether paternal dietary methyl donor intake influences behavioral and cognitive functions in the next generation. Here, we report that elevated paternal dietary methyl donor intake in a mouse model, transiently applied prior to mating, resulted in offspring animals (methyl donor-rich diet (MD) F1 mice) with deficits in hippocampus-dependent learning and memory, impaired hippocampal synaptic plasticity and reduced hippocampal theta oscillations. Gene expression analyses revealed altered expression of the methionine adenosyltransferase Mat2a and BK channel subunit Kcnmb2, which was associated with changes in Kcnmb2 promoter methylation in MD F1 mice. Hippocampal overexpression of Kcnmb2 in MD F1 mice ameliorated altered spatial learning and memory, supporting a role of this BK channel subunit in the MD F1 behavioral phenotype. Behavioral and gene expression changes did not extend into the F2 offspring generation. Together, our data indicate that paternal dietary factors influence cognitive and neural functions in the offspring generation.

Towards allele-level human leucocyte antigens genotyping - assessing two next-generation sequencing platforms: Ion Torrent Personal Genome Machine and Illumina MiSeq.

Human leucocyte antigens (HLA) typing has been a challenge due to extreme polymorphism of the HLA genes and limitations of the current technologies and protocols used for their characterization. Recently, next-generation sequencing techniques have been shown to be a well-suited technology for the complete characterization of the HLA genes. However, a comprehensive assessment of the different platforms for HLA typing, describing the limitations and advantages of each of them, has not been presented. We have compared the Ion Torrent Personal Genome Machine (PGM) and Illumina MiSeq, currently the two most frequently used platforms for diagnostic applications, for a number of metrics including total output, quality score per position across the reads and error rates after alignment which can all affect the accuracy of HLA genotyping. For this purpose, we have used one homozygous and three heterozygous well-characterized samples, at HLA-A, HLA-B, HLA-C, HLA-DRB1 and HLA-DQB1. The total output of bases produced by the MiSeq was higher, and they have higher quality scores and a lower overall error rate than the PGM. The MiSeq also has a higher fidelity when sequencing through homopolymer regions up to 9 bp in length. The need to set phase between distant polymorphic sites was more readily achieved with MiSeq using paired-end sequencing of fragments that are longer than those obtained with PGM. Additionally, we have assessed the workflows of the different platforms for complexity of sample preparation, sequencer operation and turnaround time. The effects of data quality and quantity can impact the genotyping results; having an adequate amount of good quality data to analyse will be imperative for confident HLA genotyping. The overall turnaround time can be very comparable between the two platforms; however, the complexity of sample preparation is higher with PGM, while the actual sequencing time is longer with MiSeq.

Identification and characterization of a novel HLA-B hybrid allele, B*08:132 with Next Generation Sequencing.

The new allele is a hybrid between B*08:01:01 and B*42:01:01.

Minimally invasive surgery technique for re-enclavation of prepupillary iris-claw IOLs.

BACKGROUND: Disenclavation is a common complication of prepupillary iris-claw intraocular lenses (IOL). We present a new minimally invasive revision surgery technique for reenclavation of prepupillary iris-claw IOLs using standard 23 Gauge (G) vitrectomy instruments. HISTORY AND SIGNS: Three cases of revision surgery by unilaterally dislocated prepupillary iris-claw IOLs are presented. THERAPY AND OUTCOME: Two 20 G sideports 90 degrees apart were constructed. Healon 10® was injected to maintain the anterior chamber. A standard enclavation needle was introduced to rotate the optic into correct position and a 23 G endgrasping forceps was used to grasp and stabilize the IOL for enclavation. The reenclavation was successful in all three cases and the mean visual acuity improved from preoperatively 0.1 (range counting fingers [CF] to 0.25) to 0.6 (range 0.4 to 0.8) with no significant induction of astigmatism. CONCLUSIONS: This minimally invasive reenclavation technique for repositioning of the prepupillary iris claw IOL appears to lead to successful and rapid visual rehabilitation.

Evaluation of stapled hemorrhoidopexy for hemorrhoidal disease: 14-year experience from 800 cases.

