The effect of coolant flow on surface roughness in metal cutting

The object of this thesis is to evaluate the effect of flood and spray mist application of a cutting fluid upon surface roughness in metal cutting

Deep-intronic ATM mutation detected by genomic resequencing and corrected in vitro by antisense morpholino oligonucleotide (AMO)

Recent development of next-generation DNA sequencing (NGS) techniques is changing the approach to search for mutations in human genetic diseases. We applied NGS to study an A-T patient in which one of the two expected mutations was not found after DHPLC, cDNA sequencing and MLPA screening. The 160-kb ATM genomic region was divided into 31 partially overlapping fragments of 4–6 kb and amplified by long-range PCR in the patient and mother, who carried the same mutation by segregation. We identified six intronic variants that were shared by the two genomes and not reported in the dbSNP(132) database. Among these, c.1236-405C>T located in IVS11 was predicted to be pathogenic because it affected splicing. This mutation creates a cryptic novel donor (5′) splice site (score 1.00) 405 bp upstream of the exon 12 acceptor (3′) splice site. cDNA analysis showed the inclusion of a 212-bp non-coding ‘pseudoexon' with a premature stop codon. We validated the functional effect of the splicing mutation using a minigene assay. Using antisense morpholino oligonucleotides, designed to mask the cryptic donor splice-site created by the c.1236-405C>T mutation, we abrogated the aberrant splicing product to a wild-type ATM transcript, and in vitro reverted the functional ATM kinase impairment of the patients' lymphoblasts. Resequencing is an effective strategy for identifying rare splicing mutations in patients for whom other mutation analyses have failed (DHPLC, MLPA, or cDNA sequencing). This is especially important because many of these patients will carry rare splicing variants that are amenable to antisense-based correction

The de Morton Mobility Index (DEMMI) provides a valid method for measuring and monitoring the mobility of patients making the transition from hospital to the community: an observational study

QuestionIsthe de Morton Mobility Index (DEMMI) valid for measuring the mobility of patients making the transition from hospital to the community?DesignObservational cohort study.Participants696 consecutive patients admitted to 11 Transition Care Programs for multidisciplinary care in Victoria and Tasmania during a 6-month period. The DEMMI and Modified Barthel Index were administered within 5 working days of admission and discharge from the Transition Care Program.Outcome measuresThe DEMMI and Modified Barthel Index.ResultsNeither the DEMMI nor the Modified Barthel Index had a floor or ceiling effect. Similar evidence of convergent, discriminant and known-groups validity were obtained for each instrument. The DEMMI was significantly more responsive to change than the Modified Barthel Index using criterion- and distribution-based methods. The minimum clinically important difference estimates represented similar proportions of the scale width for the DEMMI and Modified Barthel Index and were similar using criterion- and distribution-based estimates. Rasch analysis identified the DEMMI as essentially unidimensional in a Transition Care Program cohort and therefore can be applied to obtain interval level measurement. Rasch analysis demonstrated that the DEMMI was administered similarly by physiotherapists and allied health assistants under the direction of a physiotherapist.ConclusionThe DEMMI and Modified Barthel Index are both valid measures of activity limitation for Transition Care Program patients. The DEMMI has a broader scale width, provides interval level measurement, and is significantly more responsive to change than the Modified Barthel Index for measuring the mobility of Transition Care Program patient

Analysis of the DNA methylation pattern of the promoter region of calcitonin gene-related peptide 1 gene in patients with episodic migraine: An exploratory case-control study