AIM: The object of the present study was to assess results and document both the need for reoperations and the long-term outcomes. METHODS: Eight hundred patients with symptomatic grades II-IV hemorroidal disease (HD), mean age 52.3 years, were surgically managed from January 1999 to March 2013. One hundred and eight displayed comorbidity of other anal pathology. All patients underwent stapled hemorrhoidopexy (SH) or double SH, combined with accessory anal procedures in 90 cases. Distance from dentate line to staple line and width of resected doughnut were recorded. Postoperative pain was measured. RESULTS: Mean measured distance of staple to dentate line was 2.6 cm. Mean hospital stay was 1.2 days. All patients were clinically examined at 1, 4 and 12 weeks, scheduled to be monitored annually for three years and instructed thereafter to contact us for any anorectal problem. Early procedure-related complications that required reintervention occurred in 20 patients (2.5%). Patient satisfaction at 12 weeks was high (98.5%). Thirty-two patients (4%) developed late procedure-related complications that required surgery, with 24 (3%) displaying the most important recurrence. Quite low stapling caused severe pain or stenosis; inadequate mucosectomy was related to stenosis or recurrence. A learning curve was observed over time leading to significant reduction in late reoperations. CONCLUSION: Considerable experience of SH in the treatment of grades II-IV HD confirms it as safe and effective procedure with sustained favorable results. Meticulous technique is essential to avoid complications and improve outcomes in terms of low recurrence and reintervention rates.

Determining performance characteristics of an NGS-based HLA typing method for clinical applications.

This study presents performance specifications of an in-house developed human leukocyte antigen (HLA) typing assay using next-generation sequencing (NGS) on the Illumina MiSeq platform. A total of 253 samples, previously characterized for HLA-A, -B, -C, -DRB1 and -DQB1 were included in this study, which were typed at high-resolution using a combination of Sanger sequencing, sequence-specific primer (SSP) and sequence-specific oligonucleotide probe (SSOP) technologies and recorded at the two-field level. Samples were selected with alleles that cover a high percentage of HLA specificities in each of five different race/ethnic groups: European, African-American, Asian Pacific Islander, Hispanic and Native American. Sequencing data were analyzed by two software programs, Omixon's target and GenDx's NGSengine. A number of metrics including allele balance, sensitivity, specificity, precision, accuracy and remaining ambiguity were assessed. Data analyzed by the two software systems are shown independently. The majority of alleles were identical in the exonic sequences (third field) with both programs for HLA-A, -B, -C and -DQB1 in 97.7% of allele determinations. Among the remaining discrepant genotype calls at least one of the analysis programs agreed with the reference typing. Upon additional manual analysis 100% of the 2530 alleles were concordant with the reference HLA genotypes; the remaining ambiguities did not exceed 0.8%. The results demonstrate the feasibility and significant benefit of HLA typing by NGS as this technology is highly accurate, eliminates virtually all ambiguities, provides complete sequencing information for the length of the HLA gene and forms the basis for utilizing a single methodology for HLA typing in the immunogenetics labs.

Mouse cathepsin F: cDNA cloning, genomic organization and chromosomal assignment of the gene.

A murine cysteine protease of the papain family was identified by dbEST-database search. A 1.87kb full-length cDNA encoding a predicted polypeptide of 462 amino acids was sequenced. Since the encoded polypeptide shows more than 80% sequence identity with human cathepsin F, it is most likely that this cDNA represents the murine homologue of cathepsin F, and it was therefore named accordingly. Murine cathepsin F exhibits a domain structure typical for papain-like cysteine proteases, a 20 amino acid N-terminal hydrophobic signal sequence followed by an extraordinarily long propeptide of 228 amino acids and the domain of the mature protease comprising 214 amino acids. The mature region contains all features characteristic of a papain-like cysteine protease, including the highly conserved cysteine, histidine and asparagine residues of the 'catalytic triad'. Genomic clones covering the murine cathepsin F gene were isolated. The mouse cathepsin F gene consists of 14 exons and 13 introns and spans 5.8kb. Murine cathepsin F was mapped to chromosome 19, a region with synteny homology to a region of human chromosome 11 to which human cathepsin F has been mapped previously. Northern blot analysis of RNA from multiple tissues revealed a ubiquitous expression of cathepsin F in mouse and man.

Ion-selective electrodes for the H2-receptor antagonists cimetidine and ranitidine.

Liquid-membrane and polyvinyl chloride (PVC)-matrix ion-selective electrodes (ISE) that respond to the cationic forms of cimetidine and ranitidine are described. The ion-exchangers were the salts of cimetidine and ranitidine with tetrakis(m-chlorophenyl)borate dissolved in p-nitrocumene or entrapped in PVC polymer in the presence of 2-nitrophenyl octyl ether as plasticizer. The electrodes exhibited a near-Nernstian response in the range 10(-2)-10(-6)M (working pH range 2-7) for ranitidine, and 10(-2)-2 X 10(-5)M (pH 2-6) for cimetidine. Very small PVC-matrix ISE with internal diameters as small as 0.035 inches were constructed and used in combination with small cuvettes, so that measurements could be carried out in 250 muL of stirred solution. The electrodes were applied successfully for the determination of the pKa of the protonated bases and for the determination of the drugs in pharmaceutical preparations. New selective and effective solid-state extraction procedures are described for the extraction of ranitidine from urine and serum samples. Potentiometric methods were developed for the determination of ranitidine in urine and serum samples during a pharmacokinetic experiment.

Dissolution studies of drug formulations using ion-selective electrodes as sensors in an air-segmented continuous flow analyzer.

The application of drug ion-selective electrodes as sensors for the direct determination of the released drug in a continuous-flow analyzer for automated dissolution studies is described. Flow-through electrodes, selective to chlorpromazine, amitriptyline, propantheline, cimetidine, and ranitidine, have been constructed and used for the dissolution studies of 18 dosage forms using the rotating basket apparatus. The dissolution profiles are obtained in the form of potential peaks versus time.

[Smartphone-operated detonation of a firecracker resulting in a macular hole]. / Makulaforamen durch Smartphone-gesteuerte Fernzündung eines Feuerwerkskörpers.

A 22-year-old man presented to the emergency room on New Years Eve after a firecracker had exploded next to his left eye. Besides injuries to the eyelid, conjunctival and corneal trauma, a commotio retinae at the temporal periphery and the central retina were revealed funduscopically. A small full macular hole was detected by spectral domain optical coherence tomography scan (OCT). After topical treatment with tobramycin, fluorometholone and lubrication eye drops the best-corrected vision improved from 0.2 to 0.8 within 3 months and the follow-up OCT revealed a spontaneous closure of the macular hole.

Bimanual anterior segment revision surgery for anterior capsule contraction syndrome associated with anterior flexion of intraocular lens haptics.

PURPOSE: To report the incidence of anterior capsule contraction syndrome (ACCS) and to present a novel minimally invasive bimanual technique for anterior segment revision surgery associated with ACCS with anterior flexion of the intraocular lens haptics. METHODS: A consecutive cohort of 268 eyes of 161 patients undergoing phacoemulsification and implantation of the same type of hydrophilic acrylic aspheric intraocular lens cohort were analysed and a novel technique of minimally invasive bimanual technique for anterior segment revision surgery is described. RESULTS: We identified four eyes (1.5%) of three patients with advanced ACCS. Successful restoration of a clear visual axis with minimal induction of astigmatism and rapid visual rehabilitation was achieved in all four cases. CONCLUSION: This technique is a safe and minimally invasive alternative to laser or vitrector-cut capsulotomy to restore a clear visual axis. In cases of advanced ACCS, it offers the option for haptic reposition or amputation.

Filling the gaps - the generation of full genomic sequences for 15 common and well-documented HLA class I alleles using next-generation sequencing technology.

Many common and well-documented (CWD) HLA alleles have only been partially characterized. The DNA sequence of these incomplete alleles, as published in the IMGT/HLA database, is most often limited to exons that code for the extracellular domains of the mature protein. Here we describe the application of next-generation sequencing technology to obtain full length genomic sequence from a single long-range PCR amplicon for 15 common and well-documented HLA Class I alleles. This technology is well suited to fill in the gaps of the current HLA allele sequence database which is largely incomplete. A more comprehensive catalog of HLA allele sequences would be beneficial in the evaluation of mismatches in transplantation, studies of population genetics, the evolution of HLAs, regulatory mechanisms and HLA expression, and issues related to the genomic organization of the MHC.

Bone marrow lesions: evaluation with fat-suppression turbo spin echo MR imaging at 0.5 T.

The purpose of this study was the assessment of the diagnostic value of fat-suppression T2-weighted images for a variety of bone marrow lesions. We performed 40 studies of the axial or appendicular skeleton in 33 patients (age range 4-80 years) with neoplastic, inflammatory or traumatic lesions with a 0.5 T system (Gyroscan T5, Philips Medical Systems, Best, The Netherlands). Fat-suppression T2-weighted images [turbo spin echo (TSE) with spectral presaturation with inversion recovery (SPIR)] were obtained in addition to the routine T1-weighted SE and T2-weighted TSE sequences. Fat-suppression TSE T2-weighted images were better than standard TSE T2-weighted images in 25 studies. In 11 of them demonstration and characterization of the lesions (known from T1-weighted images) was possible only after fat suppression. In the other 14 patients demonstration of the full extent of the lesion especially to the nearby soft tissues was possible only after fat suppression. In 13 studies no advantage was conferred by SPIR, whereas in two instances T2-weighted images were better. Fat-suppression T2-weighted images are diagnostically useful in a variety of lesions of the musculoskeletal system, but their limitations should be known.