Recent studies suggested that epigenetic mechanisms, including DNA methylation, may be involved in migraine pathogenesis. The calcitonin gene-related peptide (CGRP), encoded by calcitonin gene-related peptide 1 (CALCA) gene, plays a key role in the disease. The aim of the study was to evaluate DNA methylation of CALCA gene in patients with episodic migraine. 22 patients with episodic migraine (F/M 15/7, mean age 39.7 ± 13.4 years) and 20 controls (F/M 12/8, mean age 40.5 ± 14.8 years) were recruited. Genomic DNA was extracted from peripheral blood. Cytosine-to-thymine conversion was obtained with sodium bisulfite. The methylation pattern of two CpG islands in the promoter region of CALCA gene was analyzed. No difference of methylation of the 30 CpG sites at the distal region of CALCA promoter was observed between migraineurs and controls. Interestingly, in patients with episodic migraine the methylation level was lower in 2 CpG sites at the proximal promoter region (CpG −1461, p = 0.037, and −1415, p = 0.035, respectively). Furthermore, DNA methylation level at different CpG sites correlates with several clinical characteristics of the disease, as age at onset, presence of nausea/vomiting, depression and anxiety (p < 0.05). In conclusion, we found that DNA methylation profile in two CpG sites at the proximal promoter region of CALCA is lower in migraineurs when compared to controls. Intriguingly, the −1415 hypomethylated unit is located at the CREB binding site, a nuclear transcription factor. In addition, we found a correlation between the level of CALCA methylation and several clinical features of migraine. Further studies with larger sample size are needed to confirm these results

Postnatal Changes in K+/Cl- Cotransporter-2 Expression in the Forebrain of Mice Bearing a Mutant Nicotinic Subunit Linked to Sleep-Related Epilepsy

ABSTRACT The Na+/K+/Cl- cotransporter-1 (NKCC1) and the K+/Cl- cotransporter-2 (KCC2) set the transmembrane Cl- gradient in the brain, and are implicated in epileptogenesis. We studied the postnatal distribution of NKCC1 and KCC2 in wild-type (WT) mice, and in a mouse model of sleep-related epilepsy, carrying the mutant \u3b22-V287L subunit of the nicotinic acetylcholine receptor (nAChR). In WT neocortex, immunohistochemistry showed a wide distribution of NKCC1 in neurons and astrocytes. At birth, KCC2 was localized in neuronal somata, whereas at subsequent stages it was mainly found in the somatodendritic compartment. The cotransporters\u2019 expression was quantified by densitometry in the transgenic strain. KCC2 expression increased during the first postnatal weeks, while the NKCC1 amount remained stable, after birth. In mice expressing \u3b22-V287L, the KCC2 amount in layer V of prefrontal cortex (PFC) was lower than in the control littermates at postnatal day 8 (P8), with no concomitant change in NKCC1. Consistently, the GABAergic excitatory to inhibitory switch was delayed in PFC layer V of mice carrying \u3b22-V287L. At P60, the amount of KCC2 was instead higher in mice bearing the transgene. Irrespective of genotype, NKCC1 and KCC2 were abundantly expressed in the neuropil of most thalamic nuclei since birth. However, KCC2 expression decreased by P60 in the reticular nucleus, and more so in mice expressing \u3b22-V287L. Therefore, a complex regulatory interplay occurs between heteromeric nAChRs and KCC2 in postnatal forebrain. The pathogenetic effect of \u3b22-V287L may depend on altered KCC2 amounts in PFC during synaptogenesis, as well as in mature thalamocortical circuits

New insights into potocki-shaffer syndrome: Report of two novel cases and literature review

Potocki-Shaffer syndrome (PSS) is a rare non-recurrent contiguous gene deletion syndrome involving chromosome 11p11.2. Current literature implies a minimal region with haploinsufficiency of three genes, ALX4 (parietal foramina), EXT2 (multiple exostoses), and PHF21A (craniofacial anomalies, and intellectual disability). The rest of the PSS phenotype is still not associated with a specific gene. We report a systematic review of the literature and included two novel cases. Because deletions are highly variable in size, we defined three groups of patients considering the PSS-genes involved. We found 23 full PSS cases (ALX4, EXT2, and PHF21A), 14 cases with EXT2-ALX4, and three with PHF21A only. Among the latter, we describe a novel male child showing developmental delay, caf&eacute;-au-lait spots, liner postnatal overgrowth and West-like epileptic encephalopathy. We suggest PSS cases may have epileptic spasms early in life, and PHF21A is likely to be the causative gene. Given their subtle presentation these may be overlooked and if left untreated could lead to a severe type or deterioration in the developmental plateau. If our hypothesis is correct, a timely therapy may ameliorate PSS phenotype and improve patients&rsquo; outcomes. Our analysis also shows PHF21A is a candidate for the overgrowth phenotype